**2. Antimicrobial activity**

Chitin and chitosan have interesting physicochemical, biological, and mechanical properties. One such property of chitosan is related to its antimicrobial activity. There are several studies demonstrating the antimicrobial and antifungal properties of chitosan and many derivatives [6–11]. Recently, the effect of the physical form of chitosan on its antibacterial activity against pathogenic bacteria was studied. Researchers examined chitosan coating as an inhibitor of *Listeria monocytogenes* on vacuum-packed pork fillets and fresh cheese. The antibacterial effect is reported to be generally rapid, eliminating bacteria within a few hours. As for the physical properties of chitosan, these are mainly governed by two factors: deacetylation

**265**

cidal for three pathogenic microorganisms at pH 5.0 [6].

Several hypotheses have been proposed about the antimicrobial function of chitosan. Ionic interactions occcuring between the positive charges of amino groups and negative bacterial surface molecules under acid conditions change the membrane permeability which leads to cellular lysis. Interaction with necessary nutrients

*Chitosan for Using Food Protection*

are all influencing factors [12].

*DOI: http://dx.doi.org/10.5772/intechopen.99247*

degree (DD) and molecular weight (MW). Natural origin, as well as variability in chemical composition, can affect the properties of chitosan and have an impact on its industrial uses. Some studies have revealed that DD correlates with the antimicrobial activity of chitosan. The effect of chitosan as an antimicrobial in the agriculture and food industry has been studied. According to these studies, the antimicrobial activity of chitosan depends on several external and internal factors as well as a number of environmental factors. The type of microorganism, physiological state, pH, temperature, ionic strength, metal ions, ethylenediaminetetraacetic acid (EDTA), the presence of organic matter, MW, DD, solvent, and concentration

Chitosan is a commercial biopolymer produced predominantly from crab and shrimp residues. The physicochemical properties of chitosan affect the functional properties that differ according to crustacean type and preparation methods. Chitosan has been studied to compare the functionality of commercial products obtained from crustacean and insect chitosan as antimicrobials. The results indicated differences between commercial insect chitosan and crustacean chitosan with regard to their antimicrobial capacity. Generally speaking, crustacean chitosan with a pH of 5,0 during a 49-hour incubation period displayed a greater antimicrobial capacity than insect chitosan at the same pH. This behavior was seen mostly in Salmonella cases where crustacean chitosan resulted in more than 4 logarithmic decreases, whereas insect chitosan was only bacteriostatic resulting in about a 1 logarithmic decrease. The similar behavior was noticed for *Escherichia coli*, despite the smaller differences in antimicrobial influence in Salmonella cases. As noted, some studies have pointed out potential differences between the functions and physical properties of chitosan in different species of crustaceans. This may be even more pronounced among chitosan obtained from various sources such as crustaceans and insects [6]. Antimicrobial activity can be adversely affected by pH, and as such pH plays an important role in the antimicrobial capacity of chitosan. Low pH chitosan appears to have more antimicrobial activity than high pH chitosan [13]. A study was conducted to determine the effect of two different concentrations of chitosan at pH 6,5 and 5,5 on different pathogenic microorganisms, including *Salmonella* Typhimurium, *E. coli*, and *L. monocytogenes*. The author concluded that chitosan with a pH of 6.5 had a rather weak effect on pathogenic microorganisms and could not inhibit *L. monocytogenes*. At pH 5,5; there was inhibition of the microorganisms tested for 24 to 72 hours of storage at 30°C. The researcher concluded that chitosan acts better at pH 5.5 than at pH 6.5 [14]. Another researcher examined the antibacterial activity of chitosan of different MW at various pH levels (pH 4, 4.5, and 5) on *L. monocytogenes* strains. The results also indicated that, with the exception of two *L. monocytogenes* strains, chitosan with a pH of 5 had the greatest bacterial reduction effect during a 24-hour incubation period [15]. In another study, two pH levels were tested at a concentration of 0.15% (w/v) chitosan. Later an 8-hour incubation, the antibacterial effect was found to be higher at pH 5,0 than pH 6,2 for *S.* Typhimurium, but the opposite for *E. coli* and *Listeria monocytogenes*, where the antimicrobial effect of chitosan at pH 6.2 was stronger than it was at pH 5.0. The effect of chitosan at both pH levels seemed to be dependent on the microorganism. Differences were observed in chitosan at both pH levels of acetic acid compared to control. Chitosan exhibited a pronounced antimicrobial activity at both pH values, particularly on *L. monocytogenes*. Chitosan obtained from both sources, crustaceans and insects, was bacteriostatic or bacteri-

### *Chitosan for Using Food Protection DOI: http://dx.doi.org/10.5772/intechopen.99247*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

prevents the uptake of toxic metals in plants [4].

from which chitosan can be produced [5].

**2. Antimicrobial activity**

1960s, studies were conducted on the ability of chitosan to bind red blood cells. In the same year, chitosan was also considered as a hemostatic agent. In the next 30 years, chitosan was used in treatment plants to provide asepsis water. In the last 20 years, research on chitosan has intensified due to its many important properties [1]. Today, chitosan has many industrial applications and after cellulose, it is the most common polysaccharide chitin in the world. As one of the most important derivatives of chitin, chitosan is a polycationic biopolymer obtained by partial or complete deacetylation (removal of an acetyl functional group from an organic compound) of chitin in an alkaline environment [2]. The only difference between cellulose and chitosan biopolymer is the presence of the acetyl (-NH2) functional group instead of the hydroxyl (-OH) functional group in the cellulose structure. This difference ensures that the chain structure of the chitosan biopolymer is polycationic. Many superior properties of chitosan arise from this polycationic structure. In addition to this advantage, the presence of both –OH and –NH2 groups in the chain structure of chitosan and the fact that these groups can be modified in different ways is a situation that highlights its uses [3]. Chitosan, which can be obtained in large quantities from many natural sources containing chitin, such as the exoskeleton of mushrooms, crayfish, shrimp, and crabs, is more advantageous than other biopolymers including chitin in terms of non-toxicity to organisms, easy biodegradability, and biocompatibility. For these reasons, chitosan is a natural, safe, cheap, raw material biopolymer used in many industrial areas such as food, medicine, pharmaceuticals, cosmetics, agriculture, wastewater treatment, and textiles. Besides having antiviral, antibacterial, and antifungal properties, chitosan is also an effective agent in controlling and reducing the spread of diseases by promoting the defense system of plants. In addition, chitosan is being used for improvement in agriculture because it chelates metal ions in the environment (water, soil, etc.) and

Chitosan is a natural and biodegradable biopolymer used in different industrial applications as an agent for flocculation and chelating, permeability control, and as an antimicrobial, among other processes. Predominantly produced today by the deacetylation of chitin on an industrial scale, chitosan is found in the exoskeleton of crustaceans and insects, and the cell walls of many fungi and some algae. Although the main source of chitin is crab, shrimp, crayfish, and shrimp residues, the importance of insect chitosan depends on the role insects play as a sustainable protein source. Insects are seen as an alternative to traditionally consumed proteins derived predominantly from traditional livestock (mainly cows, chickens, and pigs) and fish. In addition, using the insect as a protein source produces two by-products of interest to the industry, lipids that can be used as biofuels (30–40% total dry weight) as well as a residual material made of chitin with some bioactive properties

