With the growing global threat of antibiotics-resistant superbugs and increasing economic and environmental concerns on synthetic production of natural products and their analogs, it is imperative to developing alternate platforms for the discovery,production and development of new antimicrobial agents and to treat infections of human and animal origin. Bisindole alkaloids have received considerable interest for several years because of their broad range of biological properties. Many studies are focusing on the bisindole microbial pigments to reveal how these compounds can provide a survival advantage for certain microorganisms in natural environments.Violacein (Vio), a bisindole natural pigment isolated from several bacteria, have attracted much attention recently for its multiple important biological activities including antibacterial, antiviral, anti-tumor, and anti-oxidant activities. In order to give a better insight into violaceins bioactivity, two violacein derivatives, deoxyviolacein and oxyviolacein, were biosynthesized in good yield by wild and engineering strains respectively. The structures of the newly synthesized compounds were confirmed based on the analysis by HPLC-MS, 1H- 13C NMR. The crude extracts of violacein derivatives were evaluated for their antimicrobial activities against nine bacteria and fourteen fungi. All violacein derivatives exhibited moderate activities against Gram-positive bacteria and also showed pronounced antifungal activities. However, no molecule displayed activity against the gram-negative bacteria tested. All compounds displayed interesting antifungal activity. The overall results of the present work provide baseline information for the potential use of violacein and its derivative as an antimicrobial germicide.