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Author: search.filters.author.Pilasombut, KomkhaeSubject: search.filters.subject.Lactic acid bacteria

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    In vitro characterization of bacteriocin produced by lactic acid bacteria isolated from Nem Chua, a traditional Vietnamese fermented pork
    (2015-08-01)
    Pilasombut, Komkhae
    ;
    Rumjuankiat, Kittaporn
    ;
    Ngamyeesoon, Nualphan
    ;
    Duy, Le Nguyen Doan
    The aim of this study was to screen and In vitro characterize the properties of bacteriocin produced by lactic acid bacteria isolated from Vietnamese fermented pork (Nem chua). One hundred and fifty LAB were isolated from ten samples of Nem chua and screened for bacteriocin-producing lactic acid bacteria. Antimicrobial activity of bacteriocin was carried out by spot on lawn method against both gram positive and gram negative bacteria. One isolate, assigned as KL-1, produced bacteriocin and showed inhibitory activity against Lactobacillus sakei, Leuconostoc mesenteroides and Enterococcus faecalis. To characterize the bacteriocin-producing strain, optimum temperature, incubation period for maximum bacteriocin production and identification of bacteriocin-producing strain were determined. It was found that the optimum cultivation temperature of the strain to produce the maximum bacteriocin activity (12,800 AU/mL) was obtained at 30°C. Meanwhile, bacteriocin production at 6,400 AU/mL was found when culturing the strain at 37°C and 42°C. The isolate KL-1 was identified as L. plantarum. Antimicrobial activity of cell-free supernatant was completely inhibited by proteolytic enzyme of trypsin, alpha-chymotrypsin and proteinase K. Bacteriocin activity was stable at high temperature up to 100°C for 10 min and at 4°C storage for 2 d. However, the longer heating at 100°C and 4°C storage, its activity was reduced.
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    Purification and amino acid sequence of a bacteriocins produced by Lactobacillus salivarius K7 isolated from chicken intestine
    (2006-01-01)
    Pilasombut, Komkhae
    ;
    Sakpuaram, Thavajchai
    ;
    Wajjwalku, Worawidh
    ;
    Nitisinprasert, Sunee
    ;
    Swetwiwathana, Adisorn
    A bacteriocin-producing strain, Lactobacillus K7, was isolated from a chicken intestine. The inhibitory activity was determined by spot-on-lawn technique. Identification of the strain was performed by morphological, biochemical (API 50 CH kit) and molecular genetic (16S rDNA) basis. Bacteriocin purification processes were carried out by amberlite adsorption, cation exchange and reverse-phase high perform ance liquid chromatography. N-terminal amino acid sequences were performed by Edman degradation. Molecular mass was determined by electrospray-ionization (ESI) mass spectrometry (MS). Lactobacillus K7 showed inhibitory activity against Lactobacillus sakei subsp. sakei JCM 1157<sup>T</sup>, Leuconostoc mesenteroides subsp. mesenteroides JCM 6124<sup>T</sup> and Bacillus coagulons JCM 2257<sup>T</sup>. This strain was identified as Lb. salivarius. The antimicrobial substance was destroyed by proteolytic enzymes, indicating its proteinaceous structure designated as a bacteriocin type. The purification of bacteriocin by amberlite adsorption, cation exchange, and reverse-phase chromatography resulted in only one single active peak, which was designated FK22. Molecular weight of this fraction was 4331.70 Da. By amino acid sequence, this peptide was homology to Abp 118 beta produced by Lb. salivarius UCC118. In addition, Lb. salivarius UCC118 produced 2-peptide bacteriocin, which was Abp 118 alpha and beta. Based on the partial amino acid sequences of Abp 118 beta, specific primers were designed from nucleotide sequences according to data from GenBank. The result showed that the deduced peptide was high homology to 2-peptide bacteriocin, Abp 118 alpha and beta.