The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus

Introduction2,5-diketopiperazines are the simplest forms of cyclic dipeptides (CDPs) and have diverse frameworks with chiral side chains that are useful for drug development. Previous research has investigated the antimicrobial properties of proline-linked CDPs and their combinations in the culture...

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Main Authors: Jaeyoung Son, Yeonju Hong, Hyeri Seong, Yoon Sin Oh, Min-Kyu Kwak
Format: Article
Language:English
Published: Frontiers Media S.A. 2024-05-01
Series:Frontiers in Molecular Biosciences
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Online Access:https://www.frontiersin.org/articles/10.3389/fmolb.2024.1346598/full
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author Jaeyoung Son
Yeonju Hong
Hyeri Seong
Yoon Sin Oh
Min-Kyu Kwak
author_facet Jaeyoung Son
Yeonju Hong
Hyeri Seong
Yoon Sin Oh
Min-Kyu Kwak
author_sort Jaeyoung Son
collection DOAJ
description Introduction2,5-diketopiperazines are the simplest forms of cyclic dipeptides (CDPs) and have diverse frameworks with chiral side chains that are useful for drug development. Previous research has investigated the antimicrobial properties of proline-linked CDPs and their combinations in the culture filtrate (CF) of Lactobacillus plantarum LBP-K10 using anion exchange chromatography (AEC). However, the quantity of CDPs showcasing notable anti-influenza virus activity derived from AECs was generally lower than those originating from Lactobacillus CF.MethodsTo address this issue, the study aims to propose a more efficient method for isolating CDPs and to introduce the antiviral combinations of CDPs obtained using a new method. The study employed a novel technique entailing high-throughput C18-based solid-phase extraction with a methanol gradient (MeSPE). The MeSPE method involved increasing the methanol concentration from 5% to 50% in 5% increments.ResultsThe methanol SPE fractions (MeSPEfs) eluted with methanol concentrations between 35% and 45% evinced substantial efficacy in inhibiting the influenza A/H3N2 virus via plaque-forming assay. MeSPEf-45, the 45% MeSPEf, exhibited exceptional efficacy in preventing viral infections in Madin-Darby kidney cells, surpassing both individual CDPs and the entire set of MeSPEfs. To identify the specific antiviral components of MeSPEf-45, all MeSPEfs were further fractionated through preparative high-performance liquid chromatography (prep-HPLC). MeSPEf-45 fractions S8 and S11 presented the highest activity against multidrug-resistant bacteria and influenza A/H3N2 virus among all MeSPEfs, with 11 common fractions. Antiviral fractions S8 and S11 were identified as proline-based CDPs, specifically cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro), using gas chromatography-mass spectrometry. The combination of MeSPEf-45 fractions S8 and S11 displayed superior antibacterial and anti-influenza virus effects compared to the individual fractions S8 and S11.DiscussionHigh-throughput MeSPE-derived MeSPEfs and subsequent HPLC-fractionated fractions presents an innovative approach to selectively purify large amounts of potent antimicrobial CDPs from bacterial CF. The findings also show the effectiveness of physiologically bioactive combinations that utilize fractions not containing CDP. This study provides the initial evidence demonstrating the antimicrobial properties of CDPs acquired through high-throughput SPE techniques.
