Phytochemical and biological studies of Croton bonplandianum (Euphorbiaceae)

Abstract

The research work was carried out for the phytochemical and biological studies of Croton bonplandianum (Euphorbiaceae). Preliminary phytochemical screening revealed the presence of alkaloids, saponins, flavonoids, tannins and terpenoids while anthraquinone glycosides and cardiac glycosides were absent. The extraction of dried plant material was affected by dichloromethane and methanol successively. Both dichloromethane and methanol extracts were subjected to biological activities such as antibacterial, antifungal, antioxidant, α-chymotrypsin inhibitory, urease inhibitory, α-glucosidase inhibitory and butyrylcholinesterase inhibitory activities along with brine-shrimp toxicity, phytotoxicity against Lemna minor. Dichloromethane extract has shown in vitro α-glucosidase inhibitory activity of 97.89 % with IC50 value of 14.93 μg/ml compared to the standard acarbose, which exhibited 92.23 % inhibition with IC50 value of 38.25 μg/ml. Methanol extract appeared with potent butyrylcholinesterase inhibitory activity of 84.14 % with IC50 found to be 31.01 μg/ml compared to the standard eserine, which exhibited 82.82 % inhibition with IC50 value of 30.01 μg/ml. Methanol extract was found toxic with LD50 value of 115.76 (0.0048 - 13.76) μg/ml against Artemia salina and also showed radical scavenging activity (%RSA) of 59.62% with IC50 value of 396.20 μg/ml . Based on these results activity guided isolation of constituents from dichloromethane and methanol extracts were done. Fractionation of dichloromethane extract by column chromatography on silica gel and Sephadex LH 20 using different mobile phase systems led to the purification of compounds (A-I). The structures of these isolated compounds were established by spectroscopic technique such as UV and IR spectroscopy. Proton Nuclear Magnetic Resonance (1H NMR), 13C NMR and Mass spectrophotometry (EIMS, HRMS) were used for elucidation of structure. On the basis of physical and spectral data from literature, these compounds were identified as n-pentacosanyln- nonadeca-7′-en-9′-α-ol-1′-oate (A), n-tridecanyl n-octadec-9,12-dienoate (B), nonacosyl hexadecanoate (C), heptacosanoic acid (D), 1,3,5-trihydroxy-2-hexadecanoylamino-(6e,9e)- heptacosdiene (E), coumarin (F), betulin (G), stigmasterol (H), and 3,5-dimethoxy 4-hydroxy cinnamic acid (I) were isolated. All these compounds were screened for in vitro α-glucosidase inhibitory activity, compound F, G and I possessed significant α-glucosidase inhibitory activity in a concentration-dependent manner and explained more potent inhibitory activity with IC50 values ranging from 23.0 to 26.7 μg/ml than that of a positive control acarbose (IC50, 38.2 6 μg/ml). Fractionation of methanol extract by column chromatography on silica gel using different mobile phase system afforded five compounds (J-N). Based on spectral data the chemical structure has been established as 4-hydroxy-3,5-dimethoxybenzoic acid (J), 5,8- dihydroxycoumarin (K), stigmasterol 3-O- β -D-glucoside (L), sparsifol (M) and 6-O-β-Dglucopyranosyl- β-D-(1-O-sinapoyl,6'-O-sinapoyl)-glucopyranose (N) were isolated from methanol extract of Croton bonplandianum. The compounds J, K, L and N exhibited significant butyrylcholinesterase inhibitory activity in a concentration-dependent manner and exhibited potent inhibitory activity with IC50 values ranging from 21.0 to 36.0 μg/ml, than that of a positive control eserine (IC50, 32.0 μg/ml).