Investigation of plants used in Gabonese traditional medicine for the treatment of opportunistic infections caused by HIV

Abstract

Background: Currently, Human Immunodeficiency Virus/ Acquired Immune Deficiency Syndrome (HIV/AIDS) is one of the deadliest diseases in the world. In Gabon, the prevalence is estimated at 4.1%. Because of the several side effects of highly active antiretroviral therapy on the health of people living with HIV, an emphasis on the utilization of based plant treatments have been noticed highlighted with testimonies of patient health improvement. That led researchers to conduct ethnobotanical surveys aiming at reporting medicinal plants used to treat HIV related opportunistic infections. From the outcomes, 20 plants have been selected based on the frequency of their use but also on the absence of papers in the literature related to HIV research conducted in Gabon. This study aims at evaluating the cytotoxicity, antioxidant, antimicrobial, anti-HIV and immunomodulatory activities of the 20 selected plants and to establish the phytochemical profile of these plants. Methodology: The plants were extracted using absolute methanol and distilled water. Qualitative (phytochemical screening) and quantitative (TPC and TFC) analysis were performed on the 40 extracts obtained. The extracts were assessed for cytotoxicity on Vero and HeLa cells using MTT and dual staining/fluorescence microscopy. The antimicrobial activity of the extracts was evaluated using well diffusion assay and micro-broth dilution assay. DPPH free radical scavenging assay and ferric reducing power were used to determine the antioxidant potential of the extracts. The anti-HIV effect was evaluated on HIV infected blood using reverse transcriptase inhibitory assay and p24 antigen inhibitory assay. The immunomodulatory effects of the extracts were evaluated on HIV infected blood using IL-2, IL-6 and TNF-α ELISA. Coula edulis and T. iboga methanol extracts were selected for fractionation using column chromatography. LC-MS was used to establish the phytochemical profile of the two selected extracts. Results: The results revealed that saponins, alkaloids, terpenoids, phenols, tannins, steroids, flavonoids and cardiac glycosides were detected in almost all the plant extracts. The extracts were found to contain more phenolics than flavonoids especially the methanolic extracts. Both alcoholic and aqueous extracts of C. edulis, M. cecropioides, S. ochocoa, S. kamerounensis, U. guineensis and C. religiosa, the methanolic extract of A. hirtella and R. vomitoria and the aqueous extracts of U. klainei presented IC50 significantly lower (p<0.05) than ascorbic acid (38.87±1.54 μg/ml). Both Coula edulis methanolic and aqueous extracts, methanolic extracts of S. kamerounensis, S. gabonensis, M. monandra, U.guineensis and A. klaineana showed antioxidant activity significantly xii | P a g e (p<0.01) higher than Ascorbic acid whose EC50 was 152.16±1.09 μg/ml. For the fluorescence microscopy the extracts of interest for Vero cells inhibition were A. klaineana, T. iboga, U. klainei and C. edulis while for Hela cells the cytotoxic activity was significant (p<0.05) for the following extracts: U. klainei, U. guineensis, P. soyauxii, A. klaineana, V. conferta, M. monandra, R. vomitoria, R. africanum, C. edulis and S. ochocoa. The extracts of C. edulis, M. cecropioides, S. ochocoa, S. gabonensis, A. hirtella, R. vomitoria, S. kamerunensis, P. soyauxii, U. guineensis and T. iboga displayed the highest antimicrobial activity against all the selected bacteria. For the anti-HIV the extracts of interest were V. conferta (47%), C. lucanusianus (47%), C. religiosa (44%), A. hirtella (42%), S. kamerunensis (41%), M. puberula (41%) as well as both aqueous and methanolic extracts of T. iboga (48% and 45% respectively), U. guineensis (46% and 41% respectively) and A. klaineana (41% and 44% respectively). The results for RT inhibition assay revealed that out of 24 selected plant extracts selected only C. edulis, C. religiosa, C. lucanusianus, R. africanum, P. soyauxii and V. conferta were able to decrease RT by more than 50%. The modulatory effect of the extracts on the secretion of IL-2, IL-6 and TNF-α by HIV infected cells was found to be insignificant (p>0.05). The fractionation of T. iboga and C. edulis methanol extracts eluted 4 fractions each. The LC-MS analysis revealed at negative ionization the presence of phenolic compounds (Ellagic acid, Gallic acid, and quercetin) in C. edulis samples while alkaloids (ibogaine, ibogaline, voacangine) were mainly found in T. iboga samples at positive ionization. The two crude extracts and the fractions showed some levels of antimicrobial, antioxidant, anti-HIV and anti-inflammatory activities. Conclusion: The present study has validated the use of the selected plants in the management of AIDS in Gabon and provided an explanation of the improvement in HIV individual’s life reported. Some of these plants could constitute good candidate for promising anti-HIV molecules.