**Part 5**

**Thin Films, Membranes, Ceramic** 

588 Scanning Electron Microscopy

[30] K G Mor, O K Varghese, M Paulose, K Shankar, C A Grimes, *Solar Energy Materials &* 

[31] A Jaroenworaluck, D Regonini, C R Bowen, R Stevens, D Allsopp, J Mater Sci., 42 (2007)

[26] T Shibata and Y C Zhu, *Corrosion Science*., 37 (1995) 253. [27] K H Kim and N Ramaswamy, *Dental Mats J*., 28 (2009) 20.

[29] L Bartlett, *Optics & Laser Technol*., 38 (2006) 440.

*Solar Cells*., 90 (2006) 2011.

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[28] Metalast Technical Bulletin. (NV: Metalast International Inc, 2000).

[32] M K Kyung, S J Doo, S H Cheol, *Nanotechnology*., 22 (2011) 1-17.

**30** 

*Egypt* 

R. M. Abd El-Baky

**Application of Scanning Electron** 

**Microscopy for the Morphological** 

**Study of Biofilm in Medical Devices** 

*Microbiology Department, Faculty of Pharmacy, Minia University, Minia,* 

The widespread use of medical devices has caused a great advance in the management of many diseases. Indwelling medical devices are being increasingly used for the treatment of functional deficits in numerous medical fields. Urinary tract infections (UTIs) represent the most commonly acquired bacterial infection. The risk of developing a urinary tract infection increases significantly with the use of indwelling devices such as catheters and urethral stents/sphincters. Although these catheters are valuable, they also have complications, the major complications are: encrustation, stone formation and biofilm formation. Microbial biofilms may pose a public health problem for persons requiring catheterization as the microorganisms in biofilms are difficult or impossible to be treated

Several approaches have been studied to prevent the formation of biofilms or to eradicate biofilm associated microorganisms. Some of these depends on coating medical devices with silver, antiseptics or by producing radio-opacity in catheters by the silicone material and

One approach to overcome the antimicrobial resistance of biofilm bacteria would be to enhance the penetration of agents through the biofilm matrix. Many trials were done to increase the efficacy of antimicrobial agents using some agents such as protamine sulfate

This work was done to determine whether N- acetylcysteine could aid ciprofloxacin in

The use of biomaterials in the urinary tract dates back to ancient times, when the Egyptians described using lead and papyrus to construct urinary catheters (Bitschay and Brodny, 1956). Today, the majority of biomaterials used in urology are made from synthetic polymeric compounds, which were originally developed in the plastic injury. In the process of endourological development, a great variety of foreign bodies have been invented besides urethral catheters like ureter, prostatic stents, percutaneous nephrostomy, penile, testicular implants, and artificial urinary sphincters (AUS). Although the tendency of patients being predisposed to infections due to foreign bodies has been recognized since the fourteenth

some depends on the use of antimicrobial agents or non antimicrobial agents.

penetrating biofilm formed by some microorganisms on ureteral stents.

(anticoagulant)**,** EDTA, sodium citrate and penicillamine**.**

**1. Introduction** 

by antimicrobial agents.

## **Application of Scanning Electron Microscopy for the Morphological Study of Biofilm in Medical Devices**

R. M. Abd El-Baky *Microbiology Department, Faculty of Pharmacy, Minia University, Minia, Egypt* 

## **1. Introduction**

The widespread use of medical devices has caused a great advance in the management of many diseases. Indwelling medical devices are being increasingly used for the treatment of functional deficits in numerous medical fields. Urinary tract infections (UTIs) represent the most commonly acquired bacterial infection. The risk of developing a urinary tract infection increases significantly with the use of indwelling devices such as catheters and urethral stents/sphincters. Although these catheters are valuable, they also have complications, the major complications are: encrustation, stone formation and biofilm formation. Microbial biofilms may pose a public health problem for persons requiring catheterization as the microorganisms in biofilms are difficult or impossible to be treated by antimicrobial agents.

Several approaches have been studied to prevent the formation of biofilms or to eradicate biofilm associated microorganisms. Some of these depends on coating medical devices with silver, antiseptics or by producing radio-opacity in catheters by the silicone material and some depends on the use of antimicrobial agents or non antimicrobial agents.

One approach to overcome the antimicrobial resistance of biofilm bacteria would be to enhance the penetration of agents through the biofilm matrix. Many trials were done to increase the efficacy of antimicrobial agents using some agents such as protamine sulfate (anticoagulant)**,** EDTA, sodium citrate and penicillamine**.**

This work was done to determine whether N- acetylcysteine could aid ciprofloxacin in penetrating biofilm formed by some microorganisms on ureteral stents.

The use of biomaterials in the urinary tract dates back to ancient times, when the Egyptians described using lead and papyrus to construct urinary catheters (Bitschay and Brodny, 1956). Today, the majority of biomaterials used in urology are made from synthetic polymeric compounds, which were originally developed in the plastic injury. In the process of endourological development, a great variety of foreign bodies have been invented besides urethral catheters like ureter, prostatic stents, percutaneous nephrostomy, penile, testicular implants, and artificial urinary sphincters (AUS). Although the tendency of patients being predisposed to infections due to foreign bodies has been recognized since the fourteenth

Application of Scanning Electron Microscopy for

**2.3 Development** 

**2.3.1 Reversible attachment** 

the Morphological Study of Biofilm in Medical Devices 593

Once at the surface, different physical, chemical and biological processes take place during this initial interaction between bacteria and the surface. On the abiotic surface, primary attachment between bacteria and the surface is mediated by non-specific interactions such as electrostatic, hydrophobic, or vander waals forces. On biotic surfaces such as tissues, Primary attachment is through specific molecular adhesion by lectin or adhesin. The surface appendages and structures that bear adhesins include fimbriae, flagella, capsule, outer membrane and other appendages. Bacteria bearing adhesins can approach receptors at a distance, form complexes, and ultimately settle onto substratum (Ofek and Doyle, 1994).

In this stage, The organism must be brought into close approximation of the surface either randomly by a stream of fluid flowing over a surface or by a directed fashion via chemotaxsis and motility. Once the organisms reach critical proximity of the surface

The net sum of attractive or repulsive forces generated between the two surfaces. These forces include electrostatic and hydrophobic interactions, steric hindrance, vander waals forces, temperature and hydrodynamic forces. Electrostatic interactions tend to favor repulsion, because most bacteria and inert surfaces are negatively charged. Hydrophobic interactions probably have greater influence on the outcome of primary adhesion depending on the molecules in the conditioned film. Repulsive forces can be over come by specific molecular interactions mediated by adhesins located on structures extending from the cell

It was observed that flagellar and twitching motility are necessary for *pseudomonas aeruginosa* biofilm development. As when biofilm formation to a biotic surface of polyvinyl chloride plates (PVC) by *P. aeruginosa* and its 2 mutants is compared, one mutant is defective in flagellar mediated motility and the other is defective in biogenesis of polar type of iv pili. The biofilm of wild type is followed using phase contrast microscopy. First, the strain formed a monolayer of cells on the a biotic surface (PVC) followed by the appearance of microcolonies that were dispersed through the monolayer of cells. Then using time lapse microscopy, it showed that microcolonies were formed by aggregation of cells present in the monolayer. As observed in the wild type, the strain defective in type iv pili formed a monolayer of cells on the PVC but cells failed to develop microcolonies suggesting that these structures play an important role in microcolonies formation, while very few cells of flagellar defective non-motile mutant are attached to PVC surface even after 8 hours incubation showing that the role of flagella and/ or motility in initial cell-surface adhesion is

After binding to the surface through exopolymeric matrix, bacterial cells start the process of irreversible adhesion, proliferation and accumulation as multilayered cell clusters. These extracellular matrices, composed of a mixture of materials such as polysaccharides, proteins, nucleic acids and other substances are considered to be essential in cementing bacterial cells together in the biofilm structure, in helping to trap and retain nutrients for biofilm growth

(usually < 1nm) the final determination of adhesion depends on:

surface such as pili (Carpentier and Cerf, 1993).

very important (Arora *et al*., 1998).

**2.3.2 Irreversible attachment** 

century, the mechanisms of device-related infections are still not completely understood (Tenke *et al*., 2006).

Bacterial adherence and the growth of bacteria on solid surfaces as biofilm are both naturally occurring phenomena. Biofilm formation affects many aspects of our life and also plays an important role in medicine involving the field of urology. It is able to build up under natural circumstances, for instance on the urothelium or prostate stones and in the presence of temporarily or permanently implanted foreign bodies. The frequently used urethral catheters, double J stents and transrenal drains provide just as perfect surfaces to bacteria to adhere. Biofilms can have a positive impact as well, namely lining the healthy intestine and the female genito-urinary tract. Biofilms have significant implications for clinical pharmacology, particularly related to antibiotic resistance, drug adsorption onto and pealing off devices, and minimum inhibitory concentrations of drugs required for effective therapy (Mardis and Kroeger, 1988).

## **2. What is a biofilm**

## **2.1 Definition**

Biofilm is defined as structured communities of microbial species embedded in a biopolymer matrix on either biotic (living tissues) or a biotic (inert non living material) substrata. The general theory of biofilm predominance states that the majority of bacteria grow in matrix-enclosed biofilms adherent to surfaces in all nutrient-sufficient aquatic ecosystems and that these sessile bacterial cells differ greatly from their planktonic counterparts (Costerton *et al.,* 1978). The reason for this ubiquity is that the protective layer and distinct metabolic states of bacteria within biofilms provide them with special resistance to host defences and antimicrobials, including natural antibiotics.

#### **2.2 Estructure**

The basic structure unit of the biofilm is the microcolony. A mature biofilm is composed of cells ( 10-15% by volume) and of glycocalyx (85-90%). The cells embedded in glycocalyx form gross structure resembling towers and mushrooms, sometimes as high as a few millimeters. Open channels are interspersed between the microcolonies resembling a primitive circulatory system. Water and nutrients enter these channels and contribute to nutrition and formation of mature biofilms. Waste products are also removed through this system.

The cells composing a biofilm can be of single species or more commonly are heterogenous species of bacteria and fungi. In the latter case the metabolic by-products of one organism might serve to support the growth of another, and the adhesion of one species might provide ligands allowing the attachment of others. A mature biofilms contains thousands of bacteria (Dunne, 2002).

The glycocalyx is mainly composed of bacterial exopolysaccharides. Other components are nucleic acids, minerals and proteins. When fully hydrated, the glycocalyx is predominantly water, with an anionic charge that creates a scavenging system for trapping and concentrating essential nutrients from the environment. Glycocalyx also provides a protective layer against biocides and this is the most prominent morphologic feature addressed in this chapter.

## **2.3 Development**

592 Scanning Electron Microscopy

century, the mechanisms of device-related infections are still not completely understood

Bacterial adherence and the growth of bacteria on solid surfaces as biofilm are both naturally occurring phenomena. Biofilm formation affects many aspects of our life and also plays an important role in medicine involving the field of urology. It is able to build up under natural circumstances, for instance on the urothelium or prostate stones and in the presence of temporarily or permanently implanted foreign bodies. The frequently used urethral catheters, double J stents and transrenal drains provide just as perfect surfaces to bacteria to adhere. Biofilms can have a positive impact as well, namely lining the healthy intestine and the female genito-urinary tract. Biofilms have significant implications for clinical pharmacology, particularly related to antibiotic resistance, drug adsorption onto and pealing off devices, and minimum inhibitory concentrations of drugs required for effective

Biofilm is defined as structured communities of microbial species embedded in a biopolymer matrix on either biotic (living tissues) or a biotic (inert non living material) substrata. The general theory of biofilm predominance states that the majority of bacteria grow in matrix-enclosed biofilms adherent to surfaces in all nutrient-sufficient aquatic ecosystems and that these sessile bacterial cells differ greatly from their planktonic counterparts (Costerton *et al.,* 1978). The reason for this ubiquity is that the protective layer and distinct metabolic states of bacteria within biofilms provide them with special resistance

The basic structure unit of the biofilm is the microcolony. A mature biofilm is composed of cells ( 10-15% by volume) and of glycocalyx (85-90%). The cells embedded in glycocalyx form gross structure resembling towers and mushrooms, sometimes as high as a few millimeters. Open channels are interspersed between the microcolonies resembling a primitive circulatory system. Water and nutrients enter these channels and contribute to nutrition and formation of

The cells composing a biofilm can be of single species or more commonly are heterogenous species of bacteria and fungi. In the latter case the metabolic by-products of one organism might serve to support the growth of another, and the adhesion of one species might provide ligands allowing the attachment of others. A mature biofilms contains thousands of

The glycocalyx is mainly composed of bacterial exopolysaccharides. Other components are nucleic acids, minerals and proteins. When fully hydrated, the glycocalyx is predominantly water, with an anionic charge that creates a scavenging system for trapping and concentrating essential nutrients from the environment. Glycocalyx also provides a protective layer against biocides and this is the most prominent morphologic feature

to host defences and antimicrobials, including natural antibiotics.

mature biofilms. Waste products are also removed through this system.

(Tenke *et al*., 2006).

therapy (Mardis and Kroeger, 1988).

**2. What is a biofilm** 

**2.1 Definition** 

**2.2 Estructure** 

bacteria (Dunne, 2002).

addressed in this chapter.

## **2.3.1 Reversible attachment**

Once at the surface, different physical, chemical and biological processes take place during this initial interaction between bacteria and the surface. On the abiotic surface, primary attachment between bacteria and the surface is mediated by non-specific interactions such as electrostatic, hydrophobic, or vander waals forces. On biotic surfaces such as tissues, Primary attachment is through specific molecular adhesion by lectin or adhesin. The surface appendages and structures that bear adhesins include fimbriae, flagella, capsule, outer membrane and other appendages. Bacteria bearing adhesins can approach receptors at a distance, form complexes, and ultimately settle onto substratum (Ofek and Doyle, 1994).

In this stage, The organism must be brought into close approximation of the surface either randomly by a stream of fluid flowing over a surface or by a directed fashion via chemotaxsis and motility. Once the organisms reach critical proximity of the surface (usually < 1nm) the final determination of adhesion depends on:

The net sum of attractive or repulsive forces generated between the two surfaces. These forces include electrostatic and hydrophobic interactions, steric hindrance, vander waals forces, temperature and hydrodynamic forces. Electrostatic interactions tend to favor repulsion, because most bacteria and inert surfaces are negatively charged. Hydrophobic interactions probably have greater influence on the outcome of primary adhesion depending on the molecules in the conditioned film. Repulsive forces can be over come by specific molecular interactions mediated by adhesins located on structures extending from the cell surface such as pili (Carpentier and Cerf, 1993).

It was observed that flagellar and twitching motility are necessary for *pseudomonas aeruginosa* biofilm development. As when biofilm formation to a biotic surface of polyvinyl chloride plates (PVC) by *P. aeruginosa* and its 2 mutants is compared, one mutant is defective in flagellar mediated motility and the other is defective in biogenesis of polar type of iv pili. The biofilm of wild type is followed using phase contrast microscopy. First, the strain formed a monolayer of cells on the a biotic surface (PVC) followed by the appearance of microcolonies that were dispersed through the monolayer of cells. Then using time lapse microscopy, it showed that microcolonies were formed by aggregation of cells present in the monolayer. As observed in the wild type, the strain defective in type iv pili formed a monolayer of cells on the PVC but cells failed to develop microcolonies suggesting that these structures play an important role in microcolonies formation, while very few cells of flagellar defective non-motile mutant are attached to PVC surface even after 8 hours incubation showing that the role of flagella and/ or motility in initial cell-surface adhesion is very important (Arora *et al*., 1998).

#### **2.3.2 Irreversible attachment**

After binding to the surface through exopolymeric matrix, bacterial cells start the process of irreversible adhesion, proliferation and accumulation as multilayered cell clusters. These extracellular matrices, composed of a mixture of materials such as polysaccharides, proteins, nucleic acids and other substances are considered to be essential in cementing bacterial cells together in the biofilm structure, in helping to trap and retain nutrients for biofilm growth

Application of Scanning Electron Microscopy for

(Tebbs *et al*., 1994).

(Jansen *et al*., 1988). c. Hydrophobicity:

b. Surface charge:

**2.4.2 Host factors** 

(Pascual, 2002).

**2.4.3 Microbial factors** 

interactions (Martinez-Martinez *et al*., 1991).

**2.4.4 The suspending medium** 

the Morphological Study of Biofilm in Medical Devices 595

venous catheters (CVC) varied with different polymer materials so that bacteria preferentially adhered to surface defects within minutes after infusing the catheters with contaminated buffer solution (Locci *et al.,* 1981). Another study examined the surface of five commercially available polyurethane CVCs by scanning electron microscopy and found that the catheters with the most surface irregularities had significantly more adherent bacteria compared to catheters with smoother surfaces

Biomaterial surface charge greatly influences adherence of microorganisms. Most microorganisms exhibit a negative surface charge in an aqueous environment. Therefore, a negatively charged biomaterial surface should lead to decreased adherence of microorganisms due to a repulsion effect between both negatively charged surfaces

Bacterial cells, which tend to have hydrophobic cell surfaces, are attracted to the hydrophobic surfaces of many biomaterials currently used in IMDs (Schierholz and Beuth, 2001). This hydrophobic interaction between the microorganisms and the biomaterial leads to increased adherence and subsequent biofilm formation. An increase in the surface hydrophilicity of the polymers leads to weakened hydrophobic

The biomaterials used in IMDs result in the activation of the host immune response leading to local tissue damage and the development of an immuno-incompetent, fibro-inflammatory zone that increases the susceptibility of the IMD to infection (Schierholz and Beuth, 2001). The deposition of proteinaceous layer (including fibronectin, fibrinogen, fibrin, albumin, collagen, laminin) on the surface of the device forming conditioning film leads to the alteration of surface properties of the biomaterial and the increase of microbial adherence

The cell surface of a bacterium possesses many structures and properties that contribute to bacterial adhesion including fimbriae (pili), the cell wall (teichoic acid in gram-positive bacteria) and outer cell membrane (lipopolysaccharides in gram negative pathogens). These characteristics influence the surface charge and hydrophobicity of the bacterial cell, thereby directly affecting adherence (Bonner *et al.,* 1997). The physico-chemical characters of microbial cell surface, i.e., hydrophobicity and charge will influence adherence to biomaterial surfaces since the process is strongly governed by hydrophobic and electrostatic

The absorption of components from the suspending fluid can affect the adhesive properties of microorganisms. The ionic strength, osmolarity, and pH all influence the initial attachment of bacteria (Denstedt *et al*., 1998). In the process of adherence of microorganisms

interactions and decreased adherence (Jansen *et al*., 1988).

and in protecting cells from dehydration and the effects of antimicrobial agents (Davis and Geesey, 1995).

## **2.3.3 Maturation of biofilm formation**

Once having irreversibly attached to a surface, bacterial cells undergo phenotypic changes, and the process of biofilm maturation begins. Bacteria start to form microcolonies either by aggregation of already attached cells, clonal growth (cell division) or cell recruitment of planktonic cells or cell flocs from the bulk liquid. The attached cells generate a large amount of extracellular components which interact with organic and inorganic molecules in the immediate environment to create the glycocalyx (Jiang and Pace, 2006).

Mature biofilms consist of differentiated mushroom and pillar like structures of cells embedded in copious amounts of extracellular polymer matrix or glycocalyx, which are separated by water-filled channels and voids to allow convective flows that transport nutrients and oxygen from the interface to the interior parts of the biofilm and remove metabolic wastes (Stoodly *et al*., 2002).

There are many environment within a biofilm, each varying because of difference in local conditions such as nutrient availability, PH, oxidizing potential (redox) and so on. Cells near the surface of the biofilm are exposed to high concentrations of oxygen, while near the center oxygen is rapidly depleted to near anaerobic levels (Lewandowski, 1994). The steep oxygen gradients are paralleled by gradients for other nutrients or metabolites from the biofilm (de Beer *et al*., 1994). Apparently, biofilms display both structural and metabolic heterogenecity which provide this community the capability to resist stresses, whether from host defense systems or antimicrobial agents (Kumar and Anand, 1998).

## **2.3.4 Detachment**

At some point the biofilm reaches a critical mass and the the outermost layer begins to generate planktonic organisms that may escap from the biofilm and colonize other surfaces. Dispersion of planktonic cells can be facilitated by digestion of glycocalyx by enzymes and quorum-sensing might be required for this phenomenon (Dagostino et al., 1991).

## **2.4 Factors affect the adherence of microorganisms to a device surface**

### **2.4.1 Device related factors**

Certain materials used in the design of Indwelling medical devices (IMDs) are more conductive to microbial adherence/biofilm formation than others. In vitro studies performed by many laboratories have determined that microbial adherence to biomaterials occurs in the following order: latex> silicone > PVC > Teflon > Polyurethane > stainless steel > titanium (Darouiche, 2001).

Surface characteristics determining the adherence properties of specific materials include: (a) surface texture, (b) surface charge, and (c) hydrophobicity.

a. Surface texture:

Materials with irregular or rough surfaces tend to have enhanced microbial adherence compared to smooth surfaces. It is documented that surface irregularities in central venous catheters (CVC) varied with different polymer materials so that bacteria preferentially adhered to surface defects within minutes after infusing the catheters with contaminated buffer solution (Locci *et al.,* 1981). Another study examined the surface of five commercially available polyurethane CVCs by scanning electron microscopy and found that the catheters with the most surface irregularities had significantly more adherent bacteria compared to catheters with smoother surfaces (Tebbs *et al*., 1994).

b. Surface charge:

594 Scanning Electron Microscopy

and in protecting cells from dehydration and the effects of antimicrobial agents (Davis and

Once having irreversibly attached to a surface, bacterial cells undergo phenotypic changes, and the process of biofilm maturation begins. Bacteria start to form microcolonies either by aggregation of already attached cells, clonal growth (cell division) or cell recruitment of planktonic cells or cell flocs from the bulk liquid. The attached cells generate a large amount of extracellular components which interact with organic and inorganic molecules in the

Mature biofilms consist of differentiated mushroom and pillar like structures of cells embedded in copious amounts of extracellular polymer matrix or glycocalyx, which are separated by water-filled channels and voids to allow convective flows that transport nutrients and oxygen from the interface to the interior parts of the biofilm and remove

There are many environment within a biofilm, each varying because of difference in local conditions such as nutrient availability, PH, oxidizing potential (redox) and so on. Cells near the surface of the biofilm are exposed to high concentrations of oxygen, while near the center oxygen is rapidly depleted to near anaerobic levels (Lewandowski, 1994). The steep oxygen gradients are paralleled by gradients for other nutrients or metabolites from the biofilm (de Beer *et al*., 1994). Apparently, biofilms display both structural and metabolic heterogenecity which provide this community the capability to resist stresses, whether from

At some point the biofilm reaches a critical mass and the the outermost layer begins to generate planktonic organisms that may escap from the biofilm and colonize other surfaces. Dispersion of planktonic cells can be facilitated by digestion of glycocalyx by enzymes and

Certain materials used in the design of Indwelling medical devices (IMDs) are more conductive to microbial adherence/biofilm formation than others. In vitro studies performed by many laboratories have determined that microbial adherence to biomaterials occurs in the following order: latex> silicone > PVC > Teflon > Polyurethane > stainless steel

Surface characteristics determining the adherence properties of specific materials include:

Materials with irregular or rough surfaces tend to have enhanced microbial adherence compared to smooth surfaces. It is documented that surface irregularities in central

quorum-sensing might be required for this phenomenon (Dagostino et al., 1991).

**2.4 Factors affect the adherence of microorganisms to a device surface** 

(a) surface texture, (b) surface charge, and (c) hydrophobicity.

immediate environment to create the glycocalyx (Jiang and Pace, 2006).

host defense systems or antimicrobial agents (Kumar and Anand, 1998).

Geesey, 1995).

**2.3.3 Maturation of biofilm formation** 

metabolic wastes (Stoodly *et al*., 2002).

**2.3.4 Detachment** 

**2.4.1 Device related factors** 

> titanium (Darouiche, 2001).

a. Surface texture:

Biomaterial surface charge greatly influences adherence of microorganisms. Most microorganisms exhibit a negative surface charge in an aqueous environment. Therefore, a negatively charged biomaterial surface should lead to decreased adherence of microorganisms due to a repulsion effect between both negatively charged surfaces (Jansen *et al*., 1988).

c. Hydrophobicity:

Bacterial cells, which tend to have hydrophobic cell surfaces, are attracted to the hydrophobic surfaces of many biomaterials currently used in IMDs (Schierholz and Beuth, 2001). This hydrophobic interaction between the microorganisms and the biomaterial leads to increased adherence and subsequent biofilm formation. An increase in the surface hydrophilicity of the polymers leads to weakened hydrophobic interactions and decreased adherence (Jansen *et al*., 1988).

### **2.4.2 Host factors**

The biomaterials used in IMDs result in the activation of the host immune response leading to local tissue damage and the development of an immuno-incompetent, fibro-inflammatory zone that increases the susceptibility of the IMD to infection (Schierholz and Beuth, 2001). The deposition of proteinaceous layer (including fibronectin, fibrinogen, fibrin, albumin, collagen, laminin) on the surface of the device forming conditioning film leads to the alteration of surface properties of the biomaterial and the increase of microbial adherence (Pascual, 2002).

## **2.4.3 Microbial factors**

The cell surface of a bacterium possesses many structures and properties that contribute to bacterial adhesion including fimbriae (pili), the cell wall (teichoic acid in gram-positive bacteria) and outer cell membrane (lipopolysaccharides in gram negative pathogens). These characteristics influence the surface charge and hydrophobicity of the bacterial cell, thereby directly affecting adherence (Bonner *et al.,* 1997). The physico-chemical characters of microbial cell surface, i.e., hydrophobicity and charge will influence adherence to biomaterial surfaces since the process is strongly governed by hydrophobic and electrostatic interactions (Martinez-Martinez *et al*., 1991).

#### **2.4.4 The suspending medium**

The absorption of components from the suspending fluid can affect the adhesive properties of microorganisms. The ionic strength, osmolarity, and pH all influence the initial attachment of bacteria (Denstedt *et al*., 1998). In the process of adherence of microorganisms

Application of Scanning Electron Microscopy for

*mechanisms* (Brown *et al*., 1990):

*al*., 1993).

1987).

2000).

Resistance may be due to:

the Morphological Study of Biofilm in Medical Devices 597

stent surfaces in-vitro and in-vivo (Caldwell, 1995). In addition, resistance to antimicrobial agents and other chemicals is one of the greatest problems in the age of widely used medical devices. The problem in conventional clinical microbiology is how to treat patients in the best way when choosing antibiotics is based on bacterial cultures derived from planktonic bacterial cells which differ very from bacteria in the biofilm mode. This can stand behind the clinical failure rate of treating chronic bacterial infection (Choong and Whitfield, 2000).

*The failure of antimicrobial agents to treat biofilms has been associated with a variety of* 

1. The glycocalyx restricts access and diffusion of antimicrobial agents to the deeper lying bacteria (extrinsic resistance). In situ studies have shown that the surface film influences the transport of nutrients and interferes with the transport of antimicrobials (Nivens *et* 

2. The growth rates of bacteria within a biofilm vary widely. Slow-growing bacteria are particularly resistant to antimicrobial agents (Brown, 1997). The limitation of diffusion of nutrients in a biofilm results in spatial gradients of growth rate leading to a plethora of phenotypes within the biofilm. In general, the faster-growing, more susceptible bacteria lie superficially but the slow-growing, less susceptible bacteria being placed more deeply. The failure of antimicrobial agents to eradicate these slow-growing bacteria may exert selection pressures on the least susceptible genotype to select for a resistant population. Furthermore, antimicrobial binding proteins are poorly expressed in the slow-growing bacteria, rendering the antimicrobial agents ineffective (Cozens *et al*., 1986). Commonly, the entire biofilm is coated with a complex of a hydrophilic polymer, the glycocalyx that is typically anionic in nature where the antimicrobial agents reacts chemically with exopolymer or is adsorbed to it, then the net effect is that of having the appearance of a penetration barrier. There will be a similar effect if antimicrobials adsorb onto cells,

3. Bacteria within a biofilm are phenotypically so different from their planktonic counterparts that antimicrobial agents developed against the latter often fail to eradicate organisms in the biofilms. Bacteria within a biofilm activate many genes which alter the cell envelope and molecular targets, and alter the susceptibility to antimicrobial agents (intrinsic resistance). Current opinion is that phenotypic changes brought on by a genetic switch, when 65-80 proteins change, play a much more important role in the protection from antimicrobial agents than the external resistance

4. Bacteria within a biofilm can sense the external environment, communicate with each other and transfer genetic information and plasmids within biofilms (Trieu-Cuot *et al*.,

5. Bacteria in a biofilm can usually survive the presence of antimicrobial agents at concentrations 1000-1500 times higher than the concentrations that kill planktonic cells


perhaps dead ones, in the outer parts of the biofilm (Sutherland, 2001).

provided by the exopolysaccharide slime (Anonymous, 1999).

of the same species (Costerton, 1999).

to an implanted device, one or both entities will be exposed to a biological secretion or body fluid of host origin. The subsequent conditioning of microbial cell or/and biomaterial surface will modify the nature of both surfaces, thereby determining the outcome of the adherence process. It is observed that prior colonization of endotracheal tubes, microorganisms preferentially adhere to a biological film of human origin rather than to the constituent biomaterial itself (Poisson *et al*., 1991). Adherence of *E. coli* and *E. faecalis*, grown in Mueller- Hinton broth, was shown to increase after the biomaterial was exposed to human urine (Bonner *et al*., 1997).

## **2.4.5 Bacteria- biomaterial interaction**

The adhesion of microorganisms to biomaterial surfaces has been shown to require both non-specific reversible interactions and highly specific irreversible interactions. First, reversible adhesion of microorganisms to biomaterial surface is dependent upon the physical characteristics of the microorganisms, biomaterial and the surrounding environment (Gristina, 1987). Microorganisms randomly reach the surface of the biomaterial by several mechanisms: direct contamination, contiguous spread, hematogenous spread. Once near the surface, initial adherence of the microorganism depends upon microorganism-biomaterial interactions including van der waals forces and hydrophobic interactions (pascual, 2002). The common charges of the microorganisms and the IMD surfaces will repel each other, however the effect of van der waals forces overcome this repulsion beginning about 10 nm from the IMD surface keeping the microorganisms near the biomaterial surface (Gristina, 1987).

It has shown that hydrophobic forces are 10 to 100 times stronger than van der waals forces at 10 nm from the biomaterial surface. The hydrophobic forces easily overcome electrostatic repulsion and position the organisms 1-2 nm from IMD surface then allows irreversible adhesion to occur (Pashley *et al*., 1985). Second, irreversible adhesion occurs with the binding of specific microorganism adhesins to receptors expressed by the conditioning film . i.e., *S. aureus* and *S. epidermidis* which are the most common microorganisms causing IMDrelated infections relies on specific cell surface proteins called "microbial surface component recognizing adhesive matrix molecules" (MSCRAMM) which bind to specific host ligands that are found in the conditioning films. The most important MSCRAMMS are the fibronectin-binding proteins (FnBPs), the fibrinogen-binding proteins (clumping factors, CIf) and the collagen (Darouiche *et al*., 1997).

Cell surface proteins also play an important role in *S. epidermidis* adhesion to IMDs. Proteinaceaous autolysin and polysaccharide adhesin (PSA) are two surface proteins that play an early role in the irreversible adhesion of *S. epidermidis* to IMD surfaces. Once adherent to the biomaterial surface, cell accumulation and early biofilm formation are dependent upon the polysaccharide intercellular adhesin (PIA), which promotes intercellular adhesion (Rupp *et al*., 1999).

#### **2.5 Defense mechanisms**

The use of antibiotics is currently one of the possibilities for the prevention of biofilm formation. However, even in the presence of antibiotics bacteria can adhere, colonize and survive on implanted medical devices as has been shown for urinary catheters and ureteral

to an implanted device, one or both entities will be exposed to a biological secretion or body fluid of host origin. The subsequent conditioning of microbial cell or/and biomaterial surface will modify the nature of both surfaces, thereby determining the outcome of the adherence process. It is observed that prior colonization of endotracheal tubes, microorganisms preferentially adhere to a biological film of human origin rather than to the constituent biomaterial itself (Poisson *et al*., 1991). Adherence of *E. coli* and *E. faecalis*, grown in Mueller- Hinton broth, was shown to increase after the biomaterial was exposed to

The adhesion of microorganisms to biomaterial surfaces has been shown to require both non-specific reversible interactions and highly specific irreversible interactions. First, reversible adhesion of microorganisms to biomaterial surface is dependent upon the physical characteristics of the microorganisms, biomaterial and the surrounding environment (Gristina, 1987). Microorganisms randomly reach the surface of the biomaterial by several mechanisms: direct contamination, contiguous spread, hematogenous spread. Once near the surface, initial adherence of the microorganism depends upon microorganism-biomaterial interactions including van der waals forces and hydrophobic interactions (pascual, 2002). The common charges of the microorganisms and the IMD surfaces will repel each other, however the effect of van der waals forces overcome this repulsion beginning about 10 nm from the IMD surface keeping the microorganisms near

It has shown that hydrophobic forces are 10 to 100 times stronger than van der waals forces at 10 nm from the biomaterial surface. The hydrophobic forces easily overcome electrostatic repulsion and position the organisms 1-2 nm from IMD surface then allows irreversible adhesion to occur (Pashley *et al*., 1985). Second, irreversible adhesion occurs with the binding of specific microorganism adhesins to receptors expressed by the conditioning film . i.e., *S. aureus* and *S. epidermidis* which are the most common microorganisms causing IMDrelated infections relies on specific cell surface proteins called "microbial surface component recognizing adhesive matrix molecules" (MSCRAMM) which bind to specific host ligands that are found in the conditioning films. The most important MSCRAMMS are the fibronectin-binding proteins (FnBPs), the fibrinogen-binding proteins (clumping factors, CIf)

Cell surface proteins also play an important role in *S. epidermidis* adhesion to IMDs. Proteinaceaous autolysin and polysaccharide adhesin (PSA) are two surface proteins that play an early role in the irreversible adhesion of *S. epidermidis* to IMD surfaces. Once adherent to the biomaterial surface, cell accumulation and early biofilm formation are dependent upon the polysaccharide intercellular adhesin (PIA), which promotes

The use of antibiotics is currently one of the possibilities for the prevention of biofilm formation. However, even in the presence of antibiotics bacteria can adhere, colonize and survive on implanted medical devices as has been shown for urinary catheters and ureteral

human urine (Bonner *et al*., 1997).

**2.4.5 Bacteria- biomaterial interaction** 

the biomaterial surface (Gristina, 1987).

and the collagen (Darouiche *et al*., 1997).

intercellular adhesion (Rupp *et al*., 1999).

**2.5 Defense mechanisms** 

stent surfaces in-vitro and in-vivo (Caldwell, 1995). In addition, resistance to antimicrobial agents and other chemicals is one of the greatest problems in the age of widely used medical devices. The problem in conventional clinical microbiology is how to treat patients in the best way when choosing antibiotics is based on bacterial cultures derived from planktonic bacterial cells which differ very from bacteria in the biofilm mode. This can stand behind the clinical failure rate of treating chronic bacterial infection (Choong and Whitfield, 2000).

*The failure of antimicrobial agents to treat biofilms has been associated with a variety of mechanisms* (Brown *et al*., 1990):


Resistance may be due to:


Application of Scanning Electron Microscopy for

target (Labthavikul *et al*., 2003). **b. Non-antimicrobial agents:** 

to contact lenses (Arciola *et al*., 1998).

Giraldo *et al*., 1997).

(Olofsson *et al*., 2003).

the Morphological Study of Biofilm in Medical Devices 599

It is investigated that the antibiofilm effects by incubating ciprofloxacin with *P. aeruginosa* or in combination with macrolides. At twice the minimum bactericidal concentration of ciprofloxacin, 85% of the population of *P. aeruginosa* within the biofilm survived. In contrast, the killing effect of ciprofloxacin was greatly enhanced when combined with clarithromycin, erythromycin and azithromycin, but not with the 16-membered ring macrolides. It is speculated that the 14-membered and 15- membered ring macrolides posses an ability to increase the permeability of biofilms, there by facilitating the penetration of quinolone antibiotics. Tigecycline was observed to inhibit the growth of *S. epidermidis* which indicates that tigecycline is able to diffuse through the biofilm and act normally against its cellular

It is observed that some drugs other than antimicrobial agents such as anti-inflammatory or antiseptic compounds reduce the adherence of bacteria. Coating the catheter with acetylsalicylic acid or sodium salicylate reduces or inhibits microbial adherence, Bandazac lysine (non steroidal anti-inflammatory) was found to prevented the adherence of bacteria

Some mucolytics substances such as EDTA, sodium citrate and penicillamine may disperse the biofilms formed by *P*. *aeruginosa* (Gordon *et al*., 1991). It is observed also that Nacetylcysteine (NAC) (a non antibiotic drug that has antibacterial properties (bacteriostatic) and a mucolytic agent that disrupts disulphide bonds in mucus and reduces the viscosity of secretions) decreases biofilm formation and therefore may be an effective alternative for preventing infections by *S. epidermidis* and other coagulase negative staphylococci (Pérez-

It is observed that NAC not only reduced the adhesion but in fact also detached adhered cells from a steel surface. This has some importance since the initial adhesion often develops into a stronger interaction with time (bond ageing) (Meinders *et al*., 1995). The reduction in the amount of exopolysaccharides (EPS) in the presence of NAC may have many explanations. The direct effects of NAC include a possible reaction of its sulfhydryl group with disulfide bonds of enzymes involved in EPS production or excretion, which renders these molecules less active, or competitive inhibition of cysteine utilization. Also, the possibility of interference of NAC with control or signaling systems that direct the EPS production at translation or at the enzymatic level cannot be excluded. The fact that NAC is an anti-oxidant may have indirect effects on cell metabolism and EPS production. NAC increases the wettability of surfaces. Moreover, NAC detached bacteria that were adhering to steel surfaces. Growth of various bacteria, as monocultures or in multi-species community, was inhibited at different concentrations of NAC. It is also found that there was no detectable degradation of EPS by NAC, indicating that NAC reduced the production of EPS in most bacteria tested, even at concentrations at which growth was not affected

Aspirin (acetylsalicylic acid) has a short half life in circulating blood (about 20 min) and is rapidly deacetylated to form salicylic acid in-vivo. Sodium salicylate and related compounds such as aspirin are known to have a variety of effects on microorganisms. Growth of certain bacteria in the presence of salicylate can induce multiple resistance to


## **2.6 Treatment and prevention of biofilms**

Strategies for prevention of these infections include: (i) minimizing tissue destruction and removal of all extraneous biomaterials and devitalized tissues during surgery. (ii) development of biomaterials that resist the initial adherence of bacteria by surface characteristic or by promoting bactericidal, bacteriostatic or phagocytic activity at their surfaces. (iii) further study of the microstructure and chemical nature of the adherence mechanism and development of analogs and enzymes that might block the initial adherence by modification of receptors and ligands (Khardori and Yassien, 1995) .

Several approaches have been studied to prevent the formation of biofilms and to eradicate biofilms associated bacteria. Some of that depends on the use of antimicrobial agents or non antimicrobial agents.

#### **a. Antimicrobial agents:**

In the case of the use of antimicrobial agents, it was found that some antibiotic at sub-MIC inhibit the initial adherence. Dicloxacillin is the antibiotic that found to prevent the adherence to the greatest extent when it is used alone at 1/2 of the MIC (Cerca *et al*., 2005). Also clindamycin at subinhibitory concentrations inhibits the adherence of *Pseudomonas aeruginosa*, *Staphylococcus aureus*, Bacteroids spp., *Escherichia coli* to bone surfaces (Lambe *et al*., 1987).

Norfloxacin, ciprofloxacin, ofloxacin and azithromycin at sub inhibitory concentrations reduced the glycocalyx production and inhibited the adherence of *Staphylococcus epidermidis* and *Pseudomonas aeruginosa* (Pézer-Giraldo *et al*., 1989 ; Yassien *et al*., 1995). Ciprofloxacin was reported to eradicate the performed biofilms of *P. aeruginosa* (Reid *et al*., 1994). It was reported also that 1/2 MIC of ciprofloxacin, 1/4 MIC of ofloxacin and 1/32 of levofloxacin caused significant inhibition of adherence of some uropathogenic strains of *E. coli* to periuretheral epithelial cells (Baskin *et al*., 2002).

Macrolides are generally bacteriostatic in-vitro and in-vivo, and have useful activity versus gram-positive bacteria. Macrolides have been evaluated to affect the adherence of gramnegative bacteria at sub-MIC concentrations by 50 to 70% as it is found to affect the production of virulence determinants such as secreted virulence factor, motility, quorum sensing and biofilm production (Vranes, 2000). It is discovered that sub-MIC level of clarithromycin inhibits the twitching motility of *P. aeruginosa*, they do not affect the production of pili but inhibit their assembly on the surface of bacteria that should affect some steps in biofilm formation (Wozniak and keyser, 2004).

It is investigated that the antibiofilm effects by incubating ciprofloxacin with *P. aeruginosa* or in combination with macrolides. At twice the minimum bactericidal concentration of ciprofloxacin, 85% of the population of *P. aeruginosa* within the biofilm survived. In contrast, the killing effect of ciprofloxacin was greatly enhanced when combined with clarithromycin, erythromycin and azithromycin, but not with the 16-membered ring macrolides. It is speculated that the 14-membered and 15- membered ring macrolides posses an ability to increase the permeability of biofilms, there by facilitating the penetration of quinolone antibiotics. Tigecycline was observed to inhibit the growth of *S. epidermidis* which indicates that tigecycline is able to diffuse through the biofilm and act normally against its cellular target (Labthavikul *et al*., 2003).

## **b. Non-antimicrobial agents:**

598 Scanning Electron Microscopy



Strategies for prevention of these infections include: (i) minimizing tissue destruction and removal of all extraneous biomaterials and devitalized tissues during surgery. (ii) development of biomaterials that resist the initial adherence of bacteria by surface characteristic or by promoting bactericidal, bacteriostatic or phagocytic activity at their surfaces. (iii) further study of the microstructure and chemical nature of the adherence mechanism and development of analogs and enzymes that might block the initial adherence

Several approaches have been studied to prevent the formation of biofilms and to eradicate biofilms associated bacteria. Some of that depends on the use of antimicrobial agents or non

In the case of the use of antimicrobial agents, it was found that some antibiotic at sub-MIC inhibit the initial adherence. Dicloxacillin is the antibiotic that found to prevent the adherence to the greatest extent when it is used alone at 1/2 of the MIC (Cerca *et al*., 2005). Also clindamycin at subinhibitory concentrations inhibits the adherence of *Pseudomonas aeruginosa*, *Staphylococcus aureus*, Bacteroids spp., *Escherichia coli* to bone surfaces (Lambe *et al*., 1987).

Norfloxacin, ciprofloxacin, ofloxacin and azithromycin at sub inhibitory concentrations reduced the glycocalyx production and inhibited the adherence of *Staphylococcus epidermidis* and *Pseudomonas aeruginosa* (Pézer-Giraldo *et al*., 1989 ; Yassien *et al*., 1995). Ciprofloxacin was reported to eradicate the performed biofilms of *P. aeruginosa* (Reid *et al*., 1994). It was reported also that 1/2 MIC of ciprofloxacin, 1/4 MIC of ofloxacin and 1/32 of levofloxacin caused significant inhibition of adherence of some uropathogenic strains of *E. coli* to

Macrolides are generally bacteriostatic in-vitro and in-vivo, and have useful activity versus gram-positive bacteria. Macrolides have been evaluated to affect the adherence of gramnegative bacteria at sub-MIC concentrations by 50 to 70% as it is found to affect the production of virulence determinants such as secreted virulence factor, motility, quorum sensing and biofilm production (Vranes, 2000). It is discovered that sub-MIC level of clarithromycin inhibits the twitching motility of *P. aeruginosa*, they do not affect the production of pili but inhibit their assembly on the surface of bacteria that should affect

by modification of receptors and ligands (Khardori and Yassien, 1995) .

(De Kievit *et al*., 2001).

**2.6 Treatment and prevention of biofilms** 

periuretheral epithelial cells (Baskin *et al*., 2002).

some steps in biofilm formation (Wozniak and keyser, 2004).

2004).

antimicrobial agents.

**a. Antimicrobial agents:** 

It is observed that some drugs other than antimicrobial agents such as anti-inflammatory or antiseptic compounds reduce the adherence of bacteria. Coating the catheter with acetylsalicylic acid or sodium salicylate reduces or inhibits microbial adherence, Bandazac lysine (non steroidal anti-inflammatory) was found to prevented the adherence of bacteria to contact lenses (Arciola *et al*., 1998).

Some mucolytics substances such as EDTA, sodium citrate and penicillamine may disperse the biofilms formed by *P*. *aeruginosa* (Gordon *et al*., 1991). It is observed also that Nacetylcysteine (NAC) (a non antibiotic drug that has antibacterial properties (bacteriostatic) and a mucolytic agent that disrupts disulphide bonds in mucus and reduces the viscosity of secretions) decreases biofilm formation and therefore may be an effective alternative for preventing infections by *S. epidermidis* and other coagulase negative staphylococci (Pérez-Giraldo *et al*., 1997).

It is observed that NAC not only reduced the adhesion but in fact also detached adhered cells from a steel surface. This has some importance since the initial adhesion often develops into a stronger interaction with time (bond ageing) (Meinders *et al*., 1995). The reduction in the amount of exopolysaccharides (EPS) in the presence of NAC may have many explanations. The direct effects of NAC include a possible reaction of its sulfhydryl group with disulfide bonds of enzymes involved in EPS production or excretion, which renders these molecules less active, or competitive inhibition of cysteine utilization. Also, the possibility of interference of NAC with control or signaling systems that direct the EPS production at translation or at the enzymatic level cannot be excluded. The fact that NAC is an anti-oxidant may have indirect effects on cell metabolism and EPS production. NAC increases the wettability of surfaces. Moreover, NAC detached bacteria that were adhering to steel surfaces. Growth of various bacteria, as monocultures or in multi-species community, was inhibited at different concentrations of NAC. It is also found that there was no detectable degradation of EPS by NAC, indicating that NAC reduced the production of EPS in most bacteria tested, even at concentrations at which growth was not affected (Olofsson *et al*., 2003).

Aspirin (acetylsalicylic acid) has a short half life in circulating blood (about 20 min) and is rapidly deacetylated to form salicylic acid in-vivo. Sodium salicylate and related compounds such as aspirin are known to have a variety of effects on microorganisms. Growth of certain bacteria in the presence of salicylate can induce multiple resistance to

Application of Scanning Electron Microscopy for

(control) (× 3500).

the Morphological Study of Biofilm in Medical Devices 601

a wire loop on trypticase soy agar with 5% sheep blood and MacConkey agar. Organisms

Antibiotic susceptibility and MICs were determined for the isolated microorganisms by the agar dilution method, according to clinical laboratory standards institute (CLSI) (2007)

The isolated microorganisms were tested for their ability to form biofilm by tissue culture plate method (TCP). Effect of different concentrations of ciprofloxacin, N-acetylcysteine each alone and in combination on the bacterial adherence to plastic surfaces were determined by tissue culture plate assay (Christensen *et al,* 1985). The Effect of different concentrations of ciprofloxacin, N-acetylcysteine each alone and in combination on the bacterial adherence to the surface of ureteral catheter were determined by Static adhesion assay (Reid *et al.,* 1994)

Fig. 1. Scanning electron micrograph of an empty ureteral stents incubated in saline for 24 h

Fig. 2. Scanning electron micrograph showed the lumen of the ureteral stent (× 35) blocked

with a dense mass of biofilm containing bacteria.

were then identified by routine microbiological techniques (Sherertz *et al*., 1990).

and their effects were determined also using scanning electron microscope.

antibiotic. Paradoxically, it can also reduce resistance to some antibiotics (Price *et al*., 2000). *E. coli*, for example exhibits increased resistance to chloramphenicol, ampicillin, naldixic acid and tetracycline after such treatment. On the other hand *E. coli* cells grown in the presence of salicylate are more sensitive to aminoglycosides (Aumercier *et al*., 1990).

Sodium salicylate inhibits biofilm formation by *P. aeruginosa* and *S. epidermidis* on contact lenses and medical polymers such as polyethylene and polystyrene. It also decreases bacterial adhesion in a dose-dependent manner. Some strains of *S. epidermidis* secrete mucoid extracellular polymers (polysaccharides, proteins and teichoic acid) that promote biofilm formation and become important components of the biofilm matrix. Salicylate can inhibit the production of some of these components by as much as 95%. It has been suggested that the use of salicylate into contact lens solutions might decrease the incidence of some device- related infections (Farber and Wolff, 1992).

Chlorohexidine gluconate and silver sulfadiazine coated vascular catheter has been shown to be highly effective in decreasing catheter related infections (George *et al*., 1997).

Protamine sulfate (a surface active, basic polypeptide presently used to reverse the anticoagulant effects of heparin) could aid antibiotics in penetrating a *P. aeruginosa* biofilm (Richards, 1976). Parsons and coworkers have shown that protamine sulfate penetrates and disrupts the protective glycosaminoglycan layer. There is a significant, synergistic effect observed between protamine sulfate and ciprofloxacin as protamine sulfate may have denatured the complex extracellular polymeric structure of the *P. aeruginosa* biofilm enhancing penetration of ciprofloxacin through the biofilm (Soboh *et al*., 1995).

Gendine solution (a novel antiseptic solution) formed of gention violet and chlorohexidine has the ability to coat various polymers and devices. It has also a broad spectrum antiadherence activity and antimicrobial activity which decreases the risk of device colonization, which may in turn decreases the rates of nosocomial infection and their associated morbidity and mortality (Chaiban *et al*., 2005).

## **3. Techniques for the study of biofilm**

This work was done to detect biofilm formed on ureteral stents and to determine whether Nacetylcysteine could aid ciprofloxacin in penetrating biofilm formed by some microorganisms on ureteral stents. Several techniques were used in this study first Stents were removed by physicians and collected in sterile screw capped tubes, then cut into segments to be examined by Scanning electron microscope (SEM) and to be cultured on different media.

Catheter segment were fixed in 2.5% (vol/vol) glutaraldehyde in Dulbecco PBS (pH 7.2) for 1.5h, rinsed with Phosphate buffer saline (PBS), and then dehydrated through an ethanol series. Samples were critical point dried and gold-palladium coated. SEM examinations were made on a JSM-840 SEM (JEOL Ltd., Tokyo, Japan).

Urine samples were collected and streaked onto the culture media and incubated at 37°C for 24 hours (Benson, 2002). The resultant colonies were streaked and examined morphologically, microscopically and biochemically.

Catheter samples: Each catheter was placed in 10 ml of tryptic soy broth (TSB), sonicated for 1 min and then vortexed for 15 s. 0.1 ml of the sonicated broth were surface plated by using

antibiotic. Paradoxically, it can also reduce resistance to some antibiotics (Price *et al*., 2000). *E. coli*, for example exhibits increased resistance to chloramphenicol, ampicillin, naldixic acid and tetracycline after such treatment. On the other hand *E. coli* cells grown in the

Sodium salicylate inhibits biofilm formation by *P. aeruginosa* and *S. epidermidis* on contact lenses and medical polymers such as polyethylene and polystyrene. It also decreases bacterial adhesion in a dose-dependent manner. Some strains of *S. epidermidis* secrete mucoid extracellular polymers (polysaccharides, proteins and teichoic acid) that promote biofilm formation and become important components of the biofilm matrix. Salicylate can inhibit the production of some of these components by as much as 95%. It has been suggested that the use of salicylate into contact lens solutions might decrease the incidence

Chlorohexidine gluconate and silver sulfadiazine coated vascular catheter has been shown

Protamine sulfate (a surface active, basic polypeptide presently used to reverse the anticoagulant effects of heparin) could aid antibiotics in penetrating a *P. aeruginosa* biofilm (Richards, 1976). Parsons and coworkers have shown that protamine sulfate penetrates and disrupts the protective glycosaminoglycan layer. There is a significant, synergistic effect observed between protamine sulfate and ciprofloxacin as protamine sulfate may have denatured the complex extracellular polymeric structure of the *P. aeruginosa* biofilm

Gendine solution (a novel antiseptic solution) formed of gention violet and chlorohexidine has the ability to coat various polymers and devices. It has also a broad spectrum antiadherence activity and antimicrobial activity which decreases the risk of device colonization, which may in turn decreases the rates of nosocomial infection and their

This work was done to detect biofilm formed on ureteral stents and to determine whether Nacetylcysteine could aid ciprofloxacin in penetrating biofilm formed by some microorganisms on ureteral stents. Several techniques were used in this study first Stents were removed by physicians and collected in sterile screw capped tubes, then cut into segments to be examined

Catheter segment were fixed in 2.5% (vol/vol) glutaraldehyde in Dulbecco PBS (pH 7.2) for 1.5h, rinsed with Phosphate buffer saline (PBS), and then dehydrated through an ethanol series. Samples were critical point dried and gold-palladium coated. SEM examinations

Urine samples were collected and streaked onto the culture media and incubated at 37°C for 24 hours (Benson, 2002). The resultant colonies were streaked and examined

Catheter samples: Each catheter was placed in 10 ml of tryptic soy broth (TSB), sonicated for 1 min and then vortexed for 15 s. 0.1 ml of the sonicated broth were surface plated by using

to be highly effective in decreasing catheter related infections (George *et al*., 1997).

enhancing penetration of ciprofloxacin through the biofilm (Soboh *et al*., 1995).

by Scanning electron microscope (SEM) and to be cultured on different media.

presence of salicylate are more sensitive to aminoglycosides (Aumercier *et al*., 1990).

of some device- related infections (Farber and Wolff, 1992).

associated morbidity and mortality (Chaiban *et al*., 2005).

were made on a JSM-840 SEM (JEOL Ltd., Tokyo, Japan).

morphologically, microscopically and biochemically.

**3. Techniques for the study of biofilm** 

a wire loop on trypticase soy agar with 5% sheep blood and MacConkey agar. Organisms were then identified by routine microbiological techniques (Sherertz *et al*., 1990).

Antibiotic susceptibility and MICs were determined for the isolated microorganisms by the agar dilution method, according to clinical laboratory standards institute (CLSI) (2007)

The isolated microorganisms were tested for their ability to form biofilm by tissue culture plate method (TCP). Effect of different concentrations of ciprofloxacin, N-acetylcysteine each alone and in combination on the bacterial adherence to plastic surfaces were determined by tissue culture plate assay (Christensen *et al,* 1985). The Effect of different concentrations of ciprofloxacin, N-acetylcysteine each alone and in combination on the bacterial adherence to the surface of ureteral catheter were determined by Static adhesion assay (Reid *et al.,* 1994) and their effects were determined also using scanning electron microscope.

Fig. 1. Scanning electron micrograph of an empty ureteral stents incubated in saline for 24 h (control) (× 3500).

Fig. 2. Scanning electron micrograph showed the lumen of the ureteral stent (× 35) blocked with a dense mass of biofilm containing bacteria.

Application of Scanning Electron Microscopy for

the Morphological Study of Biofilm in Medical Devices 603

Fig. 5. Scanning electron micrograph showed the lumen of the ureteral stent (×5000). It

Fig. 6. Scanning electron micrograph showed the surface of the ureteral stent (× 3500). It showed a dense mass of biofilm containing microorganisms and a high level of encrustation.

ciprofloxacin and ofloxacin was shown by *C. freundii* (78.6% each).

The resistance pattern to cefotaxime, augmentin, ciprofloxacin, levofloxacin and ofloxacin revealed that the highest incidence of resistance to cefotaxime was shown by *K. oxytocae* (54.2%). Also the highest incidence of resistance to augmentin and levofloxacin was shown by Pseudomonas spp. (80 and 72.7%, respectively), while the highest resistance to

Biofilm production was found in 84.6% of the isolates using TCP. Pseudomonas spp. were the highest biofilm producing microorganism. A dose related decrease in biofilm formation was observed by both ciprofloxacin and N-acetylcysteine. This was detected by a decrease in the optical density of the biofilm layer on microtiter plates and the number of viable cells attached to the catheter surfaces in comparison to controls. It was found

showed a dense mass of biofilm (rods and cocci bacteria).

Fig. 3. Scanning electron micrograph showed the lumen of the ureteral stent covered with a densed mass of biofilm containing bacteria (*S. aureus* and *P. rettgeri*) and crystalline patches (× 5000).

In the present work, 292 strains were isolated and identified from 284 samples. As out of 100 urine samples (before catheterization), 76 (76%) were positive for bacterial growth. Out of 92 urine samples (after stent removal), 80 (86.95%) were positive for bacterial growth and out of 92 stent samples, 84 (91.3%) were positive for bacterial growth. Stents collected from patients were examined for biofilm using SEM and it was found that all stents positive for microbial growth were showing biofilm upon their examination.

Fig. 4. Scanning electron micrograph showed the lumen of a ureteral stent obtained from patients treated with cefotaxime (× 35). It showed a dense mass of biofilm and a high level of encrustation.

Klebseilla spp. was the most prevalent (21.9%) microorganism followed by Pseudomonas spp. (18.8%), Staphylococci spp. (18.2%), *E. coli* (17.8%), Proteus spp. (11.3%), *Providencia rettgeri* (4.8%) *Citrobacter freundii* (4.8%) and *Serratia marcescens* (2.8%). Mixed infection represented 22.9%. All *S. aureus* and coagulase negative staphylococci isolates were polymicrobial with Klebseilla spp., Pseudomonas spp., *Providencia rettgeri* and *S. marcescens*.

Fig. 3. Scanning electron micrograph showed the lumen of the ureteral stent covered with a densed mass of biofilm containing bacteria (*S. aureus* and *P. rettgeri*) and crystalline patches

In the present work, 292 strains were isolated and identified from 284 samples. As out of 100 urine samples (before catheterization), 76 (76%) were positive for bacterial growth. Out of 92 urine samples (after stent removal), 80 (86.95%) were positive for bacterial growth and out of 92 stent samples, 84 (91.3%) were positive for bacterial growth. Stents collected from patients were examined for biofilm using SEM and it was found that all stents positive for

Fig. 4. Scanning electron micrograph showed the lumen of a ureteral stent obtained from patients treated with cefotaxime (× 35). It showed a dense mass of biofilm and a high level of

Klebseilla spp. was the most prevalent (21.9%) microorganism followed by Pseudomonas spp. (18.8%), Staphylococci spp. (18.2%), *E. coli* (17.8%), Proteus spp. (11.3%), *Providencia rettgeri* (4.8%) *Citrobacter freundii* (4.8%) and *Serratia marcescens* (2.8%). Mixed infection represented 22.9%. All *S. aureus* and coagulase negative staphylococci isolates were polymicrobial with Klebseilla spp., Pseudomonas spp., *Providencia rettgeri* and *S. marcescens*.

microbial growth were showing biofilm upon their examination.

(× 5000).

encrustation.

Fig. 5. Scanning electron micrograph showed the lumen of the ureteral stent (×5000). It showed a dense mass of biofilm (rods and cocci bacteria).

Fig. 6. Scanning electron micrograph showed the surface of the ureteral stent (× 3500). It showed a dense mass of biofilm containing microorganisms and a high level of encrustation.

The resistance pattern to cefotaxime, augmentin, ciprofloxacin, levofloxacin and ofloxacin revealed that the highest incidence of resistance to cefotaxime was shown by *K. oxytocae* (54.2%). Also the highest incidence of resistance to augmentin and levofloxacin was shown by Pseudomonas spp. (80 and 72.7%, respectively), while the highest resistance to ciprofloxacin and ofloxacin was shown by *C. freundii* (78.6% each).

Biofilm production was found in 84.6% of the isolates using TCP. Pseudomonas spp. were the highest biofilm producing microorganism. A dose related decrease in biofilm formation was observed by both ciprofloxacin and N-acetylcysteine. This was detected by a decrease in the optical density of the biofilm layer on microtiter plates and the number of viable cells attached to the catheter surfaces in comparison to controls. It was found

Application of Scanning Electron Microscopy for

Fig. 9.

the Morphological Study of Biofilm in Medical Devices 605

CIP/NAC

NAC

CIP

control

**Low conc. high concentration**

Fig. 10. a. Scanning electron micrograph of *S. aureus* biofilm on the surface. (a uretral stent

incubated with *S. aureus* suspension for 24h as a control) (× 5000).

also that CIP/NAC combinations have the highest inhibitory effect on the initial adherence (84-100% of the controls) and the highest disruptive effect to mature biofilms (87-100% of the controls).

Fig. 7. Scanning electron micrograph showed the surface of a ureteral stent covered with high densed crystalline biofilm (× 5000).

Fig. 8. Scanning electron micrograph showed the lumen of a ureteral stent covered with a big mass of biofilm containing bacteria (rods and cocci) (*K. pneumoniae* and *S. aureus*) (× 5000).

The inhibitory effects of the tested agents were also verified by (SEM). Scanning electron micrographs showed the morphological response of the tested organisms to ciprofloxacin and N-acetylcysteine. They showed also the decrease in the extent of biofilm formation in the presence of the tested agents.

also that CIP/NAC combinations have the highest inhibitory effect on the initial adherence (84-100% of the controls) and the highest disruptive effect to mature biofilms

Fig. 7. Scanning electron micrograph showed the surface of a ureteral stent covered with

Fig. 8. Scanning electron micrograph showed the lumen of a ureteral stent covered with a big mass of biofilm containing bacteria (rods and cocci) (*K. pneumoniae* and *S. aureus*) (×

The inhibitory effects of the tested agents were also verified by (SEM). Scanning electron micrographs showed the morphological response of the tested organisms to ciprofloxacin and N-acetylcysteine. They showed also the decrease in the extent of biofilm formation in

(87-100% of the controls).

high densed crystalline biofilm (× 5000).

5000).

the presence of the tested agents.

**Low conc. high concentration** Fig. 9.

Fig. 10. a. Scanning electron micrograph of *S. aureus* biofilm on the surface. (a uretral stent incubated with *S. aureus* suspension for 24h as a control) (× 5000).

Application of Scanning Electron Microscopy for

scattered (× 5000).

surface.

the Morphological Study of Biofilm in Medical Devices 607

Fig. 10. d. Scanning electron micrograph showed the effect of ciprofloxacin-N-acetylcysteine

Scanning electron micrographs showed the effect of Ciprofloxacin, N-acetylcysteine each alone and in combination on a performed *S. aureus* biofilm developed *in-vitro* on stent

combination on a performed *S. aureus* biofilm. Cell appeared swollen, disrupted and

Fig. 11. a. Scanning electron micrograph showing the morphological responses of Pseudomonas spp. and *S. epidermidis* grown in the presence of sub-MIC concentration of

ciprofloxacin. Cells appeared swolled and scattered with no biofilm mass.

Fig. 10. b. Scanning electron micrograph showed the morphological response of *S. aureus* performed biofilm on the surface of a uretral stent exposed to sub-MIC concentration (CIP 4 µg/ml). there was a decrease in the amount of biofilm mass adhered to stent surface.

Fig. 10. c. Scanning electron micrograph showed the effect of N-acetylcysteine on a performed *S. aureus* biofilm. Cotton like mass disappeared and cells appeared swollen with disrupted cell wall (× 5000).

Fig. 10. b. Scanning electron micrograph showed the morphological response of *S. aureus* performed biofilm on the surface of a uretral stent exposed to sub-MIC concentration (CIP 4 µg/ml). there was a decrease in the amount of biofilm mass adhered to stent surface.

Fig. 10. c. Scanning electron micrograph showed the effect of N-acetylcysteine on a

disrupted cell wall (× 5000).

performed *S. aureus* biofilm. Cotton like mass disappeared and cells appeared swollen with

Fig. 10. d. Scanning electron micrograph showed the effect of ciprofloxacin-N-acetylcysteine combination on a performed *S. aureus* biofilm. Cell appeared swollen, disrupted and scattered (× 5000).

Scanning electron micrographs showed the effect of Ciprofloxacin, N-acetylcysteine each alone and in combination on a performed *S. aureus* biofilm developed *in-vitro* on stent surface.

Fig. 11. a. Scanning electron micrograph showing the morphological responses of Pseudomonas spp. and *S. epidermidis* grown in the presence of sub-MIC concentration of ciprofloxacin. Cells appeared swolled and scattered with no biofilm mass.

Application of Scanning Electron Microscopy for

disrupted cell wall.

the Morphological Study of Biofilm in Medical Devices 609

Fig 11. D. Scanning electron micrograph showed the effect of CIP/NAC combination (2 MIC/ 8 mg/ml) on the ability of *S. epidermidis* and pseudomonas spp. To form biofilm. A high decrease in the nimber of adherent cells observed. Cells appeared large, swollen and with

Scanning electron micrographs showed the morphological response and the ability of *S. epidermidis* and Pseudomonas spp. grown in the presence of Ciprofloxacin, N-acetylcysteine

Fig. 12. a. Scanning electron micrograph showed the morphological response of *S. aureus*

concentration. Cells appeared swollen, enlarged, with irregular cell wall, some showed v-

and pseudomonas spp. cells grown in the presence of ciprofloxacin at sub-MIC

shaped cells and small amount of biofilm mass observed.

and their combinations to form biofilm on stent surfaces.

Fig 11. b. Scanning electron micrograph showing the effect of N-acetylcysteine on the biofilm formed by *S. epidermidis* and pseudomonas spp.. Cells showed membrane disorganization, appeared swolled and with disrupted outer membrane.

Fig 11. c. Scanning electron micrograph showed the effect of CIP/NAC (MIC/4mg/ml) on the ability of *S. epidermidis* and Pseudomonas spp. to form biofilm. Cells appeared scattered, elongated, swollen, with disorganized (irregular) membrane and with no cotton like mass (biofilm) around cells.

Fig 11. b. Scanning electron micrograph showing the effect of N-acetylcysteine on the biofilm formed by *S. epidermidis* and pseudomonas spp.. Cells showed membrane

Fig 11. c. Scanning electron micrograph showed the effect of CIP/NAC (MIC/4mg/ml) on the ability of *S. epidermidis* and Pseudomonas spp. to form biofilm. Cells appeared scattered, elongated, swollen, with disorganized (irregular) membrane and with no cotton like mass

(biofilm) around cells.

disorganization, appeared swolled and with disrupted outer membrane.

Fig 11. D. Scanning electron micrograph showed the effect of CIP/NAC combination (2 MIC/ 8 mg/ml) on the ability of *S. epidermidis* and pseudomonas spp. To form biofilm. A high decrease in the nimber of adherent cells observed. Cells appeared large, swollen and with disrupted cell wall.

Scanning electron micrographs showed the morphological response and the ability of *S. epidermidis* and Pseudomonas spp. grown in the presence of Ciprofloxacin, N-acetylcysteine and their combinations to form biofilm on stent surfaces.

Fig. 12. a. Scanning electron micrograph showed the morphological response of *S. aureus* and pseudomonas spp. cells grown in the presence of ciprofloxacin at sub-MIC concentration. Cells appeared swollen, enlarged, with irregular cell wall, some showed vshaped cells and small amount of biofilm mass observed.

Application of Scanning Electron Microscopy for

observed on the surface of stent.

**4. Conclusion** 

concentration dependent.

**5. Acknowledgment** 

the extracellular polysaccharide matrix of biofilm.

antimicrobial and anti adherent agents treatments.

the Morphological Study of Biofilm in Medical Devices 611

Fig. 12. d. Scanning electron micrograph showed the effect of CIP/NAC combination of (2 MIC/8 mg/ml) on *S. aureus* and pseudomonas spp. ability to form biofilm. No biofilm

Scanning electron micrographs showed the morphological response and the ability of *S. aureus* and Pseudomonas spp. grown in the presence of Ciprofloxacin (sub-MIC), N-

The presence of non antimicrobial agent such as N-acetylcysteine (NAC), caused significant decrease in biofilm formation by a variety of bacteria and reduces the production of extracellular polysaccharide matrix while promoting the disruption of mature biofilms. It was found that the inhibitory effect of both ciprofloxacin and N-acetylcysteine was

CIP/NAC combinations were found to show the highest effect on bacterial adherence inhibition and on the disruption of the already formed biofilms. As N-acetylcysteine increase the therapeutic activity of ciprofloxacin when used in combination by degrading

In the chapter, Scanning Electron Microscope is used for the evaluation of medical implants, detection of biofilm and studying the effect of different biofilm inhibitory agents. This technique provides excellent visualization of glycocalyx, which is one of the most prominent features of biofilms and a crucial research subject in the searching for alternative

Thanks for my professor doctors: Mohamed Ali Mohamed El-Feky, Mostafa Said Khalil El-Rehewy, Mona Amin Hassan(Department of microbiology), Faculty of medicine, Assuit

acetylcysteine and their combinations to form biofilm on stent surfaces.

