Cocoa and milk powder samples were taken from a confectionery and tested for presence of microbes (harmful and pathogenic).Biochemical characterization of isolated microbes was carried out for confirmation. Lactobacillus was isolated from milk powder. When a culture of Lactobacillus sp. was inoculated into milk and incubated at room temperature, it multiplies and converts lactose to lactic acid. Fourier transform infrared spectroscopy was used to study the variation of functional group peaks in milk by the action of Lactobacillus sp. The spectral changes were also observed. Our main aim of this project is the production of bacteriocin from isolated lactobacillus species; it showed broad range of antibacterial activity against some food borne pathogens like staphyloccus, Ecoli, streptococcus, Enterococcus etc. The bacteriocin is purified by ammonium sulfate precipitate and dialysis. Biochemically it was pure protein moiety. Maximum bacteriocin concentration was found after dialysis. Project revealed the possibility of using bacteriocin as food preservative. © 2013 IJDDR, Ramalingam C et al.