Ebenezer O. Farombi

Antioxidative and Chemopreventive Properties of Vernonia amygdalina and Garcinia biflavonoid

Recently, considerable attention has been focused on dietary and medicinal phytochemicals that inhibit, reverse or retard diseases caused by oxidative and inflammatory processes. Vernonia amygdalina is a perennial herb belonging to the Asteraceae family. Extracts of the plant have been used in various folk medicines as remedies against helminthic, protozoal and bacterial infections with scientific support for these claims. Phytochemicals such as saponins and alkaloids, terpenes, steroids, coumarins, flavonoids, phenolic acids, lignans, xanthones, anthraquinones, edotides and sesquiterpenes have been extracted and isolated from Vernonia amygdalina. These compounds elicit various biological effects including cancer chemoprevention. Garcinia kola (Guttiferae) seed, known as “bitter kola”, plays an important role in African ethnomedicine and traditional hospitality. It is used locally to treat illnesses like colds, bronchitis, bacterial and viral infections and liver diseases. A number of useful phytochemicals have been isolated from the seed and the most prominent of them is the Garcinia bioflavonoids mixture called kolaviron. It has well-defined structure and an array of biological activities including antioxidant, antidiabetic, antigenotoxic and hepatoprotective properties. The chemopreventive properties of Vernonia amygdalina and Garcinia biflavonoids have been attributed to their abilities to scavenge free radicals, induce detoxification, inhibit stress response proteins and interfere with DNA binding activities of some transcription factors.

Monosodium glutamate-induced oxidative damage and genotoxicity in the rat: modulatory role of vitamin C, vitamin E and quercetin

Monosodium glutamate (MSG) continues to function as a flavor enhancer in West African and Asian diets. The present study examines the modulatory effects of dietary antioxidant vitamin C (VIT C), vitamin E (VIT E) and quercetin on MSG-induced oxidative damage in the liver, kidney and brain of rats. In addition, the effect of these antioxidants on the possible genotoxicity of MSG was investigated in a rat bone marrow micronuclei model. MSG administered intraperitoneally at a dose of 4 mg/g body wt markedly increase malondialdehyde (MDA) formation in the liver, the kidney and brain of rats. Simultaneous administration of VIT C, VIT E and quercetin to MSG-treated rats significantly reduced this increase in MDA induced by MSG. VIT E reduced lipid peroxidation most in the liver followed by VIT C and then quercetin, while VIT C and quercetin showed a greater ability to protect the brain from membrane damage than VIT E. The decreased glutathione (GSH) level elicited by MSG in the three organs corresponded with marked increase in the activity of glutathione-S-transferase (GST). While MSG increased (P B / 0.001) the activities of superoxide dismutase and catalase in the liver, it decreased significantly the activities of these enzymes in the kidney and the brain. The three antioxidants were effective at ameliorating the effects of MSG on GSH levels and the enzymes in the three organs examined. While MSG increased the activity of glucose-6-phosphatase in the liver and kidneys of rats (P B / 0.001), the activity of the enzyme was abysmally low in the brain. There were marked increases in the activities of alanine aminotransferase, aspartate aminotransferase and g-glutamyl transferase in rats treated with MSG. The antioxidants tested protected against MSG-induced liver toxicity significantly. MSG at a dose of 4 mg/g significantly (P B / 0.01) induced the formation of micronucleated polychromatic erythrocytes (MNPCEs). Co-treatment of rats with VIT C and quercetin inhibited the induction of MNPCEs by MSG (P B / 0.001). VIT E failed to protect against MSG-induced genotoxicity. The results indicate that dietary antioxidants have protective potential against oxidative stress induced by MSG and, in addition, suggest that active oxygen species may play an important role in its genotoxicity.

Possible anti‐atherogenic effect of kolaviron (a Garcinia kola seed extract) in hypercholesterolaemic rats

1. The hypolipidaemic effect of kolaviron, a mixture of Garcinia biflavonoid 1 (GB1), Garcinia biflavonoid 2 (GB2) and kolaflavanone, used in the treatment of various ailments in southern Nigeria, was investigated in rats. The ability of Questran (Bristol‐Myers Squibb, Hounslow, UK), a hypolipidaemic therapeutic drug, to attenuate hypercholesterolaemia in rats was also examined.

Hepatoprotective effects of Vernonia amygdalina (astereaceae) in rats treated with carbon tetrachloride.

