Polarity of extracts and fractions of four Combretum (Combretaceae) species used to treat infections and gastrointestinal disorders in southern African traditional medicine has a major effect on different relevant in vitro activities

Abstract

ETHNOPHARMACOLOGICAL IMPORTANCE : Infections and gastrointestinal (GIT) disorders such as diarrhoea causes many problems in human health and animal production. Many Combretum species are used in traditional medicine to treat various diseases by rural people in Africa and Asia. Much of the work done to date on some species to validate their ethnopharmacological use was on the non-polar or intermediate polarity components. Many species are yet to be studied against relevant disease parameters using more polar extracts. AIMS : The polar components were extracted and fractionated by solvent-solvent fractionation to yield fractions of different polarities. The activity of these fractions on different parameters that could be involved in infectious and gastrointestinal track (GIT) disorders was investigated. The cytotoxic activities of the extracts were also determined to evaluate the potential of these extracts to combat diarrhoea in production animals. MATERIALS AND METHODS : Phenolic-enriched leaf extracts of Combretum bracteosum (Cob), Combretum padoides (Cop), Combretum vendae (Cov) and Combretum woodii (Cow) were obtained by extracting with a mixture of 70% acetone acidified with 1% HCl and n-hexane. The extract was sequentially treated by solvent-solvent fractionation with dichloromethane, ethyl acetate, and butanol to yield fractions with a large variation in polarity. The phenolic constituents of the extracts and fractions were determined using standard procedures. The antioxidant activities were determined using various standard methods. The minimum inhibitory concentrations (MICs) of the crude extracts and fractions against four bacterial and three fungal strains were assessed with a microplate serial dilution method. Cyclooxygenase (COX) and lipoxygenase (LOX) enzyme inhibitory assays and cytotoxicity studies against Vero cells were also carried out. RESULT : Some of the fractions had much higher antioxidant activity than the positive controls. The average EC50 values of the extracts for the DPPH and ABTS antioxidant assays were 0.21-12 μg/ml (Cop), 0.25-16 μg/ml (Cov), 0.33-9.41 μg/ml (Cow) and 4.97-85 μg/ml (Cob) respectively while the mean EC50 values for the positive controls ascorbic acid and trolox were 1.28-1.51 and 1.02-1.19 μg/ml respectively. All the crude extracts inhibited lipid peroxidation of linoleic acid by more than 80% at a concentration of 64 μg/ml. Even though some crude extracts had relatively low antimicrobial activity, fractions from these had high activity. Cop had the highest antibacterial activity with MICs ranging between 19-2500 μg/ml,followed by Cov with MICs ranging between 39-625 μg/ml. Cop also had the highest antifungal activity with MICs between 19-625 μg/ml. The MIC for Cow and Cov ranged from 19 to 1250 μg/ml. The extracts had no activity against COX 1 and 2 enzymes in the anti-inflammatory assay but had good lipoxygenase inhibition. The crude extracts had high concentrations of hydrolysable tannin (gallotannin). A good correlation (R2= 0.99) was found between the antioxidant activity and total tannin content indicating that, gallotannins may be responsible for the antioxidant activity. CONCLUSION : The results obtained in this study provided a scientific basis for the use of leaf extracts from these plant species to treatinfectious and GIT disorders. In general non-polar fractions had a high antimicrobial activity and polar fractions had a high antioxidant activity.