Fig. 12. b. Scanning electron micrograph showed the effect of N-acetylcysteine (4 mg/ml) on biofilm formation by *S. aureus* and Pseudomonas spp.. Cells appeared swollen, irregular in shape and small microcolonies observed scattered. A decrease in the numer of adherent cells was observed.

Fig. 12. c. Scanning electron micrograph showed the effect of CIP/NAC combination of (MIC/4mg/ml) on *S. aureus* and pseudomonas spp. ability to form biofilm. Cells appeared elongated, enlarged and scattered with no biofilm mass observed on the surface.

Fig. 12. d. Scanning electron micrograph showed the effect of CIP/NAC combination of (2 MIC/8 mg/ml) on *S. aureus* and pseudomonas spp. ability to form biofilm. No biofilm observed on the surface of stent.

Scanning electron micrographs showed the morphological response and the ability of *S. aureus* and Pseudomonas spp. grown in the presence of Ciprofloxacin (sub-MIC), Nacetylcysteine and their combinations to form biofilm on stent surfaces.

## **4. Conclusion**

610 Scanning Electron Microscopy

Fig. 12. b. Scanning electron micrograph showed the effect of N-acetylcysteine (4 mg/ml) on biofilm formation by *S. aureus* and Pseudomonas spp.. Cells appeared swollen, irregular in shape and small microcolonies observed scattered. A decrease in the numer of adherent cells

Fig. 12. c. Scanning electron micrograph showed the effect of CIP/NAC combination of (MIC/4mg/ml) on *S. aureus* and pseudomonas spp. ability to form biofilm. Cells appeared

elongated, enlarged and scattered with no biofilm mass observed on the surface.

was observed.

The presence of non antimicrobial agent such as N-acetylcysteine (NAC), caused significant decrease in biofilm formation by a variety of bacteria and reduces the production of extracellular polysaccharide matrix while promoting the disruption of mature biofilms. It was found that the inhibitory effect of both ciprofloxacin and N-acetylcysteine was concentration dependent.

CIP/NAC combinations were found to show the highest effect on bacterial adherence inhibition and on the disruption of the already formed biofilms. As N-acetylcysteine increase the therapeutic activity of ciprofloxacin when used in combination by degrading the extracellular polysaccharide matrix of biofilm.

In the chapter, Scanning Electron Microscope is used for the evaluation of medical implants, detection of biofilm and studying the effect of different biofilm inhibitory agents. This technique provides excellent visualization of glycocalyx, which is one of the most prominent features of biofilms and a crucial research subject in the searching for alternative antimicrobial and anti adherent agents treatments.

## **5. Acknowledgment**

Thanks for my professor doctors: Mohamed Ali Mohamed El-Feky, Mostafa Said Khalil El-Rehewy, Mona Amin Hassan(Department of microbiology), Faculty of medicine, Assuit

Application of Scanning Electron Microscopy for

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**31** 

*Turkey* 

**Interrelated Analysis of Performance and** 

**Fouling Behaviors in Forward Osmosis** 

**by Ex-Situ Membrane Characterizations** 

Coskun Aydiner\*, Semra Topcu, Caner Tortop, Ferihan Kuvvet,

In membrane processes, flux decline takes place as an inherent result of membrane fouling that varies with specificity in their implementations. The membrane fouling having uncontrollable or unexplainable complexity in many cases leads to somewhat loss of process efficiency which results mainly in costly pretreatment, higher operating pressure requirement, limited recoveries, feed water loss, frequent chemical cleaning and short lifetimes of membranes as the factors increasing the water and energy costs (Aydiner, 2010, as cited in Tu et al., 2005; Hoek et al., 2008; Van der Bruggen et al., 2008). In recent years, more economical operation of membrane processes is to be taken into account based on lower energy and membrane costs in practice. At this point, understanding the reasons lying under the fouling phenomena as related with a membrane's performance is to be rather valuable task in terms of scientific and technological developments of these processes (Danis & Aydiner, 2009). However, non-generalization course of the fouling during a membrane filtration necessitates the use of either modeling tools or specific analyses for clarifying meaningful performance-fouling relationships in each specific application. The modeling solutions are widely utilized not only to expose these relations but to put forward performance dynamics intended for a main aim of systematic representation and reasoning. A specific modeling study for lab-scale researches mostly results in simulation deficiencies or key limitations in attainment of a definitive solution when compared to that for realworld implementations. As a matter of fact, the development of simple, accurate and effective models needs to produce a solution relying on a "solution-directed focus" approach which includes full-scale consideration of theoretical and practical issues of the events. But, it is explicit that successive synchronization of model assemblies with real-time could not be entirely accomplished by the community of membrane scientists and technologists at this time. In that sense, it can be said that specific membrane analyses based on either in-situ or ex-situ characterizations could be foreseen as a progressive tool on the purpose of obviating interruption or non-coordination of transition among small and large

**1. Introduction** 

\* Corresponding Author

Didem Ekinci, Nadir Dizge and Bulent Keskinler *Gebze Institute of Technology, Faculty of Engineering, Department of Environmental Engineering, Gebze, Kocaeli,* 


## **Interrelated Analysis of Performance and Fouling Behaviors in Forward Osmosis by Ex-Situ Membrane Characterizations**

Coskun Aydiner\*, Semra Topcu, Caner Tortop, Ferihan Kuvvet, Didem Ekinci, Nadir Dizge and Bulent Keskinler *Gebze Institute of Technology, Faculty of Engineering, Department of Environmental Engineering, Gebze, Kocaeli, Turkey* 

#### **1. Introduction**

616 Scanning Electron Microscopy

Trieu-Cuot, P.; Carlier, C.; Martin, P. and Courvalin, P. (1987): Plasmid transfer by

Vranes, J. (2000): Effect of sub minimal inhibitory concentrations of azithromycin on adherence of *pseudomonas aeruginosa* to polystyrene. J. Chemother., 12: 280-285. Wozniak, D. and Keyser, R. (2004): Effects of subinhibitory concentrations of macrolide

Yassien, M.A.; Khardori, N.; Ahmedy, A. and Toama, M. (1995): Modulation of biofilms

antibiotics on *pseudomonas aeruginosa*. Chest., 125: 62-69.

48: 289-94.

2268.

conjugation from Escherichia coli to gram-positive bacteria. FEMS Microbiol. Lett.,

*pseudomonas aeruginosa* by quinolones. Antimicrob. Agents Chemother. 39: 2262-

In membrane processes, flux decline takes place as an inherent result of membrane fouling that varies with specificity in their implementations. The membrane fouling having uncontrollable or unexplainable complexity in many cases leads to somewhat loss of process efficiency which results mainly in costly pretreatment, higher operating pressure requirement, limited recoveries, feed water loss, frequent chemical cleaning and short lifetimes of membranes as the factors increasing the water and energy costs (Aydiner, 2010, as cited in Tu et al., 2005; Hoek et al., 2008; Van der Bruggen et al., 2008). In recent years, more economical operation of membrane processes is to be taken into account based on lower energy and membrane costs in practice. At this point, understanding the reasons lying under the fouling phenomena as related with a membrane's performance is to be rather valuable task in terms of scientific and technological developments of these processes (Danis & Aydiner, 2009). However, non-generalization course of the fouling during a membrane filtration necessitates the use of either modeling tools or specific analyses for clarifying meaningful performance-fouling relationships in each specific application. The modeling solutions are widely utilized not only to expose these relations but to put forward performance dynamics intended for a main aim of systematic representation and reasoning. A specific modeling study for lab-scale researches mostly results in simulation deficiencies or key limitations in attainment of a definitive solution when compared to that for realworld implementations. As a matter of fact, the development of simple, accurate and effective models needs to produce a solution relying on a "solution-directed focus" approach which includes full-scale consideration of theoretical and practical issues of the events. But, it is explicit that successive synchronization of model assemblies with real-time could not be entirely accomplished by the community of membrane scientists and technologists at this time. In that sense, it can be said that specific membrane analyses based on either in-situ or ex-situ characterizations could be foreseen as a progressive tool on the purpose of obviating interruption or non-coordination of transition among small and large

<sup>\*</sup> Corresponding Author

Interrelated Analysis of Performance and Fouling Behaviors

2009; Ling et al., 2010).

membrane type and operating mode.

**2. Materials and methods** 

**2.1 Materials** 

in Forward Osmosis by Ex-Situ Membrane Characterizations 619

development depending on desalination needs. It has remarkably lower cost due to no hydraulic pressure operation, nearly complete rejection of many contaminants, potentially low membrane fouling tendency (Holloway et al., 2007; Cornelissen et al., 2008; Y. Xu et al., 2010; Wang et al., 2010; Chung et al., 2011). But, there are still some constrains in front of becoming widespread of FO's industrial applications. Major drawback is the lack of a membrane having a relatively high flux compared to commercial reverse osmosis (RO) membranes, as an inherent result of the membrane fouling or internal concentration polarization significantly limiting flux efficiency (McCutcheon et al., 2006; Tang, et al., 2010; Wang et al., 2010). The other is the requirement to provide high osmotic pressure difference under continuously operating conditions in which draw solution needs to be concentrated for producing clean water by a complementary process such as reverse osmosis (RO), membrane distillation, membrane osmotic distillation, decomposition with heating, and magnetic separation (Cath et al., 2005a, 2005b; McCutcheon et al., 2005; Martinetti et al.,

In spite of its development potential in membrane science and technology, very few publications on the membrane fouling in FO systems were presented in the literature when compared to those in pressure-driven membrane systems. Hence, FO process was especially preferred in this study to interrelate the performance and internal fouling with membrane surface characteristics. Besides, cheese whey was selected as the feed solution due to its high pollutant capacity with rich nutrient content. In order to render the operation of the system as independent of membrane type and operation mode, the process was employed at normal and reverse operation modes using one each of FO and RO membranes. First, the FO system performance was investigated, and internal membrane fouling was estimated by modeling of the performance data. Thereafter, internal fouling was associated with the results of ex−situ membrane surface characterizations. Solute resistivity which is defined as the internal fouling was estimated depending on salt permeability coefficient at the end of the modeling of the performance data. Ex-situ characterizations of the studied membranes were carried out by SEM, AFM, contact angle, and FTIR measurements. Afterwards, the relation equations among external fouling and each one of performance response parameter which comprises of the water flux, specific water fluxes, salt flux, and net and effective osmotic pressure differences were individually evolved by using contact angle determined as illustrative parameter for external fouling from the relation equations among external fouling and internal fouling in ex-situ characterizations. By this way, in light of significant perspectives obtained by joint interpretations of the results, whether or not the membrane fouling can be correlated with the whole performance was put forward. As concluding remarks, the individual mathematical representations of the relationships of internal and external foulings with the performance were elucidated as regards each dynamics of the whole performance. The prospects of more effective treatment of a membrane process were straightforwardly presented in special to osmotically−driven system as independent of

In the experiments, two different membrane materials in a form of flat sheet were used, one being cellulose triacetate (CTA) FO membrane (Hydration Technologies Inc., OR) and the

scale operations, especially for emerging membrane technologies such as forward osmosis and membrane distillation.

The applications on the membrane fouling characterization falls into two categories: *(i)* laboratory researches involving in-situ monitoring, and *(ii)* field-level studies employing on-line ex-situ scaling observation. In-situ monitoring techniques are to be used for analysing the membrane fouling comprising concentration polarization and cake formation, and are evaluated as an annotation tool in understanding the fouling characteristics (Huang et al., 2010). The most widely used techniques for in situ monitoring of concentration polarization are light deflection techniques (shadowgraphy and refractometry), magnetic resonance imaging, radio isotope labeling, electron diode array microscope, and direct pressure measurements. Whereas, the fouling analyses based on particle deposition or cake layer formation can be carried out by the techniques such as laser triangulometry, optical laser sensor, ultrasonic time-domain reflectometry, electrical impedance spectroscopy, and small-angle neutron scattering (Chen et al., 2004). The main advantage of monitoring-based techniques over traditional lab-scale systems is the ability to visually observe what really happens on the membrane surface simultaneously in realtime (Huang et al., 2010). Ex-situ fouling and scaling detectors are utilized as another important means for understanding of fouling-performance relationships. By the studies under this type of characterization, various observation detectors or fouling simulators can be developed with the intention of controlling the membrane fouling in real-time applications (Uchymiak et al., 2007). Also, various ex-situ membrane investigations based on different analytical techniques such as spectroscopic ellipsometry, x-ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM), atomic force microscope (AFM), Fourier transform infrared (FTIR) spectroscopy, and contact angle etc. can be effectively used for associating the fouling dynamics with the performance (Darton et al., 2004; P. Xu et al., 2010). At the end of a general evaluation of literature on characterizing the membrane fouling, it can be stated that the interrelation of ex-situ characterizations of the membrane fouling concurrently with both process performance and various model response parameters would be a viable simulative tool oriented to removing the transition problems from lab-scale toward real-world. Already, the presence of many techniques and theoretical models developed during the past two decades could makes a sense to better comprehend the interactions between foulants and membrane, develop more viable membranes and employ the process more effectively.

Forward osmosis (FO) is an osmotically−driven membrane process that works spontaneously by osmosis across a semi−permeable membrane. The process possesses a water flow through the membrane from the solution having low concentration (feed solution) toward the solution having high concentration (draw solution) due to the osmotic pressure difference between the solutions (Cath et al., 2006). Along last decade, FO process can be favorably utilized in many applications such as electricity production (Aaberg, 2003; Gormly et al., 2011), power generation (Loeb, 2007; McGinnis et al., 2007), water or wastewater reclamation (Holloway et al., 2007; Cornelissen et al., 2008), seawater desalination or brine concentration (McCutcheon et al., 2006; Low, 2009), concentration of liquid foods (Petrotos & Lazarides, 2001; Dova et al., 2007), protein enrichment and concentration (Yang et al., 2009; Wang et al., 2011), and water purification and reuse in space (Cath et al., 2005a, 2005b). FO process as one of the foremost among processes which have been recently increasingly explored in separation science and technology pursues its

scale operations, especially for emerging membrane technologies such as forward osmosis

The applications on the membrane fouling characterization falls into two categories: *(i)* laboratory researches involving in-situ monitoring, and *(ii)* field-level studies employing on-line ex-situ scaling observation. In-situ monitoring techniques are to be used for analysing the membrane fouling comprising concentration polarization and cake formation, and are evaluated as an annotation tool in understanding the fouling characteristics (Huang et al., 2010). The most widely used techniques for in situ monitoring of concentration polarization are light deflection techniques (shadowgraphy and refractometry), magnetic resonance imaging, radio isotope labeling, electron diode array microscope, and direct pressure measurements. Whereas, the fouling analyses based on particle deposition or cake layer formation can be carried out by the techniques such as laser triangulometry, optical laser sensor, ultrasonic time-domain reflectometry, electrical impedance spectroscopy, and small-angle neutron scattering (Chen et al., 2004). The main advantage of monitoring-based techniques over traditional lab-scale systems is the ability to visually observe what really happens on the membrane surface simultaneously in realtime (Huang et al., 2010). Ex-situ fouling and scaling detectors are utilized as another important means for understanding of fouling-performance relationships. By the studies under this type of characterization, various observation detectors or fouling simulators can be developed with the intention of controlling the membrane fouling in real-time applications (Uchymiak et al., 2007). Also, various ex-situ membrane investigations based on different analytical techniques such as spectroscopic ellipsometry, x-ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM), atomic force microscope (AFM), Fourier transform infrared (FTIR) spectroscopy, and contact angle etc. can be effectively used for associating the fouling dynamics with the performance (Darton et al., 2004; P. Xu et al., 2010). At the end of a general evaluation of literature on characterizing the membrane fouling, it can be stated that the interrelation of ex-situ characterizations of the membrane fouling concurrently with both process performance and various model response parameters would be a viable simulative tool oriented to removing the transition problems from lab-scale toward real-world. Already, the presence of many techniques and theoretical models developed during the past two decades could makes a sense to better comprehend the interactions between foulants and membrane, develop more viable

Forward osmosis (FO) is an osmotically−driven membrane process that works spontaneously by osmosis across a semi−permeable membrane. The process possesses a water flow through the membrane from the solution having low concentration (feed solution) toward the solution having high concentration (draw solution) due to the osmotic pressure difference between the solutions (Cath et al., 2006). Along last decade, FO process can be favorably utilized in many applications such as electricity production (Aaberg, 2003; Gormly et al., 2011), power generation (Loeb, 2007; McGinnis et al., 2007), water or wastewater reclamation (Holloway et al., 2007; Cornelissen et al., 2008), seawater desalination or brine concentration (McCutcheon et al., 2006; Low, 2009), concentration of liquid foods (Petrotos & Lazarides, 2001; Dova et al., 2007), protein enrichment and concentration (Yang et al., 2009; Wang et al., 2011), and water purification and reuse in space (Cath et al., 2005a, 2005b). FO process as one of the foremost among processes which have been recently increasingly explored in separation science and technology pursues its

and membrane distillation.

membranes and employ the process more effectively.

development depending on desalination needs. It has remarkably lower cost due to no hydraulic pressure operation, nearly complete rejection of many contaminants, potentially low membrane fouling tendency (Holloway et al., 2007; Cornelissen et al., 2008; Y. Xu et al., 2010; Wang et al., 2010; Chung et al., 2011). But, there are still some constrains in front of becoming widespread of FO's industrial applications. Major drawback is the lack of a membrane having a relatively high flux compared to commercial reverse osmosis (RO) membranes, as an inherent result of the membrane fouling or internal concentration polarization significantly limiting flux efficiency (McCutcheon et al., 2006; Tang, et al., 2010; Wang et al., 2010). The other is the requirement to provide high osmotic pressure difference under continuously operating conditions in which draw solution needs to be concentrated for producing clean water by a complementary process such as reverse osmosis (RO), membrane distillation, membrane osmotic distillation, decomposition with heating, and magnetic separation (Cath et al., 2005a, 2005b; McCutcheon et al., 2005; Martinetti et al., 2009; Ling et al., 2010).

In spite of its development potential in membrane science and technology, very few publications on the membrane fouling in FO systems were presented in the literature when compared to those in pressure-driven membrane systems. Hence, FO process was especially preferred in this study to interrelate the performance and internal fouling with membrane surface characteristics. Besides, cheese whey was selected as the feed solution due to its high pollutant capacity with rich nutrient content. In order to render the operation of the system as independent of membrane type and operation mode, the process was employed at normal and reverse operation modes using one each of FO and RO membranes. First, the FO system performance was investigated, and internal membrane fouling was estimated by modeling of the performance data. Thereafter, internal fouling was associated with the results of ex−situ membrane surface characterizations. Solute resistivity which is defined as the internal fouling was estimated depending on salt permeability coefficient at the end of the modeling of the performance data. Ex-situ characterizations of the studied membranes were carried out by SEM, AFM, contact angle, and FTIR measurements. Afterwards, the relation equations among external fouling and each one of performance response parameter which comprises of the water flux, specific water fluxes, salt flux, and net and effective osmotic pressure differences were individually evolved by using contact angle determined as illustrative parameter for external fouling from the relation equations among external fouling and internal fouling in ex-situ characterizations. By this way, in light of significant perspectives obtained by joint interpretations of the results, whether or not the membrane fouling can be correlated with the whole performance was put forward. As concluding remarks, the individual mathematical representations of the relationships of internal and external foulings with the performance were elucidated as regards each dynamics of the whole performance. The prospects of more effective treatment of a membrane process were straightforwardly presented in special to osmotically−driven system as independent of membrane type and operating mode.

## **2. Materials and methods**

#### **2.1 Materials**

In the experiments, two different membrane materials in a form of flat sheet were used, one being cellulose triacetate (CTA) FO membrane (Hydration Technologies Inc., OR) and the

Interrelated Analysis of Performance and Fouling Behaviors

conducted with 360 min duration time.

Fig. 1. Experimental setup of lab-scale FO system

**2.2.2 Analytical procedure** 

instruments, respectively.

**2.2.3 FO performance calculations** 

in Forward Osmosis by Ex-Situ Membrane Characterizations 621

that the active or selective layer of the membrane was faced on draw solution and whey, respectively. Cross−flow membrane module was a custom made cell with equivalent flow channel at both sides of the membrane. The membrane module which was made from Delrin acetal resin material (DuPont, Wilmington, Delaware) has an effective membrane area of 140 cm2. The system was employed with 3 L volumes for both the feed (whey) and draw sides. Hydrodynamic flow at the channels was co−currently run to reduce strain on the suspended membrane. Two speed controllable peristaltic pumps (EW 77111−67, Cole Parmer, IL) were used to pump whey liquor and draw solution. Cross−flow velocities on both faces of the membrane were kept constant with 5 L/min (0.5 m/s). The setup was also equipped with a constant temperature water bath (462−7028, VWR Scientific, IL) to maintain the same temperature (25±0.5 oC) at both solutions during FO tests. Each experiment was

Total protein, fat, fat−free dry matter (SNF), total solids, lactose and minerals contents of cheese whey samples were measured by Lactostar instrument equipped with thermal and optic sensors (Funke Gerber Company, Germany). The pH, conductivity, and temperature measurements were done by using WTW Multi 340i Meter (WTW, Weilheim, Germany). For density measurements in the samples, DA−130N density meter (KEM Co., Ltd., Kyoto, Japan) was used. Osmolalities of the samples were determined in duplicate for each data point by Advanced Osmometer instrument (Model 3250−Advanced Instruments Inc., USA) in accordance with freezing point depression method after completing the entire experiment. The analyses of water qualities in whey and draw solution were carried out in accordance with standard methods (American Public Health Association, 2005). Besides, nitrite, nitrate, and TOC, TN parameters were analyzed using GBC UV–visible Cintra 20 spectrometer (Cintra, Australia), and HACH IL 550 TOC–TN (Hach Lange Ltd., Germany)

The permeated water volume, *V* was determined from the osmolality differences of both solutions. First, the osmolalities of solutions were individually measured at definite time intervals along the experiments, and thereafter *V* was determined from the differences of

other being composite polyamide (CPA-3) RO membrane (Hydranautics Inc., CA). FO and RO membranes have salt rejection rates of about 95 and 99.6%, respectively. Their pure water permeabilities, *A* were determined to be approximately 8.1×10<sup>−</sup>3 m3/m2⋅h⋅bar and 3.305×10<sup>−</sup>3 m3/m2⋅h⋅bar, respectively, by means of pure water permeation experiments in a pressure−driven cross−flow membrane system operated at transmembrane pressures of 5, 10, 15, 20 and 25 bar with a constant temperature of 25 °C. FO draw solution used was prepared by dissolving 3 M NaCl (Prolabo, >99%) into the distilled water in order to obtain a high net osmotic pressure difference in the system. Cheese whey was obtained from industrial facilities of Cayirova Milk&Milk Products Inc., located at Kocaeli, Turkey. The characteristics of the raw and FO concentrated cheese whey were presented in Table 1, together with the average water quality values measured in draw solutions after the processing.


Table 1. Characteristics of the raw and FO concentrated cheese whey and the average water quality observed in draw solutions

It should be noted in Table 1 that COD and TOC parameters were measured as soluble COD and soluble TOC in cheese whey samples, and dash means that the relevant parameters were not measured in draw solution samples.

#### **2.2 Methods**

#### **2.2.1 Experimental procedure**

The experiments were carried out by a lab−scale FO system shown in Fig. 1. The process was operated in both normal and reverse orientation mode with a closed loop, which means

other being composite polyamide (CPA-3) RO membrane (Hydranautics Inc., CA). FO and RO membranes have salt rejection rates of about 95 and 99.6%, respectively. Their pure water permeabilities, *A* were determined to be approximately 8.1×10<sup>−</sup>3 m3/m2⋅h⋅bar and 3.305×10<sup>−</sup>3 m3/m2⋅h⋅bar, respectively, by means of pure water permeation experiments in a pressure−driven cross−flow membrane system operated at transmembrane pressures of 5, 10, 15, 20 and 25 bar with a constant temperature of 25 °C. FO draw solution used was prepared by dissolving 3 M NaCl (Prolabo, >99%) into the distilled water in order to obtain a high net osmotic pressure difference in the system. Cheese whey was obtained from industrial facilities of Cayirova Milk&Milk Products Inc., located at Kocaeli, Turkey. The characteristics of the raw and FO concentrated cheese whey were presented in Table 1, together with the average water quality values measured in draw solutions after the

cheese whey

pH 4.97±0.09 4.7±0.1 5.7±1.0 conductivity mS/cm 6.73±0.08 9.8±3.1 179.8±17.5 Cl<sup>−</sup> mg/L 950±28 1,896±1,103 78,776±10,345 COD mg/L 58,220±12,509 82,043±30,774 727±905 TOC mg/L 39,261±2,611 49,013±13,649 30±35 NH4-N mg/L 142±8 167±24 2.0±1.1 NO2-N mg/L 0.04±0.02 0.2±0.2 0 NO3-N mg/L 254±15 280±34 0 TKN mg/L 1,353±139 1,659±343 6.1±5.6 Org-N mg/L 1,211±138 1,492±321 4.1±6.0 TN mg/L 1,607±146 1,939±371 6.1±5.6 PO4-P mg/L 370±10 464±116 7.3±13.3 TP mg/L 470±98 569±154 12.6±23.0 total protein % 2.38±0.18 3.23±1.16 − fat % 0.37±0.07 0.50±0.12 − SNF (fat−free dry matter) % 6.39±0.22 8.82±3.04 − total solid content % 6.76±0.29 9.32±3.14 − lactose % 3.05±0.27 4.48±1.61 − minerals % 0.99±0.06 1.39±0.42 −

Table 1. Characteristics of the raw and FO concentrated cheese whey and the average water

It should be noted in Table 1 that COD and TOC parameters were measured as soluble COD and soluble TOC in cheese whey samples, and dash means that the relevant parameters

The experiments were carried out by a lab−scale FO system shown in Fig. 1. The process was operated in both normal and reverse orientation mode with a closed loop, which means

FO concentrated cheese whey

FO draw solution

parameter unit raw

quality observed in draw solutions

**2.2.1 Experimental procedure** 

**2.2 Methods** 

were not measured in draw solution samples.

processing.

that the active or selective layer of the membrane was faced on draw solution and whey, respectively. Cross−flow membrane module was a custom made cell with equivalent flow channel at both sides of the membrane. The membrane module which was made from Delrin acetal resin material (DuPont, Wilmington, Delaware) has an effective membrane area of 140 cm2. The system was employed with 3 L volumes for both the feed (whey) and draw sides. Hydrodynamic flow at the channels was co−currently run to reduce strain on the suspended membrane. Two speed controllable peristaltic pumps (EW 77111−67, Cole Parmer, IL) were used to pump whey liquor and draw solution. Cross−flow velocities on both faces of the membrane were kept constant with 5 L/min (0.5 m/s). The setup was also equipped with a constant temperature water bath (462−7028, VWR Scientific, IL) to maintain the same temperature (25±0.5 oC) at both solutions during FO tests. Each experiment was conducted with 360 min duration time.

Fig. 1. Experimental setup of lab-scale FO system

## **2.2.2 Analytical procedure**

Total protein, fat, fat−free dry matter (SNF), total solids, lactose and minerals contents of cheese whey samples were measured by Lactostar instrument equipped with thermal and optic sensors (Funke Gerber Company, Germany). The pH, conductivity, and temperature measurements were done by using WTW Multi 340i Meter (WTW, Weilheim, Germany). For density measurements in the samples, DA−130N density meter (KEM Co., Ltd., Kyoto, Japan) was used. Osmolalities of the samples were determined in duplicate for each data point by Advanced Osmometer instrument (Model 3250−Advanced Instruments Inc., USA) in accordance with freezing point depression method after completing the entire experiment. The analyses of water qualities in whey and draw solution were carried out in accordance with standard methods (American Public Health Association, 2005). Besides, nitrite, nitrate, and TOC, TN parameters were analyzed using GBC UV–visible Cintra 20 spectrometer (Cintra, Australia), and HACH IL 550 TOC–TN (Hach Lange Ltd., Germany) instruments, respectively.

#### **2.2.3 FO performance calculations**

The permeated water volume, *V* was determined from the osmolality differences of both solutions. First, the osmolalities of solutions were individually measured at definite time intervals along the experiments, and thereafter *V* was determined from the differences of

Interrelated Analysis of Performance and Fouling Behaviors

frame of the model calculations (see the section 2.2.4).

operation mode (Tan & Ng, 2008).

**2.2.4 Modeling framework** 

Tan & Ng, 2008).

in Forward Osmosis by Ex-Situ Membrane Characterizations 623

the right−hand side was utilized for obtaining theoretical salt permeability coefficient in

The effective osmotic pressure is the principal driven force that essentially governs osmotic water permeation into the draw solution. It takes place as an inherent event of internal concentration polarization which forms the most significant portion of the whole membrane fouling. It is theoretically predicted from the difference of osmotic pressures at the draw and feed sides of membrane's active layer using Eq. (7), as independent of the membrane

> Δ= − ππ

The water flux, *J*w is represented as a function of the effective osmotic pressure difference according to the osmotic−pressure model given as (Cornelissen et al., 2008; Tan & Ng, 2008)

> , , ( ) *w dw <sup>f</sup> <sup>w</sup> J A*=⋅ − π

where *A* is the pure water permeability of the membrane. In the process, another flux term needs to be taken into account due to salt transport taking place simultaneously with water transport. Dissolved salt ions transport from the draw toward the feed, as being in reverse direction to the water flow. It should be designated that mutually transport dynamics goes on across the selective layer of the membrane until system reaches equilibrium. This second flux parameter known to be the salt flux, *J*s is determined by Eq. (9) (Cornelissen et al., 2008;

The modeling of FO process is widely carried out based on water flux from the osmotic pressure difference among the draw and feed solutions which is usually theoretically determined using the data obtained from the experiments. In an osmotically−driven membrane process, water and solute transport in a membrane matrix takes place across a selective interface inside the membrane by convection and diffusion. The fouling phenomena in the membrane become together simultaneously with the transport events, in which two distinctive behaviors known as internal and external concentration polarizations occurs inside and outside the matrix, respectively. The restrictiveness of the fouling on the process efficiency comes from its weakening effect on the effective osmotic pressure of which internal fouling portion plays dominant role. Hence, the main subject in assessing the process performance by the modeling is to be the molar salt concentrations at both sides of the membrane wall which are admitted as a key factor for driven force (Cath et al., 2006; McCutcheon & Elimelech 2006). But, different operation modes (normal and reverse modes) of FO process lead to quite differentiation of the effect of internal concentration polarization depending on the placement of the membrane's active layer toward the draw or feed solution. In a FO process being operated at normal mode in which membrane's selective layer is faced on the draw solution, concentrative internal concentration polarization is valid. Whereas, at reverse mode which means that the selective layer faced on the feed solution, dilutive concentration polarization governs the process. As the indicator of internal

 π

> π

*eff d w*, , *<sup>f</sup> <sup>w</sup>* (7)

, , ( ) *s dw <sup>f</sup> <sup>w</sup> J BC C* =⋅ − (9)

(8)

sequential time points by calculations on osmolarity−based mass balance. These results were also made valid by their verification from time−dependent variations of total solid content (*TSC*) of cheese whey in the feed. The water flux, *J*w was determined from the volume increase in the draw solution using Eq. (1).

$$J\_w = \frac{1}{A\_m} \cdot \frac{\Delta V\_t}{\Delta t} \tag{1}$$

where *A*m is the membrane area, *t* the time, and *V*t the volumetric water permeation at any time. The salt flux, *J*s flowed in reverse direction from the water flux between both solutions was calculated using the following equation (Cornelissen et al., 2008).

$$J\_S = \frac{1}{A\_m} \cdot \frac{\Delta \{C\_t \cdot V\_t\}}{\Delta t} \tag{2}$$

where *C*t is the salt concentration at any time. In addition to *J*w and *J*s, the specific water fluxes as the water flux per net osmotic pressure difference (*J*w′), and the water flux per net osmotic pressure difference normalized with respect to the osmotic pressure of the feed solution (*J*w\*) were also calculated from Eqs. (3), and (4), respectively.

$$J\_w \, ^\circ = \frac{J\_w}{\Delta \pi\_{net}} \tag{3}$$

$$J\_w \stackrel{\*}{=} \frac{J\_w \cdot \pi\_{f,b}}{\Delta \pi\_{net}} \tag{4}$$

Time−dependent variations of two osmotically−driven forces comprising net osmotic pressure difference (Δπnet) and effective osmotic pressure difference (Δπeff) were examined for various operating conditions. The net osmotic pressure refers to the osmotic pressure difference between the draw and feed solutions in the system. It was determined from difference of osmotic pressures of both solutions after analytically measurement of each solution osmolality by (Cornelissen et al., 2008; Tan & Ng, 2008)

$$
\Delta \pi\_{net} = \pi\_{d,b} - \pi\_{f,b} \tag{5}
$$

where πd,b and πf,b are the osmotic pressures of the draw and feed solutions, respectively. The osmotic pressures in the solutions were calculated in accordance with the van't Hoff equation.

$$\mathbf{R} = \mathbf{R} \cdot \mathbf{T} \cdot \begin{bmatrix} \ m & \cdot & d \end{bmatrix} = \mathbf{R} \cdot \mathbf{T} \cdot \begin{bmatrix} \ i \cdot \mathbf{C} \ \end{bmatrix} \tag{6}$$

where *R* is the ideal gas constant (8.314 J/K⋅mol), *T* the absolute temperature (K), *m* the osmolality (mosm/kg), *d* the solution density (kg/L), *i* the van't Hoff factor, and *C* the molar concentration. It should be noted that the multiplication term in square bracket at left−hand side is referred to as the osmolarity (mosm/L) representing total solute concentration in the solution. The equivalence on the left−hand side of the equation was used to determine the osmotic pressure of the draw solution during the experiments. Whereas, the equivalence on the right−hand side was utilized for obtaining theoretical salt permeability coefficient in frame of the model calculations (see the section 2.2.4).

The effective osmotic pressure is the principal driven force that essentially governs osmotic water permeation into the draw solution. It takes place as an inherent event of internal concentration polarization which forms the most significant portion of the whole membrane fouling. It is theoretically predicted from the difference of osmotic pressures at the draw and feed sides of membrane's active layer using Eq. (7), as independent of the membrane operation mode (Tan & Ng, 2008).

$$
\Delta \pi\_{\rm eff} = \pi\_{d,w} - \pi\_{f,w} \tag{7}
$$

#### **2.2.4 Modeling framework**

622 Scanning Electron Microscopy

sequential time points by calculations on osmolarity−based mass balance. These results were also made valid by their verification from time−dependent variations of total solid content (*TSC*) of cheese whey in the feed. The water flux, *J*w was determined from the volume

> <sup>1</sup> *<sup>t</sup> <sup>w</sup> m <sup>V</sup> <sup>J</sup> A t*

where *A*m is the membrane area, *t* the time, and *V*t the volumetric water permeation at any time. The salt flux, *J*s flowed in reverse direction from the water flux between both solutions

*m*

'*<sup>w</sup> <sup>w</sup>*

*w*

*<sup>J</sup> <sup>J</sup>*

*<sup>J</sup> <sup>J</sup>* π

\* *w f* ,*b*

Time−dependent variations of two osmotically−driven forces comprising net osmotic

for various operating conditions. The net osmotic pressure refers to the osmotic pressure difference between the draw and feed solutions in the system. It was determined from difference of osmotic pressures of both solutions after analytically measurement of each

> Δ= − πππ

The osmotic pressures in the solutions were calculated in accordance with the van't Hoff

where *R* is the ideal gas constant (8.314 J/K⋅mol), *T* the absolute temperature (K), *m* the osmolality (mosm/kg), *d* the solution density (kg/L), *i* the van't Hoff factor, and *C* the molar concentration. It should be noted that the multiplication term in square bracket at left−hand side is referred to as the osmolarity (mosm/L) representing total solute concentration in the solution. The equivalence on the left−hand side of the equation was used to determine the osmotic pressure of the draw solution during the experiments. Whereas, the equivalence on

net) and effective osmotic pressure difference (Δ

*<sup>J</sup> A t*

where *C*t is the salt concentration at any time. In addition to *J*w and *J*s, the specific water fluxes as the water flux per net osmotic pressure difference (*J*w′), and the water flux per net osmotic pressure difference normalized with respect to the osmotic pressure of the feed

1 ( ) *t t*

*C V*

*net*

*net*

f,b are the osmotic pressures of the draw and feed solutions, respectively.

= ⋅⋅ ⋅ = ⋅⋅ ⋅ *RT m d RT iC* [ ] [ ] (6)

π

π

was calculated using the following equation (Cornelissen et al., 2008).

solution (*J*w\*) were also calculated from Eqs. (3), and (4), respectively.

solution osmolality by (Cornelissen et al., 2008; Tan & Ng, 2008)

π

*S*

<sup>Δ</sup> = ⋅ <sup>Δ</sup> (1)

Δ ⋅ = ⋅ <sup>Δ</sup> (2)

<sup>=</sup> <sup>Δ</sup> (3)

<sup>⋅</sup> <sup>=</sup> <sup>Δ</sup> (4)

*net d b*, , *<sup>f</sup> <sup>b</sup>* (5)

π

eff) were examined

increase in the draw solution using Eq. (1).

pressure difference (Δ

π

where πd,b and

equation.

π

The water flux, *J*w is represented as a function of the effective osmotic pressure difference according to the osmotic−pressure model given as (Cornelissen et al., 2008; Tan & Ng, 2008)

$$J\_w = A \cdot (\pi\_{d,w} - \pi\_{f,w}) \tag{8}$$

where *A* is the pure water permeability of the membrane. In the process, another flux term needs to be taken into account due to salt transport taking place simultaneously with water transport. Dissolved salt ions transport from the draw toward the feed, as being in reverse direction to the water flow. It should be designated that mutually transport dynamics goes on across the selective layer of the membrane until system reaches equilibrium. This second flux parameter known to be the salt flux, *J*s is determined by Eq. (9) (Cornelissen et al., 2008; Tan & Ng, 2008).

$$J\_s = B \cdot (\mathbb{C}\_{d,w} - \mathbb{C}\_{f,w}) \tag{9}$$

The modeling of FO process is widely carried out based on water flux from the osmotic pressure difference among the draw and feed solutions which is usually theoretically determined using the data obtained from the experiments. In an osmotically−driven membrane process, water and solute transport in a membrane matrix takes place across a selective interface inside the membrane by convection and diffusion. The fouling phenomena in the membrane become together simultaneously with the transport events, in which two distinctive behaviors known as internal and external concentration polarizations occurs inside and outside the matrix, respectively. The restrictiveness of the fouling on the process efficiency comes from its weakening effect on the effective osmotic pressure of which internal fouling portion plays dominant role. Hence, the main subject in assessing the process performance by the modeling is to be the molar salt concentrations at both sides of the membrane wall which are admitted as a key factor for driven force (Cath et al., 2006; McCutcheon & Elimelech 2006). But, different operation modes (normal and reverse modes) of FO process lead to quite differentiation of the effect of internal concentration polarization depending on the placement of the membrane's active layer toward the draw or feed solution. In a FO process being operated at normal mode in which membrane's selective layer is faced on the draw solution, concentrative internal concentration polarization is valid. Whereas, at reverse mode which means that the selective layer faced on the feed solution, dilutive concentration polarization governs the process. As the indicator of internal

Interrelated Analysis of Performance and Fouling Behaviors

spectrophotometer.

in Forward Osmosis by Ex-Situ Membrane Characterizations 625

active and support layers of the membranes studied, respectively. The infrared spectra were recorded in a wave number range of 4000–650 cm−1 on a Bio Rad FTS 175C

Fig. 2. Computational methodology used in the modeling of experimental data

membrane fouling, solute resistivity resulted from soluble concomitant ions inside the active layer can be estimated by Eqs. (10), and (11), respectively (Loeb et al. 1997; Cath et al., 2006; McCutcheon & Elimelech 2006).

$$K = \left(\frac{1}{f\_w}\right) \cdot \left[\ln\left(\frac{B + A \cdot \pi\_{d,w} - f\_w}{B + A \cdot \pi\_{f,b}}\right)\right] \tag{10}$$

$$K = \left(\frac{1}{J\_w}\right) \cdot \left[\ln\left(\frac{B + A \cdot \pi\_{d,b}}{B + J\_w + A \cdot \pi\_{f,w}}\right)\right] \tag{11}$$

In accordance all the equations given above for performance and fouling, performance data related to the water and salt fluxes were simultaneously modeled as a result of which actual internal membrane fouling was defined as solute resistivity at the scope of computational methodology briefly outlined in Fig. 2. The modeling was based upon a framework rendering time−dependent estimation of FO data that was individually fulfilled for each selected time point of each time−dependent data set obtained experimentally. In the computations, theoretical salt permeability coefficient, *B*theoretical was determined using Eq. (12) which is obtained by the equalization of Eqs. (8), and (9), together with the use of the formula on right−hand side of Eq. (6).

$$B = \frac{-J\_s \cdot i \cdot A \cdot R \cdot T}{J\_w} \tag{12}$$

Osmotic pressures at the membrane's wall were predicted using π−*C* relationship given in Eq. (13) which was determined experimentally for NaCl solution up to 4 M concentration.

$$\{\pi\} = \left(5.8062 \times \text{C2}\right) + \left(40.091 \times \text{C}\right) + 0.7289 \ \left(r^2 = 1.000\right) \tag{13}$$

#### **2.2.5 Ex-situ membrane characterizations**

Ex-situ membrane investigations were carried out with the surface characterizations of active and support layers of the studied membranes by SEM, AFM, FTIR, and contact angle. Prior to analytical measurements, representative samples were washed twice with pure water and dried at room temperature. The views of membrane's active and support layers were observed by AFM (NanoScope IV AFM) and SEM (Philips XL30 SFEG). The membrane surface roughness was measured in contact mode by AFM. The mean surface roughness (*R*A), the root mean square error (*R*RMSE) of the average height of surface peaks, and the mean difference in height among the five highest peaks and the five lowest valleys (*R*Z) were established to compare the roughnesses of FO and RO membranes depending on the membrane fouling. After Au coating of the samples, SEM images were taken at 5 kV to view the surface fouling. The contact angle was measured using a goniometry instrument (KSV Instruments, CAM 101) as an indicator for the hydrophilicity or wettability of the membrane surfaces. In the analysis, 2 μL of pure water in a tight syringe was dropped on the surface according to the sessile−drop technique. The results were obtained as the average values of contact angles at both sides of each drop fall on four arbitrary places of the samples. FTIR technique was employed to examine the interactions between whey components with the

membrane fouling, solute resistivity resulted from soluble concomitant ions inside the active layer can be estimated by Eqs. (10), and (11), respectively (Loeb et al. 1997; Cath et al., 2006;

> <sup>1</sup> ln *dw w w f b BA J <sup>K</sup> J BA*

 +⋅ − = ⋅ + ⋅

> <sup>1</sup> ln *d b w w fw*

In accordance all the equations given above for performance and fouling, performance data related to the water and salt fluxes were simultaneously modeled as a result of which actual internal membrane fouling was defined as solute resistivity at the scope of computational methodology briefly outlined in Fig. 2. The modeling was based upon a framework rendering time−dependent estimation of FO data that was individually fulfilled for each selected time point of each time−dependent data set obtained experimentally. In the computations, theoretical salt permeability coefficient, *B*theoretical was determined using Eq. (12) which is obtained by the equalization of Eqs. (8), and (9), together with the use of the

 + ⋅ = ⋅ + +⋅

*J BJ A*

*s*

Eq. (13) which was determined experimentally for NaCl solution up to 4 M concentration.

Ex-situ membrane investigations were carried out with the surface characterizations of active and support layers of the studied membranes by SEM, AFM, FTIR, and contact angle. Prior to analytical measurements, representative samples were washed twice with pure water and dried at room temperature. The views of membrane's active and support layers were observed by AFM (NanoScope IV AFM) and SEM (Philips XL30 SFEG). The membrane surface roughness was measured in contact mode by AFM. The mean surface roughness (*R*A), the root mean square error (*R*RMSE) of the average height of surface peaks, and the mean difference in height among the five highest peaks and the five lowest valleys (*R*Z) were established to compare the roughnesses of FO and RO membranes depending on the membrane fouling. After Au coating of the samples, SEM images were taken at 5 kV to view the surface fouling. The contact angle was measured using a goniometry instrument (KSV Instruments, CAM 101) as an indicator for the hydrophilicity or wettability of the membrane surfaces. In the analysis, 2 μL of pure water in a tight syringe was dropped on the surface according to the sessile−drop technique. The results were obtained as the average values of contact angles at both sides of each drop fall on four arbitrary places of the samples. FTIR technique was employed to examine the interactions between whey components with the

 *C* 2) + (40.091

*w J iART <sup>B</sup> J*

×

*K*

Osmotic pressures at the membrane's wall were predicted using

×

*B A*

, ,

π

π

, ,

π

− ⋅⋅ ⋅ ⋅ <sup>=</sup> (12)

π

 *C*) + 0.7289 (*r*2=1.000) (13)

−*C* relationship given in

(10)

(11)

π

McCutcheon & Elimelech 2006).

formula on right−hand side of Eq. (6).

π

**2.2.5 Ex-situ membrane characterizations** 

] = (5.8062

[

active and support layers of the membranes studied, respectively. The infrared spectra were recorded in a wave number range of 4000–650 cm−1 on a Bio Rad FTS 175C spectrophotometer.

Fig. 2. Computational methodology used in the modeling of experimental data

Interrelated Analysis of Performance and Fouling Behaviors

influence of lower internal fouling.

0

0.0 0.5 1.0 1.5 2.0 2.5 3.0 (c)

solute resistivity)

40

80

Δπ

*B* (L/m2.h)

net (bar)

120

160

200

0123456

0123456

*t* (h)

FO-normal FO-reverse RO-normal RO-reverse

*t* (h)

in Forward Osmosis by Ex-Situ Membrane Characterizations 627

In the process, net driven force decreased in company with the amount of water permeated into the draw solution. Quite different time variations were observed for the model parameters. FO membrane exhibited relatively higher salt permeability even at normal mode depending mainly on higher salt flux compared to that of RO membrane. Figs. 3-(a) and 4-(d) clearly indicated that the water flux performance of the process occurred as an inherent consequence of concentrative or dilutive concentration polarization influence of the membrane's active layer in different operation modes. Despite higher salt passages, FO membrane's inner was fouled less than RO membrane's by organic and inorganic solutes. These results gave some hints on the development of novel FO membranes. So, a FO membrane should preferably be as much thinly as possible designed by enabling high rejection rates especially for monovalent solutes and low molecular weight dissolved organics. In that circumstance, more effective operation of the FO system can be anticipated in terms of the operability with a longer period of time at high performance levels under the

(a)

Δπeff (bar)

*K* (h/m)

Fig. 4. Results for three modeling parameters via net driven force ((a) net osmotic pressure difference, (b) effective osmotic pressure difference, (c) salt permeability coefficient, and (d)

0.0

0

800

1600

2400

3200

4000

(d)

0.8

1.6

2.4

3.2

4.0

0123456

0123456

*t* (h)

*t* (h)

(b)

## **3. Results and discussion**

## **3.1 Analysis of FO Performance**

#### **3.1.1 Water permeation and fluxes**

Time-dependent variations of water permeation and both fluxes depending on two different operation modes of both membranes were depicted in Fig. 3. In the figure, dotted lines are the results estimated from the modeling of experimental water and salt fluxes, whereas solid lines are the non-linear fitting curves for water permeation.

Fig. 3. FO performance at normal and reverse modes for both membranes ((a) volumetric water permeation, (b) water flux, and (c) salt flux)

The process instituted rather different performances at the conditions operated with FO and RO membranes. The system reached a steady state at the end of 360 min except for FO-reverse condition. A conspicuous superiority for reverse operation mode of FO membrane in whey concentration was established in which water more than half of initial water volume of whey was withdrawn as being approximately five times higher than other three varieties. But, FO membrane performed lower salt rejections due to relatively higher salt passages into the whey. It can be accordingly said that novel FO membranes need to be developed to remove the partial deficiency in rejecting salt ions as well as to allow higher water flux performance when also compared to those of RO membrane systems.

### **3.1.2 Driven forces,** *B* **and** *K* **variations**

The effective osmotic pressure difference, salt permeability coefficient and solute resistivity were estimated by means of the model calculations based on computational methodology of experimental data presented in Fig. 2. Time evolutions of the estimated parameters via net driven force determined experimentally were presented in Fig. 4. In the figure, solid lines are the non-linear fitting curves, while dotted lines and symbols are the calculated and theoretical values of *B*, respectively.

Time-dependent variations of water permeation and both fluxes depending on two different operation modes of both membranes were depicted in Fig. 3. In the figure, dotted lines are the results estimated from the modeling of experimental water and salt fluxes, whereas solid

0123456

Fig. 3. FO performance at normal and reverse modes for both membranes ((a) volumetric

The process instituted rather different performances at the conditions operated with FO and RO membranes. The system reached a steady state at the end of 360 min except for FO-reverse condition. A conspicuous superiority for reverse operation mode of FO membrane in whey concentration was established in which water more than half of initial water volume of whey was withdrawn as being approximately five times higher than other three varieties. But, FO membrane performed lower salt rejections due to relatively higher salt passages into the whey. It can be accordingly said that novel FO membranes need to be developed to remove the partial deficiency in rejecting salt ions as well as to allow higher water flux performance when also compared to those of RO membrane

The effective osmotic pressure difference, salt permeability coefficient and solute resistivity were estimated by means of the model calculations based on computational methodology of experimental data presented in Fig. 2. Time evolutions of the estimated parameters via net driven force determined experimentally were presented in Fig. 4. In the figure, solid lines are the non-linear fitting curves, while dotted lines and symbols are the calculated and

(b)

*t* (h) *t* (h)

0123456

(c)

*J*

s (g/m2.h)

**3. Results and discussion 3.1 Analysis of FO Performance 3.1.1 Water permeation and fluxes** 

> FO-normal FO-reverse RO-normal RO- reverse

0123456

*t* (h)

**3.1.2 Driven forces,** *B* **and** *K* **variations** 

theoretical values of *B*, respectively.

water permeation, (b) water flux, and (c) salt flux)

0

systems.

400

800

1200

*V* (mL)

1600

2000

lines are the non-linear fitting curves for water permeation.

(a)

*J*

0

5

10

15

20

25

30

w (L/m2.h)

In the process, net driven force decreased in company with the amount of water permeated into the draw solution. Quite different time variations were observed for the model parameters. FO membrane exhibited relatively higher salt permeability even at normal mode depending mainly on higher salt flux compared to that of RO membrane. Figs. 3-(a) and 4-(d) clearly indicated that the water flux performance of the process occurred as an inherent consequence of concentrative or dilutive concentration polarization influence of the membrane's active layer in different operation modes. Despite higher salt passages, FO membrane's inner was fouled less than RO membrane's by organic and inorganic solutes. These results gave some hints on the development of novel FO membranes. So, a FO membrane should preferably be as much thinly as possible designed by enabling high rejection rates especially for monovalent solutes and low molecular weight dissolved organics. In that circumstance, more effective operation of the FO system can be anticipated in terms of the operability with a longer period of time at high performance levels under the influence of lower internal fouling.

Fig. 4. Results for three modeling parameters via net driven force ((a) net osmotic pressure difference, (b) effective osmotic pressure difference, (c) salt permeability coefficient, and (d) solute resistivity)

Interrelated Analysis of Performance and Fouling Behaviors

in Forward Osmosis by Ex-Situ Membrane Characterizations 629

Fig. 5. Top view AFM images and SEM microphotographs of active layer of clean and fouled membranes ((a) FO clean, (b) FO−normal, (c) FO−reverse, (d) RO clean, (e) RO−normal, and

(f) RO−reverse)

(a)

(b)

(c)

(d)

(e)

(f)

## **3.2 Evaluation of ex-situ fouling**

#### **3.2.1 AFM, SEM and contact angle**

Visual inspection of a membrane are employed by micro- or nano-scale membrane autopsy in which some properties such as microstructure and morphology can be observed to find any physical damage or changing on the surface. By means of analyses based on these observations, significant information are obtained with the intents of determining the membrane characteristics, analyzing the intensity or course of the membrane fouling, and improving the membrane properties for obtaining better performance. In that sense, top view AFM and SEM images of the surfaces of the studied membranes were shown in Figs. 5 and 6 for the active and support layers, respectively. In addition, contact angle and roughness results measured on the membrane surfaces were presented in Table 2.


Table 2. Contact angle (θ °) and roughness (*R*, nm) of the active and support surfaces of FO and RO membranes

Table 2 indicates the general fouling trends in the FO system including organic and inorganic foulants on each opposite side as independent of orientation mode of the membranes. As can be seen from Figs. 5 and 6 and Table 2, RO membrane was fouled more intensively, and wettability and roughness on the active surface of both membranes increased. However, higher increasing rates in external fouling of FO membrane were observed as more than those of RO membrane. Wettability and roughness increased on the support layer of FO membrane, whereas decreased on the support layer of RO membrane in spite of increased fouling. AFM and SEM images and contact angle results pointed out that the surface foulings formed on the layers of FO and RO membranes resulted from different fouling dynamics depending on solute-membrane and solute-solute interactions.

#### **3.2.2 FTIR Spectra**

FTIR technique is widely used to characterize functional groups on a membrane surface via attenuated total reflection (ATR). As is known, organic and inorganic compounds absorb the infrared radiation energy corresponding to the vibrational energy of atomic bonds. By this unique property in the absorption spectrum, the functional groups of a specific compound can be identified by means of FTIR spectra. Hence, the variations of functional chemistry on both surfaces of FO and RO membranes were investigated by FTIR spectroscopy, and thereby general behaviors in the membrane fouling were comparatively evaluated.

Visual inspection of a membrane are employed by micro- or nano-scale membrane autopsy in which some properties such as microstructure and morphology can be observed to find any physical damage or changing on the surface. By means of analyses based on these observations, significant information are obtained with the intents of determining the membrane characteristics, analyzing the intensity or course of the membrane fouling, and improving the membrane properties for obtaining better performance. In that sense, top view AFM and SEM images of the surfaces of the studied membranes were shown in Figs. 5 and 6 for the active and support layers, respectively. In addition, contact angle and

active layer support layer

θ

°) and roughness (*R*, nm) of the active and support surfaces of FO

° *<sup>R</sup>*A

(nm)

*R*Z (nm) *R*RMSE (nm)

*R*RMSE (nm)

roughness results measured on the membrane surfaces were presented in Table 2.

(nm)

FO clean 67.9±10.7 2.2 3.4 12.5 93.5±4.0 2.8 3.6 12.8 RO clean 62.5±2.9 30.1 50.2 191.6 43.0±8.1 74.5 93.4 368.2 FO−normal 43.8±12.4 19.8 30.6 103.1 61.5±3.4 30.1 35.2 115.4 FO−reverse 50.0±2.3 58.4 76.0 258.7 64.5±14.2 23.9 32.3 125.8 RO−normal 43.5±10.8 96.1 116.8 425.8 68.0±5.8 9.7 14.5 42.0 RO−reverse 41.3±17.8 76.2 96.6 326.9 72.8±4.9 36.8 44.8 181.7

Table 2 indicates the general fouling trends in the FO system including organic and inorganic foulants on each opposite side as independent of orientation mode of the membranes. As can be seen from Figs. 5 and 6 and Table 2, RO membrane was fouled more intensively, and wettability and roughness on the active surface of both membranes increased. However, higher increasing rates in external fouling of FO membrane were observed as more than those of RO membrane. Wettability and roughness increased on the support layer of FO membrane, whereas decreased on the support layer of RO membrane in spite of increased fouling. AFM and SEM images and contact angle results pointed out that the surface foulings formed on the layers of FO and RO membranes resulted from different

FTIR technique is widely used to characterize functional groups on a membrane surface via attenuated total reflection (ATR). As is known, organic and inorganic compounds absorb the infrared radiation energy corresponding to the vibrational energy of atomic bonds. By this unique property in the absorption spectrum, the functional groups of a specific compound can be identified by means of FTIR spectra. Hence, the variations of functional chemistry on both surfaces of FO and RO membranes were investigated by FTIR spectroscopy, and

fouling dynamics depending on solute-membrane and solute-solute interactions.

thereby general behaviors in the membrane fouling were comparatively evaluated.

**3.2 Evaluation of ex-situ fouling 3.2.1 AFM, SEM and contact angle** 

θ

θ

° *<sup>R</sup>*A (nm) *<sup>R</sup>*<sup>Z</sup>

membran− mode

Table 2. Contact angle (

and RO membranes

**3.2.2 FTIR Spectra** 

Fig. 5. Top view AFM images and SEM microphotographs of active layer of clean and fouled membranes ((a) FO clean, (b) FO−normal, (c) FO−reverse, (d) RO clean, (e) RO−normal, and (f) RO−reverse)

Interrelated Analysis of Performance and Fouling Behaviors

fouled by severe adsorption of proteins, lactose and fats in whey.

**3.3 Interrelated analysis of performance and fouling** 

**3.3.1 Modeling-based approach** 

π

parameters (Δ

in Forward Osmosis by Ex-Situ Membrane Characterizations 631

To determine surface groups responsible for external fouling, the results obtained by FTIR analyses were depicted in Figs. 7 and 8 for active and support layers of the studied membranes, respectively. In FTIR spectroscopy, whey containing rather different chemical groups absorbs infrared radiation energy at different band levels. Absorption bands at 1600−1700 and 1520−1565 cm<sup>−</sup>1 are responsible for amide I and amide II groups of protein structures, respectively. Besides, the bands at 1725−1745 and 1400−800 cm<sup>−</sup>1 are assigned to C−O stretching of fats, and coupled stretching and bending of carbohydrates, respectively. The vibration bands of clean FO membrane at 1730−1745, 1370−1390, 1235−1245, 1040−1100, and 880−995 cm<sup>−</sup>1 were possibly indicative for C=O stretching of carboxylic acid, symmetric CH3 bending, C−O stretching, C−O stretching of carboxylic acid or C−N stretching, and asymmetric =C−H or =CH2 stretching, respectively. Whereas, those of clean RO membrane at 1630–1680, 1500–1600, 1485, and 1040–1100 cm<sup>−</sup>1 were attributed to the stretchings of C=C, C=C ring, CH2, and C−O, respectively. The bands at around 3445 and 3272 cm-1 corresponded to asymmetric and symmetric O−H stretching of water molecules on the surface due to deficient drying of the samples. As independent of membrane operation mode, lower transmission intensities on the surfaces of RO membrane indicated more severe fouling than those of FO membrane. It was also ascertained that similar transmission peaks corresponded to comparable foulings on the surfaces of both membranes. All of the peaks extended to a range of 700-1750 cm<sup>−</sup>1 proved that the surfaces of both membranes were

This approach, namely "*modeling-based performance analysis*" was focused on the membrane fouling results obtained by the modeling of the performance results. Herein, internal fouling-performance relationships can be explicitly derived with mathematical equations as related to two different groups of performance parameters, one being the measurable

*J*w\* and *B*). At the basis of the membrane solute resistivity, non-linear relationships were evolved according to the statistical power law analysis. The equations belonging to data set at the end of the process were shown in Eqs. (14), and (15), while those belonging to the whole time−dependent data set were given in Eqs. (16) and (17). In these equations, internal fouling was represented separately for the measurable and the prevailing estimated

[ ] [ ] [] 2.780 0.368 0.083

*J B* <sup>−</sup>

[ ] [ ] [] 1.836 0.919 0.015

*J B* <sup>−</sup> <sup>−</sup>

*J J* − − = ⋅Δ ⋅ ⋅ (*r*2=1.000) (14)

= ⋅Δ ⋅ ⋅ (*r*2=1.000) (15)

*J J* <sup>−</sup> = ⋅Δ ⋅ ⋅ (*r*2=0.993; *S*=60.3) (16)

= ⋅Δ ⋅ ⋅ (*r*2=0.989; *S*=73.0) (17)

responses. Standard error of estimate (*S*) and correlation (*r*2) were also presented.

*<sup>m</sup>* (0.00091) *K net w <sup>s</sup>* π

 [ ] 0.018 0.810 \* 0.028 (192.4) *Kpe eff <sup>w</sup>* π

π

*<sup>m</sup>* (0.127) *K net <sup>w</sup> <sup>s</sup>* π

[ ] 0.121 0.779 \* 0.017 (132.7) *Kpe eff <sup>w</sup>*

net, *J*w and *J*s), and the other being the prevailing estimated parameters (Δ

πeff,

Fig. 6. Top view AFM images and SEM microphotographs of support layer of clean and fouled membranes ((a) FO clean, (b) FO−normal, (c) FO−reverse, (d) RO clean, (e) RO−normal, and (f) RO−reverse)

Fig. 6. Top view AFM images and SEM microphotographs of support layer of clean and fouled membranes ((a) FO clean, (b) FO−normal, (c) FO−reverse, (d) RO clean, (e)

RO−normal, and (f) RO−reverse)

(a)

(b)

(c)

(d)

(e)

(f)

To determine surface groups responsible for external fouling, the results obtained by FTIR analyses were depicted in Figs. 7 and 8 for active and support layers of the studied membranes, respectively. In FTIR spectroscopy, whey containing rather different chemical groups absorbs infrared radiation energy at different band levels. Absorption bands at 1600−1700 and 1520−1565 cm<sup>−</sup>1 are responsible for amide I and amide II groups of protein structures, respectively. Besides, the bands at 1725−1745 and 1400−800 cm<sup>−</sup>1 are assigned to C−O stretching of fats, and coupled stretching and bending of carbohydrates, respectively. The vibration bands of clean FO membrane at 1730−1745, 1370−1390, 1235−1245, 1040−1100, and 880−995 cm<sup>−</sup>1 were possibly indicative for C=O stretching of carboxylic acid, symmetric CH3 bending, C−O stretching, C−O stretching of carboxylic acid or C−N stretching, and asymmetric =C−H or =CH2 stretching, respectively. Whereas, those of clean RO membrane at 1630–1680, 1500–1600, 1485, and 1040–1100 cm<sup>−</sup>1 were attributed to the stretchings of C=C, C=C ring, CH2, and C−O, respectively. The bands at around 3445 and 3272 cm-1 corresponded to asymmetric and symmetric O−H stretching of water molecules on the surface due to deficient drying of the samples. As independent of membrane operation mode, lower transmission intensities on the surfaces of RO membrane indicated more severe fouling than those of FO membrane. It was also ascertained that similar transmission peaks corresponded to comparable foulings on the surfaces of both membranes. All of the peaks extended to a range of 700-1750 cm<sup>−</sup>1 proved that the surfaces of both membranes were fouled by severe adsorption of proteins, lactose and fats in whey.

#### **3.3 Interrelated analysis of performance and fouling**

#### **3.3.1 Modeling-based approach**

This approach, namely "*modeling-based performance analysis*" was focused on the membrane fouling results obtained by the modeling of the performance results. Herein, internal fouling-performance relationships can be explicitly derived with mathematical equations as related to two different groups of performance parameters, one being the measurable parameters (Δπnet, *J*w and *J*s), and the other being the prevailing estimated parameters (Δπeff, *J*w\* and *B*). At the basis of the membrane solute resistivity, non-linear relationships were evolved according to the statistical power law analysis. The equations belonging to data set at the end of the process were shown in Eqs. (14), and (15), while those belonging to the whole time−dependent data set were given in Eqs. (16) and (17). In these equations, internal fouling was represented separately for the measurable and the prevailing estimated responses. Standard error of estimate (*S*) and correlation (*r*2) were also presented.

$$K\_m = \text{(0.00091)} \cdot \left[ \Delta \pi\_{net} \right]^{2.780} \cdot \left[ f\_w \right]^{-0.368} \cdot \left[ f\_s \right]^{-0.083} \quad \text{(} \text{ $r$ =1.000)} \tag{14}$$

$$K\_{pe} = \text{(192.4)} \cdot \left[ \Delta \pi\_{eff} \right]^{0.018} \cdot \left[ \left[ \begin{smallmatrix} \* \\ w \end{smallmatrix} \right]^{-0.810} \cdot \left[ B \right]^{0.028} \quad \text{( $r$ =1.000)} \tag{15}$$

$$K\_m = \text{(0.127)} \cdot \left[\Delta\pi\_{net}\right]^{1.836} \cdot \left[f\_w\right]^{-0.919} \cdot \left[f\_s\right]^{0.015} \quad \text{( $r$ =0.993; S=60.3)}\tag{16}$$

$$\mathcal{K}\_{pe} = \text{(132.7)} \cdot \left[ \Delta \pi\_{\text{eff}} \right]^{-0.121} \cdot \left[ J\_w \right]^{-0.779} \cdot \left[ B \right]^{0.017} \quad \text{(\$r^2 = 0.989; S = 73.0)} \tag{17}$$

Interrelated Analysis of Performance and Fouling Behaviors

**3.3.2 Ex-situ-based approach** 

in Forward Osmosis by Ex-Situ Membrane Characterizations 633

Eqs. (14)-(17) proved that internal fouling was remarkably interrelated one by one with both groups of the performance variables. The internal fouling estimates were considerably reliable for simulating the FO performance due to very good agreements. With respect to final performances, the internal fouling increased by increasing of driven forces and salt permeability, while decreased by increasing of water and salt fluxes. The major restricting factor for the performance distinctly seemed to be net pressure difference. Water and salt fluxes were determined to be the second and the third factors in the system efficiency, respectively. According to these results, a FO process has to be employed with a draw solution to be yielded high net driven force. It should be however emphasized that this choice would not be alone sufficient for the expected efficacy in practice. Furthermore, the modeling-based analysis indisputably revealed that the membranes to be specifically designed with the intents of higher driven forces and lower salt permeability would be able to be operated with enough high performances. In other words, a FO membrane having a high salt rejection (>99%) under relatively high effective driven force would be effectively utilized by higher clean water production rates even under a reasonable net driven force.

Ex-situ-based approach can be described as "*ex-situ membrane characterization-based performance analysis*" of which the methodology is principally relied on an illustrative surface parameter to be representative for the internal membrane fouling. The approach follows a solution-directed strategy that includes a focal point toward predicting each performance component as associated individually with representative surface parameter. Accordingly, firstly roughness and contact angle of the membrane surfaces were associated with the internal fouling by appropriate one of linear and non-linear curve-fitting techniques to be given the highest correlation for the relation equations. Thereafter, each performance parameter was related to contact angle determined as representative for the internal fouling. The best relationships obtained in the first step of ex-situ-based performance analysis were shown in Fig. 9. The surface morphologies of the fouled membranes did not give meaningful correlations for which belong to the membrane surface coated by foulants not to the membrane itself. Unlike this case, wettability of both surfaces of FO and RO membranes were relevant to the internal fouling. However, a high correlation of *r*2=0.985 could only be obtained for the wettability of the active surface. This meant that the internal fouling was directly related with the hydrophilicity of the active layer, but not with that of the support layer. Fig. 9-(b) also showed that wettability of the active surface increased as internal fouling increased. Accordingly, internal and external foulings were interrelated with each other by the relation equations. More importantly, the increase in external fouling concurrently led to the increasing of internal fouling. Thereupon, it can be said that a novel FO membrane should be designed in a form of enabling less external fouling on its active

surface without compromising the compatibility with net driven force.

At the second step of the approach, contact angles of the surfaces were interrelated to the performance responses comprising osmotically−driven forces, salt and water passages, and the results were depicted in Figs. 10, 11, and 12, respectively. Equating procedure of wettability of the support layer was determined as practically rather complex of which the relations performed distribution-type variations. Thus, performance simulations were relied on the wettability of the active layer possessed good agreements with the experimental data.

