Effect of Aqueous Extract of Unripe Musa Paradisiaca Linn on Parameters Affecting Reproduction in Rats.

OBJECTIVE: Aqueous extract of unripe Musa paradisiaca fruit is commonly used for the treatment of ulcers in eastern Nigeria. This study aimed to assess the acute and subacute effects of an aqueous extract of unripe fruit on male and female fertility in rats. METHODS: Aqueous extracts obtained by maceration were analyzed for acute and subacute toxicity and for the presence of phytochemical constituents using standard procedures. The extract (100, 500, and 1000 mg/kg) was administered daily to rats of both sexes for 28 d. Blood samples collected on days 0 and 28 were assessed for follicle-stimulating hormone (FSH), luteinizing hormone (LH), catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA). Testes and ovaries were harvested for histopathological analysis. Sperm were also collected to determine the sperm count and motility. RESULTS: Phytochemical screening revealed the presence of saponins, tannins, alkaloids, and resins. After an oral dose of up to 5000 mg/kg, there were no deaths in the acute toxicity test. The extract (500 mg/kg) significantly (P < .05) enhanced sperm count and motility relative to the untreated control; significantly (P < .05) reduced SOD, CAT, and glutathione levels, while significantly (P < .05) elevated LH, FSH, and MDA levels in male and female rats. Histological examination revealed significant structural damage to the ovaries. CONCLUSION: Unripe Musa paradisiaca fruit exhibited an adverse toxicological profile following prolonged administration and caused oxidative stress in rodents.

Influence of Nigerian Jelly Ear Culinary-Medicinal Mushroom, Auricularia auricula-judae (Agaricomycetes), on Humoral and Cellular Immunity.

The fruiting body of Auricularia auricula-judae has received attention in folk medicine due to its possible medicinal values. Therefore, this study evaluated the immunomodulatory effects of the hot aqueous extract (AAAJ) and the ß-D-glucan-rich polysaccharide fraction of A. auricula-judae (BGPA) on specific and nonspecific humoral and cell mediated immune responses in immunocompetent and immunosuppressed mice. Oral supplementation with AAAJ or BGPA (100, 200, or 400 mg/kg) produced significantly high titers of total OVA specific or TT specific IgG1 and IgG2a compared with the levels in untreated control. Oral administration of AAAJ or BGPA (100, 200, or 400 mg/kg) evoked a significant increase in carbon clearance at all doses, indicating stimulation of the reticuloendothelial system, and potentiated the delayed-type hypersensitivity reaction induced by sheep red blood cells (SRBC) compared with the untreated mice. Total lymphocyte count, neutrophil count, and lymphocytes count increased significantly (P < 0.05) at all doses, following acute administration of AAAJ or BGPA (100, 200, or 400 mg/kg), showing increased protection toward cyclophosphamide induced myelosuppression compared with the untreated negative control group. In the hemolytic complement assay, AAAJ and BGPA at all doses significantly (P < 0.05) inhibited the hemolytic activity of the complement proteins on the sensitized SRBC. The present study reveals that the extract holds promise as an immunomodulatory agent and strengthens the rationale for its use in traditional medicine.

Anti-plasmodial and anti-inflammatory activities of cyclotide-rich extract and fraction of Oldenlandia affinis (R. & S.) D.C. (Rubiaceae).

