大蒜精油对熏马肠中德氏乳杆菌产腐胺的影响机制
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摘要
研究不同质量浓度大蒜精油对熏马肠中德氏乳杆菌产腐胺的影响机制。利用反转录实时荧光定量聚合酶链式反应技术分析供试菌在不同大蒜精油质量浓度下的胍基丁胺脱亚胺酶相关基因表达情况,使用超高效液相色谱连续48 h监测纯菌体系中的腐胺产量;并将德氏乳杆菌接入到含不同质量浓度大蒜精油的熏马肠中发酵28 d,期间取样测其菌落总数、pH值及腐胺积累量。结果表明:无论是在纯菌培养还是在熏马肠发酵过程中,大蒜精油都能明显抑制细菌的增长,在纯菌体系中会明显抑制pH值的下降,但对熏马肠的最终pH值影响不明显;大蒜精油对调节基因aguR的转录影响不明显,但能极显著抑制操纵子aguBDAC的转录(P<0.01);增加纯菌体系以及熏马肠发酵过程中的大蒜精油质量浓度均能明显减少腐胺的生成。
The effect of different concentrations of garlic essential oil on putrescine accumulation by Lactobacillus delbrueckii isolated from smoked horse meat sausages were explored, and the underlying mechanism was elucidated.Reverse transcription quantitative polymerase chain reaction(RT-qPCR) was used to analyze the gene expression of agmatine deiminase in Lactobacillus delbrueckii when cultured in the presence of different concentrations of garlic essential oil. Ultra performance liquid chromatography was used to monitor putrescine produced by the pure culture of Lactobacillus delbrueckii for 48 h. Additionally, the strain was incubated into smoked horse meat sausages with different concentrations of garlic essential oil and cultured for 28 days. Samples were collected during the fermentation period to measure the total number of bacteria, pH, and putrescine accumulation. Results showed that for both culture modes, garlic essential oil could signi?cantly inhibit bacterial growth. It signi?cantly inhibited the decrease in pH in the pure culture, but had little effect on the ?nal pH of smoked horsemeat sausages. Garlic essential oil had little impact on the transcription of the regulatory gene aguR, but signi?cantly inhibited the transcription of the operon aguBDAC(P < 0.01). Increasing garlic essential oil concentration can signi?cantly reduce the generation of putrescine both in pure culture and during the fermentation process of smoked horse meat sausages.
The effect of different concentrations of garlic essential oil on putrescine accumulation by Lactobacillus delbrueckii isolated from smoked horse meat sausages were explored, and the underlying mechanism was elucidated.Reverse transcription quantitative polymerase chain reaction(RT-qPCR) was used to analyze the gene expression of agmatine deiminase in Lactobacillus delbrueckii when cultured in the presence of different concentrations of garlic essential oil. Ultra performance liquid chromatography was used to monitor putrescine produced by the pure culture of Lactobacillus delbrueckii for 48 h. Additionally, the strain was incubated into smoked horse meat sausages with different concentrations of garlic essential oil and cultured for 28 days. Samples were collected during the fermentation period to measure the total number of bacteria, pH, and putrescine accumulation. Results showed that for both culture modes, garlic essential oil could signi?cantly inhibit bacterial growth. It signi?cantly inhibited the decrease in pH in the pure culture, but had little effect on the ?nal pH of smoked horsemeat sausages. Garlic essential oil had little impact on the transcription of the regulatory gene aguR, but signi?cantly inhibited the transcription of the operon aguBDAC(P < 0.01). Increasing garlic essential oil concentration can signi?cantly reduce the generation of putrescine both in pure culture and during the fermentation process of smoked horse meat sausages.
引文
[1]SHALABY A R. Signi?cance of biogenic amines to food safety and human health[J]. Food Research International, 1996, 29(7):675-690.DOI:10.1016/S0963-9969(96)00066-X.
[2]GARDINIF,?ZOGULY,SUZZIG,etal.Technologicalfactors affectingbiogenicaminecontentinfoods:areview[J].Frontiersin Microbiology, 2016, 7:1-8. DOI:10.3389/fmicb.2016.01218.
[3]LANDETEJM,ARENAME,PARDOI,etal.Theroleoftwo families of bacterial enzymes in putrescine synthesis from agmatine via agmatine deiminase[J]. International Microbiology, 2010, 13(4):169-177. DOI:10.2436/20.1501.01.123.
