植物乳杆菌C010产细菌素的分离纯化及理化稳定性分析  

Purification and physicochemical stability analysis of bacteriocin produced by Lactobacillus plantarum C010

作　　者：张漫敏 曾祥益 方利敏 徐静宇 程新[1] 姚明印 黄林[1,2] ZHANG Manmin;ZENG Xiangyi;FANG Limin;XU Jingyu;CHENG Xin;YAO Mingyin;HUANG Lin(School of Biological Sciences and Engineering,Jiangxi Provincial Engineering Laboratory of Agricultural Microbial Resources Development and Utilization,Institute of Applied Microbiology,Jiangxi Agricultural University(Jiangxi Agricultural University),Nanchang 330045,China;Key Experiment on Modern Agricultural Equipment,Jiangxi 330045,China)

机构地区：[1]生物科学与工程学院,江西省农业微生物资源开发与利用工程实验室,江西农业大学应用微生物研究所(江西农业大学),江西南昌330045 [2]江西省现代农业装备重点实验,江西南昌330045

出　　处：《食品与发酵工业》2023年第3期31-37,共7页Food and Fermentation Industries

基　　金：国家自然科学基金地区科学基金项目(31560482);江西农业大学研究生创新专项资金项目(YC2020-S250)。

摘　　要：为从植物乳杆菌C010 (Lactobacillus plantarum C010)发酵液中分离提纯出细菌素,实验采用硫酸铵沉淀、有机试剂萃取、pH吸附解析3种方法对L.plantarum C010发酵液进行粗步提取,通过Sephadex G-25、Sephadex LH-20凝胶层析结合半制备液相色谱法分离出单一抑菌活性组分物质,并对其结构及理化稳定性进行分析。结果表明,L.plantarum C010发酵液的3种粗提方法中,有机试剂萃取显著优于硫酸铵盐析和吸附解析(P<0.05);再经凝胶层析和半制备液相色谱分离得到单一组分抑菌活性物质,经EI质谱(electron ionization, EI)和核磁共振图谱分析,该活性物质分子质量为260.116 1 Da的环(L-苯丙氨酰-反式-4-羟基-L-脯氨酸)二肽,为首次从乳酸菌中分离提取到的细菌素(命名为Plantarum C010)。进一步理化稳定性分析,该细菌素对多类蛋白酶耐受性强,耐热性强、耐强酸,但pH>5.0时,对韩国假单胞菌PS1和梭状芽胞杆菌J4抑菌活性显著下降(P<0.05)甚至失活。该结果为L.plantarum C010所产细菌素的提纯及应用开发提供数据基础。To purify bacteriocin from the fermentation broth of Lactobacillus plantarum C010. The experiment compared three crude extraction methods from L. plantarum C010 fermentation broth including ammonium sulfate precipitation, organic reagent extraction and pH absorption-desorption. The single antibacterial active substance were obtained by Sephadex G-25 and Sephadex LH-20 gel chromatography combined with semi-preparative liquid chromatography, which were analyzed their chemical structural formula and physicochemical stability. The results showed that the organic reagent extraction method was significantly better than the other two methods(P<0.05). And the single bacteriostatic active substance was separated and obtained by combining gel chromatography with semi-preparative liquid chromatography, which was identified as bacteriocin(Plantarum C010) of Cyclo dipeptide(L-Phe-trans-4-hydroxy-L-Pro) with a molecular mass of 260.116 1 Da by electron ionization spectroscopy and nuclear magnetic resonance spectrum. This bacteriocin was first extracted from lactic acid bacteria, which was stable under high temperature, and protease attack(catalase, pepsin, trypsin, proteinase K, lipase, amylase). The bacteriocin activity has highly resistant to strong acids, when pH exceed 5.0, the inhibitory activity against Pseudomonas koreensis PS1and Bacillus fusiformis J4 was significantly decreased(P<0.05) or even inactivated. These results provide data basis for the purification and application development of the bacteriocin from L. plantarum C010.

关 键 词：植物乳杆菌C010 分离纯化 细菌素 环二肽 理化稳定性 

分 类 号：TS201.3[轻工技术与工程—食品科学]