INTRODUCTION:

Polymer nanoparticles are particles of less than 1 mm diameter that are prepared from natural or synthetic polymers. Nanoparticles have become an important area of research in the field of drug delivery because they have been extensively used to deliver drugs, genes, diagnostics, and vaccines into specific cells or tissues. The strategy of using nanoparticles as a carrier system for drug and gene delivery has attracted increasing interest. The main target of many pharmaceutical delivery systems is to deliver the drug to the specific cell types and is successful only when the drug through its delivery vehicle is internalized into cells. Owing to their small size, prolonged circulation time, and sustained drug release profile, nano-sized polymeric nanoparticles bearing drugs have received an increasing amount of attention for their ability to improve the efficacy of potent drugs. It has been reported that nano-sized drug carriers composed of natural and synthetic polymers maintain a prolonged circulation time in the body by avoiding the reticuloendothelial system (RES), as such reduced liver and spleen uptake has been exploited in cancer therapies¹.

Recently, polymer nanoparticles have been widely investigated as a carrier for drug delivery. Among them, much attention has been paid to the nanoparticles made of biodegradable polymers such as chitosan (CS) due to its good biocompatibility, biodegradability, and novel drug release behavior. Chitosan nanoparticles are potential delivery systems for vaccines, genes, and anticancer agents.

It has been reported that chitosan nanoparticles are having little particle size and enhanced zeta potential. Chitosan is a polysaccharide, similar in structure to cellulose. Both are made by linear h-(1Y4)-linked monosaccharides^2-7. However, an important difference to cellulose is that chitosan is composed of 2-amino-2-deoxy-h-d-glucan combined with glycosidic linkages. The primary amine groups render special properties that make chitosan very useful in pharmaceutical applications. Compared to many other natural polymers, chitosan has a positive charge and is mucoadhesive. Therefore, it is used extensively in drug delivery applications. Chitosan is obtained from the deacetylation of chitin, a naturally occurring and abundantly available (in marine crustaceans) biocompatible polysaccharide. However, applications of chitin are limited compared to chitosan because chitin is structurally similar to cellulose, but chemically inert. Acetamide group of chitin can be converted into amino group to give chitosan, which is carried out by treating chitin with concentrated alkali solution. Chitin and chitosan represent long-chain polymers having molecular mass up to several million Daltons. Chitosan is relatively reactive and can be produced in various forms such as powder, paste, film, fiber, etc^8-11.

Commercially available CS has an average molecular weight ranging between 3800 and 20,000 Daltons and is 66% to 95% deacetylated. Chitosan, being a cationic polysaccharide in neutral or basic pH conditions, contains free amino groups and hence, is insoluble in water. In acidic pH, amino groups can undergo protonation thus, making it soluble in water. Solubility of CS depends upon the distribution of free amino and N-acetyl groups. Usually 1-3 % aqueous acetic acid solutions are used to solubilize CS. Chitosan is biocompatible with living tissues since it does not cause allergic reactions and rejection. It breaks down slowly to harmless products (amino sugars), which are completely absorbed by the human body. Chitosan degrades under the action of ferments, it is nontoxic and easily removable from the organism without causing concurrent side reactions^12,13. It possesses antimicrobial property and absorbs toxic metals like mercury, cadmium, lead, etc. In addition, it has good adhesion, coagulation ability, and immunostimulating activity. If degree of deacetylation and molecular weight of CS can be controlled, then it would be a material of choice for developing micro/nanoparticles. Chitosan has many advantages, particularly for developing micro/nanoparticles. These include: its ability to control the release of active agents, it avoids the use of hazardous organic solvents while fabricating particles since it is soluble in aqueous acidic solution, it is a linear polyamine containing a number of free amine groups that are readily available for crosslinking, its cationic nature allows for ionic crosslinking with multivalent anions, it has mucoadhesive^14-17character, which increases residual time at the site of absorption, and so on. Chitin and CS have very low toxicity; LD50 of CS in laboratory mice is 16 g/kg body weight, which is close to sugar or salt. Chitosan is proven to be safe in rats up 10% in the diet. Various sterilization methods such as ionizing radiation, heat, steam and chemical methods can be suitably adopted for sterilization of CS in clinical applications. In view of the above-mentioned properties, CS is extensively used in developing drug delivery systems. Particularly, CS has been used in the preparation of mucoadhesive formulations improving the dissolution rate of the poorly soluble drugs drug targeting and enhancement of peptide absorption. However, the micro/nanoparticulate drug delivery systems offer numerous advantages over the conventional dosage forms. These include improved efficacy, reduced toxicity and improved patient compliance. The present review addresses the preparation of chitosan nanoparticles by ionotropic gelation method^18-21.

Fig 1. Schematic representation of preparation of chitosan nanoparticles by ionotropic gelation method

Methods of preparation of chitosan nanoparticles

Different methods such as ionotropic gelation, emulsion cross-linking, nanoprecipitation, salting out etc have been used to prepare CS particulate systems. Selection of any of the methods depends upon factors such as particle size requirement, thermal and chemical stability of the active agent, reproducibility of the release kinetic profiles, stability of the final product and residual toxicity associated with the final product. Since we are concerned only with the ionotropic gelation method, we will restrict our discussions only on these aspects.