Chitin and chitosan have interesting physicochemical, biological, and mechanical properties. One such property of chitosan is related to its antimicrobial activity. There are several studies demonstrating the antimicrobial and antifungal properties of chitosan and many derivatives [6–11]. Recently, the effect of the physical form of chitosan on its antibacterial activity against pathogenic bacteria was studied. Researchers examined chitosan coating as an inhibitor of *Listeria monocytogenes* on vacuum-packed pork fillets and fresh cheese. The antibacterial effect is reported to be generally rapid, eliminating bacteria within a few hours. As for the physical properties of chitosan, these are mainly governed by two factors: deacetylation

**264**

degree (DD) and molecular weight (MW). Natural origin, as well as variability in chemical composition, can affect the properties of chitosan and have an impact on its industrial uses. Some studies have revealed that DD correlates with the antimicrobial activity of chitosan. The effect of chitosan as an antimicrobial in the agriculture and food industry has been studied. According to these studies, the antimicrobial activity of chitosan depends on several external and internal factors as well as a number of environmental factors. The type of microorganism, physiological state, pH, temperature, ionic strength, metal ions, ethylenediaminetetraacetic acid (EDTA), the presence of organic matter, MW, DD, solvent, and concentration are all influencing factors [12].

Chitosan is a commercial biopolymer produced predominantly from crab and shrimp residues. The physicochemical properties of chitosan affect the functional properties that differ according to crustacean type and preparation methods. Chitosan has been studied to compare the functionality of commercial products obtained from crustacean and insect chitosan as antimicrobials. The results indicated differences between commercial insect chitosan and crustacean chitosan with regard to their antimicrobial capacity. Generally speaking, crustacean chitosan with a pH of 5,0 during a 49-hour incubation period displayed a greater antimicrobial capacity than insect chitosan at the same pH. This behavior was seen mostly in Salmonella cases where crustacean chitosan resulted in more than 4 logarithmic decreases, whereas insect chitosan was only bacteriostatic resulting in about a 1 logarithmic decrease. The similar behavior was noticed for *Escherichia coli*, despite the smaller differences in antimicrobial influence in Salmonella cases. As noted, some studies have pointed out potential differences between the functions and physical properties of chitosan in different species of crustaceans. This may be even more pronounced among chitosan obtained from various sources such as crustaceans and insects [6].

Antimicrobial activity can be adversely affected by pH, and as such pH plays an important role in the antimicrobial capacity of chitosan. Low pH chitosan appears to have more antimicrobial activity than high pH chitosan [13]. A study was conducted to determine the effect of two different concentrations of chitosan at pH 6,5 and 5,5 on different pathogenic microorganisms, including *Salmonella* Typhimurium, *E. coli*, and *L. monocytogenes*. The author concluded that chitosan with a pH of 6.5 had a rather weak effect on pathogenic microorganisms and could not inhibit *L. monocytogenes*. At pH 5,5; there was inhibition of the microorganisms tested for 24 to 72 hours of storage at 30°C. The researcher concluded that chitosan acts better at pH 5.5 than at pH 6.5 [14]. Another researcher examined the antibacterial activity of chitosan of different MW at various pH levels (pH 4, 4.5, and 5) on *L. monocytogenes* strains. The results also indicated that, with the exception of two *L. monocytogenes* strains, chitosan with a pH of 5 had the greatest bacterial reduction effect during a 24-hour incubation period [15]. In another study, two pH levels were tested at a concentration of 0.15% (w/v) chitosan. Later an 8-hour incubation, the antibacterial effect was found to be higher at pH 5,0 than pH 6,2 for *S.* Typhimurium, but the opposite for *E. coli* and *Listeria monocytogenes*, where the antimicrobial effect of chitosan at pH 6.2 was stronger than it was at pH 5.0. The effect of chitosan at both pH levels seemed to be dependent on the microorganism. Differences were observed in chitosan at both pH levels of acetic acid compared to control. Chitosan exhibited a pronounced antimicrobial activity at both pH values, particularly on *L. monocytogenes*. Chitosan obtained from both sources, crustaceans and insects, was bacteriostatic or bactericidal for three pathogenic microorganisms at pH 5.0 [6].

Several hypotheses have been proposed about the antimicrobial function of chitosan. Ionic interactions occcuring between the positive charges of amino groups and negative bacterial surface molecules under acid conditions change the membrane permeability which leads to cellular lysis. Interaction with necessary nutrients for bacteria could be another mechanism. Chitosan's bactericidal effect may also be affected by the inoculum size to the bacteria growth [6]. In some studies, all compounds tested after 4 hours of incubation for an inoculum size of 103 cells/mL were bactericidal at any concentration of chitosan tested. In contrast, at a higher initial inoculum concentration, 0.1% (w/v) chitosan was only bacteriostatic. Regardless of the inoculum level, any chito-oligosaccharide mixture of 0.25% (w/v) was sufficient to reduce the starting population of *E. coli* by at least 3 log cycles. However, the results regarding the effect of inoculum size are not conclusive because they vary with pH and type of microorganism. Therefore, it is not possible to predict higher antimicrobial activity at a given inoculum size in all cases [16, 17].

Included in the peptidoglycan layer on the cell surface, teichoic acid is vital for the growth of Gram-positive bacteria as well as for cell division. Chitosan and its derivatives can bind to teichoic acid on the surface of Gram-positive bacteria noncovalently. Chitosan's effect on the cell membrane has not been clearly discovered yet; however, it is well-known that it affects the cell membrane because it has a greater hydrodynamic diameter than peptidoglycans' pore size. Strangely, chitosan with a MW of 5 kDa suppresses DNA synthesis and promotes *Bacillus megaterium* filamentation, which suggests that the chitosan's MW plays a role in its potential to affect cell membrane permeability [18]. In addition, the effect of chitosan on teichoic acid has been demonstrated by testing *Staphylococcus aureus* mutant strains lacking the genes needed for teichoic acid biosynthesis [19]. Mutant strains were found to be more resistant to the environmental conditions than the wild-type strain, which indicates that anionic teichoic acid improves chitosan's antibacterial properties against Gram-positive bacteria. Teichoic acid has, interestingly, many functions. It controls activities of enzymes, helps to cope with environmental stress, and manages the cationic concentration in the cell cover by binding to the cell surface and the cell receptor. The mechanism of antimicrobial effect of chitosan on Gram-positive bacteria is due to electrostatic effect with teichoic acid, resulting in disruption and death of cell [18]. Two different mechanisms mediate the interactions between chitosan and the outer membrane of Gram-negative bacteria. The first mechanism involves chelating chitosan with various cations when pH is higher than pKa, resulting in a breakdown in the uptake of essential nutrients and a breakdown in cell wall integrity. The second mechanism involves electrostatic interactions between chitosan and anions associated with lipopolysaccharides in the outer membrane. Chitosan also creates disruptions in the inner membrane, causing intracellular content to leak. In addition, chitosan can pass through the cell membrane of Gram-negative bacteria where it likely interferes with DNA/ RNA synthesis and triggers an intracellular response in cells. Thus, the electrostatic interactions between chitosan and the anionic surface are crucial to chitosan's antimicrobial properties against Gram-negative bacteria. Moreover, chitosan can bind non-covalently to the cell membrane of Gram-negative bacteria, suggesting it plays an important role in antimicrobial activity [20, 21]. The difference between Gram-positive and Gram-negative bacteria is more obvious compared to chitosanresistant fungi and chitosan-sensitive fungi. Chitosan is belived to interact with a phospholipid component of chitosan-sensitive fungi electrostatically, thereby breaking it down and participating the cell, fnally leading to the prevention of DNA/RNA as well as protein synthesis. However, chitosan is unable to make the cell wall of chitosan-resistant fungi permeable due to its variable fluidity, so it remains on the cell surface and forms a polymer to function as a barrier against oxygen and necessary nutrients, ultimately resulting in cell death. The lowered antimicrobial activity of chitosan was also seen in a *Neurospora crassa* mutant strain, explaining the lower levels of unsaturated fatty acids relative to the wild-type strain. Thus, the antibacterial effect of chitosan on fungi is greatly affected by the fluency of the

