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"Biotechnologia Acta" V. 9, No 6, 2016
https://doi.org/10.15407/biotech9.06.039
Р. 39-49, Bibliography 28, English
Universal Decimal Classification:  577.114.083:543.544.153:543.545.2

 6 2016

HONEYBEE (Apis mellifera) CHITOSAN: PURIFICATION, HETEROGENEITY AND HEMOCOAGULATING ACTIVITY

M. D. Lootsik 1, R. O. Bilyy 2, M. M. Lutsyk 2, N. O. Manko 3, S. A. Navytka 3, V. I. Kutsiaba 1, R. S. Stoika 1

1 Institute of Cell Biology of the National Academy of Sciences of Ukraine, Lviv
2 Danylo Halytsky Lviv National Medical University, Ukraine
3 Ivan Franko National University, Lviv, Ukraine

The aim of the research was to elaborate the method of chitosan preparation obtaining from honeybee corpses. It  included the following stages: 1) washing of bee corpses with hot water; 2) delipidation of powdered material with petrol ether; 3) decalcification by EDTA at pH 4; 4) deproteination by 3 fold treatment with 5% NaOH 1 h at 70 oC; 5) bleaching of chitin with sodium hypochlorite; 6) deacetylation of chitin in 40% NaOH solution at 115 oC for 3 h; 7) purification of chitosan by its dissolving in 3% acetic acid and precipitation with ammonia at pH 8,5; 8) separation into three fractions precipitated at pH 6.4, 7.0 and 8.6. The yield of chitosan from dry bee powder was 8.5–10.0. A distinct diversity in the molecular mass of different chitosan fractions was revealed in the range of 80–320 kDa. Heterogeneity of chitosan samples was studied using gel permeation chromatography on Acrylex P-150 and electrophoresis in a slab of polyacrylamide gel with a stepwise gradient of acrylamide concentration 5, 10, 15, 20% in pH 4.5 buffer system. High molecular mass chitosan possessed blood coagulating activity, while low molecular mass fractions were poorly active. The rate of blood clot formation induced by active honeybee chitosan was 3 times lower comparing with that of chitosans obtained from crabs or shrimps.

Ключевые слова: bee subpestilence, chitosan.

© Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, 2016