The possible modulatory effect of methanolic extract of Vernonia amygdalina (MEVA), a plant widely consumed in the tropics and used locally in the treatment of fever, jaundice, stomach disorders and diabetes on the toxicity of CCl(4), was investigated in male rats. Oral administration of CCl(4) at a dose of 1.2g/kg body weight 3 times a week for 3 weeks significantly induced marked hepatic injury as revealed by increased activity of the serum enzymes ALT, AST, SALP and gamma-GT. Methanolic extract of V. amygdalina administered 5 times a week for 2 weeks before CCl(4) treatment at 250 and 500 mg/kg doses of the extract ameliorated the increase in the activities of these enzymes. Likewise the methanolic extract of V. amygdalina reduced the CCl(4)-induced increase in the concentrations of cholesterol, triglyceride and phospholipid by 37.8%, 30.6% and 8.5%, respectively, and a reduction in the cholesterol/phospholipids ratio. These parameters were however increased at 750 mg/kg extract pretreatment. CCl(4)-induced lipid peroxidation was likewise attenuated by 57.2% at 500 mg/kg dose of the methanolic extract of V. amygdalina. Similarly, administration of the extract increased the activities of the antioxidant enzymes: superoxide dismutase, glutathione S-transferase and reduced glutathione concentration significantly at 500 mg/kg (P<0.05) and catalase activity at 500-1000 mg/kg doses. These results suggest that methanolic extract of V. amygdalina leaves posseses protective effect against CCl(4)-induced hepatotoxicity by the antioxidant mechanism of action.

Evaluation of Antioxidant and Free Radical Scavenging Capacities of Some Nigerian Indigenous Medicinal Plants

Methanolic extracts of 10 selected Nigerian medicinal plants-Psidium guajava, Alstonia boonei, Cassia alata, Newbouldia laevis, Spondias mombin, Globimetula cupulatum, Chromolaena odorata, Securidaca longepedunculata, Ocimum gratissimum, and Morinda lucida-widely used in ethnomedicine, were assessed for phytochemical constituents and antioxidant and free radical scavenging activities using seven different antioxidant assay methods. Phytochemical screening gave positive tests for steroids, terpenoids, and cardiac glycosides, alkaloids, saponins, tannins, and flavonoids contained in the extracts. P. guajava contained the highest amount of total phenolics (380.08 +/- 4.40 mg/L gallic acid equivalents), and the highest amounts of total flavonoids were found in the leaf extracts of C. alata (275.16 +/- 1.62 microg/mL quercetin equivalents [QE]), C. odorata (272.12 +/- 2.32 microg/mL QE), and P. guajava (269.72 +/- 2.78 microg/mL QE). Percentage 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity was highest in S. mombin (88.58 +/- 3.04%) and P. guajava (82.79 +/- 2.84%) and compared with values obtained for ascorbic acid and gallic acid. All the extracts, generally, had low nitric oxide radical scavenging activities, and G. cupulatum had the highest hydroxyl radical scavenging activity (63.84 +/- 0.97%). The extracts in general demonstrated high lipid peroxidation inhibitory activity, with only M. lucida (38.74 +/- 1.99%) and A. boonei (47.16 +/- 0.59%) being exceptions. The reductive potential was highest in P. guajava (0.79 +/- 0.04) and least in S. longepedunculata (0.26 +/- 0.00). DPPH assay correlated well with total phenolic contents (r(2) = 0.76) and reductive potential (r(2) = 0.81) and fairly with lipid peroxidation inhibitory activity (r(2) = 0.51). There was a good correlation between total phenolic contents and reductive potential (r(2) = 0.79) and a fair correlation between total phenolic contents and lipid peroxidation inhibitory activity (r(2) = 0.55). These results suggest that the methanolic extracts of the studied plant parts possess significant antioxidant and radical scavenging activities that may be due to the phytochemical content of the plants and as such make them potential candidates as natural chemoprophylactic agents. In addition, multiple assay methods should be used in comparing antioxidant capacities of samples to have a reliable result.

Tetracycline-induced reproductive toxicity in male rats: Effects of vitamin C and N-acetylcysteine