BACKGROUND: Oldenlandia affinis, commonly called 'kalata-kalata', a versatile plant used locally to treat malaria fever in some parts of sub-Saharan Africa was investigated for anti-plasmodial and anti-inflammatory activities. OBJECTIVE: The study was designed to evaluate the antiplasmodial as well as anti-inflammatory activities of whole extract and cyclotide-rich fraction of Oldenlandia affinis. METHOD: The dichloromethane-methanol extract (ODE) of the plant, O. affinis was investigated for suppressive and curative antiplasmodial activities against Plasmodium berghei in mice. ODE and the cyclotide-rich fraction (CRF) was investigated for chronic and acute anti-inflammatory activities in rat models of inflammation. Inhibition of pro-inflammatory mediators was studied in RAW264.7 macrophages. RESULTS: ODE exhibited significant (p<0.05) reduction in mean parasitaemia in both the suppressive and curative models of Plasmodium berghei infection in mice.Administration of ODE(100, 200, or 400 mg/kg) and CRF (100, 200, or 400 mg/kg) produced significant inhibition of rodent models of acute and chronic inflammation . This observation is supported by the significant (P<0.05) inhibition of pro-inflammatory mediators, inducible nitric oxide (iNO) and tumour necrosis factor-alpha (TNF-α), and the reactive radical scavenging activities in RAW264.7 macrophages. CONCLUSION: These findings could explain, at least in part, the successes reported in the use of the herb, Oldenlandia affinis in the traditional treatment of malaria fever.

In vitro and in vivo antioxidant potentials of Alchornea floribunda leaf extract, fractions and isolated bioactive compounds.

OBJECTIVE: Alchornea floribunda leaves are widely used in ethnomedicine for the management of immuno-inflammatory disorders. We investigated the in vivo and in vitro antioxidant activity of the leaf extract, fractions and isolated compounds of A. floribunda. MATERIALS AND METHODS: The ethyl acetate fraction of the methanol leaf extract was subjected to several chromatographic separations to isolate compounds 1-4. The structures of the isolated compounds were elucidated by a combination of 1D and 2D NMR and mass spectrometry. Oxidative stress was induced with carbon tetrachloride (CCl4). Further analysis on the isolated phenolic compounds were done using 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and hydrogen peroxide scavenging activity tests. RESULTS: The ethyl acetate fraction at 200 mg/kg produced significant (p<0.05) elevations of catalase enzyme activity and a significant (p<0.05) reduction in serum malondialdehyde. The chemical investigation of the ethyl acetate fraction led to the isolation of three flavans, (-) cathechin (1), (-) epicathechin (2), (+) epicathechin (3) and a flavanone, 2R, 3R dihydroquercitin (4). In hydrogen peroxide scavenging assay, (-) epicathechin exhibited an EC50 value of 8 µg/ml, similar to the standard ascorbic acid (EC50 = 8 µg/ml). (-) epicathechin showed scavenging of DPPH radical with EC50 value of 19 µg/ml while in the FRAP assay, it had EC50 value of 46 µg/ml which was lower than that of the standard, ascobic acid (EC50 = 66 µg/ml). CONCLUSION: The medicinal uses of A. floribunda may be due to the antioxidant activities of its phenolic compounds.

Modulation of intracellular expression of IFNγ and IL-2 in culture of splenic T lymphocytes by some flavonoid glycosides of Alchornea floribunda.

Context Alchornea floribunda Müll. Arg. (Euphorbiaceae) leaves are widely used in ethnomedicine for the management of rheumatism, arthritis and toothache. Objective In this study, flavonoid glycosides isolated from Alchornea floribunda were screened for their effect on the intracellular expression of interferon-gamma (IFNγ) and interleukin-2 (IL-2) type-1 cytokines. Materials and methods Chromatographic purification of the ethyl acetate fraction of the methanol leaf extract led to the isolation of seven flavonoid glycosides (1-7). Their structures were elucidated by 1D and 2D nuclear magnetic resonance and mass spectrometry. Splenocytes were treated with graded concentrations of the compounds (6.25-25 µg/mL) and incubated for 24 h. Thereafter, their effect on the expression of IFNγ and IL-2 by CD4(+ )and CD8(+ )T-lymphocytes was evaluated using intracellular cytokine staining and FACS analysis. Results Compounds 1-7 (6.25-25 µg/mL) caused the up-regulation of activated CD8(+ )(57.85-72.45% versus 57.85% for untreated control) and, to a lesser extent, activated CD4(+ )(3.21-7.21% versus 2.75% for the untreated control) T-lymphocytes that were both largely interferon-gamma-releasing in treated mouse T lymphocytes relative to untreated control. FACS data analysis showed that stimulation with all the compounds increased the proportion of CD8(+)/IFNγ(+ )and CD4(+)/IFNγ(+ )T lymphocytes up to two-fold when compared with the cells in untreated control wells. Intracellular IL-2 secretion by treated T cells was not detected. Conclusion This recorded T-lymphocyte-specific immune-modulatory property may contribute to explain in part the dynamics associated with the ethnomedicine of Alchornea floribunda, and may find relevance as a necessary cellular immune response precursor to infection-associated disease management.

