Effect of molecular structure of chitosan on protein delivery properties of chitosan nanoparticles

https://doi.org/10.1016/S0378-5173(02)00548-3Get rights and content

Abstract

Chitosan nanoparticles (CS NP) with various formations were produced based on ionic gelation process of tripolyphosphate (TPP) and chitosan. They were examined with diameter 20–200 nm and spherical shape using TEM. FTIR confirmed tripolyphosphoric groups of TPP linked with ammonium groups of chitosan in the nanoparticles. Factors affecting delivery properties of bovine serum albumin (BSA) as model protein have been tested, they included molecular weight (Mw) and deacetylation degree (DD) of chitosan, the concentration of chitosan and initial BSA, and the presence of polyethylene glycol (PEG) in encapsulation medium. Increasing Mws of chitosan from 10 to 210 kDa, BSA encapsulation efficiency was enhanced about two times, BSA total release in PBS (phosphate buffer saline) pH 7.4 in 8 days was reduced from 73.9 to 17.6%. Increasing DD from 75.5 to 92% promoted slightly the encapsulation efficiency and decelerated the release rate. The encapsulation efficiency was highly decreased by increase of initial BSA and chitosan concentration; higher loading capacity of BSA speeded the BSA release from the nanoparticles. Adding PEG hindered the BSA encapsulation and accelerated the release rate.

Introduction

The hydrophilic nanoparticles have received much attention to deliver therapeutic peptide, protein, antigen, oligonucleotide and gene by intravenous, oral, and mucosal administration (Janes et al., 2001a). The information has emphasized the importance of size, and revealed the advantages of nanoparticles over the microspheres (Meclean et al., 1998), it has been observed that the number of nanoparticles that cross the epithelium is greater than the number of microspheres. Chitosan is a biodegradable and bioadhesive polysaccharide. It has been shown that chitosan is non-toxic and soft tissue compatible in a range of toxicity tests (Aspden et al., 1997). It has been widely used in pharmaceutical research and in industry as a carrier for drug delivery and as biomedical material (Mao et al., 2001). Chitosan was selected for nanoparticles because of its recognized mucoadhesivity and ability to enhance the penetration of large molecules across mucosal surface (Illum, 1998).

More recently, researches have attempted to study chitosan nanoparticles as follows: preparation, modification, properties of loading various drugs and their physiological characters, such as chitosan nanoparticles coated PLGA (Vila et al., 2002) and PEO-PPO (Calvo et al., 1997), nanoparticles loaded insulin (Fernandez-Urrusuno et al., 1999), DNA (Mao et al., 2001) and anticancer drug doxorubicin (Janes et al., 2001b). But some important factors affecting drug properties have not always been investigated, such as basic molecular parameters of chitosan, molecular weight (Mw) and deacetylation degree (DD), were seldom evaluated in protein delivery system of nanoparticles. Janes stated the Mws of chitosan might have a role in the protein or peptide release in the review about chitosan nanoparticles as delivery systems for macromolecules (Janes et al., 2001a), but the relative paper has not been reported. The introduction of a second ingredient may increase their versatility in terms of the encapsulation and delivery of proteins and their susceptibility to interact with biological surface. Polyethylene glycol (PEG) coated nanoparticles have been found to be important potential therapeutic application for controlled release of drugs and site-specific drug delivery (Quellec et al., 1998). Few studies have attempted to investigate chitosan nanoparticles coated with PEG. Therefore, we investigate a series of factors affecting delivery properties, the influence of Mw and DD of chitosan, concentration of chitosan and initial bovine serum albumin (BSA), and PEG introduction are all evaluated.

The purpose of the current study was to examine the influence of a number of factors on the encapsulation of a model protein BSA and release properties during incubation in phosphate buffer saline (PBS) of pH 7.4, and investigate the physicochemical structure of nanoparticles by FTIR and TEM. A series of chitosan nanoparticles with various molecular parameters were prepared, effect of Mw and DD of chitosan on protein delivery was studied systematically. Effect of initial BSA and chitosan concentration and PEG presence was also evaluated. Thus, we can modulate their encapsulation capability and release rate by adjusting the molecular and formation parameters.

Section snippets

Material

Chitosan (CS) from a shrimp shell was purchased from Yuhuan Ocean Biochemical Co. (Zhejiang, China), DD was 92%, and Mw was 210 kDa. Chitosan of DD 92% with different Mws (115, 80, 48, 10 kDa) and chitosan of Mw 210 kDa with different DDs (75.2, 85.5, 92%) were prepared according the reference (Qin et al., 2002, George and Frances, 2001). The Mws were measured by a gel permeation chromatography (GPC), the DDs were determined by elemental analysis. BSA with Mw 68 kDa, PEG with Mw 20 kDa, and

Physicochemical characterizations of nanoparticles

TEM of the nanoparticles and their surface morphology are shown in Fig. 1, they are about 20 nm in size and spherical in shape. Compared with chitosan nanoparticles, the surfaces of nanoparticles contained PEG exhibits fluffy. It has been previously reported that the incorporation of PEG in gel system is through intermolecular hydrogen bonding between the electro-positive amino hydrogen of chitosan and the electro negative oxygen atom of PEG, thus forming a CS/PEG semi-interpenetration network (

Conclusion

In the present study, the physicochemical structure of chitosan nanoparticles is different from matrix chitosan, inter and intramolecular action is enhanced due to tripolyphosphoric groups of TPP gelation with ammonium groups of chitosan. Formation and molecular parameters of chitosan nanoparticles play an important role in protein delivery. Altering concentration of BSA from 0.2 to 2 mg/ml and chitosan from 3 to 1 mg/ml enhance significantly encapsulation capacity of BSA. Nanoparticles

Acknowledgements

We gratefully acknowledge financial support from the National Natural Science Foundation of China.

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