Bioassay guided fractionation of marine streptomyces sp. GMY01 and antiplasmodial assay using microscopic and flow cytometry method

Abstract

Marine-derived Streptomyces sp. GMY01 is a potential actinobacteria which have anticancer activities. In previous studies, several anticancer agents exhibited antiplasmodial activity. This study aimed to evaluate antiplasmodial activity of GMY01 extract using bioassay guided fractionation. Crude extract of ethyl acetate was obtained from supernatant of 11 days fermentation in starch nitrate medium. Crude extract was fractionated using n-hexane and methanol solvent. Re-fractionation was applied using flash chromatography and column chromatography. Antiplasmodial assay was performed on Plasmodium falciparum FCR3 and P. falciparum 3D7 by microscopic method using thin blood smear + Giemsa stain, and was confirmed by flow cytometry method using SYBR Green I stain. Toxicity assay was performed on Vero cells line (normal cells). 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=3.96μg/mL) with very low toxicity on Vero cells (IC50=30, 072μg/mL). Bioassay guided fractionation resulted F4.7 which has the highest Plasmodium inhibition (94.3% at 5μg/mL). Main constituent analysis showed C10H13NO (163.09971 Da) and C11H15NO13 (209.10519 Da) as two mayor compounds in F4.7. This study guided the isolation and structure elucidation of potential compound in further research.