**267**

*Chitosan for Using Food Protection*

*DOI: http://dx.doi.org/10.5772/intechopen.99247*

prophage induction in lysogenic culture [22].

*lambica* at pH 4 than pH 6 [18].

cell membrane and the type of mushrooms [18]. Chitosan inhibited the growth of *Aspergillus flavus* and aflatoxin in liquid culture, pre-harvest corn and peanuts, and increased the production of phytoalexin in germinating peanuts. Chitosan has become the first compound in the list of basic substances approved by the European Union for plant protection in agricultural practices, both for organic agriculture and for integrated pest control (Tes. EU 66 2014/563). Thus, chitosan can be used as a biodegradable fungicide. In addition, chitosan shows antiviral activity against plant viruses. It has been demonstrated that chitosan inhibits productive infection caused by bacteriophages. The efficacy of bacteriophage inhibition is directly dependent on the final concentration in the medium. The main factors by which chitosan suppresses phage infections are phage particle inactivation and inhibition of bacteriophage growth at the cellular level. Chitosan can be used for induction of phagoresistance in industrial microorganism cultures to prevent unwanted phagolysis caused by inoculum contamination with virulent bacteriophages or spontaneous

As stated previously, pH can play a key role in chitosan's antimicrobial activity, and the pKa of chitosan sequences from 6,3 to 6,5 [23]. Chitosan only dissolves in acidic aqueous environment where it becomes polycationic when the pH value is lower than the pKa amount. Polycationic chitosan molecules react with negatively charged cell wall molecules, including proteins, phospholipids, polysaccharides, and fatty acids because of the high intensity of amino groups found on the polymer surface, ultimately causing intracellular materials to leak. Chitosan exhibits higher antimicrobial activity at low pH values (< 6) because its amino group is ionized at low pH rates. Moreover, the positive charge of chitosan improves at low pH values, increasing the absorption of chitosan at the bacterial cell wall. Moreover, at upper pH values (> 6) the amino group of chitosan becomes aprotic, which may lead to precipitation from solution [24]. One study informed that chitosan's antimicrobial activities against *Klebsiella pneumoniae* partly resulted from the polycationic nature of chitosan, thus being associated with the protonation of the amino group. The protonation of the amino group is related to the degree of polymerization as well as the pH of the environment. For example, chitosan is more effective against *Candida* 

In the early 1960s, chitosan's ability to bind to red blood cells was investigated. At that time, it was also seen as a hemostatic agent. Chitosan has been used in water purification for the last 30 years. Since then, numerous studies have been conducted to find ways to use these materials. Today, chitosan is known as a dietary supplement for weight loss. In fact, it has been marketed for this purpose in Japan as well as Europe for about 20 years. Many people even call it "anti-fat" [25]. Chitosan has attracted great attention because of its increasing demand as a highly beneficial biopolymer in recent times. Chitosan, which is obtained by deacetylation of chitin with sodium hydroxide (NaOH), can be extracted from a variety of fungi, insects, and crustaceans. Basically, chitosan is a polymer consisting of randomly distributed units of N-acetyl-D-glucosamine and D-glucosamine with different deacetylation degree, acetylation type, and molecular weight which could be chemically modified to its derivatives. These derivatives affect antibacterial influence of chitosan and its solubility in acidic solutions. Chitosan's three reactive functional groups are: the amino group at the C-6 position, the primary hydroxyl group at the C-6 position, and the secondary hydroxyl group at the C-3 position. The amino group at the C-6 position differs from chitosan obtained from chitin due to its chemical, physical, and biological functions [18]. Chitosan is a very useful and attractive biopolymer due to its diverse chemical structure. Structural diversity can be seen in MW ranging from low (100 kDa) to high (300 kDa) as well as DD ranging from chitin (< 60%) to chitosan (> 60%). The wide range of chitosan samples described

### *Chitosan for Using Food Protection DOI: http://dx.doi.org/10.5772/intechopen.99247*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

pounds tested after 4 hours of incubation for an inoculum size of 103

antimicrobial activity at a given inoculum size in all cases [16, 17].

for bacteria could be another mechanism. Chitosan's bactericidal effect may also be affected by the inoculum size to the bacteria growth [6]. In some studies, all com-

bactericidal at any concentration of chitosan tested. In contrast, at a higher initial inoculum concentration, 0.1% (w/v) chitosan was only bacteriostatic. Regardless of the inoculum level, any chito-oligosaccharide mixture of 0.25% (w/v) was sufficient to reduce the starting population of *E. coli* by at least 3 log cycles. However, the results regarding the effect of inoculum size are not conclusive because they vary with pH and type of microorganism. Therefore, it is not possible to predict higher

