Piper retrofractum Vahl (long pepper) is in the Piperaceae family which can be been used as a Thai medicinal herb and ingredient in Thai curry pastes. Studies investigating bioactive metabolites extracted from fruit P. retrofractum using organic solvent cascade from low polarity to high polarity. The goals of this study were screened for antimicrobial activity against Bacillus subtilis, Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella typhi, Vibrio parahaemolyticus and, Candida albicans. Acetonitrile and methanol showed an effective clear zone against bacterial than other solvents. Minimal inhibition concentration (MIC) values were determined by colorimetric assay due to pigmentations of the crude extracts that obscured visual growth turbidity inspection. This study suggested that extracted with acetonitrile shown better antimicrobial susceptibility all microbial assays. Overall, these data suggest that certain bioactive metabolites from P. retrofractum may have potential use for the treatment of several Gram-positive and Gram-negative bacterial infections.