Doctoral Degrees (Plant Sciences)

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Browsing Doctoral Degrees (Plant Sciences) by Author "Ashafa, A. O. T."

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    Antioxidant, antidiabetic and cardioprotective activities of Dicoma anomala (sond.) used in the Basotho traditional medicine
    (University of the Free State, 2016) Balogun, Fatai Oladunni; Ashafa, A. O. T.
    𝑬𝒏𝒈𝒍𝒊𝒔𝒉 Dicoma anomala (Sond.) belongs to the Asteraceae family and locally called Hloenya (South Sotho), fever or stomach bush (Afrikaans). The plant is used in the management of various diseases, particularly diabetes mellitus among the Basotho tribe of eastern Free State Province, South Africa. The study evaluates the antioxidant, antidiabetic and cardioprotective potentials of the plant as a way of validating the folkloric usage. The result of in vitro antioxidant assays [2, 2- azino-bis (3-ethylbenzothiazoline-6-) sulfonic acid (ABTS), reducing power, superoxide anion, hydroxyl radicals, 1,1-diphenyl-2-picryl hydrazyl (DPPH) radicals, etc.] as well as phytochemicals (such as total phenol, total flavonoids and total antioxidant capacity) in various concentrations (1.56-25 µg/ml) tested using water, ethanol, hydro-ethanol and methanol extracts of the plant’s root revealed that the water extract exhibited the best activity with half maximal inhibitory concentration (IC50: 15.20, 11.70, and 0.84 µg/mL) in DPPH, hydroxyl radical, and superoxide anion radicals respectively. The four extracts also possessed high phenolic contents, total antioxidant capacity with lower total flavonoids content. The effect of treatment with 125, 250 and 250 mg/kg body weight (b.w.) aqueous roots extract of Dicoma anomala (AQRED) was investigated in vivo in CCl4- induced hepatotoxic rats in a 15-day curative and prophylactic study. The result revealed that pre-treatment and treatment with AQRED lowers the elevated serum activities of aspartate transaminase (AST), alanine aminotransferase (ALT) and the level of thiobarbituric acid reactive species (TBARS) while restoring the activities of liver antioxidant enzymes such as catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD) towards normal control in a dose-dependent manner. This result proved the antioxidant and hepatoprotective activity of the plant The in vitro antidiabetic potential of D. anomala was investigated via the inhibition of αamylase and α-glucosidase using same extracts (as above) at the range of 1.56 - 25.00 µg/mL concentrations. All the tested extracts of the plant were active against both enzymes, although, the most potent against α-amylase and α -glucosidase was hydro-ethanol (IC50: 9.00 µg/mL) and water (IC50: 27.41 µg/mL) respectively. Similarly, aqueous extract of the D. anomala displayed competitive and non-competitive inhibition of α -amylase and α - glucosidase respectively using Lineweaver-Burk plot. Treatment with AQRED at concentration 125, 250 and 500 mg/kg b.w. in Wistar rats reversed towards control the elevated blood glucose levels, lipid peroxidation, lipid profile, glycosylated haemoglobin and activities of gluconeogenesis enzymes, with concomitant reduction in the activities of enzymatic antioxidants, glycolytic enzymes as well as the high-density lipoprotein – cholesterol (HDL-c) brought about by streptozotocin induction. Thus, the study proved the antihyperglycaemic activity of the plant. Additionally, AQRED at 125, 250 and 500 mg/kg b.w. was evaluated for its ameliorative activity against isoproterenol (ISP) –induced cardiotoxicity in an animal model. The results from the evaluated biochemical parameters revealed significant (p< 0.05) in these parameters was observed. The data obtained indicate that the lethal dose (LD50) of AQRED is in excess of 2000 mg/kg and its oral administration for 90 days is unlikely to cause any toxic effects. In conclusion, the results from this study proved the antioxidant, antihyperglycaemic and cardioprotective potentials of AQRED. The results further validate the folkloric usage of the plant in the management of diabetes mellitus among the Basotho tribe of Eastern Free State Province, South Africa. ___________________________________________________________________
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    Biological and pharmacological activities of root extracts and isolated compounds of hermann/a geniculata Adeniran
    (University of the Free State, 2017) Ariyo, Adeniran Lateef; Ashafa, A. O. T.
