Research Articles
Formulation pH modulates the interaction of insulin with chitosan nanoparticles

https://doi.org/10.1002/jps.10149Get rights and content

Abstract

Previous studies on chitosan‐insulin nanoparticles have reported diverse encapsulation efficiency and insulin release profiles despite similar formulation and preparation procedures. This study examined the efficiency and mechanism of association of insulin with chitosan nanoparticles in the pH range of 2.3 to 6.3. Nanoparticles of 237 to 235 nm were prepared by ionotropic gelation of chitosan with tripolyphosphate counterions. Insulin was quantified by an RP‐HPLC method. The insulin association efficiency (AE) spanned a broad range from 2 to 85%, and was highly sensitive to formulation pH. Highest AE was measured at insulin loading concentrations ≥ 4.28 U/mL and pH 6.1, close to the pI of native insulin and the pKa of chitosan. This association, attributed to physical adsorption of insulin through hydrophobic interactions with chitosan, was labile, and the associated insulin rapidly and completely released by dilution of the nanoparticles in aqueous media of pH 2 to 7.4. AE obtained at pH 5.3 was less than half that measured at pH 6.1 at corresponding insulin concentration, but the association at pH 5.3 appeared to be based on stronger interactions, because the release of insulin was pH‐dependent and recovery was less than 25% even upon disintegration of the chitosan matrix. Interaction of insulin with the chitosan nanoparticles rendered the protein more susceptible to acid and enzymatic hydrolyses, the effects being more predominant in nanoparticles prepared at pH 5.3 than at pH 6.1.

Section snippets

INTRODUCTION

Chitosan is a polysaccharide derived from chitin by alkaline deacetylation. Generally regarded as biocompatible, biodegradable, and nontoxic, chitosan is an interesting biomaterial because of its ability to mediate the permeation of molecules across absorptive epithelia. It has been reported to enhance the nasal absorption of insulin in rabbits1 and sheep,2 and the mucosal permeation of other drug molecules.3 The mechanism has been ascribed to a combination of mucoadhesion and widening of the

Materials

Chitosan (CS) (MW) 1.86 (± 0.16) × 105, degree of deacetylation 84.71 ± 0.59%, Aldrich Chemical Co., Milwaukee, WI), insulin (from porcine pancreas, 1 mg was equivalent to 27.8 USP units, Sigma Chemical Co., St. Louis, MO), lysozyme (from chicken egg white, 48,800 U/mg, Sigma Chemical Co.), chitosanase‐RD (from Bacillus SP. PI‐7S, 0.15–0.35 U/mg, Pias Corporation, Osaka, Japan) and pentasodium tripolyphosphate (TPP) (Merck, Darmstadt, Germany) were used as received. All organic solvents were of

RESULTS

Chitosan–insulin nanoparticles were prepared in the pH range of 2.3 to 6.3, achieved by careful control of the acidity and alkalinity of the chitosan and TPP solutions, respectively. Nanoparticle formation was effected by mixing the chitosan and TPP solutions. Insulin was premixed with either the TPP solution or the chitosan solution prior to nanoparticle formation, and the resultant nanoparticles are designated as TPP‐np and CS‐np, respectively. The initial insulin loading concentration was

DISCUSSION

The chitosan–insulin nanoparticles were produced by ionotropic gelation with the counterion, TPP. Chitosan, which has an apparent pKa of 6.3, assumes a stiff extended conformation in aqueous media of low pH due to the charge repulsion of highly protonated amino groups, but it becomes increasingly globular as pH increases, and can precipitate at pH 6.6 or lower, depending on the ionic strength of the solution.14 In this study, precipitation was apparent at pH ≥ 6.3, while the particle yield was

CONCLUSION

Chitosan nanoparticles (237 to 325 nm) with insulin association efficiency that varied from 2 to 85% could be prepared by the iontropic gelation method with careful control of the formulation pH from 2.3 to 6.3. The insulin association was very sensitive to formulation pH, and was also dependent on insulin loading concentration. High association (≥ 80%), attributed to physical adsorption predominated by hydrophobic interactions with chitosan, occurred at insulin loading concentrations ≥ 4.28

Acknowledgements

This study is supported by a National University of Singapore grant (R148‐000‐023‐112). Zengshuan Ma is grateful to the National University of Singapore for financial support of his graduate studies.

REFERENCES (25)

  • N.G.M. Schipper et al.

    Chitosan as absorption enhancers for poorly absorbable drugs 2: Mechanism of absorption enhancement

    Pharm Res

    (1997)
  • P. Artursson et al.

    Effect of chitosan on permeability of monolayers of intestinal epithelial cells (Caco‐2)

    Pharm Res

    (1994)
  • Cited by (0)

    View full text