Included in the peptidoglycan layer on the cell surface, teichoic acid is vital for the growth of Gram-positive bacteria as well as for cell division. Chitosan and its derivatives can bind to teichoic acid on the surface of Gram-positive bacteria noncovalently. Chitosan's effect on the cell membrane has not been clearly discovered yet; however, it is well-known that it affects the cell membrane because it has a greater hydrodynamic diameter than peptidoglycans' pore size. Strangely, chitosan with a MW of 5 kDa suppresses DNA synthesis and promotes *Bacillus megaterium* filamentation, which suggests that the chitosan's MW plays a role in its potential to affect cell membrane permeability [18]. In addition, the effect of chitosan on teichoic acid has been demonstrated by testing *Staphylococcus aureus* mutant strains lacking the genes needed for teichoic acid biosynthesis [19]. Mutant strains were found to be more resistant to the environmental conditions than the wild-type strain, which indicates that anionic teichoic acid improves chitosan's antibacterial properties against Gram-positive bacteria. Teichoic acid has, interestingly, many functions. It controls activities of enzymes, helps to cope with environmental stress, and manages the cationic concentration in the cell cover by binding to the cell surface and the cell receptor. The mechanism of antimicrobial effect of chitosan on Gram-positive bacteria is due to electrostatic effect with teichoic acid, resulting in disruption and death of cell [18]. Two different mechanisms mediate the interactions between chitosan and the outer membrane of Gram-negative bacteria. The first mechanism involves chelating chitosan with various cations when pH is higher than pKa, resulting in a breakdown in the uptake of essential nutrients and a breakdown in cell wall integrity. The second mechanism involves electrostatic interactions between chitosan and anions associated with lipopolysaccharides in the outer membrane. Chitosan also creates disruptions in the inner membrane, causing intracellular content to leak. In addition, chitosan can pass through the cell membrane of Gram-negative bacteria where it likely interferes with DNA/ RNA synthesis and triggers an intracellular response in cells. Thus, the electrostatic interactions between chitosan and the anionic surface are crucial to chitosan's antimicrobial properties against Gram-negative bacteria. Moreover, chitosan can bind non-covalently to the cell membrane of Gram-negative bacteria, suggesting it plays an important role in antimicrobial activity [20, 21]. The difference between Gram-positive and Gram-negative bacteria is more obvious compared to chitosanresistant fungi and chitosan-sensitive fungi. Chitosan is belived to interact with a phospholipid component of chitosan-sensitive fungi electrostatically, thereby breaking it down and participating the cell, fnally leading to the prevention of DNA/RNA as well as protein synthesis. However, chitosan is unable to make the cell wall of chitosan-resistant fungi permeable due to its variable fluidity, so it remains on the cell surface and forms a polymer to function as a barrier against oxygen and necessary nutrients, ultimately resulting in cell death. The lowered antimicrobial activity of chitosan was also seen in a *Neurospora crassa* mutant strain, explaining the lower levels of unsaturated fatty acids relative to the wild-type strain. Thus, the antibacterial effect of chitosan on fungi is greatly affected by the fluency of the

cells/mL were

**266**

cell membrane and the type of mushrooms [18]. Chitosan inhibited the growth of *Aspergillus flavus* and aflatoxin in liquid culture, pre-harvest corn and peanuts, and increased the production of phytoalexin in germinating peanuts. Chitosan has become the first compound in the list of basic substances approved by the European Union for plant protection in agricultural practices, both for organic agriculture and for integrated pest control (Tes. EU 66 2014/563). Thus, chitosan can be used as a biodegradable fungicide. In addition, chitosan shows antiviral activity against plant viruses. It has been demonstrated that chitosan inhibits productive infection caused by bacteriophages. The efficacy of bacteriophage inhibition is directly dependent on the final concentration in the medium. The main factors by which chitosan suppresses phage infections are phage particle inactivation and inhibition of bacteriophage growth at the cellular level. Chitosan can be used for induction of phagoresistance in industrial microorganism cultures to prevent unwanted phagolysis caused by inoculum contamination with virulent bacteriophages or spontaneous prophage induction in lysogenic culture [22].

As stated previously, pH can play a key role in chitosan's antimicrobial activity, and the pKa of chitosan sequences from 6,3 to 6,5 [23]. Chitosan only dissolves in acidic aqueous environment where it becomes polycationic when the pH value is lower than the pKa amount. Polycationic chitosan molecules react with negatively charged cell wall molecules, including proteins, phospholipids, polysaccharides, and fatty acids because of the high intensity of amino groups found on the polymer surface, ultimately causing intracellular materials to leak. Chitosan exhibits higher antimicrobial activity at low pH values (< 6) because its amino group is ionized at low pH rates. Moreover, the positive charge of chitosan improves at low pH values, increasing the absorption of chitosan at the bacterial cell wall. Moreover, at upper pH values (> 6) the amino group of chitosan becomes aprotic, which may lead to precipitation from solution [24]. One study informed that chitosan's antimicrobial activities against *Klebsiella pneumoniae* partly resulted from the polycationic nature of chitosan, thus being associated with the protonation of the amino group. The protonation of the amino group is related to the degree of polymerization as well as the pH of the environment. For example, chitosan is more effective against *Candida lambica* at pH 4 than pH 6 [18].

In the early 1960s, chitosan's ability to bind to red blood cells was investigated. At that time, it was also seen as a hemostatic agent. Chitosan has been used in water purification for the last 30 years. Since then, numerous studies have been conducted to find ways to use these materials. Today, chitosan is known as a dietary supplement for weight loss. In fact, it has been marketed for this purpose in Japan as well as Europe for about 20 years. Many people even call it "anti-fat" [25]. Chitosan has attracted great attention because of its increasing demand as a highly beneficial biopolymer in recent times. Chitosan, which is obtained by deacetylation of chitin with sodium hydroxide (NaOH), can be extracted from a variety of fungi, insects, and crustaceans. Basically, chitosan is a polymer consisting of randomly distributed units of N-acetyl-D-glucosamine and D-glucosamine with different deacetylation degree, acetylation type, and molecular weight which could be chemically modified to its derivatives. These derivatives affect antibacterial influence of chitosan and its solubility in acidic solutions. Chitosan's three reactive functional groups are: the amino group at the C-6 position, the primary hydroxyl group at the C-6 position, and the secondary hydroxyl group at the C-3 position. The amino group at the C-6 position differs from chitosan obtained from chitin due to its chemical, physical, and biological functions [18]. Chitosan is a very useful and attractive biopolymer due to its diverse chemical structure. Structural diversity can be seen in MW ranging from low (100 kDa) to high (300 kDa) as well as DD ranging from chitin (< 60%) to chitosan (> 60%). The wide range of chitosan samples described

in different studies is surprising. Moreover, there are various conflicts regarding the use of chitosan in different biological applications [26].

Speaking of the synthesis of chitosan derivatives, the most beneficial advantage of chitosan is that it can be chemically modified into a wide variety of derivatives. Due to the presence of a primary alcohol group and an amino group, N, O-modified chitosan, as well as O-modified chitosan, can be modified to N-modified chitosan. The main reason for the synthesis of different chitosan derivatives is to improve certain properties. For example, quaternized chitosan derivatives have improved antimicrobial activity and water solubility, while phosphorylated chitosan derivatives have improved solubility, and N-benzyl/N-alkyl chitosan derivatives show improved antimicrobial activity [27]. Today, chitosan can be modified using two methods: Selective and non-selective modifications. The hydroxyl group is less nucleophilic than the amino group; however, both groups can still interact with electrophiles, including isothiocyanates and acids. These reactions lead to the selective O-chitosan derivative to be synthesized by a one-point reaction, while the non-selective N, O-chitosan derivative is synthesized. An acidic solution like sulfuric acid (H2SO4) can be used in production of the O-chitosan derivative. The amino group is protonated by using an acidic solution, which makes the alcohol functional group more reactive. This reaction preserves 90–95% of the amino acids; it is also a very effective and easy way of obtaining the O-modified chitosan derivative. On the other hand, the selective chitosan derivative equiped using this method is just limited to electrophiles and can only react with the amino group [28–30].

