**3. Collagen from jumbo squid by-products**

Collagen is the main fibrous component of the connective tissue and the single most abundant protein in all organisms, since it represents up to 30% of the total protein in vertebrates and about 1-12% in aquatic organisms (Brinckmann, 2005). In general, collagen fibrils in the muscle of fish, form a delicate network structure with varying in the different connective tissues and is responsible for the integrity of the fillets (Shahidi, 2006). Moreover, the distribution of collagen may reflect the swimming behavior of the species (Sikorski et al., 1994). In several species of fish the weakening of the connective tissues may lead to serious quality deterioration that manifests itself by disintegration of the fillets. Also, thermal changes in collagen contributes to the desirable texture of the meat, however when heating is conducting under not controlled conditions, this may lead to serious losses due to the reduction in the breaking strength of the tissues (Sikorski et al., 1986).

The name collagen is in fact a generic term for a genetically distinct family of molecules that share a unique basic structure: three polypeptide chains coiled together to form a triple helix. About 19 different types of collagen molecules have been isolated and these not only varies in their molecular assembly, but also in their size, function, and tissue distribution (Table 1) (Exposito et al., 2002; Brinckmann, 2005).


\* Fibril associated collagen with interrupted triple helices Adapted from: Friess, 1998; Brinckmann, 2005.

Table 1. Chain composition, subfamily and body distribution of the main collagen types found in animal tissue.

The most abundant and widespread family of collagens, it is represented by the fibril-forming collagens, especially types I, III and V (Sato et al., 1989). Their basic structure consists of three protein chains that are supercoiled around a central axis in a right-handed manner to form a triple helix, called tropocollagen, which is a cylindrical protein of about 280 nm in length and 1.5 nm in diameter. Each chain, called α chain, contains about 1000 amino acids and it has a molecular weight of 100 kDa, depending on the source. Tropocollagen molecules may be formed by three identical chains (homotrimers) as in collagens II and III, or by two or more different chains (heterotrimers) as in collagen types I, IV, and V.

The three α chains are perfectly intertwined throughout the tropocollagen molecule to form the tripe helix except for the ends, where helical behavior is lost, since in these regions, called telopeptides, globular proteins that are involved with intermolecular crosslinking with other adjacent molecules are found (Engel & Bachinger, 2005). A structural prerequisite for the assembly of a continuous triple helix, the most typical conformation of collagen, is that every third position along the polypeptide chain is occupied by a glycine residue. Being glycine the smallest amino acid and lacking of a side chain, the collagen α chains can coil so tightly because glycine can be easily accommodated in the middle of a steric smooth superhelix and form stable packed structures; this would be very difficult with the bulkier residues. Further stabilization of the triple helix is attained by the formation of hydrogen bonds that are formed between the amino groups of glycine residues and the carbonyl groups of residues from other chains.

The structural constraints that make the collagen triple helix unique among proteins are given by its unusual amino acid content. Besides glycine as the major residue, a repeated sequence that characterizes the collagenous domains of all collagens is the triplet (Gly-X-Y), where X and Y are often proline and hydroxiproline, respectively (Figure 2). Depending on the collagen type, specific proline and lysine residues are modified by post-translational enzymatic hydroxylation. These imino acids permit the sharp twisting of the collagen helix and are associated with the stability and thermal behavior of the triple helical conformation.

Source: Branden & Tooze, 1999; Voet & Voet, 1995.

Fig. 2. Spatial conformation of a typical tropocollagen molecule.

#### **3.1 Sources and extraction**

22 Recent Advances in Plasticizers

(42%) is usually used, which is later primarily marketed fresh, frozen or pre-cooked (Luna-Raya et al., 2006). Some of the by-products produced after filleting, like fins and heads, are utilized but huge amounts are wasted. Fortunately, a number of studies have reported that this waste is an excellent raw material to obtain important by-products with high commercial value, such as collagen (Gómez-Guillen et al., 2002; Kim et al., 2005, Shahidi,