Landolphia owariensis leaf extracts reduce parasitemia in Plasmodium berghei-infected mice.

Context Landolphia owariensis P. Beauv. (Apocyanaceae) leaf is used in southeast Nigeria to treat malaria. Objective This study evaluated the antiplasmodial activity of L. owariensis leaf extract and fractions, also the phytoconstituents were standardized and analyzed. Methods The effects of daily, oral administrations of 200, 400 and 800 mg/kg of L. owariensis leaf extract (LOE), its hexane (LOHF), ethyl acetate (LOEF) and methanol (LOMF) fractions on early, established and residual infections in Plasmodium berghei-infected albino mice were evaluated in vivo. The extract and fractions were subjected to phytochemical analysis and HPLC fingerprinting, and the acute toxicity of LOE was evaluated. Results The extract and fractions elicited 29-86, 18-95 and 75-96% significant (p < 0.001) suppression of parasitemia in early, established and residual infections, respectively. The ED50 values for suppressive activity of LOE, LOHF, LOEF and LOMF were 266.56, 514.93, 392.95 and 165.70 mg/kg, respectively. The post-day 30-survival index was 16.7-50, 16.7, 16.7-66.7 and 50-83.3% for LOE, LOHF, LOEF, and LOMF, respectively. Extract-treated mice significantly (p < 0.001) gained weight and had reduced mortality compared with negative control (untreated) mice. An oral LD50 value >5000 mg/kg in mice was established for LOE. The LOMF showed the greatest antiplasmodial activity in all the models, suggesting that the antimalarial activity of the plant may be attributed to alkaloids, flavonoids, saponins and tannins present in the fraction. Conclusion Results demonstrate the antiplasmodial activity of L. owariensis leaf, and provide a pharmacological rationale for its ethnomedicinal use as an antimalarial agent.

New neolignan glycoside and an unusual benzoyl malic acid derivative from Maytenus senegalensis leaves.

Further investigation of the methanol leaf extract of Maytenus senegalensis led to the isolation of six compounds, including mayselignoside (1) and an unusual benzoyl malic acid derivative, benzoyl R-(+)-malic acid (2). Two known lignan derivatives (+)-lyoniresinol (3) and (-)-isolariciresinol (4), a known neolignan derivative dihydrodehydrodiconiferyl alcohol (5) and the triterpenoid, ß-amyrin (6) were also isolated. The structures of these compounds were elucidated by a combination of 1D and 2D NMR and mass spectroscopy. All compounds were tested for cytotoxicity against mouse lymphoma cell line (L5178Y) and for antimicrobial activity against strains of bacteria and fungi. None of the compounds showed promising cytotoxic and/or antimicrobial activities.

Hepato- and nephroprotective activities of a Nigerian local king tuber oyster mushroom, Pleurotus tuberregium (higher Basidiomycetes), in chemically induced organ toxicities in rats.

