Abstract: In this study, spoilage microorganisms were separated and characterized from firm tofu in bulk and boxed lactone tofu, the relationship between spoilage phenomena and putrefaction-making bacteria was preliminarily studied through pathogens inoculation experiments. Meanwhile, four food additives with bacteriostatic function were chosen to inhibit the spoilage microorganisms, including glycerol monolaurate (GML) , sucrose monolaurate, glucono-δ-lactone and Nisin. The minimum inhibitory concentrations (MIC) of each antibacterial agent and synergistic effects between GML and other antimicrobial agents were measured. Result showed that there were four kinds of spoilage microorganisms which led to corruption of tofu:Brevibacillus, Enterobacter, Lactobacillus and Leuconostoc. The first three strains could cause clamminess, sack expanding and softening of tofu respectively. Antibacterial activities of GML or Nisin against each spoilage microorganism mentioned above were better than others. The synergistic effect of GML and Nisin was the best, especially for Brevibacillus, Lactobacillus and Leuconostoc. This paper would lay a foundation of future study on new and efficient preservative added to tofu to prolong the shelf life.