Collagen is the main fibrous component of the connective tissue and the single most abundant protein in all organisms, since it represents up to 30% of the total protein in vertebrates and about 1-12% in aquatic organisms (Brinckmann, 2005). In general, collagen fibrils in the muscle of fish, form a delicate network structure with varying in the different connective tissues and is responsible for the integrity of the fillets (Shahidi, 2006). Moreover, the distribution of collagen may reflect the swimming behavior of the species (Sikorski et al., 1994). In several species of fish the weakening of the connective tissues may lead to serious quality deterioration that manifests itself by disintegration of the fillets. Also, thermal changes in collagen contributes to the desirable texture of the meat, however when heating is conducting under not controlled conditions, this may lead to serious losses due to the

The name collagen is in fact a generic term for a genetically distinct family of molecules that share a unique basic structure: three polypeptide chains coiled together to form a triple helix. About 19 different types of collagen molecules have been isolated and these not only varies in their molecular assembly, but also in their size, function, and tissue distribution

**Type Chain composition Subfamily Tissue distribution** 

I [(α1(I))2α2(I)] Fibrillar Skin, tendon, bone, ligament, vessel II [(α1(II))3] Fibrillar Hyaline cartilage, vitreous III [(α1(III))3] Fibrillar Skin, vessel, intestine, uterus IV [(α1(IV))2α2(IV)] Network Basement membranes V [α1(V)α2(V)(3(V)] Fibrillar Bone, skin, cornea, placenta VI [α1(VI)α2(VI)α3(VI)] Network Interstitial tissue VII [(α1(VII))3] Anchoring fibrils Epithelial tissue

VIII [α1(VIII) α2(VIII)] Network Endothelial tissue, descement's

IX [α1(IX)α2(IX)α3(IX)] FACIT\* Cartilage, cornea, vitreous X [(α1(X))3] Network Hypertrophic and mineralizing

XI [α1(XI)α2(XI)α3(XI)] Fibrillar Cartilage, intervertebral disc XII [(α1(XII))3] FACIT\* Skin, tendon, cartilage

Table 1. Chain composition, subfamily and body distribution of the main collagen types

membrane

cartilage

2006; Torres-Arreola et al., 2008; Gimenez et al., 2009).

reduction in the breaking strength of the tissues (Sikorski et al., 1986).

**3. Collagen from jumbo squid by-products** 

(Table 1) (Exposito et al., 2002; Brinckmann, 2005).

\* Fibril associated collagen with interrupted triple helices

Adapted from: Friess, 1998; Brinckmann, 2005.

found in animal tissue.

The main sources of industrial collagen are limited to the skin and bones of pigs and cattle. However, as a possible alternative to the problems associate to transmissible bovine

By-Products From Jumbo Squid (*Dosidicus gigas*): A New Source of Collagen Bio-Plasticizer? 25

under non-denaturing conditions have been reported and found to be quite effective.

medical and it is widely use in the form of collagen casings.

and degraded by heat and chemicals to produce a soluble form.

**3.2 Collagen as biomaterial and applications** 

In the case of the mantle of *Dosidicus gigas*, Uriarte-Montoya et al. (2010) used urea 6M and the three solvent systems described above to isolate collagen from this species. They found that collagen from the mantle was 37% soluble in a neutral salt solvent, 23% in a dilute acid solvent and 25% in a dilute acid solvent containing pepsin, whereas the remaining 15% of the connective tissue was insoluble. They concluded that depending on the ultimate application, each collagen fraction warrants sufficient importance and might be used for different purposes rather than the traditional industrial collagen applications. As it will be mentioned in the next subsection, collagen has a wide variety of application, from food to

Nowadays, collagen has several industrial and biomedical applications. In the former, collagen has been used from long time ago, whereas the latter have placed collagen as an object of intense research in the last years. Industrially, collagen has been used for leather processing and gelatin production. Both products consist mainly of collagen but they greatly differ in the chemical form of the collagen used (Meena et al., 1999). Leather is basically chemically treated animal skin, while gelatin is an animal connective tissue that is denatured