Fig. 7. FTIR spectra of active layers of FO (a) and RO (b) membranes

Fig. 8. FTIR spectra of support layers of FO (a) and RO (b) membranes

Eqs. (14)-(17) proved that internal fouling was remarkably interrelated one by one with both groups of the performance variables. The internal fouling estimates were considerably reliable for simulating the FO performance due to very good agreements. With respect to final performances, the internal fouling increased by increasing of driven forces and salt permeability, while decreased by increasing of water and salt fluxes. The major restricting factor for the performance distinctly seemed to be net pressure difference. Water and salt fluxes were determined to be the second and the third factors in the system efficiency, respectively. According to these results, a FO process has to be employed with a draw solution to be yielded high net driven force. It should be however emphasized that this choice would not be alone sufficient for the expected efficacy in practice. Furthermore, the modeling-based analysis indisputably revealed that the membranes to be specifically designed with the intents of higher driven forces and lower salt permeability would be able to be operated with enough high performances. In other words, a FO membrane having a high salt rejection (>99%) under relatively high effective driven force would be effectively utilized by higher clean water production rates even under a reasonable net driven force.

## **3.3.2 Ex-situ-based approach**

632 Scanning Electron Microscopy

4000 3500 3000 2500 2000 1500 1000 500

4000 3500 3000 2500 2000 1500 1000 500

4000 3500 3000 2500 2000 1500 1000 500

wave number (1/cm)

4000 3500 3000 2500 2000 1500 1000 500

(a)

(b)

(a)

wave number (1/cm)

RO-reverse (b)

transmittance (%)

transmittance (%)

transmittance (%)

transmittance (%)

> RO clean RO-normal

> RO clean RO-normal RO-reverse

 FO clean FO-normal FO-reverse

Fig. 7. FTIR spectra of active layers of FO (a) and RO (b) membranes

Fig. 8. FTIR spectra of support layers of FO (a) and RO (b) membranes

 FO clean FO-normal FO-reverse

> Ex-situ-based approach can be described as "*ex-situ membrane characterization-based performance analysis*" of which the methodology is principally relied on an illustrative surface parameter to be representative for the internal membrane fouling. The approach follows a solution-directed strategy that includes a focal point toward predicting each performance component as associated individually with representative surface parameter. Accordingly, firstly roughness and contact angle of the membrane surfaces were associated with the internal fouling by appropriate one of linear and non-linear curve-fitting techniques to be given the highest correlation for the relation equations. Thereafter, each performance parameter was related to contact angle determined as representative for the internal fouling.

> The best relationships obtained in the first step of ex-situ-based performance analysis were shown in Fig. 9. The surface morphologies of the fouled membranes did not give meaningful correlations for which belong to the membrane surface coated by foulants not to the membrane itself. Unlike this case, wettability of both surfaces of FO and RO membranes were relevant to the internal fouling. However, a high correlation of *r*2=0.985 could only be obtained for the wettability of the active surface. This meant that the internal fouling was directly related with the hydrophilicity of the active layer, but not with that of the support layer. Fig. 9-(b) also showed that wettability of the active surface increased as internal fouling increased. Accordingly, internal and external foulings were interrelated with each other by the relation equations. More importantly, the increase in external fouling concurrently led to the increasing of internal fouling. Thereupon, it can be said that a novel FO membrane should be designed in a form of enabling less external fouling on its active surface without compromising the compatibility with net driven force.

> At the second step of the approach, contact angles of the surfaces were interrelated to the performance responses comprising osmotically−driven forces, salt and water passages, and the results were depicted in Figs. 10, 11, and 12, respectively. Equating procedure of wettability of the support layer was determined as practically rather complex of which the relations performed distribution-type variations. Thus, performance simulations were relied on the wettability of the active layer possessed good agreements with the experimental data.

Interrelated Analysis of Performance and Fouling Behaviors

substances will be improved.

0

driven forces

50

100

150

Δπ

net (bar)

200

250

Δπnet=-0.952\*θ

(*r*² = 0.948)

2

+79.119\*θ-1,492.74

θ (active) θ (support)

300

350

in Forward Osmosis by Ex-Situ Membrane Characterizations 635

As can be seen from the graphs, the hydrophilicity of the active layer was the main representative for the whole system performance in second order polynomial structure. Fig. 10 proved that the performances based on net and effective driven forces realized as being under the predominant influences of external and internal foulings, respectively. The increase in the hydrophilicity yielded by the net force increase was very likely owing to the fact that soluble hydrophilic foulants increased on the surface by ionic binding or adsorption. As to the effective force increase, the hydrophilicity decrease pointed out an external fouling with the fact that soluble or insoluble hydrophobic organics were predominantly bound or attached on the surface as also supported by FTIR analyses. In FOreverse mode, weakening in interactions among organic and inorganic solutes brought about a performance result of higher net force and excessive salt passage, whereby soluble whey organics did not notably penetrated inside the membrane's active layer. Hence, it can be said that targeting a too high rejection rate for organics, especially for ones with lowmolecular weight should be seriously taken into consideration, if a FO membran devoted to the objective implementation of the process to water or wastewater containing organic

contact angle (θ

35 40 45 50 55 60 65 70 75

Δπnet ( fitting) Δπ eff ( fitting)

0 )

Δπeff=0.021\*θ

2

(*r*² = 0.998)


35 40 45 50 55 60 65 70 75

0 ) 0.0

0.5

1.0

1.5

2.0

Δπ

eff (bar)

2.5

3.0

3.5

contact angle (θ

According to Fig. 11, the active surface became more hydrophobic as both salt flux and membrane's salt permeability increased. This concluded that the fouling on the active

Fig. 10. Interrelationships of wettability of active and support layers with osmotically−

Fig. 9. Relationships of surface roughness (a) and wettability (b) to internal fouling

*R*RMSE=0.0403\**K*+215.23, (*r*² = 0.193)

*R*RMSE (support)

fitting

0 500 1000 1500 2000 2500 3000 3500 4000

(b) θ (active)

0 500 1000 1500 2000 2500 3000 3500 4000

*K* (h/m)

, (*r*² = 0.985)

Fig. 9. Relationships of surface roughness (a) and wettability (b) to internal fouling

*R*RMSE=0.0154\**K*+92.03, (*r*² = 0.157)

θ=0.0029\**K*+62.181, (*r*² = 0.766)

(a) *R*RMSE (active)

*K* (h/m)

0

10

20

30

θ=60.903\**K*(-0.047)

 θ (support) fitting

40

50

contact angle, (

θ)

60

70

80

90

100

100

200

surface roughness,

*R*

RMSE (nm)

300

400

500

600

As can be seen from the graphs, the hydrophilicity of the active layer was the main representative for the whole system performance in second order polynomial structure. Fig. 10 proved that the performances based on net and effective driven forces realized as being under the predominant influences of external and internal foulings, respectively. The increase in the hydrophilicity yielded by the net force increase was very likely owing to the fact that soluble hydrophilic foulants increased on the surface by ionic binding or adsorption. As to the effective force increase, the hydrophilicity decrease pointed out an external fouling with the fact that soluble or insoluble hydrophobic organics were predominantly bound or attached on the surface as also supported by FTIR analyses. In FOreverse mode, weakening in interactions among organic and inorganic solutes brought about a performance result of higher net force and excessive salt passage, whereby soluble whey organics did not notably penetrated inside the membrane's active layer. Hence, it can be said that targeting a too high rejection rate for organics, especially for ones with lowmolecular weight should be seriously taken into consideration, if a FO membran devoted to the objective implementation of the process to water or wastewater containing organic substances will be improved.

Fig. 10. Interrelationships of wettability of active and support layers with osmotically− driven forces

According to Fig. 11, the active surface became more hydrophobic as both salt flux and membrane's salt permeability increased. This concluded that the fouling on the active

Interrelated Analysis of Performance and Fouling Behaviors

0

10

*J*

w (L/m2.h) or *J*

w

\*

(L/m2.h.bar)

20

30

*J* <sup>w</sup>=0.204\*θ 2

*J*

*J*

*J* w \*

40

50

in Forward Osmosis by Ex-Situ Membrane Characterizations 637

0 )

contact angle (θ

35 40 45 50 55 60 65 70 75

<sup>w</sup> ( fitting)

<sup>w</sup>' ( fitting)

( fitting)

35 40 45 50 55 60 65 70 75

*J* w

**3.3.3 Performance simulation by integration of the approaches** 

10% in terms of *NRMSE* values, except for the membrane salt permeability.

(*r*² = 1.000)


θ (active) θ (support)

\* =0.049\*θ 2

(*r*² = 1.000)

0 )


2

(*r*² = 1.000)


*J* <sup>w</sup>'=0.003\*θ

0.0

0.2

0.4

0.6

*J*

w ' (L/m2.h.bar)

0.8

1.0

contact angle (θ

Final performances were individually predicted for each response parameter using the second order polynomial relations obtained from the results of ex-situ-based approach (Figs. 10-12). By means of the predicted performance responses, analogical solute resistivities, *K*<sup>m</sup> and *K*pe were separately calculated for the measurable and the prevailing estimated parameters by Eqs. (14) and (15), respectively, to compare their consistency with internal fouling (actual solute resistivity, *K*). The consistency graph was shown in Fig. 13 in which dotted lines were the variation ranges of ±10 and ±20%. In the calculations, deviations from actual values of the responses involving driven forces, water and salt passages were determined by normalized root mean square error (Fig 14). As in modeling-based approach, the use of either the measurable or the prevailing estimated parameters in ex-situ-based approach sufficed for meaningfully describing the relationships of the fouling with the performance. The consistencies of actual and analogical solute resistivities were very close to each other. In order to estimate the performance, analogical internal foulings from the joint solution of the approaches were separately conformed by actual fouling in an acceptable variation range of almost ±10 and ±20% for the prevailing estimated and the measured parameters, respectively. In that sense, Fig. 14 presented that ex-situ-modeling-performance triple with the combination of both approaches was determined as having a capability of

Fig. 12. Interrelationships of wettability of active and support layers with water passage

surface by hydrophobic foulants in whey brought increasing of internal fouling as also supported by Fig. 9-(b). That's why, for more effective operation of a FO system involving organic-inorganic binary, the membrane has to be devised by not only achievement of high organic rejection rate but also acquirement of low hydrophobic fouling on the selective surface. To this end, it can be foreseen that the process would become more economically viable with a higher performance by means of the arrangements to be made on the active surface.

Fig. 11. Interrelationships of wettability of active and support layers with salt passage

Fig. 12 presented that each one of parameters belonging to water passage was well fitted with the active contact angles as in that of driven forces and salt passage. The decrease in the surface wettability accounted for the increase in water permeation performance of the membrane. This meant that, a FO membrane for organic-inorganic binary system have to be an optimum solution for the wettability on the active layer so that the surface should have a hydrophobic domain enough to enable as much as possible of water passage. In light of the results founded on the development of novel FO membranes, matters of fact or opinions deemed to require a thinner membrane than CTA FO membrane of about 50 µm in order to increase the corresponding effective osmotic pressure difference. It can be such that increasing membrane thickness (averagely 200-250 µm for a RO membrane) may be the major triggering factor for lower FO performance by simultaneous contribution of the increase of internal fouling and the decrease of effect of net driven force.

surface by hydrophobic foulants in whey brought increasing of internal fouling as also supported by Fig. 9-(b). That's why, for more effective operation of a FO system involving organic-inorganic binary, the membrane has to be devised by not only achievement of high organic rejection rate but also acquirement of low hydrophobic fouling on the selective surface. To this end, it can be foreseen that the process would become more economically viable with a higher performance by means of the arrangements to be made on the active

> 35 40 45 50 55 60 65 70 75 80 contact angle (θ

> > <sup>s</sup>( fitting) *B* ( fitting)

> > > *B*=2.72\*10-5

\*θ 2 -2.22\*10-3

(*r*² = 0.992)

0 )

 B F B Data1B

35 40 45 50 55 60 65 70 75 80

0 ) 0.000

0.001

0.002

0.003

salt permeability,

*B*

(L/m2.h)

\*θ+4.53\*10-2

0.004

0.005

contact angle (θ

Fig. 12 presented that each one of parameters belonging to water passage was well fitted with the active contact angles as in that of driven forces and salt passage. The decrease in the surface wettability accounted for the increase in water permeation performance of the membrane. This meant that, a FO membrane for organic-inorganic binary system have to be an optimum solution for the wettability on the active layer so that the surface should have a hydrophobic domain enough to enable as much as possible of water passage. In light of the results founded on the development of novel FO membranes, matters of fact or opinions deemed to require a thinner membrane than CTA FO membrane of about 50 µm in order to increase the corresponding effective osmotic pressure difference. It can be such that increasing membrane thickness (averagely 200-250 µm for a RO membrane) may be the major triggering factor for lower FO performance by simultaneous contribution of the

Fig. 11. Interrelationships of wettability of active and support layers with salt passage

increase of internal fouling and the decrease of effect of net driven force.

surface.

0

2

4

salt flux, *J*

s (g/m2.h)

6

8

*J* s =0.083\*θ 2

*J*


θ (active) θ (support)

(*r*² = 1.000)

10

Fig. 12. Interrelationships of wettability of active and support layers with water passage

#### **3.3.3 Performance simulation by integration of the approaches**

Final performances were individually predicted for each response parameter using the second order polynomial relations obtained from the results of ex-situ-based approach (Figs. 10-12). By means of the predicted performance responses, analogical solute resistivities, *K*<sup>m</sup> and *K*pe were separately calculated for the measurable and the prevailing estimated parameters by Eqs. (14) and (15), respectively, to compare their consistency with internal fouling (actual solute resistivity, *K*). The consistency graph was shown in Fig. 13 in which dotted lines were the variation ranges of ±10 and ±20%. In the calculations, deviations from actual values of the responses involving driven forces, water and salt passages were determined by normalized root mean square error (Fig 14). As in modeling-based approach, the use of either the measurable or the prevailing estimated parameters in ex-situ-based approach sufficed for meaningfully describing the relationships of the fouling with the performance. The consistencies of actual and analogical solute resistivities were very close to each other. In order to estimate the performance, analogical internal foulings from the joint solution of the approaches were separately conformed by actual fouling in an acceptable variation range of almost ±10 and ±20% for the prevailing estimated and the measured parameters, respectively. In that sense, Fig. 14 presented that ex-situ-modeling-performance triple with the combination of both approaches was determined as having a capability of 10% in terms of *NRMSE* values, except for the membrane salt permeability.

Interrelated Analysis of Performance and Fouling Behaviors

membrane characterizations were summarized below:

internal foulings, respectively.

**4. Conclusion** 

in Forward Osmosis by Ex-Situ Membrane Characterizations 639

In this study, two different approaches grounded on modeling- and ex-situ membrane characterization-based performance analysis were applied for the evaluation of performance-fouling relationships in a FO system operated as independent of the kind and orientation mode of FO and RO membranes. The prominent findings from normative interpretations together of the results related to the performance, modeling and ex-situ

i. FO membrane operated at reverse mode exhibited better performance due to lower internal fouling compared to RO membrane. But its high salt permeability and higher increasing rate in external fouling were the main drawbacks to its more effective use. ii. The modeling-based approach was admirably applied in interrelating internal fouling and process performance by statistical power law analyses of time-dependent and timeindependent data. The approach proved that the performance restricting factors were, in decreasing order, the net pressure difference and water and salt fluxes, respectively. iii. The ex-situ-based approach was successfully implemented in interrelating both internal-external foulings by non-linear curve-fitting and external fouling-process performance by second order polynomial curve fitting for time-independent data using the contact angle of active membrane surface. It was comprehended by the approach that the increase in external fouling increased the internal fouling. The performance of net and effective driven forces was determined to be in association with external and

iv. Some useful knowledge was obtained on the improvement of FO membranes by thorough evaluation of the results belonging to both the process performance analyses and the individual application of the approaches. According to the performance results, there need to be developed a novel FO membrane to be able to be employed with better salt rejection, higher water flux, and thinner membrane thickness. The modeling-based approach justified the inferences of the stand-alone performance analysis. Whereas, knowledge obtained by the ex-situ-based approach was to some extent greater. As exclusive of others two, without compromising the compatibility with net driven force, a novel FO membrane should have less external foulings on both surfaces for both organic and inorganic foulants to provide the continuity of lower internal fouling. The integrated implementation of both approaches was also successfully carried out intended for the process simulation. The integrated approach was ascertained as a novel and progressive tool in both elucidating the performance-fouling relationships and simulating the whole performance and its components. But, the proposed methodology should amply be examined and confirmed by its experiencing in the performance predictions of full-scale membrane systems. In that sense, for the future works, testing the integrated approach for applications with real-time based on the fouling control may be offered to membrane

At the end of this chapter, it can be essentially said that, despite low fouling tendencies of FO membranes being easily removed by membrane cleaning, development of superior membranes having distinctive properties for various dewatering/purification applications would gain more prominence world-wide in the near future when technical and commercial inadequacies in their diversity are started to be considered more widely. By this means,

specialists to be oriented to ex-situ-based performance simulations.

Fig. 13. Consistencies for solutions based on "*ex-situ-modeling-performance*" integration

Fig. 14. Deviations from actual values of each performance response parameter with regard to the consistencies of the integrated approaches

## **4. Conclusion**

638 Scanning Electron Microscopy

0 800 1600 2400 3200 4000

net (net osmotic pressure difference)

eff (effective osmotic pressure difference)

Performance parameter

*J*

*J*

w

Fig. 14. Deviations from actual values of each performance response parameter with regard

w'

*J*

\*

w

*J*

s

*B*

*K* (h/m)

Fig. 13. Consistencies for solutions based on "*ex-situ-modeling-performance*" integration

<sup>w</sup> (water flux)

(salt flux)

<sup>w</sup>' (specific water flux)

<sup>w</sup>\* (specific water flux)

*B* (salt permeability coefficient)

 Δπ

 Δπ

*J*

*J*

*J*

 *J* s

> Δπeff

Δπ net

to the consistencies of the integrated approaches

0

0.0

0.2

0.4

0.6

*NRMSE*

(Normalized root mean square error)

0.8

1.0

800

1600

*K*i (h/m)

2400

3200

4000

 *K*<sup>m</sup> *K*pe - - - ± 10% -.-.- ± 20%

In this study, two different approaches grounded on modeling- and ex-situ membrane characterization-based performance analysis were applied for the evaluation of performance-fouling relationships in a FO system operated as independent of the kind and orientation mode of FO and RO membranes. The prominent findings from normative interpretations together of the results related to the performance, modeling and ex-situ membrane characterizations were summarized below:


The integrated implementation of both approaches was also successfully carried out intended for the process simulation. The integrated approach was ascertained as a novel and progressive tool in both elucidating the performance-fouling relationships and simulating the whole performance and its components. But, the proposed methodology should amply be examined and confirmed by its experiencing in the performance predictions of full-scale membrane systems. In that sense, for the future works, testing the integrated approach for applications with real-time based on the fouling control may be offered to membrane specialists to be oriented to ex-situ-based performance simulations.

At the end of this chapter, it can be essentially said that, despite low fouling tendencies of FO membranes being easily removed by membrane cleaning, development of superior membranes having distinctive properties for various dewatering/purification applications would gain more prominence world-wide in the near future when technical and commercial inadequacies in their diversity are started to be considered more widely. By this means,

Interrelated Analysis of Performance and Fouling Behaviors

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#### **5. Acknowledgment**

This study was financially supported with a national project (No: 109Y300) by the TUBITAK, the Scientific and Technological Research Council of Turkey. Authors would like to thank to the Cayirova Milk&Milk Products Inc., especially to Mr. Seyhmuz Aslan and Ms. Neslihan Genal, for the cheese whey supplement. Authors would also like to thank to Hydration Technologies Inc., and Hydranautics Inc., due to the membrane supplements.

#### **6. References**


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**5. Acknowledgment** 

**6. References** 

7388

press)

0304-3894


**32** 

**Biodegradation of Pre-Aged** 

**Modified Polyethylene Films** 

*2Laboratory of Scanning Electron Microscopy,* 

*1Department of Biochemistry,* 

 *University of Silesia,* 

*Poland* 

Bożena Nowak1, Jolanta Pająk1 and Jagna Karcz2

Synthetic polymers, which are ubiquitous in modern industrial society, contribute to improving comfort and quality of our life. Currently more than 260 million tonnes of plastics are being produced each year (O'Brine & Thompson, 2010). Among them polyolefins constitute the majority of consumed thermoplastics. Polyolefin materials, such as low-density polyethylene (LDPE), due to the exceptional mechanical and thermal properties, ease of fabrication and low cost, find diverse applications in many fields. Polyethylenes represent 64% of materials used for various applications such as containers, bottles, tubing, plastic bags, greenhouses, mulching films, which are usually discarded after only brief use (Peacock, 2000). These mostly one-trip applications lead to a large quantity of plastic waste accumulating, at the rate of 25 million tons per year (Meenakshi et al., 2002) in landfill and in natural habitats (Thompson et al., 2009). Thus, plastics - the most visible form of trash - have become ubiquitous in our environment, leading to long-term environment, economic and waste management problems (Koutny et al., 2006). Since plastic waste are often soiled by biological substances, physical recycling of these materials turned out impractical and generally undesirable (El-Naggar & Farag, 2010). Incineration of plastics, in turn, has various environmental and social constraints. It seems that the use of plastics, which can re-enter the biological life cycle through biodegradation will be the best choice

The term ''biodegradation" indicates the predominance of biological activity in this phenomenon. Until recently, biodegradation was perceived as a decomposition of substances solely by the action of microorganisms. At present, when the complexity of biodegradation of many substances, especially polymeric materials, is better understood, a new definition of this process divide it into several steps and the process can stop at each stage (Lucas et al., 2008). During the first, extracellular step, which is called biodeterioration, the action of microorganisms in combination with other decomposer organisms or/and abiotic factors fragment polymeric materials into small fractions. To go across the plasmic membrane these small fractions of materials must be depolymerised to low molecular weight products. Due to the mixed action of abiotic factors and microbial communities, which secrete enzymes and free radicals, long polymeric chains are cleaved and small

**1. Introduction**

(Sivan, 2011; Soni et al., 2009).


## **Biodegradation of Pre-Aged Modified Polyethylene Films**

Bożena Nowak1, Jolanta Pająk1 and Jagna Karcz2 *1Department of Biochemistry, 2Laboratory of Scanning Electron Microscopy, University of Silesia, Poland* 

## **1. Introduction**

642 Scanning Electron Microscopy

McGinnis, R.L.; McCutcheon, J.R. & Elimelech, M. (2007). A Novel Ammonia–Carbon

Petrotos, K.B. & Lazarides, H.N. (2001). Osmotic Processing of Liquid Foods, Journal of

Tan,C.H. & Ng, H.Y. (2008). Modified Models to Predict Flux Behavior in Forward Osmosis

Tu, S.C.; Ravindran, V. & Pirbazari, M. (2005). A Pore Diffusion Transport Model for

Uchymiak, M.; Rahardianto, A.; Lyster, E.; Glater, J. & Cohen, Y. (2007). A Novel RO Ex situ

Van der Bruggen, B.; Manttari, M. & Nystrom, M. (2008). Drawbacks of Applying

Wang, K.Y.; Teoh, M.; Nugroho, A. & Chung, T.S. (2011). Integrated Forward Osmosis–

Xu, P.; Bellona, C. & Drewes, J.E. (2010). Fouling of Nanofiltration and Reverse Osmosis

Xu, Y.; Peng, X.; Tang, C.Y.; Fu, Q.S. & Nie, S. (2010). Effect of Draw Solution Concentration

Yang, Q.; Wang, K.Y. & Chung, T.S. (2009). A Novel Dual-Layer Forward Osmosis

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Concentration Polarization and Fouling on Flux Behavior of Forward Osmosis Membranes During Humic Acid Filtration, Journal of Membrane Science, Vol.354,

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Nanofiltration and How to Avoid Them: A Review, Separation and Purification

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Membrane for Protein Enrichment and Concentration, Separation and Purification

Synthetic polymers, which are ubiquitous in modern industrial society, contribute to improving comfort and quality of our life. Currently more than 260 million tonnes of plastics are being produced each year (O'Brine & Thompson, 2010). Among them polyolefins constitute the majority of consumed thermoplastics. Polyolefin materials, such as low-density polyethylene (LDPE), due to the exceptional mechanical and thermal properties, ease of fabrication and low cost, find diverse applications in many fields. Polyethylenes represent 64% of materials used for various applications such as containers, bottles, tubing, plastic bags, greenhouses, mulching films, which are usually discarded after only brief use (Peacock, 2000). These mostly one-trip applications lead to a large quantity of plastic waste accumulating, at the rate of 25 million tons per year (Meenakshi et al., 2002) in landfill and in natural habitats (Thompson et al., 2009). Thus, plastics - the most visible form of trash - have become ubiquitous in our environment, leading to long-term environment, economic and waste management problems (Koutny et al., 2006). Since plastic waste are often soiled by biological substances, physical recycling of these materials turned out impractical and generally undesirable (El-Naggar & Farag, 2010). Incineration of plastics, in turn, has various environmental and social constraints. It seems that the use of plastics, which can re-enter the biological life cycle through biodegradation will be the best choice (Sivan, 2011; Soni et al., 2009).

The term ''biodegradation" indicates the predominance of biological activity in this phenomenon. Until recently, biodegradation was perceived as a decomposition of substances solely by the action of microorganisms. At present, when the complexity of biodegradation of many substances, especially polymeric materials, is better understood, a new definition of this process divide it into several steps and the process can stop at each stage (Lucas et al., 2008). During the first, extracellular step, which is called biodeterioration, the action of microorganisms in combination with other decomposer organisms or/and abiotic factors fragment polymeric materials into small fractions. To go across the plasmic membrane these small fractions of materials must be depolymerised to low molecular weight products. Due to the mixed action of abiotic factors and microbial communities, which secrete enzymes and free radicals, long polymeric chains are cleaved and small

Biodegradation of Pre-Aged Modified Polyethylene Films 645

sufficient energy to cleave C-C bond (Mark et al., 1986) is not absorbed by pure LDPE. Its

Thermo-oxidative degradation is an exposition of polymer to high temperatures. The sensitivity of polyolefins towards thermal oxidation is largely due to the presence of

Though both types of abiotic oxidation produce functional macromolecules susceptible to random cleavage with the formation of low molecular weight oxygenated products containing carbonyl residues (Chiellini et al., 2007), the main difference between photo- and thermal oxidation is that photochemical reactions occur only on the surface of the polymer sample, whereas thermal reactions occur throughout the bulk (Briassoulis et al., 2004).

Hydrolysis is another way by which polymers can undergo degradation (Sahebnazar et al., 2010). However, mechanism of the process strongly depend on polymer structure. Some polymer materials are hydrolysed via both bulk degradation and surface erosion, others

Initial abiotic oxidation and/or hydrolysis of polyethylene is an important stage as it determines the rate of the further biodegradation process. At a second stage of environmental degradation polymer with increased bioavailability and biodegradability should be consumed by microorganisms in soil or during composting (Abrusci et al., 2011). Another of the possible ways to accelerate biodegradation rate of polyethylene in the environment is copolymerisation, blending or grafting with functional polymers and

Few additives having hydrophilic groups make plastic less hydrophobic and susceptible for photo-, chemical and microbial degradation. The most desirable effect of this approach is microbial assimilation of the filler, serving as initial point of microbial attack, resulted in the increase of the surface area of the synthetic bulk material rendering it more susceptible not only to biotic but also abiotic oxidation (Chiellini et al., 2003; Singh & Sharma, 2008). As a result the remaining inert components should disintegrate and disappear (Chandra &

Among biodegradable plastics which can serve as degradable fillers are products of microbial fermentation (e.g., polyesters), modified natural products (e.g., starch, cellulose) and plastics based on chemical synthesis (e.g. polylactic acid, polycaprolactone) (El-Naggar & Farag, 2010). Despite advantages, such as good biodegradability, the commercial production of these polymers is 2.5–10 times more expensive than conventional polymers. Moreover, their physical or chemical properties often restrict their use (Ojeda et al., 2009). Therefore combination of their biodegradability with the excellent properties of conventional materials, such as low-priced LDPE, seems to be a promising alternative (Rosa et al., 2007). Poly(butylene succinate) (PBS) is, next to poly(butylene succinate-co-butylene adipate) (PBSA) and poly(ethylene succinate) (PESu), a member of group of biodegradable aliphatic polyesters trademarked 'Bionolle' invented by Showa Denko (Japan) in 1990, and produced through a polycondensation reaction of glycols with aliphatic dicarboxylic acids and their derivatives (Tserki et al., 2006a). Among these three polyesters, PBS is the only one which is available commercially. However some studies revealed that the biodegradation of PBS is

photo-oxidation can only be induced by some impurities (Briassoulis et al., 2004).

impurities, hydroperoxides and carbonyl groups (Briassoulis et al., 2004).

hydrolyse through only one of the mechanisms.

compounds (Corti et al., 2010; Huang et al., 2005).

much slower than that of PBSA (H-S. Kim et al., 2006).

Rustgi, 1998).

oligomers are generated. After being transported into the cytoplasm the small molecules integrate the metabolism pathways. This so called assimilation is the essential step to produce microbial energy, biomass and primary and secondary metabolites. As mineralisation takes place CO2, N2, CH4, H2O and different salts from completely oxidised metabolites are released in the extracellular environment (Lucas et al., 2008).

According to the new definition, biodegradation of plastics results from combination of biotic and abiotic factors which act synergistically to decompose organic matter. Such interaction of environmental factors is not only beneficial but, in some cases, required to degrade particularly stable compounds.

It is also noteworthy that a frequent source of misunderstanding concerning the term 'biodegradability of polymer' between the polymer scientists and microbiologists originates from the fact that for polymer scientists, degradation means the loss of physical properties, whereas microbiologists are interested in the complete mineralisation of the material (Koutny et al., 2006).

Polyethylene, like many conventional plastics, is resistant to degradation. Its recalcitrance to degradation is traced back to physical and chemical properties that limit chemical reactivity in general (Koutny et al., 2006). In addition, stabilizers contained in industrial polyethylene prevent degradation during processing and usage (Reddy et al., 2008).

Ohtake et al., estimated that it takes about 300 years to degrade LDPE films with thickness of 60µm (Ohtake et al., 1998). However, according to many published reports this estimation implies unrealistic constant rate of LDPE biodegradation. Conventional supposition assumes, that it takes rather thousands than hundreds years for the LDPE to be completely degraded (Kyrikou & Briassoulis, 2007). Formerly, some authors claimed that the poor biodegradability of synthetic substances is a consequence of their short time of presence the environment, so that the enzymes capable of degrading e.g. plastics are not available (Müeller, 2006). However, the research on the microbial metabolism of xenobiotics showed that the evolution of specifity of oxidoreductases secreted by microorganisms is relatively fast (Koutny et al., 2006; Wojcieszyńska et al., 2011). These enzymes involved in the transformation of the molecular edifices increase the polarity of the molecule (Lucas et al., 2008). Nowadays, it also seems that even hydrophobic nature of polyethylene resulted from carbon-only backbone is not a hindrance during biodegradation, since fungi, due to their ability to form hydrophobic proteins, can easily attach to the polymer surface (Sahebnazar et al., 2010). Contrary to previously described features of polyethylene, the high molecular weight itself represents a serious problem because, as a molecule of this size cannot cross a cell wall and a cytoplasmic membrane, it is inaccessible to intracellular metabolic pathways (Koutny et al., 2006). It is therefore necessary to reduce its molecular weight drastically by some predegradation method prior to biological attack. It has been shown that linear paraffin molecules having a molecular weight below 500, or n-alkanes up to C44, can be utilized as a carbon source by microorganisms (Jakubowicz et al., 2006).

Many sources clearly indicate that biodegradation of LDPE could be accelerated by polymer pretreatments, such as photo-oxidation, thermo-oxidation and chemical-oxidation (Sahebnazar et al., 2010).

Photo-oxidative degradation is the process of decomposition of the material by the action of light. It is well known that radiation in the wavelength region of 290–400 nm, which have

oligomers are generated. After being transported into the cytoplasm the small molecules integrate the metabolism pathways. This so called assimilation is the essential step to produce microbial energy, biomass and primary and secondary metabolites. As mineralisation takes place CO2, N2, CH4, H2O and different salts from completely oxidised

According to the new definition, biodegradation of plastics results from combination of biotic and abiotic factors which act synergistically to decompose organic matter. Such interaction of environmental factors is not only beneficial but, in some cases, required to

It is also noteworthy that a frequent source of misunderstanding concerning the term 'biodegradability of polymer' between the polymer scientists and microbiologists originates from the fact that for polymer scientists, degradation means the loss of physical properties, whereas microbiologists are interested in the complete mineralisation of the material

Polyethylene, like many conventional plastics, is resistant to degradation. Its recalcitrance to degradation is traced back to physical and chemical properties that limit chemical reactivity in general (Koutny et al., 2006). In addition, stabilizers contained in industrial polyethylene

Ohtake et al., estimated that it takes about 300 years to degrade LDPE films with thickness of 60µm (Ohtake et al., 1998). However, according to many published reports this estimation implies unrealistic constant rate of LDPE biodegradation. Conventional supposition assumes, that it takes rather thousands than hundreds years for the LDPE to be completely degraded (Kyrikou & Briassoulis, 2007). Formerly, some authors claimed that the poor biodegradability of synthetic substances is a consequence of their short time of presence the environment, so that the enzymes capable of degrading e.g. plastics are not available (Müeller, 2006). However, the research on the microbial metabolism of xenobiotics showed that the evolution of specifity of oxidoreductases secreted by microorganisms is relatively fast (Koutny et al., 2006; Wojcieszyńska et al., 2011). These enzymes involved in the transformation of the molecular edifices increase the polarity of the molecule (Lucas et al., 2008). Nowadays, it also seems that even hydrophobic nature of polyethylene resulted from carbon-only backbone is not a hindrance during biodegradation, since fungi, due to their ability to form hydrophobic proteins, can easily attach to the polymer surface (Sahebnazar et al., 2010). Contrary to previously described features of polyethylene, the high molecular weight itself represents a serious problem because, as a molecule of this size cannot cross a cell wall and a cytoplasmic membrane, it is inaccessible to intracellular metabolic pathways (Koutny et al., 2006). It is therefore necessary to reduce its molecular weight drastically by some predegradation method prior to biological attack. It has been shown that linear paraffin molecules having a molecular weight below 500, or n-alkanes up to C44, can be

metabolites are released in the extracellular environment (Lucas et al., 2008).

prevent degradation during processing and usage (Reddy et al., 2008).

utilized as a carbon source by microorganisms (Jakubowicz et al., 2006).

Many sources clearly indicate that biodegradation of LDPE could be accelerated by polymer pretreatments, such as photo-oxidation, thermo-oxidation and chemical-oxidation

Photo-oxidative degradation is the process of decomposition of the material by the action of light. It is well known that radiation in the wavelength region of 290–400 nm, which have

degrade particularly stable compounds.

(Koutny et al., 2006).

(Sahebnazar et al., 2010).

sufficient energy to cleave C-C bond (Mark et al., 1986) is not absorbed by pure LDPE. Its photo-oxidation can only be induced by some impurities (Briassoulis et al., 2004).

Thermo-oxidative degradation is an exposition of polymer to high temperatures. The sensitivity of polyolefins towards thermal oxidation is largely due to the presence of impurities, hydroperoxides and carbonyl groups (Briassoulis et al., 2004).

Though both types of abiotic oxidation produce functional macromolecules susceptible to random cleavage with the formation of low molecular weight oxygenated products containing carbonyl residues (Chiellini et al., 2007), the main difference between photo- and thermal oxidation is that photochemical reactions occur only on the surface of the polymer sample, whereas thermal reactions occur throughout the bulk (Briassoulis et al., 2004).

Hydrolysis is another way by which polymers can undergo degradation (Sahebnazar et al., 2010). However, mechanism of the process strongly depend on polymer structure. Some polymer materials are hydrolysed via both bulk degradation and surface erosion, others hydrolyse through only one of the mechanisms.

Initial abiotic oxidation and/or hydrolysis of polyethylene is an important stage as it determines the rate of the further biodegradation process. At a second stage of environmental degradation polymer with increased bioavailability and biodegradability should be consumed by microorganisms in soil or during composting (Abrusci et al., 2011).

Another of the possible ways to accelerate biodegradation rate of polyethylene in the environment is copolymerisation, blending or grafting with functional polymers and compounds (Corti et al., 2010; Huang et al., 2005).

Few additives having hydrophilic groups make plastic less hydrophobic and susceptible for photo-, chemical and microbial degradation. The most desirable effect of this approach is microbial assimilation of the filler, serving as initial point of microbial attack, resulted in the increase of the surface area of the synthetic bulk material rendering it more susceptible not only to biotic but also abiotic oxidation (Chiellini et al., 2003; Singh & Sharma, 2008). As a result the remaining inert components should disintegrate and disappear (Chandra & Rustgi, 1998).

Among biodegradable plastics which can serve as degradable fillers are products of microbial fermentation (e.g., polyesters), modified natural products (e.g., starch, cellulose) and plastics based on chemical synthesis (e.g. polylactic acid, polycaprolactone) (El-Naggar & Farag, 2010). Despite advantages, such as good biodegradability, the commercial production of these polymers is 2.5–10 times more expensive than conventional polymers. Moreover, their physical or chemical properties often restrict their use (Ojeda et al., 2009). Therefore combination of their biodegradability with the excellent properties of conventional materials, such as low-priced LDPE, seems to be a promising alternative (Rosa et al., 2007).

Poly(butylene succinate) (PBS) is, next to poly(butylene succinate-co-butylene adipate) (PBSA) and poly(ethylene succinate) (PESu), a member of group of biodegradable aliphatic polyesters trademarked 'Bionolle' invented by Showa Denko (Japan) in 1990, and produced through a polycondensation reaction of glycols with aliphatic dicarboxylic acids and their derivatives (Tserki et al., 2006a). Among these three polyesters, PBS is the only one which is available commercially. However some studies revealed that the biodegradation of PBS is much slower than that of PBSA (H-S. Kim et al., 2006).

Biodegradation of Pre-Aged Modified Polyethylene Films 647

clockwise direction. After four days, when a complete change of the position of samples

Films were positioned 15 cm from the lamp then UV-irradiated using a low-pressure mercury vapor lamp generating energy between 280 nm and 370 nm (TUV 6W, Philips,

Photothermal degradation of polymers proceeded with simultaneous action of UV radiation and temperature under the same conditions as described for the individual

Filamentous fungi *Aspergillus niger*, *Aspergillus terreus*, *Aureobasidium pullulans*, *Paecilomyces varioti*, *Penicillium funiculosum*, *Penicillium ochrochloron*, *Scopulariopsis brevicaulis*, *Trichoderma viride* and their mixture were employed for the biodegradation. *Aspergillus niger* and *Penicillium funiculosum* were isolated from dump in Sosnowiec and their identification was carried out by Institute for Ecology of Industrial Areas in Katowice, Poland. The others were purchased from Institute of Fermentation Technology

Fungi were maintained in test tubes containing Czapek-Doxa medium (NaNO3, 2g; KH2PO4, 0,7g; K2HPO4, 0,3g; KCl, 0,5g; MgSO4x7H2O, 0,5g; FeSO4x7H2O, 0,01g; sucrose, 30g; Bacto Agar (Difco), 20g; distilled water, 1000ml; pH 6.0). Cultures were incubated at 28°C. After completion of sporulation, spores of fungi were separated from hyphae by centrifugation at 4000 rpm and resuspended in SDS solution. The spore suspension at a concentration of 106 spores ml-1 were either used for biodegradation tests or transferred to glycerol solution (50%

Squares of each film (5 replicate samples) unexposed and pre-exposed to abiotic oxidation or/and hydrolysis were sterilised in 70% ethyl alcohol, rinsed with sterile distilled water and aseptically placed in Petri dishes containing modified sucrose-free Czapek-Doxa medium. Each film was covered with 0,1 ml spore suspension. Biodegradation was carried out at 28°C and relative humidity of > 90% for 84 days. Loss of water during incubation was

After the incubation period, polymer samples were delicately removed from the soils and sterilised by immersion in 1% mercuric chloride for 5 minutes, rinsed in water and dried in a

The samples were subjected to dark heated exposure at 50°C in air atmosphere.

within the chamber took place, they were inverted*.* 

Holland) in air atmosphere at room temperature.

**2.2.2.1 Photodegradation procedure** 

**2.2.2.2 Thermodegradation procedure** 

**2.3 Biodegradation experiments** 

and Microbiology in Łódź, Poland.

v/v) before storage at -20°C.

**2.3.2 Biodegradation procedure** 

supplemented with sterile distilled water.

desiccator until a constant weight was obtained.

**2.3.1 Strains of fungi** 

processes.

**2.2.2.3 Photothermodegradation procedure** 

So a few years ago, we started testing the biodegradability PBSA. We are the only team which, in order to accelerate the biodegradation of synthetic polymers, applied modification of LDPE with poly(butylene succinate-co-butylene adipate) (PBSA). Since then we are investigating mechanisms of biodegradation of LDPE/PBSA compositions under different environmental conditions.

It is well known, that even though the mechanism of oxidation of LDPE is more or less understood, the knowledge of the behaviour of this polymer in blends with other materials is not sufficient. It was confirmed that the lack of additivity of component properties in blends is a main reason for difficulties in predicting their life-time (Ołdak & Kaczmarek, 2005).

The purpose of this study was to examine the synergistic or antagonistic effects on the oxidation, hydrolysis and biodegradation of a commercial PELD films filled with PBSA copolyester. Investigations were conducted by first exposing polymeric films to the abiotic oxidation (action of photo- and/or thermal degradation), followed by abiotic hydrolysis under mild conditions, and subsequently to microbial biodegradation.

Several techniques were employed to elucidate the chemical and physical polymers structure. Changes in chemical structure of polymeric films caused by various types of degradation were interpreted on the basis of IR spectra analysis. The scanning electron microscope (SEM) was used to examine these polymers morphologically (surface topography); the excellent resolution provided by the SEM makes it one of the best tools for this purpose.

## **2. Experimentals**

## **2.1 Film preparation**

Low-density polyethylene (LDPE, type "FGNX23-D022", MFR of 2,2g/10min) was purchased from POLICHEM in Kędzierzyn-Koźle. Bionolle® (type #3001, MFR of 1,5g/10min.) was obtained from Showa Denko (Europe) GmbH. The LDPE and Bionolle® were homogenised in a Co-Knetter Buss high-speed mixer at 170°C. The homogenised material was further processed on a PLV 151 type Plasti-Corder extruder for the production of thin films. The compositions 85/15, 70/30, 40/60 LDPE/ Bionolle® were prepared with ratio of 33 rpm and 220, 230, 230, and 235°C set temperatures. Polyethylene film without any additives 100/0 was used as a control material. The films were extruded at the Institute for Engineering Polymer Materials and Dies in Gliwice (Poland). Each film was cut into 40 mm *x* 40 mm squares.

#### **2.2 Abiotic treatment**

#### **2.2.1 Hydrolytic aging**

Hydrolysis of films was carried out in phosphate buffer (pH 7.4) with sodium azide to prevent growth of microorganisms for 84 days.

## **2.2.2 Oxidative aging**

The photo- and/or thermal degradation of polymers was achieved by placing the samples in an adapted oven for 16 days. Every 24 hours, the location of the samples was changed in a clockwise direction. After four days, when a complete change of the position of samples within the chamber took place, they were inverted*.* 

#### **2.2.2.1 Photodegradation procedure**

646 Scanning Electron Microscopy

So a few years ago, we started testing the biodegradability PBSA. We are the only team which, in order to accelerate the biodegradation of synthetic polymers, applied modification of LDPE with poly(butylene succinate-co-butylene adipate) (PBSA). Since then we are investigating mechanisms of biodegradation of LDPE/PBSA compositions under different

It is well known, that even though the mechanism of oxidation of LDPE is more or less understood, the knowledge of the behaviour of this polymer in blends with other materials is not sufficient. It was confirmed that the lack of additivity of component properties in blends is

The purpose of this study was to examine the synergistic or antagonistic effects on the oxidation, hydrolysis and biodegradation of a commercial PELD films filled with PBSA copolyester. Investigations were conducted by first exposing polymeric films to the abiotic oxidation (action of photo- and/or thermal degradation), followed by abiotic hydrolysis

Several techniques were employed to elucidate the chemical and physical polymers structure. Changes in chemical structure of polymeric films caused by various types of degradation were interpreted on the basis of IR spectra analysis. The scanning electron microscope (SEM) was used to examine these polymers morphologically (surface topography); the excellent resolution provided by the SEM makes it one of the best tools for

Low-density polyethylene (LDPE, type "FGNX23-D022", MFR of 2,2g/10min) was purchased from POLICHEM in Kędzierzyn-Koźle. Bionolle® (type #3001, MFR of 1,5g/10min.) was obtained from Showa Denko (Europe) GmbH. The LDPE and Bionolle® were homogenised in a Co-Knetter Buss high-speed mixer at 170°C. The homogenised material was further processed on a PLV 151 type Plasti-Corder extruder for the production of thin films. The compositions 85/15, 70/30, 40/60 LDPE/ Bionolle® were prepared with ratio of 33 rpm and 220, 230, 230, and 235°C set temperatures. Polyethylene film without any additives 100/0 was used as a control material. The films were extruded at the Institute for Engineering Polymer Materials and Dies in Gliwice (Poland). Each film was cut into 40 mm

Hydrolysis of films was carried out in phosphate buffer (pH 7.4) with sodium azide to

The photo- and/or thermal degradation of polymers was achieved by placing the samples in an adapted oven for 16 days. Every 24 hours, the location of the samples was changed in a

a main reason for difficulties in predicting their life-time (Ołdak & Kaczmarek, 2005).

under mild conditions, and subsequently to microbial biodegradation.

environmental conditions.

this purpose.

**2. Experimentals 2.1 Film preparation** 

*x* 40 mm squares.

**2.2 Abiotic treatment 2.2.1 Hydrolytic aging** 

**2.2.2 Oxidative aging** 

prevent growth of microorganisms for 84 days.

Films were positioned 15 cm from the lamp then UV-irradiated using a low-pressure mercury vapor lamp generating energy between 280 nm and 370 nm (TUV 6W, Philips, Holland) in air atmosphere at room temperature.

#### **2.2.2.2 Thermodegradation procedure**

The samples were subjected to dark heated exposure at 50°C in air atmosphere.

#### **2.2.2.3 Photothermodegradation procedure**

Photothermal degradation of polymers proceeded with simultaneous action of UV radiation and temperature under the same conditions as described for the individual processes.

### **2.3 Biodegradation experiments**

#### **2.3.1 Strains of fungi**

Filamentous fungi *Aspergillus niger*, *Aspergillus terreus*, *Aureobasidium pullulans*, *Paecilomyces varioti*, *Penicillium funiculosum*, *Penicillium ochrochloron*, *Scopulariopsis brevicaulis*, *Trichoderma viride* and their mixture were employed for the biodegradation. *Aspergillus niger* and *Penicillium funiculosum* were isolated from dump in Sosnowiec and their identification was carried out by Institute for Ecology of Industrial Areas in Katowice, Poland. The others were purchased from Institute of Fermentation Technology and Microbiology in Łódź, Poland.

Fungi were maintained in test tubes containing Czapek-Doxa medium (NaNO3, 2g; KH2PO4, 0,7g; K2HPO4, 0,3g; KCl, 0,5g; MgSO4x7H2O, 0,5g; FeSO4x7H2O, 0,01g; sucrose, 30g; Bacto Agar (Difco), 20g; distilled water, 1000ml; pH 6.0). Cultures were incubated at 28°C. After completion of sporulation, spores of fungi were separated from hyphae by centrifugation at 4000 rpm and resuspended in SDS solution. The spore suspension at a concentration of 106 spores ml-1 were either used for biodegradation tests or transferred to glycerol solution (50% v/v) before storage at -20°C.

## **2.3.2 Biodegradation procedure**

Squares of each film (5 replicate samples) unexposed and pre-exposed to abiotic oxidation or/and hydrolysis were sterilised in 70% ethyl alcohol, rinsed with sterile distilled water and aseptically placed in Petri dishes containing modified sucrose-free Czapek-Doxa medium. Each film was covered with 0,1 ml spore suspension. Biodegradation was carried out at 28°C and relative humidity of > 90% for 84 days. Loss of water during incubation was supplemented with sterile distilled water.

After the incubation period, polymer samples were delicately removed from the soils and sterilised by immersion in 1% mercuric chloride for 5 minutes, rinsed in water and dried in a desiccator until a constant weight was obtained.

Biodegradation of Pre-Aged Modified Polyethylene Films 649

Among others, Fourier transform infrared (FTIR) spectroscopy is most widely used in determining the structural changes in macromolecules. Since it is known that degradation of polymers can proceed *via* both hydrolysis and oxidation, with this tool it is possible to estimate the extend of modification of polymer main chain due to the action of abiotic or biotic factors. It is assumed, that the mechanism of polymer degradation can be determined by measuring the levels of ketone carbonyl, ester carbonyl and internal double bond absorbance peaks (Gilan (Orr) et al., 2004; Jakubowicz et al., 2006; Sudhakar

Scanning electron microscopy (SEM) is a useful imaging approach for the visualization of different polymers, because it provides a consistent picture of the polymer morphology as a non-uniform structure characterised by variable thickness and variable polymer density. This technique allows to illustrate the surface topography of polymers with high resolution. Due to its high lateral resolution, its great depth of focus and its facility for x-ray microanalysis (SEM/EDX), SEM is often used in material science – including polymer sciences to elucidate the microscopic structure of polymers. In SEM, the surface of nonconductive samples must be coated with a thin layer of gold or platinum. Sometimes, a surface pretreatment (ion sputtering or chemical etching) is carried out to reveal structural details. Moreover, brittle fracture of samples (in liquid nitrogen in cryo-SEM) can give information about the internal morphology of bulk specimens. SEM micrographs indicate that polymers are characterised by different surface features and heterogenous local density of chemical components. They also show surface defects such as cracks, etching residues,

Currently, a number of different SEM techniques and sample preparation methods have been employed for study of polymers structure, including ultra-high resolution field emission SEM (UHR FE-SEM), scanning transmission SEM (STEM), low-vacuum SEM (LVSEM/cryo SEM) and environmental SEM (ESEM). In LVSEM mode, the delicate polymer samples are observed in frozen state, whereas in an ESEM mode the specimens can contain liquids. SEM equipped with an energy dispersive X-ray spectrometry profiling (SEM/EDX) is widely used to characterize the variation of chemical composition of polymers interface. STEM is used to analyze lamellar arrangements in polymers, their dimensions and crystallography. Especially the recent development of ultra-high resolution field emission scanning electron microscopy has opened new opportunities in polymer study at the molecular scale. These SEM techniques provide complementary data to

transmission electron microscopy (TEM) and scanning probe microscopy (SPM).

In polymer studies the following applications of SEM have been made: study of surface microstructure of polymer films, fibres and powders (amorphous and crystalline); investigation of liquid crystals; control of the polymerization process; study of the structure of copolymers, polymer blends and networks (investigation of miscibility and adhesion of components); observation of structural defects and sample roughness; changes in the structure of polymers during stretching and upon loading (formation of crazes and cracks; fracture surface morphology; chemical agent transport processes through membranes (porosity of membranes) and after biotic (microbiological) treatment (Bonhomme et al., 2003; Borghei et al., 2010; González et al., 2006; Guise et al., 2011; Šašek et al., 2006; Vezie et al., 1995).

et al., 2008).

differential swelling, depressions and perforations.

### **2.4 Assessment of degradation**

Sample weight loss was determined gravimetrically on an analytical balance (Mettler Toledo, AB 240-S).

The tensile strength (Rm) and elongation at brake (εr) tests were carried out in accordance with EN ISO 527-3: 1995; EN ISO 527-1: 1996; EN ISO 527-2: 1996. Tests were performed on a tensile tester (INSTRON 4466). Results of mechanical strength evaluation were averaged over 5 replicate specimen.

Infrared spectra of the films were recorded on an FTS 40A spectrophotometer (BIO-RAD) over a range of 3700-700 cm-1 at a resolution of 2 cm-1 and over 32 scans. Samples were dissolved in a mixture of decahydronaphthalene and dimethylformamide at 70°C and analysed as thin films on the NaCl cell surfaces after evaporation of the solvent. Carbonyl index (CI) and terminal double bond index, were used as a parameters to monitor the degree of degradation of films. Carbonyl index is the ratio between the absorbance of the carbonyl peak (1712 cm−1) and the absorbance of the CH2 groups at 1465 cm−1. Terminal double bond index is the ratio between the absorbance of the terminal double-bond peak (908 cm−1) and the absorbance of the CH2 groups at 1465 cm−1 (Gilan (Orr) et al., 2004).

The pieces of control and treated polyethylene films were cut with a sharp blade to obtain a small cube (5 mm). The cube samples were mounted on an aluminium stubs with doublesided adhesive carbon tape, and sputter-coated in Pelco SC-6 sputter coater (25 mA and 0,8 hPa) for 30 seconds with a thin film of gold to improve the electrically conducting properties of the sample surface. After sputtering the samples were imaged by the Tesla BS 340 scanning electron microscope (SEM) in a high-vacuum mode operating at 20 kV with secondary electron detector (ESD), and working distance (WD) of 10 mm. Collected images were compared with those recorded for the original untreated samples.

## **3. Results and discussion**

The term degradation with respect to decomposition of polymeric materials has not been explicitly specified. The main problem is to determine the susceptibility of the polymer material to degradation in the environment and the length of time during which process will last. Several methods can be used to estimate polymer deterioration. Frequently used methods rely on gravimetric, spectroscopic and microscopic techniques, mostly in combination with each other (Sudhakar et al., 2008).

A simple and quick way to measure the degradation of polymers is by determining the weight variations. However, this measurement itself cannot be a reliable indicative of material degradability, since both an increase in weight and a weight loss of polymer sample, not directly related to the breakdown of polymer chain, may occur. A good example is an increase in weight due to accumulation of microorganism, whereas loss of weight can be due to the vanishing of volatile and soluble impurities (Lucas et al., 2008).

Deterioration of polymers can be also evaluated by change in their rheological properties. Contrary to the weight measurement, these properties directly depend on molecular weight of polymers, their crystallinity and the presence of branches and crosslinkings effects (Briassoulis et al., 2004).

Sample weight loss was determined gravimetrically on an analytical balance (Mettler

The tensile strength (Rm) and elongation at brake (εr) tests were carried out in accordance with EN ISO 527-3: 1995; EN ISO 527-1: 1996; EN ISO 527-2: 1996. Tests were performed on a tensile tester (INSTRON 4466). Results of mechanical strength evaluation were averaged

Infrared spectra of the films were recorded on an FTS 40A spectrophotometer (BIO-RAD) over a range of 3700-700 cm-1 at a resolution of 2 cm-1 and over 32 scans. Samples were dissolved in a mixture of decahydronaphthalene and dimethylformamide at 70°C and analysed as thin films on the NaCl cell surfaces after evaporation of the solvent. Carbonyl index (CI) and terminal double bond index, were used as a parameters to monitor the degree of degradation of films. Carbonyl index is the ratio between the absorbance of the carbonyl peak (1712 cm−1) and the absorbance of the CH2 groups at 1465 cm−1. Terminal double bond index is the ratio between the absorbance of the terminal double-bond peak (908 cm−1) and the absorbance of the CH2 groups at 1465 cm−1 (Gilan (Orr) et al., 2004).

The pieces of control and treated polyethylene films were cut with a sharp blade to obtain a small cube (5 mm). The cube samples were mounted on an aluminium stubs with doublesided adhesive carbon tape, and sputter-coated in Pelco SC-6 sputter coater (25 mA and 0,8 hPa) for 30 seconds with a thin film of gold to improve the electrically conducting properties of the sample surface. After sputtering the samples were imaged by the Tesla BS 340 scanning electron microscope (SEM) in a high-vacuum mode operating at 20 kV with secondary electron detector (ESD), and working distance (WD) of 10 mm. Collected images

The term degradation with respect to decomposition of polymeric materials has not been explicitly specified. The main problem is to determine the susceptibility of the polymer material to degradation in the environment and the length of time during which process will last. Several methods can be used to estimate polymer deterioration. Frequently used methods rely on gravimetric, spectroscopic and microscopic techniques, mostly in

A simple and quick way to measure the degradation of polymers is by determining the weight variations. However, this measurement itself cannot be a reliable indicative of material degradability, since both an increase in weight and a weight loss of polymer sample, not directly related to the breakdown of polymer chain, may occur. A good example is an increase in weight due to accumulation of microorganism, whereas loss of weight can

Deterioration of polymers can be also evaluated by change in their rheological properties. Contrary to the weight measurement, these properties directly depend on molecular weight of polymers, their crystallinity and the presence of branches and crosslinkings effects

be due to the vanishing of volatile and soluble impurities (Lucas et al., 2008).

were compared with those recorded for the original untreated samples.

**2.4 Assessment of degradation** 

Toledo, AB 240-S).

over 5 replicate specimen.

**3. Results and discussion** 

(Briassoulis et al., 2004).

combination with each other (Sudhakar et al., 2008).

Among others, Fourier transform infrared (FTIR) spectroscopy is most widely used in determining the structural changes in macromolecules. Since it is known that degradation of polymers can proceed *via* both hydrolysis and oxidation, with this tool it is possible to estimate the extend of modification of polymer main chain due to the action of abiotic or biotic factors. It is assumed, that the mechanism of polymer degradation can be determined by measuring the levels of ketone carbonyl, ester carbonyl and internal double bond absorbance peaks (Gilan (Orr) et al., 2004; Jakubowicz et al., 2006; Sudhakar et al., 2008).

Scanning electron microscopy (SEM) is a useful imaging approach for the visualization of different polymers, because it provides a consistent picture of the polymer morphology as a non-uniform structure characterised by variable thickness and variable polymer density. This technique allows to illustrate the surface topography of polymers with high resolution. Due to its high lateral resolution, its great depth of focus and its facility for x-ray microanalysis (SEM/EDX), SEM is often used in material science – including polymer sciences to elucidate the microscopic structure of polymers. In SEM, the surface of nonconductive samples must be coated with a thin layer of gold or platinum. Sometimes, a surface pretreatment (ion sputtering or chemical etching) is carried out to reveal structural details. Moreover, brittle fracture of samples (in liquid nitrogen in cryo-SEM) can give information about the internal morphology of bulk specimens. SEM micrographs indicate that polymers are characterised by different surface features and heterogenous local density of chemical components. They also show surface defects such as cracks, etching residues, differential swelling, depressions and perforations.

Currently, a number of different SEM techniques and sample preparation methods have been employed for study of polymers structure, including ultra-high resolution field emission SEM (UHR FE-SEM), scanning transmission SEM (STEM), low-vacuum SEM (LVSEM/cryo SEM) and environmental SEM (ESEM). In LVSEM mode, the delicate polymer samples are observed in frozen state, whereas in an ESEM mode the specimens can contain liquids. SEM equipped with an energy dispersive X-ray spectrometry profiling (SEM/EDX) is widely used to characterize the variation of chemical composition of polymers interface. STEM is used to analyze lamellar arrangements in polymers, their dimensions and crystallography. Especially the recent development of ultra-high resolution field emission scanning electron microscopy has opened new opportunities in polymer study at the molecular scale. These SEM techniques provide complementary data to transmission electron microscopy (TEM) and scanning probe microscopy (SPM).

In polymer studies the following applications of SEM have been made: study of surface microstructure of polymer films, fibres and powders (amorphous and crystalline); investigation of liquid crystals; control of the polymerization process; study of the structure of copolymers, polymer blends and networks (investigation of miscibility and adhesion of components); observation of structural defects and sample roughness; changes in the structure of polymers during stretching and upon loading (formation of crazes and cracks; fracture surface morphology; chemical agent transport processes through membranes (porosity of membranes) and after biotic (microbiological) treatment (Bonhomme et al., 2003; Borghei et al., 2010; González et al., 2006; Guise et al., 2011; Šašek et al., 2006; Vezie et al., 1995).

Biodegradation of Pre-Aged Modified Polyethylene Films 651

100/0 16,40 516,0 85/15 14,02 518,4 70/30 10,80 336,8 40/60 20,50 485,0

The changes in tensile strength and elongation at break of films after biodegradation with selected fungi are shown in Figure 1. It has been reported that the changes in tensile strength

Although little weight loss of polyethylene was observed after 84 days of biodegradation, a marked reduction in tensile strength imply that both *Penicillium funiculosum* and mixed fungi population excrete enzymes able to cleave macromolecules of LDPE. Slight increase in the elongation at break in samples incubated with mixed population of fungi, could be tentatively attributed to a sort of plasticisation effect exerted by low molecular weight fractions produced in the first stage of the biodegradation of the polymer matrix (Chiellini et al., 2003). After 84 days of biodegradation, the tensile strength of film containing 30% Bionolle® decreased by about 20%. At this time, elongation at break changed by about 74% and 52% after incubation with *Penicillium funiculosum* and mixed fungi population, respectively. Far-advanced decomposition of 40/60 blend, prevented determination of its

Data obtained from FTIR spectra (spectra are presented elsewhere (Łabużek et al.) of examined films showed an increase in carbonyl and double bond indices, except for LDPE (decrease in carbonyl index by 57%), incubated with mixed population of fungi (Łabużek et al., 2006a). Increase in the internal double bond index is in accordance with the biodegradation mechanism suggested by Albertsson et al. who reported on the formation of terminal double bonds as a result of exposure to biotic environment (Albertsson et al., 1987). This can be attributed to biotic dehydrogenation (Chiellini et al., 2003). However, in contrast

and elongation loss are excellent indicators of degradation (Reddy et al., 2008).

Mechanical properties Rm, MPa εr,%

LDPE/Bionolle® film compositions

Table 2. Mechanical properties of control films

Fig. 1. Mechanical properties of films after biodegradation

mechanical properties.

## **3.1 Biodegradation**

The percentage weight loss of LDPE and LDPE/Bionolle® compositions after biodegradation with different filamentous fungi is shown in Table 1 (Łabużek et al., 2006a; Nowak et al., 2010).


Table 1. Weight loss of films after biodegradation

It was found that pure LDPE and the blends 85/15 and 70/30 showed little loss of mass, which probably reflected the inertness of LDPE towards biological degradation. Unfortunately, even 30% Bionolle® wasn't enough to observe sufficient weight loss of film. Probably, the polyethylene matrix prevented microbes from accessing the polyester in the depth of the film. Similar relationships were observed for the polyolefins modified with 6- 15% starch content, where only the surface of the material was susceptible to biodegradation (Nakamura et al., 2005). In a separate study, Rosa et al. found that the pure LDPE and the blends 25PHB/75LDPE and 50PHB/50LDPE showed little or no loss of mass during aging in simulated soil (Rosa et al., 2007). Also Lee et al. discovered that polystyrene (PS) in the P(3HB-co-3HV)/PS (95/5 by wt%) blend acts as a retardant of enzymatic attack to the surface of the blend film (Lee et al., 2003).

This phenomenon is likely due to the high-molecular weight hydrophobic chains of the synthetic polymer preventing enzymes from accessing the biodegradable polymers contained within the material (Nowak et al., 2011).

Among polymeric compositions, film containing 60% Bionolle® had the most obvious reduction in weight. Film lost 24%, 60% and 58% of its initial mass after incubation with *Aspergillus niger*, *Penicillium funiculosum* and mixed population of fungi, respectively. Other fungi caused slight film weight loss ranging from 0,25% to 1,17%. Taken together with our previous studies, the present findings show, that only fungi which were able to decompose Bionolle® (80-100% for 90 days), were also able to degrade 40/60 composition (Nowak et al., 2010).

Mechanical properties of the investigated films before degradation are shown in Table 2.

The percentage weight loss of LDPE and LDPE/Bionolle® compositions after biodegradation with different filamentous fungi is shown in Table 1 (Łabużek et al., 2006a;

*Aspergillus niger* 0,12 0,16 1,29 24,0 *Aspergillus terreus* 0,08 0,08 0,17 0,35 *Aureobasidium pullulans* 0,04 0,05 0,28 0,25 *Paecilomyces varioti* 0,07 0,06 0,22 0,29 *Penicillium funiculosum* 0,15 0,22 1,24 60,0 *Penicillium ochrochloron* 0,06 0,04 0,29 0,27 *Scopulariopsis brevicaulis* 0,06 0,10 0,47 1,17 *Trichoderma viride* 0,02 0,04 0,18 0,34 Mixed fungal population 0,11 0,15 1,22 58,0

It was found that pure LDPE and the blends 85/15 and 70/30 showed little loss of mass, which probably reflected the inertness of LDPE towards biological degradation. Unfortunately, even 30% Bionolle® wasn't enough to observe sufficient weight loss of film. Probably, the polyethylene matrix prevented microbes from accessing the polyester in the depth of the film. Similar relationships were observed for the polyolefins modified with 6- 15% starch content, where only the surface of the material was susceptible to biodegradation (Nakamura et al., 2005). In a separate study, Rosa et al. found that the pure LDPE and the blends 25PHB/75LDPE and 50PHB/50LDPE showed little or no loss of mass during aging in simulated soil (Rosa et al., 2007). Also Lee et al. discovered that polystyrene (PS) in the P(3HB-co-3HV)/PS (95/5 by wt%) blend acts as a retardant of enzymatic attack to the

This phenomenon is likely due to the high-molecular weight hydrophobic chains of the synthetic polymer preventing enzymes from accessing the biodegradable polymers

Among polymeric compositions, film containing 60% Bionolle® had the most obvious reduction in weight. Film lost 24%, 60% and 58% of its initial mass after incubation with *Aspergillus niger*, *Penicillium funiculosum* and mixed population of fungi, respectively. Other fungi caused slight film weight loss ranging from 0,25% to 1,17%. Taken together with our previous studies, the present findings show, that only fungi which were able to decompose Bionolle® (80-100% for 90 days), were also able to degrade 40/60 composition (Nowak et al.,

Mechanical properties of the investigated films before degradation are shown in Table 2.

LDPE/Bionolle® film compositions 100/0 85/15 70/30 40/60 Weight loss, %

**3.1 Biodegradation** 

Nowak et al., 2010).

Filamentous fungi

Table 1. Weight loss of films after biodegradation

surface of the blend film (Lee et al., 2003).

2010).

contained within the material (Nowak et al., 2011).


Table 2. Mechanical properties of control films

The changes in tensile strength and elongation at break of films after biodegradation with selected fungi are shown in Figure 1. It has been reported that the changes in tensile strength and elongation loss are excellent indicators of degradation (Reddy et al., 2008).

Fig. 1. Mechanical properties of films after biodegradation

Although little weight loss of polyethylene was observed after 84 days of biodegradation, a marked reduction in tensile strength imply that both *Penicillium funiculosum* and mixed fungi population excrete enzymes able to cleave macromolecules of LDPE. Slight increase in the elongation at break in samples incubated with mixed population of fungi, could be tentatively attributed to a sort of plasticisation effect exerted by low molecular weight fractions produced in the first stage of the biodegradation of the polymer matrix (Chiellini et al., 2003). After 84 days of biodegradation, the tensile strength of film containing 30% Bionolle® decreased by about 20%. At this time, elongation at break changed by about 74% and 52% after incubation with *Penicillium funiculosum* and mixed fungi population, respectively. Far-advanced decomposition of 40/60 blend, prevented determination of its mechanical properties.

