http://hdl.handle.net/123456789/93 2026-04-13T10:38:38Z http://hdl.handle.net/123456789/1584 ISOLATION OF BIOACTIVE COMPOUNDS FROM SELECTED NIGERIAN MEDICINAL PLANTS FOR MANAGEMENT OF LETROZOLE-INDUCED POLYCYSTIC OVARIAN SYNDROME IN RATS OGUNLAKIN, AKINGBOLABO DANIEL 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. 2021-02-01T00:00:00Z http://hdl.handle.net/123456789/1582 MOLECULAR AND BIOLOGICAL STANDARDISATION OF Alstonia boonei DE WILD. AND Alstonia congensis ENGL. LEAVES AND STEM-BARKS OMITOLA, Opeyemi Josephine Alstonia boonei (AB) and Alstonia congensis (AC), belonging to the family Apocynaceae are different but closely related species, indigenous to Africa. The two plants possess varied biological activities including antimalarial, antihypertensive and antidiarrhoeal. However, the apparent similarities between the two species could lead to misidentification and inappropriate medicinal utilisation, necessitating the need to establish diagnostic characters for each species for proper identification. This study, was therefore, aimed at evaluating molecular, pharmacognostic, antispasmodic and antidiarrhoeal profiles of the two plants for their pharmacopoeial standardisation. Deoxyribonucleic acid was isolated from nine accessions of AB and AC leaves (ABL and ACL) collected from southwestern Nigeria and amplified, using Internal Transcribed Spacer (ITS) region. Micromorphological diagnostic characters of leaves and stem-barks were studied by light microscopy. Physico-chemical and elemental analysis of the powdered samples were determined using incineration method and Atomic Absorption Spectroscopy, respectively. Aqueous extraction of plant samples was done, extracts were concentrated in vacuo and freeze dried. The freeze-dried extracts were partitioned successively into dichloromethane (DCM), ethyl acetate and aqueous fractions. Antispasmodic activities of the extracts and fractions were evaluated on high-potassium induced and spontaneous contractions on isolated rat ileum. The AB and AC stem-barks (ABSb, ACSb) extracts and their dichloromethane fractions (DCM-ABSb, DCM ACSb) showing high antispasmodic activity were evaluated for in vivo antidiarrhoeal activities in mice (22- 25 g b.w). Loperamide (5mg/kg) was used as standard. Compounds were isolated from the most active fraction (DCM-ABSb) using chromatographic techniques (Column, TLC). Structures of the isolated compounds were elucidated using spectroscopic techniques (NMR and MS) and their antispasmodic activities evaluated. Data were analysed using descriptive statistics, unweighted pair group method with arithmetic mean and one-way ANOVA and Tukey’s multiple comparison at α0.05. The amplification of the ITS region discriminated between the two Alstonia species. The adaxial epidermal cells of both species were polygonal, straight anticlinal walls and leaves were hypostomatic. The vascular bundle of ABL mid-rib was arc-shaped, while that of ACL was V-shaped. Moisture content values of stem bark of AB (6.4±0.06%) was significantly different from that of AC (7.2±0.4%), while the values were not significantly different for the leaves. Elemental analysis revealed that calcium was the most abundant mineral in the leaves of AB (68.2±3.2 mg/g) and AC (65.7 ±1.0 mg/g). The stem-bark extracts of AB and AC were antispasmodic with IC50 of 0.03±0.2, 0.12±0.01 mg/mL and 1.15±0.1, 1.05±0.8 mg/mL on high-potassium induced and spontaneous contractions, respectively. The DCM-ABSb fraction exhibited significant antispasmodic activity (IC50: 0.02±0.05, 0.31±0.02 mg/mL) on high-potassium induced and spontaneous contractions, respectively. The DCM-ABSb (200 mg/kg b.w.) showed significant antidiarrhoeal activity (87.3%) comparable to Loperamide (87.5%). The isolated compounds from DCM-ABSb were β-amyrin and boonein with only boonein exhibiting