Regarding the biomedical uses of collagen, probably one of the main attraction of collagen as a biomaterial is its low immunogenicity. Moreover, collagen can be processed into various presentations, such as sheets, tubes, sponges, powders, injectable solutions and dispersions, making it a functional component for specific applications in ophthalmology, wounds and burns, tumor treatment, engineering and tissue regeneration, among others areas (Kim & Mendis, 2006). Table 2 summarizes the main advantages and disadvantages of collagen as a biomaterial. Another recent application of collagen in biomedicine is found in the formulation of membranes and hydrogels, products in which collagen interacts with

achieve a 1:10 ratio between the enzyme and the dry weight of the tissue to be extracted. The effectiveness of this system lies in its ability to solubilize native collagen (Miller & Rhodes, 1982). Despite its great efficacy, an essential and adequate control is needed for these systems, since pepsin is an exogenous proteolytic enzyme and if not controlled, it may contribute to the total disorganization or degradation of the collagen molecules. After the different soluble collagen fractions are attained, the insoluble collagen remains. This fraction is characterized by numerous crosslinks within the molecule, which in turn prevent its disintegration and make it more resistant to proteolysis. Physically, insoluble collagen is an opalescent fibrous material that can only be degraded using either strong denaturing agents or mechanical fragmentation under acidic conditions (Friess, 1998). The insolubility of collagen is attributed to the modification of the forces that hold in together the α-chains. Its presence is also related to the age of the animal and to its frozen storage, due to the protein aggregation that results from the removal of water molecules from the stromal proteins (Montero et al., 2000). Finally, once all the soluble and insoluble collagen fractions are collected, a selective neutral or dilute acid- salt precipitation procedure is usually performed in order to initially purify and recover the individual collagen types that may be present in extracts from various tissues. In the case that a final purification and resolution of the collagen molecules is required, different chromatographic techniques

spongiform encephalopathy (BSE) and foot and mouth disease (FMD), as well as religious barriers, new alternatives are being sought for collagen sources (Jongjareonraka et al., 2005; Nagai, 2004). As a result, the extraction of collagen and their derivatives from marine sources has considerably increased in the last years since it represents an appropriate alternative source to land animals with promising functional properties (Shen et al., 2007).

Collagen typically exists in a concentration from 3 to 11.1% in the mantle of some squid species like *Illex* and *Loligo* (Sikorski & Kolodziejska, 1986), whereas in *Dosidicus gigas*, collagen was found in a concentration up to 18.33% (Torres-Arreola et al., 2008). This variability among squid species may be attributed to the high degree of protein turnover that takes place in the muscle of cephalopods, which are fast-growing species as they usually reach their maximum maturity in one year or two. In general, the collagen from aquatic organisms, is highly soluble in salt solutions, dilute acids and acid buffers, unlike collagen from land mammals that are poorly soluble in such solvents (Kolodziejska et al., 1999). Consequently, in order to extract collagen from marine organisms, the most common solvent systems that have been found to be generally useful and convenient are described below; however, there is no single standard method for the isolation of collagen (Miller & Rhodes, 1982).

Prior to the actual collagen extraction procedure, it is a prerequisite to wash or digest the original tissue with a dilute alkali (*e.g*. 0.1 M NaOH) in order to remove non-collagenous proteins and to prevent the effect of endogenous proteases on collagen. This procedure can also be achieved by using caothropic solutes such as urea in high concentrations (6 M). These organic solutes are capable of increasing the ionic strength of the medium and breakdown the structure of water, which causes the disruption of the hydrogen bonds among the α chains, which induces the unfolding and solubilization of hydrophobic residues inside the protein molecule, only affecting the soluble protein fractions (myofibrilar and sarcoplasmic) but leaving the stromal proteins intact (Usha & Ramasami, 2004). Once the connective tissue have been isolated, the following collagen extraction procedures are usually conducted at low temperatures (4-8°C) with the aim to minimize bacterial growth, enhance the solubility of native collagens, and to ensure the retention of native conformation on the part of the solubilized collagens (Miller & Rhodes, 1982):


