Isolation,Identification,Characterization And Purification Of Bacteriocins Of Lactobacillus Casei From The Starter Of Chinese Fermented Vegetables

Bacteriocins are heat-stable antimicrobial peptides produced by many organisms,including lactic acid bacteria(LAB),have the potential to kill or inhibit other bacteria.LAB are Gram-positive bacteria with great biotechnological potential in food industry.LAB produce a diverse group of bacteriocins.Keeping in view the consumer demand for less processed and free of chemical preservatives,bacteriocins of LAB received much attention as natural food preservatives.Moreover,these bacteriocins might warrant serious consideration as an alternative of the conventional antibiotics due to its broad-spectrum antimicrobial activities.In this study,Lactobacillus casei(L.casei)was derived from a traditional Chinese fermented food(named Jiangshui-Cai).A total of 16 strains of LAB from Chinese fermented vegetable food were isolated and tested for their antimicrobial activity against the indicator strains.The most effective strain against the indicators was selected for further identification by 16 S rRNA.Results of NCBI blast showed 100% similarities with L.casei(GenBank accession No.KP966438.1).A modified tomato base medium was optimized for the growth of L.casei taking care to exclude constituents that might compromise bacteriocin purification.Purified bacteriocins(Sephadax G-25 > HiPrep QXL 16/10 column of AKTA > RPC column of AKTA > reverse phase HPLC)from L.casei was named LiN333 which showed antimicrobial activity at a wide range of temperatures(60,80,100,121oC)and pH values(pH 3,4,5,6,7,8,9).The enzymatic treatment proved its proteinases nature as α-trypsin significantly decrease its inhibitory activity.Growth curve of L.casei showed a peak of strain production at 24 h cultured at 37°C.The antimicrobial activity of LiN333 was tested against Gram-positive and Gram-negative indictor strains including Listeria in this work.MALDI-TOF-MS confirmed the molecular weight of LiN333(4890Da).Scanning electron microscopy was used to analyze the mechanism of action of LiN333,while minimum inhibition concentration was recorded at 15 μg/ml.Mechanism of action,stability at a range of pH and temperatures highlights the potential of LiN333 as a bio-control agent for food preservation.Although many studies characterized bacteriocins biochemically,genetically,and their mechanism of action,but still many aspects are unknown.Therefore,the aim of the current work was to participate in the race of the detail characterization of novel bacteriocins.The results of the current work suggested that L.casei have potential applications in the control of both Grampositive and Gram-negative bacteria in the food industry.However,further detail study is suggested for the complete sequence of the LiN333,in vivo study for antimicrobial activity,antifungal,and anti-viral activity,to recommend it as an alternate of chemical food preservative and conventional antibiotics.