Characterization of chito-oligosaccharides prepared by chitosanolysis with the aid of papain and Pronase, and their bactericidal action against Bacillus cereus and Escherichia coli

Acharya, B. Vishu Kumar and Varadaraj, M. C. and Lalitha, R. Gowda and Tharanathan, R. N. (2005) Characterization of chito-oligosaccharides prepared by chitosanolysis with the aid of papain and Pronase, and their bactericidal action against Bacillus cereus and Escherichia coli. Biochemical Journal, 391 (2). pp. 167-175.

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Abstract

<p align="justify">Papain (from papaya latex; EC 3.4.22.2) and Pronase (from Streptomyces griseus; EC 3.4.24.31) caused optimum depoly-merization of chitosan at pH 3.5 and 37 °C, resulting in LMMC (low molecular mass chitosan) and chito-oligomeric-monomeric mixture. The yield of the latter was 14-16% and 14-19% re-spectively for papain- and Pronase-calalysed reactions, depending on the reaction time (1-5 h). HPLC revealed the presence of monomer(s) and oligomers of DP (degree of polymerization) 2-6, which was also confirmed by matrix-assisted laser-desorption ionization-time-of-llighl MS. Along with the chito-oligomers, the appearance of only GlcNAc (N-acetylglucosamine) in Pronase-catalysed chitosanolysis was indicative of its different action pattern compared with papain. Fourier-transform infrared, liquid-state l3C-NMR spectra and CD analyses of chito-oligomeric-monomeric mixture indicated the release of GlcNAc/GlcNAc-rich oligomers. The monomeric sequence at the non-reducing ends of chito-oligomers was elucidated using N-acelylglucosaminidase. The chito-oligomeric-monomeric mixture showed better growth inhibitory activity towards Bacillus cereus and Escherichia coli compared with native chitosan. Optimum growth inhibition was observed with chito-oligomers of higher DP having low degree of acetylation. The latter caused pore formation and permeabiliz-ation of the cell wall of B. cereus, whereas blockage of nutrient flow due to the aggregation of chito-oligomers-monomers was responsible for the growth inhibition and lysis of E. coli, which were evidenced by scanning electron microscopy analysis. The spillage of cytoplasmic enzymes and native PAGE of the cell-free supernatant of B. cereus treated with chito-oligomeric-monomeric mixture further confirmed bactericidal activity of the latter. Use of papain and Pronase, which are inexpensive and easily available, for chitosanolysis, is of commercial importance, as the products released are of considerable biomedical value.</p>

Item Type:	Article
Uncontrolled Keywords:	bactericidal activity chitosan chitosanolysis oligomer papain Pronase
Subjects:	600 Technology > 08 Food technology > 11 Food Biochemistry 500 Natural Sciences and Mathematics > 07 Life Sciences > 03 Biochemistry & Molecular Biology > 07 Enzyme Biochemistry 500 Natural Sciences and Mathematics > 07 Life Sciences > 03 Biochemistry & Molecular Biology > 18 Phytochemistry
Divisions:	Dept. of Biochemistry
Depositing User:	Food Sci. & Technol. Information Services
Date Deposited:	29 Dec 2005
Last Modified:	16 May 2012 06:47
URI:	http://ir.cftri.res.in/id/eprint/284

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