PEMURNIAN DAN KARAKTERISASI PEDIOSIN YANG DIPRODUKSI OLEH Pediococcus pentosaceus 2A2

Abstract

Pediocin production by Pediococcus pentosaceus 2A2 was done in 30% molasses based medium at 37°C for 24 hours. The culture was then centrifuged to separate the cells and yield cell-free supernatant (CFS). Purification of the CFS was carried out by stepwise ammonium sulfate precipitation to achieve a 90% concentrate, followed by dialysis using membrane with a molecular weight cut off 2.0 kDa resulting in pediocin crude extract (PCE). The PCE was purified by cation exchange chromatography using SP Se-phadex C-25 and eluted with sodium acetat buffer at pH of 5.5; 6.0; 6.5; and 7.0 successively resulting in 4 fractions (F1, F2, F3, F4). Each purification step resulted in the increase in antimicrobial activity against Listeria monocytogenes ATCC 7644. The PCE had specific antimicrobial activity up to 5 fold higher than the CFS. F4 fraction, which showed the largest inhibition zone among the other fractions, had a very high specific activity up to 435 fold higher than the CFS. The SDS-PAGE analysis suggested that the molecular weight of F4 was approximately 5.9 kDa. The minimum inhibitory concentration (MIC) of F4 to inhibit 1 log (90%) of L. monocytogenes ATCC 7644 was 143 ppm. The inhibitory phenomenon observed under scanning electron microscope (SEM) showed that L. monocytogenes’s cells exposed to F4 experience morphological changes such as cells shrinkage.

Keywords

• characterization
• molasses
• pediocin
• pediococcus pentosaceus 2a2
• purification