Due to its low cost, biocompatibility, absence of toxicity, and biodegradability, chitosan has applications in various fields such as tissue engineering, cosmetics, biomedicine, and biotechnology. Chitosan can be used to clarify agent wastewater and remove dye or metal ions due to its potential to protonate the amino group [31]. It can widely be used in the food industry as a browning inhibitor in juices, an antioxidant in sausages, a purifying agent in apple juices, and an antimicrobial agent. Chitosan can also be used to deliver transmucosal proteins and peptides thanks to its ability to adhere to the mucosa and open epithelial cell connections. Finally, it can be used as a carrier of macromolecular drugs. Conventionaly, chitosan has been used in its natural form with some limitations such as low surface area, low porosity, and low solubility at neutral pH. The functionality of chitosan can be increased by producing different derivatives through various chemical and physical processes [18].

Today, while preserving the organoleptic and nutritional properties of food products, great importance is attached to microbiological food safety. To accommodate these processes, the food industry must use special packaging materials that protect the quality and safety of food. Moreover, new generation food packaging materials are expected to have antimicrobial properties which create an environment that delays or completely prevents microbial growth, thus extending the shelf life of food products. Antimicrobial materials can be classified into two broad categories: organic materials and inorganic materials [32, 33]. Of particular interest as inorganic materials are metals, metal phosphates, and metal oxides considered safe for human and/or animal use. Inorganic substances are stable under severe conditions. However, examples of organic antimicrobial materials include halogenated compounds, quaternary ammonium salts, and phenols. Also, recent studies have found that natural polymers like chitosan and its derivatives have antibacterial activities. Thus, chitosan is promising substance that can be used in food packaging due to its ability to prevent gas or aroma in dry status and to form an excellent film [18] and for this purposes chitosan is used in various foods to extend shelf life mentioned in **Table 1**.

The antibacterial function of chitosan and its derivatives can be affected by different food ingredients. Charges and electrostatic forces on chitosan are the key

**269**

**Table 1.**

*Chitosan for Using Food Protection*

chitosan, Lychee fruit

lemon, and orange juice

**Juices**

*DOI: http://dx.doi.org/10.5772/intechopen.99247*

Apple, banana, citrus, mango, peach, carrot and lettuce coated with chitosan, Strawberry coated with

Use of soluble chitosan as a purifier in apple, grape,

**Applications in Meat and Meat Products**

**Applications in Dairy Products**

**Applications in Eggs**

*The effect of chitosan on some food groups.*

**Applications in Fruits and Vegetables**

**Food Impact / Finding**

Strawberry coated with chitosan It has been observed that shelf life increases

Lychee fruit The browning is delayed by preventing the increase

To control the acidity of carrots and apple juice It was observed to cause a significant decrease in

Apple and pear juice It has also been indicated it prevent enzymatic

Beef It was determined that the value of thiobarbutyric

Beef, fowl It was determined that the addition of 3% chitosan-

Pork products It was determined that chitosanglutamate used at

Sausage It has been determined that chitosan reduces the

Cheese *It has been reported that it inhibits the growth of* 

Mozzarella It has been determined that when used with the

Coated with chitosan (3% chitosan in 1% acetic acid) Reported at least 2 weeks longer shelf life of eggs at

Coated with chitosan-lysozyme mixture *Growth inhibition of L. monocytogenes, Salmonella* 

Decreased respiratory rate and ethylene production, caries control and softening delay were observed.

due to its antifungal properties and / or its ability to stimulate defense enzymes (chitinase

Fruit juices are purer than bentonite and gelatin, and the acceptance of fruit juices has increased.

acid (TBA) decreased by 70% compared to the control sample and had a positive effect on maintaining the red color of the meat during

glutamate reduced the development of Clostridium

0.3% level and 0.6% was an effective preservative and the total number of bacteria, yeast, mold and lactic acid bacteria decreased to 3 records as a result

use of sodium nitrite in sausage by half (150 ppm) without affecting quality and storage stability, and has also been found to reduce the amount of

*L. monocytogenes and S. aureus, but does not affect Gram-negative Pseudomonas aeroginosa.*

Lysozyme enzyme for film and coating purposes, it inhibits the growth of E. coli, L. monocytogenes, *Pseudomonas fluorescens* and yeast and molds and

*enterica, coliforms, yeast and mold, delayed moisture* 

25° C according to the control sample.

*loss and pH changes have been reported.*

and-1,3-glucanase).

titration acidity.

browning.

storage.

perfringens spores.

residual nitrite.

improves shelf life.

of storage at 4° C for 18 days.

in polyphenol oxidase activity.

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

use of chitosan in different biological applications [26].

in different studies is surprising. Moreover, there are various conflicts regarding the

limited to electrophiles and can only react with the amino group [28–30].

Due to its low cost, biocompatibility, absence of toxicity, and biodegradability, chitosan has applications in various fields such as tissue engineering, cosmetics, biomedicine, and biotechnology. Chitosan can be used to clarify agent wastewater and remove dye or metal ions due to its potential to protonate the amino group [31]. It can widely be used in the food industry as a browning inhibitor in juices, an antioxidant in sausages, a purifying agent in apple juices, and an antimicrobial agent. Chitosan can also be used to deliver transmucosal proteins and peptides thanks to its ability to adhere to the mucosa and open epithelial cell connections. Finally, it can be used as a carrier of macromolecular drugs. Conventionaly, chitosan has been used in its natural form with some limitations such as low surface area, low porosity, and low solubility at neutral pH. The functionality of chitosan can be increased by producing different derivatives through various chemical and physical processes [18]. Today, while preserving the organoleptic and nutritional properties of food products, great importance is attached to microbiological food safety. To accommodate these processes, the food industry must use special packaging materials that protect the quality and safety of food. Moreover, new generation food packaging materials are expected to have antimicrobial properties which create an environment that delays or completely prevents microbial growth, thus extending the shelf life of food products. Antimicrobial materials can be classified into two broad categories: organic materials and inorganic materials [32, 33]. Of particular interest as inorganic materials are metals, metal phosphates, and metal oxides considered safe for human and/or animal use. Inorganic substances are stable under severe conditions. However, examples of organic antimicrobial materials include halogenated compounds, quaternary ammonium salts, and phenols. Also, recent studies have found that natural polymers like chitosan and its derivatives have antibacterial activities. Thus, chitosan is promising substance that can be used in food packaging due to its ability to prevent gas or aroma in dry status and to form an excellent film [18] and for this purposes chitosan is used in various foods to extend shelf life

The antibacterial function of chitosan and its derivatives can be affected by different food ingredients. Charges and electrostatic forces on chitosan are the key

Speaking of the synthesis of chitosan derivatives, the most beneficial advantage of chitosan is that it can be chemically modified into a wide variety of derivatives. Due to the presence of a primary alcohol group and an amino group, N, O-modified chitosan, as well as O-modified chitosan, can be modified to N-modified chitosan. The main reason for the synthesis of different chitosan derivatives is to improve certain properties. For example, quaternized chitosan derivatives have improved antimicrobial activity and water solubility, while phosphorylated chitosan derivatives have improved solubility, and N-benzyl/N-alkyl chitosan derivatives show improved antimicrobial activity [27]. Today, chitosan can be modified using two methods: Selective and non-selective modifications. The hydroxyl group is less nucleophilic than the amino group; however, both groups can still interact with electrophiles, including isothiocyanates and acids. These reactions lead to the selective O-chitosan derivative to be synthesized by a one-point reaction, while the non-selective N, O-chitosan derivative is synthesized. An acidic solution like sulfuric acid (H2SO4) can be used in production of the O-chitosan derivative. The amino group is protonated by using an acidic solution, which makes the alcohol functional group more reactive. This reaction preserves 90–95% of the amino acids; it is also a very effective and easy way of obtaining the O-modified chitosan derivative. On the other hand, the selective chitosan derivative equiped using this method is just

**268**

mentioned in **Table 1**.


