ISOLATION OF BIOACTIVE COMPOUNDS FROM SELECTED NIGERIAN MEDICINAL PLANTS FOR MANAGEMENT OF LETROZOLE-INDUCED POLYCYSTIC OVARIAN SYNDROME IN RATS

Abstract

Polycystic Ovary Syndrome (PCOS) is an endocrine disorder with a global prevalence of 5-10% among women of reproductive age. Women with PCOS have a 2.7-fold increased risk of developing ovarian and cervical cancers. Ethnobotanical survey revealed that plants including Kigelia africana (Lam.) Benth. (KA), Basella alba L. (BA), Tetracera potatoria G. Don (TP) and Mormodica charantia L. (MC) are used for treatment of PCOS in southwestern Nigeria. However, there is no reported scientific evidence to validate these claims. This study was, therefore, designed to investigate the effect of the four plants in alleviating polycystic ovary conditions in rats and the associated risk of ovarian and cervical cancers. Letrozole (1 mg/kg) was used for the induction of PCOS in thirty female albino rats (180-200 g, n=5). Methanol leaf extracts of KA, BA, TP and MC (100 mg/kg b.w.) and Clomiphene citrate (1 mg/kg b.w., standard drug) were administered for 15 days. Histopathological evaluation of the ovaries was done microscopically. Luteinizing hormone (LH), follicle stimulating hormone (FSH) and estradiol were measured using ELISA. Extracts of the most active plants, KA (FHI-111350) and TP (IFE-17794) were successively partitioned into n-hexane, dichloromethane and ethyl acetate fractions, and screened for PCOS inhibitory activity. Compounds were isolated from the active fractions using chromatographic techniques. Structures of isolated compounds were elucidated using spectroscopic techniques. Anti-proliferative effect of fractions, isolated compounds and derivatised cinnamic acid analogues were determined on cervix adenocarcinoma (HeLa) and Chinese Hamster ovarian (CHO 1) cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Doxorubicin was used as standard. Data were analysed using one-way ANOVA followed by Student’s t-test at α0.05. Animals treated with KA showed normal ovarian stroma with moderate fibroblastic tissues. The levels of FSH in KA, BA, TP and MC treated groups were 0.96±0.08, 1.10±0.23, 0.81±0.04 and 0.69±0.01 mIU/mL, respectively compared to control group (0.93±0.19 mIU/mL). The TP (0.19±0.05 mIU/mL) significantly reduced the level of LH compared with the control group (0.22±0.0l mIU/mL). Hexane and ethyl acetate fractions displayed selective moderate inhibitory effect (IC50 = 5.3±1.10 µg/mL) on CHO 1 cell line compared to doxorubicin (IC50 = 0.8±0.01 µg/mL). Compounds 3-(3, 4-dimethoxyphenyl) acrylic acid (1), sitosterol (2), methyl 3-(3,4- dihydroxyphenyl) acrylate (3), 2, 3-dihydro-5-(hydroxymethyl) furan-2, 3, 4-triol (4), p-coumaric acid (5) and caffeic acid (6) were isolated from KA, while apigenin (7) was isolated from TP. iii Compound 1 displayed moderate anti-proliferative activity against HeLa cell line (IC50 = 33.5±0.60 µg/mL). Compound 7 inhibited proliferation of HeLa cell line (IC50 = 6.2 µg/mL) and had moderate inhibitory effect on CHO l cell line (IC50 = 22.2 µg/mL). Derivatised compound N'-(2, 6- dimethoxybenzylidene)-3-(4-methoxyphenyl) acrylohydrazide (8) inhibited both CHO l (IC50 = l8.4±4.l µg/mL) and HeLa (IC50 = 22.4±0.4 µg/mL) cells. The extracts of Kigelia africana and Tetracera potatoria had inhibitory effects on polycystic ovary condition, irregular oestrual cycle and hormonal imbalance in female albino rats. The phenylpropanoids and derivatised analogues exhibited antiproliferative effect on ovarian and cervical cancer cell lines, which could contribute to their use for management of polycystic ovary syndrome.