J Microbiol. 2013 Dec;51(6):836-843.

Cyclic dipeptides from lactic acid bacteria inhibit proliferation of the influenza A virus.

Min-Kyu Kwak, Rui Liu, Jun-Oh Kwon, Min-Kyu Kim, Andrew HyoungJin Kim, Sa-Ouk Kang.

Laboratory of Biophysics, School of Biological Sciences, and Institute of Microbiology, Seoul National University, Seoul 151-747, Republic of Korea.

 

Abstract

We isolated Lactobacillus plantarum LBP-K10 from the traditional Korean fermented food kimchi. When organic acids were removed, the culture filtrate of this isolate showed high antiviral activity (measured using a plaque-forming assay) against the influenza A (H3N2) virus. Two fractions that were active against influenza A virus were purified from the culture filtrate using a C18 column with high-performance liquid chromatography. These active fractions were crystallized and identified to be the cyclic dipeptides cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) using gas chromatography-mass spectrometry; this identification was confirmed by X-ray crystallography. These cyclic dipeptides were identified in the culture filtrate of other lactic acid bacteria, including Lactobacillus spp., Leuconostoc spp., Weissella spp., and Lactococcus lactis.

 

Summary

Lactic acid bacteria and their culture supernatant are useful tools as food preservation and the protectors by inhibiting environmental unwanted microbes. Particularly, their culture supernatants have been regarded as the important inhibitor of the proliferation of Gram-positive and Gram-negative bacteria, pathogenic fungi and virus as immune enhancer. Around 400 isolated strains of lactic acid bacteria were isolated from kimchi, using various vegetables including leaves and stems of mustard, sedum, and Chinese cabbage and identified using 16S rDNA sequencing methods with PCR amplification. Furthermore, we confirmed Lactobacillus plantarum LBP-K10 and its three day-cultured supernatant was revealed to have the most potent antimicrobial and antiviral activities among isolated lactic acid bacterial strains. Antimicrobial activity of the culture supernatant of Lb. plantarum LBP-K10 was stable as non-proteinous substances when several proteolytic enzymes were treated and also was resistant to high temperatures. Seventeen fractions from the culture supernatant of Lb. plantarum LBP-K10 were collected by C18 hydrophobic column using high performance liquid chromatography (FIG1) and confirmed by gas chromatography-mass spectroscopy (FIG2) or X-ray crystallography (FIG3). The fractions were observed to be seven kinds of proline-containing cyclic dipeptides, one kind of non-proline-containing cyclic dipeptide and one kind of small molecules. Among these fractions, two kinds of cyclic dipeptides significantly inhibited the proliferation of the influenza A virus (H3N2). This is the first report on antiviral activity of cyclic dipeptides. Moreover, the cyclic dipeptides complex compound, which was removed of organic acids, showed higher antiviral activity compared to single cyclic dipeptide, significantly. Surprisingly, Pediococcus pentosaceus, Lb. sakei LBP-S01, Lb. plantarum/pentos LBP-S02, Lactococcus lactis LBP-S03, Leuconostoc citreum LBP-S05, Leuconostoc mesenteroides LBP-K06, Lb. plantarum LBP-K10, Weissella cibaria LBP-K15 and W. confusa LBP-K16 have been thought to produce identical antiviral cyclic dipeptides inhibitory to influenza A virus. Furthermore, we observed that Korean traditional foods including kimchi and shrimp-fermented jeotgal had abundant these compounds. From this result, Korean people have taken natural antibiotics what we found in this study for a long time without causing any side effects and toxicity.

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Dr. Sa-Ouk Kang

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