Characterization of Antimicrobial Peptide fraction producing Lactobacillus spp. based on LC/MS-MS and determination of ACE-inhibition activity in Kefir

Öz

Bu çalışmada, Kefir üretiminde farklı Lactobacillus (Lactobacillus delbrueckii ssp. bulgaricus ATCC 11842, Lactobacillus helveticus ATCC 15009 ve Lactobacillus plantarum ATCC 14917) ilave kültür kullanımının ACE inhibisyon aktivitesi ve patojenik bakterilere karşı antimikrobiyal aktivite gibi biyoaktif özellikleri üzerine etkisi araştırılmış ve proteoliz özellikleri (HPLC peptit profilleri, HPLC serbest amino asit profilleri) incelenmiştir. Antimikrobiyal aktivite, kefir ekstraktları ve LC-MS/MS ile karakterize edilen ve öncü ve ürün iyonları belirlenen peptid fraksiyonunda (< 3kDa, Fraksiyon 2 olarak adlandırılan) araştırılmıştır. Tüm kefir örneklerinde Escherichia coli gibi patojenik bakterilere karşı antimikrobiyal aktivite gözlenmiştir. Fakat sonuçlar, Lactobacillus kullanılarak üretilen kefirden ayrılan F2 fraksiyonunun, kontrol örneklerinden daha güçlü antibakteriyel etkiye sahip olduğunu ortaya koymuştur. Bu fraksiyonun S. aureus, S. warneri 95052 ve S. hominis'e karşı antimikrobiyal aktiviteye sahip olduğu belirlenmiştir. Kefir A, B, C ve kontrol numunelerinin ACE-i aktivitesi sırasıyla % 76.47, % 84.95, % 87.33 ve % 85.57 olarak bulunmuştur. Lactobacillus kullanımıyla ACE-i aktivitesinde önemli bir (P <0.01) artış meydana gelmiştir. Bu çalışmayla, Lactobacillus yardımcı kültür kullanımının kefirin fonksiyonel değerine katkıda bulunduğu sonucuna varılmıştır.

Characterization of Antimicrobial Peptide Fraction Producing Lactobacillus spp. Based on LC/MS-MS and Determination of ACE-inhibitory Activity in Kefir

Abstract

In the present study, bioactive properties such as ACE-I activity and antimicrobial activity of kefir using different Lactobacillus (Lactobacillus delbrueckii ssp. bulgaricus ATCC 11842, Lactobacillus helveticus ATCC 15009 and Lactobacillus plantarum ATCC 14917) on some pathogen and Gram-positive bacteria during 28 day-storage periods was investigated and proteolysis (RP-HPLC peptide profiles, RP-HPLC amino acid profiles) were studied. The antimicrobial activity was investigated in kefir extract and separated peptide fractions (<3 kDa, named F2) which were characterized by LC-MS/MS and precursor and product ions were determined. Antimicrobial activity has been observed against pathogenic bacteria such as Escherichia coli (E. coli) in all samples. But the results revealed that the F2 fraction separated from kefir manufactured using Lactobacillus had a stronger antibacterial effect than control samples. It was determined that the F2 fraction has antimicrobial activity against S. aureus, S. warneri 95052 and S. hominis. The ACE-I activity of samples A, B, C and K were 76.47%, 84.95%, 87.33% and 85.57%, respectively. In the kefir using Lactobacillus has increase of ACE-I activity and was significant (P<0.01). In this study, it was concluded that the using of adjunct Lactobacillus contributed to the functional value of kefir.

Keywords

• Bioactive peptides
• In-Vitro antimicrobial peptides
• ACE- I activity
• purification
• LC-MS/MS

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