Bioassay Guided Fractionation of Marine Streptomyces sp. GMY01 and Antiplasmodial Assay using Microscopic and Flow Cytometry Method

Abstract

Genome mining study showed that marine-derived Streptomyces sp. GMY01 is a potential actinobacteria with abundant of secondary metabolite and anticancer activity. The study aimed to evaluate bioassay guided fractionation for antiplasmodial screening of GMY01 extract and to predict compound on active fractions. Ethyl acetate fraction was obtained from 11 days fermentation product of GMY01 and then it was fractionated using n-hexane and methanol solvent. Refractionated was applied using flash chromatography and column chromatography. Antiplasmodial assay was performed on Plasmodium falciparum FCR3 by microscopic method using thin blood smear + Giemsa stain, and flow cytometry method using SYBR Green I stain. Toxicity assay was performed on Vero cells line. Main constituent of active fraction was analyzed using LCMS/MS. The result of the study showed that ethyl acetate-methanol fraction has high antiplasmodial activity (IC50=3.96 µg/mL) with very low toxicity on Vero cells (IC50=30,072 µg/mL). Bioassay guided fractionation resulted F4.7 as highest Plasmodium inhibition (94.3% at 5 µg/mL) and was confirmed by microscopic and flow cytometry assay. Main constituent analysis showed C10H13NO (163.09971 Da) as mayor compound and predicted as nonribosomal polyketide synthetase (NRPS) secondary metabolite.

Keywords

• drug discovery
• malaria
• antiplasmodial assay
• actinomycetes