Indian Black Scorpion (Heterome- trus bengalensis) Venom Action Neutralization by Indian Medici- nal Plants in Experimental Ani- mals

The anti scorpion venom activity of the Indian medicinal plant (Hemidesmus indicus, Pluchea indica and Aristolochia indica) root extracts (aqueous and methanol) was established in experimental animal models. Adult black scorpions (Heterometrous bengalensis) of both sexes were collected and the Scorpion Venom (SV) was collected by electrical stimulation, pooled, lyophilized and stored at 4 °C. Scorpion venom was expressed in terms of dry weight. Plant root extracts of Hemidesmus indicus (H.I) Pluchea indica (P.I) and Aristolochia indica (A.I) were (aqueous/methanol) prepared and expressed in terms of dry weight. Anti scorpion venom activity was evaluated using various in vivo and in vitro models (lethality, edema, urinary changes, plasma recalcification, cardiotoxicity, neurotoxicity, etc.). The aqueous/ methanol plant root extracts (H.I, P.I and A.I) could neutralize scorpion venom induced lethality, edema, urinary changes, plasma recalcification, cardiotoxicity, neurotoxicity, in experimental animal models. The aqueous root extracts were more effective in neutralizing scorpion venom actions as compared to methanol plant root extracts. These observations confirmed the ethnopharmacological value of the Indian medicinal plants active against scorpion venom and warrants further detailed studies. Figure 1: Indian black scorpion Heterometrus bengalensis. Citation: Das R, Ghosh S, Gomes A. Indian Black Scorpion (Heterometrus bengalensis) Venom Action Neutralization by Indian Medicinal Plants in Experimental Animals. J Toxins. 2016;3(2): 7 J Toxins 3(2): 7 (2016) Page 02 ISSN: 2328-1723 otherwise mentioned. Petroleum ether, Chloroform, Methanol (Spectrochem, India), Potassium Chloride (SRL, India), Calcium Chloride (Merck, India), Magnesium Chloride (Qualigen, India), Sodium bicarbonate (SRL, India), Sodium biphosphate (SRL, India), Glucose (Qualigen, India). Collection of scorpion and scorpion venom Adult black scorpions (Heterometrous bengalensis) of both sexes were collected from Burdwan district of West Bengal, India, during the rainy seasons (June-September) and were kept in a wire mesh cage (Figure 1). They were provided with food (live cockroach, an egg), water ad libitum. The Scorpion Venom (SV) was collected once in a month by applying square wave electrical stimulation (15 V, 1 ms) to the telson. The venom was pooled, lyophilized and stored at 4 °C in amber colour bottle, until further use. Before use, SV was weighed, dissolve in 0.9% saline/ phosphate buffer 0.01 M, pH 7.2 and was expressed in terms of dry weight. Collection of plant materials The plant (Hemidesmus indicus, Pluchea indica and Aristolochia indica) roots were collected commercially from M/s United Chemicals and Allied Products, Kolkata and were identified by Prof N. Paria, Department of Botany, University of Calcutta. A voucher specimen (AG100699, AG100799, AG100) has been deposited at the Department of Botany, University of Calcutta, Kolkata, India. Preparation of plant root extract a) Aqueous plant extract: The air dried grounded plant roots (2 gm) were soaked in distilled water overnight at 8 °C. It was filtered and the filtrate was centrifuged at 2000 rpm X 10 mins. The supernatant was used for further testing and kept at 8 °C. The plant extract was expressed in terms of dry weight. b) Methanol plant extract: Air dried plant roots were grounded and extracted by refluxing with petroleum ether (60-80 °C), chloroform (6065 °C) and methanol (64-65 °C) in a soxhlet apparatus. The materials were then dried in a desiccators at room temperature for further use. Before use, the extracted material was dissolved in 0.9% saline and centrifuged at 2000 rpm X 10 mins at room temperature. Supernatant was used for further investigation and kept at 4 °C. The plant extract was expressed in terms of dry weight. Animals used Male Swiss albino mice (20±2 gm), male guineapig (180±10 gm) and Wister albino rats (100±10 gm) were obtained from enlisted supplier of Calcutta University and were maintained in standard laboratory conditions, diet and water ad libitum. All animal experiments were approved by the University Animal Ethics Committee, Department of Physiology, University of Calcutta, Kolkata, India and were in accordance with the guidelines of the committee for the purpose of Control and Supervision of Experiments on Animal (CPCSEA), Government of India (Ref. No.: 820/04/ac/ CPCSEA dated 06.08.2004). Scorpion venom neutralization (in vivo) Neutrlization of lethal activity: MLD (Minimum Lethal Dose) of SV was defined as the minimum dose of SV, which when injected in tail vein (i.v) of male albino mice (20±2 g), caused death of the animal within 24 hrs [17]. Antilethal activity of plant root extracts was studied by the neutralization method [18] using incubate of certain Aqueous extracts (mg/kg) SV mg (no. of animals died/total animals) MLD Fold Protection (%) H.I (100) 0.4(0/6) 1 100