spongiform encephalopathy (BSE) and foot and mouth disease (FMD), as well as religious barriers, new alternatives are being sought for collagen sources (Jongjareonraka et al., 2005; Nagai, 2004). As a result, the extraction of collagen and their derivatives from marine sources has considerably increased in the last years since it represents an appropriate alternative source to land animals with promising functional properties (Shen et al., 2007). Collagen typically exists in a concentration from 3 to 11.1% in the mantle of some squid species like *Illex* and *Loligo* (Sikorski & Kolodziejska, 1986), whereas in *Dosidicus gigas*, collagen was found in a concentration up to 18.33% (Torres-Arreola et al., 2008). This variability among squid species may be attributed to the high degree of protein turnover that takes place in the muscle of cephalopods, which are fast-growing species as they usually reach their maximum maturity in one year or two. In general, the collagen from aquatic organisms, is highly soluble in salt solutions, dilute acids and acid buffers, unlike collagen from land mammals that are poorly soluble in such solvents (Kolodziejska et al., 1999). Consequently, in order to extract collagen from marine organisms, the most common solvent systems that have been found to be generally useful and convenient are described below; however, there is no

Prior to the actual collagen extraction procedure, it is a prerequisite to wash or digest the original tissue with a dilute alkali (*e.g*. 0.1 M NaOH) in order to remove non-collagenous proteins and to prevent the effect of endogenous proteases on collagen. This procedure can also be achieved by using caothropic solutes such as urea in high concentrations (6 M). These organic solutes are capable of increasing the ionic strength of the medium and breakdown the structure of water, which causes the disruption of the hydrogen bonds among the α chains, which induces the unfolding and solubilization of hydrophobic residues inside the protein molecule, only affecting the soluble protein fractions (myofibrilar and sarcoplasmic) but leaving the stromal proteins intact (Usha & Ramasami, 2004). Once the connective tissue have been isolated, the following collagen extraction procedures are usually conducted at low temperatures (4-8°C) with the aim to minimize bacterial growth, enhance the solubility of native collagens, and to ensure the retention of native

a. Neutral salt solvents (*e.g.* 1 M NaC1, 0.05 M Tris, pH 7.5): The high ionic strength of this solvent system allows to solubilize newly synthesized or young collagen molecules (Friess, 1998). Moreover, this solvent is also capable of solubilize more components beyond the stromal fractions, such as remaining myofibrillar proteins. Therefore, this

b. Dilute acid solvents (e.g. 0.5 M acetic acid): Dilute acids such as acetic acid, hydrochloric acid, or citrate buffer are widely used to dissolve some pure collagen, although they are only limited to the portion of non-crosslinked collagen (Jongjareonrak et al., 2005). The pH of these solvents is about 3.0, which exhibits a sufficient capacity to induce swelling of most tissues promoting the solubilization of the triple helix junctions. Collagen extracted using these solvents usually have greater industrial applications in comparison with other soluble fractions because of the facility to incorporate the biomolecule with other

c. Dilute acid solvents containing pepsin (*e.g.* 0.5 M acetic acid plus EC 3.4.23.1): These systems are one of the most versatile and widely used procedures for the extraction of collagen. The enzyme is usually added to the acidic solvent in sufficient quantities to

single standard method for the isolation of collagen (Miller & Rhodes, 1982).

conformation on the part of the solubilized collagens (Miller & Rhodes, 1982):

solvent shows the least ability to solubilize pure collagen.

polymers in an acidic medium (Garcia et al., 2007).

achieve a 1:10 ratio between the enzyme and the dry weight of the tissue to be extracted. The effectiveness of this system lies in its ability to solubilize native collagen (Miller & Rhodes, 1982). Despite its great efficacy, an essential and adequate control is needed for these systems, since pepsin is an exogenous proteolytic enzyme and if not controlled, it may contribute to the total disorganization or degradation of the collagen molecules.

After the different soluble collagen fractions are attained, the insoluble collagen remains. This fraction is characterized by numerous crosslinks within the molecule, which in turn prevent its disintegration and make it more resistant to proteolysis. Physically, insoluble collagen is an opalescent fibrous material that can only be degraded using either strong denaturing agents or mechanical fragmentation under acidic conditions (Friess, 1998). The insolubility of collagen is attributed to the modification of the forces that hold in together the α-chains. Its presence is also related to the age of the animal and to its frozen storage, due to the protein aggregation that results from the removal of water molecules from the stromal proteins (Montero et al., 2000). Finally, once all the soluble and insoluble collagen fractions are collected, a selective neutral or dilute acid- salt precipitation procedure is usually performed in order to initially purify and recover the individual collagen types that may be present in extracts from various tissues. In the case that a final purification and resolution of the collagen molecules is required, different chromatographic techniques under non-denaturing conditions have been reported and found to be quite effective.

