Bacteriocin producing Lactobacillus rhamnosus GP1 was isolated from Grape peel and characterized. The bacteriocin had wide spectrum of inhibitory activity against test strains of pathogenic and food spoilage micro organisms. The cell-free supernatant of L. rhamnosus inhibited the grow th of Bacillus brevis, B. pumilus, B. subtilis, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Vibrio harveyi, Acinetobacter sp. and Arthrobacter sp. The influence of culture conditions on the production of the bacteriocin by L. rhamnosus was evaluated. Maximum bacteriocin activity of 1200 AU/mL was obtained in MRS broth at 72 h at an initial pH of 6.0 in 30ºC. Increased bacteriocin production by L. rhamnosus GP1 was noted when MRS broth was supplemented with yeast extract (0.1%), tween 80 (0.1%), MgSO₄ (0.02% to 0.04%) and dextrose (2.0 to 3.0%). Addition of bacteriological peptone, beef extract, NaCl and SDS to the culture medium however, reduced the production of bacteriocin. Results of sensitivity studies revealed remarkable stability of the bacteriocin at different storage conditions, temperatures and pH tested.