*1.2.2 Crystalline structure*

Chitin exists in three different polymorphic forms, which are α-chitin, β-chitin, and γ-chitin (**Figure 2**). The interaction between C═O⋯NH and C═O⋯OH maintaining the strength of the polymeric network chain [13]. α-chitin appears in its antiparallel structure and the chain is interacting through both inter-and intramolecular hydrogen bond. β-chitin has a parallel structure, which leads to the formation of the intramolecular hydrogen bond. γ-chitin consists of both antiparallel and parallel structure, as it is the combination of α-chitin and β-chitin [21].

β-Chitin exhibits better water solubility but less common as compared to α-chitin [14]. It has been shown that α-chitosan has a higher crystallinity index as compared to β-chitin. However, the crystallinity index for both forms is lower than the raw chitosan [21]. The crystallinity index will increase when the degree of deacetylation of chitosan increases [16].

The β-chitin has a high affinity to the organic solvent due to its structural flexibility [14]. It exhibits higher reactivity than α-chitosan due to a lack of hydrogen bond. This form has a high capability to swell between crystalline structures while losing its crystalline fraction [21]. The swelling of β-chitin sometimes disrupts the polymeric chain and crystalline structure.

**Figure 2.** *The different conformational structure between* α*-chitin,* β*-chitin and* γ*-chitin (adapted from [14]).*

**55**

*Chitosan-Based Oral Drug Delivery System for Peptide, Protein and Vaccine Delivery*

The particle size of chitosan plays a major role in developing an efficient carrier for peptide drugs [22]. Monodisperse preparation of nanoparticles is desirable to provide better bioavailability and low toxicity [23]. Polydispersity leads to a larger size distribution, which interferes with the tendency for the nanocarrier to accumu-

PDI increases with an increase in molecular weight. However, it decreases as the degree of deacetylation increases [23]. The increase of amino group protonation and removal of the acetyl group from chitosan structure lead to the enhancement of repulsive forces between molecules and stretch the chitosan to become larger in size [11, 24]. Therefore, the development of chitosan with the optimum degree of deacetylation is needed to minimise the risk of polydispersity. This can be achieved by modifying the time and temperature of the de-*N*-acetylation process [18]. The degree of entanglement for HMWC nanoparticles is higher than LMWC. Therefore, HMWC nanoparticle has a high tendency to aggregate with each other and disrupt the uniformity of the system [25]. However, LMWC cannot be loaded into nanoparticles with smaller size due to its limitation to entangle to the structures of the system [26]. Therefore, maintaining the particle size of the chitosan is crucial

Modified chitosan shows greater advantages as compared to unmodified chitosan. The modification of chitosan either chemically or physically may improve its solubility, properties of gelling, and biocompatibility. This modification can be done through cross-linking or substitution [27]. The presence of the various reactive functional groups in the chitosan structure makes it available in many derivatives with different

A quaternary ammonium salt is a hydrophilic group with a permanent positive charge. Therefore, quaternary chitosan does not need an acidic condition to undergo protonation [12, 28]. It allows chitosan quaternary ammonium salts to be soluble in both acidic and basic pH. This is a good approach to increase chitosan

The high strength of the positive charge will weaken the hydrogen bond. However, this activity depends on the degree of substitution. The higher the degree of substitution, the higher the water solubility of chitosan. This will improve the quality of chitosan to act as a mucoadhesive agent that aids the penetration into

Trimethyl chitosan (TMC) is an example of a chitosan derivative from quaternisation. This modification is effective in enhancing the bioavailability of antibacterial drugs with antibacterial properties. Moreover, quaternised chitosan also exhibits antibacterial properties by the interaction of its positive charge with the

Polydispersity describes the degree of non-uniformity of size distribution between molecules due to the aggregation or agglomeration of the polymeric network. It can be estimated using the polydispersity index (PDI), where the ideal index for chitosan nanoparticles is below 0.3 [22, 23]. The degree of deacetylation and molecular weight of chitosan have been proven to influence the polydispersity

*DOI: http://dx.doi.org/10.5772/intechopen.95771*

late in the target tissue [23].

of the system [24].

stability properties.