#### **Table 1.**

*The effect of chitosan on some food groups.*

factors enabling its antibacterial property; therefore, any food ingredient that can affect these factors inhibits chitosan's antimicrobial activity. For instance, inorganic cations (Mg2+) inhibit the adhesion of *E. coli* to hexadecane via chitosan as a result of disruption of the electrostatic interaction liable for chitosan adsorption to the organism cell surface. Also, the addition of a metal ion lessened the antimicrobial influence of the chitosan derivative against *Staphylococcus aureus*. It has also been informed that starch, α-lactalbumin and β-lactoglobulin (whey proteins), and sodium chloride (NaCl) have a negative effect on antibacterial function of chitosan; however, fat had no effect [34, 35].

Chitosan is used as a food additive in many countries, including Japan, Korea, and Italy, due to its many properties. Today, customers demand safe and quality food products. The food industry's need to extend the shelf life of food products has pushed research to identify improved preservation strategies [36]. The food industry is an area where important applications of chitosan are widely used. Reducing or preventing the number of chemicals in food is highly demanded in food industry. To meet this growing demand, chitosan can be used as an additive in food products. Chitosan can react with metals and prevent the initiation of lipid oxidation; therefore, it can be used as a secondary antioxidant. What's more, the antioxidant effect of chitosan can be increased by combining it with many other naturally occurring ingredients. For example, combining chitosan with glucose enhances its antioxidant property, but it does not affect its antibacterial influence against *E. coli*, *S. aureus*, *Bacillus subtilis*, and *Pseudomonas*. Chitosan can also be bound to other naturally occurring substances such as xylan to improve their antibacterial and antioxidant properties [37, 38]. In addition, the low oxygen permeability of chitosan can decrease the contact of food with oxygen, thereby reducing the oxidation rate. Chitosan and its derivatives can be used as a promising substance to extend the shelf life of various food products. For example, when a chitosan-based substance is used to coat certain food products, it can decrease bread hardness, retrogradation, weight loss, and bacterial development. The surface of eggs and fruits can be coated with chitosan to create a protective barrier that can decrease respiration and sweating rates, as well as prevent the transfer of gas and moisture from albumin through eggshells. Thus, chitosan can be used to improve the structure and quality of food products as well as prevent microbial growth and color changes [18]. It is known that cattle act as a native reservoir for the *E. coli* O157:H7 agent that causes most foodborne diseases. Unfortunately, the inhibition of *E. coli* O157:H7 contamination on meat and meat products has not been successful. Controlling the contamination of these pathogens is very important during processing level and to reduce the contamination of *E. coli* O157:H7 in cattle to an acceptable value. The effect of chitosan on *E. coli* O157:H7 infected calves was researched and the results defined that the time of fecal contamination was remarkably decreased in chitosan-treated animals compared to untreated animals. Also, chitosan administration did not cause any ration profitability or abnormal behavior [39].

One of the factors affecting the antimicrobial activity of chitosan is the DD. An increase in DD means an increased number of amino groups on chitosan. As a result, chitosan has an increasing number of protonated amino groups in an acidic condition and is fully soluble in water, which increases the likelihood of interaction between chitosan and negatively charged cell walls of microorganisms. A variation of the deacetylation process resulted in the variation of MW as well as significant differences in the % DD of chitosan. It has been proven that chitosans with low MW (< 10 kDa) have more antimicrobial activity than natural chitosans. Low MW fractions have little or no activity. Chitosan with a MW ranging from 10,000 to 100,000 Da will be useful in inhibiting bacterial growth. In addition, chitosan with an average MW of 9300 Da, was effective against *E. coli*. One researcher reported

**271**

*Chitosan for Using Food Protection*

effective [36].

*DOI: http://dx.doi.org/10.5772/intechopen.99247*

controversial use as a weight loss agent [42].

supplement suitable for human diets [7].

that while D-glucosamine hydrochloride (chitosan monomer) did not exhibit any growth inhibition against several bacteria, chitosan was effective. This suggests that the antimicrobial activity of chitosan is not only related to its cationic nature but also its chain length. However, another researcher found that 10,000 Da chitosan was least effective in bactericidal activities, while 220,000 Da chitosan was most

Chitosan is also used as an encapsulation material to improve food processing. Encapsulation is an attractive technology for protecting chemicals to prevent unwanted changes. Encapsulation materials can be formed with one or more compounds, such as chitosan, maltodextrin, acacia gum, hydroxypropyl methylcellulose phthalate gelatin, and starch, which can be used as a mixture or alone, among others. Chitosan has also attracted attention due to its applications in food and pharmacy. The antimicrobial and antifungal activities of chitosan are some of the most intriguing properties for improving food preservation and reducing the use of chemical preservatives. One study reported the use of chitosan in combination with essential oils, using nanoencapsulation processes, which have the potential to be applied in food industries. Due to the fact that essential oils such as thymol, eugenol, and carvacrol found in thyme, clove, and thyme essential oils easily degrade in light, air, and high temperatures, nanoencapsulation has recently been developed as

an effective technique to protect them from evaporation and oxidation [40].