Data obtained from FTIR spectra (spectra are presented elsewhere (Łabużek et al.) of examined films showed an increase in carbonyl and double bond indices, except for LDPE (decrease in carbonyl index by 57%), incubated with mixed population of fungi (Łabużek et al., 2006a). Increase in the internal double bond index is in accordance with the biodegradation mechanism suggested by Albertsson et al. who reported on the formation of terminal double bonds as a result of exposure to biotic environment (Albertsson et al., 1987). This can be attributed to biotic dehydrogenation (Chiellini et al., 2003). However, in contrast

Biodegradation of Pre-Aged Modified Polyethylene Films 653

experiments were conducted in minimal solid medium, it is obvious that solid surface of

b)

 Fig. 3. SEM micrographs of neat LDPE film after biodegradation with a) *Aspergillus niger*

As a result of fungal degradation, peeling and cracking in texture of film containing 15% Bionolle® were visible (Figure 4). The entire surface of film was densely covered with spores belonging to *Aspergillus niger* (Figure 4a) and *Aureobasidium pullulans* (Figure 4b). Scarce hyphae of *Paecilomyces varioti* (Figure 4c), *Penicillium funiculosum* (Figure 4d) and mixed population of fungi (Figure 4f) colonised both the edges and the surface of film. Agglomerations of *Trichoderma viride* conidiophores (Figure 4e) inhabited primarily the

d)

The destructive process of biodegradation was very prominent in film containing 30% Bionolle®. Small holes and cracks, surface irregularities, peeling and exfoliation appeared. *Aspergillus niger* (Figure 5a), *Paecilomyces varioti* (Figure 5c), *Penicillium funiculosum* (Figure 5d) and mixed population of fungi (Figure 5h) produced a well-developed mycelium over the entire surface of the film. Hyphae and conidiophores of *Penicillium ochrochloron* (Figure 5e), *Scopulariopsis brevicaulis* (Figure 5f) and *Trichoderma viride* (Figure 5g) was scattered on the film surface. *Aspergillus terreus* (Figure 5b), almost unable to colonise 85/15 composition, introduced deep cracks and holes, suggesting that the fungi penetrate into the sample

After 84-day incubation with filamentous fungi the most intense changes were found on the surface of 40/60 composition. Hyphae and conidiophores of *Aspergillus niger* (Figure 6a), *Aspergillus terreus* (Figure 6b), *Aureobasidium pullulans* (Figure 6c) and *Trichoderma viride*

b) *Aureobasidium pullulans* c) *Paecilomyces varioti* d) *Penicillium funiculosum*

edges of the sample.

a)

c)

matrix during degradation.

LDPE, at least for some fungi, served as the source of carbon and energy.

with study of Albertsson et al., we found an increase in the amount of carbonyl residues (up to 525%) in LDPE after 84 days of incubation with *Penicillium funiculosum* (Albertsson et al., 1987). Carbonyl residues have been reported as major products formed in the presence oxidoreductases (Sudhakar et al., 2008).

Important aspect during the biodegradation of a material is the sustained growth of microorganisms during the entire process (Abrusci et al., 2011). The changes on the surface of polymers as a result of biodegradation are no less important (Nowak et al., 2011).

Figure 2 shows the micrographs of control films.

Fig. 2. SEM micrographs showing surface of control films a) LDPE b) composition 85/15 c) composition 70/30 d) composition 40/60

The photomicrograph shows that the surface of non-degraded material is smooth, without cracks and holes.

Neat polymer samples after biodegradation with filamentous fungi are presented in Figures 3-6.

*Paecilomyces varioti* (Figure 3c) and *Penicillium funiculosum* (Figure 3d) expanded their colonies over the entire surface of neat LDPE. Apart from hyphae and conidiophores, samples were covered with characteristic spores. *Aspergillus niger* (Figure 3a), *Aspergillus terreus*, *Aureobasidium pullulans* (Figure 3b), *Penicillium ochrochloron*, *Scopulariopsis brevicaulis*, *Trichoderma viride* and mixed fungal population grew less rapidly and primarily on the edges of the sample. Considerable change in mechanical properties (Figure 1) and FTIR spectrum of LDPE proved that growth of fungi cannot be considered only as a result of the surface moistness (Sahebnazar et al., 2010). Moreover, knowing that the biodegradation

with study of Albertsson et al., we found an increase in the amount of carbonyl residues (up to 525%) in LDPE after 84 days of incubation with *Penicillium funiculosum* (Albertsson et al., 1987). Carbonyl residues have been reported as major products formed in the presence

Important aspect during the biodegradation of a material is the sustained growth of microorganisms during the entire process (Abrusci et al., 2011). The changes on the surface

of polymers as a result of biodegradation are no less important (Nowak et al., 2011).

b)

 Fig. 2. SEM micrographs showing surface of control films a) LDPE b) composition 85/15 c)

The photomicrograph shows that the surface of non-degraded material is smooth, without

d)

Neat polymer samples after biodegradation with filamentous fungi are presented in Figures

*Paecilomyces varioti* (Figure 3c) and *Penicillium funiculosum* (Figure 3d) expanded their colonies over the entire surface of neat LDPE. Apart from hyphae and conidiophores, samples were covered with characteristic spores. *Aspergillus niger* (Figure 3a), *Aspergillus terreus*, *Aureobasidium pullulans* (Figure 3b), *Penicillium ochrochloron*, *Scopulariopsis brevicaulis*, *Trichoderma viride* and mixed fungal population grew less rapidly and primarily on the edges of the sample. Considerable change in mechanical properties (Figure 1) and FTIR spectrum of LDPE proved that growth of fungi cannot be considered only as a result of the surface moistness (Sahebnazar et al., 2010). Moreover, knowing that the biodegradation

oxidoreductases (Sudhakar et al., 2008).

composition 70/30 d) composition 40/60

cracks and holes.

3-6.

c)

a)

Figure 2 shows the micrographs of control films.

experiments were conducted in minimal solid medium, it is obvious that solid surface of LDPE, at least for some fungi, served as the source of carbon and energy.

Fig. 3. SEM micrographs of neat LDPE film after biodegradation with a) *Aspergillus niger* b) *Aureobasidium pullulans* c) *Paecilomyces varioti* d) *Penicillium funiculosum*

As a result of fungal degradation, peeling and cracking in texture of film containing 15% Bionolle® were visible (Figure 4). The entire surface of film was densely covered with spores belonging to *Aspergillus niger* (Figure 4a) and *Aureobasidium pullulans* (Figure 4b). Scarce hyphae of *Paecilomyces varioti* (Figure 4c), *Penicillium funiculosum* (Figure 4d) and mixed population of fungi (Figure 4f) colonised both the edges and the surface of film. Agglomerations of *Trichoderma viride* conidiophores (Figure 4e) inhabited primarily the edges of the sample.

The destructive process of biodegradation was very prominent in film containing 30% Bionolle®. Small holes and cracks, surface irregularities, peeling and exfoliation appeared. *Aspergillus niger* (Figure 5a), *Paecilomyces varioti* (Figure 5c), *Penicillium funiculosum* (Figure 5d) and mixed population of fungi (Figure 5h) produced a well-developed mycelium over the entire surface of the film. Hyphae and conidiophores of *Penicillium ochrochloron* (Figure 5e), *Scopulariopsis brevicaulis* (Figure 5f) and *Trichoderma viride* (Figure 5g) was scattered on the film surface. *Aspergillus terreus* (Figure 5b), almost unable to colonise 85/15 composition, introduced deep cracks and holes, suggesting that the fungi penetrate into the sample matrix during degradation.

After 84-day incubation with filamentous fungi the most intense changes were found on the surface of 40/60 composition. Hyphae and conidiophores of *Aspergillus niger* (Figure 6a), *Aspergillus terreus* (Figure 6b), *Aureobasidium pullulans* (Figure 6c) and *Trichoderma viride*

Biodegradation of Pre-Aged Modified Polyethylene Films 655

b)

d)

e) f)

g) h)

Fig. 5. SEM micrographs of neat 70/30 composition after biodegradation with a) *Aspergillus niger* b) *Aspergillus terreus* c) *Paecilomyces varioti* d) *Penicillium funiculosum* e) *Penicillium ochrochloron* f) *Scopulariopsis brevicaulis* g) *Trichoderma viride* h) mixed fungal

population

a)

c)

(Figure 6g) grew out directly from the polymer sample. However, changes induced by the action of these microorganisms deep and distinct. Dense network of fractures was particularly visible after incubation with *Aspergillus terreus*. *Aspergillus niger* and *Aureobasidium pullulans* caused massive exfoliation of the plastic edges. Although *Penicillium ochrochloron* (Figure 6e) and *Scopulariopsis brevicaulis* (Figure 6f) created mycelium, a considerable part of the film surface was found to be unaffected. Loss of integrity of film resulted into fragile surface entirely covered by dense mycelia of *Penicillium funiculosum* (Figure 6d) and mixed population of fungi (Figure 6h).

Fig. 4. SEM micrographs of neat 85/15 composition after biodegradation with a) *Aspergillus niger* b) *Aureobasidium pullulans* c) *Paecilomyces varioti* d) *Penicillium funiculosum* e) *Trichoderma viride* f) mixed fungal population

(Figure 6g) grew out directly from the polymer sample. However, changes induced by the action of these microorganisms deep and distinct. Dense network of fractures was particularly visible after incubation with *Aspergillus terreus*. *Aspergillus niger* and *Aureobasidium pullulans* caused massive exfoliation of the plastic edges. Although *Penicillium ochrochloron* (Figure 6e) and *Scopulariopsis brevicaulis* (Figure 6f) created mycelium, a considerable part of the film surface was found to be unaffected. Loss of integrity of film resulted into fragile surface entirely covered by dense mycelia of *Penicillium funiculosum*

b)

d)

*niger* b) *Aureobasidium pullulans* c) *Paecilomyces varioti* d) *Penicillium funiculosum*

e) *Trichoderma viride* f) mixed fungal population

e) f)

Fig. 4. SEM micrographs of neat 85/15 composition after biodegradation with a) *Aspergillus* 

(Figure 6d) and mixed population of fungi (Figure 6h).

a)

c)

Fig. 5. SEM micrographs of neat 70/30 composition after biodegradation with a) *Aspergillus niger* b) *Aspergillus terreus* c) *Paecilomyces varioti* d) *Penicillium funiculosum* e) *Penicillium ochrochloron* f) *Scopulariopsis brevicaulis* g) *Trichoderma viride* h) mixed fungal population

Biodegradation of Pre-Aged Modified Polyethylene Films 657

Weight variations of LDPE film and LDPE/Bionolle® compositions recorded after exposing

Photodegradation 0,06 0,05 0,07 0,06 Thermodegradation 0,02 0,03 0,13 0,1 Photo- and thermodegradation 0,03 0,04 0,09 0,26 Hydrolysis 0 0,04 0,07 0,38 Photodegradation and hydrolysis 0,02 0,05 0,07 0,27 Thermodegradation and hydrolysis 0,02 0,03 0,08 0,34 Photo- thermodegradation and hydrolysis 0 0,04 0,09 0,31

Regardless of the type and combination of abiotic aging factors, there were no significant differences in weight loss observed for LDPE and its compositions containing up to 30% Bionolle®. More significant decrease of the weight in film containing 60% polyester was recorded not till then it was subjected to hydrolysis or when more than one aging factor was used during degradation experiments. Oxidative aging of films did not accelerate their loss of mass, most likely due to the presence of crosslinks evolved under the action of radiation and/or heat (Ojeda et al., 2011). It is often reported that, in LDPE films, crosslinking competes with the chain scission mechanism depending on the oxygen concentration at the

Changes in some mechanical properties of films after abiotic degradation are presented in

Values obtained by measuring elongation at break of polyethylene after photodegradation, thermodegradation and photothermodegradation slightly increased. As it was reported above, due to the formation of cross-linking bonds between the polyethylene chains. However, reduction in the mechanical properties after hydrolysis was observed. This decrease can be attributed to the chain scission of the polymer which, in this case, most intensively proceeded in films exposed earlier to both UV radiation and heat. This macromolecular chain scission is the cause of embrittlement of films (Abrusci et al., 2011). Amongst modified films, sensitivity toward abiotic treatment in terms of loss in the mechanical properties can be arranged as follows: 40/60>85/15>70/30. It seems that the exposure of the films containing 15% and 30% polyester to abiotic oxidation has accelerated their subsequent hydrolysis. On the contrary, in film consisted in most part of polyester, the most significant decrease in tensile strength by about 81% and elongation at break reduced by 99% was observed in samples subjected only to hydrolysis. From the above results it can be concluded that the samples with low content of Bionolle® behaved more like polyethylene while 40/60 composition more like polyester. It is known, that in hydrolytic degradation of biodegradable polyesters, elongation at break is the most sensitive property

LDPE/Bionolle® film compositions 100/0 85/15 70/30 40/60 Weight loss, %

**3.2 Abiotic degradation** 

to abiotic treatment are shown in Table 3.

Table 3. Weight loss of films after abiotic aging

reaction site (Feuilloley et al., 2005).

among the tensile properties (Tsuji et al., 2006).

Figure 7.

**Abiotic degradation processes** 

Fig. 6. SEM micrographs of neat 40/60 composition after biodegradation with a) *Aspergillus niger* b) *Aspergillus terreus* c) *Aureobasidium pullulans* d) *Penicillium funiculosum* e) *Penicillium ochrochloron* f) *Scopulariopsis brevicaulis* g) *Trichoderma viride* h) mixed fungal population

## **3.2 Abiotic degradation**

656 Scanning Electron Microscopy

b)

d)

e) f)

g) h)

*Aspergillus niger* b) *Aspergillus terreus* c) *Aureobasidium pullulans* d) *Penicillium funiculosum* e) *Penicillium ochrochloron* f) *Scopulariopsis brevicaulis* g) *Trichoderma viride* h) mixed fungal

Fig. 6. SEM micrographs of neat 40/60 composition after biodegradation with a)

population

a)

c)

Weight variations of LDPE film and LDPE/Bionolle® compositions recorded after exposing to abiotic treatment are shown in Table 3.


Table 3. Weight loss of films after abiotic aging

Regardless of the type and combination of abiotic aging factors, there were no significant differences in weight loss observed for LDPE and its compositions containing up to 30% Bionolle®. More significant decrease of the weight in film containing 60% polyester was recorded not till then it was subjected to hydrolysis or when more than one aging factor was used during degradation experiments. Oxidative aging of films did not accelerate their loss of mass, most likely due to the presence of crosslinks evolved under the action of radiation and/or heat (Ojeda et al., 2011). It is often reported that, in LDPE films, crosslinking competes with the chain scission mechanism depending on the oxygen concentration at the reaction site (Feuilloley et al., 2005).

Changes in some mechanical properties of films after abiotic degradation are presented in Figure 7.

Values obtained by measuring elongation at break of polyethylene after photodegradation, thermodegradation and photothermodegradation slightly increased. As it was reported above, due to the formation of cross-linking bonds between the polyethylene chains. However, reduction in the mechanical properties after hydrolysis was observed. This decrease can be attributed to the chain scission of the polymer which, in this case, most intensively proceeded in films exposed earlier to both UV radiation and heat. This macromolecular chain scission is the cause of embrittlement of films (Abrusci et al., 2011). Amongst modified films, sensitivity toward abiotic treatment in terms of loss in the mechanical properties can be arranged as follows: 40/60>85/15>70/30. It seems that the exposure of the films containing 15% and 30% polyester to abiotic oxidation has accelerated their subsequent hydrolysis. On the contrary, in film consisted in most part of polyester, the most significant decrease in tensile strength by about 81% and elongation at break reduced by 99% was observed in samples subjected only to hydrolysis. From the above results it can be concluded that the samples with low content of Bionolle® behaved more like polyethylene while 40/60 composition more like polyester. It is known, that in hydrolytic degradation of biodegradable polyesters, elongation at break is the most sensitive property among the tensile properties (Tsuji et al., 2006).

Biodegradation of Pre-Aged Modified Polyethylene Films 659

Fig. 8. FTIR spectra of films before and after degradation with selected abiotic and biotic

factors a) LDPE b) 70/30 composition c) 40/60 composition

Fig. 7. Changes in some mechanical properties of films after abiotic degradation A) and B) LDPE C) and D) 85/15 blend E) and F) 70/30 blend G) and H) 40/60 blend

Figure 8 shows FTIR spectra of samples before and after exposing to selected abiotic and biotic factors.

Fig. 7. Changes in some mechanical properties of films after abiotic degradation A) and

Figure 8 shows FTIR spectra of samples before and after exposing to selected abiotic and

B) LDPE C) and D) 85/15 blend E) and F) 70/30 blend G) and H) 40/60 blend

biotic factors.

Fig. 8. FTIR spectra of films before and after degradation with selected abiotic and biotic factors a) LDPE b) 70/30 composition c) 40/60 composition

Biodegradation of Pre-Aged Modified Polyethylene Films 661

Compared to the smooth control film (Figure 2), only some irregularities were visible on the surface of 70/30 composition exposed to UV radiation (Figure 9a). After thermodegradation numerous and well distributed oval cavities with a diameter 1-8 mm were seen all over the surface (Figure 9b). Observations of changes resulting from the individual oxidative processes (photo- and thermodegradation) were helpful in interpreting the data obtained after simultaneous action of UV radiation and heat (Figure 9c). It is evident that both processes act antagonistically. This observation confirms the results obtained by other methods. The smallest weight loss and negligible increase in carbonyl index, probably resulted from the fact that sites of potential chain oxidation (macro radicals) were involved in the crosslinks formation (H-S. Kim & H-J Kim, 2008). Hence, observed reinforcement of plastics after photothermodegradation. However, the most pronounced changes were observed after hydrolysis of the film (Figure 9d). Holes on the surface were less in number but bigger in size than cavities observed after thermodegradation. Studies conducted by other researchers suggest that hydrolysis, in contrast to the photodegradation, occured mainly in the amorphous region of polymer (Bikiaris et al., 2006; Tsuji et al., 2006). Judging by the size, shape and distribution of holes, identical to that observed after thermodegradation, it can be concluded, that during the heat-treatment, the amorphous phase breaks down in the first place. This is due to the fact that, under the impact of warmth, the mobility of macromolecules within amorphous region increases more

significantly, therefore they become more prone to degradation.

a)

c)

b)

There were no visible changes of film texture after photo- and photothermodegradation (Figure 10a and c). Again, after another analysis of 40/60 composition, it could be stated that the behaviour of the composition under the influence of different factors is affected mainly by a large quantity of polyester in polyethylene. These findings are in agreement with our previous study on photodegradation of PBSA (Łabużek et al., 2006b). Thermodegradation of

d)

Fig. 10. SEM micrographs of 40/60 composition after a) photodegradation

b) thermodegradation c) photothermodegradation d) hydrolysis

The data showed that the area corresponding to the carbonyl region has grown after abiotic degradation of all films indicating the formation of low molecular weight carbonyl compounds as a result of oxidation and/or hydrolysis. Carbonyl index of LDPE was increased by 25%(photothermodegradation)-650%(thermodegradation). As for polyethylene, the greatest impact on the oxidation of 70/30 composition had thermodegradation (230%) while the smallest - photothermodegradation (196%). Similar correlations were also observed for internal double bond index, which after thermodegradation of polyethylene and 70/30 composition increased by 167% and 316%, respectively. For film containing 60% Bionolle®, the largest increase in carbonyl index, amounting 50%, was found after hydrolysis. Such low degree of oxidation resulted probably from the fact that low-molecular fractions of polymer diffused out of the polymer matrix during the hydrolysis of film (Göpferich, 1996). The above findings suggest that single processes, especially thermo-oxidation, are more efficient in polymer degradation than simultaneous action of UV radiation and heat. These findings are in agreement with (Ram et al., 1980) who claims that the presence of oxygen in conjunction with high temperatures, plays more significant role in the increase of carbonyl concentration than when combined with UV exposure. Moreover, the air temperature as a critical factor increases the rate of various chemical reactions associated with degradation (Briassoulis et al., 2004). Additionally, as mentioned earlier, thermodegradation occurs throughout the bulk of polymer, not only on its surface.

Micrographs of films exposed to selected abiotic factors are presented in Figures 9 and 10.

Fig. 9. SEM micrographs of 70/30 composition after a) photodegradation b) thermodegradation c) photothermodegradation d) hydrolysis

The data showed that the area corresponding to the carbonyl region has grown after abiotic degradation of all films indicating the formation of low molecular weight carbonyl compounds as a result of oxidation and/or hydrolysis. Carbonyl index of LDPE was increased by 25%(photothermodegradation)-650%(thermodegradation). As for polyethylene, the greatest impact on the oxidation of 70/30 composition had thermodegradation (230%) while the smallest - photothermodegradation (196%). Similar correlations were also observed for internal double bond index, which after thermodegradation of polyethylene and 70/30 composition increased by 167% and 316%, respectively. For film containing 60% Bionolle®, the largest increase in carbonyl index, amounting 50%, was found after hydrolysis. Such low degree of oxidation resulted probably from the fact that low-molecular fractions of polymer diffused out of the polymer matrix during the hydrolysis of film (Göpferich, 1996). The above findings suggest that single processes, especially thermo-oxidation, are more efficient in polymer degradation than simultaneous action of UV radiation and heat. These findings are in agreement with (Ram et al., 1980) who claims that the presence of oxygen in conjunction with high temperatures, plays more significant role in the increase of carbonyl concentration than when combined with UV exposure. Moreover, the air temperature as a critical factor increases the rate of various chemical reactions associated with degradation (Briassoulis et al., 2004). Additionally, as mentioned earlier, thermodegradation occurs throughout the bulk of

Micrographs of films exposed to selected abiotic factors are presented in Figures 9 and 10.

b)

d)

Fig. 9. SEM micrographs of 70/30 composition after a) photodegradation

b) thermodegradation c) photothermodegradation d) hydrolysis

polymer, not only on its surface.

a)

c)

Compared to the smooth control film (Figure 2), only some irregularities were visible on the surface of 70/30 composition exposed to UV radiation (Figure 9a). After thermodegradation numerous and well distributed oval cavities with a diameter 1-8 mm were seen all over the surface (Figure 9b). Observations of changes resulting from the individual oxidative processes (photo- and thermodegradation) were helpful in interpreting the data obtained after simultaneous action of UV radiation and heat (Figure 9c). It is evident that both processes act antagonistically. This observation confirms the results obtained by other methods. The smallest weight loss and negligible increase in carbonyl index, probably resulted from the fact that sites of potential chain oxidation (macro radicals) were involved in the crosslinks formation (H-S. Kim & H-J Kim, 2008). Hence, observed reinforcement of plastics after photothermodegradation. However, the most pronounced changes were observed after hydrolysis of the film (Figure 9d). Holes on the surface were less in number but bigger in size than cavities observed after thermodegradation. Studies conducted by other researchers suggest that hydrolysis, in contrast to the photodegradation, occured mainly in the amorphous region of polymer (Bikiaris et al., 2006; Tsuji et al., 2006). Judging by the size, shape and distribution of holes, identical to that observed after thermodegradation, it can be concluded, that during the heat-treatment, the amorphous phase breaks down in the first place. This is due to the fact that, under the impact of warmth, the mobility of macromolecules within amorphous region increases more significantly, therefore they become more prone to degradation.

Fig. 10. SEM micrographs of 40/60 composition after a) photodegradation b) thermodegradation c) photothermodegradation d) hydrolysis

There were no visible changes of film texture after photo- and photothermodegradation (Figure 10a and c). Again, after another analysis of 40/60 composition, it could be stated that the behaviour of the composition under the influence of different factors is affected mainly by a large quantity of polyester in polyethylene. These findings are in agreement with our previous study on photodegradation of PBSA (Łabużek et al., 2006b). Thermodegradation of

Biodegradation of Pre-Aged Modified Polyethylene Films 663

Changes in some mechanical properties of film samples after the abiotic degradation and

A B

C D

E F

 Photodegradation and biodegradation Thermodegradation and biodegradation Photothermodegradation and biodegradation Hydrolysis and biodegradation

Fig. 11. Changes in some mechanical properties of films after abiotic and biotic degradation

A significant increase in elongation of LDPE after thermo-, photo-thermo- and biodegradation with both *Penicillium funiculosum* and mixed populations of fungi was noted. These values were significantly higher than observed after biodegradation of neat polyethylene. The highest impact of abiotic pretreatment on the subsequent biodegradability was observed for 85/15 composition which elongation decreased by 9,26%- 16,82% as compared to 3,09%-8,49% for the film subjected only to biodegradation. As seen from Figure 11 there is a difference concerning the influence of abiotic aging to biodegradation of modified films between mixed fungi population and *Penicillium* 

A) and B) LDPE C) and D) 85/15 blend E) and F) 70/30 blend.

Control *Penicillium funiculosum* Mixed fungal population

biodegradation are shown in Figure 11.

film containing 60% Bionolle® resulted in the formation of regular holes in small quantities (Figure 10b). The most significant surface fragmentation was observed after hydrolysis of the film (Figure 10d). As is evident from SEM, amorphous regions are preferably hydrolysed, revealing the crystalline ones which in turn affects the mechanical properties. It is postulated that mechanical properties, especially elongation at break (see Figure 7G and H), depend more on changes occurring in the amorphous phase (Briassoulis et al., 2004).

## **3.3 Influence of abiotic degradation on the rate of biodegradation**

*Penicillium funiculosum* and mixed population of fungi were used in studies on biodegradation of abiotically aged films, since, in previous stage of experiments, they exhibited the greatest ability to degrade examined plastics.

Table 4 shows the percentage weight loss of pre-aged LDPE film and LDPE/Bionolle® compositions after biodegradation with *Penicillium funiculosum* and mixed fungal population.


Table 4. Weight loss of pre-aged films after biodegradation

Based on material weight loss it cannot be clearly determined what impact have abiotic processes had on subsequent biodegradation of the material. Contrary to LDPE, photodegradation, thermodegradation and photothermodegradation of modified polyethylene films accelerated their biodegradation. Percentage weight loss of LDPE films modified with 30% Bionolle® after biodegradation with *Penicillium funiculosum* and mixed population of fungi increased by 15% and 13%; 19% and 28%; 17% and 4% in comparison to weight loss of neat films, respectively. The biodegradative propensity of abiotically degraded 40/60 composition was also markedly affected. The pre-treated film after being exposed to *Penicillium funiculosum* and mixed population of fungi exhibit a mass loss of 15- 19% and 4-28% higher than observed for control film. However, the biggest mass loss was observed after hydrolytic aging and biodegradation with *Penicillium funiculosum*. Biodegradation of films containing 15%, 30% and 60% polyester after prior hydrolysis increased by 104.5%, 75.8% and 24%, respectively. Considering percentage weight loss after hydrolysis and biodegradation of 40/60 composition, we have to remark, that any amount greater than 60% meant the degradation of polyethylene.

film containing 60% Bionolle® resulted in the formation of regular holes in small quantities (Figure 10b). The most significant surface fragmentation was observed after hydrolysis of the film (Figure 10d). As is evident from SEM, amorphous regions are preferably hydrolysed, revealing the crystalline ones which in turn affects the mechanical properties. It is postulated that mechanical properties, especially elongation at break (see Figure 7G and H), depend more on changes occurring in the amorphous phase (Briassoulis et al., 2004).

*Penicillium funiculosum* and mixed population of fungi were used in studies on biodegradation of abiotically aged films, since, in previous stage of experiments, they

Table 4 shows the percentage weight loss of pre-aged LDPE film and LDPE/Bionolle® compositions after biodegradation with *Penicillium funiculosum* and mixed fungal

Based on material weight loss it cannot be clearly determined what impact have abiotic processes had on subsequent biodegradation of the material. Contrary to LDPE, photodegradation, thermodegradation and photothermodegradation of modified polyethylene films accelerated their biodegradation. Percentage weight loss of LDPE films modified with 30% Bionolle® after biodegradation with *Penicillium funiculosum* and mixed population of fungi increased by 15% and 13%; 19% and 28%; 17% and 4% in comparison to weight loss of neat films, respectively. The biodegradative propensity of abiotically degraded 40/60 composition was also markedly affected. The pre-treated film after being exposed to *Penicillium funiculosum* and mixed population of fungi exhibit a mass loss of 15- 19% and 4-28% higher than observed for control film. However, the biggest mass loss was observed after hydrolytic aging and biodegradation with *Penicillium funiculosum*. Biodegradation of films containing 15%, 30% and 60% polyester after prior hydrolysis increased by 104.5%, 75.8% and 24%, respectively. Considering percentage weight loss after hydrolysis and biodegradation of 40/60 composition, we have to remark, that any amount

LDPE/Bionolle® film compositions 100/0 85/15 70/30 40/60 Weight loss, %

*P. funiculosum* 0,13 0,26 1,43 69,19 Mixed fungal population 0,07 0,18 1,38 65,61

*P. funiculosum* 0,28 0,28 1,48 71,61 Mixed fungal population 0,04 0,23 1,56 74,14

*P. funiculosum* 0,08 0,14 1,45 70,16

*P. funiculosum* 0,17 0,45 2,18 74,62 Mixed fungal population 0,13 0,32 1,86 59,32

Mixed fungal population 0,12 0,15 1,27 60,38

**3.3 Influence of abiotic degradation on the rate of biodegradation** 

exhibited the greatest ability to degrade examined plastics.

Table 4. Weight loss of pre-aged films after biodegradation

greater than 60% meant the degradation of polyethylene.

Process Fungi

population.

Photodegradation and biodegradation

Thermodegradation and

thermodegradation and

biodegradation

biodegradation

Hydrolysis and biodegradation

Photo- and

Changes in some mechanical properties of film samples after the abiotic degradation and biodegradation are shown in Figure 11.

Fig. 11. Changes in some mechanical properties of films after abiotic and biotic degradation A) and B) LDPE C) and D) 85/15 blend E) and F) 70/30 blend.

A significant increase in elongation of LDPE after thermo-, photo-thermo- and biodegradation with both *Penicillium funiculosum* and mixed populations of fungi was noted. These values were significantly higher than observed after biodegradation of neat polyethylene. The highest impact of abiotic pretreatment on the subsequent biodegradability was observed for 85/15 composition which elongation decreased by 9,26%- 16,82% as compared to 3,09%-8,49% for the film subjected only to biodegradation. As seen from Figure 11 there is a difference concerning the influence of abiotic aging to biodegradation of modified films between mixed fungi population and *Penicillium* 

Biodegradation of Pre-Aged Modified Polyethylene Films 665

b)

 Fig. 12. SEM micrographs of films after exposing to different abiotic factors and subsequent biodegradation with mixed fungal population a) LDPE after thermo- and biodegradation b) 85/15 composition after photo- and biodegradation c) 70/30 composition after photothermo- and biodegradation d) 40/60 composition after hydrolysis and biodegradation

d)

Massive erosion (holes with diameter about 200 mm) of film 70/30 and dense network of fungal hyphae indicated that, after abiotic degradation, surface of film become at least as

It was impossible to separate fungal hyphae from the residual 40/60 composition after 84 days of hydrolysis and subsequent biodegradation. As revealed earlier (Table 4), weight loss of film exposed to both factors increased slightly compared to film subjected only to biodegradation. In order to describe the possible mechanism of degradation of this material, it is essential that a few facts should be given. Firstly, hydrolytic degradation (abiotic or biotic) of the polymer occurs predominantly in the amorphous regions (H-S. Kim & H-J Kim, 2008) Secondly, hydrolysis of the crystalline material is slow, because of the limited water diffusion rates into the crystalline domains and stereochemical limitations (Bikiaris et al., 2006). In our study, after inoculation of the composition with fungi, developing hyphae at first assimilated low-molecular products of polymer hydrolysis then attacked its crystalline region (Tserki et al., 2006)). Moreover, by taking into account the weight loss of the composition (59-75%), it can be assumed that fungi (long before the end of the experiment) completely assimilated polyester. Remaining LDPE fibres are clearly visible in the micrographs. Hence, decrease in the rate of biodegradation could be the result of (i) slow biodegradation of crystalline phase of polyester (ii) complex biodegradation of LDPE and

available to microorganism as unaged 40/60 blend.

a)

c)

(iii) penetration of fungi into the depths of the polymer matrix.

*funiculosum*. Contrary to mixed fungal population, assimilation of products of polymer oxidation during incubation with *Penicillium funiculosum* occurred less efficiently than depolymerisation of long LDPE chains. Far-advanced decomposition of 40/60 blend, prevented determination of its mechanical properties.

The amount of carbonyl groups, resulting from thermodegradation, decreased by 83% as a consequence of the assimilation of the degradation products by *Penicillium funiculosum*. Similar mechanism of biodegradation of abiotically aged polymers is repeatedly reported. Also Albertsson et al. (1987) found a synergistic effect between photooxidation and the biodegradation of polyethylene. Carbonyl residues completely disappeared after the incubation of the polymer samples in the presence of *Arthrobacter paraffineus*. Decrease of carbonyl index by 30-35% with respect to the starting materials was described for LDPE film samples containing pro-oxidant additives exposed to thermal- and biological degradation (Chiellini et al., 2007). The spectroscopic investigations led by Roy et al. revealed that the bacterial consortium consisted of *Bacillus pumilus*, *Bacillus halodenitrificans* and *Bacillus cereus* preferentially consumed the oxygenated products, thus leading to a decrease in the carbonyl index from 1,29 to 0,31 (Roy et al., 2008). In our study, internal double bonds also proved to be equally easily digested, since their amount after thermo- and biodegradation decreased by 92%. Interestingly, our study also shown that the number of carbonyl residues was about 80% lower than that observed during the biodegradation alone. The amount of carbonyl groups in the film with 30% content of polyester after thermo- and biodegradation compared to control film increased by 96%, while in relation to the film after thermodegradation decreased by 41%. As shown on Figure 8 carbonyl index of aged 40/60 composition after hydrolysis and subsequent biodegradation was lower by 40%, 93% and 60% than the value obtained for the control, after biodegradation and after hydrolysis, respectively.

These results implies that the filamentous fungi having at their disposal oxidised degradation products of polyethylene become less effective in degradation of macromolecules. Decrease in both indices (carbonyl and internal double bond) is a clear evidence that microorganisms use other set of enzymes during biodegradation of aged films than when they grow on high-molecular neat LDPE and polyester. Another conclusion is that the main enzymes involved in degradation of polymers are not constitutive proteins, expressed and secreted by microorganism independently of the substrate. On the contrary, the difference in mechanism of biodegradation clearly indicates the participation of inducible enzymes expressed only under specific conditions.

Micrographs of films exposed to selected abiotic factors and subsequent biodegradation are presented in Figure 12.

Compared to the film exposed only to biodegradation, observations of LDPE after thermoand biodegradation revealed filamentous fungi growing over the entire surface of the film (Figure 12a). Rough, peeling surface of the material was seen at higher magnification. It is likely that the fungi inhabiting film, used the degradation products of LDPE as a carbon and energy source. This surveillance was supported by a decrease of carbonyl index (Figure 8) and elongation at break (Figure 11) indicating fungal assimilation of low-molecular weight fractions.

SEM micrographs of 85/15 composition revealed deep cracks and holes with diameter 5-50 mm. The cavities on the surface suggested that microorganisms penetrated the polymer matrix during the degradation process.

*funiculosum*. Contrary to mixed fungal population, assimilation of products of polymer oxidation during incubation with *Penicillium funiculosum* occurred less efficiently than depolymerisation of long LDPE chains. Far-advanced decomposition of 40/60 blend,

The amount of carbonyl groups, resulting from thermodegradation, decreased by 83% as a consequence of the assimilation of the degradation products by *Penicillium funiculosum*. Similar mechanism of biodegradation of abiotically aged polymers is repeatedly reported. Also Albertsson et al. (1987) found a synergistic effect between photooxidation and the biodegradation of polyethylene. Carbonyl residues completely disappeared after the incubation of the polymer samples in the presence of *Arthrobacter paraffineus*. Decrease of carbonyl index by 30-35% with respect to the starting materials was described for LDPE film samples containing pro-oxidant additives exposed to thermal- and biological degradation (Chiellini et al., 2007). The spectroscopic investigations led by Roy et al. revealed that the bacterial consortium consisted of *Bacillus pumilus*, *Bacillus halodenitrificans* and *Bacillus cereus* preferentially consumed the oxygenated products, thus leading to a decrease in the carbonyl index from 1,29 to 0,31 (Roy et al., 2008). In our study, internal double bonds also proved to be equally easily digested, since their amount after thermo- and biodegradation decreased by 92%. Interestingly, our study also shown that the number of carbonyl residues was about 80% lower than that observed during the biodegradation alone. The amount of carbonyl groups in the film with 30% content of polyester after thermo- and biodegradation compared to control film increased by 96%, while in relation to the film after thermodegradation decreased by 41%. As shown on Figure 8 carbonyl index of aged 40/60 composition after hydrolysis and subsequent biodegradation was lower by 40%, 93% and 60% than the value obtained for the

These results implies that the filamentous fungi having at their disposal oxidised degradation products of polyethylene become less effective in degradation of macromolecules. Decrease in both indices (carbonyl and internal double bond) is a clear evidence that microorganisms use other set of enzymes during biodegradation of aged films than when they grow on high-molecular neat LDPE and polyester. Another conclusion is that the main enzymes involved in degradation of polymers are not constitutive proteins, expressed and secreted by microorganism independently of the substrate. On the contrary, the difference in mechanism of biodegradation clearly indicates the participation of

Micrographs of films exposed to selected abiotic factors and subsequent biodegradation are

Compared to the film exposed only to biodegradation, observations of LDPE after thermoand biodegradation revealed filamentous fungi growing over the entire surface of the film (Figure 12a). Rough, peeling surface of the material was seen at higher magnification. It is likely that the fungi inhabiting film, used the degradation products of LDPE as a carbon and energy source. This surveillance was supported by a decrease of carbonyl index (Figure 8) and elongation at break (Figure 11) indicating fungal assimilation of low-molecular weight

SEM micrographs of 85/15 composition revealed deep cracks and holes with diameter 5-50 mm. The cavities on the surface suggested that microorganisms penetrated the polymer

prevented determination of its mechanical properties.

control, after biodegradation and after hydrolysis, respectively.

inducible enzymes expressed only under specific conditions.

presented in Figure 12.

matrix during the degradation process.

fractions.

Fig. 12. SEM micrographs of films after exposing to different abiotic factors and subsequent biodegradation with mixed fungal population a) LDPE after thermo- and biodegradation b) 85/15 composition after photo- and biodegradation c) 70/30 composition after photothermo- and biodegradation d) 40/60 composition after hydrolysis and biodegradation

Massive erosion (holes with diameter about 200 mm) of film 70/30 and dense network of fungal hyphae indicated that, after abiotic degradation, surface of film become at least as available to microorganism as unaged 40/60 blend.

It was impossible to separate fungal hyphae from the residual 40/60 composition after 84 days of hydrolysis and subsequent biodegradation. As revealed earlier (Table 4), weight loss of film exposed to both factors increased slightly compared to film subjected only to biodegradation. In order to describe the possible mechanism of degradation of this material, it is essential that a few facts should be given. Firstly, hydrolytic degradation (abiotic or biotic) of the polymer occurs predominantly in the amorphous regions (H-S. Kim & H-J Kim, 2008) Secondly, hydrolysis of the crystalline material is slow, because of the limited water diffusion rates into the crystalline domains and stereochemical limitations (Bikiaris et al., 2006). In our study, after inoculation of the composition with fungi, developing hyphae at first assimilated low-molecular products of polymer hydrolysis then attacked its crystalline region (Tserki et al., 2006)). Moreover, by taking into account the weight loss of the composition (59-75%), it can be assumed that fungi (long before the end of the experiment) completely assimilated polyester. Remaining LDPE fibres are clearly visible in the micrographs. Hence, decrease in the rate of biodegradation could be the result of (i) slow biodegradation of crystalline phase of polyester (ii) complex biodegradation of LDPE and (iii) penetration of fungi into the depths of the polymer matrix.

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biodegradation and mechanical properties of high-content starch/low-density

## **4. Conclusion**

In the environment plastics decompose under the influence of different abiotic and biotic factors. The abiotic factors such as radiation, temperature, humidity, chemical pollution and wind can act synergistically or antagonistically causing various types of structural and chemical changes in the polymer. Microorganisms, especially bacteria or fungi, play a crucial role in biological degradation of polymers.

Scanning electron microscopy (SEM) is an invaluable tool for polymer analysis, since it is extensively used to study changes in the texture and composition of biodegradable polymer materials exposed to various environmental factors. It allows for the exploration of large surfaces with excellent resolution of topographic features.

Especially the new generation of SEM technology, known as ultra-high resolution field emission SEM (UHR FE-SEM) presents a promising technique for polymer morphological characterization, and provides complementary data to other microscopic methods.

One of the possible ways to accelerate the degradation of so-called "stable polymers" in the environment is their blending with polymers containing ester, ether, carboxyl and hydroxyl groups that are susceptible to hydrolytic attack of microorganisms. The chapter describes studies on biodegradation of LDPE/Bionolle® blends. It was shown that the addition of polyester significantly accelerated biodegradation of material.

Another way to sensitize the polymer is to expose it to the abiotic aging (action of abiotic factors) followed by the action of microbes. Abiotically modified surface of plastics promotes growth of microorganisms thus accelerates biodegradation.

Indeed, examined films underwent rapid biodegradation, but the course of the process was significantly different from that seen earlier. Further investigation on the mechanism of biodegradation revealed that fungi secreted different sets of enzymes depending on molecular weight of substrate. Unlike some other researchers, we have shown that filamentous fungi used in our study, were capable of efficient oxidation and degradation of the high-molecular weight LDPE. However, when pre-degraded, low-molecular weight chains of polyethylene served as a source of carbon and energy, microorganisms assimilated only short oligomers resulted from prior abiotic degradation. They did not show strong oxidising activity.

In our opinion, LDPE/Bionolle® blends can be used in the production of environmentally degradable packagings. Particularly noteworthy is composition containing 60% Bionolle®, which decompose several dozen percent within 84 days.

## **5. References**


In the environment plastics decompose under the influence of different abiotic and biotic factors. The abiotic factors such as radiation, temperature, humidity, chemical pollution and wind can act synergistically or antagonistically causing various types of structural and chemical changes in the polymer. Microorganisms, especially bacteria or fungi, play a

Scanning electron microscopy (SEM) is an invaluable tool for polymer analysis, since it is extensively used to study changes in the texture and composition of biodegradable polymer materials exposed to various environmental factors. It allows for the exploration of large

Especially the new generation of SEM technology, known as ultra-high resolution field emission SEM (UHR FE-SEM) presents a promising technique for polymer morphological

One of the possible ways to accelerate the degradation of so-called "stable polymers" in the environment is their blending with polymers containing ester, ether, carboxyl and hydroxyl groups that are susceptible to hydrolytic attack of microorganisms. The chapter describes studies on biodegradation of LDPE/Bionolle® blends. It was shown that the addition of

Another way to sensitize the polymer is to expose it to the abiotic aging (action of abiotic factors) followed by the action of microbes. Abiotically modified surface of plastics

Indeed, examined films underwent rapid biodegradation, but the course of the process was significantly different from that seen earlier. Further investigation on the mechanism of biodegradation revealed that fungi secreted different sets of enzymes depending on molecular weight of substrate. Unlike some other researchers, we have shown that filamentous fungi used in our study, were capable of efficient oxidation and degradation of the high-molecular weight LDPE. However, when pre-degraded, low-molecular weight chains of polyethylene served as a source of carbon and energy, microorganisms assimilated only short oligomers resulted from prior abiotic degradation. They did not show strong

In our opinion, LDPE/Bionolle® blends can be used in the production of environmentally degradable packagings. Particularly noteworthy is composition containing 60% Bionolle®,

Abrusci, C.; Pablos, J.L.; Corrales, T.; López-Marín, J.; Marín, I. & Catalina, F. (2011).

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characterization, and provides complementary data to other microscopic methods.

**4. Conclusion** 

oxidising activity.

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surfaces with excellent resolution of topographic features.

polyester significantly accelerated biodegradation of material.

which decompose several dozen percent within 84 days.

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**33** 

*1Singapore 2,3India* 

**Surface Analysis Studies** 

**on Polymer Electrolyte Membranes** 

**and Atomic Force Microscope** 

M. Ulaganathan1,2,\*, R. Nithya3 and S. Rajendran1,\*

*3Dept. of Physics, Shanmuganathan Engineering College,* 

*Arasampatti, Pudukkottai, Tamil Nadu,* 

*2School of Physics, Alagappa University, Karaikudi, Tamil Nadu,* 

**Using Scanning Electron Microscope** 

*1Energy Research Institute @ NTU, Nanyang Technological University, Singapore* 

A battery generally consists of three important parts namely, anode, cathode and electrolyte. The batteries further classified into primary and secondary batteries. Among the different kinds of batteries, Li-ion batteries are plays a very important role in the development of modern technologies especially in the portable electronic device industries and in heavy electrical vehicles because of its advantages such as high theoretical capacity, improved safety, lower material costs, ease of fabrication into flexible geometries, and the absence of electrolyte leakage. In the battery system different kinds of electrolytes were used for promoting the ions from anode to cathode (during charge) and cathode to anode (during discharge). For this purpose, liquid electrolyte is identified as suitable electrolytes which facilitate higher ionic conductivity (10-2 Scm-1) than other electrolyte systems. However, it has several disadvantages namely gas formation during the operation, leakage, difficult to utilise for portable applications and etc. To overcome these difficulties, many attempts were made on solid polymer electrolyte systems. The main objective of the researchers is to improve the ambient temperature ionic conductivity, mechanical stability, thermal and interfacial stability of the electrolytes. However, it is difficult task for the researchers, inorder to improve these basic requirements of the electrolytes simultaneously because the ionic conductivity and mechanical strength of a polymer electrolyte are disparate to each

other, i.e., mechanical strength of the electrolyte decreases as conductivity increases.

In recent years, Polymer electrolytes have been attracted scientific and technological importance because of their potential applications in many areas such as Li-ion polymer batteries, super capacitor, electro chromic devices and etc. The idea of preparation of

**1. Introduction** 

 \*

Corresponding Authors


## **Surface Analysis Studies on Polymer Electrolyte Membranes Using Scanning Electron Microscope and Atomic Force Microscope**

M. Ulaganathan1,2,\*, R. Nithya3 and S. Rajendran1,\*

*1Energy Research Institute @ NTU, Nanyang Technological University, Singapore 2School of Physics, Alagappa University, Karaikudi, Tamil Nadu, 3Dept. of Physics, Shanmuganathan Engineering College, Arasampatti, Pudukkottai, Tamil Nadu, 1Singapore 2,3India* 

#### **1. Introduction**

670 Scanning Electron Microscopy

Sivan, A. (2011). New perspectives in plastic biodegradation*. Current Opinion in Biotechnology*, Vol.22, No.3, (June 2011), pp. 422-426, ISSN 0958-1669 Soni, R.; Kapri, A.; Zaidi, M.G.H. & Goel, R. (2009). Comparative Biodegradation Studies of

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Wojcieszyńska, D.; Hupert-Kocurek, K.; Greń, I. & Guzik, U. (2011). High activity catechol

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ISSN 1566-2543

1973-1976, ISSN 1471-2970

pp. 853-858, ISSN 0964-8305

(February 2006), pp.367-376, ISSN 0141-3910

Non-poronized and Poronized LDPE Using Indigenous Microbial Consortium. *Journal of Polymers and the Environment*, Vol.17, No.4, (December 2009), pp. 233-239,

biodegradation of low- and high-density polyethylenes. *International Biodeterioration* 

*Transactions of the Royal Society Biological Sciences*, Vol.364, No.1526, (July 2009), pp.

aliphatic polyesters. Part I. Properties and biodegradation of poly(butylene succinate-*co*-butylene adipate). *Polymer Degradation and Stability*, Vol.91, No.2,

A comprehensive study on poly(L-lactide) and poly(ε-caprolactone). *Polymer Degradation and Stability*, Vol.91, No.5, (May 2006), pp.1128-1137, ISSN 0141-3910 Viezie, D.L.; Thomas, E.L. & Adams, W.W. (1995). Low-voltage, high-resolution scanning

electron microscopy: a new characterization technique for polymer morphology.

2,3-dioxygenase from the cresols – Degrading *Stenotrophomonas maltophilia* strain KB2. *International Biodeterioration & Biodegradation*, Vol.65, No.6, (September 2011), A battery generally consists of three important parts namely, anode, cathode and electrolyte. The batteries further classified into primary and secondary batteries. Among the different kinds of batteries, Li-ion batteries are plays a very important role in the development of modern technologies especially in the portable electronic device industries and in heavy electrical vehicles because of its advantages such as high theoretical capacity, improved safety, lower material costs, ease of fabrication into flexible geometries, and the absence of electrolyte leakage. In the battery system different kinds of electrolytes were used for promoting the ions from anode to cathode (during charge) and cathode to anode (during discharge). For this purpose, liquid electrolyte is identified as suitable electrolytes which facilitate higher ionic conductivity (10-2 Scm-1) than other electrolyte systems. However, it has several disadvantages namely gas formation during the operation, leakage, difficult to utilise for portable applications and etc. To overcome these difficulties, many attempts were made on solid polymer electrolyte systems. The main objective of the researchers is to improve the ambient temperature ionic conductivity, mechanical stability, thermal and interfacial stability of the electrolytes. However, it is difficult task for the researchers, inorder to improve these basic requirements of the electrolytes simultaneously because the ionic conductivity and mechanical strength of a polymer electrolyte are disparate to each other, i.e., mechanical strength of the electrolyte decreases as conductivity increases.

In recent years, Polymer electrolytes have been attracted scientific and technological importance because of their potential applications in many areas such as Li-ion polymer batteries, super capacitor, electro chromic devices and etc. The idea of preparation of

<sup>\*</sup> Corresponding Authors

Surface Analysis Studies on Polymer Electrolyte

**1.2.3 Solvent free polymer electrolytes** 

electrolytes iii) Composite polymer electrolytes.

batteries to be constructed [Quartarone et al., 1998].

al., 1994 and Slane & Salomon et al., 1995].

*c) Composite polymer electrolytes* 

*a) Solid polymer electrolytes* 

*b) Gel polymer electrolytes* 

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 673

The polymer –salt complexes are formed by complexes between salts of alkali metals and polymer containing solvating heteroatoms such as O, N, S, etc. The most common examples are complexation between poly (ethylene oxide) (PEO) and alkali metal salts. The polymer salt complexes are further classified into: i) Solid polymer electrolytes ii) Blend polymer electrolytes iii) Gel polymer electrolytes iv) Composite polymer electrolytes. Among the various polymer electrolytes which are used in Li-ion batteries, solvent free polymer electrolytes are the most favourable for device fabrications. The solvent free polymer salt complexes are further classified into: i) Solid polymer electrolytes ii) Gel polymer

Solid polymer electrolytes (SPEs) have an ionic conductivity when modified by dissolving alkali salts in suitable polymer matrix. SPEs are typically thin films, which have a wide range of electrochemical applications such as batteries and electrochromic devices. They have several advantages when used in a battery and can be formed into thin films of large surface area giving high power levels. The flexibility of the films allows space-efficient

Plasticizers incorporated polymer- salt complex is called gel polymer electrolytes. The addition of plasticizers into the polymer matrix softens the polymers and they increase free volumes which are used for ion migration. Addition of plasticizer also increases the chain flexibility, reduces crystallinity, decreases the glass transition temperature and hence increases the ionic conductivity. The conductivity of PEO: LiBF4 is of the order of x10-6 Scm-1 which has been increased to the order of x10-4Scm-1 when the complex is plasticized at 25ºC [Chiodelli et al., 1988] this is mainly due to the specific nature of the plasticizers and the prepared gels has both the cohesive properties of solids and the diffusive property of liquids. Even though the gel polymer electrolyte exhibits high ionic conductivity, its thermal and mechanical stability are poor and it has higher reactivity towards the electrode. Gel electrolytes may undergo solvent exudation upon long storage, especially under open atmosphere conditions. This phenomenon is known as 'Synerisis effect', and has been encountered in many systems such as PAN: EC: PC: LiClO4, PAN: EC: PC: LiAsF6 [Groce et

This is another approach in which both the ionic conductivity and the mechanical stability of the electrolytes were considerably enhanced simultaneously. Composite polymer electrolytes are prepared by the addition of high surface inorganic fillers such as Al2O3, SiO2, MgO, LiAlO2, TiO2, BaTiO3 and Zeolite powders. The mechanical strength and stiffness of the complex systems were improved appreciably when the fillers are incorporated into the polymer matrix. However the main advantages of the composite electrolyte is the enhancement of room temperature ionic conductivity and an improved stability at the electrode electrolyte interface. The inert fillers due to its large surface area prevent the local chain reorganization with the result of locking in at ambient temperature, a high degree of disorder characteristic of the amorphous phase, which is more favour for the high ionic

polymer electrolytes was first proposed by Wright and Fenton et.al in 1973 [wright et al., 1973] but their technological significances are fulfilled and appreciated by Armand et.al few years later [Armand et al., 1998]. Poly (ethylene oxide) was the first solvating polymer to be proposed and studied in solid polymer electrolyte (SPE) Li-rechargeable batteries. Most of the solid polymer electrolytes (SPEs) are prepared by dissolving lithium salts in a solvating polymer using common solvent or by diffusion in the solid (or) molten state. An effort is also made to fix or immobilize the anion on the polymer matrix by covalent bonding (or) another chemical or physical process.

In general, Polymer electrolytes are plastic materials that can be modified and processed by conventional techniques. If the polymer chains are helped for a charge transport of the ionic type, often called "polymer electrolyte". Solid polymer electrolytes (SPEs) afford two important roles in Li-ion battery. 1) It is used as a separator in the battery system because of it rigid structure, at the same time to avoid the electrical contact between the anode and the cathode; 2) it is the medium in which the ions are transported between the anode and cathode during the cell operations. As a result, the polymer electrolyte should act as good electrical insulator but at the same time it should has high ionic conductivity.

## **1.1 Classification of polymer electrolyte systems**

The polymer electrolyte systems could be classified into three categories, namely, i) Polyelectrolyte, ii) Solvent swollen polymer electrolyte and iii) solvent free polymer electrolytes.

## **1.2.1 Polyelectrolyte's**

Polyelectrolytes are polymers which have their own ion-generating groups chemically bound to the macromolecular chain and the presence of a counter-ion maintains the electroneutrality of the salt. This class of materials either positively or negatively charged ions covalently attached to the polymer backbone and therefore only the unattached counter ion has long range mobility. The conductivity of these polymers is very low (10-10-10-15 Scm-1) in dry conditions but hydrated polyelectrolytes achieve high conductivity in the presence of high dielectric constant solvent (e.g. Water). In hydrated polyelectrolytes, ionic transport takes place through the aqueous medium in which the polymer is dispersed. Slade et.al. [Slade et al., 1983] reported high ambient temperature conductivity of 10-2 Scm-1 in hydrated Nafion.

## **1.2.2 Solvent swollen polymer electrolytes**

In solvent swollen polymers, solvents (aqueous/non-aqueous) swell the basic polymer host [like poly(vinyl alcohol) or poly(vinyl pyrrolidone)]. The dopant ionic solutes like H3PO4 are accommodated in the swollen lattice which permits the ionic motion in solvent rich swollen region of the polymer host. These materials are, in general, unstable and their conductivity depends on the concentration of the solvent in the swollen region. The properties of such polymers depend on the pre-treatment, structure of the sample, temperature, relative humidity, etc.

Examples: Nafian, complexes of poly(vinyl alcohol) (PVA) with H3PO4 [Polak et al., 1986] poly (silamine) with H3PO4[Rekukawa et al.,, 1996], PEI with H3SO4 [Daniel et al., 1988].

#### **1.2.3 Solvent free polymer electrolytes**

The polymer –salt complexes are formed by complexes between salts of alkali metals and polymer containing solvating heteroatoms such as O, N, S, etc. The most common examples are complexation between poly (ethylene oxide) (PEO) and alkali metal salts. The polymer salt complexes are further classified into: i) Solid polymer electrolytes ii) Blend polymer electrolytes iii) Gel polymer electrolytes iv) Composite polymer electrolytes. Among the various polymer electrolytes which are used in Li-ion batteries, solvent free polymer electrolytes are the most favourable for device fabrications. The solvent free polymer salt complexes are further classified into: i) Solid polymer electrolytes ii) Gel polymer electrolytes iii) Composite polymer electrolytes.

#### *a) Solid polymer electrolytes*

672 Scanning Electron Microscopy

polymer electrolytes was first proposed by Wright and Fenton et.al in 1973 [wright et al., 1973] but their technological significances are fulfilled and appreciated by Armand et.al few years later [Armand et al., 1998]. Poly (ethylene oxide) was the first solvating polymer to be proposed and studied in solid polymer electrolyte (SPE) Li-rechargeable batteries. Most of the solid polymer electrolytes (SPEs) are prepared by dissolving lithium salts in a solvating polymer using common solvent or by diffusion in the solid (or) molten state. An effort is also made to fix or immobilize the anion on the polymer matrix by covalent bonding (or)

In general, Polymer electrolytes are plastic materials that can be modified and processed by conventional techniques. If the polymer chains are helped for a charge transport of the ionic type, often called "polymer electrolyte". Solid polymer electrolytes (SPEs) afford two important roles in Li-ion battery. 1) It is used as a separator in the battery system because of it rigid structure, at the same time to avoid the electrical contact between the anode and the cathode; 2) it is the medium in which the ions are transported between the anode and cathode during the cell operations. As a result, the polymer electrolyte should act as good

The polymer electrolyte systems could be classified into three categories, namely, i) Polyelectrolyte, ii) Solvent swollen polymer electrolyte and iii) solvent free polymer

Polyelectrolytes are polymers which have their own ion-generating groups chemically bound to the macromolecular chain and the presence of a counter-ion maintains the electroneutrality of the salt. This class of materials either positively or negatively charged ions covalently attached to the polymer backbone and therefore only the unattached counter ion has long range mobility. The conductivity of these polymers is very low (10-10-10-15 Scm-1) in dry conditions but hydrated polyelectrolytes achieve high conductivity in the presence of high dielectric constant solvent (e.g. Water). In hydrated polyelectrolytes, ionic transport takes place through the aqueous medium in which the polymer is dispersed. Slade et.al. [Slade et al., 1983]

In solvent swollen polymers, solvents (aqueous/non-aqueous) swell the basic polymer host [like poly(vinyl alcohol) or poly(vinyl pyrrolidone)]. The dopant ionic solutes like H3PO4 are accommodated in the swollen lattice which permits the ionic motion in solvent rich swollen region of the polymer host. These materials are, in general, unstable and their conductivity depends on the concentration of the solvent in the swollen region. The properties of such polymers depend on the pre-treatment, structure of the sample, temperature, relative

Examples: Nafian, complexes of poly(vinyl alcohol) (PVA) with H3PO4 [Polak et al., 1986] poly (silamine) with H3PO4[Rekukawa et al.,, 1996], PEI with H3SO4 [Daniel et al., 1988].

reported high ambient temperature conductivity of 10-2 Scm-1 in hydrated Nafion.

electrical insulator but at the same time it should has high ionic conductivity.

**1.1 Classification of polymer electrolyte systems** 

**1.2.2 Solvent swollen polymer electrolytes** 

another chemical or physical process.

electrolytes.

humidity, etc.

**1.2.1 Polyelectrolyte's** 

Solid polymer electrolytes (SPEs) have an ionic conductivity when modified by dissolving alkali salts in suitable polymer matrix. SPEs are typically thin films, which have a wide range of electrochemical applications such as batteries and electrochromic devices. They have several advantages when used in a battery and can be formed into thin films of large surface area giving high power levels. The flexibility of the films allows space-efficient batteries to be constructed [Quartarone et al., 1998].

#### *b) Gel polymer electrolytes*

Plasticizers incorporated polymer- salt complex is called gel polymer electrolytes. The addition of plasticizers into the polymer matrix softens the polymers and they increase free volumes which are used for ion migration. Addition of plasticizer also increases the chain flexibility, reduces crystallinity, decreases the glass transition temperature and hence increases the ionic conductivity. The conductivity of PEO: LiBF4 is of the order of x10-6 Scm-1 which has been increased to the order of x10-4Scm-1 when the complex is plasticized at 25ºC [Chiodelli et al., 1988] this is mainly due to the specific nature of the plasticizers and the prepared gels has both the cohesive properties of solids and the diffusive property of liquids. Even though the gel polymer electrolyte exhibits high ionic conductivity, its thermal and mechanical stability are poor and it has higher reactivity towards the electrode. Gel electrolytes may undergo solvent exudation upon long storage, especially under open atmosphere conditions. This phenomenon is known as 'Synerisis effect', and has been encountered in many systems such as PAN: EC: PC: LiClO4, PAN: EC: PC: LiAsF6 [Groce et al., 1994 and Slane & Salomon et al., 1995].

#### *c) Composite polymer electrolytes*

This is another approach in which both the ionic conductivity and the mechanical stability of the electrolytes were considerably enhanced simultaneously. Composite polymer electrolytes are prepared by the addition of high surface inorganic fillers such as Al2O3, SiO2, MgO, LiAlO2, TiO2, BaTiO3 and Zeolite powders. The mechanical strength and stiffness of the complex systems were improved appreciably when the fillers are incorporated into the polymer matrix. However the main advantages of the composite electrolyte is the enhancement of room temperature ionic conductivity and an improved stability at the electrode electrolyte interface. The inert fillers due to its large surface area prevent the local chain reorganization with the result of locking in at ambient temperature, a high degree of disorder characteristic of the amorphous phase, which is more favour for the high ionic

Surface Analysis Studies on Polymer Electrolyte

and cyclic voltametry.

analysis.

**1.4.2 PVC based electrolytes** 

**1.4.3 PAN based polymer electrolytes** 

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 675

the composite polymer electrolytes using poly(ethylene oxide)/ poly(triethylene glycol) benzoate, BaTiO3 and lithium imides. They estimated the ionic conductivity value as 1.6x10-3 Scm-1 at 80ºC and the electrochemical stability window as 4.0V. The membrane was also characterized by TG/DTA and it is thermally stable upto 307ºC. Novel effect of organic acids such as malonic, maleic and carboxylic acids on PEO/LiClO4/Al2O3 complexes was studied by Park et al. [Park et al., 2006]. It was noted that the ionic conductivity of the film consisting of PEO/LiClO4: Citric acid (99.95:0.05 wt%) was 3.25x10-4Scm-1 at 30ºC and it was further improved to 3.1x10-3 Scm-1 at 20ºC by adding 20wt% of Al2O3 filler. The prepared membranes were also characterized by Brewster Angle Microscopy (BAM), thermal analysis

PVdF/PVC blend composite polymer electrolyte was prepared by Aravindhan et al. [Aravindhan et al., 2007] incorporating lithium bis(oxalate) borate and ZrO2. All the prepared membranes were subjected to SEM, XRD and ac impedance studies. The maximum ionic conductivity (1.53x10-3 Scm-1) was obtained for 2.5wt% of ZrO2 at 343K. The ionic conductivity and FTIR studies on plasticized polymer electrolyte based on PVC and PMMA as host polymer were studied by Manuel Stephan et al. It was found that LiBF4 based PVC/PMMA/EC/PC complexes exhibited higher ionic conductivity compared to that with LiClO4. The thermal stability of the films was also ascertained using TG/DTA

Kim et.al [Kim,D.W. Sun,Y.K. 2001] prepared highly porous polymer electrolyte employing P(VdF-co-HFP) and PAN with a view to attain high ionic conductivity and good mechanical strength. Lithium-ion polymer battery using these gel polymer electrolytes was assembled, and its charge-discharge characteristics were also reported. Panero et al. [Panero et al., 2002] studied the characteristics and the properties of a polymer electrolyte formed by trapping LiPF6-PC solution in a poly(acrylonitrile) matrix with the addition of Al2O3. They reported the ionic conductivity value as 0.8x10-2Scm-1 at 25ºC. The performances of the electrolyte were found to be promising in terms of cycle life and basic energy density content. Very recently, Moreno et.al [Moreno et al., 2010] reported a series of composite electrolytes basically constituted by poly(acrylonitrile), Clay and montmorillonite as filler. The structural and complex formations of the CPEs were also studied. However, the composite

based on PAN system showed poor ionic conductivity of the order of x10-6 Scm-1.

Tsutsumi and Kitagawa [Tsutsumi and Kitagawa et al., 2010] synthesized a new type polymer electrolyte films based on poly(acrylonitrile), and Cyanoethylated poly(vinyl alcohol) (CN-PVA) and its conductivity behaviour was also investigated. They found the ionic conductivity value as 14.6x10-3Scm-1 at 30ºC for PAN (10)-CN-PVA (10) - LiClO4 (8)-PC (4) complex system. The interactions of Li+-ion and nitrile groups of PAN in the matrix were confirmed by FTIR analysis. The ionic conductivity and FTIR studies were carried out on PAN based gel electrolytes with EC: PC and EC: DMC mixtures as plasticizers, LiClO4 or LiBF4 as the salt by Amaral et al [Amaral et al., 2007]. The high ionic conductivity (1.47x10-3 Scm-1) was estimated for 20:28:45:7 molar ratio of PAN-PVA: EC: DMC: LiBF4 system.

transport [G. Nagasubramanian and S. Di Stefano, 1990, Peter P Chu,P.P. Reddy, M.J., 2003]. The nano sized BaTiO3 incorporated PEO composite electrolytes exhibits ionic conductivity of the order of x10-3 Scm-1 and good electrochemical stability (4.0V).

## **1.3 Blend polymer electrolytes**

Blend is a mixture of two or more polymers. Mixing of two polymers is a well established strategy for the purpose of obtaining materials with combined superior properties or avoiding the need to synthesize novel structures constitutes an attractive research area. As many emerging applications are of limited volume and require specific property profiles not suitable for broad application utility, polymer blend technology is often the only viable method to deliver the desired material. These polymer blends have some unique properties that are different from the basic polymers from which these have been produced. To improve the processing behaviour for end use, one polymer blending with another polymer is a common practice. The exploitation of certain unique set of properties of individual polymer for the benefit of the overall properties of a multi component system forms the basis of polymer blending. Hence blending of polymers has resulted in the development of polymeric materials with desirable combination of properties. Polymer blend electrolytes are developed in such a way that they remain structurally stable during manufacturing, cell assembling, storage and usages as well as to prevent leakage from the cell container or without the cell.

### **1.4 Some commonly available polymer electrolytes for lithium ion batteries**

#### **1.4.1 PEO based electrolytes**

PEO is a crystalline polymer. The oxygen in PEO acts as a donor for the cation and the anion generally of large dimension stabilizes the PEO alkali salt complex.

The polymer electrolytes composed of a blend of poly (ethylene oxide) (PEO) and poly (vinylidenefluoride-hexa fluoropropylene) as a host polymer, mixture of EC and PC as plasticizer and LiClO4 as a salt were prepared by Fan et al. [Fan et al., 2002]. The ionic conductivity of various compositions of blend polymers was found to be in the order of x10- 4 Scm-1 at 30ºC. On increasing the PEO content in the matrix, the conductivity decreased due to its high crystalline nature. The mechanical strength of the polymer electrolytes was measured from stress-strain tests. The electrolytes were also characterized by SEM, XRD and thermal analysis techniques. Xi et al. [Xi., 2006] aimed to improve ionic conductivity with a novel approach using PEO and PVdF as host polymers by phase inversion technique. The room temperature conductivity was measured as a function of PEO content. As the weight ratio of PEO was increased from 40 to 50%, the ionic conductivity increased more than one magnitude from 0.15 to 1.96 x10-1Scm-1 which is mainly due to the increasing of pore connectivity. This is very important for the transport of charge carriers in microporous polymer membrane. The plasticizer effect on PEO-salt complex was studied by Fan et al [Fan et al., 2008] using succinitrile (SN) as a plasticizer, LiClO4, LiPF6 and LiCF3SO3 as lithium salts. They found that the addition of plasticizer was responsible for high ionic conductivity which could be attributed to the high polarity and diffusivity of succinitrile. This, in turn, decreased the crystallinity of PEO polymer. Activation energy of the electrolytes was also estimated from Arrhenius plot. Itoh et al. [Itoh et al., 2003] prepared the composite polymer electrolytes using poly(ethylene oxide)/ poly(triethylene glycol) benzoate, BaTiO3 and lithium imides. They estimated the ionic conductivity value as 1.6x10-3 Scm-1 at 80ºC and the electrochemical stability window as 4.0V. The membrane was also characterized by TG/DTA and it is thermally stable upto 307ºC. Novel effect of organic acids such as malonic, maleic and carboxylic acids on PEO/LiClO4/Al2O3 complexes was studied by Park et al. [Park et al., 2006]. It was noted that the ionic conductivity of the film consisting of PEO/LiClO4: Citric acid (99.95:0.05 wt%) was 3.25x10-4Scm-1 at 30ºC and it was further improved to 3.1x10-3 Scm-1 at 20ºC by adding 20wt% of Al2O3 filler. The prepared membranes were also characterized by Brewster Angle Microscopy (BAM), thermal analysis and cyclic voltametry.