*1.3.1 Quaternisation*

solubility in water [28].

mucus [29].

*1.2.3 Polydispersity and particle size of chitosan*

in the development of a chitosan nanoparticle.

**1.3 Modification of chitosan as biomaterial**

negative charge of Gram-negative bacteria [28–30].

*Chitosan-Based Oral Drug Delivery System for Peptide, Protein and Vaccine Delivery DOI: http://dx.doi.org/10.5772/intechopen.95771*

## *1.2.3 Polydispersity and particle size of chitosan*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

to be spontaneous.

of amino groups [20].

*1.2.2 Crystalline structure*

deacetylation of chitosan increases [16].

polymeric chain and crystalline structure.

usable energy in the system. Loss of energy means the reaction in the system tends

The α-chitin crystalline form exhibit lower water solubility as compared to β-chitin. The shorter polymeric chain of low molecular weight chitosan (LMWC) is unlikely to aggregate [11]. Interaction between molecules declines due to the formation of the hydrogen bonds is limited. A short chitosan chain contains a low number

Chitin exists in three different polymorphic forms, which are α-chitin, β-chitin, and γ-chitin (**Figure 2**). The interaction between C═O⋯NH and C═O⋯OH maintaining the strength of the polymeric network chain [13]. α-chitin appears in its antiparallel structure and the chain is interacting through both inter-and intramolecular hydrogen bond. β-chitin has a parallel structure, which leads to the formation of the intramolecular hydrogen bond. γ-chitin consists of both antiparallel and

parallel structure, as it is the combination of α-chitin and β-chitin [21].

β-Chitin exhibits better water solubility but less common as compared to α-chitin [14]. It has been shown that α-chitosan has a higher crystallinity index as compared to β-chitin. However, the crystallinity index for both forms is lower than the raw chitosan [21]. The crystallinity index will increase when the degree of

The β-chitin has a high affinity to the organic solvent due to its structural flexibility [14]. It exhibits higher reactivity than α-chitosan due to a lack of hydrogen bond. This form has a high capability to swell between crystalline structures while losing its crystalline fraction [21]. The swelling of β-chitin sometimes disrupts the

*The different conformational structure between* α*-chitin,* β*-chitin and* γ*-chitin (adapted from [14]).*

**54**

**Figure 2.**

The particle size of chitosan plays a major role in developing an efficient carrier for peptide drugs [22]. Monodisperse preparation of nanoparticles is desirable to provide better bioavailability and low toxicity [23]. Polydispersity leads to a larger size distribution, which interferes with the tendency for the nanocarrier to accumulate in the target tissue [23].

Polydispersity describes the degree of non-uniformity of size distribution between molecules due to the aggregation or agglomeration of the polymeric network. It can be estimated using the polydispersity index (PDI), where the ideal index for chitosan nanoparticles is below 0.3 [22, 23]. The degree of deacetylation and molecular weight of chitosan have been proven to influence the polydispersity of the system [24].

PDI increases with an increase in molecular weight. However, it decreases as the degree of deacetylation increases [23]. The increase of amino group protonation and removal of the acetyl group from chitosan structure lead to the enhancement of repulsive forces between molecules and stretch the chitosan to become larger in size [11, 24]. Therefore, the development of chitosan with the optimum degree of deacetylation is needed to minimise the risk of polydispersity. This can be achieved by modifying the time and temperature of the de-*N*-acetylation process [18].

The degree of entanglement for HMWC nanoparticles is higher than LMWC. Therefore, HMWC nanoparticle has a high tendency to aggregate with each other and disrupt the uniformity of the system [25]. However, LMWC cannot be loaded into nanoparticles with smaller size due to its limitation to entangle to the structures of the system [26]. Therefore, maintaining the particle size of the chitosan is crucial in the development of a chitosan nanoparticle.

## **1.3 Modification of chitosan as biomaterial**

Modified chitosan shows greater advantages as compared to unmodified chitosan. The modification of chitosan either chemically or physically may improve its solubility, properties of gelling, and biocompatibility. This modification can be done through cross-linking or substitution [27]. The presence of the various reactive functional groups in the chitosan structure makes it available in many derivatives with different stability properties.