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spelling doaj-art-e3d69b4817c84dbda18eb097919c404c2025-08-20T02:16:10ZengFrontiers Media S.A.Frontiers in Molecular Biosciences2296-889X2024-05-011110.3389/fmolb.2024.13465981346598The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virusJaeyoung Son0Yeonju Hong1Hyeri Seong2Yoon Sin Oh3Min-Kyu Kwak4Laboratory of Microbial Physiology and Biotechnology, Department of Food and Nutrition, Institute of Food and Nutrition Science, College of Bio-Convergence, Eulji University, Seongnam, Republic of KoreaLaboratory of Microbial Physiology and Biotechnology, Department of Food and Nutrition, Institute of Food and Nutrition Science, College of Bio-Convergence, Eulji University, Seongnam, Republic of KoreaLaboratory of Microbial Physiology and Biotechnology, Department of Food and Nutrition, Institute of Food and Nutrition Science, College of Bio-Convergence, Eulji University, Seongnam, Republic of KoreaDepartment of Food and Nutrition, Institute of Food and Nutrition Science, College of Bio-Convergence, Eulji University, Seongnam, Republic of KoreaLaboratory of Microbial Physiology and Biotechnology, Department of Food and Nutrition, Institute of Food and Nutrition Science, College of Bio-Convergence, Eulji University, Seongnam, Republic of KoreaIntroduction2,5-diketopiperazines are the simplest forms of cyclic dipeptides (CDPs) and have diverse frameworks with chiral side chains that are useful for drug development. Previous research has investigated the antimicrobial properties of proline-linked CDPs and their combinations in the culture filtrate (CF) of Lactobacillus plantarum LBP-K10 using anion exchange chromatography (AEC). However, the quantity of CDPs showcasing notable anti-influenza virus activity derived from AECs was generally lower than those originating from Lactobacillus CF.MethodsTo address this issue, the study aims to propose a more efficient method for isolating CDPs and to introduce the antiviral combinations of CDPs obtained using a new method. The study employed a novel technique entailing high-throughput C18-based solid-phase extraction with a methanol gradient (MeSPE). The MeSPE method involved increasing the methanol concentration from 5% to 50% in 5% increments.ResultsThe methanol SPE fractions (MeSPEfs) eluted with methanol concentrations between 35% and 45% evinced substantial efficacy in inhibiting the influenza A/H3N2 virus via plaque-forming assay. MeSPEf-45, the 45% MeSPEf, exhibited exceptional efficacy in preventing viral infections in Madin-Darby kidney cells, surpassing both individual CDPs and the entire set of MeSPEfs. To identify the specific antiviral components of MeSPEf-45, all MeSPEfs were further fractionated through preparative high-performance liquid chromatography (prep-HPLC). MeSPEf-45 fractions S8 and S11 presented the highest activity against multidrug-resistant bacteria and influenza A/H3N2 virus among all MeSPEfs, with 11 common fractions. Antiviral fractions S8 and S11 were identified as proline-based CDPs, specifically cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro), using gas chromatography-mass spectrometry. The combination of MeSPEf-45 fractions S8 and S11 displayed superior antibacterial and anti-influenza virus effects compared to the individual fractions S8 and S11.DiscussionHigh-throughput MeSPE-derived MeSPEfs and subsequent HPLC-fractionated fractions presents an innovative approach to selectively purify large amounts of potent antimicrobial CDPs from bacterial CF. The findings also show the effectiveness of physiologically bioactive combinations that utilize fractions not containing CDP. This study provides the initial evidence demonstrating the antimicrobial properties of CDPs acquired through high-throughput SPE techniques.https://www.frontiersin.org/articles/10.3389/fmolb.2024.1346598/fullcyclic dipeptidescis-cyclo(L-Leu-L-Pro)cis-cyclo(L-Phe-L-Pro)solid-phase extractioninfluenza A virusLactobacillus plantarum LBP-K10
spellingShingle Jaeyoung Son
Yeonju Hong
Hyeri Seong
Yoon Sin Oh
Min-Kyu Kwak
The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
Frontiers in Molecular Biosciences
cyclic dipeptides
cis-cyclo(L-Leu-L-Pro)
cis-cyclo(L-Phe-L-Pro)
solid-phase extraction
influenza A virus
Lactobacillus plantarum LBP-K10
title The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
title_full The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
title_fullStr The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
title_full_unstemmed The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
title_short The high-throughput solid-phase extraction of cis-cyclo(L-Leu-L-Pro) and cis-cyclo(L-Phe-L-Pro) from Lactobacillus plantarum demonstrates efficacy against multidrug-resistant bacteria and influenza A (H3N2) virus
title_sort high throughput solid phase extraction of cis cyclo l leu l pro and cis cyclo l phe l pro from lactobacillus plantarum demonstrates efficacy against multidrug resistant bacteria and influenza a h3n2 virus
topic cyclic dipeptides
cis-cyclo(L-Leu-L-Pro)
cis-cyclo(L-Phe-L-Pro)
solid-phase extraction
influenza A virus
Lactobacillus plantarum LBP-K10
url https://www.frontiersin.org/articles/10.3389/fmolb.2024.1346598/full
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