antispasmodic activities on both high-potassium induced (IC50: 0.09±0.01 µg/mL) and spontaneous (0.29±0.05 µg/mL) contractions. Alstonia boonei and Alstonia congensis were identified as two distinct species. The diagnostic indices of the two plants provided pharmacopoeial standards for their identification. The isolated boonein could serve as a template for the development of antidiarrhoeal drugs 2021-02-01T00:00:00Z http://hdl.handle.net/123456789/1580 ISOLATION AND CHARACTERISATION OF ACETYLCHOLINESTERASE AND PROLYL ENDOPEPTIDASE INHIBITORS FROM SELECTED MEDICINAL PLANTS ONOJA, OJOGBANE JOEL Acetylcholinesterase (AChE) and Prolyl endopeptidases (PEP) are involved in the catalytic hydrolysis of acetylcholine and cleavage of neuropeptides influencing the activity of the brain, leading to some symptoms of Alzheimer’s disease (AD).Inhibition of AChE and PEP is considered a promising strategy for AD management.There are no effective drugs for the management of AD which necessitated the search for new potent neurotherapeutic agents.This study, was therefore, aimed at isolating and identifying inhibitors of AChE and PEP from selected medicinal plants identified from Nigerian ethnomedicine. Ten plants selected were macerated in methanol and their extracts evaluated for AChE inhibitory activity using Ellman colorimetric in vitro assay. The most active plant extracts: Phyllanthus muellerianus (PM) leaves (FHI 111339), Tinospora cordifolia(TC) stem (FHI 112287) and Cola hispida(CH) seed (FHI 111321) were successively partitioned into n-hexane, dichloromethane, ethyl acetate and aqueous fractions. The fractionswere assessed for AChE inhibitory activity using eserine as standard. In vitro analyses were used to evaluate the antioxidant potential of the plant extracts. Active fractions were subjected to chromatographic separations to isolate and purify bioactive compounds. Structures of isolated compounds were determined by spectroscopic analyses. Compounds were screened for AChE and PEP inhibition using in vitro colorimetric assays and metal chelating potential, respectively. Bacitracin was used as standard for PEP. Molecular docking was done using software (MOE 2015.010) to anticipate the binding affinity between drug candidates with protein targetsusing PDB ID: 3IVM (Prolyl endopeptidase) and 10CE (Acetylcholinesterase). Standard curves were generated and calculation of IC50 values was done using Microsoft Excel. Data were analysed using One way ANOVA followed by Dunnet’s Multiple Comparison testat α0.05 Methanol extracts of PM (IC50 of 1.29±0.70 mg/mL) and CH (IC50 of0.87±0.40 mg/mL) gave promisingAChE inhibitory activity at 5 mg/mL. Ethyl acetate fraction of PM (0.74±0.12 mg/mL) and CH(0.66±0.24 mg/mL) gave the highestAChE inhibitory activity compared to eserine. The dichloromethane fraction of TC exhibited good metal chelating activity (IC50 of 0.20±0.08 mg/mL). Beta-sitosterol, daucosterol, 1-octacosanol, rel-(2s,3s,4r,16e)-2-[(2'r)-2'-hydroxynonadecanoylamino]-heneicosadec-16-ene-1,3,4-triol, four clerodane diterpenoids and five alkaloids were isolated from TC.Two steroids, 5-hydroxymethylfurfural, and 2-hydroxyquinoline-4-carboxylic acidwere isolated from CH. Stigmasterol and β-sitosterol glucoside were isolated from PM. Oxoglaucine (Oxoaporphinoid alkaloid) from TC exhibited the highest AChE inhibitory activity (IC50 of 0.80±0.09 mg/mL) compared to eserine (IC50=0.53±0.34 mg/mL) at 1 mg/mL, while stigmasterol from PM demonstrated the highest PEP inhibitory activity (IC50= 0.77±2.9 mM) compared to bacitracin (IC50= 0.13±1.5 Mm) at 1 mM.Oxoglaucine also gave the highest metal chelating potential (IC50= 0.22±0.00 mg/mL) compared to EDTA (IC50 of 0.05±0.11 mg/mL). Molecular docking revealed hydrophobic, hydrogen bonding and π -stacking interactions among tested compounds and the active site of AChE andPEPresidue, which indicates their ability to mitigate the symptoms associated with AD. Oxoglaucine and stigmasterol isolated from Tinospora cordifoliaand Phyllanthus muellerianus, respectivelyshowed high acetylcholinesterase and prolyl endopeptidase inhibitory activities. The two biomolecules could serve as potential leads for novel drug development for the management of Alzheimer’s disease. 