In the case of the mantle of *Dosidicus gigas*, Uriarte-Montoya et al. (2010) used urea 6M and the three solvent systems described above to isolate collagen from this species. They found that collagen from the mantle was 37% soluble in a neutral salt solvent, 23% in a dilute acid solvent and 25% in a dilute acid solvent containing pepsin, whereas the remaining 15% of the connective tissue was insoluble. They concluded that depending on the ultimate application, each collagen fraction warrants sufficient importance and might be used for different purposes rather than the traditional industrial collagen applications. As it will be mentioned in the next subsection, collagen has a wide variety of application, from food to medical and it is widely use in the form of collagen casings.

#### **3.2 Collagen as biomaterial and applications**

Nowadays, collagen has several industrial and biomedical applications. In the former, collagen has been used from long time ago, whereas the latter have placed collagen as an object of intense research in the last years. Industrially, collagen has been used for leather processing and gelatin production. Both products consist mainly of collagen but they greatly differ in the chemical form of the collagen used (Meena et al., 1999). Leather is basically chemically treated animal skin, while gelatin is an animal connective tissue that is denatured and degraded by heat and chemicals to produce a soluble form.

Regarding the biomedical uses of collagen, probably one of the main attraction of collagen as a biomaterial is its low immunogenicity. Moreover, collagen can be processed into various presentations, such as sheets, tubes, sponges, powders, injectable solutions and dispersions, making it a functional component for specific applications in ophthalmology, wounds and burns, tumor treatment, engineering and tissue regeneration, among others areas (Kim & Mendis, 2006). Table 2 summarizes the main advantages and disadvantages of collagen as a biomaterial. Another recent application of collagen in biomedicine is found in the formulation of membranes and hydrogels, products in which collagen interacts with

By-Products From Jumbo Squid (*Dosidicus gigas*): A New Source of Collagen Bio-Plasticizer? 27

In its native form, the chitosan is insoluble in water and in the majority of organic solvents; nevertheless, at pH values <6.0 chitosan easily dissolve in diluted acidic solutions due to the protonation of the amino groups, turning it into a soluble cationic polysaccharide (Agulló et al., 2004; Raafat & Sahl, 2009; Sharma et al., 2009). Commercially, chitosan is available in several grades of purity, molecular weight, distribution and length of chain, deacetylation degree, density, viscosity, solubility, water retention capacity and the distribution of the amino/acetamide groups. All these characteristics affect the physicochemical properties and

Among their biological properties, the antibacterial, antifungal, antiviral and insecticide activity of chitosan and its derivatives (Badawy & Rabea, 2005; Badawy et al., 2005) have been explored for agricultural applications (Daayf et al., 2010). Due to its peculiar

Biomedicine It promotes the growth of tissues, wound healing (bandages, sutures),

Agriculture Preservation of fruits and vegetables, soils supplements, release of

Paper and textile Flexibility and resistance. It improves stickiness of inks and clothes

protects against microbial damage.

flocculent, homeostatic, bacteriostatic/fungistatic, spermicide, antitumoral, anticholesterolemic, immunoadjuvant, sedative of the central nervous system, acceleration of the osteoblast's formation.

agrochemicals and nutrients supply, improves the seed's germination,

Bleaching of waste water, removal of heavy metals, water clarification.

stains, stabilizes the color and resistance of cloths. It improves the paper sheen, the resistance to the microbial and enzymatic

of photographic paper, improves the paper air-tightness.

Water retention, reduces the lipids and cholesterol absorption, antiacid agent, food fibre, food formulation for babies, emulsifier, preservative, antioxidant, gellificant, clarficant of fruit juices,

It protects and preserves food, decreases the production of ethylene and CO2, antimicrobial, reduces the water permeability, controls

Cosmetics Solar and moisturizing protector, decrease the expression lines, useful

in contact glasses and hair conditioners.

