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A series of substituted 1,2,3,4-tetrahydropyrazino [1,2-a] indole derivatives have been synthesized and tested against the Gram positive and Gram negative strains of bacteria namely Staphylococcus aureus (MTCCB 737), Salmonella typhi (MTCCB 733), Pseudomonas aeruginosa (MTCCB 741), Streptomyces thermonitrificans (MTCCB 1824) and Escherichia coli (MTCCB 1652). All synthesized compounds showed mild to moderate activity. However, compounds 4d–f were found to have potent activity against pathogenic bacteria used in the study. Their MIC ranged from 3.75 to 60μg/disc. In vitro toxicity tests demonstrated that toxicity of 4d–f was not significantly different than that of gentamycin. However, at higher concentration (1000–4000μg/ml) difference was highly significant.