Chronic kidney disease, end-stage renal failure, and liver diseases are increasing worldwide and constitute a huge burden on health care costs, with attendant high morbidity and debility. Despite advances in modern medicine, there are still no licensed drugs that satisfactorily restore lost kidney or hepatic functions. In this study the chemoprotective effects of the hot aqueous extract of a local edible oyster mushroom, Pleurotus tuberregium (APTR), was evaluated in experimental liver and kidney toxicities. The effect of APTR on carbon tetrachloride (CCl4)- and paracetamol (PCM)-induced hepatotoxicity in rats was investigated by determining serum concentrations of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP). Short-term oral treatment with APTR (100 and 250 mg/kg) significantly reduced (P < 0.05) the increased concentrations of AST, ALT, and ALP induced in both PCM and CCl4 models of liver toxicity. APTR (100 and 250 mg/kg) decreased the mean serum AST concentrations by as much as 73.00% and 99.37%, respectively, in PCM-treated rats. Nephroprotection was assessed by determining the serum concentrations of creatinine and urea, as well as antioxidant enzymes, in kidney tissue homogenates after a repeated high dose of gentamicin. APTR (100 and 250 mg/kg) produced a significant decrease (P < 0.05) in the escalated serum concentrations of creatinine and urea by as much as 48.36% and 41.53%, respectively, compared to control. Similarly, levels of the antioxidant enzymes catalase, glutathione peroxidase, and superoxide dismutase in kidney tissue were increased in a dose-related manner in groups that received oral APTR supplementation. The results of this study suggest that the consumption of our local edible mushroom, P. tuberregium, could, in addition to its high nutritive value, protect the liver and kidneys from oxidative damage caused by drugs and toxicants such as CCl4 and high doses of gentamicin and PCM.

beta-Amyrin and alpha-amyrin acetate isolated from the stem bark of Alstonia boonei display profound anti-inflammatory activity.

CONTEXT: Alstonia boonei De Wild (Apocyanaceae) is used in ethnomedicine for the management of malaria, ulcer, rhematic pain, toothache, and inflammatory disorders. OBJECTIVE: To investigate the anti-inflammatory potential of ß-amyrin and α-amyrin acetate isolated from the stem bark of Alstonia boonei using animal models. MATERIALS AND METHODS: Chromatographic purification of the crude methanol extract led to the isolation and structure elucidation of ß-amyrin and α-amyrin acetate. Their anti-inflammatory activities were evaluated in rodents using egg albumen-induced paw edema and xylene-induced ear edema models. The gastric ulcerogenic, in vivo leucocyte migration, and RBC membrane stabilization tests were also investigated. RESULTS: α-Amyrin acetate at 100 mg/kg showed significant (p < 0.05) inhibition of egg albumen-induced paw edema with % inhibition of 40 at the 5th hour. Oral administration up to 100 mg/kg did not produce significant (p > 0.01) irritation of the gastric mucosa while significant (p < 0.01) ulceration was recorded for indomethacin at 40 mg/kg compared with the negative control. At 100 µg/mL, both ß-amyrin and α-amyrin acetate inhibited heat-induced hemolysis to as much 47.2 and 61.5%, respectively, while diclofenac sodium (100 µg/mL) evoked only 40.5% inhibition. Both compounds at 100 µg/ear produced significant (p < 0.01) inhibition of ear edema in mice by 39.4 and 55.5%, respectively. Also at 100 mg/kg (p.o.) α-amyrin acetate evoked 60.3% reduction in total leucocyte count and significant (p < 0.05) suppression (47.9%) of neutrophil infiltration. DISCUSSION AND CONCLUSION: This study generally provided evidence of profound anti-inflammatory activity of ß-amyrin and α-amyrin acetate isolated from the Alstonia boonei stem bark.

Isolation and structure elucidaton of polyphenols from Loranthus micranthus Linn. parasitic on Hevea brasiliensis with antiinflammatory property.