The ion binding character of chitosan is another important quality. Chitosan has proven to have the best chelating properties among other natural polymers. Although hydroxyl groups may also be involved in absorption, the amino groups of chitosan are responsible for compound formation, in which nitrogen is a donor of electron pairs. The mechanism for collaborating the reactive groups with metal ions is very different and can link to the ion pattern, pH, and also the key ingredients of the solution. The constitution of compounds can also be reported based on Lewis' acid–base theory: the metal ion (acting as an acid) is the acceptor of the double electron given by the chitosan (acting as the base) [41]. With regard to food applications of chitosan, information on the selective binding of essential metal ions to chitosan is important for its application as a cholesterol-lowering agent and its more

Recently, researchers are increasingly interested in active food packaging materials, and there has been more interest in finding materials that provide biological activity to thin films as well as improving their properties. With the widespread use of non-fragile petroleum-based plastics, environmental pollution has become increasingly apparent. Most countries have placed restrictions on plastics,

and there is an increasing demand for biodegradable functional packaging materials. Among the many natural biopolymers, chitosan has gained increasing attention thanks to its non-toxicity, biodegradability, biocompatibility, antibacterial activity, and excellent film-forming ability. Chitosan is a native cationic linear polysaccharide created of D-glucosamine and N-acetyl-D-glucosamine units prepared by partial deacetylation of chitin. Chitosan has excellent features that enable it to be used as wound dressing in the medical area, for tissue engineering, and as food packaging in the industrial area [8]. As a result, chitosan is one of the most important edible films used worldwide, produced by the deacetylation of chitin. Many native biopolymers can be used to compose edible films; however, among them chitosan attracts the attention for its excellent film-forming activity, flexibility, stability, biocompatibility, non-toxicity, biodegradability, and commercial usability. Chitosan, which is a traditionally available polysaccharide with the deacetylation of chitin, was generally accepted as safe by FDA (United States Food and Drug Administration) in 2005 and was confirmed for use as a food

#### *Chitosan for Using Food Protection DOI: http://dx.doi.org/10.5772/intechopen.99247*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

however, fat had no effect [34, 35].

any ration profitability or abnormal behavior [39].

One of the factors affecting the antimicrobial activity of chitosan is the DD. An increase in DD means an increased number of amino groups on chitosan. As a result, chitosan has an increasing number of protonated amino groups in an acidic condition and is fully soluble in water, which increases the likelihood of interaction between chitosan and negatively charged cell walls of microorganisms. A variation of the deacetylation process resulted in the variation of MW as well as significant differences in the % DD of chitosan. It has been proven that chitosans with low MW (< 10 kDa) have more antimicrobial activity than natural chitosans. Low MW fractions have little or no activity. Chitosan with a MW ranging from 10,000 to 100,000 Da will be useful in inhibiting bacterial growth. In addition, chitosan with an average MW of 9300 Da, was effective against *E. coli*. One researcher reported

factors enabling its antibacterial property; therefore, any food ingredient that can affect these factors inhibits chitosan's antimicrobial activity. For instance, inorganic cations (Mg2+) inhibit the adhesion of *E. coli* to hexadecane via chitosan as a result of disruption of the electrostatic interaction liable for chitosan adsorption to the organism cell surface. Also, the addition of a metal ion lessened the antimicrobial influence of the chitosan derivative against *Staphylococcus aureus*. It has also been informed that starch, α-lactalbumin and β-lactoglobulin (whey proteins), and sodium chloride (NaCl) have a negative effect on antibacterial function of chitosan;

Chitosan is used as a food additive in many countries, including Japan, Korea, and Italy, due to its many properties. Today, customers demand safe and quality food products. The food industry's need to extend the shelf life of food products has pushed research to identify improved preservation strategies [36]. The food industry is an area where important applications of chitosan are widely used. Reducing or preventing the number of chemicals in food is highly demanded in food industry. To meet this growing demand, chitosan can be used as an additive in food products. Chitosan can react with metals and prevent the initiation of lipid oxidation; therefore, it can be used as a secondary antioxidant. What's more, the antioxidant effect of chitosan can be increased by combining it with many other naturally occurring ingredients. For example, combining chitosan with glucose enhances its antioxidant property, but it does not affect its antibacterial influence against *E. coli*, *S. aureus*, *Bacillus subtilis*, and *Pseudomonas*. Chitosan can also be bound to other naturally occurring substances such as xylan to improve their antibacterial and antioxidant properties [37, 38]. In addition, the low oxygen permeability of chitosan can decrease the contact of food with oxygen, thereby reducing the oxidation rate. Chitosan and its derivatives can be used as a promising substance to extend the shelf life of various food products. For example, when a chitosan-based substance is used to coat certain food products, it can decrease bread hardness, retrogradation, weight loss, and bacterial development. The surface of eggs and fruits can be coated with chitosan to create a protective barrier that can decrease respiration and sweating rates, as well as prevent the transfer of gas and moisture from albumin through eggshells. Thus, chitosan can be used to improve the structure and quality of food products as well as prevent microbial growth and color changes [18]. It is known that cattle act as a native reservoir for the *E. coli* O157:H7 agent that causes most foodborne diseases. Unfortunately, the inhibition of *E. coli* O157:H7 contamination on meat and meat products has not been successful. Controlling the contamination of these pathogens is very important during processing level and to reduce the contamination of *E. coli* O157:H7 in cattle to an acceptable value. The effect of chitosan on *E. coli* O157:H7 infected calves was researched and the results defined that the time of fecal contamination was remarkably decreased in chitosan-treated animals compared to untreated animals. Also, chitosan administration did not cause

**270**

that while D-glucosamine hydrochloride (chitosan monomer) did not exhibit any growth inhibition against several bacteria, chitosan was effective. This suggests that the antimicrobial activity of chitosan is not only related to its cationic nature but also its chain length. However, another researcher found that 10,000 Da chitosan was least effective in bactericidal activities, while 220,000 Da chitosan was most effective [36].

Chitosan is also used as an encapsulation material to improve food processing. Encapsulation is an attractive technology for protecting chemicals to prevent unwanted changes. Encapsulation materials can be formed with one or more compounds, such as chitosan, maltodextrin, acacia gum, hydroxypropyl methylcellulose phthalate gelatin, and starch, which can be used as a mixture or alone, among others. Chitosan has also attracted attention due to its applications in food and pharmacy. The antimicrobial and antifungal activities of chitosan are some of the most intriguing properties for improving food preservation and reducing the use of chemical preservatives. One study reported the use of chitosan in combination with essential oils, using nanoencapsulation processes, which have the potential to be applied in food industries. Due to the fact that essential oils such as thymol, eugenol, and carvacrol found in thyme, clove, and thyme essential oils easily degrade in light, air, and high temperatures, nanoencapsulation has recently been developed as an effective technique to protect them from evaporation and oxidation [40].

The ion binding character of chitosan is another important quality. Chitosan has proven to have the best chelating properties among other natural polymers. Although hydroxyl groups may also be involved in absorption, the amino groups of chitosan are responsible for compound formation, in which nitrogen is a donor of electron pairs. The mechanism for collaborating the reactive groups with metal ions is very different and can link to the ion pattern, pH, and also the key ingredients of the solution. The constitution of compounds can also be reported based on Lewis' acid–base theory: the metal ion (acting as an acid) is the acceptor of the double electron given by the chitosan (acting as the base) [41]. With regard to food applications of chitosan, information on the selective binding of essential metal ions to chitosan is important for its application as a cholesterol-lowering agent and its more controversial use as a weight loss agent [42].

