The aim of this study was to investigate the antimicrobial activity of Bacillus subtilis LS 1-2 grown on citrus juice waste (CJW). Citrus-juice waste (CJW) was obtained from the residue of squeezed citrus fruits. To use CJW as a raw material for the growth of B. sutilitis, a citrus-juice medium (CJM) was prepared by treating CJW with Ca(OH)2. No antimicrobial activity was observed either in the culture broth of Luria Broth medium or in CJM itself. The maximum antimicrobial activity was obtained after 24 hr of cultivation (culture) of Bacillus sutilitis LS 1-2 in CJM. The culture supernatant exhibited inhibitory activity against E. coli O-157 (140 AU/mL), Staphylococcus aureus (180 AU/mL), and Candida albicans (260 AU/mL), respectively. Bacillus subtilus LS 1-2 also produced protease (3,600 U/ml) and amylase (290 U/ml) in CJM. Antimicrobial activity of the culture broth was stable for 1 hr at 100 ℃, pH 2-10, and bile acid (concentration needs, 1 mM TDOC and 0.27 mM DOCmM), respectively. These results indicate the potential of CJW as a novel bioresource and the scope of probiotic applications of Bacillus subtilis LS 1-2 in various industrial applications.