## **1.4.2 PVC based electrolytes**

674 Scanning Electron Microscopy

transport [G. Nagasubramanian and S. Di Stefano, 1990, Peter P Chu,P.P. Reddy, M.J., 2003]. The nano sized BaTiO3 incorporated PEO composite electrolytes exhibits ionic conductivity

Blend is a mixture of two or more polymers. Mixing of two polymers is a well established strategy for the purpose of obtaining materials with combined superior properties or avoiding the need to synthesize novel structures constitutes an attractive research area. As many emerging applications are of limited volume and require specific property profiles not suitable for broad application utility, polymer blend technology is often the only viable method to deliver the desired material. These polymer blends have some unique properties that are different from the basic polymers from which these have been produced. To improve the processing behaviour for end use, one polymer blending with another polymer is a common practice. The exploitation of certain unique set of properties of individual polymer for the benefit of the overall properties of a multi component system forms the basis of polymer blending. Hence blending of polymers has resulted in the development of polymeric materials with desirable combination of properties. Polymer blend electrolytes are developed in such a way that they remain structurally stable during manufacturing, cell assembling, storage and usages as well as to prevent leakage from the cell container or

**1.4 Some commonly available polymer electrolytes for lithium ion batteries** 

generally of large dimension stabilizes the PEO alkali salt complex.

PEO is a crystalline polymer. The oxygen in PEO acts as a donor for the cation and the anion

The polymer electrolytes composed of a blend of poly (ethylene oxide) (PEO) and poly (vinylidenefluoride-hexa fluoropropylene) as a host polymer, mixture of EC and PC as plasticizer and LiClO4 as a salt were prepared by Fan et al. [Fan et al., 2002]. The ionic conductivity of various compositions of blend polymers was found to be in the order of x10- 4 Scm-1 at 30ºC. On increasing the PEO content in the matrix, the conductivity decreased due to its high crystalline nature. The mechanical strength of the polymer electrolytes was measured from stress-strain tests. The electrolytes were also characterized by SEM, XRD and thermal analysis techniques. Xi et al. [Xi., 2006] aimed to improve ionic conductivity with a novel approach using PEO and PVdF as host polymers by phase inversion technique. The room temperature conductivity was measured as a function of PEO content. As the weight ratio of PEO was increased from 40 to 50%, the ionic conductivity increased more than one magnitude from 0.15 to 1.96 x10-1Scm-1 which is mainly due to the increasing of pore connectivity. This is very important for the transport of charge carriers in microporous polymer membrane. The plasticizer effect on PEO-salt complex was studied by Fan et al [Fan et al., 2008] using succinitrile (SN) as a plasticizer, LiClO4, LiPF6 and LiCF3SO3 as lithium salts. They found that the addition of plasticizer was responsible for high ionic conductivity which could be attributed to the high polarity and diffusivity of succinitrile. This, in turn, decreased the crystallinity of PEO polymer. Activation energy of the electrolytes was also estimated from Arrhenius plot. Itoh et al. [Itoh et al., 2003] prepared

of the order of x10-3 Scm-1 and good electrochemical stability (4.0V).

**1.3 Blend polymer electrolytes** 

without the cell.

**1.4.1 PEO based electrolytes** 

PVdF/PVC blend composite polymer electrolyte was prepared by Aravindhan et al. [Aravindhan et al., 2007] incorporating lithium bis(oxalate) borate and ZrO2. All the prepared membranes were subjected to SEM, XRD and ac impedance studies. The maximum ionic conductivity (1.53x10-3 Scm-1) was obtained for 2.5wt% of ZrO2 at 343K. The ionic conductivity and FTIR studies on plasticized polymer electrolyte based on PVC and PMMA as host polymer were studied by Manuel Stephan et al. It was found that LiBF4 based PVC/PMMA/EC/PC complexes exhibited higher ionic conductivity compared to that with LiClO4. The thermal stability of the films was also ascertained using TG/DTA analysis.

### **1.4.3 PAN based polymer electrolytes**

Kim et.al [Kim,D.W. Sun,Y.K. 2001] prepared highly porous polymer electrolyte employing P(VdF-co-HFP) and PAN with a view to attain high ionic conductivity and good mechanical strength. Lithium-ion polymer battery using these gel polymer electrolytes was assembled, and its charge-discharge characteristics were also reported. Panero et al. [Panero et al., 2002] studied the characteristics and the properties of a polymer electrolyte formed by trapping LiPF6-PC solution in a poly(acrylonitrile) matrix with the addition of Al2O3. They reported the ionic conductivity value as 0.8x10-2Scm-1 at 25ºC. The performances of the electrolyte were found to be promising in terms of cycle life and basic energy density content. Very recently, Moreno et.al [Moreno et al., 2010] reported a series of composite electrolytes basically constituted by poly(acrylonitrile), Clay and montmorillonite as filler. The structural and complex formations of the CPEs were also studied. However, the composite based on PAN system showed poor ionic conductivity of the order of x10-6 Scm-1.

Tsutsumi and Kitagawa [Tsutsumi and Kitagawa et al., 2010] synthesized a new type polymer electrolyte films based on poly(acrylonitrile), and Cyanoethylated poly(vinyl alcohol) (CN-PVA) and its conductivity behaviour was also investigated. They found the ionic conductivity value as 14.6x10-3Scm-1 at 30ºC for PAN (10)-CN-PVA (10) - LiClO4 (8)-PC (4) complex system. The interactions of Li+-ion and nitrile groups of PAN in the matrix were confirmed by FTIR analysis. The ionic conductivity and FTIR studies were carried out on PAN based gel electrolytes with EC: PC and EC: DMC mixtures as plasticizers, LiClO4 or LiBF4 as the salt by Amaral et al [Amaral et al., 2007]. The high ionic conductivity (1.47x10-3 Scm-1) was estimated for 20:28:45:7 molar ratio of PAN-PVA: EC: DMC: LiBF4 system.

Surface Analysis Studies on Polymer Electrolyte

1 for upto 20 cycles at different current densities.

**1.4.7 PVAc based polymer electrolytes** 

was confirmed by XRD and SEM analysis.

performance.

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 677

polymer PVdF-co-HFP/ PC/ DEC/ LiClO4 and PVdF/ PC/ DEC/ LiClO4 separately. The co-polymer complex showed higher ionic conductivity and transport number compared to PVdF system. The higher conductivity of the polymer electrolyte based on copolymer was attributed to its higher amorphousity. Wang et.al [Wang et al, 2004] reported that the polymer electrolyte composed of poly(methyl methacrylate-co-acrylonitrile-co-lithium methacrylate) (PMAML) and poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-co-HFP) exhibited high ionic conductivity and good electrochemical stability. The ionic conductivity was about 2*.*6×10-3 Scm−1 at ambient temperature and the electrochemical window of the polymer electrolyte was about 4.6V. Charge –discharge test results revealed that lithium ion batteries with these gel polymer electrolytes have good electrochemical

Manuel Stephan et.al [Manuel Stephan et al., 2006] prepared the composite polymer electrolyte membranes, comprising Poly(vinylidene fluoride–hexafluoropropylene), Aluminum Oxyhydroxide of two different sizes 7nm/14 nm and LiN(C2F5SO2)2 as lithium salt and they found that the incorporation of the inert filler not only reduces the crystallinity of the polymer host but also acts as 'solid plasticizer' capable of enhancing the transport properties and also provides a better interfacial property towards lithium metal anode. Nam-Soon Choi et.al [Choi et al., 2001] reported that the interfacial stability between the polymer electrolyte and the lithium electrode was enhanced by blending PVAc with P(VdFco-HFP)(Kynar 2801). The ionic conductivity of the polymer electrolyte based on the Kynar 2801: PVAc (7:3, w:w) blend was 2.3×10-3 S cm-1 at 25°C. Kim et.al [Kim et al., 2005] prepared and characterized gel polymer electrolytes consisting 25wt% PVdF-co-HFP/65 (EC+PC)/10 wt% LiN(CF3SO2)2.They reported the ionic conductivity value as 1.2x10-3Scm-1. The electrochemical stability window of the membrane was obtained at around 4.8V Vs Li/Li+ using linear sweep voltametry technique. The Charge – discharge behaviour of the membrane was also studied and they estimated the specific discharge capacity as 140 mAhg-

Though variety of polymer electrolytes were characterized for the fast four decades, a limited number of studies were made on PVAc based polymer electrolytes. PVAc polymer has a large dipole moment and high relaxation time. Baskaran et al [Baskaran et al., 2006] prepared the polymer electrolyte comprising of PVAc-PMMA and reported the conductivity value as 1.76x10-3Scm-1 at 303K. The DSC thermograms of the blend electrolytes showed two Tg's and they decreased with an increase of LiClO4 concentration. The structural and complex formations of the electrolytes were confirmed by X-ray diffraction analysis. They established that the optimized blend ratio of PVAc: PMMA: LiClO4 is suitable for lithium battery applications. Structural, thermal and transport properties of PVAc-LiClO4 base complexes were studied by Baskaran et al [Baskaran et al., 2007]. The bulk conductivity of PVAc: LiClO4 system was found to vary between 7.6x10-7Scm and 6.2x10-5 Scm-1 at 303K with an increase in the salt concentration. The amorphous nature of the polymer complexes

Surface morphology and ionic conductivity of the membrane based on P(EO)/PVAc /LiClO4 were studied by Animitsa et al [Animitsa et al., 1998]. They reported the conductivity value as 10-5 Scm-1 for lower concentration of PVAc at 25ºC. Baskaran et al.

Charge/discharge performance of the maximum ionic conductivity complex was also studied. The practical performance and thermal stability of Li-ion polymer batteries with LiNi0.8Co0.2O2, mesocarbon microbead-based graphite, and poly (acrylonitrile) (PAN) based gel electrolytes were reported by Akashi et al. [Akashi et al., 2002].

## **1.4.4 PVdF based polymer electrolytes**

PVdF is a semicrystalline polymer and the electrolytes based on PVdF are expected to have high anodic stabilities due to strong electron withdrawing functional groups (-C-F). It also has high permittivity, relatively low dissipation factor and high dielectric constant (ε=8.4) which assist in high ionization of lithium salts, providing a high concentration of charge carriers. Choe et al. [Cheo et al., 1995] reported that the PVdF based electrolytes plasticized with a solution of LiN(SO2CF3)2 in PC had a conductivity of 1.74x10-3 Scm-1 at 30ºC and has a oxidatively stable potential limits between 3.9 and 4.3V vs Li+/Li. Nicotera et al. [Nicotera et al., 2006] measured the ionic conductivity and the lithium salt diffusion coefficient of PMMA/PVdF based blend electrolytes with EC/PC as plasticizers and lithium perchlorate as salt by the PFG-NMR method, which revealed maximum lithium mobility for the composition PMMA 60%-PVdF 40%. Raman spectroscopic study confirmed the change of interaction between the lithium cations and the plasticizer molecules for different PMMA/PVdF ratios. Wang et al. [Wang et al., 2007] prepared the nanocomposite polymer electrolytes comprising of poly (vinylidene fluoride) (PVdF) as a host polymer, lithium perchlorate (LiClO4) as salt and TiO2 used as a filler by solvent-casting technique. The prepared films were characterized by XRD, DSC and SEM. The conductivity value was found to be of the order of 10-3 Scm-1 for the sample with 10% TiO2.

#### **1.4.5 PMMA based electrolytes**

Ali et al. [Ali et al., 2007] reported the electrical properties of polymer electrolytes comprising PMMA, PC, EC as plasticizer and different lithium salts LiCF3SO3 and LiN(CF3SO2)2. The polymer electrolytes exhibited high ionic conductivity at room temperature in the range of 10-6 to 10-4 Scm-1. The temperature dependence studies confirmed that the conduction in electrolyte is only by ions and seemed to obey the VTF rule. FTIR spectroscopy studies confirmed the polymer-salt interactions. FTIR spectroscopic investigations coupled with ionic conductivity and viscosity measurements on lithium imide LiN(CF3SO2)2–propylene carbonate (PC)–poly(methyl methacrylate) (PMMA) based liquid and gel electrolytes over a wide range of salt (0.025–3 M) and polymer (5–25 wt.%) concentration were reported by Deepa et al. [Deepa et al., 2004] and found that the high ionic conductivity occurs at salt concentrations ≥1.25 M.

#### **1.4.6 PVdF-co-HFP based polymer electrolytes**

In recent years, the studies on PVdF-co-HFP based systems are electrochemically stable and indispensable for the electrode properties. The PVdF-co-HFP based electrolyte system shows high electrochemical stability in the range 4V. Fan et al. [Fan et al., 2002] studied the thermal, electrical and mechanical properties of EC/PC/LiClO4 based PEO/P(VdF-co-HFP) blends. They concluded that the polymers have good compatibility and PVdF-HFP hinders the crystallinity of PEO. Saika and Kumar [Saika and Kumar, 2004] made systematic studies on the ionic conductivity and transport properties of polymer electrolytes comprising of co-

Charge/discharge performance of the maximum ionic conductivity complex was also studied. The practical performance and thermal stability of Li-ion polymer batteries with LiNi0.8Co0.2O2, mesocarbon microbead-based graphite, and poly (acrylonitrile) (PAN) based

PVdF is a semicrystalline polymer and the electrolytes based on PVdF are expected to have high anodic stabilities due to strong electron withdrawing functional groups (-C-F). It also has high permittivity, relatively low dissipation factor and high dielectric constant (ε=8.4) which assist in high ionization of lithium salts, providing a high concentration of charge carriers. Choe et al. [Cheo et al., 1995] reported that the PVdF based electrolytes plasticized with a solution of LiN(SO2CF3)2 in PC had a conductivity of 1.74x10-3 Scm-1 at 30ºC and has a oxidatively stable potential limits between 3.9 and 4.3V vs Li+/Li. Nicotera et al. [Nicotera et al., 2006] measured the ionic conductivity and the lithium salt diffusion coefficient of PMMA/PVdF based blend electrolytes with EC/PC as plasticizers and lithium perchlorate as salt by the PFG-NMR method, which revealed maximum lithium mobility for the composition PMMA 60%-PVdF 40%. Raman spectroscopic study confirmed the change of interaction between the lithium cations and the plasticizer molecules for different PMMA/PVdF ratios. Wang et al. [Wang et al., 2007] prepared the nanocomposite polymer electrolytes comprising of poly (vinylidene fluoride) (PVdF) as a host polymer, lithium perchlorate (LiClO4) as salt and TiO2 used as a filler by solvent-casting technique. The prepared films were characterized by XRD, DSC and SEM. The conductivity value was

Ali et al. [Ali et al., 2007] reported the electrical properties of polymer electrolytes comprising PMMA, PC, EC as plasticizer and different lithium salts LiCF3SO3 and LiN(CF3SO2)2. The polymer electrolytes exhibited high ionic conductivity at room temperature in the range of 10-6 to 10-4 Scm-1. The temperature dependence studies confirmed that the conduction in electrolyte is only by ions and seemed to obey the VTF rule. FTIR spectroscopy studies confirmed the polymer-salt interactions. FTIR spectroscopic investigations coupled with ionic conductivity and viscosity measurements on lithium imide LiN(CF3SO2)2–propylene carbonate (PC)–poly(methyl methacrylate) (PMMA) based liquid and gel electrolytes over a wide range of salt (0.025–3 M) and polymer (5–25 wt.%) concentration were reported by Deepa et al. [Deepa et al., 2004] and found that the high

In recent years, the studies on PVdF-co-HFP based systems are electrochemically stable and indispensable for the electrode properties. The PVdF-co-HFP based electrolyte system shows high electrochemical stability in the range 4V. Fan et al. [Fan et al., 2002] studied the thermal, electrical and mechanical properties of EC/PC/LiClO4 based PEO/P(VdF-co-HFP) blends. They concluded that the polymers have good compatibility and PVdF-HFP hinders the crystallinity of PEO. Saika and Kumar [Saika and Kumar, 2004] made systematic studies on the ionic conductivity and transport properties of polymer electrolytes comprising of co-

gel electrolytes were reported by Akashi et al. [Akashi et al., 2002].

found to be of the order of 10-3 Scm-1 for the sample with 10% TiO2.

ionic conductivity occurs at salt concentrations ≥1.25 M.

**1.4.6 PVdF-co-HFP based polymer electrolytes** 

**1.4.4 PVdF based polymer electrolytes** 

**1.4.5 PMMA based electrolytes** 

polymer PVdF-co-HFP/ PC/ DEC/ LiClO4 and PVdF/ PC/ DEC/ LiClO4 separately. The co-polymer complex showed higher ionic conductivity and transport number compared to PVdF system. The higher conductivity of the polymer electrolyte based on copolymer was attributed to its higher amorphousity. Wang et.al [Wang et al, 2004] reported that the polymer electrolyte composed of poly(methyl methacrylate-co-acrylonitrile-co-lithium methacrylate) (PMAML) and poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-co-HFP) exhibited high ionic conductivity and good electrochemical stability. The ionic conductivity was about 2*.*6×10-3 Scm−1 at ambient temperature and the electrochemical window of the polymer electrolyte was about 4.6V. Charge –discharge test results revealed that lithium ion batteries with these gel polymer electrolytes have good electrochemical performance.

Manuel Stephan et.al [Manuel Stephan et al., 2006] prepared the composite polymer electrolyte membranes, comprising Poly(vinylidene fluoride–hexafluoropropylene), Aluminum Oxyhydroxide of two different sizes 7nm/14 nm and LiN(C2F5SO2)2 as lithium salt and they found that the incorporation of the inert filler not only reduces the crystallinity of the polymer host but also acts as 'solid plasticizer' capable of enhancing the transport properties and also provides a better interfacial property towards lithium metal anode. Nam-Soon Choi et.al [Choi et al., 2001] reported that the interfacial stability between the polymer electrolyte and the lithium electrode was enhanced by blending PVAc with P(VdFco-HFP)(Kynar 2801). The ionic conductivity of the polymer electrolyte based on the Kynar 2801: PVAc (7:3, w:w) blend was 2.3×10-3 S cm-1 at 25°C. Kim et.al [Kim et al., 2005] prepared and characterized gel polymer electrolytes consisting 25wt% PVdF-co-HFP/65 (EC+PC)/10 wt% LiN(CF3SO2)2.They reported the ionic conductivity value as 1.2x10-3Scm-1. The electrochemical stability window of the membrane was obtained at around 4.8V Vs Li/Li+ using linear sweep voltametry technique. The Charge – discharge behaviour of the membrane was also studied and they estimated the specific discharge capacity as 140 mAhg-1 for upto 20 cycles at different current densities.

## **1.4.7 PVAc based polymer electrolytes**

Though variety of polymer electrolytes were characterized for the fast four decades, a limited number of studies were made on PVAc based polymer electrolytes. PVAc polymer has a large dipole moment and high relaxation time. Baskaran et al [Baskaran et al., 2006] prepared the polymer electrolyte comprising of PVAc-PMMA and reported the conductivity value as 1.76x10-3Scm-1 at 303K. The DSC thermograms of the blend electrolytes showed two Tg's and they decreased with an increase of LiClO4 concentration. The structural and complex formations of the electrolytes were confirmed by X-ray diffraction analysis. They established that the optimized blend ratio of PVAc: PMMA: LiClO4 is suitable for lithium battery applications. Structural, thermal and transport properties of PVAc-LiClO4 base complexes were studied by Baskaran et al [Baskaran et al., 2007]. The bulk conductivity of PVAc: LiClO4 system was found to vary between 7.6x10-7Scm and 6.2x10-5 Scm-1 at 303K with an increase in the salt concentration. The amorphous nature of the polymer complexes was confirmed by XRD and SEM analysis.

Surface morphology and ionic conductivity of the membrane based on P(EO)/PVAc /LiClO4 were studied by Animitsa et al [Animitsa et al., 1998]. They reported the conductivity value as 10-5 Scm-1 for lower concentration of PVAc at 25ºC. Baskaran et al.

Surface Analysis Studies on Polymer Electrolyte

Fig. 1. Preparation of Polymer electrolyte

(Fig.1.).

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 679

own advantages and disadvantages. Among the various method of preparation solvent costing is the cost effective one and easier to control the moistures during the preparation of electrolytes as well as in the cell assembling. However, when assembling the cells, these membranes show poor cyclic behaviour. As a result many research groups have been mainly focused on phase inversion method for preparing the suitable electrolytes. Preparation of polymer electrolyte system is explained using the following flow chart

In the present study, all the electrolytes were prepared using solvent casting technique. The polymers PVAc, PVdF-co-HFP, and the salt LiClO4 were dissolved in a common solvent (tetrahydrofuran) separately. All three solutions were mixed together and starrier continuously using magnetic stirrer until got a homogeneous mixture. The low molecular weight plasticizer and the inorganic fillers were added into the matrix inorder to get the gel and composite polymer electrolytes systems. In the present study, ethylene carbonate(EC) and barium titanate (BaTiO3) were used a plasticizer and fillers respectively. Thus the obtained homogenous slurry was degassed to remove air bubbles for about five minutes and the slurry was poured on a well cleaned glass plate. The casted slurry was allowed to evaporate the solution at room temperature about 5h followed by the electrolyte membranes were heated using hot air oven at a temperature of 60 ºC for 6h in order to removing the

[Baskaran et al., 2006] studied the ac impedance and dielectric properties of PVdF/PVAc blend electrolytes. They reported that the blend ratio (75:25) of PVAc/PVdF exhibited a maximum ionic conductivity value of the order of 6.4x10-4Scm-1 at 343K. The ionic transference number of mobile ions was also estimated by Wagner's polarization method. The complex formation and thermal behaviour of the electrolytes were also studied by FTIR and DSC analysis respectively.

## **2. Basic requirements of polymers and the salt for polymer electrolytes**

Atoms or groups of atoms with sufficient electron donor power to form coordinate bonds with cations.


#### **Electron pair donicity (DN)**

DN measures the ability of the solvent to donate electrons to solvate the cations considered as a Lewis acid. So the polymer host should have high DN number.

#### **Acceptor number (AN)**

The acceptor number quantifies the possibility of anion salvation. It should be less for an inorganic salt so that cationic salvation is high compared to anionic salvation.

#### **Entropy term**

The entropy term depends on the optimal spatial disposition of the solvating units which should be high for the polymer host.

#### **2.1 Importance of Li<sup>+</sup> cation**

Last four decades, many alkali salts consisting Li+, Na+, K+, Ag+, Mg+, NH4 + cations were mixed with the polymers (PEO), (PPO) etc in the preparation of polymer electrolytes. Among the various cations in the periodic table, Li+ is the most electropositive. Lithium easily gives up electrons to form a positive Li+ which has small ionic radii (0.6Å). Lithium is promising candidate for high energy density batteries because of its high specific capacity of 3860 Ah/Kg, its light weight and high electrochemical reduction potential [Scrosati et al., 1994; Abraham et al., 1993; Dell, 2000].

#### **2.2 Preparation of polymer blend electrolytes**

Polymer blend electrolytes have been prepared using various approaches namely, Phase inversion method, Hot pressed method, Solvent casting method and etc. Each method has

[Baskaran et al., 2006] studied the ac impedance and dielectric properties of PVdF/PVAc blend electrolytes. They reported that the blend ratio (75:25) of PVAc/PVdF exhibited a maximum ionic conductivity value of the order of 6.4x10-4Scm-1 at 343K. The ionic transference number of mobile ions was also estimated by Wagner's polarization method. The complex formation and thermal behaviour of the electrolytes were also studied by FTIR

**2. Basic requirements of polymers and the salt for polymer electrolytes** 

intrapolymers ion bonds appears to be important

movement and yields high ionic conductivity. • The lattice energy of the salt should be low. • High electrochemical reduction potential

• Low glass transition temperature to increase the segmental motion.

as a Lewis acid. So the polymer host should have high DN number.

inorganic salt so that cationic salvation is high compared to anionic salvation.

Atoms or groups of atoms with sufficient electron donor power to form coordinate bonds

• Low barriers to bond rotation so that segmental motion of the polymer chain can take

• A suitable distance between coordinating centres for the formation of multiple

• The polymer should have amorphous phase which lowers the barrier for ionic

DN measures the ability of the solvent to donate electrons to solvate the cations considered

The acceptor number quantifies the possibility of anion salvation. It should be less for an

The entropy term depends on the optimal spatial disposition of the solvating units which

Last four decades, many alkali salts consisting Li+, Na+, K+, Ag+, Mg+, NH4+ cations were mixed with the polymers (PEO), (PPO) etc in the preparation of polymer electrolytes. Among the various cations in the periodic table, Li+ is the most electropositive. Lithium easily gives up electrons to form a positive Li+ which has small ionic radii (0.6Å). Lithium is promising candidate for high energy density batteries because of its high specific capacity of 3860 Ah/Kg, its light weight and high electrochemical reduction potential [Scrosati et al.,

Polymer blend electrolytes have been prepared using various approaches namely, Phase inversion method, Hot pressed method, Solvent casting method and etc. Each method has

and DSC analysis respectively.

with cations.

place readily.

**Electron pair donicity (DN)** 

**Acceptor number (AN)** 

**2.1 Importance of Li<sup>+</sup>**

should be high for the polymer host.

1994; Abraham et al., 1993; Dell, 2000].

**2.2 Preparation of polymer blend electrolytes** 

 **cation** 

**Entropy term** 

own advantages and disadvantages. Among the various method of preparation solvent costing is the cost effective one and easier to control the moistures during the preparation of electrolytes as well as in the cell assembling. However, when assembling the cells, these membranes show poor cyclic behaviour. As a result many research groups have been mainly focused on phase inversion method for preparing the suitable electrolytes. Preparation of polymer electrolyte system is explained using the following flow chart (Fig.1.).

Fig. 1. Preparation of Polymer electrolyte

In the present study, all the electrolytes were prepared using solvent casting technique. The polymers PVAc, PVdF-co-HFP, and the salt LiClO4 were dissolved in a common solvent (tetrahydrofuran) separately. All three solutions were mixed together and starrier continuously using magnetic stirrer until got a homogeneous mixture. The low molecular weight plasticizer and the inorganic fillers were added into the matrix inorder to get the gel and composite polymer electrolytes systems. In the present study, ethylene carbonate(EC) and barium titanate (BaTiO3) were used a plasticizer and fillers respectively. Thus the obtained homogenous slurry was degassed to remove air bubbles for about five minutes and the slurry was poured on a well cleaned glass plate. The casted slurry was allowed to evaporate the solution at room temperature about 5h followed by the electrolyte membranes were heated using hot air oven at a temperature of 60 ºC for 6h in order to removing the

Surface Analysis Studies on Polymer Electrolyte

and lithium salt.

stretching vibration modes of blend electrolytes.

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 681

all the blends, some peaks are found above 3000 cm-1, which correspond to the C-H

In addition, some new peaks are present and some of them are absent in the blend electrolytes. Thus the spectral analysis confirms the complexation of these two polymers

Fig. 2. FTIR analysis of LiClO4, PVAc, PVdF-co-HFP and their complexes

XRD patterns of LiClO4, PVdF-co-HFP, PVAc and their complexes A, B, C are shown in Fig. 3. The presence of characteristic peaks corresponding to the lithium salt reveals the high crystalline nature of the salt. Three peaks found at 2θ = 17.3, 18.59 and 38.78° for PVdF-co-

**3.2 X-ray diffraction analysis** 

residual solvent present in the electrolyte films. Finally, the harvested electrolyte films were stored in highly evacuated desiccators to avoid the moistures absorption.

## **3. Characterization of polymer blend electrolytes**

Ac impedance analysis was carried out with the help of stainless steel blocking electrodes by using a computer controlled micro auto lab type III Potentiostat/Galvanostat of frequency range 1 Hz–300 KHz in the temperature range 303–373 K. The XRD equipment used in this study was X'pert PRO PANlytical X-ray diffractometer. FTIR spectroscopy studies were carried out using SPECTRA RXI, Perkin Elmer spectro-photometer in the range 400–4000 cm−1. FTIR spectroscopy studies were carried out for confirming the complex formation using SPECTRA RXI, Perkin-Elmer spectro-photometer in the range 400–4000 cm−1. TG/DTA thermal analysis of the film having maximum ionic conductivity was studied using PYRIS DIAMOND under air atmosphere with the scan rate of 10 ◦C min−1. The electrolyte film having maximum ionic conductivity was subjected to atomic force microscopy [model Veeco-diCP-II]. The pore size and the root mean square (rms) roughness of the film were measured from the topography image. Secondary electron images of the sample were examined by using JEOL, JSM-840A scanning electron microscope

## **3.1 FTIR analysis**

Infrared spectral (IR) analysis is a powerful tool for identifying the nature of bonding and different functional groups present in a sample by monitoring the vibrational energy levels of the molecules, which are essentially the fingerprint of different molecules [Nagatomo et al., 1987]. Fig.2. depicts the FTIR transmittance spectra in the range 400-4000 cm-1 for polymers, the LiClO4 salt, the blend electrolyte with the incorporation of plasticizer ethylene carbonate and the filler BaTiO3.

The vibrational bands observed at 2933 cm-1 and 2465 cm-1 are ascribed to –CH3 asymmetric and symmetric stretching vibrations of PVAc respectively. The strong absorbance at 1734 cm-1 represents the C=O stretching vibration mode of PVAc polymer. The existence of C-O band has been confirmed by the strong absorbance band at around 1033 cm-1. The strong band at 1373 cm-1 is ascribed to -CH3 symmetric bending vibration of pure PVAc. The band at 1243 cm-1 is assigned to C-O-C symmetric stretching mode of vibration. The peak at 947 cm-1 is ascribed to CH bending vibration and the peak at 609 cm-1 is assumed to be linked with CH3 (C-O) group. The C-H wagging mode of vibration has been confirmed by the presence of a band at 799 cm-1 [Baskaran et al., 2004]. The vibrational peaks at 502 and 416 cm-1 are assigned to bending and wagging vibrations of –CF2 of PVdF-co-HFP polymer respectively. Crystalline phase of the PVdF-co-HFP polymer is identified by the vibrational bands at 985, 763, and 608 cm-1 and the amorphous phase of the co-polymer is confirmed by the presence of vibrational band at 872 cm-1 [Rajendran et al., 2002].

The strong absorption peak appeared at 1173 cm-1 is assigned to the symmetrical stretching of –CF2 group. The peak appeared at 1390 cm-1 is assigned to the CH2 groups [Rajendran et al., 2002; Singh Missan et al., 2006]. Table.1. shows the comparison of band spectra of pure polymers and their blends with different blend ratios. From the table, it is clear that the band assignments of FTIR spectra of blend samples are shifted from their pure spectra. For

residual solvent present in the electrolyte films. Finally, the harvested electrolyte films were

Ac impedance analysis was carried out with the help of stainless steel blocking electrodes by using a computer controlled micro auto lab type III Potentiostat/Galvanostat of frequency range 1 Hz–300 KHz in the temperature range 303–373 K. The XRD equipment used in this study was X'pert PRO PANlytical X-ray diffractometer. FTIR spectroscopy studies were carried out using SPECTRA RXI, Perkin Elmer spectro-photometer in the range 400–4000 cm−1. FTIR spectroscopy studies were carried out for confirming the complex formation using SPECTRA RXI, Perkin-Elmer spectro-photometer in the range 400–4000 cm−1. TG/DTA thermal analysis of the film having maximum ionic conductivity was studied using PYRIS DIAMOND under air atmosphere with the scan rate of 10 ◦C min−1. The electrolyte film having maximum ionic conductivity was subjected to atomic force microscopy [model Veeco-diCP-II]. The pore size and the root mean square (rms) roughness of the film were measured from the topography image. Secondary electron images of the

stored in highly evacuated desiccators to avoid the moistures absorption.

sample were examined by using JEOL, JSM-840A scanning electron microscope

the presence of vibrational band at 872 cm-1 [Rajendran et al., 2002].

Infrared spectral (IR) analysis is a powerful tool for identifying the nature of bonding and different functional groups present in a sample by monitoring the vibrational energy levels of the molecules, which are essentially the fingerprint of different molecules [Nagatomo et al., 1987]. Fig.2. depicts the FTIR transmittance spectra in the range 400-4000 cm-1 for polymers, the LiClO4 salt, the blend electrolyte with the incorporation of plasticizer ethylene

The vibrational bands observed at 2933 cm-1 and 2465 cm-1 are ascribed to –CH3 asymmetric and symmetric stretching vibrations of PVAc respectively. The strong absorbance at 1734 cm-1 represents the C=O stretching vibration mode of PVAc polymer. The existence of C-O band has been confirmed by the strong absorbance band at around 1033 cm-1. The strong band at 1373 cm-1 is ascribed to -CH3 symmetric bending vibration of pure PVAc. The band at 1243 cm-1 is assigned to C-O-C symmetric stretching mode of vibration. The peak at 947 cm-1 is ascribed to CH bending vibration and the peak at 609 cm-1 is assumed to be linked with CH3 (C-O) group. The C-H wagging mode of vibration has been confirmed by the presence of a band at 799 cm-1 [Baskaran et al., 2004]. The vibrational peaks at 502 and 416 cm-1 are assigned to bending and wagging vibrations of –CF2 of PVdF-co-HFP polymer respectively. Crystalline phase of the PVdF-co-HFP polymer is identified by the vibrational bands at 985, 763, and 608 cm-1 and the amorphous phase of the co-polymer is confirmed by

The strong absorption peak appeared at 1173 cm-1 is assigned to the symmetrical stretching of –CF2 group. The peak appeared at 1390 cm-1 is assigned to the CH2 groups [Rajendran et al., 2002; Singh Missan et al., 2006]. Table.1. shows the comparison of band spectra of pure polymers and their blends with different blend ratios. From the table, it is clear that the band assignments of FTIR spectra of blend samples are shifted from their pure spectra. For

**3. Characterization of polymer blend electrolytes** 

**3.1 FTIR analysis** 

carbonate and the filler BaTiO3.

all the blends, some peaks are found above 3000 cm-1, which correspond to the C-H stretching vibration modes of blend electrolytes.

In addition, some new peaks are present and some of them are absent in the blend electrolytes. Thus the spectral analysis confirms the complexation of these two polymers and lithium salt.

Fig. 2. FTIR analysis of LiClO4, PVAc, PVdF-co-HFP and their complexes

#### **3.2 X-ray diffraction analysis**

XRD patterns of LiClO4, PVdF-co-HFP, PVAc and their complexes A, B, C are shown in Fig. 3. The presence of characteristic peaks corresponding to the lithium salt reveals the high crystalline nature of the salt. Three peaks found at 2θ = 17.3, 18.59 and 38.78° for PVdF-co-

Surface Analysis Studies on Polymer Electrolyte

polymers.

conductivity.

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 683

For AC the ratio gives an analogous quantity, the impedance, Z, also measured in ohms. The impedance contains four main contributions; these are from resistance, capacitance, constant phase elements, and inductance. The induction is not an important factor for the polymer electrolytes although it can play a role in other electrochemical applications of

Measurement of the impedance as a function of frequency is called impedance spectroscopy. In general, impedance is complex quantity, in which the real and the imaginary parts are labelled Z' and Z" respectively. In the complex impedance plot, the real quantity Z' (X-axis) is plotted against Z" (Y-axis) which displayed the polymer electrolytes characteristics as an arc followed by the linear spike is straight line inclined to the real axis. From the plotted

The complex impedance plot of the PVAc/PVdF-co-HFP/LiClO4 electrolyte is shown in Fig.4a. Figure shows the semicircular portion which is mainly due to the parallel combination of the geometrical capacitance, Cg and the bulk resistance, R. When adding the plasticizer and the filler (Fig.4b.) into the electrolyte matrix, the impedance spectra shows only a linear spike which corresponds to the lower frequency region. It confirms the idea that the current carriers are ions and the majority of the conduction only by the ions not by the electrons. And the disappearance of the semicircular portion is may be due to the fact the corresponding characteristic frequency is higher than the frequency 300kHz and it is mainly depends on the instrument limit. From the obtained bulk resistance value, we can estimate the ionic conductivity value of the electrolyte system using the relation σ = l/Rb A , where, Rb is the bulk resistance of the electrolyte film, A is the area of the electrode surface and is the thickness of the electrolyte medium and σ is the ionic

It is noted from the spectra that the addition of plasticizer (ethylene carbonate) in to the polymer salt matrix greatly being reduced the bulk resistance of the system; this is because of the high dielectric nature of the low molecular weight plasticizer. The addition of plasticizer would considerably enhance the amorphous phase of the polymer electrolyte which will improve the ionic conductivity of the system. However, the gain in conductivity is adversely associated with a loss of the mechanical properties and by a loss of the compatibility with the lithium electrode, both effects resulting in serious problems since they affect the battery cycle life and increase the safety hazard. So it is necessary to identify the solid additives which would not affect the mechanical stability and interfacial stability of the electrolyte, at the same time will enhances the ionic conductivity. The addition of solid additives should improve the amorphicity of the electrolyte at room temperature. The addition of solid additives in the present study, such as nano filler BaTiO3 highly being enhanced the amourphicity of the electrolyte medium, hence the room temperature ionic conductivity and the interfacial stability of the electrode-electrolyte interface is increased. The ceramic dispersed electrolyte shows good thermal stability. The BaTiO3 incorporated sample is thermally stable up to 320 ºC. The temperature dependence of the conductivity is given by the Vogel–Tamman–Fulcher (VTF) equation σ = σ0 exp (-B/T-T0), where σ0 is the pre-exponential factor, B should not be confused with an activation energy in the Arrhenius expression and T0 is related to the so called thermodynamic Tg. Plots of logs vs 1/T are

graph, we can easily read the bulk resistance of the electrolyte system.

curved because of the reduced temperature (T-T0).

HFP confirms the partial crystallization of PVdF units present in the copolymer, to give an overall semi-crystalline morphology for PVdF-co-HFP [Saika, Kumar, 2004]. Presence of broad humps in the XRD pattern of PVAc confirms the complete amorphous nature of the polymer. It is observed that the characteristic peaks corresponding to the lithium salt in their respective electrolyte systems (A, B and C) were absent and it confirms the complete dissolution of the lithium salts in the complex matrix which implies that the salt do not have any separate phase in the electrolytes. The addition of plasticizer in the blend complex enhances the amorphous region thus permitting the free flow of ions from one site to another site; hence the overall ionic conductivity of the electrolyte has been significantly improved. According to Hodge et al. [Hodge et al., 1996] the ionic conduction in the polymer electrolytes occurs mostly in the amorphous region and it has been achieved by the addition of low molecular weight plasticizer. Further addition of inorganic filler into the polymer salt matrix would increase the dissolution of the charge carriers in the matrix; hence, the ionic conductivity was improved. The XRD pattern of the sample contains the filler BaTiO3 shows a broad hump confirms the further enhancement of the amorphous region in the polymer electrolyte complex systems.

Fig. 3. X-ray diffraction analysis of LiClO4, PVdF-co-HFP, PVAc and their complexes.

#### **3.3 Ac impedance analysis**

Ac studies are similar to the DC techniques in that the ratio of voltage to current is measured. For DC, this ratio provides the value of the resistance, R, measured in ohms.

HFP confirms the partial crystallization of PVdF units present in the copolymer, to give an overall semi-crystalline morphology for PVdF-co-HFP [Saika, Kumar, 2004]. Presence of broad humps in the XRD pattern of PVAc confirms the complete amorphous nature of the polymer. It is observed that the characteristic peaks corresponding to the lithium salt in their respective electrolyte systems (A, B and C) were absent and it confirms the complete dissolution of the lithium salts in the complex matrix which implies that the salt do not have any separate phase in the electrolytes. The addition of plasticizer in the blend complex enhances the amorphous region thus permitting the free flow of ions from one site to another site; hence the overall ionic conductivity of the electrolyte has been significantly improved. According to Hodge et al. [Hodge et al., 1996] the ionic conduction in the polymer electrolytes occurs mostly in the amorphous region and it has been achieved by the addition of low molecular weight plasticizer. Further addition of inorganic filler into the polymer salt matrix would increase the dissolution of the charge carriers in the matrix; hence, the ionic conductivity was improved. The XRD pattern of the sample contains the filler BaTiO3 shows a broad hump confirms the further enhancement of the amorphous

Fig. 3. X-ray diffraction analysis of LiClO4, PVdF-co-HFP, PVAc and their complexes.

Ac studies are similar to the DC techniques in that the ratio of voltage to current is measured. For DC, this ratio provides the value of the resistance, R, measured in ohms.

region in the polymer electrolyte complex systems.

**3.3 Ac impedance analysis** 

For AC the ratio gives an analogous quantity, the impedance, Z, also measured in ohms. The impedance contains four main contributions; these are from resistance, capacitance, constant phase elements, and inductance. The induction is not an important factor for the polymer electrolytes although it can play a role in other electrochemical applications of polymers.

Measurement of the impedance as a function of frequency is called impedance spectroscopy. In general, impedance is complex quantity, in which the real and the imaginary parts are labelled Z' and Z" respectively. In the complex impedance plot, the real quantity Z' (X-axis) is plotted against Z" (Y-axis) which displayed the polymer electrolytes characteristics as an arc followed by the linear spike is straight line inclined to the real axis. From the plotted graph, we can easily read the bulk resistance of the electrolyte system.

The complex impedance plot of the PVAc/PVdF-co-HFP/LiClO4 electrolyte is shown in Fig.4a. Figure shows the semicircular portion which is mainly due to the parallel combination of the geometrical capacitance, Cg and the bulk resistance, R. When adding the plasticizer and the filler (Fig.4b.) into the electrolyte matrix, the impedance spectra shows only a linear spike which corresponds to the lower frequency region. It confirms the idea that the current carriers are ions and the majority of the conduction only by the ions not by the electrons. And the disappearance of the semicircular portion is may be due to the fact the corresponding characteristic frequency is higher than the frequency 300kHz and it is mainly depends on the instrument limit. From the obtained bulk resistance value, we can estimate the ionic conductivity value of the electrolyte system using the relation σ = l/Rb A , where, Rb is the bulk resistance of the electrolyte film, A is the area of the electrode surface and is the thickness of the electrolyte medium and σ is the ionic conductivity.

It is noted from the spectra that the addition of plasticizer (ethylene carbonate) in to the polymer salt matrix greatly being reduced the bulk resistance of the system; this is because of the high dielectric nature of the low molecular weight plasticizer. The addition of plasticizer would considerably enhance the amorphous phase of the polymer electrolyte which will improve the ionic conductivity of the system. However, the gain in conductivity is adversely associated with a loss of the mechanical properties and by a loss of the compatibility with the lithium electrode, both effects resulting in serious problems since they affect the battery cycle life and increase the safety hazard. So it is necessary to identify the solid additives which would not affect the mechanical stability and interfacial stability of the electrolyte, at the same time will enhances the ionic conductivity. The addition of solid additives should improve the amorphicity of the electrolyte at room temperature. The addition of solid additives in the present study, such as nano filler BaTiO3 highly being enhanced the amourphicity of the electrolyte medium, hence the room temperature ionic conductivity and the interfacial stability of the electrode-electrolyte interface is increased. The ceramic dispersed electrolyte shows good thermal stability. The BaTiO3 incorporated sample is thermally stable up to 320 ºC. The temperature dependence of the conductivity is given by the Vogel–Tamman–Fulcher (VTF) equation σ = σ0 exp (-B/T-T0), where σ0 is the pre-exponential factor, B should not be confused with an activation energy in the Arrhenius expression and T0 is related to the so called thermodynamic Tg. Plots of logs vs 1/T are curved because of the reduced temperature (T-T0).

Surface Analysis Studies on Polymer Electrolyte

**3.4 TG/DTA analysis** 

**3.5 SEM analysis** 

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 685

Thermo gravimetric analysis /differential thermal analysis have been used widely to study all physical processes involving the weight changes. It is also used to investigate the thermal degradation, phase transitions and crystallization of polymers. Fig.5(a-c) shows the TG/DTA curves of PVAc/PVdF-co-HFP/LiClO4, PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3 polymer electrolytes. From the thermogram, it is observed that the sample A is thermally stable up to 238ºC. The sample starts to decompose at 238ºC, beyond which, there is a gradual weight loss of 20% in the temperature range 240-280ºC. DTA curve of the sample shows an exothermic peak at 265ºC, which is well correlated with the weight loss of the sample observed in TG curve. It is also observed that the complete decomposition of the sample takes place between 490-510ºC with the corresponding weight loss of about 80-90%. After 520ºC, there is no appreciable weight loss (Fig.5a). The remaining residue around 15% may be due to the formation of impure crystalline metal oxide and lithium fluoride. It is also observed from DTA curves that the exothermic peaks at 90, 225, and 445 ºC are concurrent with the weight losses observed in the TG trace. The sample (Fig.5b.) exhibit gradual weight loss of about 10-15%, which is due to the removal of the residual solvent and the moisture from the electrolyte sample in the temperature range 90-115 ºC. From the TG curve of the sample PVAc/PVdF-co-HFP/LiClO4/EC, it is observed that the decomposition occurred at 229ºC with the weight loss of about 20%. After the second decomposition, there is sudden weight loss of 40-45% in the temperature range 446-460 ºC for the electrolyte. The thermogram of the sample having BaTiO3 inert filler is shown Fig.5c. From the themogram, it is observed that the sample is thermally stable up to 320ºC. The sample exhibits gradual weight loss of about 8% in the temperature range 100-110 ºC, which is due to the removal of the residual solvent and the moisture. DTA curve of the sample shows an exothermic peak at 320ºC, which is well correlated with the weight loss of the sample observed in the TG curve. The remaining residue around 30% may be due to the presence of BaTiO3. It is noted from the above analysis that the additions of plasticizer into the polymer blend-salt matrix slightly influence the thermal stability of the electrolyte medium; however it has enhanced the ionic conductivity of the electrolyte. But, the addition of nano composite in to the matrix greatly being increased the thermal stability and the room temperature conductivity simultaneously. It is concluded that the incorporation/dispersion of inorganic filler in the

electrolyte significantly increased the thermal stability of the electrolyte membrane.

The scanning electron microscope (SEM) is one of the most versatile instruments available for the examination and analysis of the microstructure morphology of the conducting surfaces. Scanning electron microscope (SEM) image of PVAc-LiClO4, PVdF-co-HFP-LiClO4, PVAc/PVdF-co-HFP/LiClO4, PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4 /EC+PC/BaTiO3 electrolyte films are shown in Fig .6(a-e). Fig.6a. clearly shows smooth and uniform surface morphology of the PVAc- LiClO4. This smooth morphology confirms the complete amorphous nature of PVAc polymer and complete dissolution of the lithium salt, which also coincides with the XRD result. Fig.6b shows the photograph of PVdF-co-HFP-LiClO4 salt complex with maximum number of pores giving rise to high ionic conductivity. Presence of the spherical grains in the microstructure image

Fig. 4a. Room temperature complex ac impedance spectrum of PVAc/PVdF-co-HFP/LiClO4electrolyte

Fig. 4b. Room temperature complex ac impedance spectra of PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3 electrolyte

## **3.4 TG/DTA analysis**

684 Scanning Electron Microscopy

Fig. 4a. Room temperature complex ac impedance spectrum of PVAc/PVdF-co-

Fig. 4b. Room temperature complex ac impedance spectra of PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3 electrolyte

HFP/LiClO4electrolyte

Thermo gravimetric analysis /differential thermal analysis have been used widely to study all physical processes involving the weight changes. It is also used to investigate the thermal degradation, phase transitions and crystallization of polymers. Fig.5(a-c) shows the TG/DTA curves of PVAc/PVdF-co-HFP/LiClO4, PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3 polymer electrolytes. From the thermogram, it is observed that the sample A is thermally stable up to 238ºC. The sample starts to decompose at 238ºC, beyond which, there is a gradual weight loss of 20% in the temperature range 240-280ºC. DTA curve of the sample shows an exothermic peak at 265ºC, which is well correlated with the weight loss of the sample observed in TG curve. It is also observed that the complete decomposition of the sample takes place between 490-510ºC with the corresponding weight loss of about 80-90%. After 520ºC, there is no appreciable weight loss (Fig.5a). The remaining residue around 15% may be due to the formation of impure crystalline metal oxide and lithium fluoride. It is also observed from DTA curves that the exothermic peaks at 90, 225, and 445 ºC are concurrent with the weight losses observed in the TG trace. The sample (Fig.5b.) exhibit gradual weight loss of about 10-15%, which is due to the removal of the residual solvent and the moisture from the electrolyte sample in the temperature range 90-115 ºC. From the TG curve of the sample PVAc/PVdF-co-HFP/LiClO4/EC, it is observed that the decomposition occurred at 229ºC with the weight loss of about 20%. After the second decomposition, there is sudden weight loss of 40-45% in the temperature range 446-460 ºC for the electrolyte. The thermogram of the sample having BaTiO3 inert filler is shown Fig.5c. From the themogram, it is observed that the sample is thermally stable up to 320ºC. The sample exhibits gradual weight loss of about 8% in the temperature range 100-110 ºC, which is due to the removal of the residual solvent and the moisture. DTA curve of the sample shows an exothermic peak at 320ºC, which is well correlated with the weight loss of the sample observed in the TG curve. The remaining residue around 30% may be due to the presence of BaTiO3. It is noted from the above analysis that the additions of plasticizer into the polymer blend-salt matrix slightly influence the thermal stability of the electrolyte medium; however it has enhanced the ionic conductivity of the electrolyte. But, the addition of nano composite in to the matrix greatly being increased the thermal stability and the room temperature conductivity simultaneously. It is concluded that the incorporation/dispersion of inorganic filler in the electrolyte significantly increased the thermal stability of the electrolyte membrane.

#### **3.5 SEM analysis**

The scanning electron microscope (SEM) is one of the most versatile instruments available for the examination and analysis of the microstructure morphology of the conducting surfaces. Scanning electron microscope (SEM) image of PVAc-LiClO4, PVdF-co-HFP-LiClO4, PVAc/PVdF-co-HFP/LiClO4, PVAc/PVdF-co-HFP/LiClO4/EC and PVAc/PVdF-co-HFP/LiClO4 /EC+PC/BaTiO3 electrolyte films are shown in Fig .6(a-e). Fig.6a. clearly shows smooth and uniform surface morphology of the PVAc- LiClO4. This smooth morphology confirms the complete amorphous nature of PVAc polymer and complete dissolution of the lithium salt, which also coincides with the XRD result. Fig.6b shows the photograph of PVdF-co-HFP-LiClO4 salt complex with maximum number of pores giving rise to high ionic conductivity. Presence of the spherical grains in the microstructure image

Surface Analysis Studies on Polymer Electrolyte

confirmed from FTIR analysis.

**3.6 AFM analysis** 

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 687

increased number of porosity leads to entrapment of large volumes of the liquid in the pores accounting for the increased conductivity. The interconnected microspores in the membrane helped in absorbing liquid electrolytes and hence the ionic conductivity of the membrane is enhanced. The presence of pores in the microstructure is mainly due to the solvent removal and increased amorphous region and solvent retention ability in the electrolyte system. Surface images of the samples BaTiO3 is shown in Fig.6e. The pores in the complexes are responsible for entrapping the large volume of the solution (plasticizer +salt) in the cavities accounting for the enhanced ionic conductivity. It is observed from image that the membrane has numerous number of randomly distributed spherical grains and shows maximum number of pores with very small size of the order of 50-100nm. The smooth surface of the sample reveals that the polymers and salt used in this study have a good compatible nature and the light gray region indicates the presence of plasticizer rich medium which assists for ionic motion. It is also studied that the content BaTiO3 increases beyond certain percentage the film surface becomes rough above the optimum level the grain size increases, with a reduction in the number of grain aggregates, that tend to restrict the ionic movement. Finally, the SEM photograph of the polymer electrolyte indicates good compatibility of these two polymers and the other constituents which are used in the electrolyte preparation. The enhancement of the amorphous region in the matrix has also been confirmed from the images. The miscibility of these two polymers has also been

An AFM is a mechanical imaging instrument, which is used to obtain the three dimensional topography images of the samples. In the present study, the scanning probe spectroscopic method was used to measure the pore size of the prepared sample as well as the roughnees factor of the sample. The two dimensional and three dimensional topography images of PVAc/PVdF-co-HFP/LiClO4 complex are shown in Fig.7a. This image clearly shows the presence of pores within the scanned area of 3x3µm and the measured pore size of the sample is approximately 600nm. The size of the chain segment is also obtained and it is in the order of 688 nm. The root mean square (rms) roughness of the topography image over the scanned area is found to be 122 nm. The topography image of the sample PVAc/PVdF-co-HFP/LiClO4/EC is shown in Fig.7b. The two dimensional image shows smooth surface. The modified surface image of the sample is mainly due to the addition of plasticizer which increases the amorphous phase of the matrix and hence the ionic conductivity. In addition, small pores are also observed in the surface entrapping the liquid solution, which are responsible for easy ionic movement. From the topography image we have determined the pore size of the order of 100nm which is in close agreement with the value obtained from SEM photograph. In addition, the rms roughness of the sample over the scanned area 1.4x1.4μm has been estimated and it is of the order of 53nm, it is quiet low when compare with sample without plasticizers. The micropores, amorphous phase and the chain segments of the plasticized polymer electrolytes are responsible for the enhancement of ionic conductivity. Two and three dimensional topographic images of the sample having BaTiO3 are shown in Fig.7c. The image shows the dispersion of the fillers and it also contains small pore with a size of 100nm entrapping the ionic liquid which assists for fast ionic motion. In addition, the rms roughness of the sample over the scanned area 1x1μm has been obtained and it is of the order of 4nm. It is noted that the sample contains BaTiO3 showed lower roughness value than the other two

Fig. 5. TG/DTA Thermal analysis of a) PVAc/PVdF-co-HFP/LiClO4; b) PVAc/PVdF-co-HFP/LiClO4/EC; c) PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3

of the samples A and B (Fig.6b and d) are belongs to the co-polymer and it means that the copolymer do not dissolve completely in the matrix which results, the membrane gets brittle nature. The appearance of number of uniform tracks of few micrometer sizes is responsible for the appreciable ionic conductivity of the electrolyte (Fig.6c). The maximum ionic conductivity of the polymer blend electrolyte also depends on the segmental motion of the PVAc and PVdF-co-HFP. The better miscibility of these two polymers can be depicted from the microstructural photograph. Fig.6d shows the scanning electron micrographs of PVAc/PVdF–co-HFP/LiClO4/EC-based electrolyte system. The micrograph shows the spherical grains, and they are uniformly distributed in the electrolyte system. It is observed that the numerous pores (dark region) with the size of 1–10μm are responsible for the high conductivity of the sample, i.e., the membrane shows highly porous structure. This increased number of porosity leads to entrapment of large volumes of the liquid in the pores accounting for the increased conductivity. The interconnected microspores in the membrane helped in absorbing liquid electrolytes and hence the ionic conductivity of the membrane is enhanced. The presence of pores in the microstructure is mainly due to the solvent removal and increased amorphous region and solvent retention ability in the electrolyte system. Surface images of the samples BaTiO3 is shown in Fig.6e. The pores in the complexes are responsible for entrapping the large volume of the solution (plasticizer +salt) in the cavities accounting for the enhanced ionic conductivity. It is observed from image that the membrane has numerous number of randomly distributed spherical grains and shows maximum number of pores with very small size of the order of 50-100nm. The smooth surface of the sample reveals that the polymers and salt used in this study have a good compatible nature and the light gray region indicates the presence of plasticizer rich medium which assists for ionic motion. It is also studied that the content BaTiO3 increases beyond certain percentage the film surface becomes rough above the optimum level the grain size increases, with a reduction in the number of grain aggregates, that tend to restrict the ionic movement. Finally, the SEM photograph of the polymer electrolyte indicates good compatibility of these two polymers and the other constituents which are used in the electrolyte preparation. The enhancement of the amorphous region in the matrix has also been confirmed from the images. The miscibility of these two polymers has also been confirmed from FTIR analysis.

#### **3.6 AFM analysis**

686 Scanning Electron Microscopy

(a)

(b) (c) Fig. 5. TG/DTA Thermal analysis of a) PVAc/PVdF-co-HFP/LiClO4; b) PVAc/PVdF-co-

of the samples A and B (Fig.6b and d) are belongs to the co-polymer and it means that the copolymer do not dissolve completely in the matrix which results, the membrane gets brittle nature. The appearance of number of uniform tracks of few micrometer sizes is responsible for the appreciable ionic conductivity of the electrolyte (Fig.6c). The maximum ionic conductivity of the polymer blend electrolyte also depends on the segmental motion of the PVAc and PVdF-co-HFP. The better miscibility of these two polymers can be depicted from the microstructural photograph. Fig.6d shows the scanning electron micrographs of PVAc/PVdF–co-HFP/LiClO4/EC-based electrolyte system. The micrograph shows the spherical grains, and they are uniformly distributed in the electrolyte system. It is observed that the numerous pores (dark region) with the size of 1–10μm are responsible for the high conductivity of the sample, i.e., the membrane shows highly porous structure. This

HFP/LiClO4/EC; c) PVAc/PVdF-co-HFP/LiClO4/EC+PC/BaTiO3

An AFM is a mechanical imaging instrument, which is used to obtain the three dimensional topography images of the samples. In the present study, the scanning probe spectroscopic method was used to measure the pore size of the prepared sample as well as the roughnees factor of the sample. The two dimensional and three dimensional topography images of PVAc/PVdF-co-HFP/LiClO4 complex are shown in Fig.7a. This image clearly shows the presence of pores within the scanned area of 3x3µm and the measured pore size of the sample is approximately 600nm. The size of the chain segment is also obtained and it is in the order of 688 nm. The root mean square (rms) roughness of the topography image over the scanned area is found to be 122 nm. The topography image of the sample PVAc/PVdF-co-HFP/LiClO4/EC is shown in Fig.7b. The two dimensional image shows smooth surface. The modified surface image of the sample is mainly due to the addition of plasticizer which increases the amorphous phase of the matrix and hence the ionic conductivity. In addition, small pores are also observed in the surface entrapping the liquid solution, which are responsible for easy ionic movement. From the topography image we have determined the pore size of the order of 100nm which is in close agreement with the value obtained from SEM photograph. In addition, the rms roughness of the sample over the scanned area 1.4x1.4μm has been estimated and it is of the order of 53nm, it is quiet low when compare with sample without plasticizers. The micropores, amorphous phase and the chain segments of the plasticized polymer electrolytes are responsible for the enhancement of ionic conductivity. Two and three dimensional topographic images of the sample having BaTiO3 are shown in Fig.7c. The image shows the dispersion of the fillers and it also contains small pore with a size of 100nm entrapping the ionic liquid which assists for fast ionic motion. In addition, the rms roughness of the sample over the scanned area 1x1μm has been obtained and it is of the order of 4nm. It is noted that the sample contains BaTiO3 showed lower roughness value than the other two

Surface Analysis Studies on Polymer Electrolyte

Membranes Using Scanning Electron Microscope and Atomic Force Microscope 689

Fig. 7. AFM images of a) PVAc/LiClO4; b) PVdF-co-HFP-LiClO4; c) PVAc/PVdF-co-

HFP/LiClO4; d) PVAc/PVdF-co-HFP/LiClO4/EC; e) PVAc/PVdF-co-HFP/LiClO4/EC+PC

samples which means that the incorporation of the fillers and the plasticizers are significantly improve d the amorphous phase in the matrix is helpful for the ionic movement.

(a) (b)

(c) (d)

Fig. 6. SEM images of a) PVAc/LiClO4; b) PVdF-co-HFP-LiClO4; c) PVAc/PVdF-co-HFP/LiClO4;d) PVAc/PVdF-co-HFP/LiClO4/EC; e) PVAc/PVdF-co-HFP/LiClO4/EC+PC

samples which means that the incorporation of the fillers and the plasticizers are significantly

(a) (b)

(c) (d)

(e)

HFP/LiClO4;d) PVAc/PVdF-co-HFP/LiClO4/EC; e) PVAc/PVdF-co-HFP/LiClO4/EC+PC

Fig. 6. SEM images of a) PVAc/LiClO4; b) PVdF-co-HFP-LiClO4; c) PVAc/PVdF-co-

improve d the amorphous phase in the matrix is helpful for the ionic movement.

Fig. 7. AFM images of a) PVAc/LiClO4; b) PVdF-co-HFP-LiClO4; c) PVAc/PVdF-co-HFP/LiClO4; d) PVAc/PVdF-co-HFP/LiClO4/EC; e) PVAc/PVdF-co-HFP/LiClO4/EC+PC

Surface Analysis Studies on Polymer Electrolyte

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## **4. Conclusion**

All the polymer electrolytes were prepared using solvent costing technique. The specific interactions of the constituents were confirmed using FTIR analysis. The enhanced amorphous region of the polymer electrolyte has been identified from X-ray diffraction analysis. The porous natures of the samples were identified using scanning electron microscope. The Atomic force microscope study was used to estimate the roughness factors of the scanned area. The thermal stability of the electrolyte samples were estimated using TG/DTA analysis. It is concluded that the addition of plasticizer (Ethylene Carbonate) and the dispersion of inorganic filler into the PVAc/PVdF-co-HFP/LiClO4 electrolyte system significantly improve the amourphicity of the medium, which will helped for easy ionic motion. These enhanced regions have been confirmed from the impedance and the surface image studies. The change in bulk resistance of the electrolytes mainly due to the interactions of the basic constituents which cause produce more amorphous phase in the matrix. It is no doubt about that the addition of plasticizers and the fillers are greatly enhanced the amorphous phase of the electrolyte, hence the ionic conductivity is improved. To conclude, polymer electrolyte for possible applications as in high energy density batteries has been identified in terms of parameters such as conductivity, thermal stability.

#### **5. References**


All the polymer electrolytes were prepared using solvent costing technique. The specific interactions of the constituents were confirmed using FTIR analysis. The enhanced amorphous region of the polymer electrolyte has been identified from X-ray diffraction analysis. The porous natures of the samples were identified using scanning electron microscope. The Atomic force microscope study was used to estimate the roughness factors of the scanned area. The thermal stability of the electrolyte samples were estimated using TG/DTA analysis. It is concluded that the addition of plasticizer (Ethylene Carbonate) and the dispersion of inorganic filler into the PVAc/PVdF-co-HFP/LiClO4 electrolyte system significantly improve the amourphicity of the medium, which will helped for easy ionic motion. These enhanced regions have been confirmed from the impedance and the surface image studies. The change in bulk resistance of the electrolytes mainly due to the interactions of the basic constituents which cause produce more amorphous phase in the matrix. It is no doubt about that the addition of plasticizers and the fillers are greatly enhanced the amorphous phase of the electrolyte, hence the ionic conductivity is improved. To conclude, polymer electrolyte for possible applications as in high energy density batteries has been identified in terms of parameters such as

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**34** 

*México* 

**Characterization of Ceramic Materials** 

**Synthesized by Mechanosynthesis** 

1Claudia A. Cortés-Escobedo1, Félix Sánchez-De Jesús2,\*,

*4Centro de Investigación y Estudios Avanzados del IPN, Unidad Querétaro,* 

*1Centro de Investigación e Innovación Tecnológica del IPN, 2Universidad Autónoma del Estado de Hidalgo-AACTyM, 3Instituto de Investigaciones en Materiales-UNAM,* 

Gabriel Torres-Villaseñor3, Juan Muñoz-Saldaña4 and Ana M. Bolarín-Miró2

The close relationship between processing, structure and properties of materials is well known. Some of the most useful tools to elucidate the best choice in processing for a given application are scanning electron microscopy (SEM), transmission electron microscopy (TEM), scanning transmission electron microscopy (STEM), selected area electron diffraction (SAED) and x-ray diffraction (XRD). In this chapter we will focus on the application of these techniques to the characterization of ceramic materials processed by mechanosynthesis,

The ceramics that are the focus of this chapter, lanthanum manganites, have a Perovskitestructure (ABO3), with the general chemical formula R1–xAxMnO3 (where R3+ = La, and A2+ = Ca and Sr). Perovskite structures have been extensively studied for almost 50 years (Coey & Viret, 1999). Since the initial discovery of their electrical and magnetic properties, the interest in these compounds has remained high, and they have been the focus of significant scientific activity throughout the past decade. This kind of ceramic material has a wide variety of applications due to its ionic conduction, magnetic, thermal and mechanical properties, etc. Furthermore, it is known that the physical properties of Perovskite manganites depend on many factors, such as external pressure (Hwang & Palstra, 1995; Neumeier et al., 1995), magnetic field (Asamitsu et al., 1995; Kuwahara et al., 1995), structure (Tokura et al., 1994) and chemical composition (Hwang et al., 1995; Mahesh et al., 1995; Schiffer et al., 1995). For example, the ionic conduction of lanthanum manganates has led to their use in oxygen sensors and solid oxide fuel cells (Shu et al., 2009). Their ionic conduction is due to punctual defects, in the form of oxygen vacancies, in the crystalline structure of the Perovskite lattice. The generation of punctual defects is activated by the addition of doping atoms, as well as changing synthesis precursors and by applying mechanical energy, all of which are typical processes going on during the high-energy ball-

evaluating the effect of the milling process on their physical properties.

milling process. These effects are described in detail in this chapter.

**1. Introduction** 

\*1Corresponding Author

**for Energy Applications** 

Xi, J. Qiu, X. Li, J. Tang, X. Zhu, W. Chen, L. PVDF–PEO blends based microporous polymer electrolyte: Effect of PEO on pore configurations and ionic conductivity, *J.Power Sources*, Vol.157, No.1, (June 2006), pp.501-506, ISSN 0378-7753

## **Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications**

1Claudia A. Cortés-Escobedo1, Félix Sánchez-De Jesús2,\*, Gabriel Torres-Villaseñor3, Juan Muñoz-Saldaña4 and Ana M. Bolarín-Miró2 *1Centro de Investigación e Innovación Tecnológica del IPN, 2Universidad Autónoma del Estado de Hidalgo-AACTyM, 3Instituto de Investigaciones en Materiales-UNAM, 4Centro de Investigación y Estudios Avanzados del IPN, Unidad Querétaro, México* 

## **1. Introduction**

694 Scanning Electron Microscopy

Xi, J. Qiu, X. Li, J. Tang, X. Zhu, W. Chen, L. PVDF–PEO blends based microporous polymer

*Sources*, Vol.157, No.1, (June 2006), pp.501-506, ISSN 0378-7753

electrolyte: Effect of PEO on pore configurations and ionic conductivity, *J.Power* 

The close relationship between processing, structure and properties of materials is well known. Some of the most useful tools to elucidate the best choice in processing for a given application are scanning electron microscopy (SEM), transmission electron microscopy (TEM), scanning transmission electron microscopy (STEM), selected area electron diffraction (SAED) and x-ray diffraction (XRD). In this chapter we will focus on the application of these techniques to the characterization of ceramic materials processed by mechanosynthesis, evaluating the effect of the milling process on their physical properties.

The ceramics that are the focus of this chapter, lanthanum manganites, have a Perovskitestructure (ABO3), with the general chemical formula R1–xAxMnO3 (where R3+ = La, and A2+ = Ca and Sr). Perovskite structures have been extensively studied for almost 50 years (Coey & Viret, 1999). Since the initial discovery of their electrical and magnetic properties, the interest in these compounds has remained high, and they have been the focus of significant scientific activity throughout the past decade. This kind of ceramic material has a wide variety of applications due to its ionic conduction, magnetic, thermal and mechanical properties, etc. Furthermore, it is known that the physical properties of Perovskite manganites depend on many factors, such as external pressure (Hwang & Palstra, 1995; Neumeier et al., 1995), magnetic field (Asamitsu et al., 1995; Kuwahara et al., 1995), structure (Tokura et al., 1994) and chemical composition (Hwang et al., 1995; Mahesh et al., 1995; Schiffer et al., 1995). For example, the ionic conduction of lanthanum manganates has led to their use in oxygen sensors and solid oxide fuel cells (Shu et al., 2009). Their ionic conduction is due to punctual defects, in the form of oxygen vacancies, in the crystalline structure of the Perovskite lattice. The generation of punctual defects is activated by the addition of doping atoms, as well as changing synthesis precursors and by applying mechanical energy, all of which are typical processes going on during the high-energy ballmilling process. These effects are described in detail in this chapter.

