Exploring nature's antidote: unveiling the inhibitory potential of selected medicinal plants from Kisumu, Kenya against venom from some snakes of medical significance in sub-Saharan Africa

Abstract

Background: The present study investigated the efficacy of Conyza bonariensis, Commiphora africana, Senna obtusifolia, Warburgia ugandensis, Vernonia glabra, and Zanthoxylum usambarense against Bitis arietans venom (BAV), Naja ashei venom (NAV), and Naja subfulva venom (NSV). Methods: 40 extracts and fractions were prepared using n-hexane, dichloromethane, ethyl acetate, and methanol. In vitro efficacy against snake venom phospholipase A₂ (svPLA₂) was determined in 96-well microtiter and agarose-egg yolk coagulation assays. in vivo efficacy against venom-induced cytotoxicity was determined using Artemia salina. Two commercial antivenoms were used for comparison. Results: The 96-well microtiter assay revealed poor svPLA₂ inhibition of BAV by antivenom (range: 20.76% ± 13.29% to 51.29% ± 3.26%) but strong inhibition (>90%) by dichloromethane and hexane fractions of C. africana, hexane and ethyl acetate extracts and fraction of W. ugandensis, dichloromethane fraction of V. glabra, and the methanol extract of S. obtusifolia. The methanol extract and fraction of C. africana, and the hexane extract of Z. usambarense strongly inhibited (>90%) svPLA₂ activity in NAV. The hexane and ethyl acetate fractions of V. glabra and the dichloromethane, ethyl acetate, and methanol extracts of C. africana strongly inhibited (>90%) svPLA₂ in NSV. The agarose egg yolk coagulation assay showed significant inhibition of BAV by the dichloromethane fraction of C. africana (EC₅₀ = 3.51 ± 2.58 μg/mL), significant inhibition of NAV by the methanol fraction of C. africana (EC₅₀ = 7.35 ± 1.800 μg/mL), and significant inhibition of NSV by the hexane extract of V. glabra (EC₅₀ = 7.94 ± 1.50 μg/mL). All antivenoms were non-cytotoxic in A. salina but the methanol extract of C. africana and the hexane extracts of V. glabra and Z. usambarense were cytotoxic. The dichloromethane fraction of C. africana significantly neutralized BAV-induced cytotoxicity, the methanol fraction and extract of C. africana neutralized NAV-induced cytotoxicity, while the ethyl acetate extract of V. glabra significantly neutralized NSV-induced cytotoxicity. Glycosides, flavonoids, phenolics, and tannins were identified in the non-cytotoxic extracts/fractions. Conclusion: These findings validate the local use of C. africana and V. glabra in snakebite but not C. bonariensis, S. obtusifolia, W. ugandensis, and Z. usambarense. Further work is needed to isolate pure compounds from the effective plants and identify their mechanisms of action.

Keywords: Artemia salina bioassay; Bitis arietans; Naja ashei; Naja subfulva; medicinal plants; preclinical efficacy evaluation; snake venom.

FIGURE 1

Photos of Bitis arietans (A), Naja ashei (B), and Naja subfulva (C). Photos by Mitchel Okumu.

FIGURE 2

Photos of Commiphora africana (A. Rich) Engl (A), Conyza bonariensis (L.)., Cronquist (B), Senna obtusifolia (L.) Irwin and Barneby (C), Vernonia glabra (Streetz) (D), Warburgia ugandensis (Sprague) (E), and Zanthoxylum usambarense (Engl.) Kokwaro (F) used in this study.

FIGURE 3

The effects of extracts, fractions, and antivenoms on the minimum phospholipase concentration of Bitis arietans venom. BA: Bitis arietans, VG (DCM-M): Dichloromethane fraction of Vernonia glabra leaves, CA SB (HEX-M): Hexane fraction of Commiphora africana stem bark, CA SB (DCM-M): Dichloromethane fraction of Commiphora africana stem bark. SO (MEOH-S): Methanol extract of Senna obtusifolia leaves, WU (HEX-S): Hexane extract of Warburgia ugandensis leaf stalk, WU (EA-M): Ethyl acetate fraction of Warburgia ugandensis leaf stalk, VBA: Vins bioproducts antivenom, IBA: Inoserp biopharma antivenom.

FIGURE 4

The effects of extracts, fractions, and antivenoms on the minimum phospholipase concentration of Naja ashei venom. NA: Naja ashei, CA B (MEOH-S): Methanol extract of Commiphora africana bark, ZU R (HEX-S): Hexane extract of Zanthoxylum usambarense root, CA SB (MEOH-S): Methanol extract of Commiphora africana stem bark, CA SB (MEOH-M): Methanol extract of Commiphora africana stem bark, VBA: Vins bioproducts antivenom, IBA: Inoserp biopharma antivenom.

FIGURE 5

The effects of extracts, fractions, and antivenoms on the minimum phospholipase concentration of Naja subfulva venom in the snake venom phospholipase A₂ agarose-egg yolk assay. NS: Naja subfulva, VG (DCM-S): Dichloromethane extract of Vernonia glabra leaves, VG (HEX-S): Hexane extract of Vernonia glabra leaves, VG (EA-M), Ethyl acetate fraction of Vernonia glabra, VG (HEX-M): Hexane fraction of Vernonia glabra, VG (MEOH-S): Methanol extract of Vernonia glabra leaves, VG (EA-S): Ethyl acetate extract of Vernonia glabra leaves, CA SB (MEOH-S): Methanol extract of Commiphora africana stem bark, VBA: Vins bioproducts antivenom, IA: Inoserp antivenom.