#### *1.3.1 Quaternisation*

A quaternary ammonium salt is a hydrophilic group with a permanent positive charge. Therefore, quaternary chitosan does not need an acidic condition to undergo protonation [12, 28]. It allows chitosan quaternary ammonium salts to be soluble in both acidic and basic pH. This is a good approach to increase chitosan solubility in water [28].

The high strength of the positive charge will weaken the hydrogen bond. However, this activity depends on the degree of substitution. The higher the degree of substitution, the higher the water solubility of chitosan. This will improve the quality of chitosan to act as a mucoadhesive agent that aids the penetration into mucus [29].

Trimethyl chitosan (TMC) is an example of a chitosan derivative from quaternisation. This modification is effective in enhancing the bioavailability of antibacterial drugs with antibacterial properties. Moreover, quaternised chitosan also exhibits antibacterial properties by the interaction of its positive charge with the negative charge of Gram-negative bacteria [28–30].

#### *1.3.2 Sulfonated chitosan*

Sulfonated chitosan is water-soluble anionic chitosan, which was derived with N-benzyl disulfonated derivative [31]. This modification of chitosan has been shown to be effective, not only as antiviral and antibacterial but also as anticoagulant properties. Sulfonated chitosan interferes with the interaction between the envelope glycoprotein (gp120) and its receptor on the CD4 cells' surface. Therefore, It inhibits the replication of HIV [32].

Sulfonated chitosan has been developed to carry anticoagulant drugs such as reviparin and enhance the anticoagulant activity. Sulfonated chitosan nanoparticles interact with factor Xa and inhibit their function in the blood clotting mechanism [31, 33]. Low molecular weight sulfoethyl chitosan acts as capping of nanoparticles [33]. A capping agent is needed in the nanoparticulate system to prevent agglomeration.

Amphotericin B is used to treat fungal infection by binding to ergosterol on the cell membrane of fungal. It depolarises the membrane and alters its permeability [34]. Sulfonated chitosan has been used in the formulation of amphotericin B to reduce the side effects of the drug by making sure the drug specifically targets the ergosterol of fungal [35].

#### *1.3.3 Thiolation*

The structure of mucin that coats the intestinal epithelial cell contains the cysteine-rich domain. This domain easily forms a disulfide bond with a thiolated derivative of chitosan. The bond formations increase the residence time for the chitosan to the mucus and increase the mucoadhesive property of the chitosan [30, 36]. When chitosan covalently bonded with any thiolated moiety, watersoluble carbodiimide is required as a cross-linker [30]. Carbodiimide increases the number of thiol groups. This enhances the immobilisation phenomena, which is the formation of disulfide bonds due to the activation of carboxylic groups.

The thiolation of TMC with the conjugation of cysteine residue increases the strength of covalent bonding between mucin. The covalent bond formation of chitosan with thioglycolic acid (TGA) is an effective carrier in delivering trimethoprim for urinary tract infection [37]. The preparation of chitosan-TGA nanoparticles should be stabilised by covalent cross-linking with polyanion, such as tripolyphosphate [38]. The cross-linking minimise the risk of particle aggregation, increases the disulfide bond, and strengthens the mechanical force between networks, which allows trimethoprim to be released slowly [37].

The conjugation of chitosan with glutathione will protect peptide drugs from aminopeptidase in the GIT [39]. Glutathione has thiol groups which exhibit strong electron-donating properties. It forms an α-peptide bond with cysteine moiety of aminopeptidase [30]. Glutathione also acts as an antioxidant which reduces oxidative stress and increases the adhesion of formulation to the cell [30].

#### *1.3.4 Carboxy alkyl chitosan*

The poor water solubility of chitosan makes them less effective as a permeation enhancer. The addition of the carboxyalkyl group will transform the molecule into amphoteric in nature and allow them to react in both basic and acidic conditions [40]. The interaction between the carboxyl group and the primary amino group of chitosan exhibits a promising approach in developing controlled drug release.