<sup>\*1</sup>Corresponding Author

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 697

A-sites of the Perovskite structure. Results of crystal structure analyses of calcium-doped lanthanum manganites, La1-xCaxMnO3 have been previously reported. The calcium-tolanthanum ratio *x* was varied from 0 to 1 in increments of 0.1, allowing the study of changes in crystal structure with different degrees of calcium substitution, from LaMnO3 (x=0) to CaMnO3 (x=1) (Lira-Hernández et al., 2010). Here we present a study to increase the calcium content while maintaining the orthorhombic in order to obtain the best ion-electronic conduction. The repercussions of the doping on the microstructural characteristics were

The third part of the chapter is dedicated to the process of consolidating manganite powder in order to see how the compaction and sintering process affects the physical properties of the ceramic. A study of the relationship between the crystalline structure, the mixed ionicelectronic conductivity and calcium content in calcium-doped lanthanum manganites, La1-

Finally, in the fourth part of the chapter, the interaction between lanthanum manganites and yttria-stabilized zirconia, which are used as cathode and electrolyte, respectively, in both solid oxide fuel cells and sensors is discussed. A reduction in ionic conductivity is observed at certain temperatures, due to the formation of a very stable pyrochlore phase (lanthanum zirconate). Diffusion from manganite to zirconia to form zirconate has been well demonstrated by backscattered electron microscopy, showing the microstructure of cubic zirconia, in contrast with tetragonal zirconia, lanthanum manganite and lanthanum

Mechanochemical processing (MCP), or mechanosynthesis, is a synthesis method that uses mechanical energy (e.g., ball milling) to activate chemical reactions and structural changes in powder mixtures. In some cases, milling is followed by a low-temperature heat treatment to complete the reaction. In this study, lanthanum manganites were produced from oxide precursors La2O3, Mn2Ox and CaO or SrO by means of high-energy ball millling, with a

Lanthanum manganites, La1-xMxMnO3 (0 < x < 1), where M was Ca+2 or Sr+2, were prepared using a SPEX 8000 D high-energy ball mill. Powders of Mn2Ox and La2O3 were mixed in stoichiometric proportion for obtaining different level of doped manganite. MCP was carried out at room temperature in air atmosphere, in hardened-steel vials with steel balls as milling elements; ball-to-powder weight ratio was 10:1 at different milling times. X-ray diffraction (XRD) was used to evaluate phase transformations as function of milling time. Morphology and particle size of the manganites were characterized by scanning electron microscopy (SEM). Particle size distribution was measured by a zeta size analyzer. Selected area electron diffraction (SAED) patterns, obtained using a transmission electron microscope, were indexed to identify the resultant crystalline phase. Rietveld refinements of the XRD patterns were performed with the purpose of identifying and quantifying the phases and to identify the structural changes in the starting oxides until the formation of

For applications as sensors, as well as cathodes in solid oxide fuel cells, ionic conductivity of lanthanum manganites is desired. However, ionic conductivity is favored by punctual

**2. Theoretical aspects of mechanochemical processing (MCP)** 

studied by means of XRD and TEM /SAED.

xCaxMnO3, is presented.

subsequent heat treatment.

zirconate.

manganites.

There are several methods to synthesize lanthanum manganites. The traditional method is via solid-state reaction of the components (mixed oxides route). Controlling the temperature during the solid state reaction is the major challenge to obtaining homogeneity in the stoichiometry, grain size, porosity and purity. Alternatively, chemical methods such as solgel (Zhou et al., 2010), solution combustion (Shinde et al., 2010), co-precipitation (Uskokovic & Drofenik, 2007), and others (Jafari et al., 2010) have been used, resulting in lanthanum manganites with a wide variety of physical properties.

Another technique, high-energy ball-milling, used to promote mechanosynthesis of nanostructured manganites, such as La1-xCaxMnO3, by mechanical activation of chloride and oxide compounds, has shown excellent results (Muroi et al., 2000; Bolarín et al., 2006, 2007). Zhang et al. (Zhang & Saito, 2000, Zhang et al., 2000) synthesized LaMnO3 Perovskites at room temperature by milling a mixture of Mn2O3 and La2O3 powders using a planetary ball mill. Additionally, K. Sato et al. (Sato et al., 2006) used an alternative mechanical synthesis route to produce fine LaMnO3 powder – compression and shear stress were repeatedly applied to a mixture of La2O3 and Mn3O4 using an attrition type milling apparatus.

Fig. 1. Variables used to maximize punctual defects, and therefore ionic conduction in doped lanthanum manganites.

The chapter is divided in four main parts. In the first, details of the mechanosynthesis of lanthanum manganites, showing the process parameters that are necessary to carry out the synthesis of this ceramic material without a posterior annealing treatment are presented. The effect of these process parameters on the crystalline structure of the Perovskite-type manganites and mixed ionic-electronic conductivity will be presented and discussed (Figure 1). Specifically, the synthesis of lanthanum manganese oxide (LaMnO3) by solid-state reaction using high-energy reactive ball milling from manganese oxides (MnO, Mn2O3, MnO2) mixed with lanthanum oxide in stoichiometric ratios, for different milling times is reported. This is a useful route used to maximize punctual defects while preserving the initial structure by using manganese oxides with different oxidation numbers for manganese (Mn2+, Mn3+ and Mn4+). The mechanical energy of the mechanosynthesis processs induces a higher level of intrinsic defects than can be created by other synthesis methods (Cortés-Escobedo et al., 2008). In addition, a comparative study of mechanosynthesis with the solid-state reaction at high temperature is presented. In this section, we demonstrate the usefulness of electronic microscopy and x-ray diffraction to provide a structural description of the synthesized material.

In the second part of the chapter, the effects of doping on the structure and physical properties of lanthanum manganite is emphasized, using Ca+2 and Sr+2 as doping atoms in

There are several methods to synthesize lanthanum manganites. The traditional method is via solid-state reaction of the components (mixed oxides route). Controlling the temperature during the solid state reaction is the major challenge to obtaining homogeneity in the stoichiometry, grain size, porosity and purity. Alternatively, chemical methods such as solgel (Zhou et al., 2010), solution combustion (Shinde et al., 2010), co-precipitation (Uskokovic & Drofenik, 2007), and others (Jafari et al., 2010) have been used, resulting in lanthanum

Another technique, high-energy ball-milling, used to promote mechanosynthesis of nanostructured manganites, such as La1-xCaxMnO3, by mechanical activation of chloride and oxide compounds, has shown excellent results (Muroi et al., 2000; Bolarín et al., 2006, 2007). Zhang et al. (Zhang & Saito, 2000, Zhang et al., 2000) synthesized LaMnO3 Perovskites at room temperature by milling a mixture of Mn2O3 and La2O3 powders using a planetary ball mill. Additionally, K. Sato et al. (Sato et al., 2006) used an alternative mechanical synthesis route to produce fine LaMnO3 powder – compression and shear stress were repeatedly

> Doped level of La1-xCaxMnO3

Compacts: physical properties

porosity

mixed conductivity

0<x<1

applied to a mixture of La2O3 and Mn3O4 using an attrition type milling apparatus.

Doping atoms in A site

Sr2+ Ca+2

Fig. 1. Variables used to maximize punctual defects, and therefore ionic conduction in

provide a structural description of the synthesized material.

The chapter is divided in four main parts. In the first, details of the mechanosynthesis of lanthanum manganites, showing the process parameters that are necessary to carry out the synthesis of this ceramic material without a posterior annealing treatment are presented. The effect of these process parameters on the crystalline structure of the Perovskite-type manganites and mixed ionic-electronic conductivity will be presented and discussed (Figure 1). Specifically, the synthesis of lanthanum manganese oxide (LaMnO3) by solid-state reaction using high-energy reactive ball milling from manganese oxides (MnO, Mn2O3, MnO2) mixed with lanthanum oxide in stoichiometric ratios, for different milling times is reported. This is a useful route used to maximize punctual defects while preserving the initial structure by using manganese oxides with different oxidation numbers for manganese (Mn2+, Mn3+ and Mn4+). The mechanical energy of the mechanosynthesis processs induces a higher level of intrinsic defects than can be created by other synthesis methods (Cortés-Escobedo et al., 2008). In addition, a comparative study of mechanosynthesis with the solid-state reaction at high temperature is presented. In this section, we demonstrate the usefulness of electronic microscopy and x-ray diffraction to

In the second part of the chapter, the effects of doping on the structure and physical properties of lanthanum manganite is emphasized, using Ca+2 and Sr+2 as doping atoms in

manganites with a wide variety of physical properties.

doped lanthanum manganites.

Oxidation number for manganese in B site (precursor-Mn2Ox: M+2, Mn+3 y Mn+4)

Mechanosynthesis of LaMnO3

milling time

A-sites of the Perovskite structure. Results of crystal structure analyses of calcium-doped lanthanum manganites, La1-xCaxMnO3 have been previously reported. The calcium-tolanthanum ratio *x* was varied from 0 to 1 in increments of 0.1, allowing the study of changes in crystal structure with different degrees of calcium substitution, from LaMnO3 (x=0) to CaMnO3 (x=1) (Lira-Hernández et al., 2010). Here we present a study to increase the calcium content while maintaining the orthorhombic in order to obtain the best ion-electronic conduction. The repercussions of the doping on the microstructural characteristics were studied by means of XRD and TEM /SAED.

The third part of the chapter is dedicated to the process of consolidating manganite powder in order to see how the compaction and sintering process affects the physical properties of the ceramic. A study of the relationship between the crystalline structure, the mixed ionicelectronic conductivity and calcium content in calcium-doped lanthanum manganites, La1 xCaxMnO3, is presented.

Finally, in the fourth part of the chapter, the interaction between lanthanum manganites and yttria-stabilized zirconia, which are used as cathode and electrolyte, respectively, in both solid oxide fuel cells and sensors is discussed. A reduction in ionic conductivity is observed at certain temperatures, due to the formation of a very stable pyrochlore phase (lanthanum zirconate). Diffusion from manganite to zirconia to form zirconate has been well demonstrated by backscattered electron microscopy, showing the microstructure of cubic zirconia, in contrast with tetragonal zirconia, lanthanum manganite and lanthanum zirconate.

## **2. Theoretical aspects of mechanochemical processing (MCP)**

Mechanochemical processing (MCP), or mechanosynthesis, is a synthesis method that uses mechanical energy (e.g., ball milling) to activate chemical reactions and structural changes in powder mixtures. In some cases, milling is followed by a low-temperature heat treatment to complete the reaction. In this study, lanthanum manganites were produced from oxide precursors La2O3, Mn2Ox and CaO or SrO by means of high-energy ball millling, with a subsequent heat treatment.

Lanthanum manganites, La1-xMxMnO3 (0 < x < 1), where M was Ca+2 or Sr+2, were prepared using a SPEX 8000 D high-energy ball mill. Powders of Mn2Ox and La2O3 were mixed in stoichiometric proportion for obtaining different level of doped manganite. MCP was carried out at room temperature in air atmosphere, in hardened-steel vials with steel balls as milling elements; ball-to-powder weight ratio was 10:1 at different milling times. X-ray diffraction (XRD) was used to evaluate phase transformations as function of milling time. Morphology and particle size of the manganites were characterized by scanning electron microscopy (SEM). Particle size distribution was measured by a zeta size analyzer. Selected area electron diffraction (SAED) patterns, obtained using a transmission electron microscope, were indexed to identify the resultant crystalline phase. Rietveld refinements of the XRD patterns were performed with the purpose of identifying and quantifying the phases and to identify the structural changes in the starting oxides until the formation of manganites.

For applications as sensors, as well as cathodes in solid oxide fuel cells, ionic conductivity of lanthanum manganites is desired. However, ionic conductivity is favored by punctual

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 699

/30min 23 23 23 23

3 23 23

*LaMnO La O Mn O LaMnO*

+ + →

Fig. 2. X-ray diffraction patterns obtained from La2O3 + MnO powder mixtures before milling, milled for 630 min, and heat treated at 1000°C for 600 min (L: La2O3, JCPDS No. 05- 0602; MII: MnO, JCPDS No. 07-0230; LM: La1-xMn1-zO3). The pattern for MnII+L milled for

Fig. 3*.* Left: X-ray diffraction patterns obtained from a mixture of La2O3 and Mn2O3 powder for various milling times (MIII: Mn2O3, JCPDS No. 24-0508). Right: mol fraction of phases as analyzed using Rietveld refinement of the x-ray diffraction patterns. LM corresponds to the lanthanum manganite phases irrespective of the structure (Cortés-Escobedo et al., 2007).

630 min was rescaled for easier comparison (Cortés-Escobedo et al., 2007).

( )

0.5 0.5 ( )

*La O Mn O La O Mn O*

+ →+

*MCP*

*MCP remnant*

*LaMnO La O Mn O*

( )

→ ++

3 23 23 3

*MCP remnant fine grained*

/210min

*fine grained*

*fine grained*

(3)

/60min

defects in the structure of the Perovskite ceramic. For magnetic applications, it has been discussed that punctual defects have an effect on the colossal magnetoresistance of the material. For these reasons, punctual defects were induced, firstly by changing the oxidation number of manganese in precursor manganese oxide in Mn+2, Mn+3 and Mn+4, it is to say, by inducing intrinsic defects and secondly by inserting calcium or strontium atoms which will suit in A sites (in place of lanthanum), inducing extrinsic defects in the atomic structure.

#### **3. Defects in ceramic materials**

In general, properties of solid ionics and in particular ceramic materials depend on crystalline structure and chemical bonding, and sometimes these properties can be improved by means of the processing or synthesis route employed. One of the ways to improve mechanical, electrical, magnetic, and structural properties is by inducing defects in crystalline structure. Defects are deviations from ideality in crystals and can be volumetric (3-dimensional), surface (2-dimensional), or punctual (1-dimensional). Punctual defects in ionic solids can be intrinsic, this is, due to disorder in atomic arrangement. Cations may be shifted to interstitial sites (Frenkel defect) or there may be a lack of an anion-cation pair (Schottky defect), both cases leading to a vacancy. Punctual defects can also be extrinsic, due to a substitution of an atom with another that has a different size and shell configuration, resulting in vacancy generation to maintain electroneutrality. The presence of intrinsic or extrinsic defects in ceramics promotes mobility of ions driven by local electrical charges present in vacancies, improving ionic conduction properties.

#### **3.1 Intrinsic defects in ceramic materials**

Intrinsic defects can be promoted by different synthesis or processing routes. One way to promote intrinsic defects is synthesizing by mechanical milling. This process generates nanometric-sized particles and disorder at the atomic level. It is important to consider that vacancies in the crystalline structure can also be induced by using precursors composed of cations with non-stoichiometric oxidation numbers.

With the intention of promoting intrinsic defects, a stoichiometric mixture of MnO and La2O3 were milled to follow the reaction:

$$\text{La}\_2\text{O}\_3 + 2\text{MnO} \xrightarrow{\text{MCP}} 2\text{La}\text{MnO}\_3 + 2\text{Mn}'\_{\text{Mn}} + 2V\_\text{O} \tag{1}$$

After 630 min of milling, the x-ray diffraction pattern (Figure 2) reveals the amorphization of the precursor mixture. But after 600 min of heat treatment at 1000°C, orthorhombic LaMnO3 is formed. This can be represented by the following reaction:

$$\begin{array}{rcl} 0.5 \text{La}\_2\text{O}\_3 + \text{MnO} + 0.5 \text{O}\_2 & \stackrel{\text{MCP}/630 \text{min}}{\rightarrow} & \text{(0.5La}\_2\text{O}\_3 + \text{MnO})\_{\text{fnc}-\text{gained}} + 0.5 \text{O}\_2 & \stackrel{1000^\circ \text{C}/600 \text{min}}{\rightarrow} & \text{LaMnO}\_3 & \text{(2)} \end{array}$$

A stoichiometric mixture of precursors Mn2O3 and La2O3 were milled from 0 to 540 min, and x-ray diffraction patterns are shown in Figure 3. Formation of lanthanum manganite can be detected after 60 min, and the precursors were completely consumed after 210 min of milling, following the reaction:

defects in the structure of the Perovskite ceramic. For magnetic applications, it has been discussed that punctual defects have an effect on the colossal magnetoresistance of the material. For these reasons, punctual defects were induced, firstly by changing the oxidation number of manganese in precursor manganese oxide in Mn+2, Mn+3 and Mn+4, it is to say, by inducing intrinsic defects and secondly by inserting calcium or strontium atoms which will suit in A sites (in place of lanthanum), inducing extrinsic defects in the

In general, properties of solid ionics and in particular ceramic materials depend on crystalline structure and chemical bonding, and sometimes these properties can be improved by means of the processing or synthesis route employed. One of the ways to improve mechanical, electrical, magnetic, and structural properties is by inducing defects in crystalline structure. Defects are deviations from ideality in crystals and can be volumetric (3-dimensional), surface (2-dimensional), or punctual (1-dimensional). Punctual defects in ionic solids can be intrinsic, this is, due to disorder in atomic arrangement. Cations may be shifted to interstitial sites (Frenkel defect) or there may be a lack of an anion-cation pair (Schottky defect), both cases leading to a vacancy. Punctual defects can also be extrinsic, due to a substitution of an atom with another that has a different size and shell configuration, resulting in vacancy generation to maintain electroneutrality. The presence of intrinsic or extrinsic defects in ceramics promotes mobility of ions driven by local electrical charges

Intrinsic defects can be promoted by different synthesis or processing routes. One way to promote intrinsic defects is synthesizing by mechanical milling. This process generates nanometric-sized particles and disorder at the atomic level. It is important to consider that vacancies in the crystalline structure can also be induced by using precursors composed of

With the intention of promoting intrinsic defects, a stoichiometric mixture of MnO and

After 630 min of milling, the x-ray diffraction pattern (Figure 2) reveals the amorphization of the precursor mixture. But after 600 min of heat treatment at 1000°C, orthorhombic LaMnO3

*MCP C La O MnO O La O MnO fine grained <sup>O</sup> LaMnO* °

++ → + + → <sup>−</sup> (2)

2 3 <sup>2</sup> 2 3 <sup>2</sup> <sup>3</sup> 0.5 0.5 (0.5 ) 0.5

A stoichiometric mixture of precursors Mn2O3 and La2O3 were milled from 0 to 540 min, and x-ray diffraction patterns are shown in Figure 3. Formation of lanthanum manganite can be detected after 60 min, and the precursors were completely consumed after 210 min of

��� ������� � ������ � ���

/630min 1000 /600min

∙

(1)

���

atomic structure.

**3. Defects in ceramic materials** 

present in vacancies, improving ionic conduction properties.

**3.1 Intrinsic defects in ceramic materials** 

La2O3 were milled to follow the reaction:

milling, following the reaction:

cations with non-stoichiometric oxidation numbers.

����� � ����

is formed. This can be represented by the following reaction:

$$\begin{array}{rcl} 0.5 \text{La}\_2\text{O}\_3 + 0.5 \text{Mn}\_2\text{O}\_3 & \stackrel{\text{MCP}/30 \text{min}}{\rightarrow} & \text{(La}\_2\text{O}\_3 + \text{Mn}\_2\text{O}\_3\text{)}\_{\text{fine grain}} \\ \text{ } & \stackrel{\text{MCP}/60 \text{min}}{\rightarrow} & \text{La} \text{MnO}\_3 + \text{(La}\_2\text{O}\_3 + \text{Mn}\_2\text{O}\_3)^{\text{reeman}mt}\_{\text{fine grain}} \\ \text{ } & \text{La} \text{MnO}\_3 + \text{(La}\_2\text{O}\_3 + \text{Mn}\_2\text{O}\_3)^{\text{reram}mt}\_{\text{fine grain}} & \stackrel{\text{MCP}/210 \text{min}}{\rightarrow} & \text{La} \text{MnO}\_3 \end{array} \tag{3}$$

Fig. 2. X-ray diffraction patterns obtained from La2O3 + MnO powder mixtures before milling, milled for 630 min, and heat treated at 1000°C for 600 min (L: La2O3, JCPDS No. 05- 0602; MII: MnO, JCPDS No. 07-0230; LM: La1-xMn1-zO3). The pattern for MnII+L milled for 630 min was rescaled for easier comparison (Cortés-Escobedo et al., 2007).

Fig. 3*.* Left: X-ray diffraction patterns obtained from a mixture of La2O3 and Mn2O3 powder for various milling times (MIII: Mn2O3, JCPDS No. 24-0508). Right: mol fraction of phases as analyzed using Rietveld refinement of the x-ray diffraction patterns. LM corresponds to the lanthanum manganite phases irrespective of the structure (Cortés-Escobedo et al., 2007).

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 701

Thermodynamic calculations for ΔG and ΔH were made using HSC Chemistry 5.112 software (Figure 9) in order to know the feasibility of the formation of LaMnO3 in the three cases, and the reaction with lowest ΔG was MnO2 + La2O3 = LaMnO3, opposite to what was observed for the complete formation of the manganite in the milling time experiments. This difference can be attributed to structural and redox transformations required to obtain the

Fig. 5. Left: X-ray diffraction patterns obtained from a La2O3+MnO2 powder mixture at different milling times (MIV: MnO2, JCPDS No.72-1984). Right: mol fraction of phases as analysed by the Rietveld x-ray diffraction pattern refinement, LM corresponds to the lanthanum manganite phases irrespective to the structure. (Cortés-Escobedo et al., 2007)

Fig. 6. Typical SEM micrographs of powder mixtures from different Mn precursors and milling times: LM2 1.5 h: La2O3+MnO for 90 min; LM2 6 h: La2O3+MnO for 360 min; LM3 1.5 h: La2O3+Mn2O3 at 90 min; LM3 6 h: La2O3+Mn2O3 at 360 min; LM4 1.5 h: La2O3+MnO2 at 90

min; LM4 6 h: La2O3+MnO2 at 360 min. (Cortés-Escobedo et al., 2007).

2 HSC Chemistry 5.11, Copyright (C) Outokumpu Research Oy, Pori, Finland, A. Roine.

final Perovskite structure.

Figure 4 shows the electron diffraction patterns for lanthanum manganite obtained from its stoichiometric precursors, and the rings correspond to reflections from the orthorhombic and rhombohedral structure of lanthanum manganite.

Fig. 4. Electron diffraction patterns of LaMnO3 milled for 7 h, with the measured interplanar spacings (d) noted (Bolarín et al., 2007).

Finally, milling was performed using a mixture according to the following reaction:

$$2La\_2O\_3 + 2MnO\_2 \xrightarrow{MCP} 2LaMnO\_3 + \frac{1}{2}O\_2 + 2Mn^{\cdot}\_{Mn} + V\_O^{\times} \tag{4}$$

After 270 of milling time LaMnO3 is completely formed following the steps:

$$\begin{array}{rcl} \text{0.5La}\_2\text{O}\_3 + \text{MnO}\_2 & \rightarrow & \text{(La}\_2\text{O}\_3 + \text{MnO}\_2\text{)}\_{\text{fine gained}} & \rightarrow & \text{(La}\_2\text{O}\_3 + \text{MnO}\_2\text{)}\_{\text{fine gained}}\\ + \text{(LaMnO}\_3\text{)}\_{\text{fine gained}} & \rightarrow & \text{LaMnO}\_3 \end{array} \tag{5}$$

The corresponding x-ray diffraction patterns are shown in Figure 5. After Rietveld refinement, three different structures of lanthanum manganite were identified: orthorhombic, rhombohedral and cubic, in varying proportions.

A discussion of the structures can be read in (Cortés-Escobedo et al., 2008). The particle size and morphology of the milled powders as a function of time and precursor is shown in Figure 6. The decrease in particle size as a function of time is consistent with the diminishing crystallite size observed with x-ray diffraction (Figures 3-5).

Figure 7 shows the particle size as measured by laser diffraction and image analysis of the SEM photomicrographs. In both cases, after 100 min of milling time particles of 500 nm are predominant.

In Figure 8 photomicrographs of the crystallite size for the La2O3+Mn2O3 mixture milled for 90 min show crystallites from 5 to 20 nm in size and the deformation of the particles.

Figure 4 shows the electron diffraction patterns for lanthanum manganite obtained from its stoichiometric precursors, and the rings correspond to reflections from the orthorhombic

Fig. 4. Electron diffraction patterns of LaMnO3 milled for 7 h, with the measured interplanar

��� ������� <sup>+</sup> �

/90min /120min 23 2 23 2 23 2

The corresponding x-ray diffraction patterns are shown in Figure 5. After Rietveld refinement, three different structures of lanthanum manganite were identified:

A discussion of the structures can be read in (Cortés-Escobedo et al., 2008). The particle size and morphology of the milled powders as a function of time and precursor is shown in Figure 6. The decrease in particle size as a function of time is consistent with the

Figure 7 shows the particle size as measured by laser diffraction and image analysis of the SEM photomicrographs. In both cases, after 100 min of milling time particles of 500 nm are

In Figure 8 photomicrographs of the crystallite size for the La2O3+Mn2O3 mixture milled for

90 min show crystallites from 5 to 20 nm in size and the deformation of the particles.

0.5 ( ) ( )

*La O MnO La O MnO La O MnO*

+→+ → +

�

�� + �����

<sup>∙</sup> + ��

*fine grained fine grained*

� (4)

(5)

Finally, milling was performed using a mixture according to the following reaction:

���

After 270 of milling time LaMnO3 is completely formed following the steps:

*MCP MCP*

and rhombohedral structure of lanthanum manganite.

spacings (d) noted (Bolarín et al., 2007).

*fine grained*

+ →

( )

predominant.

����� + �����

/270min 3 3

orthorhombic, rhombohedral and cubic, in varying proportions.

diminishing crystallite size observed with x-ray diffraction (Figures 3-5).

*MCP*

*LaMnO LaMnO*

Thermodynamic calculations for ΔG and ΔH were made using HSC Chemistry 5.112 software (Figure 9) in order to know the feasibility of the formation of LaMnO3 in the three cases, and the reaction with lowest ΔG was MnO2 + La2O3 = LaMnO3, opposite to what was observed for the complete formation of the manganite in the milling time experiments. This difference can be attributed to structural and redox transformations required to obtain the final Perovskite structure.

Fig. 5. Left: X-ray diffraction patterns obtained from a La2O3+MnO2 powder mixture at different milling times (MIV: MnO2, JCPDS No.72-1984). Right: mol fraction of phases as analysed by the Rietveld x-ray diffraction pattern refinement, LM corresponds to the lanthanum manganite phases irrespective to the structure. (Cortés-Escobedo et al., 2007)

Fig. 6. Typical SEM micrographs of powder mixtures from different Mn precursors and milling times: LM2 1.5 h: La2O3+MnO for 90 min; LM2 6 h: La2O3+MnO for 360 min; LM3 1.5 h: La2O3+Mn2O3 at 90 min; LM3 6 h: La2O3+Mn2O3 at 360 min; LM4 1.5 h: La2O3+MnO2 at 90 min; LM4 6 h: La2O3+MnO2 at 360 min. (Cortés-Escobedo et al., 2007).

<sup>2</sup> HSC Chemistry 5.11, Copyright (C) Outokumpu Research Oy, Pori, Finland, A. Roine.


$$2La\_2O\_3 + 2Mn\_2O\_3 \xrightarrow{CaU} 2LaMnO\_3 + 2Ca\_{La}^{\prime} + V\_o^{\text{xx}} \tag{6}$$

$$\frac{1}{2}\frac{(1-\chi)}{2}La\_2O\_3 + \frac{1}{2}Mn\_2O\_3 + \chi CaO - \frac{\chi}{4}O\_2 \xrightarrow{MCP} La\_{1-\chi}Ca\_2MnO\_3 \tag{7}$$

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 705

Figures 17-19 show the morphology of the nanoparticles aggregated after 4.5 and 7 h of milling of the (Mn2O3+CaO+La2O3) mixtures. The morphology is the same for compositions x = 0.3 and 0.4, but for x = 0.8 after 7 h of milling a different morphology is observed - larger

Figure 20 shows the morphology for the extreme case of x = 1, the Mn2O3+CaO mixture after 4.5 h of milling, and the absence of large agglomerates is noticeable; the particle size is on

> Fig. 13. Lanthanum oxide precursor for the reaction of formation of La(1-x)CaxMnO3.

> > milling.

of milling (x = 0.4).

Fig. 14. Manganesum oxide precursor (Mn2O3) for the reaction of formation of

La(1-x)CaxMnO3.

Fig. 16. (Mn2O3+La2O3) mixture after 7 h of

Fig. 18. (Mn2O3+CaO+La2O3) mixture after 7 h

and continuous particles with softer edges.

Fig. 15. (Mn2O3+La2O3) mixture after 4.5 h of

Fig. 17. (Mn2O3+CaO+La2O3) mixture after

4.5 h of milling (x = 0.3).

the order of nanometers.

Fig. 12. Calcium oxide precursor for the reaction of formation of La(1-x)CaxMnO3.

milling.

After 7 h of milling, there is a shift to the right of the mean peak of the lanthanum manganite as the amount of doping is increased, indicating a decrease in the interplanar distance, corresponding to deformation of the network as proposed in Figure 11.

In Figures 12- 14*,* morphology of the oxide precursors is shown. In the micrographs particle sizes of up to 20 μm before milling can be observed.

After 4.5 h of milling, particle sizes have decreased to around 1 μm with aggregates. Figures 15 and 16 show the aggregates and particles after 4.5 and 7 h of milling, respectively, for the stoichiometric mixture used to form LaMnO3.

Fig. 10. X-ray powder diffraction patterns of different mixtures milled at 7 h, modifying the level of doping from x = 0 to x = 1 (Lira-Hernández et al., 2010).

Fig. 11. Distortion of the La(1-x)CaxMnO3 unit cell.

Figures 17-19 show the morphology of the nanoparticles aggregated after 4.5 and 7 h of milling of the (Mn2O3+CaO+La2O3) mixtures. The morphology is the same for compositions x = 0.3 and 0.4, but for x = 0.8 after 7 h of milling a different morphology is observed - larger and continuous particles with softer edges.

Figure 20 shows the morphology for the extreme case of x = 1, the Mn2O3+CaO mixture after 4.5 h of milling, and the absence of large agglomerates is noticeable; the particle size is on the order of nanometers.

Fig. 12. Calcium oxide precursor for the reaction of formation of La(1-x)CaxMnO3.

704 Scanning Electron Microscopy

After 7 h of milling, there is a shift to the right of the mean peak of the lanthanum manganite as the amount of doping is increased, indicating a decrease in the interplanar

In Figures 12- 14*,* morphology of the oxide precursors is shown. In the micrographs particle

After 4.5 h of milling, particle sizes have decreased to around 1 μm with aggregates. Figures 15 and 16 show the aggregates and particles after 4.5 and 7 h of milling, respectively, for the

Fig. 10. X-ray powder diffraction patterns of different mixtures milled at 7 h, modifying the

level of doping from x = 0 to x = 1 (Lira-Hernández et al., 2010).

Fig. 11. Distortion of the La(1-x)CaxMnO3 unit cell.

distance, corresponding to deformation of the network as proposed in Figure 11.

sizes of up to 20 μm before milling can be observed.

stoichiometric mixture used to form LaMnO3.

Fig. 13. Lanthanum oxide precursor for the reaction of formation of La(1-x)CaxMnO3.

Fig. 14. Manganesum oxide precursor (Mn2O3) for the reaction of formation of La(1-x)CaxMnO3.

Fig. 15. (Mn2O3+La2O3) mixture after 4.5 h of milling. milling.

Fig. 17. (Mn2O3+CaO+La2O3) mixture after 4.5 h of milling (x = 0.3).

Fig. 16. (Mn2O3+La2O3) mixture after 7 h of

Fig. 18. (Mn2O3+CaO+La2O3) mixture after 7 h of milling (x = 0.4).

$$2La\_2O\_3 + 2Mn\_2O\_3 \xrightarrow{SrO} 2LaMnO\_3 + 2Sr\_{La}' + V\_O^{xx} \tag{8}$$

$$\frac{(1-\chi)}{2}La\_2O\_3 + MnO \to \text{x}SrO + \frac{1}{4}(1-\chi)O\_2 \xrightarrow{MCP} La\_{1-x}Sr\_xMnO\_3 \dots \text{LSMx2} \tag{9}$$

$$\frac{(\text{1}-\text{x})}{2}La\_{2}O\_{3} + \frac{1}{2}Mn\_{2}O\_{3} + \text{x}SnO \xrightarrow{\text{MCP}} La\_{1-\text{x}}Sr\_{\text{x}}MnO\_{3} + \frac{(\text{-x})}{4}O\_{2} \dots \text{LSMx3} \tag{10}$$

$$\frac{(1-\chi)}{2}La\_2O\_3 + MnO\_2 + \text{xSrO} \xrightarrow{\text{MCP}} La\_{1-\chi}Sr\_{\text{X}}MnO\_3 + \frac{(1-\chi)}{4}O\_2 \dots \text{LSMx4} \tag{11}$$

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 709

As was mentioned previously, for applications as sensors and as cathodes in solid oxide fuel cells, the desired functionality of the material is based on the ionic conduction of oxygen through the interface between yttrium-doped zirconia (YSZ) and lanthanum manganite (LSM). This functionality can be potentiated by increasing the triple phase boundaries (TPB)

But in some conditions, an undesirable reaction occurs in the TPBs, giving rise to lanthanum zirconate formation through the diffusion of lanthanum atoms from manganite to zirconia

Fig. 26. Schematic of the triple phase boundaries in LSM-YSZ interfaces.

Fig. 27. Schematic of lanthanum zirconate formation in TPBs.

For this reason, it is important to determine the optimal conditions required to avoid the formation of lanthanum zirconate, which diminishes the ionic conduction through TPBs.

**5. YSZ-LSM interactions** 

(Figure 27).

formed between oxygen, YSZ and LSM (Figure 26).

Fig. 23. SEM images of La1-xCaxMnO3 consolidated with 10 wt% EBS at: a) 1100°C for 1 h and b) 1300°C for 3 h.

Fig. 24. SEM images of La1-xCaxMnO3 consolidated with 15 wt% EBS at: a) 1100°C for 1 h and b) 1300°C for 3 h.

Fig. 25. SEM images of La1-xCaxMnO3 consolidated with 20 wt% EBS at: a) 1100°C for 1 h and b) 1300°C for 3 h.

## **5. YSZ-LSM interactions**

708 Scanning Electron Microscopy

Fig. 23. SEM images of La1-xCaxMnO3 consolidated with 10 wt% EBS at: a) 1100°C for 1 h

Fig. 24. SEM images of La1-xCaxMnO3 consolidated with 15 wt% EBS at: a) 1100°C for 1 h

Fig. 25. SEM images of La1-xCaxMnO3 consolidated with 20 wt% EBS at: a) 1100°C for 1 h

and b) 1300°C for 3 h.

and b) 1300°C for 3 h.

and b) 1300°C for 3 h.

As was mentioned previously, for applications as sensors and as cathodes in solid oxide fuel cells, the desired functionality of the material is based on the ionic conduction of oxygen through the interface between yttrium-doped zirconia (YSZ) and lanthanum manganite (LSM). This functionality can be potentiated by increasing the triple phase boundaries (TPB) formed between oxygen, YSZ and LSM (Figure 26).

But in some conditions, an undesirable reaction occurs in the TPBs, giving rise to lanthanum zirconate formation through the diffusion of lanthanum atoms from manganite to zirconia (Figure 27).

Fig. 26. Schematic of the triple phase boundaries in LSM-YSZ interfaces.

Fig. 27. Schematic of lanthanum zirconate formation in TPBs.

For this reason, it is important to determine the optimal conditions required to avoid the formation of lanthanum zirconate, which diminishes the ionic conduction through TPBs.

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 711

In all cases, grain growth is reflected in the decrease of the peak width and increase in the intensity of the peaks. The relative intensity of the main lanthanum zirconate peak is greatest for lanthanum manganite obtained from MnO2+La2O3 without strontium and is smallest when doped with 15 at% strontium. In addition, there is a phase change for zirconia in the presence of lanthanum zirconate, tending to the cubic phase instead of its

The last point can be also observed in the backscattered electron SEM micrograph (Figure 29), in which the cubic zirconia morphology (shown with small dots and in dark gray) is clearly distinguished from the lanthanum zirconate (bright gray) and the lanthanum manganite (medium gray). From this image we can also deduce the direction of the diffusion of the atoms. That is to say, lanthanum zirconate grains tend to invade zirconate

Fig. 29. SEM micrographs of the mixture of YPSZ with lanthanum manganite obtained from undoped MnII (a), (b), (c) before and (d), (e), (f) after sintering at 1300°C (Cortés-Escobedo et al., 2008). LZ: Lanthanum zirconate; LSM: Strontium doped lanthanum manganite; ZC:

In another experiment, using spark plasma sintering, for samples heat treated at 1000°C, lanthanum zirconate is observed only for sintered LaMnO3 mechanosinthesized from MnO2+La2O3 (LM4). But by treating at 1300°C, lanthanum zirconate is detected in all the

usual tetragonal structure (without lanthanum zirconate).

grains from lanthanum manganite grains.

cubic zirconia; ZC-T: tetragonal zirconia.

samples (Figures 30-31).

Heat treatment (1000 – 1300°C) and spark plasma sintering were used to obtain the conditions in which the formation of lanthanum zirconates is avoided.

Figure 28 shows the x-ray diffraction patterns of mixtures of yttria partially stabilized tetragonal zirconia (YPSZ) with lanthanum manganites synthesized from different manganese oxide precursors (LMz) without heat-treatment (Figures 28a, 28e, and 28i) and heat-treated at 1300°C (Figures 28b-28d, 28f-28h and 28j-28l).

*ZM* = monoclinic zirconia, *ZC-T* = cubic + tetragonal zirconia, *ZT* = tetragonal zirconia, *LZ* = lanthanum zirconate, *LM* = lanthanum manganite (Cortés-Escobedo et al., 2008).

Fig. 28. X-ray diffraction patterns of mixtures of YPSZ with lanthanum manganites prepared from: (a)-(d) MnO, La2O3 and SrO by mechanosynthesis and heat treatment in air; (e)-(h) Mn2O3, La2O3 and SrO by mechanosynthesis; (i)-(l) MnO2, La2O3 and SrO by mechanosynthesis. Manganites in mixtures (a), (b), (e), (f), (i) and (j) are undoped, while manganites in mixtures (c), (g) and (k) are doped with 15 at% Sr in La sites and (d), (h) and (l) have 20 at% Sr in La sites. All mixtures were heat-treated at 1300°C except (a), (e) and (i).

Figures 28a-b, 28e-f and 28i-j correspond to mixtures of YPSZ with undoped manganites prepared from MnII, MnIII and MnIV, respectively. In these figures, increasing heat-treatment temperature is shown to result in increased intensity and a widening of the peaks (Cortés-Escobedo et al., 2008).

Heat treatment (1000 – 1300°C) and spark plasma sintering were used to obtain the

Figure 28 shows the x-ray diffraction patterns of mixtures of yttria partially stabilized tetragonal zirconia (YPSZ) with lanthanum manganites synthesized from different manganese oxide precursors (LMz) without heat-treatment (Figures 28a, 28e, and 28i) and

*ZM* = monoclinic zirconia, *ZC-T* = cubic + tetragonal zirconia, *ZT* = tetragonal zirconia, *LZ* = lanthanum

Fig. 28. X-ray diffraction patterns of mixtures of YPSZ with lanthanum manganites prepared from: (a)-(d) MnO, La2O3 and SrO by mechanosynthesis and heat treatment in air; (e)-(h)

Figures 28a-b, 28e-f and 28i-j correspond to mixtures of YPSZ with undoped manganites prepared from MnII, MnIII and MnIV, respectively. In these figures, increasing heat-treatment temperature is shown to result in increased intensity and a widening of the peaks (Cortés-

mechanosynthesis. Manganites in mixtures (a), (b), (e), (f), (i) and (j) are undoped, while manganites in mixtures (c), (g) and (k) are doped with 15 at% Sr in La sites and (d), (h) and (l) have 20 at% Sr in La sites. All mixtures were heat-treated at 1300°C except (a), (e) and (i).

Mn2O3, La2O3 and SrO by mechanosynthesis; (i)-(l) MnO2, La2O3 and SrO by

zirconate, *LM* = lanthanum manganite (Cortés-Escobedo et al., 2008).

Escobedo et al., 2008).

conditions in which the formation of lanthanum zirconates is avoided.

heat-treated at 1300°C (Figures 28b-28d, 28f-28h and 28j-28l).

In all cases, grain growth is reflected in the decrease of the peak width and increase in the intensity of the peaks. The relative intensity of the main lanthanum zirconate peak is greatest for lanthanum manganite obtained from MnO2+La2O3 without strontium and is smallest when doped with 15 at% strontium. In addition, there is a phase change for zirconia in the presence of lanthanum zirconate, tending to the cubic phase instead of its usual tetragonal structure (without lanthanum zirconate).

The last point can be also observed in the backscattered electron SEM micrograph (Figure 29), in which the cubic zirconia morphology (shown with small dots and in dark gray) is clearly distinguished from the lanthanum zirconate (bright gray) and the lanthanum manganite (medium gray). From this image we can also deduce the direction of the diffusion of the atoms. That is to say, lanthanum zirconate grains tend to invade zirconate grains from lanthanum manganite grains.

Fig. 29. SEM micrographs of the mixture of YPSZ with lanthanum manganite obtained from undoped MnII (a), (b), (c) before and (d), (e), (f) after sintering at 1300°C (Cortés-Escobedo et al., 2008). LZ: Lanthanum zirconate; LSM: Strontium doped lanthanum manganite; ZC: cubic zirconia; ZC-T: tetragonal zirconia.

In another experiment, using spark plasma sintering, for samples heat treated at 1000°C, lanthanum zirconate is observed only for sintered LaMnO3 mechanosinthesized from MnO2+La2O3 (LM4). But by treating at 1300°C, lanthanum zirconate is detected in all the samples (Figures 30-31).

Characterization of Ceramic Materials Synthesized by Mechanosynthesis for Energy Applications 713

In Figure 32 SEM images show the microstructure of spark plasma sintered LaMnO3 mechanosinthesized from MnO+La2O3 (LM2) and MnO2+La2O3 (LM4) and La0.85Sr0.15MnO3 mechanosinthesized from MnO+SrO+La2O3 (LSM152) and MnO2+SrO+La2O3 (LSM154), treated at 1000°C and 1300°C. In the case of treatment at 1000°C, pores are present in all samples, but the differentiation between grains is diffuse. Samples treated at 1300°C show

continuity in the microstructure without differentiation in grains.

Fig. 32. SEM images of the microstructure of spark plasma sintered LaMnO3

1300°C.

mechanosinthesized from MnO+La2O3 (LM2) and MnO2+La2O3 (LM4) and La0.85Sr0.15MnO3 mechanosinthesized from MnO+SrO+La2O3 (LSM152) and MnO2+SrO+La2O3 (LSM154). On the left are samples spark plasma sintered at 1000°C, and on right are samples treated at

Fig. 30. X-ray diffraction of lanthanum manganite – yttria-stabilized zirconia mixtures, which have been spark plasma sintered at 1000°C for 10 min. LM2: Lanthanum manganite from MnO+La2O3; LM4: Lanthanum manganite from MnO2+La2O3; LSM152: 15 at% Srdoped lanthanum manganite from MnO+La2O3+ SrO; LSM154: 15 at% Sr-doped lanthanum manganite from MnO2+La2O3+SrO.

Fig. 31. X-ray diffraction of lanthanum manganite – yttria-stabilized zirconia mixtures, which have been spark plasma sintered at 1300°C for 10 min. LM2: Lanthanum manganite from MnO+La2O3; LM4: Lanthanum manganite from MnO2+La2O3; LSM152: 15 at% Sr-doped lanthanum manganite from MnO+La2O3+SrO; LSM154: 15 at% Sr-doped lanthanum manganite from MnO2+La2O3+SrO.

Fig. 30. X-ray diffraction of lanthanum manganite – yttria-stabilized zirconia mixtures, which have been spark plasma sintered at 1000°C for 10 min. LM2: Lanthanum manganite from MnO+La2O3; LM4: Lanthanum manganite from MnO2+La2O3; LSM152: 15 at% Srdoped lanthanum manganite from MnO+La2O3+ SrO; LSM154: 15 at% Sr-doped lanthanum

Fig. 31. X-ray diffraction of lanthanum manganite – yttria-stabilized zirconia mixtures, which have been spark plasma sintered at 1300°C for 10 min. LM2: Lanthanum manganite from MnO+La2O3; LM4: Lanthanum manganite from MnO2+La2O3; LSM152: 15 at% Sr-doped lanthanum manganite from MnO+La2O3+SrO; LSM154: 15 at% Sr-doped

manganite from MnO2+La2O3+SrO.

lanthanum manganite from MnO2+La2O3+SrO.

In Figure 32 SEM images show the microstructure of spark plasma sintered LaMnO3 mechanosinthesized from MnO+La2O3 (LM2) and MnO2+La2O3 (LM4) and La0.85Sr0.15MnO3 mechanosinthesized from MnO+SrO+La2O3 (LSM152) and MnO2+SrO+La2O3 (LSM154), treated at 1000°C and 1300°C. In the case of treatment at 1000°C, pores are present in all samples, but the differentiation between grains is diffuse. Samples treated at 1300°C show continuity in the microstructure without differentiation in grains.

Fig. 32. SEM images of the microstructure of spark plasma sintered LaMnO3 mechanosinthesized from MnO+La2O3 (LM2) and MnO2+La2O3 (LM4) and La0.85Sr0.15MnO3 mechanosinthesized from MnO+SrO+La2O3 (LSM152) and MnO2+SrO+La2O3 (LSM154). On the left are samples spark plasma sintered at 1000°C, and on right are samples treated at 1300°C.

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**35** 

*Morocco* 

**Scanning Electron Microscopy (SEM) and** 

**of Adhesion Stage and Biofilm Formation** 

*1Laboratory of Microbial Biotechnology, Faculty of Science and Technics, Fez,* 

Soumya El Abed1,2, Saad Koraichi Ibnsouda1,2,

*3Laboratory of Valorization and Security Food Products,* 

Hassan Latrache3 and Fatima Hamadi3

*Faculty of Science and Technics, Beni Mellal,* 

**Environmental SEM: Suitable Tools for Study** 

*2Regional University Center of Interface, University Sidi Mohamed Ben Abdellah, Fez,* 

For most of the history of microbiology, microorganisms have primarily been characterized as planktonic, freely suspended cells and described on the basis of their growth characteristics in nutritionally rich culture media. The discovery of microorganisms, 1684, is usually ascribed to Antoni van Leeuwenhoek, who was the first person to publish microscopic observations of bacteria. The direct quantitative recovery techniques showed unequivocally that more than 99.9% of the bacteria grow in biofilms on a wide variety of surfaces. Although the most common mode of growth for microorganisms on earth is in surface associated communities (Stoodley et al., 2002; Sutherland, 2001), the first reported findings of microorganisms "attached in layers" were not made until the 1940s. During the 1960s and 70s the research on "microbial slimes" accelerated but the term "biofilm" was not unanimous formulated until 1984 (Bryers, 2000). Biofilm has three-dimensional (3D) structured, heterogeneous community of microbial cells enclosed in an exopolysaccharide matrix (also called glycocalyx) that are irreversibly attached to an inert or living surface. As establish, biofilm formation has a serious implications in public health and medicine. In the case of human health, a number of microbial infections are associated with surface colonization not only on live surfaces (sinusitis, pulmonary infection in cystic fibrosis patients, periodontitis, etc. (Hall-Stoodley et al., 2004) but also on medical implants (contact lenses, dental implants, intravascular catheters, urinary stents) etc. (Donlan, 2001; Hall-Stoodley et al., 2004). Biofilms affect heat exchangers, filters, etc. because they induce biocorrosion and biofouling, producing damages on metallic surfaces and the efficiency loss in industrial set-up (Dunne, 2002; Garret et al., 2008). However,biofilms have also useful applications in bioremediation (Vidali, 2001) of different environments (microorganisms degrade and convert pollutants into less toxic forms) and biolixiviation (bacteria can efficiently dissolve minerals used in

**1. Introduction** 

industry, to obtain copper and gold).

Zhou, S. M., Zhao, S. Y., He, L. F., Guo, Y.Q. & Shi, L. (2010). Facile synthesis of Ca-doped manganite nanoparticles by a nonaqueous sol–gel method and their magnetic properties. *Materials Chemistry and Physics*. Vol. 120, (March 2010), pp. (75-78), ISSN 0254-0584.

## **Scanning Electron Microscopy (SEM) and Environmental SEM: Suitable Tools for Study of Adhesion Stage and Biofilm Formation**

Soumya El Abed1,2, Saad Koraichi Ibnsouda1,2, Hassan Latrache3 and Fatima Hamadi3

*1Laboratory of Microbial Biotechnology, Faculty of Science and Technics, Fez, 2Regional University Center of Interface, University Sidi Mohamed Ben Abdellah, Fez, 3Laboratory of Valorization and Security Food Products, Faculty of Science and Technics, Beni Mellal, Morocco* 

#### **1. Introduction**

716 Scanning Electron Microscopy

Zhou, S. M., Zhao, S. Y., He, L. F., Guo, Y.Q. & Shi, L. (2010). Facile synthesis of Ca-doped

0254-0584.

manganite nanoparticles by a nonaqueous sol–gel method and their magnetic properties. *Materials Chemistry and Physics*. Vol. 120, (March 2010), pp. (75-78), ISSN

> For most of the history of microbiology, microorganisms have primarily been characterized as planktonic, freely suspended cells and described on the basis of their growth characteristics in nutritionally rich culture media. The discovery of microorganisms, 1684, is usually ascribed to Antoni van Leeuwenhoek, who was the first person to publish microscopic observations of bacteria. The direct quantitative recovery techniques showed unequivocally that more than 99.9% of the bacteria grow in biofilms on a wide variety of surfaces. Although the most common mode of growth for microorganisms on earth is in surface associated communities (Stoodley et al., 2002; Sutherland, 2001), the first reported findings of microorganisms "attached in layers" were not made until the 1940s. During the 1960s and 70s the research on "microbial slimes" accelerated but the term "biofilm" was not unanimous formulated until 1984 (Bryers, 2000). Biofilm has three-dimensional (3D) structured, heterogeneous community of microbial cells enclosed in an exopolysaccharide matrix (also called glycocalyx) that are irreversibly attached to an inert or living surface. As establish, biofilm formation has a serious implications in public health and medicine. In the case of human health, a number of microbial infections are associated with surface colonization not only on live surfaces (sinusitis, pulmonary infection in cystic fibrosis patients, periodontitis, etc. (Hall-Stoodley et al., 2004) but also on medical implants (contact lenses, dental implants, intravascular catheters, urinary stents) etc. (Donlan, 2001; Hall-Stoodley et al., 2004). Biofilms affect heat exchangers, filters, etc. because they induce biocorrosion and biofouling, producing damages on metallic surfaces and the efficiency loss in industrial set-up (Dunne, 2002; Garret et al., 2008). However,biofilms have also useful applications in bioremediation (Vidali, 2001) of different environments (microorganisms degrade and convert pollutants into less toxic forms) and biolixiviation (bacteria can efficiently dissolve minerals used in industry, to obtain copper and gold).

Scanning Electron Microscopy (SEM) and Environmental SEM:

2005) and is microenvironment-conservative (Beech, 2004).

2003).

**2.3 Maturation of biofilm** 

**3. Imaging application** 

**3.1 SEM applied of adhesion stage** 

amorphous).

for biofilm to spread (Kaplan et al., 2003).

**2.4 Detachment and dispersal of biofilm cells** 

Suitable Tools for Study of Adhesion Stage and Biofilm Formation 719

called extracellular polymeric substances (EPS). This matrix is a complex hydrogel embedding the bacteria community and building up in three dimensions. The backbone of this gel is mainly composed of polysaccharides produced by bacteria (such as colanic acid, chitosan, alginate), other components such as enzymes, DNA, RNA, nutrients, proteins, surfactants (Flemming et al., 2007). The exact role of the matrix is not yet completely elucidated but it has been demonstrated that the matrix acts as a protective layer (Fux et al.,

After the adherence of microorganism to the inert surface, the association becomes stable for micro-colonies formation (Bechmann & Eduvean, 2006; O'Toole et al., 2000). The microorganism begin to multiply while sending out chemical signals that intercommunicate among the bacterial cells. In this way, the bacteria multiply within the embedded exopolysaccharide matrix, thus giving rise to formation of a micro-colonies (Prakash et al.,

Once bacteria have irreversibly attached to a surface, the process of biofilm maturation begins. The overall density and complexity of the biofilm increase as surface-bound organisms begin to actively replicate and extracellular components generated by attached bacteria interact with organic and inorganic molecules in the immediate environment to create the glycocalyx (Carpentier & Cerf, 1993). The maturation of biofilm generate many process already having taken place, such as quorum sensing (Nadell et al., 2008), gene transfer (Molin, 2003), persister development (Lewis, 2005) etc. All of these processes contribute to the community life of the biofilm and play an important role in biofilm survival and biofilm spreading, since they allow also detachment of biofilm parts and release of free bacteria, which is the most common way

As the biofilm gets older, cells detach, disperse and colonize a new niche. This detachment can be due to various factors including, fluid dynamics and shear effects of the bulk fluid (Brugnoni et al., 2007). At some point of biofilms may partially dissolve releasing cells that

SEM is a well-established basic method to observe the morphology of bacteria adhered on a material surfaces, the morphology of the material surface, and the relationships between them (Peters et al., 1982). SEM has been used for enumeration of adhered bacteria or tissue large number of samples. It is as a key technique that provides also information about the morphology of biofilm, presence of EPS and the nature of corrosion products (crystalline or

Microbial adhesion is the first step of the formation of biofilm and an extremely complicated process that is affected by many factors. In this regard, detailed investigation of microbial

more away to other where a new cycle begins (Prakash et al., 2003; Singh et al., 2002).

In order that we may gain a greater insight into the ecology of the microorganisms that exist in biofilm, it is necessary not only to be able to isolate them by traditional culture methods but also to have some understanding of the way in which these individual microorganisms interact in situ in their environment. Different microscopic techniques for biofilm monitoring including Scanning Electron microscopy (SEM) have been proved to be suitable tools in order to follow the study of adhesion stage and biofilm formation. Scanning electron microscopy as a specialized field of science that employs the electron microscope as a tool and uses a beam of electrons to form an image of a specimen allowing imaging and quantification of surface topographic features.

The scope of this chapter is to illustrate the importance of scanning electron microscopy and environnemental scanning electron microscopy in biofilm examination and control. Furthermore, although we are conscious about the vast variety of biofilms in natural, clinical and industrial environments, this chapter will mainly concentrate on imaging application of SEM and ESEM biofilms.

## **2. Step of biofilm formation**

Planktonic cells are able to attach on the surfaces and form biofilm through a process that include several steps:

Fig. 1. Schematic illustrations of biofilm formation and development. (Filloux & Vallet, 2003).

## **2.1 Attachment/colonization**

The primary adhesion stage constitutes the beneficial contact between a conditioned surface and planktonic microorganisms. During the process of attachment, the organism must be brought into close proximity of the surface, propelled either randomly or in a directed fashion via chemotaxis and mobility (Prakash et al., 2003). This step is reversible and it is characterized by a number of physicochemical variables that defines the interaction between the microbial cell surface and the conditioned surface of interest (An et al., 2000; Liu et al., 2004; Singh et al., 2002).

#### **2.2 Irreversible adhesion**

The second step is the irreversible adhesion during which bacteria start to express adhesion protein such as curli or fimbriae to adhere to the surface. Microorganisms starts to produce intercellular connections (intercellular curli for example) and a polymeric matrix, usually called extracellular polymeric substances (EPS). This matrix is a complex hydrogel embedding the bacteria community and building up in three dimensions. The backbone of this gel is mainly composed of polysaccharides produced by bacteria (such as colanic acid, chitosan, alginate), other components such as enzymes, DNA, RNA, nutrients, proteins, surfactants (Flemming et al., 2007). The exact role of the matrix is not yet completely elucidated but it has been demonstrated that the matrix acts as a protective layer (Fux et al., 2005) and is microenvironment-conservative (Beech, 2004).

After the adherence of microorganism to the inert surface, the association becomes stable for micro-colonies formation (Bechmann & Eduvean, 2006; O'Toole et al., 2000). The microorganism begin to multiply while sending out chemical signals that intercommunicate among the bacterial cells. In this way, the bacteria multiply within the embedded exopolysaccharide matrix, thus giving rise to formation of a micro-colonies (Prakash et al., 2003).

## **2.3 Maturation of biofilm**

718 Scanning Electron Microscopy

In order that we may gain a greater insight into the ecology of the microorganisms that exist in biofilm, it is necessary not only to be able to isolate them by traditional culture methods but also to have some understanding of the way in which these individual microorganisms interact in situ in their environment. Different microscopic techniques for biofilm monitoring including Scanning Electron microscopy (SEM) have been proved to be suitable tools in order to follow the study of adhesion stage and biofilm formation. Scanning electron microscopy as a specialized field of science that employs the electron microscope as a tool and uses a beam of electrons to form an image of a specimen allowing imaging and

The scope of this chapter is to illustrate the importance of scanning electron microscopy and environnemental scanning electron microscopy in biofilm examination and control. Furthermore, although we are conscious about the vast variety of biofilms in natural, clinical and industrial environments, this chapter will mainly concentrate on imaging application of

Planktonic cells are able to attach on the surfaces and form biofilm through a process that

Fig. 1. Schematic illustrations of biofilm formation and development. (Filloux & Vallet,

The primary adhesion stage constitutes the beneficial contact between a conditioned surface and planktonic microorganisms. During the process of attachment, the organism must be brought into close proximity of the surface, propelled either randomly or in a directed fashion via chemotaxis and mobility (Prakash et al., 2003). This step is reversible and it is characterized by a number of physicochemical variables that defines the interaction between the microbial cell surface and the conditioned surface of interest (An et al., 2000; Liu et al.,

The second step is the irreversible adhesion during which bacteria start to express adhesion protein such as curli or fimbriae to adhere to the surface. Microorganisms starts to produce intercellular connections (intercellular curli for example) and a polymeric matrix, usually

quantification of surface topographic features.

SEM and ESEM biofilms.

include several steps:

2003).

**2. Step of biofilm formation** 

**2.1 Attachment/colonization** 

2004; Singh et al., 2002).

**2.2 Irreversible adhesion** 

Once bacteria have irreversibly attached to a surface, the process of biofilm maturation begins. The overall density and complexity of the biofilm increase as surface-bound organisms begin to actively replicate and extracellular components generated by attached bacteria interact with organic and inorganic molecules in the immediate environment to create the glycocalyx (Carpentier & Cerf, 1993). The maturation of biofilm generate many process already having taken place, such as quorum sensing (Nadell et al., 2008), gene transfer (Molin, 2003), persister development (Lewis, 2005) etc. All of these processes contribute to the community life of the biofilm and play an important role in biofilm survival and biofilm spreading, since they allow also detachment of biofilm parts and release of free bacteria, which is the most common way for biofilm to spread (Kaplan et al., 2003).

### **2.4 Detachment and dispersal of biofilm cells**

As the biofilm gets older, cells detach, disperse and colonize a new niche. This detachment can be due to various factors including, fluid dynamics and shear effects of the bulk fluid (Brugnoni et al., 2007). At some point of biofilms may partially dissolve releasing cells that more away to other where a new cycle begins (Prakash et al., 2003; Singh et al., 2002).

## **3. Imaging application**

SEM is a well-established basic method to observe the morphology of bacteria adhered on a material surfaces, the morphology of the material surface, and the relationships between them (Peters et al., 1982). SEM has been used for enumeration of adhered bacteria or tissue large number of samples. It is as a key technique that provides also information about the morphology of biofilm, presence of EPS and the nature of corrosion products (crystalline or amorphous).

## **3.1 SEM applied of adhesion stage**

Microbial adhesion is the first step of the formation of biofilm and an extremely complicated process that is affected by many factors. In this regard, detailed investigation of microbial

Scanning Electron Microscopy (SEM) and Environmental SEM:

pH2 pH3

on plant aerial surfaces using SEM (Gras et al., 1994).

al., 1992).

pH11

Suitable Tools for Study of Adhesion Stage and Biofilm Formation 721

pH5

Fig. 3. SEM images of *S. aureus* adhered to glass as a function of pH (Hamadi et al., 2005)

Biofilm morphology and mass are important characteristics that control the kinetics of substrate removal by biofilms. SEM is a powerful technique for revealing the fine structure of living systems and has been applied to biofilms (Eighmy et al., 1983; Richards and Turner, 1984; Weber et al.*,* 1978). It has also been of special importance in elucidating biofilm structure for understanding the physiology and ecology of these microbial systems (Blenkinsopp & Costerton 1991). For example, electron-microscopic studies proved that the biofilm is composed of bacterial cells "wrapped" in a dense "glycocalyx", i.e. exopolysaccharide matrix (Blenkinsopp & Costerton, 1991; Eighmy et al. 1983). In medical applications, for example, Storti et al., (2005) used scanning electron microscopy and reported that the extracellular biofilm matrix appears as an amorphous material on the catheter surface. In the same context, scanning electron microscopy (SEM) images of matrixenclosed microbial assemblages on leaf surfaces (Surico, 1993) have led some authors to suggest that biofilms occur in the phyllosphere (Beattie and Lindow, 1995). Morris et al., (1997) have been to observe microbial biofilms directly on leaf surfaces. Bacterial aggregates in the phyllosphere have been observed previously with SEM (Surico,1993), but most have been very small (less than 20 mm long) or have lacked an obvious exopolymeric matrix (Surico,1993). Previous studies have claimed to demonstrate the presence of biofilms in situ

Biofilm thickness is also especially important for calculation of heat exchange or diffusion rates of antimicrobials or nutrients through a biofilm and for evaluation of the mechanical properties of a biofilm (Korstgens et al., 2001). As reported elsewhere, SEM sample (freezedried cross-section of Foley bladder catheter) revealed the thickness of biofilm and also the layers of embedded of slime by different strains and species of bacterial cells (Ganderton et

In general, other application of SEM techniques may be mentioned. Akernan et al., (1993) used scanning electron microscopy of nanobacteria - novel biofilm producing organisms in

adhesion involved in the developmental process from single sessile bacteria to multicellular biofilm is crucial to elaborate strategies to control biofilm development. Moreover, submicrometer-scale cell surface polymers and appendages, such as curli, flagella, and exocellular polymers, have been shown to play essential roles during cell adhesion and biofilm formation (Busscher et al., 2008; Dufrêne, 2008; Rodrigues & Elimelech, 2009). A SEM image of such a curli is depicted in Figure 2.

Fig. 2. SEM images of *E.coli* K-12 MG 1655 ompR234 producing curli (Olsen et al., 1989)

Adhesion phenomena has been evaluated as function of substratum, liquid medium, carbone source, pH and hydrodynamics parameters including flow rate. Many of the conclusions about biofilm development, composition, distribution, and relationship to substratum have been derived from scanning electron microscopy (Bragadeewaran et al., 2010; Herald & Zottola, 1988; Pinna et al., 2000). We report here several investigations made in our laboratory used scanning electron microscopy to study adhesion phenomena. Hamadi et al., (2005) have investigated the adhesion of *Staphylococcus aureus* ATCC 25923 to glass at different pH values using scanning electron microscopy and image analysis with the Mathlab® program is shown in Figure 3.

The surface topography has been widely discussed as a parameter influencing microbial adhesion. In this regard, experiments made by Kouider et al., (2010) using SEM to determine the effect of stainless steel surface roughness on *Staphylococcus aureus* adhesion shown that adhesion level was found to largely depend on the substrate roughness with maximum at Ra = 0.025μm and minimum at Ra= 0.8μm. Mallouki et al., (2007) have studied the antiadhesive effect of fucans by SEM and a MATLAB program to determine the number and characteristics of adhered cells.

## **3.2 SEM applied of biofilm formation**

Scanning electron microscopy (SEM) is a useful technique for the investigation of surface structure of biological samples (Duckett & Ligrone, 1995; Minoura et al., 1995; Motta et al., 1994). For instance, much of the current knowledge about biofilms is due to the advances in imaging studies, especially the SEM. Early microscopic techniques used in biofilm monitoring, mainly applied during the 1980s, include scanning electron microscopy. SEM has been previously used to show a clear visualization of bacteria within a biofilm and is capable of demonstrating even a single bacterium and the relation of the biofilm to the underlying surface.

adhesion involved in the developmental process from single sessile bacteria to multicellular biofilm is crucial to elaborate strategies to control biofilm development. Moreover, submicrometer-scale cell surface polymers and appendages, such as curli, flagella, and exocellular polymers, have been shown to play essential roles during cell adhesion and biofilm formation (Busscher et al., 2008; Dufrêne, 2008; Rodrigues & Elimelech, 2009). A

Fig. 2. SEM images of *E.coli* K-12 MG 1655 ompR234 producing curli (Olsen et al., 1989)

Adhesion phenomena has been evaluated as function of substratum, liquid medium, carbone source, pH and hydrodynamics parameters including flow rate. Many of the conclusions about biofilm development, composition, distribution, and relationship to substratum have been derived from scanning electron microscopy (Bragadeewaran et al., 2010; Herald & Zottola, 1988; Pinna et al., 2000). We report here several investigations made in our laboratory used scanning electron microscopy to study adhesion phenomena. Hamadi et al., (2005) have investigated the adhesion of *Staphylococcus aureus* ATCC 25923 to glass at different pH values using scanning electron microscopy and image analysis with the

The surface topography has been widely discussed as a parameter influencing microbial adhesion. In this regard, experiments made by Kouider et al., (2010) using SEM to determine the effect of stainless steel surface roughness on *Staphylococcus aureus* adhesion shown that adhesion level was found to largely depend on the substrate roughness with maximum at Ra = 0.025μm and minimum at Ra= 0.8μm. Mallouki et al., (2007) have studied the antiadhesive effect of fucans by SEM and a MATLAB program to determine the number and

Scanning electron microscopy (SEM) is a useful technique for the investigation of surface structure of biological samples (Duckett & Ligrone, 1995; Minoura et al., 1995; Motta et al., 1994). For instance, much of the current knowledge about biofilms is due to the advances in imaging studies, especially the SEM. Early microscopic techniques used in biofilm monitoring, mainly applied during the 1980s, include scanning electron microscopy. SEM has been previously used to show a clear visualization of bacteria within a biofilm and is capable of demonstrating even a single bacterium and the relation of the biofilm to the

SEM image of such a curli is depicted in Figure 2.

Mathlab® program is shown in Figure 3.

characteristics of adhered cells.

underlying surface.

**3.2 SEM applied of biofilm formation** 

Fig. 3. SEM images of *S. aureus* adhered to glass as a function of pH (Hamadi et al., 2005)

Biofilm morphology and mass are important characteristics that control the kinetics of substrate removal by biofilms. SEM is a powerful technique for revealing the fine structure of living systems and has been applied to biofilms (Eighmy et al., 1983; Richards and Turner, 1984; Weber et al.*,* 1978). It has also been of special importance in elucidating biofilm structure for understanding the physiology and ecology of these microbial systems (Blenkinsopp & Costerton 1991). For example, electron-microscopic studies proved that the biofilm is composed of bacterial cells "wrapped" in a dense "glycocalyx", i.e. exopolysaccharide matrix (Blenkinsopp & Costerton, 1991; Eighmy et al. 1983). In medical applications, for example, Storti et al., (2005) used scanning electron microscopy and reported that the extracellular biofilm matrix appears as an amorphous material on the catheter surface. In the same context, scanning electron microscopy (SEM) images of matrixenclosed microbial assemblages on leaf surfaces (Surico, 1993) have led some authors to suggest that biofilms occur in the phyllosphere (Beattie and Lindow, 1995). Morris et al., (1997) have been to observe microbial biofilms directly on leaf surfaces. Bacterial aggregates in the phyllosphere have been observed previously with SEM (Surico,1993), but most have been very small (less than 20 mm long) or have lacked an obvious exopolymeric matrix (Surico,1993). Previous studies have claimed to demonstrate the presence of biofilms in situ on plant aerial surfaces using SEM (Gras et al., 1994).