    Hermannia geniculala is of the genus of flowering plant from the subfamily Byttnerioideae in the family Malnaceae. H. geniculata has been used in treatment of several diseases like colic, diabetes mellitus and other oxidative stress induced illnesses. The dry root material is chopped, boiled in water and taken three times daily to ameliorate blood sugar disorders, management of diarrhoea, heartburn, stomach disorder and flatulency called "leletha" in pregnant Basotho women. Phytochemical analysis revealed the presence of saponin, phenols, flavonoids, alkaloids, tannin, phytosterol, triterpenes and anthraquinone. Ethanolic extract exhibited the highest free radical scavenging capability with the lowest ICso value (0.52, 0.38, 0.59, 0.63, 0.39 mg/mL) for I, ldiphenyl- 2-picrylhydrazyl (DPPH), 2,2-Azino-bis(3 ethylbenzothiazoline-6-Sulphonic acid (ABTS), hydroxyl radical, superoxide anion radical, metal chelating abi lity which is significantly lower (p<0.05) than the standard si lymarin while hydro-ethanol has the highest reducing power showing a significant (p<0.05) ICso value of 0.24 mg/mL compared to citrate (!Cso value; 0.5 mg/mL). In antidiabetic studies, ethanolic extract was a potent inhibitor of a-glucosidase (!Cso; 0.0 I mg/mL) which is significantly lower (p<0.05) than standard acarbose ICso value (0.52 mg/mL) and hydro-ethanol decoction and aqueous extracts. It also has a milder percentage inhibition of a-amylase enzyme with lC.'io (0.57mg /mL) which is significantly higher (p<0.05) than the standard acarbose ICso (0.047 mg/mL). The mode of inhibition of a-amylase is by competitive inhibition and uncompetitive inhibition of the o.-glucosidase enzyme was observed in ethanolic extract. These find ings provide an empirical rationalization for the use of the root extract of Hermannia geniculata in the management of diabetes mellitus and other oxidative stress induced ai lments. The Vero, HepG2 and RAW 264.7 macrophage cell lines were used to determine the toxicity of the extracts on cells. Similarly, the capabilities of the extract to inhibit 5-lipoxygenase enzyme activities and overproduction of nitric oxide from LPS-activated RAW 264. 7 macrophages were evaluated. Results showed selective toxicity of the extracts with LCso values of Vero cells ranges from (0.40-0.57 mg/mL) while the LCso value of HepG2 cells varies from (0.0 16-0. 136 mg/mL). The selectivity index (SI) were (3 1.87, 18.87, 33.33 and 3.52) for ethanol, hydro-ethanol, decoction and aqueous extracts respectively. The ethanolic extract inhibited NO production in a concentration dependent manner. There was a decrease of 82% at concentration of0. I mg/mL and the LCso:3.64 mg/mL is lower and significantly different (p<0.05) compared to the reference compound quercetin with LCso value of 8.28 mg/mL. Similarly, the ethanolic extract exhibited potent inhibition of 5-lipoxygenase enzyme with the lowest ICso value of 0. 14 mg/mL which is significantly different (p<0.05) compared to all other extracts and indomethacin. The GCMS chromatograms revealed five compound (2-keto-butyric-acid, 2, 2-Bis ( 4-nitrobenzyl)-l - phenylbutane-1 ,3-dione, n-Undecane, 1,4,5,8-tetrathiadelin and imidazo-1,5-pyrimidine) which has been reported to have antioxidant, anti-inflammatory and antifungal properties. This result suggested that Hermannia geniculata roots extract is not toxic and possesses antioxidant, antinflammatory and anticancer activities which could be exploited in development of new safe and effective drugs. The chemical profiling and in vitro biological activities of flavonoids of Hermannia geniculate (FHG) roots was also investigated using High Pressure Thin Layer Chromatography (HPTLC) finger print analysis. Antioxidant, antidiabetic anti-inflammatory activities and the ability of FHG extract to inhibit the production of nitric oxide (NO) in lipopolysaccharide (LPS) activated RA W264.7 Macrophage were investigated using standard methods. The