Tetracycline, a broad-spectrum antibiotic employed clinically in the treatment of bacteria infections, is known to cause a number of biochemical dysfunctions and suspected to induce testicular damage to animals and humans, but there is paucity of data on its effect and mechanism of action on the male reproductive system. The present study therefore evaluates its spermatotoxic and testicular toxicity in male rats and the chemoprotective effects of Vitamin C (Vit C) and N-acetylcysteine (NAC). Tetracycline was administered orally at the dose level of 28.6 mg/kg body weight per day in two equal divided doses (12h interval). Vit C and NAC were also administered orally to the rats at doses of 200 and 50 mg/kg body weight per day, respectively, for the 14 days of the experiment. While there was no change in the body weights of rats, tetracycline administration caused significant decrease in the relative weights of testis, epididymis and seminal vesicles (P<0.05). Administration of tetracycline caused a reduction in the epididymal sperm motility, percentage of live spermatozoa, sperm count, and an increase in abnormal sperm morphology, as well as induction of adverse histopathologic changes in the testes. While Vit C and NAC significantly mitigated the toxic effect of tetracycline on sperm parameters, the antioxidants did not improve the adverse histopathologic changes induced by antibiotic. Treatment of rats with tetracycline significantly decreased the activities of superoxide dismutase, catalase (CAT), glucose-6-phosphate dehydrogenase, glutathione-S-transferase (GST) and the levels of GSH and serum testosterone, while the activity of gamma-glutamyltranspeptidase and the formation of malondialdehyde (MDA) increased. Both Vit C and NAC significantly attenuated the toxic effects of tetracycline to the antioxidant and testicular marker enzymes as well as markers of oxidative stress. Collectively, the results suggest that therapeutic dose of tetracycline elicits spermatotoxic and testicular toxicity in male rats through induction of oxidative stress. The chemoprotective effects of Vit C and NAC during tetracycline treatment suggest that these antioxidants may find clinical application in cellular damage involving reactive oxygen species (ROS).

Hypolipidemic and antioxidant effects of ethanolic extract from dried calyx of Hibiscus sabdariffa in alloxan-induced diabetic rats.

The present study investigated the hypolipidemic and antioxidant effects of ethanolic extract of Hibiscus sabdariffa L (HSE) in rats treated with alloxan. The results were compared with the standard hypolipidemic drug lovastatin. HSE at doses of 100 and 200 mg/kg elicited dose‐dependent effects on the biomarkers evaluated. In alloxan‐treated rats, HSE at the dose of 200 mg/kg significantly attenuated the elevated blood glucose concentration by 57%. Lovastatin (10 mg/kg) similarly reduced the glucose level in alloxan‐treated rats by 48%. HSE reduced the alloxan‐induced increases in cholesterol, very low‐density lipoprotein cholesterol (VLDL‐C), low‐density lipoprotein cholesterol (LDL‐C) and atherogenic index by 29%, 36%, 40%, and 32%, respectively while lovastatin decreased the alloxan‐induced increases in the parameters by 25%, 23%, 28%, and 31%, respectively. HSE (200 mg/kg) and lovastatin (P < 0.01) decreased the alloxan‐induced increases in the lipid profiles both in the liver and the kidneys. HSE at 200 mg/kg attenuated the alloxan‐induced decrease in the activities of superoxide dismutase (SOD), catalase (CAT) and the level of glutathione (GSH) by 36%, 44%, and 64% in the liver and by 20%, 43%, and 85% in the kidney of rats. Lovastatin similarly increased SOD, CAT and GSH by 32%, 29%, and 64% in the liver and by 17%, 26%, and 73% in the kidney of alloxan‐treated rats. HSE (200 mg/kg) significantly decreased the alloxan‐mediated increase in malondialdehyde (MDA) and protein carbonyl (PC) levels in the liver by 44% and 43% and in the kidneys by 45% and 38%, respectively, while lovastatin decreased the alloxan‐induced elevation in MDA and PC in the liver by 42% and 41% and in the kidney by 45% and 33%, respectively. While HSE at a dose of 200 mg/kg and lovastatin normalized the activity of phosphatidate phosphohydrolase in the liver, the extract and lovastatin did not elicit significant changes in the kidney enzyme activity in rats treated with alloxan. Overall, our data demonstrate that HSE possesses strong hypolipidemic as well as antioxidant properties in alloxan‐induced diabetic rats and as such Hibiscus sabdariffa could be useful in preventing the development of atherosclerosis and possible related cardiovascular pathologies associated with diabetes.

Protective effects of kolaviron and quercetin on cadmium-induced testicular damage and endocrine pathology in rats

This study evaluated the effects of kolaviron, a biflavonoid from Garcinia kola seed, and quercetin on cadmium‐induced reproductive toxicity in rats. Adult male rats were administered with either cadmium (15 mg kg−1) alone or in combination with kolaviron (200 mg kg−1) or quercetin (10 mg kg−1) daily for 5 days. Cadmium‐treated rats showed (P < 0.05) decrease in the body weight gain, testis and epididymis weights. However, upon co‐administration of kolaviron or quercetin, these changes were significantly reversed in cadmium‐treated rats. Also, administration of kolaviron or quercetin significantly prevented cadmium‐mediated decrease in sperm motility and epididymal sperm concentration and reversed the increased level of sperm abnormality to near control. In testes and sperm, cadmium treatment resulted in significant decrease in the activities of superoxide dismutase, catalase and glutathione peroxidase, whereas it increased glutathione S‐transferase activity as well as hydrogen peroxide and malondialdehyde levels. While plasma levels of triiodothyronine and tetraiodothyronine remained unaffected, the levels of testosterone, luteinising hormone and follicle stimulating hormone were decreased in cadmium‐treated rats. Cadmium treatment caused mild congestion of interstitial vessels and oedema in the testes. Taken together, kolaviron and quercetin inhibited the adverse effects of cadmium on the antioxidant enzymes, markers of oxidative stress, endocrine and testicular structure in rats.