Biofilm thickness is also especially important for calculation of heat exchange or diffusion rates of antimicrobials or nutrients through a biofilm and for evaluation of the mechanical properties of a biofilm (Korstgens et al., 2001). As reported elsewhere, SEM sample (freezedried cross-section of Foley bladder catheter) revealed the thickness of biofilm and also the layers of embedded of slime by different strains and species of bacterial cells (Ganderton et al., 1992).

In general, other application of SEM techniques may be mentioned. Akernan et al., (1993) used scanning electron microscopy of nanobacteria - novel biofilm producing organisms in

Scanning Electron Microscopy (SEM) and Environmental SEM:

for maintaining the biological samples in their natural state.

The relative humidity in an ESEM specimen chamber can be controlled (Stokes & Donald, 2000), so ESEM is particularly useful for hydrated materials (Muscariello et al., 2005; Stokes & Donald, 2000; Stokes, 2001). A gaseous secondary electron detector (GSED) exploits the gas in the specimen chamber for signal amplification. BSED operation produces positive

uncharged up by the electron beam.

Suitable Tools for Study of Adhesion Stage and Biofilm Formation 723

these conditions without rapid collapse (Heslop-Harrison, 1970) and fewer still survive (Read & Lord, 1991). Uncoated non-conductors build up local concentration of electron, referred to as-charging- that prevent the formation of usable images. Energy X-ray Spectroscopy (EDS) can be used to determine the elemental composition of surface films in the SEM, but EDS analyses must be completed prior to deposition of the thin metal coating EDS data are typically collected from an area, the specimen must be removed from the

To allow observations under the high vacuum conditions of SEM, many preparations of biological samples have been developed, e.g., glutaraldehyde fixation, negative staining, the Sputter–Cryo technique, and coating with gold or osmium (Allan-Wojtas et al., 2008; Hassan et al., 2003; Lamed et al., 1987). Moreover, these preparations have some positive effects on the biological sample; for instance, they enhance contrast, reduce damage, and are

**4. Biofilm formation: Environmental Scanning Electron Microscopy (ESEM)**  A new SEM technique is now available which allows overcoming these obstacles. a modified, low-vacuum scanning electron microscopy technique for biofilm monitoring that enables imaging of hydrated specimens, termed environmental scanning electron microscopy (ESEM) also called variable pressure SEM (VP-SEM),was introduced in the mid-1990s (Little et al., 1991). The environmental SEM (reviewed in Stokes & Donald, 2000) uses a series of pressure limiting apertures (Muscariello et al., 2005) while preventing gas leakage from the specimen chamber, which can be maintained at 1–20 Torr. The ESEM is based upon the gaseous detection device (GDD). The main feature distinguishing ESEM from conventional SEM is the presence of a gas in the specimen chamber. Gases may include nitrous oxide, helium, argon and other, but water vapour is the most efficient amplifying gas found and the most common gas used in ESEM. The ionization GDD uses the ionization of the gas for the detection of secondary electrons from the specimen surface. It is a conical electrode about 1 cm in diameter that is positioned with the apex downward and concentric with the beam at the bottom of the pole piece. Secondary electrons emitted from the sample collide with water molecules in the chamber producing additional electrons and positive ions. The positive ions are attracted to the sample surface and eliminate the charging artifacts. A proportional cascade amplification of the original secondary electron signal results. With the GDD both secondary and backscattered electron images can be produced. Detailed technical explanations about this device can be found elsewhere (Danilatos, 1990). The balance of gas flows into and out of the ESEM sample chamber determines its pressure. The multiple apertures are situated below the objective lens and separate the sample chamber from the column. This feature allows the column to remain at high vacuum while the specimen chamber may sustain pressures as high as 50 Torr. The temperature and humidity of the sample can also be manually controlled to provide a suitable environment

specimen chamber and coating with a conductive layer, and returned to the SEM.

blood. Indeed, nanoscale characterization of *Escherichia coli* Biofilm formed in the glass surface using scanning electron microscopy has been reported by Lim et al., (2008). He showed reticular structures on the surface of biofilms. The reticular structures consist of nanopores having diameter ranging from 14 nm to 100 nm.

Scanning electron microscopy (SEM) is one of the many methods available for the visual the effect of antibacterial or antifungal on biofilm development (Camargo et al., 2005; McDowell et al., 2004; Sasidharan et al., 2010; Sevinç & Hanley, 2010; Zameer & Gopal, 2010; Zeraik & Nitschke, 2010). Sasidharan et al., (2010) used SEM for studied The effects of potential antifungal extracts from natural sources in *Candida albicans* biofilm (Figure.4).

Fig. 4. Scanning electron micrograph reduction in *Candida albicans* biofilm after 36 h treatment. (a) Control and (b) Cassia spectabilis extract treated *C. albicans* cells.

#### **3.3 Advantages and disadvantages of SEM**

In part, it is true that Scanning electron microscopy (SEM) present a many advantages, the more important are: (i) higher resolution of visualization microbial biofilms (Walker et al., 2001) than other imaging techniques, typically 3.5 nm, (ii) able to measure and quantify data in three dimensions. However, this technique utilizes graded solvents (alcohol, acetone, and xylene) to gradually dehydrate the specimen prior to examination, since water of hydration is not compatible with the vacuum used with the electron beam. While any pretreatment can alter specimen morphology, drying appears to significantly alter biofilms due to EPS polymers collapsing (Fassel & Edmiston, 1999; Little et al., 1991). The dehydration process results in significant sample distortion and artifacts; the extracellular polymeric substances, which are approximately 95% water and the liquid loss led them to appear more like fibers surrounding the cells than like a gelatinous matrix (Characklis & Marshall, 1990). Several ultrastructural studies have used conventional scanning electron microscopy (SEM) to investigate the glycocalyx, but these studies (Costerton et al., 1981; Fassel et al., 1991; Marshall et al., 1971) were hampered by low resolution and also by the inability to use low voltages (<5 keV), which yield increased information from small topographical features (Pawley & Erlandsen, 1989).

Typically, SEM imaging requires a high vacuum, ≤10-8 Torr (reviewed in Stewart, 1985), having rst been chemically xed, dehydrated, and coated with a conductive material (e.g. gold) to prevent charge buildup from the electron beam. Few biological specimens tolerate

blood. Indeed, nanoscale characterization of *Escherichia coli* Biofilm formed in the glass surface using scanning electron microscopy has been reported by Lim et al., (2008). He showed reticular structures on the surface of biofilms. The reticular structures consist of

Scanning electron microscopy (SEM) is one of the many methods available for the visual the effect of antibacterial or antifungal on biofilm development (Camargo et al., 2005; McDowell et al., 2004; Sasidharan et al., 2010; Sevinç & Hanley, 2010; Zameer & Gopal, 2010; Zeraik & Nitschke, 2010). Sasidharan et al., (2010) used SEM for studied The effects of potential

antifungal extracts from natural sources in *Candida albicans* biofilm (Figure.4).

(a) (b)

Fig. 4. Scanning electron micrograph reduction in *Candida albicans* biofilm after 36 h treatment. (a) Control and (b) Cassia spectabilis extract treated *C. albicans* cells.

In part, it is true that Scanning electron microscopy (SEM) present a many advantages, the more important are: (i) higher resolution of visualization microbial biofilms (Walker et al., 2001) than other imaging techniques, typically 3.5 nm, (ii) able to measure and quantify data in three dimensions. However, this technique utilizes graded solvents (alcohol, acetone, and xylene) to gradually dehydrate the specimen prior to examination, since water of hydration is not compatible with the vacuum used with the electron beam. While any pretreatment can alter specimen morphology, drying appears to significantly alter biofilms due to EPS polymers collapsing (Fassel & Edmiston, 1999; Little et al., 1991). The dehydration process results in significant sample distortion and artifacts; the extracellular polymeric substances, which are approximately 95% water and the liquid loss led them to appear more like fibers surrounding the cells than like a gelatinous matrix (Characklis & Marshall, 1990). Several ultrastructural studies have used conventional scanning electron microscopy (SEM) to investigate the glycocalyx, but these studies (Costerton et al., 1981; Fassel et al., 1991; Marshall et al., 1971) were hampered by low resolution and also by the inability to use low voltages (<5 keV), which yield increased information from small topographical features

Typically, SEM imaging requires a high vacuum, ≤10-8 Torr (reviewed in Stewart, 1985), having rst been chemically xed, dehydrated, and coated with a conductive material (e.g. gold) to prevent charge buildup from the electron beam. Few biological specimens tolerate

nanopores having diameter ranging from 14 nm to 100 nm.

**3.3 Advantages and disadvantages of SEM** 

(Pawley & Erlandsen, 1989).

these conditions without rapid collapse (Heslop-Harrison, 1970) and fewer still survive (Read & Lord, 1991). Uncoated non-conductors build up local concentration of electron, referred to as-charging- that prevent the formation of usable images. Energy X-ray Spectroscopy (EDS) can be used to determine the elemental composition of surface films in the SEM, but EDS analyses must be completed prior to deposition of the thin metal coating EDS data are typically collected from an area, the specimen must be removed from the specimen chamber and coating with a conductive layer, and returned to the SEM.

To allow observations under the high vacuum conditions of SEM, many preparations of biological samples have been developed, e.g., glutaraldehyde fixation, negative staining, the Sputter–Cryo technique, and coating with gold or osmium (Allan-Wojtas et al., 2008; Hassan et al., 2003; Lamed et al., 1987). Moreover, these preparations have some positive effects on the biological sample; for instance, they enhance contrast, reduce damage, and are uncharged up by the electron beam.

## **4. Biofilm formation: Environmental Scanning Electron Microscopy (ESEM)**

A new SEM technique is now available which allows overcoming these obstacles. a modified, low-vacuum scanning electron microscopy technique for biofilm monitoring that enables imaging of hydrated specimens, termed environmental scanning electron microscopy (ESEM) also called variable pressure SEM (VP-SEM),was introduced in the mid-1990s (Little et al., 1991). The environmental SEM (reviewed in Stokes & Donald, 2000) uses a series of pressure limiting apertures (Muscariello et al., 2005) while preventing gas leakage from the specimen chamber, which can be maintained at 1–20 Torr. The ESEM is based upon the gaseous detection device (GDD). The main feature distinguishing ESEM from conventional SEM is the presence of a gas in the specimen chamber. Gases may include nitrous oxide, helium, argon and other, but water vapour is the most efficient amplifying gas found and the most common gas used in ESEM. The ionization GDD uses the ionization of the gas for the detection of secondary electrons from the specimen surface. It is a conical electrode about 1 cm in diameter that is positioned with the apex downward and concentric with the beam at the bottom of the pole piece. Secondary electrons emitted from the sample collide with water molecules in the chamber producing additional electrons and positive ions. The positive ions are attracted to the sample surface and eliminate the charging artifacts. A proportional cascade amplification of the original secondary electron signal results. With the GDD both secondary and backscattered electron images can be produced. Detailed technical explanations about this device can be found elsewhere (Danilatos, 1990).

The balance of gas flows into and out of the ESEM sample chamber determines its pressure.

The multiple apertures are situated below the objective lens and separate the sample chamber from the column. This feature allows the column to remain at high vacuum while the specimen chamber may sustain pressures as high as 50 Torr. The temperature and humidity of the sample can also be manually controlled to provide a suitable environment for maintaining the biological samples in their natural state.

The relative humidity in an ESEM specimen chamber can be controlled (Stokes & Donald, 2000), so ESEM is particularly useful for hydrated materials (Muscariello et al., 2005; Stokes & Donald, 2000; Stokes, 2001). A gaseous secondary electron detector (GSED) exploits the gas in the specimen chamber for signal amplification. BSED operation produces positive

Scanning Electron Microscopy (SEM) and Environmental SEM:

**5. Conclusion** 

**6. References** 

Supplement III.

No.1,(January 2008), pp.101–108.

Humana Press, Totowa, N.J.

physiology, thickness, etc.

of inorganic products within the biofilm.

Suitable Tools for Study of Adhesion Stage and Biofilm Formation 725

Interestingly, Shen et al., (2011) have been proposed a novel method for measuring an adhesion force of single yeast cell based on a nanorobotic manipulation system inside an environmental scanning electron microscope (ESEM) and Dubey & Ben-Yehuda (2011) report the identication of analogous nanotubular channels formed among bacterial cells grown on solid surface. They demonstrate that nanotubes connect bacteria of the same and different

Scanning electron microscopy is a key tool to study the effect of physicochemical properties on adhesion phenomena (pH, roughness, topography, temperature, etc). SEM plays also a paramount role for assessing the microbial populations, three-dimensional structure,

SEM proved to be an invaluable method for ultra-structural investigation, allowing imaging of the overall appearance and/or specific features of biofilms formed in different environments , e.g. microbial colonies and individual cells, the glycocalyx, and the presence

Surely, Scanning Electron Microscope (SEM) is a powerful research tool, but since it requires high vacuum conditions, the wet materials and biological samples must undergo a complex preparation that limits the application of SEM on this kind of specimen and often causes the introduction of artifacts. The introduction of Environmental Scanning Electron Microscope (ESEM), working in gaseous atmosphere, represented a new perspective in biofilm

ESEM could be useful as a complementary technique to help in the characterization of the structure and architecture of biofilms. In fact, ESEM could reveal the exact topography of intact, live and fully hydrated biofilms, with a higher magnification than the other microscopy techniques. In general, a combination of several techniques is to be recommended when investigating biofilms as the different techniques offer distinctly

Akernan, K.K. Kuronen,Ilpo. Olavi Kajander, E. (1993). Scanning electron microscopy of

Allan-Wojtas, P. Hansen, L.T. & Paulson, A.T. (2008). Microstructural studies of probiotic

An, Y. H, Dickinson, R. B. & Doyle, R. J. (2000). Mechanisms of bacterial adhesion and

Bacteria to Polystyrene Surfaces: Effect of Temperature and hydrophobicity. *Current of* 

nanobacteria - Novel biofilm producing organisms in blood. *Scanning,* Vol.15,

bacteria loaded alginate microcapsules using standard electron microscopy techniques and anhydrous fixation. *LWT-Food Science and Technology,* Vol.41,

pathogenesis of implant and tissue infections. pp. 1-27. In An, Y. H. & Friedman, R. J. (ed.), Handbook of bacterial adhesion: principles, methods, and applications.

monitoring with high resolution without prior fixing and staining.

valuable information about different aspects of biofilm development.

*Microbiology,*Vol.61, (December 2010), pp.554–559.

species, thereby providing an effective conduit for exchange of intracellular content.

ions that have the added benefit of limiting charging of non-conductive specimens (Stokes & Donald, 2000). It does not require prior fixing and staining of the biofilm, minimizes biofilm dehydration and thus preserves native morphologies including surface structures (Walker et al., 2001) and native morphologies of bacteria and biofilms (e.g. Priester et al., 2007) and is able to achieve high magnifications, comparable with SEM. Shrinkage is prevented and artefact formation is reduced.

Additional advantages of ESEM include minimal processing of samples. It results in shorter time scales and lower costs while reducing the possibility of introducing artefacts. Samples can be preserved in saline in a common refrigerator (in fresh) if examination is to be deferred a few hours (Ramírez-Camacho et al., 2008). ESEM provides spatial resolutions of 10 nm or less. Compared to SEM, ESEM produces different, perhaps complementary, information for biological specimens (Doucet et al., 2005; Surman et al., 1996). Cell structures are visible with SEM, but external polymers around cells are more apparent in ESEM (Callow et al., 2003; Doucet et al., 2005; S. Douglas & D.D. Douglas, 2001).

## **4.1 ESEM applied of biofilm formation**

Sutton et al., (1994) used this technique to study the structure of a *Streptococcus crista* CR3 biofilm. Gilpin & Sigee (1995) showed that biological samples can be imaged in the ESEM in wet or partially hydrated states with a minimum of sample damage and changes in specimen morphology. This gave the possibility to the visualization of biofilm surfaces in their natural wet anaerobic state (Darkin et al., 2001). Recently, Schwartz et al., (2009) used ESEM imaging to obtain information about the bacterial composition, matrix composition, and spatial biofilm structures of natural biofilms grown on filter materials at waterworks.

Scanning electron microscopes are frequently equipped with an energy dispersive x-ray analyser. This equipment permits elemental analysis with a high horizontal resolution of the inspected specimens. In this same context, mineral structures formed by bacterial and microalgal biofilms growing on the archaeological surface in Maltese hypogea were studied using Energy Dispersive X-Ray Spectroscopy (EDS) coupled to Environmental Scanning Electron Microscopy (ESEM), are reported by Zammit et al., (2011). These techniques have shown that mineral structures having different morphologies and chemical composition were associated with the microorganisms in the subaerophytic biofilm (Figure.5).

Fig. 5. ESEM and EDS analysis for the system under SRB-biofilm influence. (A) SEM Image of carbon steel exposed to sterile artificial seawater (supplemented with nutrients) and with SRB, (B) EDS analysis corresponding to the ESEM smooth region.

Interestingly, Shen et al., (2011) have been proposed a novel method for measuring an adhesion force of single yeast cell based on a nanorobotic manipulation system inside an environmental scanning electron microscope (ESEM) and Dubey & Ben-Yehuda (2011) report the identication of analogous nanotubular channels formed among bacterial cells grown on solid surface. They demonstrate that nanotubes connect bacteria of the same and different species, thereby providing an effective conduit for exchange of intracellular content.

## **5. Conclusion**

724 Scanning Electron Microscopy

ions that have the added benefit of limiting charging of non-conductive specimens (Stokes & Donald, 2000). It does not require prior fixing and staining of the biofilm, minimizes biofilm dehydration and thus preserves native morphologies including surface structures (Walker et al., 2001) and native morphologies of bacteria and biofilms (e.g. Priester et al., 2007) and is able to achieve high magnifications, comparable with SEM. Shrinkage is prevented and

Additional advantages of ESEM include minimal processing of samples. It results in shorter time scales and lower costs while reducing the possibility of introducing artefacts. Samples can be preserved in saline in a common refrigerator (in fresh) if examination is to be deferred a few hours (Ramírez-Camacho et al., 2008). ESEM provides spatial resolutions of 10 nm or less. Compared to SEM, ESEM produces different, perhaps complementary, information for biological specimens (Doucet et al., 2005; Surman et al., 1996). Cell structures are visible with SEM, but external polymers around cells are more apparent in ESEM

Sutton et al., (1994) used this technique to study the structure of a *Streptococcus crista* CR3 biofilm. Gilpin & Sigee (1995) showed that biological samples can be imaged in the ESEM in wet or partially hydrated states with a minimum of sample damage and changes in specimen morphology. This gave the possibility to the visualization of biofilm surfaces in their natural wet anaerobic state (Darkin et al., 2001). Recently, Schwartz et al., (2009) used ESEM imaging to obtain information about the bacterial composition, matrix composition, and spatial biofilm structures of natural biofilms grown on filter materials at waterworks. Scanning electron microscopes are frequently equipped with an energy dispersive x-ray analyser. This equipment permits elemental analysis with a high horizontal resolution of the inspected specimens. In this same context, mineral structures formed by bacterial and microalgal biofilms growing on the archaeological surface in Maltese hypogea were studied using Energy Dispersive X-Ray Spectroscopy (EDS) coupled to Environmental Scanning Electron Microscopy (ESEM), are reported by Zammit et al., (2011). These techniques have shown that mineral structures having different morphologies and chemical composition

were associated with the microorganisms in the subaerophytic biofilm (Figure.5).

Fig. 5. ESEM and EDS analysis for the system under SRB-biofilm influence. (A) SEM Image of carbon steel exposed to sterile artificial seawater (supplemented with nutrients) and with

SRB, (B) EDS analysis corresponding to the ESEM smooth region.

(Callow et al., 2003; Doucet et al., 2005; S. Douglas & D.D. Douglas, 2001).

artefact formation is reduced.

**4.1 ESEM applied of biofilm formation** 

Scanning electron microscopy is a key tool to study the effect of physicochemical properties on adhesion phenomena (pH, roughness, topography, temperature, etc). SEM plays also a paramount role for assessing the microbial populations, three-dimensional structure, physiology, thickness, etc.

SEM proved to be an invaluable method for ultra-structural investigation, allowing imaging of the overall appearance and/or specific features of biofilms formed in different environments , e.g. microbial colonies and individual cells, the glycocalyx, and the presence of inorganic products within the biofilm.

Surely, Scanning Electron Microscope (SEM) is a powerful research tool, but since it requires high vacuum conditions, the wet materials and biological samples must undergo a complex preparation that limits the application of SEM on this kind of specimen and often causes the introduction of artifacts. The introduction of Environmental Scanning Electron Microscope (ESEM), working in gaseous atmosphere, represented a new perspective in biofilm monitoring with high resolution without prior fixing and staining.

ESEM could be useful as a complementary technique to help in the characterization of the structure and architecture of biofilms. In fact, ESEM could reveal the exact topography of intact, live and fully hydrated biofilms, with a higher magnification than the other microscopy techniques. In general, a combination of several techniques is to be recommended when investigating biofilms as the different techniques offer distinctly valuable information about different aspects of biofilm development.

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**36** 

*USA* 

Mohammad Kamal Hossain

**Scanning Electron Microscopy Study of** 

*Department of Mechanical Engineering, Tuskegee University,* 

**Fiber Reinforced Polymeric Nanocomposites** 

The nanoscale materials offer the opportunity to explore new behavior beyond those established in conventional materials. It has been established that mechanical and thermal properties, moisture barrier, and flame resistance of polymeric composites can be improved by adding a small amount of nanoparticles as filler particles without compromising the density, toughness, storage life, weight or processability of the composite [Sandler et al., 2002; Bruzaud & Bourmaud, 2007]. The higher surface area is one of the most promising characteristics of nanoparticles due to their ability in creating a good interface in a composite. The dispersion of nanoparticles in the matrix is one of the most important parameters in fabricating nanophased composites. It depends on processing techniques such as solution blending, shear mixing, in-situ polymerization, ultrasonic cavitation, and high pressure mixing [Giannelis, 1998; Yasmin et al., 2003; Vaia et al., 1996]. Nanomaterials have enhanced various characteristics in a given polymer. However, these enhancements have limitations at higher loadings due to increased

During service life, composite structures might encounter high stresses resulting in crack propagation through fiber matrix interfaces. Therefore, stronger adhesion between fiber and matrix, higher strength, and higher toughened matrix are desired. Improvement of flexural strength by addition of nanofillers into a matrix is expected to be observed for several reasons. Young's modulus of the second phase dispersed particles is higher than that of the matrix and thus stress transfer from the matrix to the particles will take place. As a result, the strength of the composites is increased. Strong interfacial bonding between the fiber and matrix also contributes to higher flexural strength. Dispersed filler particles act as a mechanical interlocking between the fiber and matrix which creates a high friction coefficient. Finally, a mixed mode of fracture (flexural and shear) occurs under bending-load conditions. After an initial failure of fibers at the tensile side of the specimen, cracks are deflected parallel to the fibers and also to the applied load direction. The stress-strain curve shows a sharp increment with increasing load before reaching the maximum stress and then irregularities and staggered decrease in stress were observed for both conventional and nanophased composites [M.K. Hossain et al., 2011]. However, the initial load and the crack arrest area are higher in nanophased composites which lead to high energy absorbing

**1. Introduction** 

agglomeration causing premature failure.

mechanisms [Hussain et al., 1996].


## **Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites**

Mohammad Kamal Hossain

*Department of Mechanical Engineering, Tuskegee University, USA* 

## **1. Introduction**

730 Scanning Electron Microscopy

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The nanoscale materials offer the opportunity to explore new behavior beyond those established in conventional materials. It has been established that mechanical and thermal properties, moisture barrier, and flame resistance of polymeric composites can be improved by adding a small amount of nanoparticles as filler particles without compromising the density, toughness, storage life, weight or processability of the composite [Sandler et al., 2002; Bruzaud & Bourmaud, 2007]. The higher surface area is one of the most promising characteristics of nanoparticles due to their ability in creating a good interface in a composite. The dispersion of nanoparticles in the matrix is one of the most important parameters in fabricating nanophased composites. It depends on processing techniques such as solution blending, shear mixing, in-situ polymerization, ultrasonic cavitation, and high pressure mixing [Giannelis, 1998; Yasmin et al., 2003; Vaia et al., 1996]. Nanomaterials have enhanced various characteristics in a given polymer. However, these enhancements have limitations at higher loadings due to increased agglomeration causing premature failure.

During service life, composite structures might encounter high stresses resulting in crack propagation through fiber matrix interfaces. Therefore, stronger adhesion between fiber and matrix, higher strength, and higher toughened matrix are desired. Improvement of flexural strength by addition of nanofillers into a matrix is expected to be observed for several reasons. Young's modulus of the second phase dispersed particles is higher than that of the matrix and thus stress transfer from the matrix to the particles will take place. As a result, the strength of the composites is increased. Strong interfacial bonding between the fiber and matrix also contributes to higher flexural strength. Dispersed filler particles act as a mechanical interlocking between the fiber and matrix which creates a high friction coefficient. Finally, a mixed mode of fracture (flexural and shear) occurs under bending-load conditions. After an initial failure of fibers at the tensile side of the specimen, cracks are deflected parallel to the fibers and also to the applied load direction. The stress-strain curve shows a sharp increment with increasing load before reaching the maximum stress and then irregularities and staggered decrease in stress were observed for both conventional and nanophased composites [M.K. Hossain et al., 2011]. However, the initial load and the crack arrest area are higher in nanophased composites which lead to high energy absorbing mechanisms [Hussain et al., 1996].

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 733

Flexural tests under three-point bend configuration were performed using a Zwick Roell testing unit according to the ASTM D790-02 standard to evaluate flexural modulus and strength of each of the material systems of the polymer nanocomposites and its laminates [M.E. Hossain et al., 2011; M.K. Hossain et al., 2011]. The machines were run under displacement control mode at a crosshead speed of 2.0 mm/min and tests were performed at room temperature. The span to depth ratio was maintained at 16:1. The maximum stress at failure on the tension side of a flexural specimen was considered as the flexural strength of the material. Flexural modulus was calculated from the slope of the stress-strain plot. Five samples of each type were tested. The average values and standard deviation of flexural

SEM studies were carried out to examine change in the microstructure due to the addition of CNFs using a JEOL JSM 5800 microscope. SEM also facilitates to monitor the failure approach at micro level. Failed samples from the three point flexure test were examined to distinguish the changes in the failure mode. The samples were cut through the cross-section of the failed region. The samples were positioned on a sample holder with a silver paint and coated with gold to prevent charge build-up by the electron absorbed by the specimen. A 15 kilovolt accelerating voltage was applied to achieve desired magnification [M.E. Hossain et

Fig. 1. Flow chart of sample fabrication

strength and modulus were determined.

al., 2011; M.K. Hossain et al., 2011].

**2.4 Scanning Electron Microscopy (SEM)** 

**2.3 Flexural test** 

The morphological study of the nanoparticle dispersion in the CNF-loaded polyester nanocomposites using various processing techniques, including mechanical mixing, magnetic stirring, and sonication is evaluated through SEM studies in this article. Nanoparticles facilitate proper wetting out fibers with resin, void reduction, enhanced crosslinking, and increased friction co-efficient. Sometimes it also works as a nucleating agent in a fiber reinforced polymeric composite. Morphology of the glass/polyester-CNF composites manufactured by the VARTM process has also been studied using the SEM for various applications, including civil infrastructure, automotive, aerospace, sporting, and marine industries. Quantitative enhancement in properties is characterized through the flexural test in this article. Qualitative and visual analyses are important to support the resultant experimental quantitative data and fracture morphology evaluation of different tested specimens using SEM is inevitable in this matter. Therefore, the fracture morphology of all types of tested specimens has been evaluated in this article.

## **2. Research methodology**

#### **2.1 Neat and CNF-loaded polyester sample fabrication**

B-440 premium polyester resin, styrene, and heat treated PR-24 CNF were used as matrix, thinner, and nanoparticle, respectively. Polyester resin contains two parts: part-A (polyester resin) and part-B (MEKP- methyl ethyl ketone peroxide) as crosslinking agent. Sonication was performed in a glass beaker using a high intensity ultrasonic irradiation (Ti-horn, 20 kHz Sonics Vibra Cell, Sonics Mandmaterials, Inc, USA) for 60, 90, and 120 minutes, respectively, adding 0.1-0.4 wt.% CNFs to polyester resin while adding 10 wt.% styrene. The mixing process was carried out in a pulse mode of 30 sec. on and 15 sec. off at an amplitude of 50%. To study other types of mixing methods, CNFs were mixed using mechanical mixing and magnetic stirring methods. The mechanical mixer was run for 90 minutes at 300 rpm at room temperature. The magnetic stirring was carried out for 5 hours at 500 rpm at room temperature. To lessen the void formation, vacuum was applied using Brand Tech Vacuum system for about 90-120 minutes and 0.7 wt.% catalyst was then added to the mixer using a high-speed mechanical stirrer for about 2-3 minutes and vacuum was again applied for about 6-8 minutes to degasify the bubbles produced during the catalyst mixing. The asprepared resin was poured into the mold and kept at room temperature for 12-15 hours. Controlled polyester samples were fabricated to compare with the nanophased samples. All samples were kept in a mechanical convection oven at 110 °C for 3 hours for post curing [M.E. Hossain et al., 2011].

#### **2.2 Conventional and nanophased fiber reinforced composite sample cabrication**

Both conventional and nanophased E-glass/polyester-CNF composites were manufactured by the VARTM process. Vacuum was maintained until the end of cure to remove any volatiles generated during the polymerization process. The panels were cured for about 12- 15 hours at room temperature and then thermally post cured at 110 °C for 3 hours in a mechanical convection oven. The fiber volume fraction for the nanophased glass reinforced polyester composites fabricated by VARTM was found to be around 56%. The void content (3-4%) was also within a reasonable limit in these composites [M.K. Hossain et al., 2011]. The overall sample fabrication procedure is presented in Figure 1.

Fig. 1. Flow chart of sample fabrication

## **2.3 Flexural test**

732 Scanning Electron Microscopy

The morphological study of the nanoparticle dispersion in the CNF-loaded polyester nanocomposites using various processing techniques, including mechanical mixing, magnetic stirring, and sonication is evaluated through SEM studies in this article. Nanoparticles facilitate proper wetting out fibers with resin, void reduction, enhanced crosslinking, and increased friction co-efficient. Sometimes it also works as a nucleating agent in a fiber reinforced polymeric composite. Morphology of the glass/polyester-CNF composites manufactured by the VARTM process has also been studied using the SEM for various applications, including civil infrastructure, automotive, aerospace, sporting, and marine industries. Quantitative enhancement in properties is characterized through the flexural test in this article. Qualitative and visual analyses are important to support the resultant experimental quantitative data and fracture morphology evaluation of different tested specimens using SEM is inevitable in this matter. Therefore, the fracture morphology

B-440 premium polyester resin, styrene, and heat treated PR-24 CNF were used as matrix, thinner, and nanoparticle, respectively. Polyester resin contains two parts: part-A (polyester resin) and part-B (MEKP- methyl ethyl ketone peroxide) as crosslinking agent. Sonication was performed in a glass beaker using a high intensity ultrasonic irradiation (Ti-horn, 20 kHz Sonics Vibra Cell, Sonics Mandmaterials, Inc, USA) for 60, 90, and 120 minutes, respectively, adding 0.1-0.4 wt.% CNFs to polyester resin while adding 10 wt.% styrene. The mixing process was carried out in a pulse mode of 30 sec. on and 15 sec. off at an amplitude of 50%. To study other types of mixing methods, CNFs were mixed using mechanical mixing and magnetic stirring methods. The mechanical mixer was run for 90 minutes at 300 rpm at room temperature. The magnetic stirring was carried out for 5 hours at 500 rpm at room temperature. To lessen the void formation, vacuum was applied using Brand Tech Vacuum system for about 90-120 minutes and 0.7 wt.% catalyst was then added to the mixer using a high-speed mechanical stirrer for about 2-3 minutes and vacuum was again applied for about 6-8 minutes to degasify the bubbles produced during the catalyst mixing. The asprepared resin was poured into the mold and kept at room temperature for 12-15 hours. Controlled polyester samples were fabricated to compare with the nanophased samples. All samples were kept in a mechanical convection oven at 110 °C for 3 hours for post curing

**2.2 Conventional and nanophased fiber reinforced composite sample cabrication** 

overall sample fabrication procedure is presented in Figure 1.

Both conventional and nanophased E-glass/polyester-CNF composites were manufactured by the VARTM process. Vacuum was maintained until the end of cure to remove any volatiles generated during the polymerization process. The panels were cured for about 12- 15 hours at room temperature and then thermally post cured at 110 °C for 3 hours in a mechanical convection oven. The fiber volume fraction for the nanophased glass reinforced polyester composites fabricated by VARTM was found to be around 56%. The void content (3-4%) was also within a reasonable limit in these composites [M.K. Hossain et al., 2011]. The

of all types of tested specimens has been evaluated in this article.

**2.1 Neat and CNF-loaded polyester sample fabrication** 

**2. Research methodology** 

[M.E. Hossain et al., 2011].

Flexural tests under three-point bend configuration were performed using a Zwick Roell testing unit according to the ASTM D790-02 standard to evaluate flexural modulus and strength of each of the material systems of the polymer nanocomposites and its laminates [M.E. Hossain et al., 2011; M.K. Hossain et al., 2011]. The machines were run under displacement control mode at a crosshead speed of 2.0 mm/min and tests were performed at room temperature. The span to depth ratio was maintained at 16:1. The maximum stress at failure on the tension side of a flexural specimen was considered as the flexural strength of the material. Flexural modulus was calculated from the slope of the stress-strain plot. Five samples of each type were tested. The average values and standard deviation of flexural strength and modulus were determined.

## **2.4 Scanning Electron Microscopy (SEM)**

SEM studies were carried out to examine change in the microstructure due to the addition of CNFs using a JEOL JSM 5800 microscope. SEM also facilitates to monitor the failure approach at micro level. Failed samples from the three point flexure test were examined to distinguish the changes in the failure mode. The samples were cut through the cross-section of the failed region. The samples were positioned on a sample holder with a silver paint and coated with gold to prevent charge build-up by the electron absorbed by the specimen. A 15 kilovolt accelerating voltage was applied to achieve desired magnification [M.E. Hossain et al., 2011; M.K. Hossain et al., 2011].

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 735

Fig. 2. Flexural stress-strain plot of polyester samples with different wt.% of CNFs.

Fig. 3. Flexural stress- strain plot of GRPC laminates with different wt.% of CNF.

#### **2.5 SEM sample preparation**

SEM samples must have an appropriate size to fit in the specimen chamber and is generally mounted rigidly on a specimen holder. Specimens must be electrically conductive, specially the surface, for imaging, and electrically grounded to prevent the accumulation of electrostatic charge at the surface of the specimen during electron irradiation [Suzuki, 2002]. Metal objects require little special preparation for SEM except for cleaning and mounting on the specimen holder. Nonconductive specimens tend to charge when scanned by the electron beam, especially in the secondary electron imaging mode. This causes scanning faults and other image artifacts. They are therefore usually coated with an ultrathin electrically-conducting material, commonly gold, deposited on the sample either using a low vacuum sputtering machine or a high vacuum evaporation unit.

## **3. Results and discussion**

#### **3.1 Flexural test results**

Typical stress–strain curves of neat and nanophased polyester samples as well as their fiber reinforced laminates generated from flexural tests are illustrated in Figures 2 and 3. Flexural strength, modulus, and the strain at maximum strength for all CNF-loaded samples were larger than those of the neat samples. CNF has high aspect ratio which can prevent crack generation and propagation in the polyester matrix (Figure 2). In all cases, the samples failed rapidly after experiencing the maximum load showing induced brittle nature of failure due to the addition of CNFs. The 0.2 wt.% CNFs loading and 90 minutes sonication time were observed to be the optimal condition for this nanocomposite system. The 0.2 wt.% CNF-loaded samples enhanced the flexural strength and modulus by about 88% and 16%, respectively, compared to the neat ones. The failure strain also increased significantly with the addition of CNFs into the system. Flexural properties were slightly decreased at higher CNF content. It might be due to the creation of micro aggregates of CNFs in various regions of the polymer matrix, which act as areas of weakness [M.E. Hossain et al., 2011].

Typical stress-strain curves of conventional and nanophased glass/polyester composites presented in Figure 3 demonstrated significant improvement in the mechanical properties up to the 0.2 wt.% of CNFs loading, beyond which there was a decreasing trend. These curves showed considerable nonlinear deformation before reaching the maximum stress. This was attributed to the random fiber breakage during loading. However, more or less ductility was observed in each type of laminate sample and cracking noise was heard while the individual fiber broke or the inter-layer delaminated. No obvious yield point was found. From the resultant data, it was concluded that the 0.2 wt.% CNF was the optimum amount for this material system to achieve the maximum flexural modulus and strength. These specimens showed approximately 49% and 31% increase in the flexural strength and modulus, respectively. There are several reasons for the better mechanical properties observed in the CNF-infused glass fabric reinforced polyester laminates. First, CNFs increase the strength and modulus of the polyester matrix, which was observed in the CNFloaded polyester in this study. Second, the presence of CNFs increases the crack propagation resistance and prevents crack generation by bridging effect at the interface region of the long glass fiber, CNF, and polyester matrix. Moreover, CNF has high aspect ratio, which improves the strength and modulus [M.K. Hossain et al., 2011].

SEM samples must have an appropriate size to fit in the specimen chamber and is generally mounted rigidly on a specimen holder. Specimens must be electrically conductive, specially the surface, for imaging, and electrically grounded to prevent the accumulation of electrostatic charge at the surface of the specimen during electron irradiation [Suzuki, 2002]. Metal objects require little special preparation for SEM except for cleaning and mounting on the specimen holder. Nonconductive specimens tend to charge when scanned by the electron beam, especially in the secondary electron imaging mode. This causes scanning faults and other image artifacts. They are therefore usually coated with an ultrathin electrically-conducting material, commonly gold, deposited on the sample either using a

Typical stress–strain curves of neat and nanophased polyester samples as well as their fiber reinforced laminates generated from flexural tests are illustrated in Figures 2 and 3. Flexural strength, modulus, and the strain at maximum strength for all CNF-loaded samples were larger than those of the neat samples. CNF has high aspect ratio which can prevent crack generation and propagation in the polyester matrix (Figure 2). In all cases, the samples failed rapidly after experiencing the maximum load showing induced brittle nature of failure due to the addition of CNFs. The 0.2 wt.% CNFs loading and 90 minutes sonication time were observed to be the optimal condition for this nanocomposite system. The 0.2 wt.% CNF-loaded samples enhanced the flexural strength and modulus by about 88% and 16%, respectively, compared to the neat ones. The failure strain also increased significantly with the addition of CNFs into the system. Flexural properties were slightly decreased at higher CNF content. It might be due to the creation of micro aggregates of CNFs in various regions

of the polymer matrix, which act as areas of weakness [M.E. Hossain et al., 2011].

improves the strength and modulus [M.K. Hossain et al., 2011].

Typical stress-strain curves of conventional and nanophased glass/polyester composites presented in Figure 3 demonstrated significant improvement in the mechanical properties up to the 0.2 wt.% of CNFs loading, beyond which there was a decreasing trend. These curves showed considerable nonlinear deformation before reaching the maximum stress. This was attributed to the random fiber breakage during loading. However, more or less ductility was observed in each type of laminate sample and cracking noise was heard while the individual fiber broke or the inter-layer delaminated. No obvious yield point was found. From the resultant data, it was concluded that the 0.2 wt.% CNF was the optimum amount for this material system to achieve the maximum flexural modulus and strength. These specimens showed approximately 49% and 31% increase in the flexural strength and modulus, respectively. There are several reasons for the better mechanical properties observed in the CNF-infused glass fabric reinforced polyester laminates. First, CNFs increase the strength and modulus of the polyester matrix, which was observed in the CNFloaded polyester in this study. Second, the presence of CNFs increases the crack propagation resistance and prevents crack generation by bridging effect at the interface region of the long glass fiber, CNF, and polyester matrix. Moreover, CNF has high aspect ratio, which

low vacuum sputtering machine or a high vacuum evaporation unit.

**2.5 SEM sample preparation** 

**3. Results and discussion** 

**3.1 Flexural test results** 

Fig. 2. Flexural stress-strain plot of polyester samples with different wt.% of CNFs.

Fig. 3. Flexural stress- strain plot of GRPC laminates with different wt.% of CNF.

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 737

Agglomerations in the polyester matrix were observed from the micrographs (Figure 6) of 0.2 wt.% infused polyester samples prepared through mechanical mixing and magnetic stirring methods, respectively. These agglomerates area create stress concentration zones

 (a) (b) Fig. 6. Micrographs of acid-etched 0.2 wt.% CNF-loaded polyester: (a) Mechanical stirring

Strong attractive fiber van der Waals forces cause CNFs to agglomerate, which reduces the strength of the nanocomposite by stress concentration effect. Agglomerates of CNFs, called nanoropes, are difficult to separate and infiltrate with matrix. They entangle and form nestlike structures due to their curvature and high aspect ratios. Both disagglomeration and dispersion in resins depend on the relative van der Waals forces, curvature, and on the relative surface energy of CNFs versus that of the resin. To overcome attractive forces, researchers have been extensively using mechanical energy, intense ultrasonication, and high speed shearing. Some rebundling of the aggregates is possible even after discontinuation of the external force [Yoonessi et al., 2008]. However, optimal loading and uniform dispersion of CNFs in matrix are the key parameters to promote better nanofibermatrix interface properties to reach an efficient load transfer between two constituents of the

Uniform dispersion of 0.2 wt.% CNFs into the polyester resin was achieved using the sonication mixing method for 90 minutes. High magnification SEM micrograph in Figure 7 clearly exhibits that CNFs are well separated and uniformly embedded in the 0.2% polyester resin system. It can also be easily observed that the interfacial bonding between the CNF and matrix was very compact which would allow CNFs to be anchored in the embedding matrix. In essence, these CNFs are likely to interlock and entangle with the polymer chains in the matrix [Li et al., 2008]. Thus, addition of CNFs enhanced the crosslinking between

Figures 8 (a) and 8 (b) show the woven glass reinforced polyester laminates with 0.2 wt.% CNFs. It was found that the resin was distributed uniformly over the fabric, and the interfacial bonding between matrix and fiber was very good. Resin flow and impregnation of the glass fibers can be observed in the SEM micrographs. Clear resin matrix adhesion is present in these micrographs, and glass fibers are observed to be embedded within the

which might act as a crack initiator.

and (b) Magnetic stirring

nanocomposite [Kozey et al., 1995; Ma et al., 2003].

polymer chains and provided better interfacial bonding.

### **3.2 Microstructure and morphological analyses**

The SEM micrographs of as-received PR-24 CNF and the neat polyester matrix are shown in Figures 4 (a) and 4 (b), respectively. To investigate the dispersion properties of CNFs in polyester, drops of concentrated HNO3 acid were added on the cleavage surfaces to partly unveil the CNFs formerly covered by the polyester. From the micrograph of 0.2 wt.% CNFfilled polyester, excellent dispersion of CNFs was found (Figure 5). Only broken ends of CNFs were observed near the surface. Some CNFs broke in a brittle manner and some were pulled out.

Fig. 4. SEM micrographs of (a) as-received PR-24 CNF and (b) neat polyester matrix.

Fig. 5. SEM micrograph of acid-etched 0.2 wt.% CNF-loaded polyester at 3000X.

The SEM micrographs of as-received PR-24 CNF and the neat polyester matrix are shown in Figures 4 (a) and 4 (b), respectively. To investigate the dispersion properties of CNFs in polyester, drops of concentrated HNO3 acid were added on the cleavage surfaces to partly unveil the CNFs formerly covered by the polyester. From the micrograph of 0.2 wt.% CNFfilled polyester, excellent dispersion of CNFs was found (Figure 5). Only broken ends of CNFs were observed near the surface. Some CNFs broke in a brittle manner and some were

(a) (b)

Fig. 4. SEM micrographs of (a) as-received PR-24 CNF and (b) neat polyester matrix.

Fig. 5. SEM micrograph of acid-etched 0.2 wt.% CNF-loaded polyester at 3000X.

**3.2 Microstructure and morphological analyses** 

pulled out.

Agglomerations in the polyester matrix were observed from the micrographs (Figure 6) of 0.2 wt.% infused polyester samples prepared through mechanical mixing and magnetic stirring methods, respectively. These agglomerates area create stress concentration zones which might act as a crack initiator.

Fig. 6. Micrographs of acid-etched 0.2 wt.% CNF-loaded polyester: (a) Mechanical stirring and (b) Magnetic stirring

Strong attractive fiber van der Waals forces cause CNFs to agglomerate, which reduces the strength of the nanocomposite by stress concentration effect. Agglomerates of CNFs, called nanoropes, are difficult to separate and infiltrate with matrix. They entangle and form nestlike structures due to their curvature and high aspect ratios. Both disagglomeration and dispersion in resins depend on the relative van der Waals forces, curvature, and on the relative surface energy of CNFs versus that of the resin. To overcome attractive forces, researchers have been extensively using mechanical energy, intense ultrasonication, and high speed shearing. Some rebundling of the aggregates is possible even after discontinuation of the external force [Yoonessi et al., 2008]. However, optimal loading and uniform dispersion of CNFs in matrix are the key parameters to promote better nanofibermatrix interface properties to reach an efficient load transfer between two constituents of the nanocomposite [Kozey et al., 1995; Ma et al., 2003].

Uniform dispersion of 0.2 wt.% CNFs into the polyester resin was achieved using the sonication mixing method for 90 minutes. High magnification SEM micrograph in Figure 7 clearly exhibits that CNFs are well separated and uniformly embedded in the 0.2% polyester resin system. It can also be easily observed that the interfacial bonding between the CNF and matrix was very compact which would allow CNFs to be anchored in the embedding matrix. In essence, these CNFs are likely to interlock and entangle with the polymer chains in the matrix [Li et al., 2008]. Thus, addition of CNFs enhanced the crosslinking between polymer chains and provided better interfacial bonding.

Figures 8 (a) and 8 (b) show the woven glass reinforced polyester laminates with 0.2 wt.% CNFs. It was found that the resin was distributed uniformly over the fabric, and the interfacial bonding between matrix and fiber was very good. Resin flow and impregnation of the glass fibers can be observed in the SEM micrographs. Clear resin matrix adhesion is present in these micrographs, and glass fibers are observed to be embedded within the

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 739

Results from the SEM study substantiate the quantitative results obtained through flexural test [M.E. Hossain et al., 2011; M.K. Hossain et al., 2011]. SEM performed on the fractured samples of flexure tests revealed rough and smooth fracture surfaces in 0.2 CNF-loaded sample and neat sample, respectively (Figure 9). The bonding between polyester and CNF was seen to be strong and attributed to cause deviation in the path of crack front as it propagated, thus requiring more energy to fracture the samples. This has resulted in increased strength and stiffness of samples. The effect was most pronounced at 0.2 wt.%

From the SEM micrograph taken at higher magnification as shown in Figure 10 (a), excellent bridging effect in the interfacial region of the long glass fiber, CNF, and matrix was observed. CNF has high aspect ratio which can prevent crack propagation and crack generation resulting in improved performance. Some resin was stacked on the fractured glass fiber as shown in Figure 10 (b), which represents better adhesion due to the addition of CNFs. The presence of polyester adhering to the fiber surface also suggests that interfacial adhesion is stronger than matrix strength in nanophased composites [Hussain et al., 1996]. Thus, it is evident from these micrographs that CNFs are anchored with both resin and fiber tightly that promotes a better interfacial bonding between the matrix and fiber. Better fibermatrix interfacial bonding, and CNFs' crack generation and propagation resistance result in higher strength in nanocomposites. On the other hand, the addition of the CNFs led to an improvement in the modulus of elasticity of the nanophased composites. This is attributed to the stiffening of the matrix of these composites (Figure 2). The interfacial area between the resin matrix and CNFs was increased because of the high aspect ratio of the CNFs, which in turn led to better mechanical properties [Tsantzalis et al., 2007]. The nanoparticles also act as reinforcing element and bear the load in the composite material system [Jawahar et al., 2006]. Again, both CNFs and fibers are stronger than matrix. Thus, when load is applied to the composite structures, matrix starts to crack first and stress is then transferred from the lower modulus matrix to the CNFs to the long fiber by bridging effect and ultimately, the

(a) Controlled polyester samples (b) 0.2 CNF loaded polyester samples

Fig. 9. SEM micrographs of fracture surface (a & b) after flexural test

**3.3 Fracture morphology analysis** 

composites' properties enhance.

loading of CNFs.

matrix. Good matrix-fiber wetting was achieved and resin is also visible in between the glass fiber filaments. It appears that better interfacial bonding between the nanophased polymer matrix and glass fiber is present due to the presence of CNFs [Green et al., 2009]. The fiber volume fraction as determined from matrix digestion method for the nanophased glass reinforced polyester composites fabricated by the VARTM process was found to be around 56%.

Fig. 7. SEM micrograph of acid-etched 0.2 wt.% CNF-loaded polyester at 5000X

Fig. 8. 0.2 wt.% CNF-loaded GRPC laminates (a and b).

#### **3.3 Fracture morphology analysis**

738 Scanning Electron Microscopy

matrix. Good matrix-fiber wetting was achieved and resin is also visible in between the glass fiber filaments. It appears that better interfacial bonding between the nanophased polymer matrix and glass fiber is present due to the presence of CNFs [Green et al., 2009]. The fiber volume fraction as determined from matrix digestion method for the nanophased glass reinforced polyester composites fabricated by the VARTM process was found to be around

Fig. 7. SEM micrograph of acid-etched 0.2 wt.% CNF-loaded polyester at 5000X

(a) (b)

Fig. 8. 0.2 wt.% CNF-loaded GRPC laminates (a and b).

56%.

Results from the SEM study substantiate the quantitative results obtained through flexural test [M.E. Hossain et al., 2011; M.K. Hossain et al., 2011]. SEM performed on the fractured samples of flexure tests revealed rough and smooth fracture surfaces in 0.2 CNF-loaded sample and neat sample, respectively (Figure 9). The bonding between polyester and CNF was seen to be strong and attributed to cause deviation in the path of crack front as it propagated, thus requiring more energy to fracture the samples. This has resulted in increased strength and stiffness of samples. The effect was most pronounced at 0.2 wt.% loading of CNFs.

(a) Controlled polyester samples (b) 0.2 CNF loaded polyester samples

Fig. 9. SEM micrographs of fracture surface (a & b) after flexural test

From the SEM micrograph taken at higher magnification as shown in Figure 10 (a), excellent bridging effect in the interfacial region of the long glass fiber, CNF, and matrix was observed. CNF has high aspect ratio which can prevent crack propagation and crack generation resulting in improved performance. Some resin was stacked on the fractured glass fiber as shown in Figure 10 (b), which represents better adhesion due to the addition of CNFs. The presence of polyester adhering to the fiber surface also suggests that interfacial adhesion is stronger than matrix strength in nanophased composites [Hussain et al., 1996]. Thus, it is evident from these micrographs that CNFs are anchored with both resin and fiber tightly that promotes a better interfacial bonding between the matrix and fiber. Better fibermatrix interfacial bonding, and CNFs' crack generation and propagation resistance result in higher strength in nanocomposites. On the other hand, the addition of the CNFs led to an improvement in the modulus of elasticity of the nanophased composites. This is attributed to the stiffening of the matrix of these composites (Figure 2). The interfacial area between the resin matrix and CNFs was increased because of the high aspect ratio of the CNFs, which in turn led to better mechanical properties [Tsantzalis et al., 2007]. The nanoparticles also act as reinforcing element and bear the load in the composite material system [Jawahar et al., 2006]. Again, both CNFs and fibers are stronger than matrix. Thus, when load is applied to the composite structures, matrix starts to crack first and stress is then transferred from the lower modulus matrix to the CNFs to the long fiber by bridging effect and ultimately, the composites' properties enhance.

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 741

For a better understanding of the fracture process, fracture morphology of samples was studied using higher magnification SEM micrographs. The SEM micrographs of the fractured surfaces of the conventional and 0.2 wt.% CNF-loaded GRPC are illustrated in Figure 11. For conventional composite shown in Figure 11 (a), the surface of the fiber was clean, and no matrix adhered to the fiber. The fracture surface of the matrix was flat, and some cracks were seen in the matrix side near the fiber-matrix interface. Resin appears not to protrude from the surface of fibers. These results indicate that the interfacial bonding between the fiber and matrix was weak. The fracture surface of the nanophased composite (Figure 11 (b)) shows that the surface of the matrix was rougher than that of neat composite. CNFs were observed to be randomly but uniformly distributed in the matrix. The resin appears to cling to fibers well. The strengthened matrix held the glass fabrics together. The protrusion of the resin from the surface of the fibers accounts for the increase in fracture toughness of the samples. Moreover, the resin appears to be sticking to the fiber surface giving rise to a significant plastic deformation [Xu & Hoa, 2008]. The plastic deformation enhances mechanical properties significantly in the nanophased composites (Figure 3) [M.K.

(a) (b)

Sonication, mechanical mixing, and magnetic stirring were performed to infuse 0.1-0.4 wt.% carbon nanofibers (CNFs) into the polyester resin. CNFs were used as nanoparticle fillers in woven glass fiber-reinforced polyester composites. Better dispersion of CNFs was observed in the 0.2 wt.% CNF-loaded polyester resin while CNFs were mixed using the sonication. The fiber volume fraction for the nanophased GRPC fabricated by the VARTM process was found around 56%. The void content was also within a reasonable limit in these composites. CNFs infusion even at quite low concentrations enhanced the mechanical properties of the system. This SEM investigation visually demonstrated that CNFs can be used without difficulty to modify the conventional fiber reinforced composite materials. Thus, SEM micrographs confirm that uniform dispersion and optimal loading of nanoparticles improve

Fig. 11. Fracture morphology of (a) conventional, and (b) 0.2 wt.% CNF-loaded GRPC.

the mechanical properties of composites with the following outcomes:

Hossain et al., 2011].

**4. Conclusion** 

(a)

(b)

Fig. 10. (a) Bridging effect at the interface region of the long glass fiber, CNF and the resin and (b) 0.2 wt.% CNF-loaded polyester matrix stacked with glass fabric after fractured laminate.

For a better understanding of the fracture process, fracture morphology of samples was studied using higher magnification SEM micrographs. The SEM micrographs of the fractured surfaces of the conventional and 0.2 wt.% CNF-loaded GRPC are illustrated in Figure 11. For conventional composite shown in Figure 11 (a), the surface of the fiber was clean, and no matrix adhered to the fiber. The fracture surface of the matrix was flat, and some cracks were seen in the matrix side near the fiber-matrix interface. Resin appears not to protrude from the surface of fibers. These results indicate that the interfacial bonding between the fiber and matrix was weak. The fracture surface of the nanophased composite (Figure 11 (b)) shows that the surface of the matrix was rougher than that of neat composite. CNFs were observed to be randomly but uniformly distributed in the matrix. The resin appears to cling to fibers well. The strengthened matrix held the glass fabrics together. The protrusion of the resin from the surface of the fibers accounts for the increase in fracture toughness of the samples. Moreover, the resin appears to be sticking to the fiber surface giving rise to a significant plastic deformation [Xu & Hoa, 2008]. The plastic deformation enhances mechanical properties significantly in the nanophased composites (Figure 3) [M.K. Hossain et al., 2011].

(a) (b)

Fig. 11. Fracture morphology of (a) conventional, and (b) 0.2 wt.% CNF-loaded GRPC.

## **4. Conclusion**

740 Scanning Electron Microscopy

(a)

(b) Fig. 10. (a) Bridging effect at the interface region of the long glass fiber, CNF and the resin and (b) 0.2 wt.% CNF-loaded polyester matrix stacked with glass fabric after fractured

laminate.

Sonication, mechanical mixing, and magnetic stirring were performed to infuse 0.1-0.4 wt.% carbon nanofibers (CNFs) into the polyester resin. CNFs were used as nanoparticle fillers in woven glass fiber-reinforced polyester composites. Better dispersion of CNFs was observed in the 0.2 wt.% CNF-loaded polyester resin while CNFs were mixed using the sonication. The fiber volume fraction for the nanophased GRPC fabricated by the VARTM process was found around 56%. The void content was also within a reasonable limit in these composites. CNFs infusion even at quite low concentrations enhanced the mechanical properties of the system. This SEM investigation visually demonstrated that CNFs can be used without difficulty to modify the conventional fiber reinforced composite materials. Thus, SEM micrographs confirm that uniform dispersion and optimal loading of nanoparticles improve the mechanical properties of composites with the following outcomes:

Scanning Electron Microscopy Study of Fiber Reinforced Polymeric Nanocomposites 743

Green, K.J., Dean, D.R., Vaidya, U.K., & Nyairo, E. (2009). Multiscale fiber reinforced

Hossain, M.E., Hossain, M.K., Hosur, M.V., & Jeelani, S. (2011). Study of Mechanical

Hossain, M.K., Hossain, M.E., Hosur, M.V., & Jeelani, S. (2011). Flexural and Compression

Hussain, M., Nakahira, A., & Niihara, K. (1996). Mechanical property improvement of

Jawahar, P., Gnanamoorthy, R., & Balasubramanian, M. (2006). Tribological behavior of clay-thermoset polyester nanocomposites. *Wear*, 261, (2006), pp. (835-840). Kozey, V.V., Jiang, H., Mehta, V.R., & Kumar, S. (1995). Compressive behavior of materials, Part II: High performance fibers. *J Mater Res*., 10, 4, (1995), pp. (1044-1061). Li, X.F., Lau, K.T., & Yin, Y.S. (2008). Mechanical properties of epoxy-based composites using coiled carbon naotubes. *Comp Sci Technol*., 68, (2008), pp. (2876-2881). Ma, H., Zeng, J., Realff, M.L., Kumar, S., & Schiraldi, D.A. (2003). Processing, structure, and

Sandler, J., Werner, P., Shaffer, M.S.P., Denchuk, V., Altstadt, V., & Windle, A.H. (2002).

Suzuki, E. (2002). High-Resolution Scanning Electron Microscopy of Immunogold-Labelled

Tsantzalis, S., Karapappas, P., Vavouliotis, A., Tsotra, P., Paipetis, A., Kostopoulos, V.,

Vaia, R.A., Jandt, K.D., Kramer, E.J., & Giannelis E.P. (1996). Microstructural evolution of

Xu, Y. & Hoa, S.V. (2008). Mechanical properties of carbon fiber reinforced epoxy/clay

Yasmin, A., Abot, J.L., & Daniel, I.M. (2003). Processing of clay/epoxy nanocomposites by

nanocomposites. *Comp Sci Technol*., 68, 3-4, (2008), pp. (854-861).

Shear mixing. *Scripta Materia*, 49, 1, (2003), pp. (81–86).

pp. (1470-1475).

DOI:10.1557/opl.2011.111.

3, (1996), pp. (185-191).

63, (2003), pp. (1617-1628).

33, (2002), pp. (1033-1039).

*Chem Mater,* 8*, (*1996), pp. (2628- 2635).

(2002), pp. (153–157).

1081).

*A.*, 42, 11, (2011), pp. (1774-1782).

composites based on a carbon nanofiber/epoxy nanophased polymer matrix: Synthesis, mechanical and thermomechanical behavior. *Comp: Part A.*, 40, 9, (2009),

Responses and Thermal Expansion of CNF-modified Polyester Nanocomposites Processed by Different Mixing Systems. *Cambridge Journal Online 2011*,

Response of Woven E-Glass/Polyester–CNF Nanophased Composites. *Comp: Part* 

carbon fiber reinforced epoxy composites by Al2O3 filler dispersion. *Mater Lett*., 26,

properties of fibers from polyester/carbon naofiber composites. *Comp Sci Technol*.,

Carbon nanofibers reinforced poly (ether ether ketone) composites. *Comp Part A*,

Cells by the Use of Thin Plasma Coating of Osmium. *Journal of Microscopy* 208, 3,

& Friedrich, K. (2007). Enhancement of the mechanical performance of an epoxy resin and fiber reinforced epoxy resin composites by the introduction of CNF and PZT particles at the microscale. *Comp: Part A*, 38, 4, *(*2007), pp. (1076-

melt intercalated polymer-organically modified layered silicates nanocomposites.


## **5. Aknowledgements**

The author is truly grateful to his hardworking and dedicated graduate students: Mr. Muhammad Enayet Hossain, Mr. Mohammad Washim Dewan, Mr. Kazi Al Imran, and Mr. Chinedu Okoro for their direct and indirect contribution in this article. The author also likes to express his gratitude towards Dr. Mahesh Hosur and Dr. Vijay Rangari for their cooperation and advice. The author also appreciates the help from the staff members of Tuskegee Center for Advanced Materials (T-CAM).

The author's special thanks are extended to Dr. Shaik Jeelani, Vice President, Research and Sponsored Programs at Tuskegee University for his invaluable support and guidance throughout this work both professionally and personally.

The author acknowledges the financial support of NSF-EPSCoR Grant No. EPS-0814103 and NSF-RISE Grant No. HRD-0833158 for this research work.

Finally, special thanks go to his wife, Dr. Shamim Ara Begum, and son, Ahnaf Hossain, for their love, patience, and mental support. Above all, the author thanks to God, Almighty for His love and mercy.

## **6. References**


• SEM micrographs revealed better dispersion of CNFs in the 0.2 wt.% CNF-loaded polyester prepared by sonication mixing and agglomeration in samples prepared by

• Uniform resin flow and proper impregnation of the glass fibers were observed in the

• Uniform resin flow and proper impregnation of the glass fibers appeared to result in a better interaction between the fiber and matrix that aided to an efficient stress transfer from the continuous polymer matrix to the dispersed fiber reinforcement through the

• Excellent bridging effect in the interfacial region of the long glass fiber, CNF, and

• SEM micrographs exhibited rougher fracture surface in the CNF-loaded polyester sample compared to the neat sample due to the presence of well dispersed and well

• SEM micrographs revealed the flat and clean fracture surface of the matrix with some cracks in the matrix side of the fiber-matrix interface in the conventional GRPC whereas nanophased GRPC showed rougher fracture of the matrix with randomly but uniformly

The author is truly grateful to his hardworking and dedicated graduate students: Mr. Muhammad Enayet Hossain, Mr. Mohammad Washim Dewan, Mr. Kazi Al Imran, and Mr. Chinedu Okoro for their direct and indirect contribution in this article. The author also likes to express his gratitude towards Dr. Mahesh Hosur and Dr. Vijay Rangari for their cooperation and advice. The author also appreciates the help from the staff members of

The author's special thanks are extended to Dr. Shaik Jeelani, Vice President, Research and Sponsored Programs at Tuskegee University for his invaluable support and guidance

The author acknowledges the financial support of NSF-EPSCoR Grant No. EPS-0814103 and

Finally, special thanks go to his wife, Dr. Shamim Ara Begum, and son, Ahnaf Hossain, for their love, patience, and mental support. Above all, the author thanks to God, Almighty for

Bruzaud, S., & Bourmaud, A. (2007). Thermal degradation and (nano) mechanical behavior

Giannelis, E.P. (1998). Polymer-layered silicate nanocomposites: synthesis, properties and

of layered silicate reinforced poly(3-hydroxybutyrate-co-3-hydroxyvalerate)

distributed CNFs throughout the matrix that appeared to cling to fibers well.

mechanical and magnetic mixing methods.

Tuskegee Center for Advanced Materials (T-CAM).

throughout this work both professionally and personally.

NSF-RISE Grant No. HRD-0833158 for this research work.

nanocomposites. *Polym Test*, 26, (2007), pp. (652-659).

applications. *Appl Organomet Chem*, 12, (1998), pp. (675-680).

separated CNFs.

**5. Aknowledgements** 

His love and mercy.

**6. References** 

SEM micrographs due to the presence of CNFs.

mechanical interlocking of the CNFs with the fibers.

matrix was observed in SEM micrographs of nanophased GRPC.


**37** 

Feng Shi

*P.R. China* 

**Preparation and Characterization of Dielectric** 

Barium-strontium-zinc niobate (BSZN) [(BaxSr1-x)(Zn1/3Nb2/3)O3, where *x* is the mole fraction and 0 ≤ *x* ≤ 1] is one of the *AB B O* ( ) 13 23 3 ′ ′′ type microwave dielectric ceramics which have many advantage properties at very high microwave frequency, such as extremely low dielectric loss and near zero temperature coefficient of resonance frequency with low cost (Xu et al., 2006; Yu et al., 2006; Ianculescu et al., 2007; Huang et al., 2006; Varma et al., 2006). Due to these properties, BSZN has several potential applications in the fields of satellite communication and radar and mobile communication systems. As promising materials used in microelectronic and microwave integration circuit, microwave dielectric ceramic thin

Radio frequency (RF) magnetron sputtering is a dominant technique to grow thin films because a large quantity of thin films can be prepared at relatively high purity and low cost. The growth of thin films using (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramic as target materials has not been reported all over the world except my group (Cui et al., 2010; Shi et al., 2010) and there is very little information for direct reference. In this article, the thin films were initially prepared by radio frequency magnetron sputtering system using sintered (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramic as target. Then the deposited samples were annealed in oxygen ambience at different powers of 150 W, 200 W, and 250 W, different pressures of 0.1 Pa, 0.25 Pa, 0.5 Pa, 0.7 Pa, and 1.0 Pa, different annealing temperatures of 850 ℃, 1000 ℃, and 1150 ℃ and different annealing times of 15 min, 30 min, 45 min, and 60 min. The microstructure, components and surface morphology properties are investigated in detail, and the effect of experimental conditions on the growth of the thin

Ceramic thin films were deposited on SiO2 (110) substrates by adopting sintered (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramics as sputtering target with a size of 62- × 3 mm in a JGP-450 radio frequency magnetron sputtering system. The SiO2 (110) substrate was ultrasonically cleaned in an acetone and followed by rinsing in de-ionized

films will attract great attention in the near future (Huang et al., 2006).

**1. Introduction** 

films is studied in particular.

**2. Experimental procedure** 

**Thin Films by RF Magnetron-Sputtering with** 

**(Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target** 

*College of Physics & Electronics, Shandong Normal University,* 

Yoonessi, M., Toghiani, H., Wheeler, R., Porcar, L., Kline, S., & Pittman, C. (2008). Neutron scattering, electron microscopy and dynamic mechanical studies of carbon nanofiber/phenolic resin composites. *Carbon*, 46, (2008), pp. (577-588).

## **Preparation and Characterization of Dielectric Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target**

Feng Shi

*College of Physics & Electronics, Shandong Normal University, P.R. China* 

## **1. Introduction**

744 Scanning Electron Microscopy

Yoonessi, M., Toghiani, H., Wheeler, R., Porcar, L., Kline, S., & Pittman, C. (2008). Neutron

nanofiber/phenolic resin composites. *Carbon*, 46, (2008), pp. (577-588).

scattering, electron microscopy and dynamic mechanical studies of carbon

Barium-strontium-zinc niobate (BSZN) [(BaxSr1-x)(Zn1/3Nb2/3)O3, where *x* is the mole fraction and 0 ≤ *x* ≤ 1] is one of the *AB B O* ( ) 13 23 3 ′ ′′ type microwave dielectric ceramics which

have many advantage properties at very high microwave frequency, such as extremely low dielectric loss and near zero temperature coefficient of resonance frequency with low cost (Xu et al., 2006; Yu et al., 2006; Ianculescu et al., 2007; Huang et al., 2006; Varma et al., 2006). Due to these properties, BSZN has several potential applications in the fields of satellite communication and radar and mobile communication systems. As promising materials used in microelectronic and microwave integration circuit, microwave dielectric ceramic thin films will attract great attention in the near future (Huang et al., 2006).