The present study was carried out to evaluate the anti-inflammatory activities of polyphenols isolated from the leaves of mistletoe (Loranthus micranthus Linn.) parasitic on Hevea brasiliensis. The anti-inflammatory properties of the isolated compounds were evaluated on the basis of their ability to inhibit the production of nitric oxide (NO) and tumuor necrosis factor-α (TNF-α) in lipopolysaccharide (LPS) activated RAW 264.7 mouse macrophages. Semi-preparative HPLC separation of the ethyl acetate (EtOAc) and butanol (n-BuOH) fractions of the leaves of mistletoe (Loranthus micranthus Linn) parasitic on Hevea brasiliensis led to the isolation of four polyphenols: 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin (TMECG) (1); (-)-epicatechin-3-O-(3″-O-methyl)-gallate (ECG3″Me) (2); rutin (3) and peltatoside (4). Compounds 1-4 were isolated for the first time from this plant while 1 was isolated for the first time in nature. These compounds (1-4) were readily identified by comparison of their spectroscopic data with those reported in the literature. The polyphenols proved to have anti-inflammatory activity as evidenced by the suppression of inducible nitric oxide (iNO) and cytokine (TNF-α) levels in the culture supernatant of lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophages. However, the study showed that the quercetin diglycosides showed stronger inhibition of proinflammatory mediators than the epicatechin derivates. These data provide evidence that polyphenolic compounds isolated from the mistletoe parasitic on Hevea brasiliensis may contribute to its anti-inflammatory properties by inhibiting the expression of inducible nitric oxide and proinflammatory cytokines such as tumour necrosis factor-α.

Three hydroxylated lupeol-based triterpenoid esters isolated from the Eastern Nigeria mistletoe parasitic on Kola acuminata.

Column fractionation and purification of an n-hexane fraction led to the isolation of three lupeol-based triterpenoid esters from the leaves of the Eastern Nigeria mistletoe, Loranthus micranthus Linn parasitic on a local kola nut tree, Kola acuminata. These three compounds were adequately characterised using a combination of UV/visible, IR, NMR (¹³C-NMR and ¹H-NMR), DEPT, MS and two-dimensional correlation (H-H COSY, Hetero-nuclear Single Quantum Correlation (HSQC), HMBC, NOE and NOESY) studies as 7ß,15α-dihydroxyl-lup-20(29)-ene-3ß-esters of palmitic (I), stearic (II) and eicosanoic acids (III). The characterisation of other isolated compounds is ongoing. Remarkably, this is the first report of the existence of fatty acid esters of an unusual 7ß,15α-dihydroxylated lupeol in the Eastern Nigeria mistletoe. These isolated compounds might contribute in part to the numerous established bio-activities of the Eastern Nigeria mistletoes.

A novel sesquiterpene acid and an alkaloid from leaves of the Eastern Nigeria mistletoe, Loranthus micranthus with potent immunostimulatory activity on C57BL6 mice splenocytes and CD69 molecule.

CONTEXT: The Eastern Nigeria mistletoe, Loranthus micranthus Linn. (Loranthaceae), is used in the treatment of several diseases including immune-modifying diseases and thus there is a need to identify the immunoactive constituents. OBJECTIVE: This research isolated and characterized the immunoactive constituents in the Eastern Nigeria mistletoe. MATERIALS AND METHODS: Bioassay-guided fractionation was employed in the isolation and purification of the constituents. The characterized compounds were screened for immunostimulatory activities on isolated C57BL/6 mice splenocytes and early activation marker, CD69 at concentrations of 10, 25, and 100 µg/mL using flow cytometry techniques and compared to lipopolysaccharide (LPS; 10 µg/mL) and concanavalin A (ConA; 2 µg/mL) as standards. RESULTS: Two compounds, a novel sesquiterpene, 2, 3-dimethoxy-benzo [a, b] cyclopentenyl-3',3',5'-trimethyl pyran-4-carboxylic acid, and a known alkaloid, lupinine were isolated and characterized. The compounds (25 µg/mL) showed statistically significantly (p < 0.05) stimulatory activity on the splenocytes with values of 56.34 ± 0.26% and 69.84 ± 0.19%, respectively, compared to 7.58 ± 0.42% recorded for the unstimulated control. Similarly, the CD69 expression assay showed immunostimulation with statistically significant values (p < 0.05) of 2.31 ± 0.07% and 2.71 ± 0.03%, respectively, compared to 1.69 ± 0.05% recorded for the nonstimulated control. DISCUSSION: These data suggest that the isolated compounds possess immunomodifying abilities. In addition, the activation of the CD69 molecule is possibly one of its mechanisms of action. CONCLUSION: These compounds may be responsible in part, for the immunostimulatory activities already established for the Eastern Nigeria mistletoes.