[4]LADEROV,RATTRAYFP,MAYOB,etal.Sequencingand transcriptional analysis of the biosynthesis gene cluster of putrescineproducingLactococcuslactis[J].AppliedandEnvironmental Microbiology, 2011, 77(18):6409-6418. DOI:10.1128/AEM.05507-11.
[5]DELRIOB,LINARESDM,LADEROV,etal.Putrescine production via the agmatine deiminase pathway increases the growth ofLactococcuslactisandcausesthealkalinizationoftheculture medium[J]. Applied Microbiology&Biotechnology, 2015, 99(2):897-905. DOI:10.1007/s00253-014-6130-8.
[6]LINARESDM,DELRIOB,REDRUELLOB,etal.AguR,a transmembrane transcription activator of the putrescine biosynthesis operoninLactococcuslactis,actsinresponsetotheagmatine concentration[J].Applied&EnvironmentalMicrobiology,2015,81(18):6145-6157. DOI:10.1128/AEM.00959-15.
[7]L I N A R E SDM,P A T R I C I AAS,B E A T R I ZDR,e ta l.Implementationoftheagmatine-controlledexpressionsystemfor induciblegeneexpressioninLactococcuslactis[J].MicrobialCell Factories, 2015, 14(1):1-9. DOI:10.1186/s12934-015-0399-x.
[8]LINARESDM,DELRIOB,LADEROV,etal.Theputrescine biosynthesispathwayinLactococcuslactisistranscriptionally regulatedbycarboncatabolicrepression,mediatedbyCcpA[J].InternationalJournalofFoodMicrobiology,2013,165(1):43-50.DOI:10.1016/j.ijfoodmicro.2013.04.021.
[9]DEL RIO B, LINARES D, LADERO V, et al. Putrescine biosynthesis inLactococcuslactis,istranscriptionallyactivatedatacidic pHandcounteractsacidificationofthecytosol[J].International JournalofFoodMicrobiology,2016,236:83-89.DOI:10.1016/j.ijfoodmicro.2016.07.021.
[10]BILLING J, SHERMAN P W. Antimicrobial functions of spices:why some like it hot[J]. The Quartely Review of Biology, 1998, 73(1):3-49.DOI:10.1086/420058.
[11]SHETTY S, THOMAS B, SHETTY V, et al. An in-vitro evaluation of the ef?cacy of garlic extract as an antimicrobial agent on periodontal pathogens:amicrobiological study[J].AYU, 2014,34(4):445-451.DOI:10.4103/0974-8520.127732.
[12]张雅晴,卢士玲,黄发添,等.大蒜精油对屎肠球菌和粪肠球菌产苯乙胺和酪胺的影响[J].现代食品科技, 2016, 32(11):117-122; 84.DOI:10.13982/j.mfst.1673-9078.2016.11.018.
[13]TA?BIA,DABOURN,LAMOUREUXM,etal.Comparative transcriptome analysis of Lactococcus lactis subsp. cremoris strains underconditionssimulatingCheddarcheesemanufacture[J].International Journal of Food Microbiology, 2011, 146(3):263-275.DOI:10.1016/j.ijfoodmicro.2011.02.034.
[14]LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression datausingreal-timequantitativePCRandthe2-ΔΔCTmethod[J].Methods, 2002, 25(4):402-408. DOI:10.1006/meth.2001.1262.
[15]LUShiling,JIHua,WANGQingling,etal.Theeffectsofstarter culturesandplantextractsonthebiogenicamineaccumulationin traditionalChinesesmokedhorsemeatsausages[J].FoodControl,2015, 50:869-875. DOI:10.1016/j.foodcont.2014.08.015.
[16]马宇霞,卢士玲,李开雄,等.熏马肠中生物胺氧化酶菌株的筛选与鉴定[J].现代食品科技, 2014, 30(5):49-55. DOI:10.13982/j.mfst.1673-9078.2014.05.002.
[17]CARSONCF,MEEBJ,RILEYTV.Mechanismofactionof melaleucaalternifolia(teatree)oilonStaphylococcusaureus determined by time-kill, lysis, leakage, and salt tolerance assays and electron microscopy[J]. Antimicrobial Agents&Chemotherapy, 2002,46(6):1914-1920. DOI:10.1128/AAC.46.6.1914-1920.2002.
[18]吴楠,祖元刚,王微.大蒜精油抗菌活性研究[J].食品科学,2008,29(3):103-105. DOI:10.3321/j.issn:1002-6630.2008.03.014.