Chromatography Enzymes separation and gas chromatography.

immobilization of enzymes.

enzymatic oxidation.

Table 3. Applications of chitosan and chitosan composite films.

Preservative Antibacterial, antifungal.

deterioration; improves the paper biodegradability and the antistatic

characteristics, chitosan has potential applications in diverse fields (Table 3).

therefore, its application (Raafat & Sahl, 2009).

Pharmaceutics Controlled release of medicines.

**Industry Applications** 

Waste-water treatment

Supplement or food additive

Films and eatable recovering

Adapted from: Agulló et al., 2004

another material to form a composite. The combination of collagen with chitosan is perhaps one of the most studied composites with practical emphasis on medicine, dentistry, and pharmacology (Lima et al., 2006).


Table 2. Main Advantages and disadvantages of collagen as a biomaterial.

#### **4. Chitosan**

Chitosan, the main deacetylated derivative of chitin, is a biocompatible, non-toxic, edible, biodegradable polymer, and it possesses antimicrobial activity against bacteria, yeast and fungi, including toxigenic fungi (Cota-Arriola et al., 2011). This polymer is the only natural cationic polysaccharide, which special characteristics makes it useful in numerous applications and areas, such medicine, cosmetics, food packing, food additives, water treatment, antifungal agent, among others (Hu et al., 2009). At a commercial level, chitosan is obtained from the thermo-alkaline deacetylation of chitin from crustaceans (Kurita, 2006), and its production has taken great importance in the ecological and economic aspects, due to the use of marine by-products (Nogueira et al., 2005).

Chemically, chitosan is a linear polycationic hetero-polysaccharide (poly [β-(1,4)-2-amino-2 deoxi-D-glucopyranose]) (Figure 3) of high molecular weight, whose polymeric chain changes in size and deacetylation degree (Tharanathan & Srinivasa, 2007). The nitrogen forms a primary amine and causes N-acylation reactions and Schiff alkalis formation, while the hydroxyl (OH) and amino (NH2) groups allow the formation of hydrogen bonds (Agulló et al., 2004).

Fig. 3. Chemical structure of chitosan showing the characteristic amino (NH2) groups.

In its native form, the chitosan is insoluble in water and in the majority of organic solvents; nevertheless, at pH values <6.0 chitosan easily dissolve in diluted acidic solutions due to the protonation of the amino groups, turning it into a soluble cationic polysaccharide (Agulló et al., 2004; Raafat & Sahl, 2009; Sharma et al., 2009). Commercially, chitosan is available in several grades of purity, molecular weight, distribution and length of chain, deacetylation degree, density, viscosity, solubility, water retention capacity and the distribution of the amino/acetamide groups. All these characteristics affect the physicochemical properties and therefore, its application (Raafat & Sahl, 2009).

Among their biological properties, the antibacterial, antifungal, antiviral and insecticide activity of chitosan and its derivatives (Badawy & Rabea, 2005; Badawy et al., 2005) have been explored for agricultural applications (Daayf et al., 2010). Due to its peculiar characteristics, chitosan has potential applications in diverse fields (Table 3).


Adapted from: Agulló et al., 2004

26 Recent Advances in Plasticizers

another material to form a composite. The combination of collagen with chitosan is perhaps one of the most studied composites with practical emphasis on medicine, dentistry, and

**Advantages Disadvantages** 

more rapid release

compared with hydrolytic degradation

Biodegradable and biocompatible Hydrophilicity may lead to swelling and

Synergic with bioactive compounds Variability in enzymatic degradation rate as

Hemostatic Possible side effects such as mineralization

Chitosan, the main deacetylated derivative of chitin, is a biocompatible, non-toxic, edible, biodegradable polymer, and it possesses antimicrobial activity against bacteria, yeast and fungi, including toxigenic fungi (Cota-Arriola et al., 2011). This polymer is the only natural cationic polysaccharide, which special characteristics makes it useful in numerous applications and areas, such medicine, cosmetics, food packing, food additives, water treatment, antifungal agent, among others (Hu et al., 2009). At a commercial level, chitosan is obtained from the thermo-alkaline deacetylation of chitin from crustaceans (Kurita, 2006), and its production has taken great importance in the ecological and economic aspects, due