The Schiff base reactive gives rise to the formation of the N-carboxymethyl derivative of chitosan [41]. This modification of chitosan has been shown to improve

**57**

**Figure 3.**

*Chitosan-Based Oral Drug Delivery System for Peptide, Protein and Vaccine Delivery*

the absorption of low molecular weight heparin (LMWH). After coating into polydopamine, the conjugation of LMWH with carboxymethyl chitosan into polyurethane substrate shows excellent hemocompatibility of heparin. This modification enhances the bioavailability and improves the anticoagulant effect of heparin [42]. Propranolol hydrochloride has a short half-life and requires every 6 to 8 hours in a divided daily dose. The use of carboxymethyl chitosan will coat the drug with a polymer matrix. It controls the release of drug with zero-order kinetics, allowing the constant amount of drug will be eliminated per unit time. The hydration of carboxymethyl chitosan will form a gel layered around the drug, which essential in

The release of drugs from the chitosan nanoparticle is influenced by the hydrophilicity of chitosan and pH of the swelling solution. Chitosan release mechanism involves swelling, diffusion of drugs through the polymeric matrix and polymer erosion [45]. Due to the hydrophilicity of chitosan, chitosan nanoparticles exhibit

Acid and base act as catalysts in the degradation of polymers [46]. Therefore, the behaviour of swelling and the amount of drug released is highly dependent on the pH of the swelling solution. Hence, a modified drug release can be achieved [46]. When polymers get into contact with an aqueous medium, the water will diffuse

*DOI: http://dx.doi.org/10.5772/intechopen.95771*

**1.4 Release mechanisms of chitosan nanoparticle**

pH-dependent drug and controlled drug release system [6].

into the polymer until the polymer swells (**Figure 3**).

*The swelling of swellablepolymers in aqueous medium (adapted from [49]).*

drug release [43, 44].

*Chitosan-Based Oral Drug Delivery System for Peptide, Protein and Vaccine Delivery DOI: http://dx.doi.org/10.5772/intechopen.95771*

the absorption of low molecular weight heparin (LMWH). After coating into polydopamine, the conjugation of LMWH with carboxymethyl chitosan into polyurethane substrate shows excellent hemocompatibility of heparin. This modification enhances the bioavailability and improves the anticoagulant effect of heparin [42].

Propranolol hydrochloride has a short half-life and requires every 6 to 8 hours in a divided daily dose. The use of carboxymethyl chitosan will coat the drug with a polymer matrix. It controls the release of drug with zero-order kinetics, allowing the constant amount of drug will be eliminated per unit time. The hydration of carboxymethyl chitosan will form a gel layered around the drug, which essential in drug release [43, 44].

#### **1.4 Release mechanisms of chitosan nanoparticle**

The release of drugs from the chitosan nanoparticle is influenced by the hydrophilicity of chitosan and pH of the swelling solution. Chitosan release mechanism involves swelling, diffusion of drugs through the polymeric matrix and polymer erosion [45]. Due to the hydrophilicity of chitosan, chitosan nanoparticles exhibit pH-dependent drug and controlled drug release system [6].

Acid and base act as catalysts in the degradation of polymers [46]. Therefore, the behaviour of swelling and the amount of drug released is highly dependent on the pH of the swelling solution. Hence, a modified drug release can be achieved [46]. When polymers get into contact with an aqueous medium, the water will diffuse into the polymer until the polymer swells (**Figure 3**).

**Figure 3.** *The swelling of swellablepolymers in aqueous medium (adapted from [49]).*

*Chitin and Chitosan - Physicochemical Properties and Industrial Applications*

Sulfonated chitosan is water-soluble anionic chitosan, which was derived with N-benzyl disulfonated derivative [31]. This modification of chitosan has been shown to be effective, not only as antiviral and antibacterial but also as anticoagulant properties. Sulfonated chitosan interferes with the interaction between the envelope glycoprotein (gp120) and its receptor on the CD4 cells' surface. Therefore,

