PHYTOCHEMICAL AND SOME BIOLOGICAL STUDIES OF THE METHANOL LEAF EXTRACT OF ASPILIA AFRICANA (PERS.) C. D. ADAMS (ASTERACEAE)
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Abstract
Aspilia africana, (Pers) C.D. Adams; (Asteraceae), a semi perennial woody plant, commonly known as haemorrhage plant is widely distributed across tropical Africa and all parts of Nigeria. The plant is well known in folkloric medicine for its anti-haemorrhagic uses and for the treatment of cardiovascular diseases, diabetes, malarial fever, wounds, coughs, microbial infections and gastro-intestinal disorders.This research was undertaken inorder to isolate some of the bioactive compounds responsible for the anti-microbial and anti-diabetic activities of this plant. The dried powdered leaf material was subjected to cold maceration with 70% aqueous methanol to obtain the methanol extract which waspartitioned with n-hexane, dichloromethane, ethyl acetate and n-butanol. The methanol extract and the fractions were screened for phytochemical constituents, anti-microbial and anti-diabetic properties and then subjected to chromatographic separation. Preliminary phytochemical screening results showed the presence of carbohydrate, cardiac glycosides, flavonoids, polyphenols, saponins, steroids, tannins and terpenoids in the methanol extract and the fractions. Chemical investigation of the methanol extract and fractions of the leaves led to the isolation of three pentacyclic triterpenic acids namely; 3β – hydroxyolean – 12 – en – 28 – oic acid (Oleanolic acid), 3β – hydroxyurs – 12 – en – 28 – oic acid (ursolic acid) and 2, 3 - dihydroxyurs – 12 en – 28 – oic acid (corosolic acid) from the butanol fraction and 3β-hydroxyurs-12-en-28-oic (ursolic acid) from the dichloromethane fraction through silica gel column chromatography. The structures of the isolated compounds were elucidated based on the analysis of 1-D and 2-D NMR (including 1H, 13C, HMBC, HMQC, COSY, and DEPTexperiments), FTIR spectral data, physicochemical properties and comparison with authentic data of these compounds from literature. In Agar welldiffusion anti-microbial screening using isolated clinical strains of pathogens namelyStaphylococcus aureus, Methicillin Resistant Staphylococcus aureus (MRSA), viii Streptococcus pyogenes, Bacillus substilis, Proteus vulgaris, Salmonella typhi, Shigella dysenteriae, Escherichia coli, Klebsiella pneumoniae Candida albicans and Candida stellafoidea; the methanol extract and the butanol fraction (which was more active than other fractions) inhibited the growth of nine of the eleven bacterial strains and one of the three fungal strains with zones of inhibition ranging from 10 – 25mm. However, the activity of one of the isolated compounds (oleanolic acid) surpassed that of crude extract and n-butanol fraction; it inhibited the growth of all the bacteria and two of the fungal strains with inhibition zones ranging from 25 – 33mm compared with the standard drugs used. The MIC, MBC/MFC of the plant extracts ranged from between 5.00 and 10.00 mg/mL while that of the isolated compounds ranged between 0.0125 and 0.0500mg/mL. In the in-vitro anti-diabetic studies, the isolated oleanolic acid (IC50 = (25.45±0.45 µg/mL; 47.57±0.40 µg/mL) was more potent in the inhibition of α-amylase and α-glucosidase than the crude methanol extract (IC50 = 96.40±0.20µg/mL; 95.10±0.20µg/mL), butanol fraction (the only active fraction, IC50 = 72.50±0.65µg/mL; 72.65±3.2µg/mL) and the reference drug (acarbose) (IC50 = 46.31±0.58 µg/mL; 48.10±0.130 µg/mL) for α-amylase and α-glucosidase respectively. The results of the anti-microbial and antidiabetic studies have authenticated the traditional use of the plant to treat microbial infection and diabetes.The research work has also identified the oleanolic acid to be responsible for the antimicrobial and anti-diabetic activities of the plant.
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