[19]GUOJJ,KUOCM,CHUANGYC,etal.TheeffectsofgarlicsupplementeddietsonantibacterialactivityagainstStreptococcus iniae,andongrowthinorange-spottedgrouper,Epinephelus coioides[J].Aquaculture,2012,364/365(3):33-38.DOI:10.1016/j.aquaculture.2012.07.023.
[20]DELRIOB,REDRUELLOB,LADEROV,etal.Putrescine productionbyLactococcuslactissubsp.cremorisCECT8666is reduced by NaCl via a decrease in bacterial growth and the repression ofthegenesinvolvedinputrescineproduction[J].International JournalofFoodMicrobiology,2016,232:1-6.DOI:10.1016/j.ijfoodmicro.2016.05.010.
[21]ARENA M P, RUSSO P, CAPOZZI V, et al. Effect of abiotic stress conditionsonexpressionoftheLactobacillusbrevis,IOEB9809tyrosine decarboxylase and agmatine deiminase genes[J]. Annals of Microbiology, 2011, 61(1):179-183. DOI:10.1007/s13213-010-0089-1.
[22]LIUY,BURNERA.Multipletwo-componentsystemsof Streptococcusmutansregulateagmatinedeiminasegeneexpression and stress tolerance[J]. Journal of Bacteriology, 2009, 191(23):7363-7366. DOI:10.1128/JB.01054-09.
[23]DEL RIO B, LADERO V, REDRUELLO B, et al. Lactose-mediated carboncataboliterepressionofputrescineproductionindairy Lactococcus lactis is strain dependent[J]. Food Microbiology, 2015,48(1):163-170. DOI:10.1016/j.fm.2014.11.018.
[24]CHENG C, CHEN J, FANG C, et al. Listeria monocytogenes aguA1,but not aguA2, encodes a functional agmatine deiminase:biochemical characterization of its catalytic properties and roles in acid tolerance[J].JournalofBiologicalChemistry,2013,288(37):26606-26615.DOI:10.1074/jbc.M113.477380.
[25]FERNáNDEZM,LINARESDM,RODRíGUEZA,etal.Factors affecting tyramine production in Enterococcus durans, IPLA 655[J].AppliedMicrobiology&Biotechnology,2007,73(6):1400-1406.DOI:10.1007/s00253-006-0596-y.
[26]PEREZM,CALLES-ENRíQUEZM,NESI,etal.Tyramine biosynthesisistranscriptionallyinducedatlowpHandimproves thefitnessofEnterococcusfaecalis,inacidicenvironments[J].AppliedMicrobiology&Biotechnology,2015,99(8):3547-3558.DOI:10.1007/s00253-014-6301-7.
[27]BOZKURTH.Utilizationofnaturalantioxidants:greenteaextract and Thymbra spicata oil in Turkish dry-fermented sausage[J]. Meat Science, 2006, 73(3):442-450. DOI:10.1016/j.meatsci.2006.01.005.
[28]ELíASM,CARRASCOSAAV.CharacterisationofthePaio doAlentejo,atraditionalPortugueseIberiansausage,inrespect toitssafety[J].FoodControl,2010,21(1):97-102.DOI:10.1016/j.foodcont.2009.04.004.
[29]雷志方,谢晶,尹乐,等.温度和姜精油对金枪鱼品质影响及生物胺相关性[J].食品科学, 2017, 38(3):45-52. DOI:10.7506/spkx1002-6630-201703008.
[30]FILIPPIS F D, PENNACCHIA C, PASQUA R D, et al. Decarboxylase geneexpressionandcadaverineandputrescineproductionby Serratia proteamaculans in vitro and in beef[J]. International Journal ofFoodMicrobiology,2013,165(3):332-338.DOI:10.1016/j.ijfoodmicro.2013.05.021.
[31]L A D E R OV,C A L L E S-E N RíQ U E ZM,F E R NáN D E ZM,etal.Toxicologicaleffectsofdietarybiogenicamines[J].Current Nutrition&Food Science,2010,6(2):145-156.DOI:10.2174/157340110791233256.
[2]GARDINIF,?ZOGULY,SUZZIG,etal.Technologicalfactors affectingbiogenicaminecontentinfoods:areview[J].Frontiersin Microbiology, 2016, 7:1-8. DOI:10.3389/fmicb.2016.01218.