2021-01-01T00:00:00Z http://hdl.handle.net/123456789/1578 ANTISICKLING ACTIVITIES OF CASSIA SIEBERIANA DC (LEGUMINOSAE) FATOKUN, OMOLOLA TEMITOPE Sickle Cell Disease (SCD) affects primarily black populations. There are few drugs available for the management of SCD with deleterious side effects complicating the condition. Although many plants including Cassia sieberiana have been documented in the management of SCD, there is paucity of scientific evidence for their effectiveness and bioactive constituents. This study was designed to investigate the antisickling activities of morphological parts of C. sieberiana, isolate and characterise their constituents. The powdered samples of C. sieberiana (FHI- 112359) root, seed, pericarp and whole fruit were extracted into 70% ethanol by Soxhlet extraction and water by reflux. The extracts were used to either inhibit or reverse sodium metabisuphite induced sickling of HbSS erythrocytes from SCD patients in steady state, in vitro. The ethanol extract of the most active plant part (whole fruit) was successively partitioned to give n-hexane, dichloromethane (DCM), ethyl acetate (EtOAc) and aqueous fractions. The fractions were screened for inhibition and reversal of sickling and inhibition of haemoglobin polymerisation. The percentage of inhibition, reversal and rate of decrease in HbSS polymerisation were calculated. The DCM and EtOAc fractions were purified using column and vacuum liquid chromatographic (VLC) techniques. Chromatographic fractions were screened for inhibitory and reversal activities and the active fractions were further purified using column and preparative thin layer chromatography to isolate compounds. Structures of isolated compounds were identified using 1D and 2D NMR (1H and 13C). Data were analysed using one-way ANOVA followed by Student t-test at α0.05. The ethanol extract of C. sieberiana whole fruit (CSF) at 180 minutes, exhibited the highest inhibitory activity (83.7±1.3)% compared to seed (65.7±2.2)%, pericarp (10.9±1.2)% extracts and was significantly different from vanillic acid-reference standard (50.4 ± 0.4)% and water whole fruit (51.4±1.6)% extract, while the root ethanol extract lysed the erythrocytes. The root ethanol extract exhibited a reversal activity of (88.9±0.8)%, compared to whole fruit (82.3±0.8)%, seed (82.2±1.0)% and pericarp extracts (55.6±1.4)%; para hydroxy benzoic acid reference standard (86.0±0.6)% and water whole fruit extract (84.4±1.9)%. The CSF exhibited both iii inhibitory and reversal activities and also decreased the rate of HbSS polymerisation by (73.5±0.06)%. The DCM and EtOAc fractions exhibited comparable inhibitory (77.9±1.1)%; (77.3±2.9)% and reversal (73.5±1.4)%; (76.8±3.8)% activities, respectively. The EtOAc and aqueous partitioned fractions decreased the rate of HbSS polymerisation by (80.6±0.02)% and (84.2±0.03)%, respectively, which were significantly lower than that of ascorbic acid (92.3±0.002)%. The DCM column fractions B3 and D3 exhibited inhibitory activities of (87.9±0.1)% and (91.7±1.5)%, respectively, which were significantly higher than the activities of all other column fractions. Fractions B3 (70.7±1.6)% and D3 (86.2±1.4)% also exhibited reversal activities. Nine phenolic compounds were isolated from; EtOAc fraction (E2, E3 and E4) and DCM fraction (M2, M3, G2, G3 and G4). One of the compounds - E3 was characterised as 4′-methoxy epiafzelechin. 2019-09-01T00:00:00Z