Radio frequency (RF) magnetron sputtering is a dominant technique to grow thin films because a large quantity of thin films can be prepared at relatively high purity and low cost. The growth of thin films using (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramic as target materials has not been reported all over the world except my group (Cui et al., 2010; Shi et al., 2010) and there is very little information for direct reference. In this article, the thin films were initially prepared by radio frequency magnetron sputtering system using sintered (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramic as target. Then the deposited samples were annealed in oxygen ambience at different powers of 150 W, 200 W, and 250 W, different pressures of 0.1 Pa, 0.25 Pa, 0.5 Pa, 0.7 Pa, and 1.0 Pa, different annealing temperatures of 850 ℃, 1000 ℃, and 1150 ℃ and different annealing times of 15 min, 30 min, 45 min, and 60 min. The microstructure, components and surface morphology properties are investigated in detail, and the effect of experimental conditions on the growth of the thin films is studied in particular.

### **2. Experimental procedure**

Ceramic thin films were deposited on SiO2 (110) substrates by adopting sintered (BaxSr1-x)(Zn1/3Nb2/3)O3 microwave dielectric ceramics as sputtering target with a size of 62- × 3 mm in a JGP-450 radio frequency magnetron sputtering system. The SiO2 (110) substrate was ultrasonically cleaned in an acetone and followed by rinsing in de-ionized

Preparation and Characterization of Dielectric

(a) 150 W, (b) 200 W, (c) 250 W.

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 747

Figure 2 shows the XRD spectra obtained from the thin films deposited at the pressure of 0.25 Pa, annealing at 1150 °C for 30 min and different powers of (a) 150 W, (b) 200 W, (c) 250 W. In Figure 2 (a), there are a small number of diffraction peaks with low intensity in comparison to those of the other two samples. This phenomenon is due to the fact that the low kinetic energies of the sputtered-ejected ions are not sufficient for the arrangement and crystallization of particles on the substrates when sputtered at lower RF power (Wang et al., 2004). On the other hand, at 200 W (as shown in Figure 1 (b)), the amorphous thin films begin to crystallize, and more diffraction peaks appear in the sample with the strongest intensity. In Figure 2 (b), major peaks are identified to be Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O 5.78 as compared with JCPDS files No. 39-0265 and No. 31-0166 (International Center for

Fig. 2. X-ray diffraction patterns of the thin films deposited at 0.25 Pa and different powers:

These peaks are caused by the volatilization of ZnO during the process of sputtering and annealing; this is the same as that stated in reference (Huang et al., 2006). There is no preferential orientation for the thin films, i.e., they are randomly orientated. The diffraction peaks in every crystal plane of the films are almost complete, which indicates the grains have excellent crystal quality. When sputtered at 250 W (as shown in Figure 2 (c)), both the quantity and intensity of the thin film diffraction peaks are lower and weaker than that of the sample sputtered at 200 W. Both the crystal quality of the thin films and the intensity of the reflection peaks are at their best when the RF power is of 200 W, because the increase in kinetic energies of the sputter-ejected species accelerates the arrangement and crystallization

Diffraction Data, 2002), which is different from the target components.

water several times. The chamber pressure was maintained at 1.0 × 10-3 Pa, and the argon gas (99.999%) was then introduced into the chamber. The distance between the target and the substrates was 11 cm, the substrate was heated to 610 °C and the sputtering time is 3 hrs per time. A post-deposition annealing was needed so that the deposited films have a wellcrystallized structure. When the conventional tube furnace was heated up to a certain temperature, the quartz boat with the samples was placed into the constant temperature region. Then the flowing O2 (99.999℅) was introduced into the tube and the samples were annealed at O2 atmosphere with a flow rate of 500 ml/min. After being annealed, the samples were taken out for characterization.

A Rigaku D/max-rB X-ray diffraction (XRD) meter with Cu Kα-line, X-ray photoelectron spectroscope (XPS), a Hitachi S-450 scanning electron microscope (SEM), a Park Autoprobe CP atomic force microscope (AFM) and a Hitachi H-8010 transmission electron microscopy (TEM) were applied to characterize the microstructure, components and surface morphology properties of the thin films and to study their crystallinity. The morphology of cross-section of the thin film was examined by SEM (JEOL JSM-6390).

## **3. Results and discussion**

### **3.1 Microstructure and components analysis**

Figure 1 shows the X-ray diffraction patterns of the as-cast thin films at different powers (the sputtering pressure is 0.25 Pa).

Fig. 1. X-ray diffraction patterns of the as-cast thin films at different powers. (a) 150 W, (b) 200 W, (c) 250 W.

Figure 1 shows no diffraction peak exists, only three weak diffraction packets, which indicates that the thin films do not crystallize, but only exist in an amorphous state. The intensities and shapes are different obviously for these samples sputtered at different powers. The intensity is the strongest when sputtering power is 200 W.

water several times. The chamber pressure was maintained at 1.0 × 10-3 Pa, and the argon gas (99.999%) was then introduced into the chamber. The distance between the target and the substrates was 11 cm, the substrate was heated to 610 °C and the sputtering time is 3 hrs per time. A post-deposition annealing was needed so that the deposited films have a wellcrystallized structure. When the conventional tube furnace was heated up to a certain temperature, the quartz boat with the samples was placed into the constant temperature region. Then the flowing O2 (99.999℅) was introduced into the tube and the samples were annealed at O2 atmosphere with a flow rate of 500 ml/min. After being annealed, the

A Rigaku D/max-rB X-ray diffraction (XRD) meter with Cu Kα-line, X-ray photoelectron spectroscope (XPS), a Hitachi S-450 scanning electron microscope (SEM), a Park Autoprobe CP atomic force microscope (AFM) and a Hitachi H-8010 transmission electron microscopy (TEM) were applied to characterize the microstructure, components and surface morphology properties of the thin films and to study their crystallinity. The morphology of

Figure 1 shows the X-ray diffraction patterns of the as-cast thin films at different powers (the

cross-section of the thin film was examined by SEM (JEOL JSM-6390).

Fig. 1. X-ray diffraction patterns of the as-cast thin films at different powers.

powers. The intensity is the strongest when sputtering power is 200 W.

Figure 1 shows no diffraction peak exists, only three weak diffraction packets, which indicates that the thin films do not crystallize, but only exist in an amorphous state. The intensities and shapes are different obviously for these samples sputtered at different

samples were taken out for characterization.

**3.1 Microstructure and components analysis** 

**3. Results and discussion** 

sputtering pressure is 0.25 Pa).

(a) 150 W, (b) 200 W, (c) 250 W.

Figure 2 shows the XRD spectra obtained from the thin films deposited at the pressure of 0.25 Pa, annealing at 1150 °C for 30 min and different powers of (a) 150 W, (b) 200 W, (c) 250 W.

In Figure 2 (a), there are a small number of diffraction peaks with low intensity in comparison to those of the other two samples. This phenomenon is due to the fact that the low kinetic energies of the sputtered-ejected ions are not sufficient for the arrangement and crystallization of particles on the substrates when sputtered at lower RF power (Wang et al., 2004). On the other hand, at 200 W (as shown in Figure 1 (b)), the amorphous thin films begin to crystallize, and more diffraction peaks appear in the sample with the strongest intensity. In Figure 2 (b), major peaks are identified to be Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O 5.78 as compared with JCPDS files No. 39-0265 and No. 31-0166 (International Center for Diffraction Data, 2002), which is different from the target components.

Fig. 2. X-ray diffraction patterns of the thin films deposited at 0.25 Pa and different powers: (a) 150 W, (b) 200 W, (c) 250 W.

These peaks are caused by the volatilization of ZnO during the process of sputtering and annealing; this is the same as that stated in reference (Huang et al., 2006). There is no preferential orientation for the thin films, i.e., they are randomly orientated. The diffraction peaks in every crystal plane of the films are almost complete, which indicates the grains have excellent crystal quality. When sputtered at 250 W (as shown in Figure 2 (c)), both the quantity and intensity of the thin film diffraction peaks are lower and weaker than that of the sample sputtered at 200 W. Both the crystal quality of the thin films and the intensity of the reflection peaks are at their best when the RF power is of 200 W, because the increase in kinetic energies of the sputter-ejected species accelerates the arrangement and crystallization

Preparation and Characterization of Dielectric

the increase in annealing temperature.

( )( )( )

crystal structures are obtained accordingly.

2 22 <sup>2</sup> *d ha ka lc hkl* / // <sup>−</sup>

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 749

Figure 3 (b), because this thin film has transited from the amorphous state to the crystalline state at higher annealing temperature. When the annealing temperature increases to 1150 °C, as shown in Figure 3 (d), which is the same as Figure 2 (b), all peaks of crystal planes have appeared, which reveals that the thin film crystallizes completely, but there is no preferential growth. Moreover, the intensity of diffraction peak in Figure 3 (d) enhances greatly due to good crystallization at the highest annealing temperature. Figure 3 states there is steady growth for the strength and quantity of the thin film diffraction peaks with

Fig. 3. X-ray diffraction patterns of the as-cast thin films and the thin films annealing at

According to the (320), (410), and (330) peaks, through the formula (Wang et al., 2004)

The intensity is related with the crystallinity, the thickness of the thin films and the consistency, whereas the full width at half maximum (FWHM) value only reveals the crystallinity of the thin films. Due to different volatilization at different annealing temperatures, the thickness and consistency of peaks are different, therefore, the intensities of diffraction peaks are also different. Generally speaking, the annealing treatment can

= ++ , where *hkl <sup>d</sup>* is interplanar crystal spacing, and h, k and l are crystal indices, we can calculate the lattice constants of Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases: a=b=12.456 Å and c=3.952 Å of Ba0.5Sr0.5Nb2O6 phase and a=b=12.430 Å and c=3.913 Å of Ba0.27Sr0.75Nb2O5.78 phase. A higher annealing temperature accelerates the material migration and diminishes the crystallographic defects and dislocations, thus promots crystallization of the thin films, and therefore, the excellent

different temperatures for 30 min,(a) as-cast, (b) 850 ℃, (c) 1000 ℃, (d) 1150 ℃.

1

of particles on the substrate surface (Hsi et al., 2003). However, when the power is of 250 W, the kinetic energy of the sputter -ejected species is very high. As a result, the particles excited from the ceramic target were thrown away and not deposited on the substrate.

On the other hand, the greater the sputtering powers, the higher the kinetic energies, and the quicker the deposition rate. Deposition rate is lower at 150 W, and is higher at 250 W, i.e., appropriate deposition rate attributes to the film growth; therefore, 200 W is the optimum condition for the growth of the thin films.

In order to further investigate the growth of the thin films, the full-width at half-maximum (FWHM) of the (410) peaks of the Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases is analyzed. According to Scherrer`s formula (Klung et al., 1974), the grain size in (410) orientation can be estimated as follows:

$$L\_{\text{(410)}} = \frac{k\mathcal{X}}{\beta\_0 \cos\theta} \,' \,'$$

where *k* is a constant with a value of about 0.89 for Cu target, λ is the X-ray wavelength, β<sup>0</sup> is the FWHM of (410) peak, and θ is the diffraction angle. The estimated results are listed in Table 1.


Table 1. The grain size of Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases in the thin films estimated by Scherrer`s formula.

Therefore, with an increase in the sputtering power, the grains sizes of the thin films reach a maximum of 36.3 nm at 200 W and then decrease to 30.6 nm at 250 W.

Figure 3 shows X-ray diffraction patterns of the as-cast thin films and the samples annealing at different temperatures for 30 min (sputtered at 200 W and 0.25 Pa).

Figure 3 (a) indicates there is only a single crystal SiO2 peak instead of diffraction peaks of other phases. It is well known that the formation of crystalline phase in thin films is affected by two important factors: one is the temperature in the sputtering process and the other is the annealing temperature. Therefore, during sputtering, the temperature of SiO2 substrate, about 610 °C, is insufficient to initiate the formation of crystallization, thus the annealing treatment after sputtering is crucial. When the annealing temperature is 850 °C, as shown in Figure 3 (b), one weak peak appears, which is too weak to separate clearly and shows the thin film is in an amorphous state. But when the annealing temperature increases to 1000 °C, more and stronger peaks occur in Figure 3 (c). The main phases are also Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78, but there is no peak of (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3. The difference between the components of the films and target attributes to the volatilization of ZnO during the process of sputtering and annealing, which is the same as that stated in Figure 2. The intensity of diffraction peak in Figure 3 (c) is stronger than that of the sample shown in

of particles on the substrate surface (Hsi et al., 2003). However, when the power is of 250 W, the kinetic energy of the sputter -ejected species is very high. As a result, the particles excited from the ceramic target were thrown away and not deposited on the substrate.

On the other hand, the greater the sputtering powers, the higher the kinetic energies, and the quicker the deposition rate. Deposition rate is lower at 150 W, and is higher at 250 W, i.e., appropriate deposition rate attributes to the film growth; therefore, 200 W is the

In order to further investigate the growth of the thin films, the full-width at half-maximum (FWHM) of the (410) peaks of the Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases is analyzed. According to Scherrer`s formula (Klung et al., 1974), the grain size in (410) orientation can be

<sup>0</sup> cos

β λ

θ<sup>=</sup> ,

/0 FWHM/rad L/nm

λ

is the diffraction angle. The estimated results are

is the X-ray wavelength,

( ) 410

θ

θ

maximum of 36.3 nm at 200 W and then decrease to 30.6 nm at 250 W.

at different temperatures for 30 min (sputtered at 200 W and 0.25 Pa).

B1(150 w) (410) 29.51 0.0061 24.1 B2(200 w) (410) 29.57 0.0043 36.3 B3(250 w) (410) 29.51 0.0050 30.6

Table 1. The grain size of Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases in the thin films

Therefore, with an increase in the sputtering power, the grains sizes of the thin films reach a

Figure 3 shows X-ray diffraction patterns of the as-cast thin films and the samples annealing

Figure 3 (a) indicates there is only a single crystal SiO2 peak instead of diffraction peaks of other phases. It is well known that the formation of crystalline phase in thin films is affected by two important factors: one is the temperature in the sputtering process and the other is the annealing temperature. Therefore, during sputtering, the temperature of SiO2 substrate, about 610 °C, is insufficient to initiate the formation of crystallization, thus the annealing treatment after sputtering is crucial. When the annealing temperature is 850 °C, as shown in Figure 3 (b), one weak peak appears, which is too weak to separate clearly and shows the thin film is in an amorphous state. But when the annealing temperature increases to 1000 °C, more and stronger peaks occur in Figure 3 (c). The main phases are also Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78, but there is no peak of (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3. The difference between the components of the films and target attributes to the volatilization of ZnO during the process of sputtering and annealing, which is the same as that stated in Figure 2. The intensity of diffraction peak in Figure 3 (c) is stronger than that of the sample shown in

where *k* is a constant with a value of about 0.89 for Cu target,

Sample (hkl) 2

*<sup>k</sup> <sup>L</sup>*

optimum condition for the growth of the thin films.

<sup>0</sup> is the FWHM of (410) peak, and

estimated by Scherrer`s formula.

estimated as follows:

listed in Table 1.

β

Figure 3 (b), because this thin film has transited from the amorphous state to the crystalline state at higher annealing temperature. When the annealing temperature increases to 1150 °C, as shown in Figure 3 (d), which is the same as Figure 2 (b), all peaks of crystal planes have appeared, which reveals that the thin film crystallizes completely, but there is no preferential growth. Moreover, the intensity of diffraction peak in Figure 3 (d) enhances greatly due to good crystallization at the highest annealing temperature. Figure 3 states there is steady growth for the strength and quantity of the thin film diffraction peaks with the increase in annealing temperature.

Fig. 3. X-ray diffraction patterns of the as-cast thin films and the thin films annealing at different temperatures for 30 min,(a) as-cast, (b) 850 ℃, (c) 1000 ℃, (d) 1150 ℃.

According to the (320), (410), and (330) peaks, through the formula (Wang et al., 2004) ( )( )( ) 1 2 22 <sup>2</sup> *d ha ka lc hkl* / // <sup>−</sup> = ++ , where *hkl <sup>d</sup>* is interplanar crystal spacing, and h, k and l are crystal indices, we can calculate the lattice constants of Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 phases: a=b=12.456 Å and c=3.952 Å of Ba0.5Sr0.5Nb2O6 phase and a=b=12.430 Å and c=3.913 Å of Ba0.27Sr0.75Nb2O5.78 phase. A higher annealing temperature accelerates the material migration and diminishes the crystallographic defects and dislocations, thus promots crystallization of the thin films, and therefore, the excellent crystal structures are obtained accordingly.

The intensity is related with the crystallinity, the thickness of the thin films and the consistency, whereas the full width at half maximum (FWHM) value only reveals the crystallinity of the thin films. Due to different volatilization at different annealing temperatures, the thickness and consistency of peaks are different, therefore, the intensities of diffraction peaks are also different. Generally speaking, the annealing treatment can

Preparation and Characterization of Dielectric

appropriate for the thin films being annealed for 30 min.

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 751

improve crystallization of the thin films, but the heat energy is not excessive. That is, it is

Fig. 4. X-ray diffraction patterns of the thin films annealing at 1150 °C for different times

(sputtered at 200 w and 0.25 Pa).(a) 15 min, (b) 30 min, (c) 45 min, (d) 60 min.

Fig. 5. The relation between the FWHM values and the grain size with the different

Figure 6 shows X-ray diffraction patterns of the thin films at different pressures (sputtered

annealing times.

at 200 W, annealing at 1150 °C for 30 min).

promote atomic mobility and enhance the ability of atoms so that they can find the most energetically favored points; therefore, with the increase in the annealing temperature, the crystallographic defects including dislocations and vacancies in the thin films decrease rapidly, i.e., the appropriate annealing temperature is the key factor for a better crystal structure. The Scherrer formula (Schroeder et al., 1968)tells us that the smaller of the FWHM value, the better of the C-axis-preferred orientation, and the larger of the grain size and the better of the crystalline quality of the thin films. That is, the thin film which has the smallest FWHM value possesses the best crystalline quality and the largest grain size. The relation between FWHM and the annealing temperature is listed in Table 2.



As shown in Table 2, the value of FWHM decreases with the increase in annealing temperature. The sample annealed at 1150 °C has the smallest value of 0.125, which shows the best crystalline quality (Zheng et al., 1993), because the energy supplied from the outside environment during the annealing process is sufficient to make the films crystallize completely at higher temperature.

Figure 4 shows X-ray diffraction patterns of the thin films annealing at 1150 °C for different times (sputtered at 200 W and 0.25 Pa).

In Figure 4 (a), there is only one diffraction peak with low intensity, and the phase is Ba0.27Sr0.75Nb2O5.78. This thin film just began to crystallize at 1150 °C for 15 min, because there was no enough time for the atoms migration on the substrate surface (Zannetti et al., 1969; Yang et al., 1998). Figure 4 (b) is the same as shown in Figure 3 (d): there is no other peak, which indicates that the thin film has high purity. Figure 4 (c) shows the intensity and number of the diffraction peaks decrease greatly in comparison to that shown in Figure 4 (b) because of excess volatilization of ZnO and recrystallization of the grains at longer annealing time, and the trend is clear as seen in Figure 4 (d). A new phase has appeared, named Ba5.75Nb2.25O11.375 in Figure 4 (c) and Figure 4 (d).

Figure 5 is the relation between the FWHM values and the grain sizes with the different annealing times.

Annealing for 15 min, the samples cannot crystallize because of deficiency of the heat energy for material migration, thus the FWHM value is large; whereas after annealing for longer time, such as 45 min and 60 min, more heat energy will lead to excess volatilization of ZnO and recrystallization of the grains, thus lessening the FWHM value. As for the sample annealing for 30 min, the FWHM value is at its minimum and the grain size is the largest with the best crystallinity, as compared with the other samples annealing for 15 min, 45 min, and 60 min, because of the sufficient heat energy supplied by the outside environment

promote atomic mobility and enhance the ability of atoms so that they can find the most energetically favored points; therefore, with the increase in the annealing temperature, the crystallographic defects including dislocations and vacancies in the thin films decrease rapidly, i.e., the appropriate annealing temperature is the key factor for a better crystal structure. The Scherrer formula (Schroeder et al., 1968)tells us that the smaller of the FWHM value, the better of the C-axis-preferred orientation, and the larger of the grain size and the better of the crystalline quality of the thin films. That is, the thin film which has the smallest FWHM value possesses the best crystalline quality and the largest grain size. The relation

θ

(410) 29.6085 0.291

(410) 29.6085 0.215

(410) 29.6085 0.125

/0 FWHM/0

between FWHM and the annealing temperature is listed in Table 2.

Table 2. The relation between FWHM and annealing temperatures of the samples.

As shown in Table 2, the value of FWHM decreases with the increase in annealing temperature. The sample annealed at 1150 °C has the smallest value of 0.125, which shows the best crystalline quality (Zheng et al., 1993), because the energy supplied from the outside environment during the annealing process is sufficient to make the films crystallize

Figure 4 shows X-ray diffraction patterns of the thin films annealing at 1150 °C for different

In Figure 4 (a), there is only one diffraction peak with low intensity, and the phase is Ba0.27Sr0.75Nb2O5.78. This thin film just began to crystallize at 1150 °C for 15 min, because there was no enough time for the atoms migration on the substrate surface (Zannetti et al., 1969; Yang et al., 1998). Figure 4 (b) is the same as shown in Figure 3 (d): there is no other peak, which indicates that the thin film has high purity. Figure 4 (c) shows the intensity and number of the diffraction peaks decrease greatly in comparison to that shown in Figure 4 (b) because of excess volatilization of ZnO and recrystallization of the grains at longer annealing time, and the trend is clear as seen in Figure 4 (d). A new phase has appeared,

Figure 5 is the relation between the FWHM values and the grain sizes with the different

Annealing for 15 min, the samples cannot crystallize because of deficiency of the heat energy for material migration, thus the FWHM value is large; whereas after annealing for longer time, such as 45 min and 60 min, more heat energy will lead to excess volatilization of ZnO and recrystallization of the grains, thus lessening the FWHM value. As for the sample annealing for 30 min, the FWHM value is at its minimum and the grain size is the largest with the best crystallinity, as compared with the other samples annealing for 15 min, 45 min, and 60 min, because of the sufficient heat energy supplied by the outside environment

Sample phase (hkl) 2

Ba0.27Sr0.75Nb2O5.78

Ba0.27Sr0.75Nb2O5.78

Ba0.27Sr0.75Nb2O5.78

named Ba5.75Nb2.25O11.375 in Figure 4 (c) and Figure 4 (d).

B1(850 °C) Ba0.5Sr0.5Nb2O6

B2(1000 °C) Ba0.5Sr0.5Nb2O6

B3(1150 °C) Ba0.5Sr0.5Nb2O6

completely at higher temperature.

annealing times.

times (sputtered at 200 W and 0.25 Pa).

improve crystallization of the thin films, but the heat energy is not excessive. That is, it is appropriate for the thin films being annealed for 30 min.

Fig. 4. X-ray diffraction patterns of the thin films annealing at 1150 °C for different times (sputtered at 200 w and 0.25 Pa).(a) 15 min, (b) 30 min, (c) 45 min, (d) 60 min.

Fig. 5. The relation between the FWHM values and the grain size with the different annealing times.

Figure 6 shows X-ray diffraction patterns of the thin films at different pressures (sputtered at 200 W, annealing at 1150 °C for 30 min).

Preparation and Characterization of Dielectric

systematic error of the XRD.

Pa, 1150 oC, 30 min).

**3.2 Morphology analysis** 

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 753

peak is usually observed in binding energy area of 529-535 eV; the peaks in 529-530 eV are ascribed to the lattice oxygen, the peaks for binding energy ranging from 530.0 eV to 530.9 eV correspond to chemisorbed oxygen, and therefore, the O1s peak corresponds to lattice oxygen. The peaks of C1s attribute to the standard sample during XPS testing. There is a weak peak corresponding to the Zn element in Figure 7, indicating that a part of Zn element exists in the thin film although the other part of ZnO is evaporated during the sputtering and annealing process, which is not consistent with the results shown in XRD because of the

Fig. 7. XPS spectra obtained from the thin films deposited at 150 W, 200 W, and 250 W (0.25

Figure 8 shows the typical SEM images of the samples sputtered at different powers and

annealing at 1150 oC for 30 min (sputtering pressure: 0.25 Pa).

As seen from Figure 6, the trend is the same as that stated above in Figure 2 - Figure 4, i.e., the higher of the pressure, the lower of the intensity and the less of the number of the diffraction peaks. There is greater atomic density in vacuum chamber at higher gaseous pressure, i.e., the chances of atomic collision increase, and the energy loss of atoms is enhanced during material migration, thus lessening the quantity of the atoms reaching the substrate surface. The nucleation density decreases subsequently due to less particles; therefore, the growth of the thin films is weakened. The imbalance between Ba and Sr ions is closely related with saturated gaseous pressure, for instance, the stoichiometric ratio imbalance of Ba and Sr elements becomes inconspicuous at lower pressure as compared to the phases in Figure 6 (a), (b) and (c).

Fig. 6. X-ray diffraction patterns of the thin films at different pressures (sputtered at 200 W, annealed at 1150 °C for 30 min). (a) 0.25 Pa, (b) 0.50 Pa, (c) 0.70 Pa.

In Figure 6 (a), the grain growth is relatively complete, and therefore, more and stronger diffraction peaks appear. When the pressure increases to 0.5 Pa, the intensity and number of peaks decrease because of more collision among particles caused by higher pressure, thus material migration is inhibited and it's the same trend as the sample shown in Figure 6 (c).

According to the results of XRD, we pay much attention to the samples deposited at 150 W, 200 W, and 250 W (the sputtering pressure are of 0.25 Pa and annealed at 1150 oC for 30 min) and study their components; Figure 7 shows the XPS spectra of these samples. In Figure 7, the general scan of binding energy ranging from 0 to 1100 eV, the peaks of Sr3d, Nb3p3/2, Ba3d, C1s, Nb3d, Zn3d and O1s are observed.

As seen in Figure 7, the main elements in the thin films are Ba, Sr, Nb, Zn, and O element. Since the samples selected for XPS analysis are exposed to the air, the elements of O and C arise from the thin film solution. O1s and C1s core level peaks are always detected, and O1s peak centers at 529.8 eV. Amanullah et al., (Amanullah et al., 1998) have reported that O1s

As seen from Figure 6, the trend is the same as that stated above in Figure 2 - Figure 4, i.e., the higher of the pressure, the lower of the intensity and the less of the number of the diffraction peaks. There is greater atomic density in vacuum chamber at higher gaseous pressure, i.e., the chances of atomic collision increase, and the energy loss of atoms is enhanced during material migration, thus lessening the quantity of the atoms reaching the substrate surface. The nucleation density decreases subsequently due to less particles; therefore, the growth of the thin films is weakened. The imbalance between Ba and Sr ions is closely related with saturated gaseous pressure, for instance, the stoichiometric ratio imbalance of Ba and Sr elements becomes inconspicuous at lower pressure as compared to

Fig. 6. X-ray diffraction patterns of the thin films at different pressures (sputtered at 200 W,

In Figure 6 (a), the grain growth is relatively complete, and therefore, more and stronger diffraction peaks appear. When the pressure increases to 0.5 Pa, the intensity and number of peaks decrease because of more collision among particles caused by higher pressure, thus material migration is inhibited and it's the same trend as the sample shown in Figure 6 (c). According to the results of XRD, we pay much attention to the samples deposited at 150 W, 200 W, and 250 W (the sputtering pressure are of 0.25 Pa and annealed at 1150 oC for 30 min) and study their components; Figure 7 shows the XPS spectra of these samples. In Figure 7, the general scan of binding energy ranging from 0 to 1100 eV, the peaks of Sr3d, Nb3p3/2,

As seen in Figure 7, the main elements in the thin films are Ba, Sr, Nb, Zn, and O element. Since the samples selected for XPS analysis are exposed to the air, the elements of O and C arise from the thin film solution. O1s and C1s core level peaks are always detected, and O1s peak centers at 529.8 eV. Amanullah et al., (Amanullah et al., 1998) have reported that O1s

annealed at 1150 °C for 30 min). (a) 0.25 Pa, (b) 0.50 Pa, (c) 0.70 Pa.

Ba3d, C1s, Nb3d, Zn3d and O1s are observed.

the phases in Figure 6 (a), (b) and (c).

peak is usually observed in binding energy area of 529-535 eV; the peaks in 529-530 eV are ascribed to the lattice oxygen, the peaks for binding energy ranging from 530.0 eV to 530.9 eV correspond to chemisorbed oxygen, and therefore, the O1s peak corresponds to lattice oxygen. The peaks of C1s attribute to the standard sample during XPS testing. There is a weak peak corresponding to the Zn element in Figure 7, indicating that a part of Zn element exists in the thin film although the other part of ZnO is evaporated during the sputtering and annealing process, which is not consistent with the results shown in XRD because of the systematic error of the XRD.

Fig. 7. XPS spectra obtained from the thin films deposited at 150 W, 200 W, and 250 W (0.25 Pa, 1150 oC, 30 min).

#### **3.2 Morphology analysis**

Figure 8 shows the typical SEM images of the samples sputtered at different powers and annealing at 1150 oC for 30 min (sputtering pressure: 0.25 Pa).

Preparation and Characterization of Dielectric

longer material migration time.

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 755

Figure 8 (c). As seen in Figure 10 (b), the film is cracked and many gaps and holes exist among each fragmental structure, which attributes to recrystallization of grains due to

(a) (b) (c) Fig. 9. Typical SEM images of the as-cast thin film and the thin films sputtered at 200 W, 0.25

(a) (b)

Figure 11 shows the SEM images of the samples sputtered at 200 W and different pressures

As seen from Figure 11, surface morphology differs with the sputtering gaseous pressures. Morphology shown in Figure 11 (a) is similar to that shown in Figure 8 (b), that is, the sample in Figure 11 (a) has perfect crystalline and surface morphology, which indicates that an appropriate low pressure can contribute to the film growth. With the increase of sputtering pressures, more holes and gaps form on the surface accompanied by the decrease

Fig. 10. SEM images of the thin films sputtered at 200 W and 250 W, respectively, after

annealing at 1150 oC for 45 min. (a)200 W, (b)250 W.

of 0.10 Pa, 0.50 Pa, and 0.70 Pa, after annealing at 1150 oC for 30 min.

Pa after annealing at different temperatures. (a) as-cast, (b) 850 oC, (c) 1000 oC.

Fig. 8. Typical SEM images of the thin films sputtered at different powers. (a) 150 W, (b) 200 W, (c) 250 W.

When deposited at 150 W, the grains of the thin films have been formed with uniform and loose surface, and there are many holds on the thin film surface. During the coalescence stage of the sputtering process, fine particles do not have enough kinetic energy to grow into larger grains. After the annealing process, the thin films have changed from amorphous phase to crystal phase partially. Therefore, the intensities of diffraction peaks in XRD spectra are at their lowest, probably due to small deposition rate at 150 W. In Figure 8 (b), it is evident that the grains have well-defined boundaries around them and homogenously distribute on the substrates surface with uniform grain size. The grains are rod-like structures which form during the annealing process because the greater grains can combine with the smaller ones around them to generate rod-like grains. With higher energy gas plasma caused by higher RF power, more fine particles were sputtered from the target and these high-energy particles can migrate and combine together with each other on the substrate surface and form high quality crystalline films, which complies with the result of the XRD analysis (as shown in Figure 2 (b)). The sample in Figure 8 (c) is uniform and dense with strip shape grains, because adequate energy supplied during material migration promotes the formation of the thin films and crystallization of grains. Therefore, the thin films sputtered at higher powers possess perfect surface morphology.

Figure 9 shows the typical SEM images of the as-cast sample and the samples sputtered at 200 W, 0.25 Pa after annealing at different temperatures for 30 min.

As compared with the sample annealing at 1150 oC in Figure 8 (b), the as-cast sample (Figure 9 (a)) has a clean surface, but no grain; whereas when annealing at 850 oC, less grains appear; when the annealing temperature is 1000 oC, the amount of grains increases with fine particles covered on the surface. That is, crystallization is not complete at lower annealing temperature.

Figure 10 shows the SEM images of the samples sputtered at 200 W and 250 W, after annealing at 1150 oC for 45 min (sputtering pressure: 0.25 Pa).

In comparison to Figure 8 (b), there are more holes in Figure 10 (a) and the thin films are composed of block structures; this situation is similar to that shown in Figure 10 (b) and

(a) (b) (c) Fig. 8. Typical SEM images of the thin films sputtered at different powers. (a) 150 W, (b) 200

When deposited at 150 W, the grains of the thin films have been formed with uniform and loose surface, and there are many holds on the thin film surface. During the coalescence stage of the sputtering process, fine particles do not have enough kinetic energy to grow into larger grains. After the annealing process, the thin films have changed from amorphous phase to crystal phase partially. Therefore, the intensities of diffraction peaks in XRD spectra are at their lowest, probably due to small deposition rate at 150 W. In Figure 8 (b), it is evident that the grains have well-defined boundaries around them and homogenously distribute on the substrates surface with uniform grain size. The grains are rod-like structures which form during the annealing process because the greater grains can combine with the smaller ones around them to generate rod-like grains. With higher energy gas plasma caused by higher RF power, more fine particles were sputtered from the target and these high-energy particles can migrate and combine together with each other on the substrate surface and form high quality crystalline films, which complies with the result of the XRD analysis (as shown in Figure 2 (b)). The sample in Figure 8 (c) is uniform and dense with strip shape grains, because adequate energy supplied during material migration promotes the formation of the thin films and crystallization of grains. Therefore, the thin

Figure 9 shows the typical SEM images of the as-cast sample and the samples sputtered at

As compared with the sample annealing at 1150 oC in Figure 8 (b), the as-cast sample (Figure 9 (a)) has a clean surface, but no grain; whereas when annealing at 850 oC, less grains appear; when the annealing temperature is 1000 oC, the amount of grains increases with fine particles covered on the surface. That is, crystallization is not complete at lower annealing

Figure 10 shows the SEM images of the samples sputtered at 200 W and 250 W, after

In comparison to Figure 8 (b), there are more holes in Figure 10 (a) and the thin films are composed of block structures; this situation is similar to that shown in Figure 10 (b) and

films sputtered at higher powers possess perfect surface morphology.

200 W, 0.25 Pa after annealing at different temperatures for 30 min.

annealing at 1150 oC for 45 min (sputtering pressure: 0.25 Pa).

W, (c) 250 W.

temperature.

Figure 8 (c). As seen in Figure 10 (b), the film is cracked and many gaps and holes exist among each fragmental structure, which attributes to recrystallization of grains due to longer material migration time.

(a) (b) (c) Fig. 9. Typical SEM images of the as-cast thin film and the thin films sputtered at 200 W, 0.25 Pa after annealing at different temperatures. (a) as-cast, (b) 850 oC, (c) 1000 oC.

Fig. 10. SEM images of the thin films sputtered at 200 W and 250 W, respectively, after annealing at 1150 oC for 45 min. (a)200 W, (b)250 W.

Figure 11 shows the SEM images of the samples sputtered at 200 W and different pressures of 0.10 Pa, 0.50 Pa, and 0.70 Pa, after annealing at 1150 oC for 30 min.

As seen from Figure 11, surface morphology differs with the sputtering gaseous pressures. Morphology shown in Figure 11 (a) is similar to that shown in Figure 8 (b), that is, the sample in Figure 11 (a) has perfect crystalline and surface morphology, which indicates that an appropriate low pressure can contribute to the film growth. With the increase of sputtering pressures, more holes and gaps form on the surface accompanied by the decrease

Preparation and Characterization of Dielectric

(3 hrs per schedule).

(b) 850 oC, (c) 1000 oC, (d) 1150 oC

the thickness is about 0.9 um after sputtering for 3 hrs.

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 757

Fig. 13. The SEM micrograph of cross-section of the thin films deposited at 200 W for 3 times

It can be seen that the thin film is dense and crack-free, with the thickness of 2.8 μm. That is,

Figure 14 shows the AFM images of the as-cast thin film and the thin films deposited at 200 W, 0.25 Pa and annealing at different temperatures for 30 min (longitudinal cross-section).

Fig. 14. AFM images of the as-cas thin film and the thin films deposited at 200 W and annealed at different temperatures for 30 min (longitudinal cross-section), (a)as-cast,

(d) (c)

(a) (b)

As shown in Figure 14, the grains on the surface of the samples become greater gradually with the increase of annealing temperature, which indicates that surface morphology and

in grain size, which are shown in Figs.11 (b) and (c). During sputtering, the variation of sputtering gaseous pressure leads to the change of transmission capacity of Ba, Sr, Zn and Nb elements. The higher the sputtering gaseous pressure, the smaller the mean free path. When sputtered at lower pressures, the mean free path and transmission capacity of Ba, Sr, Zn and Nb elements increase, and particles sputtered to the substrate surface multiply rapidly (Wang et al., 2002; Liu et al., 2005). Therefore, the content of elements deposited on the surface increases at the lower pressure, and there are sufficient materials on the substrate surface which can promote the film growth and crystallization of grains. This conclusion can also be proved by Figure 12 as compared with Figure 8(c).

Fig. 11. SEM images of the thin films sputtered at 200 W and different pressures after annealing at 1150 oC for 30 min. (a) 0.10 Pa, (b) 0.50 Pa, (c) 0.70 Pa.

Fig. 12. SEM images of the thin films sputtered at 250 W and different pressures annealing at 150 oC for 30 min, (a) 0.50 Pa, (b) 0.70 Pa.

Figure 13 shows the SEM micrograph of the cross section of the thin films prepared at 200 W, 0.25 Pa, with annealing 3 times (3 hrs per schedule ) at 1150 oC for 30 min.

in grain size, which are shown in Figs.11 (b) and (c). During sputtering, the variation of sputtering gaseous pressure leads to the change of transmission capacity of Ba, Sr, Zn and Nb elements. The higher the sputtering gaseous pressure, the smaller the mean free path. When sputtered at lower pressures, the mean free path and transmission capacity of Ba, Sr, Zn and Nb elements increase, and particles sputtered to the substrate surface multiply rapidly (Wang et al., 2002; Liu et al., 2005). Therefore, the content of elements deposited on the surface increases at the lower pressure, and there are sufficient materials on the substrate surface which can promote the film growth and crystallization of grains. This

(a) (b) (c)

(a) (b) Fig. 12. SEM images of the thin films sputtered at 250 W and different pressures annealing at

Figure 13 shows the SEM micrograph of the cross section of the thin films prepared at 200

W, 0.25 Pa, with annealing 3 times (3 hrs per schedule ) at 1150 oC for 30 min.

Fig. 11. SEM images of the thin films sputtered at 200 W and different pressures after

annealing at 1150 oC for 30 min. (a) 0.10 Pa, (b) 0.50 Pa, (c) 0.70 Pa.

150 oC for 30 min, (a) 0.50 Pa, (b) 0.70 Pa.

conclusion can also be proved by Figure 12 as compared with Figure 8(c).

Fig. 13. The SEM micrograph of cross-section of the thin films deposited at 200 W for 3 times (3 hrs per schedule).

It can be seen that the thin film is dense and crack-free, with the thickness of 2.8 μm. That is, the thickness is about 0.9 um after sputtering for 3 hrs.

Figure 14 shows the AFM images of the as-cast thin film and the thin films deposited at 200 W, 0.25 Pa and annealing at different temperatures for 30 min (longitudinal cross-section).

Fig. 14. AFM images of the as-cas thin film and the thin films deposited at 200 W and annealed at different temperatures for 30 min (longitudinal cross-section), (a)as-cast, (b) 850 oC, (c) 1000 oC, (d) 1150 oC

As shown in Figure 14, the grains on the surface of the samples become greater gradually with the increase of annealing temperature, which indicates that surface morphology and

Preparation and Characterization of Dielectric

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 759

It is evident that different surface morphologies rely on sputtering powers significantly. Figure 15 (a) shows the surface morphology of the thin film at 150 W and the surface is smooth and compact with the smallest roughness. In Figure 15 (b), the film surface is obviously rough with high roughness value, and it can be seen that a few spikes grow along the c-axis that might have been caused by the agglomeration of the particles during crystallization of grains. Figure 15 shows that the films with high-quality crystalline form and the grain sizes of the thin films are at the greatest when the sputtering power is 200 W. The crystal quality is better for the sample deposited at 250 W than that of the sample deposited at 150 W, but worse than that of the sample deposited at 200 w. As seen from Figure 15 (c), the surface roughness is greater than that of the sample in Figure 15 (a), but less than that of the sample in Figure 15( b). As seen from Figure 15 (b) and (c), it can be inferred that the thin films consist of tightly packed particles with an average size of 92.8 nm and 58.5 nm, respectively. The grain size is considerably larger than that obtained by Scherrer`s formula from XRD. Obviously, each particle contains many single-crystal grains. In addition, the overall observation of the thin films indicate a superior microstructure for

the thin films sputtered at 200 W, 0.25 Pa, after annealing at 1150 oC for 30 min.

0.25 Pa and annealing at 1150 oC for different times (longitudinal cross-section).

Fig. 16. AFM images of the as-cast thin film and the thin films deposited at 200 W and annealing at 1150 oC for different times (longitudinal cross-section).(a)15 min, (b) 45 min,

(c)

(c) 60 min.

(a)

Figure 16 shows the AFM images of the as-cast thin film and the films deposited at 200 W,

(b)

crystallization of the grains depend on annealing temperature significantly – the higher the annealing temperature, the better the morphology and crystalline quality. Besides, the thin film surface becomes rougher with the increase in annealing temperature. Figure 14 also tells us the fact that the growth mechanism complies with the nucleus growth mode (Tang et al., 2003). At the beginning of the thin film growth, some atoms or molecules are deposited on the substrate surface and form the so-called nucleation phases. These atoms or molecules form some homogenous and fine atomic groups which can move and are called "islands". These islands can accept new atoms continuously and merge with other islands, and nascent islands could appear at the empty area at the same time, therefore, the amount of islands saturates quickly. The progress of formation and consolidation of islands was ongoing till these isolated islands combine into flakes, with less holes and gaps, which are filled with new atoms. Therefore, the continuous films form. The consolidation of islands ends when the thickness of the thin films is about several tens of nanometers (Tang et al., 2003).

Figure 15 shows the AFM images (horizontal cross-section) of the thin films deposited at different powers (0.25 Pa of sputtering pressure and annealing at 1150 oC for 30 min).

(a) (b)

(c)

Fig. 15. AFM images (horizontal cross-section,10 μm×10 μm) of the thin films deposited at different RF powers: (a)150 W, (b) 200 W, (c) 250 W.

crystallization of the grains depend on annealing temperature significantly – the higher the annealing temperature, the better the morphology and crystalline quality. Besides, the thin film surface becomes rougher with the increase in annealing temperature. Figure 14 also tells us the fact that the growth mechanism complies with the nucleus growth mode (Tang et al., 2003). At the beginning of the thin film growth, some atoms or molecules are deposited on the substrate surface and form the so-called nucleation phases. These atoms or molecules form some homogenous and fine atomic groups which can move and are called "islands". These islands can accept new atoms continuously and merge with other islands, and nascent islands could appear at the empty area at the same time, therefore, the amount of islands saturates quickly. The progress of formation and consolidation of islands was ongoing till these isolated islands combine into flakes, with less holes and gaps, which are filled with new atoms. Therefore, the continuous films form. The consolidation of islands ends when the thickness of

Figure 15 shows the AFM images (horizontal cross-section) of the thin films deposited at different powers (0.25 Pa of sputtering pressure and annealing at 1150 oC for 30 min).

(a) (b)

Fig. 15. AFM images (horizontal cross-section,10 μm×10 μm) of the thin films deposited at

(c)

different RF powers: (a)150 W, (b) 200 W, (c) 250 W.

the thin films is about several tens of nanometers (Tang et al., 2003).

It is evident that different surface morphologies rely on sputtering powers significantly. Figure 15 (a) shows the surface morphology of the thin film at 150 W and the surface is smooth and compact with the smallest roughness. In Figure 15 (b), the film surface is obviously rough with high roughness value, and it can be seen that a few spikes grow along the c-axis that might have been caused by the agglomeration of the particles during crystallization of grains. Figure 15 shows that the films with high-quality crystalline form and the grain sizes of the thin films are at the greatest when the sputtering power is 200 W. The crystal quality is better for the sample deposited at 250 W than that of the sample deposited at 150 W, but worse than that of the sample deposited at 200 w. As seen from Figure 15 (c), the surface roughness is greater than that of the sample in Figure 15 (a), but less than that of the sample in Figure 15( b). As seen from Figure 15 (b) and (c), it can be inferred that the thin films consist of tightly packed particles with an average size of 92.8 nm and 58.5 nm, respectively. The grain size is considerably larger than that obtained by Scherrer`s formula from XRD. Obviously, each particle contains many single-crystal grains. In addition, the overall observation of the thin films indicate a superior microstructure for the thin films sputtered at 200 W, 0.25 Pa, after annealing at 1150 oC for 30 min.

Figure 16 shows the AFM images of the as-cast thin film and the films deposited at 200 W, 0.25 Pa and annealing at 1150 oC for different times (longitudinal cross-section).

Fig. 16. AFM images of the as-cast thin film and the thin films deposited at 200 W and annealing at 1150 oC for different times (longitudinal cross-section).(a)15 min, (b) 45 min, (c) 60 min.

Preparation and Characterization of Dielectric

**4. Conclusion** 

with the nucleus growth mode.

**5.1 Introduction** 

**5.2 Experimental** 

**5. Using Zn - enriched (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 as Target** 

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 761

(a) (b) (c)

Fig. 18. (a) TEM image (bright field image) and (b) SAED spectrum of the thin film

deposited at 200 W, 0.25 Pa and annealing at 1150 oC for 30 min. (c) TEM image (dark field image) of the thin film deposited at 250 W, 0.25 Pa and annealed at 1150 oC for 30 min.

The thin films were successfully fabricated on SiO2(110) substrates through radio frequency (RF) magnetron sputtering system using sintered (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 microwave dielectric ceramics as sputtering target, and next annealing in O2 ambient. The thin films cannot crystallize without annealing and the main phases after annealing at 1150 oC for 30 min are Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 with less Ba5.75Nb2.25O11.375, and the difference between the ceramic target and the thin films arises from the Zn loss through volitilization during sputtering and annealing process. The experimental conditions of sputtering powers, annealing temperatures, annealing times and sputtering pressures have deep influence on the microstructures and morphologies of the thin films. The samples deposited at 200 W, 0.25 Pa, and annealed at 1150 oC for 30 min have the highest crystalline quality. This thin film is polycrystalline with a dense and rod-like structure. The growth mechanism complies

To obtain a dense thin film with fewer oxygen vacancies and less Zn loss, in this article, we adjusted the target component to fabricate ceramic thin films using magnetron sputtering deposition, which differs from those targets reported previously (Cui et al., 2010; Shi et al., 2010), i.e., 1 mol excess ZnO was incorporate in this stoichiometric (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 target so as to compensate the ZnO volatilization during the sputtering and annealing process.

Ceramic thin films were deposited on SiO2(110) substrates by RF magnetron sputtering, using a Zn-enriched target comprised of a homogeneous mixture of 1mol (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3

As compared with Figure 14 (d), Figure 16 reveals that the morphologies of the thin films are closely related with the annealing time and the roughness of the thin films changes greatly with the variation in the annealing times. Annealing after 15 min, surface irregularity appears but root-mean-square surface roughness (RMS, calculated from the AFM data using software) is lower than that of the sample in Figure 14 (d), because the time for material migration is not sufficient. While annealing after 45 min and 60 min, surface irregularities enhance and the roughness is larger than that of the sample in Figure 16 (a), due to availability of more time for material migration, but lower than that of the sample in Figure 14 (d), because of excess volatilization of ZnO and recrystallization of grains. That is, surface irregularities increase from 15 min to 30 min, and then decrease gradually from 30 min to 60 min, which is the same as that shown in Figure 17, the relation between RMS and annealing time.

Fig. 17. Relation between RMS and annealing time.

Figure 17 shows that the roughness of the samples annealing after different times are 0.633 nm (as-cast), 14 nm (15 min), 28.2 nm (30 min), 7.12 nm (45 min), and 5.2 nm (60 min). Therefore, annealing times can affect RMS deeply, as the results show in Figure 5.

Transmission electron microscopy (TEM) is carried out for further investigating the crystal property of the sample deposited at 200 W, 0.25 Pa and annealing at 1150 oC for 30min, as shown in Figure 18. In Figure 18 (a), the grains are relatively straight, and the shape of most grains is rod-like structure, as indicated in Figure 8 (b). Figure 18 (b) shows the selected area electron diffraction (SAED) spectrum of the sample likes a ring, which indicates that the thin films are polycrystalline, and there are many flakes like dancing butterflies in Figure 18 (c), which indicates that the shape of the sample spurtered at 250 W is different from that shown in Figure 18 (b). The grain sizes in Figure 18 (b) are greater than that of the samples sputtered at 200 W.

Fig. 18. (a) TEM image (bright field image) and (b) SAED spectrum of the thin film deposited at 200 W, 0.25 Pa and annealing at 1150 oC for 30 min. (c) TEM image (dark field image) of the thin film deposited at 250 W, 0.25 Pa and annealed at 1150 oC for 30 min.

## **4. Conclusion**

760 Scanning Electron Microscopy

As compared with Figure 14 (d), Figure 16 reveals that the morphologies of the thin films are closely related with the annealing time and the roughness of the thin films changes greatly with the variation in the annealing times. Annealing after 15 min, surface irregularity appears but root-mean-square surface roughness (RMS, calculated from the AFM data using software) is lower than that of the sample in Figure 14 (d), because the time for material migration is not sufficient. While annealing after 45 min and 60 min, surface irregularities enhance and the roughness is larger than that of the sample in Figure 16 (a), due to availability of more time for material migration, but lower than that of the sample in Figure 14 (d), because of excess volatilization of ZnO and recrystallization of grains. That is, surface irregularities increase from 15 min to 30 min, and then decrease gradually from 30 min to 60 min, which is the same as that shown in Figure 17, the relation between RMS and

Figure 17 shows that the roughness of the samples annealing after different times are 0.633 nm (as-cast), 14 nm (15 min), 28.2 nm (30 min), 7.12 nm (45 min), and 5.2 nm (60 min).

Transmission electron microscopy (TEM) is carried out for further investigating the crystal property of the sample deposited at 200 W, 0.25 Pa and annealing at 1150 oC for 30min, as shown in Figure 18. In Figure 18 (a), the grains are relatively straight, and the shape of most grains is rod-like structure, as indicated in Figure 8 (b). Figure 18 (b) shows the selected area electron diffraction (SAED) spectrum of the sample likes a ring, which indicates that the thin films are polycrystalline, and there are many flakes like dancing butterflies in Figure 18 (c), which indicates that the shape of the sample spurtered at 250 W is different from that shown in Figure 18 (b). The grain sizes in Figure 18 (b) are greater than that of the samples

Therefore, annealing times can affect RMS deeply, as the results show in Figure 5.

annealing time.

sputtered at 200 W.

Fig. 17. Relation between RMS and annealing time.

The thin films were successfully fabricated on SiO2(110) substrates through radio frequency (RF) magnetron sputtering system using sintered (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 microwave dielectric ceramics as sputtering target, and next annealing in O2 ambient. The thin films cannot crystallize without annealing and the main phases after annealing at 1150 oC for 30 min are Ba0.5Sr0.5Nb2O6 and Ba0.27Sr0.75Nb2O5.78 with less Ba5.75Nb2.25O11.375, and the difference between the ceramic target and the thin films arises from the Zn loss through volitilization during sputtering and annealing process. The experimental conditions of sputtering powers, annealing temperatures, annealing times and sputtering pressures have deep influence on the microstructures and morphologies of the thin films. The samples deposited at 200 W, 0.25 Pa, and annealed at 1150 oC for 30 min have the highest crystalline quality. This thin film is polycrystalline with a dense and rod-like structure. The growth mechanism complies with the nucleus growth mode.

## **5. Using Zn - enriched (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 as Target**

#### **5.1 Introduction**

To obtain a dense thin film with fewer oxygen vacancies and less Zn loss, in this article, we adjusted the target component to fabricate ceramic thin films using magnetron sputtering deposition, which differs from those targets reported previously (Cui et al., 2010; Shi et al., 2010), i.e., 1 mol excess ZnO was incorporate in this stoichiometric (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 target so as to compensate the ZnO volatilization during the sputtering and annealing process.

### **5.2 Experimental**

Ceramic thin films were deposited on SiO2(110) substrates by RF magnetron sputtering, using a Zn-enriched target comprised of a homogeneous mixture of 1mol (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3

Preparation and Characterization of Dielectric

estimated by the following expression:

wavelength with a value of about 1.54718 Å,

0

Fig. 20. XPS spectrum of the thin film.

100000

200000

300000

400000

500000

Intensity(a.u.)

600000

700000

800000

900000

1000000

the diffraction angle. In the Scherrer`s formula,

Nb3d3/2

Ba4p3/2

Zn3p Sr4s

the grain size in the (013) plane of the thin film is about 52.76 nm.

130 132 134 136 138 140 <sup>2000</sup>

Sr3P3/2

Sr3d

C1s

Sr3P1/2

Nb3s

Nb3p1/2

Sr3d5/2

where

κ

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 763

According to the Scherrer`s formula (Ji et al., 2011), the grain size in the (013) plane can be

0

β

β

cos κλ

β

O1s

0 200 400 600 800 1000 1200

Binding Energy/Ev

Figure 20 shows the XPS spectrum of the thin film in the binding energy range from 0 eV to 1200 eV, and the binding energies at various peaks were calibrated using the C1s (284.6 eV) as a standard sample. The inset images in Figure 20 shows the XPS spectrum of O1s peak. All the XPS spectra of Ba3d, Nb3d, Sr3d, Sr3p, and Zn2p of the thin films consist of two peaks corresponding to their angular momentum of electron. Only one spin-orbit doublet is observed for the individual element, i.e., Ba3d5/2 and Ba3d3/2 peaks at 780.67 and 795.95 eV, Nb3d5/2 and Nb3d3/2 peaks at 206.57 and 209.47 eV, Sr3d5/2 and Sr3d3/2 peaks at 133.07 and 135.02 eV, Sr3p3/2 and Sr3p1/2 peaks at 267.84 and 278.37 eV, Zn2p3/2 and Zn2p1/2 peaks at 1023.27 and 1046.51 eV, indicating that only one chemical state exists in the thin films for each element of Ba, Nb, Sr, Zn, i.e., chemical state of Ba2+, Sr2+, Nb5+, and Zn2+. A doublet structure is observed in the XPS spectrum of O1s peak. Its component peak in the spectrum is fitted to a Gaussian-type distribution with the lower binding energy of 530.45 eV and the higher binding energy of 532.65 eV, as shown in the inset spectrum, corresponding to the

θ<sup>=</sup> ,

λ

0 is the FWHM of the (013) peak, and

θ

530.45

O1s

532.65

530 535

OKLL

Zn2p1/2

Zn2p3/2

BaMNN

Ba3d3/2

0 = 0.003 rad and

20000

Ba3d5/2

50000

80000

is the X-ray

=29.065°; therefore,

θis

( ) 013

is a constant with a value of about 0.89 for the Cu target,

L

Sr3d3/2

Nb3p3/2

and 1 mol ZnO, which were synthesized by a conventional solid-state sintering technique. The monocrystal SiO2(110) substrates were placed on the substrate holder, which can rotate around a central axis to improve the homogeneity of the thin films. The thin films were deposited using an Ar-O2 gas mixture in a JGP450 RF magnetron sputtering systerm with the process parameters: 200 W of sputtering power, 0.25 Pa of sputtering pressure, and 610 °C of the substrate temperature. The Ar gas flow rate was fixed at 16 ml/min and the oxygen flow rate was fixed at 3.2 ml/min, i.e., the O2/Ar flow ratio of 0.2:1 was used as working atmosphere. The distance between the substrates and the targets was 11cm, the sputtering time was 180 min, and the base pressure of the chamber was 1.0×10-3 Pa. As-deposited films were annealed in flowing oxygen (99.999%) at 1150 °C for 30 min in a tube furnace.

#### **5.3 Results and discussion**

Figure 19 is the X-ray diffraction patterns of the thin films, exhibiting the presence of the well-crystallized thin films. No preferential orientation for the thin film is observed.The thin films contain a mixture of Ba0.5Sr0.5Nb2O6 (as compared with the JCPDS card of No.39-0265, International Center for Diffraction Data, 2002), Ba0.67Sr0.33Nb2O6 (No.73-0126), Sr0.744Ba0.247Nb2O6 (No.70-3747), and SrNb2O6 (No.72-2088), which can be written as BaxSr1 xNb2O6. And ZnO is also observed, as compared with the characteristic peak of ZnO. As we all know, the various sputtering yields of Ba, Sr, Nb, Zn, and O elements are a knotty problem in the deposition of (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 – based thin films. The discrepancy between thin film and target probably results from the volatilization of ZnO during the process of sputtering and annealing, because Zn has a significant larger vapor pressure than the other metal elements (Ba, Sr, Nb) (Tang et al., 2009). Therefore, ZnO may either not stick to or be re-evaporated from the growing surface (Tang et al., 2009), worsening the composition deviation with the target.

Fig. 19. XRD spectrum of the thin film.

According to the Scherrer`s formula (Ji et al., 2011), the grain size in the (013) plane can be estimated by the following expression:

$$\mathcal{L}\_{\text{(013)}} = \frac{\kappa \mathcal{A}}{\beta\_0 \cos \theta} \,' \,'$$

where κ is a constant with a value of about 0.89 for the Cu target, λ is the X-ray wavelength with a value of about 1.54718 Å, β0 is the FWHM of the (013) peak, and θ is the diffraction angle. In the Scherrer`s formula, β0 = 0.003 rad and θ =29.065°; therefore, the grain size in the (013) plane of the thin film is about 52.76 nm.

Fig. 20. XPS spectrum of the thin film.

762 Scanning Electron Microscopy

and 1 mol ZnO, which were synthesized by a conventional solid-state sintering technique. The monocrystal SiO2(110) substrates were placed on the substrate holder, which can rotate around a central axis to improve the homogeneity of the thin films. The thin films were deposited using an Ar-O2 gas mixture in a JGP450 RF magnetron sputtering systerm with the process parameters: 200 W of sputtering power, 0.25 Pa of sputtering pressure, and 610 °C of the substrate temperature. The Ar gas flow rate was fixed at 16 ml/min and the oxygen flow rate was fixed at 3.2 ml/min, i.e., the O2/Ar flow ratio of 0.2:1 was used as working atmosphere. The distance between the substrates and the targets was 11cm, the sputtering time was 180 min, and the base pressure of the chamber was 1.0×10-3 Pa. As-deposited films were annealed

Figure 19 is the X-ray diffraction patterns of the thin films, exhibiting the presence of the well-crystallized thin films. No preferential orientation for the thin film is observed.The thin films contain a mixture of Ba0.5Sr0.5Nb2O6 (as compared with the JCPDS card of No.39-0265, International Center for Diffraction Data, 2002), Ba0.67Sr0.33Nb2O6 (No.73-0126), Sr0.744Ba0.247Nb2O6 (No.70-3747), and SrNb2O6 (No.72-2088), which can be written as BaxSr1 xNb2O6. And ZnO is also observed, as compared with the characteristic peak of ZnO. As we all know, the various sputtering yields of Ba, Sr, Nb, Zn, and O elements are a knotty problem in the deposition of (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 – based thin films. The discrepancy between thin film and target probably results from the volatilization of ZnO during the process of sputtering and annealing, because Zn has a significant larger vapor pressure than the other metal elements (Ba, Sr, Nb) (Tang et al., 2009). Therefore, ZnO may either not stick to or be re-evaporated from the growing surface (Tang et al., 2009), worsening the

in flowing oxygen (99.999%) at 1150 °C for 30 min in a tube furnace.

**5.3 Results and discussion** 

composition deviation with the target.

Fig. 19. XRD spectrum of the thin film.

Figure 20 shows the XPS spectrum of the thin film in the binding energy range from 0 eV to 1200 eV, and the binding energies at various peaks were calibrated using the C1s (284.6 eV) as a standard sample. The inset images in Figure 20 shows the XPS spectrum of O1s peak. All the XPS spectra of Ba3d, Nb3d, Sr3d, Sr3p, and Zn2p of the thin films consist of two peaks corresponding to their angular momentum of electron. Only one spin-orbit doublet is observed for the individual element, i.e., Ba3d5/2 and Ba3d3/2 peaks at 780.67 and 795.95 eV, Nb3d5/2 and Nb3d3/2 peaks at 206.57 and 209.47 eV, Sr3d5/2 and Sr3d3/2 peaks at 133.07 and 135.02 eV, Sr3p3/2 and Sr3p1/2 peaks at 267.84 and 278.37 eV, Zn2p3/2 and Zn2p1/2 peaks at 1023.27 and 1046.51 eV, indicating that only one chemical state exists in the thin films for each element of Ba, Nb, Sr, Zn, i.e., chemical state of Ba2+, Sr2+, Nb5+, and Zn2+. A doublet structure is observed in the XPS spectrum of O1s peak. Its component peak in the spectrum is fitted to a Gaussian-type distribution with the lower binding energy of 530.45 eV and the higher binding energy of 532.65 eV, as shown in the inset spectrum, corresponding to the

Preparation and Characterization of Dielectric

results shown in Figure 21.

crystalline.

section.

have not special periodicity (Ji et al., 2011).

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 765

Figure 22. shows the thickness of the thin film is about 1.38 μm with dense structure and homogeneous grains and well-defined grain boundaries, which are consistent with the

Figure 23. shows the AFM images of the sample, with a scan area of 10 × 10 μm. As seen in Figure 23(a), the sample is regular in surface with fine and uniform grains, and a few spherical particles are observed growing along the c axis. The root-mean-square surface roughness (RMS, calculated from the AFM data using the PSI ProScan Image Processing software package version 1.0, Park Scientific Instruments, Sunnyvale, CA) of the thin films is about 27.4 nm. Figure 23(b) shows the formation of the ceramic thin film with high-quality

(a) (b)

The TEM image and the selected area electron diffraction (SAED) spectrum are shown in Figure 24. The spherical particles with the well-defined boundaries are observed in Figure 24(a). The spherical shape of the grains is consistent with the results acquired by the SEM and AFM. The SAED spectrum indicats that the thin films are polycrystalline, as shown in Figure 24(b). About the (013) grain size, there is considerable discrepancy between the measured grain size (approximate 1600 nm by TEM) and calculation size (53 nm obtains from the Scherrer`s formula), which may be because the grain size obtained through the Scherrer`s formula is the average size of the orderly arrangement grains, however, the grain size measured by the TEM is the size of the clusters consisting of many small grains, which

Fig. 23. AFM images of the thin film. (a) longitudinal cross-section; (b) horizontal cross-

lattice oxygen and the adsorbed oxygen, respectively. In general, the peak of adsorptional oxygen is much weaker than that of lattice oxygen, which would be favorable for the dielectric property of the thin films, because the oxygen vacancies result in the dielectric loss. The quantitative analysis using Ba3d, Sr3d, Zn2p, Nb3d, and O1s peaks reveals that the Ba:Sr:Zn:Nb:O atomic ratio is 3.01: 5.33: 0.96: 27.90: 62.80.

Figure 21. shows the SEM image of the thin film, which shows that fine spherical particles are distributed on the sample's surface. The surface morphology of the thin film is of dense structure with less small holes. It is evident that the grains are of uniform grain size, and are distributed homogenously on the surface of the thin film.

Fig. 21. SEM image of the thin film.

Fig. 22. Cross-sectional SEM images of the thin film.

lattice oxygen and the adsorbed oxygen, respectively. In general, the peak of adsorptional oxygen is much weaker than that of lattice oxygen, which would be favorable for the dielectric property of the thin films, because the oxygen vacancies result in the dielectric loss. The quantitative analysis using Ba3d, Sr3d, Zn2p, Nb3d, and O1s peaks reveals that the

Figure 21. shows the SEM image of the thin film, which shows that fine spherical particles are distributed on the sample's surface. The surface morphology of the thin film is of dense structure with less small holes. It is evident that the grains are of uniform grain size, and are

Ba:Sr:Zn:Nb:O atomic ratio is 3.01: 5.33: 0.96: 27.90: 62.80.

distributed homogenously on the surface of the thin film.

Fig. 21. SEM image of the thin film.

Fig. 22. Cross-sectional SEM images of the thin film.

Figure 22. shows the thickness of the thin film is about 1.38 μm with dense structure and homogeneous grains and well-defined grain boundaries, which are consistent with the results shown in Figure 21.

Figure 23. shows the AFM images of the sample, with a scan area of 10 × 10 μm. As seen in Figure 23(a), the sample is regular in surface with fine and uniform grains, and a few spherical particles are observed growing along the c axis. The root-mean-square surface roughness (RMS, calculated from the AFM data using the PSI ProScan Image Processing software package version 1.0, Park Scientific Instruments, Sunnyvale, CA) of the thin films is about 27.4 nm. Figure 23(b) shows the formation of the ceramic thin film with high-quality crystalline.

Fig. 23. AFM images of the thin film. (a) longitudinal cross-section; (b) horizontal crosssection.

The TEM image and the selected area electron diffraction (SAED) spectrum are shown in Figure 24. The spherical particles with the well-defined boundaries are observed in Figure 24(a). The spherical shape of the grains is consistent with the results acquired by the SEM and AFM. The SAED spectrum indicats that the thin films are polycrystalline, as shown in Figure 24(b). About the (013) grain size, there is considerable discrepancy between the measured grain size (approximate 1600 nm by TEM) and calculation size (53 nm obtains from the Scherrer`s formula), which may be because the grain size obtained through the Scherrer`s formula is the average size of the orderly arrangement grains, however, the grain size measured by the TEM is the size of the clusters consisting of many small grains, which have not special periodicity (Ji et al., 2011).

Preparation and Characterization of Dielectric

0925-8388

1359-6454

2626-2629, ISBN 0169-4332

Metallurgy Industry Press, Beijing, 2003.

Thin Films by RF Magnetron-Sputtering with (Ba0.3Sr0.7)(Zn1/3Nb2/3)O3 Ceramic Target 767

HSi, C.S.; Chen, C.Y.; Wu, N.C.; & Wang, M.C. (2003). Dielectric Properties of Ba(ZrxTi1–x)O3

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*Ceramics Society*, Vol. 26, No. 15, (June 2006), pp. 3211-3219, ISBN 0955-2219 Ianculescu, A.; Despax, B.; Bley, V.; & Lebey, T. (2007). Structure-Properties Correlations for

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Fig. 24. TEM image and the SAED spectrum of the thin film. (a) TEM; (b) SAED.

## **5.4 Conclusion**

The main phase of the ceramic thin film is BaxSr1-xNb2O6. The thin film presences a high crystalline quality, with few adsorbed oxygen. The surface morphology indicates that the thin film is of dense structure. The grains are polycrystalline and uniform in size with the spherical shape and well-defined grain boundaries.

## **6. Acknowledgements**

The authors would like to thank the National Natural Science Foundation of China (No.51042001). The authors are also grateful to Professor Chengshan Xue for his help in discussion our results.

## **7. References**


Fig. 24. TEM image and the SAED spectrum of the thin film. (a) TEM; (b) SAED.

spherical shape and well-defined grain boundaries.

The main phase of the ceramic thin film is BaxSr1-xNb2O6. The thin film presences a high crystalline quality, with few adsorbed oxygen. The surface morphology indicates that the thin film is of dense structure. The grains are polycrystalline and uniform in size with the

The authors would like to thank the National Natural Science Foundation of China (No.51042001). The authors are also grateful to Professor Chengshan Xue for his help in

Amanullah, F.M.; Pratap, K.I.; & Hari, V.B. (1998). Compositional Analysis and Depth

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Structure Microwave Dielectric Ceramic Thin Films Fabricated by the RF

**5.4 Conclusion** 

**6. Acknowledgements** 

98. ISBN 0921-5107

discussion our results.

**7. References** 

Magnetron-Sputtering Method, *Journal of Materials Science: Materials in Electronics*, Vol. 21, No. 4, (April 2010), pp. 349-354, ISBN 0957-4522


**Part 6** 

**Geoscience, Mineralogy** 

Annealing, *Applied Physics Letter*, Vol. 73, No.11, (September 1998). pp.1514-1517. ISBN 1882-0778