Recently, researchers are increasingly interested in active food packaging materials, and there has been more interest in finding materials that provide biological activity to thin films as well as improving their properties. With the widespread use of non-fragile petroleum-based plastics, environmental pollution has become increasingly apparent. Most countries have placed restrictions on plastics, and there is an increasing demand for biodegradable functional packaging materials. Among the many natural biopolymers, chitosan has gained increasing attention thanks to its non-toxicity, biodegradability, biocompatibility, antibacterial activity, and excellent film-forming ability. Chitosan is a native cationic linear polysaccharide created of D-glucosamine and N-acetyl-D-glucosamine units prepared by partial deacetylation of chitin. Chitosan has excellent features that enable it to be used as wound dressing in the medical area, for tissue engineering, and as food packaging in the industrial area [8]. As a result, chitosan is one of the most important edible films used worldwide, produced by the deacetylation of chitin. Many native biopolymers can be used to compose edible films; however, among them chitosan attracts the attention for its excellent film-forming activity, flexibility, stability, biocompatibility, non-toxicity, biodegradability, and commercial usability. Chitosan, which is a traditionally available polysaccharide with the deacetylation of chitin, was generally accepted as safe by FDA (United States Food and Drug Administration) in 2005 and was confirmed for use as a food supplement suitable for human diets [7].

The most prominent properties of chitosan, as a compound obtained by various methods, can be attributed to its antimicrobial and antioxidant properties. Scientific publications reporting the antimicrobial activity of chitosan are specified in **Tables 2** and **3**. Considering these properties, the use of chitosan as an edible film to extend the shelf life of foods has been studied by many researchers.


**273**

*Chitosan for Using Food Protection*

**Chitosan or its derivatives**

Chitosan solution prepared in 1% acetic

Chitosan coating solution (1% and 2% w/v in 1% acetic acid)

Chitosan coating solution (2% w/v in 1%

Chitosan coating solution prepared with 2% (w/v) chitosan in 1% acetic acid

1% (w/v) chitosan coating solution in 1% v/v acetic acid and 0.2% (w/v) bamboo leaves

Chitosan, deacetylated 2% (w/v) in acetic acid

Chitosan-based edible

Chitosan (2% w/v) prepared in 1% acetic acid added with thyme oil (1% w/v)

at 1% v/v Chitosan coating solutions with 1.5% cinnamon oil added

coatings

acetic acid)

acid

*DOI: http://dx.doi.org/10.5772/intechopen.99247*

**Preparation method and/or foods Target microorganisms** 

Chitooligosaccharides of different

Chitooligosaccharides using papaya and dissolved in 0.25% acetic acid

molecular weights: > 100 kDa, 100 to 10 kDa, 10 to 1 kDa

Chitosan (0–2.0% w/w) Surimi gel made from black catfish

*niloticus*)

*mykiss*)

*molitrix*)

*mykiss*) fillets

(*Clarias gariepinus*)

Culture tilapia (*Oreochromis* 

Rainbow trout (*Oncorhynchus* 

Silver carp (*Hypophthalmichthys* 

Rainbow trout (*Oncorhynchus* 

Deepwater pink shrimp (*Parapenaeus longirostris*)

Butterfly-shaped rainbow trout (*Oncorhynchus mykiss*)

**and/or findings**

Antimicrobial effects were attributed to the type of strains. There was no

All microorganisms tested were inhibited but a higher effect was

Bacterial growth is inhibited.

A shelf life of 6 days was observed for the control group, while a shelf life of 12 days was observed for samples

respectively, for fillets treated with 1% and 2% chitosan compared to the control group, whose shelf life was

The shelf life of hot smoked fillets with a shelf life of 14–16 days, vacuumpacked and stored at +4 °C was extended to 24 days for fillets treated

total number of aerobic organisms, psychrophilic bacteria, lactic acid bacteria, and *Enterobacteriaceae*

The total number of living beings was higher in the control group stored at

When chitosan only and chitosan with essential oil were added, the shelf life with chitosan was doubled compared

The shelf life of shrimp treated with chitosan was extended by 3 days.

Compared to the control group, the shelf life of fillets treated with chitosan was extended by more than 15 days.

association with MW.

reported for *E. coli.*

treated with chitosan.

Sardine *(Sardinella longiceps*) fillets Shelf life increased to 7 and 9 days,

Carp (*Cyprinus carpio*) A decrease was determined in the

5 days.

with chitosan.

bacteria.

4 °C for 24 days.

to the control group.

*Escherichia coli Staphylococcus aureus Salmonella typhimurium Salmonella enteritidis*

*Bacillus cereus Bacillus subtilis Staphylococcus aureus Escherichia coli Pseudomonas aeruginosa Candida albicans Saccharomyces chevalieri Macrophomina phaseolina Aspergillus niger*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

Modified chitosan Chitosan obtained from shrimp

chitosan

Kitooligosaccharides Chitin (338 kDa MW and 35%

deacetylation grade) Kitooligosaccharide (chitin hydrolyzed with HCl) Kitooligosaccharide (HCl hydrolyzed chitosan, 80% deacetylation degree)

Chitosan oligomers hydrolyzed with nitrous acid (NaNO2 + CH3COOH) and dissolved in 1% acetic acid

chitin in three particle sizes by deacetylating with different concentrations of NaOH (30%, 40%, and 50%) under microwave irradiation for 10 minutes

The most prominent properties of chitosan, as a compound obtained by various methods, can be attributed to its antimicrobial and antioxidant properties. Scientific publications reporting the antimicrobial activity of chitosan are specified in **Tables 2** and **3**. Considering these properties, the use of chitosan as an edible film to extend the shelf life of foods has been studied by many researchers.

> In 1% acetic acid, 73.68% classical deacetylated chitosan, and 83.55% ultrasound-assisted deacetylated

Chitosan obtained by treating chitin with 50% NaOH and dissolved in 1% acetic acid without modification and with modification with ultraviolet or ozone

**Preparation method and/or foods Target microorganisms** 

**and/or findings**

*Salmonella typhimurium Escherichia coli*

*Staphylococcus aureus Escherichia coli Pseudomonas aeruginosa Klebsiella pneumonia Candida albicans Candida parapsilosis*

of deacetylation.

*Staphylococcus aureus Bacillus cereus Bacillus subtilis Escherichia coli Pseudomonas aeruginosa Aspergillus niger Candida albicans*

*Candida tropicalis* ve *Rhizopus* No difference was observed in the antibacterial properties of unmodified

Chitin showed a bacteriostatic effect on *E. coli*, *V. cholerae*, *S. dysenteriae,* and *B. fragilis*, while Chitosan showed a bacteriostatic effect on all bacteria tested except *S. typhimurium*.

and modified chitosan.

*Staphylococcus aureus Bacillus subtilis Bacillus cereus Escherichia coli Pseudomonas aeruginosa Salmonella typhimurium Vibrio cholerae Shigella dysenteriae Prevotella melaninogenica Bacteroides fragilis*

*Enterobacter aerogen Enterococcus faecalis Escherichia coli Staphylococcus aureus* Inhibition was observed in the microorganisms tested, but sharp inhibition was detected against

*E. faecalis.*

The inhibitory effect was greater against *S. typhimurium* than *E. coli.*

Antimicrobial activities are directly proportional to the increasing degree

**Chitosan or its derivatives**

**272**



#### **Table 2.**

*Studies revealing the antibacterial properties of chitosan, accordind to Olatunde et al. [43].*


**Table 3.**

*Antimicrobial activity of chitosan against some organisms in foods.*