[3]LANDETEJM,ARENAME,PARDOI,etal.Theroleoftwo families of bacterial enzymes in putrescine synthesis from agmatine via agmatine deiminase[J]. International Microbiology, 2010, 13(4):169-177. DOI:10.2436/20.1501.01.123.
[4]LADEROV,RATTRAYFP,MAYOB,etal.Sequencingand transcriptional analysis of the biosynthesis gene cluster of putrescineproducingLactococcuslactis[J].AppliedandEnvironmental Microbiology, 2011, 77(18):6409-6418. DOI:10.1128/AEM.05507-11.
[5]DELRIOB,LINARESDM,LADEROV,etal.Putrescine production via the agmatine deiminase pathway increases the growth ofLactococcuslactisandcausesthealkalinizationoftheculture medium[J]. Applied Microbiology&Biotechnology, 2015, 99(2):897-905. DOI:10.1007/s00253-014-6130-8.
[6]LINARESDM,DELRIOB,REDRUELLOB,etal.AguR,a transmembrane transcription activator of the putrescine biosynthesis operoninLactococcuslactis,actsinresponsetotheagmatine concentration[J].Applied&EnvironmentalMicrobiology,2015,81(18):6145-6157. DOI:10.1128/AEM.00959-15.
[7]L I N A R E SDM,P A T R I C I AAS,B E A T R I ZDR,e ta l.Implementationoftheagmatine-controlledexpressionsystemfor induciblegeneexpressioninLactococcuslactis[J].MicrobialCell Factories, 2015, 14(1):1-9. DOI:10.1186/s12934-015-0399-x.
[8]LINARESDM,DELRIOB,LADEROV,etal.Theputrescine biosynthesispathwayinLactococcuslactisistranscriptionally regulatedbycarboncatabolicrepression,mediatedbyCcpA[J].InternationalJournalofFoodMicrobiology,2013,165(1):43-50.DOI:10.1016/j.ijfoodmicro.2013.04.021.
[9]DEL RIO B, LINARES D, LADERO V, et al. Putrescine biosynthesis inLactococcuslactis,istranscriptionallyactivatedatacidic pHandcounteractsacidificationofthecytosol[J].International JournalofFoodMicrobiology,2016,236:83-89.DOI:10.1016/j.ijfoodmicro.2016.07.021.
[10]BILLING J, SHERMAN P W. Antimicrobial functions of spices:why some like it hot[J]. The Quartely Review of Biology, 1998, 73(1):3-49.DOI:10.1086/420058.
[11]SHETTY S, THOMAS B, SHETTY V, et al. An in-vitro evaluation of the ef?cacy of garlic extract as an antimicrobial agent on periodontal pathogens:amicrobiological study[J].AYU, 2014,34(4):445-451.DOI:10.4103/0974-8520.127732.
[12]张雅晴,卢士玲,黄发添,等.大蒜精油对屎肠球菌和粪肠球菌产苯乙胺和酪胺的影响[J].现代食品科技, 2016, 32(11):117-122; 84.DOI:10.13982/j.mfst.1673-9078.2016.11.018.
[13]TA?BIA,DABOURN,LAMOUREUXM,etal.Comparative transcriptome analysis of Lactococcus lactis subsp. cremoris strains underconditionssimulatingCheddarcheesemanufacture[J].International Journal of Food Microbiology, 2011, 146(3):263-275.DOI:10.1016/j.ijfoodmicro.2011.02.034.
[14]LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression datausingreal-timequantitativePCRandthe2-ΔΔCTmethod[J].Methods, 2002, 25(4):402-408. DOI:10.1006/meth.2001.1262.
[15]LUShiling,JIHua,WANGQingling,etal.Theeffectsofstarter culturesandplantextractsonthebiogenicamineaccumulationin traditionalChinesesmokedhorsemeatsausages[J].FoodControl,2015, 50:869-875. DOI:10.1016/j.foodcont.2014.08.015.
[16]马宇霞,卢士玲,李开雄,等.熏马肠中生物胺氧化酶菌株的筛选与鉴定[J].现代食品科技, 2014, 30(5):49-55. DOI:10.13982/j.mfst.1673-9078.2014.05.002.
[17]CARSONCF,MEEBJ,RILEYTV.Mechanismofactionof melaleucaalternifolia(teatree)oilonStaphylococcusaureus determined by time-kill, lysis, leakage, and salt tolerance assays and electron microscopy[J]. Antimicrobial Agents&Chemotherapy, 2002,46(6):1914-1920. DOI:10.1128/AAC.46.6.1914-1920.2002.