selective cytotoxicity of the extract on Vero and HepG2 cells was also determined. Kaempferol (Rr0.81) was detected in the extract, its Rr value is similar and comparable with the kaempferol standard used (Rr 0.80). Other flavonoids were also present in the extracts with their Rr values of 0.08-0.95. The FHG extract showed commendable antioxidant properties with !Cso values (3.07± 0.12, 2.13± 0.67) for DPPH and ABTS radicals which was lower and significantly different (p<0.05) compared to standard silymarin with !Cso: (3.55± 0.10, 2.77± 0.75) for DPPH and ABTS respectively. The results indicated milder inhibition of a-amylase with ICso: (5.55± 0.37) which was higher and significantly different from the standard acarbose with ICso: (3.81± 0.29) Nevertheless, the extract exhibited 73% inhibition of a-glucosidase which exerted better inhibitory effect on 5- lipoxygenase enzyme than indomethacin with their respective !Cso: ( I 0.15± 0.02 and 12.03± 0.02). Inhibition of NO production was observed in LPS activated RAW 264. 7 Macrophages with the highest concentration of 0.1 mg/mL decreasing NO production by 87%. Selective toxicity of Vero and HepG2 cells with their respective LC so value of(> I and 0.02 mg/mL) was also observed. The antipro liferative potentials of the extract was confirmed with Selectivity Index of 50. This study indicated for the first time that FHG extract was non-toxic to normal cells and possess antioxidant, antidiabetic, anti-inflammatory and antiproliferative activities. The bioactive constituent and pharmaco logical activ ities of pheno ls extracted from Hermcmnia geniculata (PoHG) roots was investigated using in vitro methods. The chemical profile was determined by HPTLC analysis. Antioxidant, antidiabetic, anti-inflammatory and cytotoxic effect of PoHG on Vero and HepG2 cells was carried out using standard procedures. Phenolic compounds were detected in the sample at Rr (0.14, 0.81 and 0.95). PoHG radical scavenging capabilities on DPPH, ABTS+ and superoxide anion radicals were similar to the standard (silymarin). The ICso values were DPPH (0.12± 0.00), ABTS (0. 13± 0.01) and superoxide anion (0.20± 0.00). The values of the metal chelating activity of Po HG extract is lower and significantly different from the standard (silymarin) their respective ICso values were (0.06± 0.00 and 0.18±0.0 I). The antidiabetic effect was determined by its ability to mildly inhibit a -amylase and strongly inhibit a -glucosidase enzymes, the respective ICso values obtained were (7.52± 0.23 and 1.76 ± 0. 14). PoHG extract exhibited a commendable inhibition of 5-lipoxygenase enzyme with ICso value of (0.15 ± 0 .03) which is similar to the ICso: (011 ± 0.0 I) value for the standard (indomethacin). However, the extract was non-toxic to Vero cells with LCso value of> 1.00 mg/mL but highly toxic to HepG2 cells with LCso: 0.08 mg/mL. The selectivity index of 12.50 was recorded. The presence of phenolics/ carboxylic acids were a lso confirmed in the extract, the result of the antiox, antidiabetic and antinflammatory activities of Po HG suggested that the phenols extract may be useful in the management of oxidative stress induce diseases, type 2 diabetes and asthma. It is also safe for use and its antiproliferative activities can be exploited in search for anticancer agents. A new xanthene derivative Hermannol (9-(7-methyloctyl)-9H- xanthene-2,3-diol) was isolated from the roots of Hermannia geniculata. The structure was e lucidated by ana lysis of their ID, 2D NMR, MS and IR spectroscopic data. The compound displayed good antioxidant and antidiabetic activities. In conclusion, it is evident from the study that d ifferent crude extracts of H geniculata roots and its bioactive constituents (tlavono ids and phenols), the isolated compound (Hermanno l) is nontoxic and possess varied degree of antiox idant, antidiabetic, antiiflammatory and antiproliferative activities which can be exploited for new drug development.