Associative pullulan gels and their interaction with biological active substances

doi:10.1016/S0168-3659(02)00169-4

Journal of Controlled Release

Volume 83, Issue 1, 18 September 2002, Pages 41-51

https://doi.org/10.1016/S0168-3659(02)00169-4 Get rights and content

Abstract

This paper studies the synthesis and properties of anionic and/or amphiphilic pullulan microparticles crosslinked with epichlorohydrine or with sodium trimethaphosphate. The polysaccharide gels were physicochemically characterized and their interaction with enzymes (lysozyme) was studied with the aim to appreciate the performances for separation/purification/immobilization of the enzymes or controlled release drug systems.

Introduction

Great attention has been paid in the last years to the study of polymeric hydrophilic gels, which represent crosslinked macromolecular networks by chemical or physical methods. This interest is justified by their specific properties which recommend them in biomedical, biotechnological and pharmaceutical applications.

Polysaccharide-based gels are intensively studied as delivery systems for pharmaceutically active peptides and protein drugs [1], [2]. Self-assembled hydrogel nanoparticles of cholesterol-bearing pullulan were tested as carriers of protein drugs — insulin [3].

For the recovery of the proteins carboxymethyl [4] and sulfopropyl [5] cellulose derivatives were used. A great variety of dextran hydrogels were prepared and their interaction with proteins (lysozyme, IgG) was studied [6], [7], [8], [9], [10].

It is well known that the peptides and the proteins orally administered have a low bioavailability, due to their instability in the gastrointestinal tract and to their low permeability through intestinal mucous. The injectable forms, which maintain better the therapeutic effects are poorly complied by patients. For this reason, some alternative forms were developed, i.e. nasal, buccal, transdermal, rectal routes. Some recent studies reported the synthesis of some nanoparticles having hydrophobic backbone and hydrophilic branches as drug carriers for oral peptide delivery [11].

Our paper is devoted to the synthesis of new associative carboxymethyl pullulan microparticles substituted with long chain alkyl-amido groups and to the study of their interaction with enzymes (lysozyme). Two different crosslinking agents were used: epichlorohydrine and comparatively a less used reagent: sodium trimethaphosphate. The latter was used for crosslinking of starch [12], [13], hydroxypropyl starch [14] carboxymethyl starch [15] or guar [16], for colon-specific drug delivery. Studies on the synthesis and physicochemical characteristics of associative derivatives of linear carboxymethyl pullulan or carboxymethyl cellulose were earlier performed [17], [18]. Also, studies on the synthesis and interaction with biological active molecules of hydrophobically modified crosslinked carboxymethyl cellulose were reported in a earlier publication [19].

Section snippets

Materials

•
Polysaccharide: pullulan PI20 Hayashibara, Japan (PL), M_w: 200,000.
•
Lysozyme (M_w: 14,000)—Sigma.
•
Crosslinking agents: epichlorohydrine (ECH), sodium trimethaphosphate (TMP).
•
Reagents: sodium chloroacetate (NaClAc), sodium hydroxide.
•
Solvents: dichloroethane, methanol, dimethyl sulfoxide (DMSO), acetone, ethyl alcohol, isopropanol p.a.

Methods

The following reactions were performed:

1.
Crosslinking of pullulan with epichlorohydrine (sample: PC_ECH) or with TMP (sample: PC_TMP). To 100 ml organic suspension medium

Characterization of polymeric supports

The granulometric distribution in the dry state of a sample of carboxymethyl pullulan crosslinked with TMP is presented in Fig. 1. By maintaining the reaction parameters (amount of the stabilizer, stirrer speed, concentration of polysaccharide solutions) at the same values, the granulometric distribution varies with about ±5%. For the study only the microparticles 0.05–0.125 mm in diameter (dry) are used to assure a satisfactory flow-rate in the chromatographic column applications.

Conclusions

Microparticles of pulullan and carboxymethyl pullulan were obtained by using a new crosslinking agent: sodium trimethaphosphate. These were compared with those obtained through crosslinking with epichlorohydrine. By chemical transformation of some carboxylic groups with aliphatic long chain amine (C16) new associative crosslinked microparticles were obtained. Interaction with the lysozyme of the products was studied with the aim to appreciate their performances in

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Associative pullulan gels and their interaction with biological active substances

Abstract

Introduction

Section snippets

Materials

Methods

Characterization of polymeric supports

Conclusions

Carbohydr. Polymers

J. Controlled Release

J. Controlled Release

Adv. Drug Deliv. Rev.

Carbohydr. Polymers

Carbohydr. Polymers

J. Controlled Release

Int. J. Biol. Macromol.

Carbohydr. Polymers

Eur. Polymer J.

J. Biol. Chem.

Anal. Biochem.

Colloids Surfaces B: Biointerfaces

J. Pharm. Sci.

React. Polym.