[18]吴楠,祖元刚,王微.大蒜精油抗菌活性研究[J].食品科学,2008,29(3):103-105. DOI:10.3321/j.issn:1002-6630.2008.03.014.
[19]GUOJJ,KUOCM,CHUANGYC,etal.TheeffectsofgarlicsupplementeddietsonantibacterialactivityagainstStreptococcus iniae,andongrowthinorange-spottedgrouper,Epinephelus coioides[J].Aquaculture,2012,364/365(3):33-38.DOI:10.1016/j.aquaculture.2012.07.023.
[20]DELRIOB,REDRUELLOB,LADEROV,etal.Putrescine productionbyLactococcuslactissubsp.cremorisCECT8666is reduced by NaCl via a decrease in bacterial growth and the repression ofthegenesinvolvedinputrescineproduction[J].International JournalofFoodMicrobiology,2016,232:1-6.DOI:10.1016/j.ijfoodmicro.2016.05.010.
[21]ARENA M P, RUSSO P, CAPOZZI V, et al. Effect of abiotic stress conditionsonexpressionoftheLactobacillusbrevis,IOEB9809tyrosine decarboxylase and agmatine deiminase genes[J]. Annals of Microbiology, 2011, 61(1):179-183. DOI:10.1007/s13213-010-0089-1.
[22]LIUY,BURNERA.Multipletwo-componentsystemsof Streptococcusmutansregulateagmatinedeiminasegeneexpression and stress tolerance[J]. Journal of Bacteriology, 2009, 191(23):7363-7366. DOI:10.1128/JB.01054-09.
[23]DEL RIO B, LADERO V, REDRUELLO B, et al. Lactose-mediated carboncataboliterepressionofputrescineproductionindairy Lactococcus lactis is strain dependent[J]. Food Microbiology, 2015,48(1):163-170. DOI:10.1016/j.fm.2014.11.018.
[24]CHENG C, CHEN J, FANG C, et al. Listeria monocytogenes aguA1,but not aguA2, encodes a functional agmatine deiminase:biochemical characterization of its catalytic properties and roles in acid tolerance[J].JournalofBiologicalChemistry,2013,288(37):26606-26615.DOI:10.1074/jbc.M113.477380.
[25]FERNáNDEZM,LINARESDM,RODRíGUEZA,etal.Factors affecting tyramine production in Enterococcus durans, IPLA 655[J].AppliedMicrobiology&Biotechnology,2007,73(6):1400-1406.DOI:10.1007/s00253-006-0596-y.
[26]PEREZM,CALLES-ENRíQUEZM,NESI,etal.Tyramine biosynthesisistranscriptionallyinducedatlowpHandimproves thefitnessofEnterococcusfaecalis,inacidicenvironments[J].AppliedMicrobiology&Biotechnology,2015,99(8):3547-3558.DOI:10.1007/s00253-014-6301-7.
[27]BOZKURTH.Utilizationofnaturalantioxidants:greenteaextract and Thymbra spicata oil in Turkish dry-fermented sausage[J]. Meat Science, 2006, 73(3):442-450. DOI:10.1016/j.meatsci.2006.01.005.
[28]ELíASM,CARRASCOSAAV.CharacterisationofthePaio doAlentejo,atraditionalPortugueseIberiansausage,inrespect toitssafety[J].FoodControl,2010,21(1):97-102.DOI:10.1016/j.foodcont.2009.04.004.
[29]雷志方,谢晶,尹乐,等.温度和姜精油对金枪鱼品质影响及生物胺相关性[J].食品科学, 2017, 38(3):45-52. DOI:10.7506/spkx1002-6630-201703008.
[30]FILIPPIS F D, PENNACCHIA C, PASQUA R D, et al. Decarboxylase geneexpressionandcadaverineandputrescineproductionby Serratia proteamaculans in vitro and in beef[J]. International Journal ofFoodMicrobiology,2013,165(3):332-338.DOI:10.1016/j.ijfoodmicro.2013.05.021.
[31]L A D E R OV,C A L L E S-E N RíQ U E ZM,F E R NáN D E ZM,etal.Toxicologicaleffectsofdietarybiogenicamines[J].Current Nutrition&Food Science,2010,6(2):145-156.DOI:10.2174/157340110791233256.