Kolaviron, a Natural Antioxidant and Anti‐Inflammatory Phytochemical Prevents Dextran Sulphate Sodium‐Induced Colitis in Rats

The beneficial effects of kolaviron, a natural biflavonoid from the seeds of Garcinia kola, have been attributed mainly to its antioxidant and anti‐inflammatory effects. This study investigated these effects on dextran sulphate sodium (DSS)‐induced ulcerative colitis in rats. Sulfasalazine served as standard reference in this study. Kolaviron and sulfasalazine were separately co‐administered orally at 200 mg/kg and 500 mg/kg, respectively, to dextran sulphate sodium‐exposed rats for 5 days. The result indicated that kolaviron or sulfasalazine significantly prevented DSS‐induced body weight loss as well as the incidence of diarrhoea and bleeding in DSS‐exposed rats. Kolaviron suppressed the DSS‐mediated increase in colonic nitric oxide concentration and myeloperoxidase activity and significantly prevented the increase in inflammatory mediators, interleukin‐1β and tumour necrosis factor alpha, in the colon of DSS‐treated rats. The significant depletion in colonic antioxidant status in rats exposed to DSS alone was evident by marked reduction in colonic catalase and glutathione S‐transferase activities as well as glutathione content, leading to elevated hydrogen peroxide and lipid peroxidation levels. Histopathologically, DSS alone resulted in severe epithelial erosion, total absence of goblet cells, destruction of the crypts, necrotic and distorted glands, accompanied by marked cellular mononuclear cells infiltration. However, administration of kolaviron and sulfasalazine ameliorated DSS‐induced colitis by increasing the antioxidant status decreased hydrogen peroxide and lipid peroxidation levels and attenuated the adverse effect of DSS on colon architecture. In conclusion, the anti‐colitis effect of kolaviron is related to its intrinsic anti‐inflammatory and anti‐oxidative properties.

Induction of oxidative damage in the testes and spermatozoa and hematotoxicity in rats exposed to multiple doses of ethylene glycol monoethyl ether

The effects of ethylene glycol monoethyl ether (EGEE) on the antioxidant systems of the testes and epididymal spermatozoa were investigated in rats at dose levels of 0, 100, 200 and 400 mg kg-1 body weight (bw) administered orally by gavage for 14 consecutive days. The bw gain of the EGEE-treated rats decreased significantly at 200 and 400 mg kg- 1 bw compared with the control group. There were no significant changes in the weights of the testes, epididymis, seminal vesicles and prostate glands of the EGEE-treated rats. In the testes, while EGEE treatment resulted in significant decrease in glutathione (GSH) level, superoxide dismutase (SOD) and catalase (CAT) activities, it markedly increased the malondialdehyde (MDA) level, glutathione-S-transferase (GST) and lactate dehydrogenase (LDH) activities at 200 and 400 mg kg-1 dose levels but vitamin C content remained unaffected in all the groups. In the spermatozoa, administration of EGEE caused significant decrease in the activities of CAT, GST and LDH as well as in the levels of vitamin C and GSH but significantly increased the MDA level and SOD activity compared with the control rats. Histopathological examination showed severe degeneration of the testes, such as generalized erosion and necrosis of the germinal epithelium of the testes, but mildly affected the epididymis at 400 mg kg-1 dose only. Data on spermatozoa analysis of EGEE-treated rats revealed significant decrease in the epididymal spermatozoa number, testicular spermatozoa number, daily spermatozoa production and spermatozoa motility but significantly increased the total spermatozoa abnormalities without affecting the spermatozoa live-dead ratio at all dose levels when compared with the control group. Results of haematological examination showed that white blood cells (WBC), platelets neutrophils and mean corpuscular haemoglobin concentration (MCHC) were significantly lower whereas lymphocytes were increased in 200 and 400 mg/kg EGEE-exposed rats than in the controls. EGEE at 100 mg/kg bw produced minor effect on haematological parameters but adversely affected testes and spermatozoa. In summary, short term administration of EGEE is hematotoxic and gonadotoxic and its effects on male reproduction could be due to the induction of oxidative stress in testes and spermatozoa.