In vitro cytotoxicity assay on venom of highly venomous snakes in Sri Lanka
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Date
2019-09-12
Authors
Edirisinghe, L. E. A. D. A.
Karunathilake, K. J. K.
Dissanayake, D. A.
Rathnayaka, R. M. M. K. N.
Kularatne, S. A. M.
Rajapakse, R. P. V. J.
Journal Title
Journal ISSN
Volume Title
Publisher
University of Peradeniya
Abstract
Highly venomous snakes cause significant morbidity and mortality in Sri Lanka. Indian polyvalent antivenom currently used as treatment for snake envenomation is sometimes unsuccessful due to its non-specificity to snake venoms of Sri Lanka. Therefore, venom toxicity studies on highly venomous snakes of Sri Lanka are important. The protein concentration and IC-50 of venom extracted from snakes of Sri Lanka (Bungarus caeruleus, Daboia russelii, Echis carinatus, Hypnale hypnale, Naja naja and Trimeresurus trigonocephalus) were determined. The protein concentrations of the crude venom were determined by bicinchoninic acid protein assay and the IC-50 was determined by MTT cytotoxicity assay using L929 mouse fibroblast cell lines. Protein electrophoresis using SDS-PAGE was performed to determine the most prominent protein bands in each venom. The protein concentrations were: B. caeruleus 4213.9 mg/ml, D. russelii 3905.6 mg/ml, E. carinatus 2594.4 mg/ml, H. hypnale 3655.6 mg/ml, H. nepa 2822.2 mg/ml, H. zara 3333.3 mg/ml, N. naja 4055.6 mg/ml, and T. trigonocephalus 1461.1 mg/ml. The IC-50 values obtained were: B. caeruleus 4.9 mg/ml, D. russelii 10.5 mg/ml, E. carinatus 42.1 mg/ml, H. hypnale 16.8 mg/ml, N. naja 1.2 mg/ml, and T. trigonocephalus: 13.8 mg/ml. The most prominent protein bands observed were: B. caeruleus 086, 070, 061, 028, 020, 016, 011, 009 kDa; D. russelii 113, 041, 033, 021, 013 kDa; E. carinatus 086, 080, 070, 061, 056, 040, 035, 030, 024, 017, 014, 012 kDa; H. hypnale 069, 064, 055, 029, 013 kDa; H. nepa 070, 065, 061, 053, 046, 043, 030, 028, 023, 013 kDa; H. zara - 070, 053, 037, 030, 013 kDa; and N. naja 113, 097, 084, 068, 013 kDa. The highest and lowest cytotoxicities were observed in venom of N. naja and E. carinatus, respectively. This preliminary study is useful for developing an effective antivenom specific to venomous snakes of Sri Lanka.
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Keywords
Snake venom , In vitro cytotoxicity , IC-50 , Highly venomous snakes , Antivenom