Sulfonated chitosan has been developed to carry anticoagulant drugs such as reviparin and enhance the anticoagulant activity. Sulfonated chitosan nanoparticles interact with factor Xa and inhibit their function in the blood clotting mechanism [31, 33]. Low molecular weight sulfoethyl chitosan acts as capping of nanoparticles [33]. A capping agent is needed in the nanoparticulate system to prevent

Amphotericin B is used to treat fungal infection by binding to ergosterol on the cell membrane of fungal. It depolarises the membrane and alters its permeability [34]. Sulfonated chitosan has been used in the formulation of amphotericin B to reduce the side effects of the drug by making sure the drug specifically targets the

The structure of mucin that coats the intestinal epithelial cell contains the cysteine-rich domain. This domain easily forms a disulfide bond with a thiolated derivative of chitosan. The bond formations increase the residence time for the chitosan to the mucus and increase the mucoadhesive property of the chitosan [30, 36]. When chitosan covalently bonded with any thiolated moiety, watersoluble carbodiimide is required as a cross-linker [30]. Carbodiimide increases the number of thiol groups. This enhances the immobilisation phenomena, which is the formation of disulfide bonds due to the activation of carboxylic groups. The thiolation of TMC with the conjugation of cysteine residue increases the strength of covalent bonding between mucin. The covalent bond formation of chitosan with thioglycolic acid (TGA) is an effective carrier in delivering trimethoprim for urinary tract infection [37]. The preparation of chitosan-TGA nanoparticles should be stabilised by covalent cross-linking with polyanion, such as tripolyphosphate [38]. The cross-linking minimise the risk of particle aggregation, increases the disulfide bond, and strengthens the mechanical force between

The conjugation of chitosan with glutathione will protect peptide drugs from aminopeptidase in the GIT [39]. Glutathione has thiol groups which exhibit strong electron-donating properties. It forms an α-peptide bond with cysteine moiety of aminopeptidase [30]. Glutathione also acts as an antioxidant which reduces oxidative

The poor water solubility of chitosan makes them less effective as a permeation enhancer. The addition of the carboxyalkyl group will transform the molecule into amphoteric in nature and allow them to react in both basic and acidic conditions [40]. The interaction between the carboxyl group and the primary amino group of chitosan exhibits a promising approach in developing controlled drug release. The Schiff base reactive gives rise to the formation of the N-carboxymethyl derivative of chitosan [41]. This modification of chitosan has been shown to improve

networks, which allows trimethoprim to be released slowly [37].

stress and increases the adhesion of formulation to the cell [30].

*1.3.2 Sulfonated chitosan*

agglomeration.

*1.3.3 Thiolation*

ergosterol of fungal [35].

*1.3.4 Carboxy alkyl chitosan*

It inhibits the replication of HIV [32].

**56**

The polymeric chitosan chain will start to detangle. The swelling polymer will form pores which allow drugs to diffuse out of the nanoparticulate system [6, 43, 47]. Therefore, the water solubility of chitosan is crucial in the mechanism of drug release from the nanoparticulate system.

#### **1.5 The use of chitosan to improve drug delivery system**

Oral drug administration is the most convenient route, especially among the elderly and children. Unfortunately, some drugs and vaccines cannot withstand the physiological barrier of GIT. In the presence of mucus, proteolytic enzymes, and first-pass metabolism by the liver, drugs tend to be degraded or converted into inactive metabolites [48]. Some drugs will be excreted in the urine lead to low bioavailability.

Due to the challenges aforesaid, chitosan and its derivatives have been used in the development of nanotechnology to improve oral drug delivery [25, 30]. It encapsulates drugs to protect them from degradation in the GIT environment. As a consequence of its excellent biodegradable, biocompatibility, and non-toxic properties, chitosan promotes a stimuli-responsive release of drugs. It allows active ingredients to be released from the formulation in a controlled manner, specifically in enteric-coated drugs [38, 43].

Due to its antimicrobial properties, it was used in the delivery of oral antibiotics to eradicate Gram-negative bacteria such as *E. coli* [49]. This approach not only improves the bioavailability of antibiotic in the body but also indirectly enhance the effectiveness of the drugs in eradicating the infection [15].
