Bioassay-Guided Fractionation and in vitro Study of Antibacterial Activity of Tamarindus indica L. Stem Bark and Formulation of an Ointment.

Abstract

Background: The emergence of antibiotic resistance leads to find new antibacterial compounds. Tamarindus indica L. (T.indica L) (Siyambala) is a medicinally significant plant used in traditional medicinal systems for antibacterial activity. Objective: To perform bioassay-guided fractionation and evaluate the antibacterial activity of the crude extract of the internal stem bark of T. indica L. and formulate an ointment Methods: The plant was authenticated at the National Herbarium, Peradeniya and stem bark was shade-dried and refluxed using 80% ethanol. The freeze-dried extract was subjected to phytochemical screening (PS) and agar well diffusion (AWD) against Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922). The extract was subjected to bioassay guided fractionation with hexane, dichloromethane, and ethyl acetate and fractions were evaluated for PS and AWD. Three formulations were prepared using 2.5%, 5% and 10% of the crude extract. AWD test was conducted for each formulation and physical stability was evaluated for 14 days. Ciprofloxacin 2 mg/mL and dimethyl sulfoxide were used as positive (P) and negative (N) controls for extracts while mupirocin ointment 2% w/w (P) and ointment base (N) were used for formulation testing. Results: Crude extract indicated presence of phenols, flavonoids, saponins, terpenoids, tannins, carbohydrates, reducing sugars, triterpenoids and alkaloids. Extract showed antibacterial activity against S. aureus and E. coli with 15.47±0.40 and 16.10±0.17 mm zone of inhibition (ZOI) respectively at 16 mg/mL. Hexane fraction showed a minimum ZOI, 9.40±0.00 and 10.90±0.17 mm for S. aureus and E. coli respectively. Ethyl acetate fraction had maximum ZOI, 17.91±0.38 and 18.25±0.23 mm for S. aureus and E. coli respectively. The highest antibacterial activity was shown for 2.5% formulation against S. aureus (26.55±0.77 mm) and 10% formulation against E. coli (14.09±0.08 mm). Mupirocin ointment showed 35.63±0.37 and 20.89±0.04 mm ZOI against S. aureus and E. coli respectively. All formulations were physically stable during the tested period. Conclusion: Tested ointment formulations were physically stable and showed antibacterial activity against tested bacteria. Clinical trials to prove safety and efficacy are recommended.