Immunomodulatory activity of a lupane triterpenoid ester isolated from the eastern Nigeria mistletoe, Loranthus micranthus (Linn)

OBJECTIVE@#To provide further evidence for the ethnomedicinal use of the Eastern Nigeria mistletoe, Loranthus micranthus (L. micranthus), as an immunostimulant.@*METHODS@#Solvent fractions from the crude extract of the mistletoe plant was obtained and screened by the cell mediated delayed type hypersensitivity reaction (DTHR) model in mice. Then the immunomodulatory potentials of a major lupane triterpenoid ester isolated from an active hexane fraction of the Eastern Nigeria mistletoe was investigated. Three lupeol-based triterpenoid esters: 7β 15α-dihydroxyl-lup-20(29)-ene-3β-palmitate (I), 7β, 15α-dihydroxyl-lup-20(29)-ene-3β-stearate (II) and 7β, 15α-dihydroxyl-lup-20(29)-ene-3β-decadecanoate (III) were isolated from the plant leaves epiphyting on a local kola nut tree and were characterized. Compound 1 was subjected to cell proliferation studies using C57Bl/6 splenocytes at three dose levels (5, 25 and 100 μg/mL) in presence of controls. Furthermore, the effect of this compound on IL-8 receptor expression was evaluated at three doses (1, 5 and 10 μg/mL) using the real time polymerase chain reaction assay.@*RESULTS@#This triterpenoid ester produced some enhancement of the splenocytes at the tested doses but at doses higher than 5 μg/mL caused inhibition of the IL-8 receptor expression.@*CONCLUSIONS@#The present findings support the ethnomedicinal use of the Eastern Nigeria Mistletoe in the management of diseases affecting the immune system. The triterpenoid(s) have some immunomodulatory abilities on splenocytes and IL-8 receptors and may partly account for the overall immunomodulatory activity of this plant.

Utilization of antimicrobial agents with and without prescription by out-patients in selected pharmacies in South-eastern Nigeria.

OBJECTIVE: We conducted a study in out-patient pharmacies in South-eastern Nigeria in order to determine the extent of self-medication of antimicrobial agents in this area, assess the dosing error associated with this practice and to ascertain the extent of involvement of community pharmacies. METHOD: A survey was carried out daily in selected community pharmacies for a period of 90 days. Data were collected on the number of patients visiting these shops for antimicrobial agents, the number getting their medication with a prescription, the number getting their order without prescription and on the type and dose of antimicrobial agents received. MAIN OUT-COME MEASURE: The percentage of patients with prescription and without prescription was compared. The percentage under-dosages or over-dosages associated with how each antimicrobial agent was obtained were compared. The total DDDs of antimicrobial agents dispensed within this period with and without prescription were compared. RESULTS: A total of 4,128 outpatients visited the shops for antimicrobial agents within the period and were involved in the study. Of this number, 1,742 (42.2%) came with a prescription from qualified medical personnel and 2,386 (57.8%) came without a prescription. A total of 13,693.13 DDDs of antimicrobial agents was dispensed, of which 56.38% was dispensed with prescription and 43.62% was dispensed without prescription. The degrees of under-dosing were significantly (P < 0.05) higher in regimen filled without prescription when compared to those filled with prescription. CONCLUSION: Majority of the patients in this region still obtains their antimicrobial agents without a proper prescription, which is associated with sub-therapeutic dosing of these agents. There is a need for better regulation of antimicrobial agents dispensing and utilization in the region.