Evaluation of the Antioxidant, Antimicrobial and Anti-inflammatory Activities of Cyphostemma cyphopetalum (Fresen) Descoings ex-Wild & Drummond and Cyphostemma junceum (Webb) Descoings ex-Wild & Drummond
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Date
2018-02-02
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Addis Ababa University
Abstract
Cyphostemma cyphopetalum and Cyphostemma junceum (Family Vitaceae) are known for their various medicinal uses in different parts of Ethiopia. This study was therefore conducted to evaluate the antioxidant, antimicrobial and anti-inflammatory activities of the leaves and roots of C. cyphopetalum and C. junceum. The leaves and roots of the two plants were extracted using 80% methanol and distilled water with maceration technique and filtered using Whatman No 1 filter paper and the filtrate of 80% methanol extracts were concentrated using rotary evaporator and that of water extracts were stored at (-20oC) and lyophilized to dryness. The extracts were further fractionated with methanol, ethanol, acetone and distilled water. Acute toxicity of 80% methanol extracts were evaluated on female Swiss albino mice up to a dose of 5000mg/kg body weight and the secondary metabolites present in these extracts were identified. Compounds were isolated from eight gram ethanol extract of the leaves of C. cyphopetalum using column chromatography. The column was eluted with n-hexane, chloroform, ethyl acetate and methanol to afford 24 fractions A-X. Structure of the compounds was elucidated using nuclear magnetic resonance (NMR) and compared with the available database. The antioxidant activities of the water crude extracts and methanol solvent fractions of the leaves and roots of the two plants were evaluated using 2, 2 diphenyl-1-picrylhydrazyl hydrates (DPPH) radical scavenging assay. Antimicrobial activities of 80% methanol extracts and methanol, ethanol, and acetone solvent fractions were evaluated against pathogenic gram-negative bacteria; Escherichia coli (ATCC25922), Pseudomonas aeruginosa (ATCC27853), Shigella boydii (ATCC9207), Shigella flexineri (ATCC12022), Klebsiella pneumonia (ATCC70603) and Gram-positive bacteria; Staphylococcus aureus (ATCC29213), Staphylococcus epidermidis (ATCC12228), Listeria monocytogens (ATCC19115) and the fungal test organism, Candida albicans (ATCC10231) using agar disc and agar well diffusion methods. The minimum inhibitory concentration (MIC) of the methanol solvent fraction was determined using broth dilution method, which was followed by conducting the minimum bactericidal concentration (MBC). Ciprofloxacin and fluconazole were used as positive control for the antibacterial and antifungal test respectively. The anti-inflammatory effect of the crude methanol, water fractions and β-sitosterol isolated from the ethanol leaves extract of C. cyphopetalum was evaluated using carrageenan-induced mouse paw edema. The acute toxicity study indicated that 80% methanol leaves and roots extracts did not cause death or induce other toxicity symptoms on mice to a dose of 5000mg/kg body weight. Alkaloids, flavonoids, catechol tannins, saponins, and terpenoids were the secondary metabolites identified from 80% methanol extracts of the two plants. β-sitosterol, diterpenoid, luteolin, sitosterol glucoside, resveratrol dimer, salicylic acid, a mixture of flavonoid glycosides and sucrose were isolated from ethanol leaves extract of C. cyphopetalum. The methanol solvent fraction of the leaves of C. cyphopetalum and C. junceum showed respective inhibition of 92.2% and 87.1% at 12.5mg/mL while ascorbic acid showed inhibition of 90.9% at 12.5mg/mL on DPPH assay. The highest inhibition zone was exhibited against S. aureus by 80% methanol leaves extract of C. cyphopetalum with a diameter of 13.0±0.0mm at 500mg/mL and methanol solvent fractions of the leaves of C. cyphopetalum with a diameter of 15.0±0.0mm at 500mg/mL whereas, the acetone solvent fractions of the leaves and roots showed the lowest inhibition zone against K. pneumonia with inhibition diameter of 7.0±0.0mm. The methanol solvent fraction of the roots of C. junceum showed the highest inhibition zone (9.7±0.3mm) against S. aureus at 500mg/mL while the methanol, ethanol, and acetone solvent fractions of the leaves of C. junceum exhibited the lowest against K. pneumonia (6.7±0.3mm). The methanol solvent fractions of the leaves of C. cyphopetalum exhibited the highest antifungal activity against C. albicans with inhibition diameter of 12.3±0.3mm when compared with ethanol and acetone solvent fractions. The methanol solvent fractions of the leaves of C. cyphopetalum showed the lowest MIC (15.63mg/mL) and MBC (31.25mg/mL) against S. aureus and S. epidermidis. The highest MIC (125mg/mL) and MBC (250mg/mL) were depicted by methanol solvent fractions of the leaves of C. cyphopetalum and roots of C. junceum against K. pneumonia. On the anti-inflammatory evaluation, 80% methanol leaves extracts and water fractions of the leaves and roots of C. cyphopetalum and C. junceum markedly reduced the swelling. The percentage reduction on mice paw thickness was 95.83% for 80% methanol leaves extracts of C. cyphopetalum (500mg/kg), 97.40% for distilled water fractions of the leaves of C. cyphopetalum (300mg/kg) and 97.40% for β-Sitosterol (25mg/kg) at six hours. Likewise, the percentage reduction on mice paw thickness was 88.54% and 93.23% for 80% methanol leaves extract of C. junceum (500mg/kg) and distilled water fractions of the leaves of C. junceum (300mg/kg) respectively. The leaves of C. cyphopetalum had a significant anti-inflammatory effect compared with the indomethacin (10mg/kg) ( P < 0.05) that showed percentage paw thickness reduction of 93.23%. It can be concluded that the leaves and roots of C. cyphopetalum and C. junceum have antioxidant, antimicrobial and anti-inflammatory activity at the tested doses. Thus, these plants could be used for possible drug development.
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Acute Toxicity, Anti-Inflammatory, Antimicrobial, Antioxidant, Cyphostemma Cyphopetalum, Cyphostemma Junceum, Phytochemical Screening