Ionotropic Gelation Method

The use of complexation between oppositely charged macromolecules to prepare CS nanoparticles has attracted much attention because the process is very simple and mild. In addition, reversible physical cross-linking by electrostatic interaction, instead of chemical cross-linking, has been applied to avoid the possible toxicity of reagents and other undesirable effects. Tripolyphosphate (TPP) is a polyanion, which can interact with the cationic CS by electrostatic forces. After Bodmeier et al, reported the preparation of TPP–CS complex by dropping CS droplets into a TPP solution, many researchers have explored its potential pharmaceutical usage. In the ionic gelation method, CS is dissolved in aqueous acidic solution to obtain the cation of CS. This solution is then added dropwise under constant stirring to polyanionic TPP solution. The chitosan molecules has abundant NH₃group which can react with negatively charged phosphoric ions of TPP to form cross-linked chitosan nanoparticles. During the process of cross-linking and hardening process water was extruded from the particles, which may help in sustaining the release of drug. Three kinds of phenomena were observed: solution, aggregation and opalescent suspension while preparing the nanoparticles. The last stage indicates the completion of the process. Insulin-loaded CS nanoparticles have been prepared by mixing insulin with TPP solution and then adding this to CS solution under constant stirring. Two types of CS in the form of hydrochloride salt (SeacureR 210 Cl and ProtasanR 110 Cl), varying in their molecular weight and degree of deacetylation, were utilized for nanoparticle preparation. For both types of CS, TPP concentration was adjusted to get a CS/TPP ratio of 3.6:1. Chitosan nanoparticles thus obtained were in the size range of 300–390 nm with a positive surface charge ranging from +34 to +45 mV. Using this method, insulin loading was modulated reaching the values up to 55%. Efficiency of the method was dependent upon the deacetylation of CS, since it involves the gelation of protonated amino groups of CS^1,18,22,23.

There are many ongoing investigations, which demonstrate the improved oral bioavailability of peptide and protein formulations. Bioadhesive polysaccharide CS nanoparticles would seem to further enhance their intestinal absorption. Yan et al. prepared the insulin-loaded CS nanoparticles by ionotropic gelation of CS with TPP anions. Particle size distribution and zeta potential were determined by photon correlation spectroscopy. The ability of CS nanoparticles to enhance the intestinal absorption of insulin and the relative pharmacological bioavailability of insulin was investigated by monitoring the plasma glucose level of alloxan-induced diabetic rats after the oral administration of various doses of insulin-loaded CS nanoparticles. The positively charged, stable CS nanoparticles showed particle size in the range of 250–400 nm. Insulin association was up to 80%. The in vitro release experiments indicated initial burst effect, which is pH-sensitive. The CS nanoparticles enhanced the intestinal absorption of insulin to a greater extent than the aqueous solution of CS in vivo. After administration of 21 I.U./kg insulin in the CS nanoparticles, hypoglycemia was prolonged over 15 h. The average pharmacological bioavailability relative to s.c. injection of insulin solution was up to 14.9%^2,4,6,8.

Fig 2. Interactions of chitosan with TPP (A) Deprotonation, (B) Ionic cross linking

Lifeng Qi et al. have prepared the chitosan nanoparticles by ionotropic gelation method and also evaluated the antibacterial activity of chitosan nanoparticles. They have also characterized the particles by SEM,AFM and their MIC value were less than 0.25 μg/ml, and the MBC values of nanoparticles reached 1 μg/ml .AFM revealed that the exposure of S.choleraesuis to the chitosan nanoparticles lead to the disruption to the cell membranes and the leakage of cytoplasm²¹.

Zengshuan Ma et al. have prepared chitosan insulin nanoparticles by ionotropic gelation of chitosan with TPP anions. The ability of chitosan nanoparticles to enhance the intestinal absorption. The relative pharmacological activity and bioavailability of insulin were investigated by monitoring the plasma glucose level of administration of various dose of insulin loaded chitosan nanoparticles²⁴.

Fig 3. Chitosan nanoparticles prepared by ionotropic gelation method

Physicochemical, Biological and Pharmacological Properties of

Chitosan

Physical Properties: Particle Size 30mm

Density 1.35-1.40 g/cc

pH 6.5-7.5

Solubility Insoluble in water but soluble in acids

Chemical properties

o Cationic Polyamine

o High charge density at pH 6.6

o Adheres to negatively charge surfaces

o Forms gels with polyanions

o High molecular weight, linear polyelectrolyte

o Viscosity- high to low chelates certain transitional metals

o Reactive hydroxyl/ amino group

Biological Properties

o Nontoxicity

o Biocompatibility

o Biodegradability

Pharmacological Properties

o Hypocholesterolemic action

o Wound-healing properties

o Antacid and antiulcer activity

o Antifungal and antibacterial activity

Table 1. Properties of chitosan

Advantages of ionotropic gelation method^1,5,7

· The method is very economic and simple

· The method requires less equipment and time

· In addition, reversible physical cross-linking by electrostatic interaction, instead of chemical cross-linking, has been applied to avoid the possible toxicity of reagents and other undesirable effects.

· No use of organic solvent.

Disadvantages of ionotropic gelation method⁷

The only disadvantage of TPP/CS nanoparticles is their poor mechanical strength

CONCLUSION:

Chitosan has the desired properties for safe use as a pharmaceutical excipient. This has prompted accelerated research activities worldwide on chitosan micro and nanoparticles as drug delivery vehicles. These systems have great utility in controlled release and targeting studies of almost all class of bioactive molecules. Recently, chitosan is also extensively explored in gene delivery and ionotropic gelation method is one of the best and simple methods to prepare chitosan nanoparticles.

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Received on 10.11.2010 Modified on 04.01.2011

Research J. Pharm. and Tech. 4(4): April 2011; Page 492-495