Chemically, chitosan is a linear polycationic hetero-polysaccharide (poly [β-(1,4)-2-amino-2 deoxi-D-glucopyranose]) (Figure 3) of high molecular weight, whose polymeric chain changes in size and deacetylation degree (Tharanathan & Srinivasa, 2007). The nitrogen forms a primary amine and causes N-acylation reactions and Schiff alkalis formation, while the hydroxyl (OH) and amino (NH2) groups allow the formation of hydrogen bonds (Agulló

Fig. 3. Chemical structure of chitosan showing the characteristic amino (NH2) groups.

from living organisms High cost of pure type collagen Non-antigenic and non-toxic Variability of isolated collagen

presentations Complex handling properties

Table 2. Main Advantages and disadvantages of collagen as a biomaterial.

pharmacology (Lima et al., 2006).

Available in abundance and easily purified

Formulated in a number of different

**4. Chitosan** 

et al., 2004).

Easily modifiable to produce other materials Plasticity due to high tensile strength Compatible with synthetic polymers Adapted from: Friess, 1998; Lee et al., 2001.

to the use of marine by-products (Nogueira et al., 2005).

Table 3. Applications of chitosan and chitosan composite films.

By-Products From Jumbo Squid (*Dosidicus gigas*): A New Source of Collagen Bio-Plasticizer? 29

It improves the tensile force, the thermal degradation and the glass

Increased flexibility and barrier properties; decreased rigidity; increased in solubility; affected color.

Improve the elastic properties, enhanced the protein adsorption, cell adhesion, growth and proliferation.

Decrease in thermal properties, increase in percentage of elongation and CO2 and water permeability

High plasticizer contents caused decrease in both tensile and modulus.

Recently, the potential action of acid soluble collagen from jumbo squid (ASC) as a plasticizer in chitosan films was evaluated (Uriarte-Montoya et al., 2010; Arias-Moscoso et al., 2011). In general, the use of acid soluble collagen from jumbo squid by-products, in amounts equal or lower than 50 % in the production of chitosan biocomposites, produces films with reasonable tensile properties (Arias-Moscoso et al., 2011). More details about it

Biomaterials are increasingly being used in several fields, like packaging material (Tharanathan, 2003), tissue engineering (Nalwa, 2005), medical and pharmaceutical

Collagen, as a natural polymer, has very weak mechanical properties, especially in aqueous media (Zhang et al., 1997). On the other hand, as it was mentioned previously, chitosan poses excellent film-forming property, antimicrobial activity, and unique coagulating ability with metal and other lipid and protein complexes due to the presence of a high density of amino groups and hydroxyl groups in the polymeric structure of chitosan (Dutta et al., 2002; Li et al., 1992; Shahidi et al., 1999). Although, mechanical properties of chitosan films are comparable to those of many medium-strength commercial polymers, it is considered that

equal to that of HDPE and LDPE films

Tensile strength values

Improves ductility, increase in strain and decrease in stress, and increases

Good flexibility and mechanical force. (Fernández et al.,

(Chen et al., 2009)

(Bourtoom, 2007)

(Suyatama et al.,

(Zhang et al., 2002)

(Arvanitoyannis et al.,

(Butler et al., 1996).

2005)

2004)

1997)

**Film/Plasticizer Physicochemical properties Reference** 

film hydrophilicity. Propylene glycol exposes antiplasticization phenomenon.

increase.

Table 4. Plasticizer effect on the properties of chitosan films.

applications (Bures et al., 2001) due to their functional properties.

will discuss in the following subsections.

**5. Chitosan and collagen films** 

transition temperatures.

Chitosan Lignine

Starch-chitosan film/ Sorbitol; glycerol; polyethylene glycol

Glycerol; ethylene glicol; poly (ethylene glycol); propylene glycol

Glycerol; sorbitol; i-erythritol

Poly(vinyl alcohol; sorbitol;

Chitosan film/

Chitosan film/

Chitosan film/ Poly (ethylene glycol)

Chitosan film/

Chitosan films/ Glycerol

sucrose
