Isolation And Identification Of Antibacterial Substance Prouduced By Bacillus Coagulans And Apply It In Large Yellow Croaker Preservation

Flesh of large yellow croaker (Pseudosciaena crocea) is a great source of nutrition, and could be an important part of people’s diet structure. However, due to the high content of the proteins, degradation and degeneration will easily happen by the effects of proteases of spoilage bacteria in the process of storage, leading to fish nutrition loss, decline in sensory characteristics and freshness. The spoilage will cause large economic losses.This study investigated physiochemical properties of antibacterial substance produced by the probiotic B. coagulans LL1103isolated and stored in our lab. Then we purified and identified the antibacterial substance for application in large yellow croaker preservation during cold storage. The major conclusions are described below:1、In the present study, the relationship among growth condition, pH and bacteriostatic activity of probiotics Bacillus coagulans LL1103was examined, and the optimal fermentation time for antibacterial substance producing is18h. The antibacterial crude extract was then prepared for antibacterial spectrum measuring. The results showed that the extract has a broad spectrum of significant bacteriostatic activities, including activities against gram-negative and-positive bacteria. In addition, the maximal bacteriostatic activity of this extract was observed by incubating B. coagulans LL1103in MRS medium at37℃, pH6.0.2、We found that antibacterial product was sensitive to protease K, trypsin and pepsin, but not hydrogen oxidation enzymes. Thus, we speculate that the main ingredients of antibacterial product is a kind of bacteriocin which is sensitive to proteases.3、Crude extracting of antibacterial substance Lactocin LL1103was demonstrated by ammonium sulfate grading precipitation. Tricine-SDS-PAGE showed that the molecular weight of Lactocin LL1103is about2.4kDa. Furthermore, Lactocin LL1103was purified by gel filtration and HPLC. Finally, the N-terminal amino acid sequence of Lactocin LL1103was obtained by Edman degradation, which is GEPGPAGAVGPAGKQGPPGPQGPTGPP.4、Lactocin LL1103crude extracts with different concentrations were used for large yellow croaker preservation during cold storage. We found that Lactocin LL1103crude extracts with all the testing concentrations can inhibit the growth of spoilage organisms and prolong the shelf life of the large yellow croaker by measuring TVB-N, TBA, K value, sensory index, pH and microbial index. The optimal concentration was also confirmed as0.8mg/ml.