南山牧场牛乳中具有抑菌活性乳酸菌的筛选及鉴定
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摘要
该研究采用湖南南山高山牧场的鲜牛奶为原料,李斯特菌为指示菌,抑菌圈直径为评价指标,从中筛选抑菌特性较高的乳酸菌,通过形态观察和分子生物技术对其进行鉴定。同时,对菌株所产的抑菌物质的成分进行初步分析,并研究pH、温度及NaCl对其抑菌效果的影响。结果表明,从湖南南山高山牧场的鲜牛奶中共分离出28株乳酸菌,其中7株对李斯特菌具有抑菌活性,且菌株C15的抑菌能力最强,抑菌圈直径为6.17 mm。菌株C15被鉴定为乳酸乳球菌(Lactococcal lactis),初步判定其所产的抑菌物质成分为蛋白类或多肽类物质,该抑菌物质在酸性及中性条件下稳定,且具有较好的热稳定性和耐盐性,经80℃处理20 min后,抑菌活性保留54.35%;经10%NaCl(1 mol/L)处理2 h后,抑菌活性保留86.76%。
Using fresh milk from Nanshan pasture in Hunan province as raw material, Listeria as indicator bacteria, and inhibition zone diameter as evaluation indicator, the lactic acid bacteria with higher antibacterial properties were screened and identified by morphological observation and molecular biotechnology. At the same time, the components of the bacteriostatic substances produced by the strain were preliminary analyzed. The effects of p H, temperature and NaCl on the antibacterial effect were studied. The results showed that 28 strains of lactic acid bacteria were isolated from the fresh milk of Nanshan pasture in Hunan province, 7 of which had antibacterial activity against Listeria, strain C15 had the strongest antibacterial activity, and the inhibition zone diameter was 6.17 mm. The strain C15 was identified as Lactococcal lactis, and the antibacterial substance was protein or polypeptide substance. The bacteriostatic substance was stable under acidic and neutral conditions, and had good thermal stability and salt tolerance. After treatment at 80 ℃ for 20 min, the bacteriostatic substance activity retained 54.35%, after treated with 10% NaCl(1 mol/L) for 2 h, the antibacterial activity was 86.76%.
Using fresh milk from Nanshan pasture in Hunan province as raw material, Listeria as indicator bacteria, and inhibition zone diameter as evaluation indicator, the lactic acid bacteria with higher antibacterial properties were screened and identified by morphological observation and molecular biotechnology. At the same time, the components of the bacteriostatic substances produced by the strain were preliminary analyzed. The effects of p H, temperature and NaCl on the antibacterial effect were studied. The results showed that 28 strains of lactic acid bacteria were isolated from the fresh milk of Nanshan pasture in Hunan province, 7 of which had antibacterial activity against Listeria, strain C15 had the strongest antibacterial activity, and the inhibition zone diameter was 6.17 mm. The strain C15 was identified as Lactococcal lactis, and the antibacterial substance was protein or polypeptide substance. The bacteriostatic substance was stable under acidic and neutral conditions, and had good thermal stability and salt tolerance. After treatment at 80 ℃ for 20 min, the bacteriostatic substance activity retained 54.35%, after treated with 10% NaCl(1 mol/L) for 2 h, the antibacterial activity was 86.76%.
引文
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[18]任士菊,王俊钢,周红,等.冷水鱼肠道抑菌活性乳酸菌的筛选及其生理特性分析[J].食品工业科技,2014,35(14):178-181,185.
[19]O'SULLIVAN L,ROSS R P,HILL C.Potential of bacteriocin producing lactic acid bacteria for improvements in food safety and quality[J].Biochimie,2002,84(5/6):593-604.
[20]L譈X,LI L H,DANG J,et al.Purification of novel bacteriocin produced by Lactobacillus coryniformis MXJ 32 for inhibiting bacterial foodborne pathogens including antibiotic-resistant microorganisms[J].Food Control,2014,46:264-271.
[2]张香美,赵玉星,闫晓晶,等.1株具抑菌和抗氧化活性乳酸菌的筛选及鉴定[J].食品科学,2018,39(2):93-98.
[3]WIDYASTUTI Y,ROHMATUSSOLIHAT,FEBRISIANTOSA A.The role of lactic acid bacteria in milk fermentation[J].Food Nutr Sci,2014,5(4):435-442.
[4]孙强正,徐建国.乳酸乳球菌食品级表达载体的研究进展[J].中国微生态学杂志,2006,18(3):260-261.
[5]辛灵莹.乳酸乳球菌胞外多糖磷酸化及其生物活性研究[D].南京:南京师范大学,2012.
[6]杜琨.乳酸链球菌素的稳定性及抑菌特性研究[J].食品工业,2012,33(1):99-101.
[7]吕淑霞,白泽朴,代义,等.乳酸链球菌素(nisin)抑菌作用及其抑菌机理的研究[J].中国酿造,2008,5(9):87-91.
[8]倪萍,黄洁洁,纪文营,等.1株具广谱抑菌活性的乳酸乳球菌的分离鉴定与保鲜效果[J].食品与发酵工业,2013,39(4):53-57.
[9]廖荣华.南山牧场区域生态文化研究[D].长沙:中南林业科技大学,2007.
[10]谭才邓,朱美娟,杜淑霞,等.抑菌试验中抑菌圈法的比较研究[J].食品工业,2016,37(11):122-125.
[11]LIU G,REN L,SONG Z,et al.Purification and characteristics of Bifidocin A,a novel bacteriocin produced by Bifidobacterium animals BB04 from centenarians'intestine[J].Food Control,2015,50:889-895.
[12]房华,汪瑞忠,张晓雪,等.6种中草药对多重耐药菌的抗菌活性及逆转耐药性作用的体外研究[J].国际检验医学杂志,2018,39(4):396-399.
[13]ARASU M V,AL-DHABI N A,REJINIEMON T S,et al.Identification and characterization of Lactobacillus brevis P68 with antifungal,antioxidant and probiotic functional properties[J].Ind J Microbiol,2015,55(1):19-28.
[14]付晓艳,霍贵成.利用16S r DNA部分序列聚类分析鉴定乳球菌[J].中国乳品工业,2005,33(8):7-9.
[15]于微,高学军,马春丽,等.干酪乳杆菌产细菌素的生物学特性分析[J].安徽农业科学,2014,42(32):11542-11543,11570.
[16]唐春梅,陈俊亮,任广跃.一株乳酸乳球菌所产细菌素的生物学特性[J].食品科学,2013,34(1):248-251.
[17]侯运华,孔健,郝运伟,等.一株乳酸菌产类细菌素enteriocin LK-S1的初步研究[J].山东大学学报(理学版),2002,37(5):463-466,470.
[18]任士菊,王俊钢,周红,等.冷水鱼肠道抑菌活性乳酸菌的筛选及其生理特性分析[J].食品工业科技,2014,35(14):178-181,185.
[19]O'SULLIVAN L,ROSS R P,HILL C.Potential of bacteriocin producing lactic acid bacteria for improvements in food safety and quality[J].Biochimie,2002,84(5/6):593-604.
[20]L譈X,LI L H,DANG J,et al.Purification of novel bacteriocin produced by Lactobacillus coryniformis MXJ 32 for inhibiting bacterial foodborne pathogens including antibiotic-resistant microorganisms[J].Food Control,2014,46:264-271.