牦牛酸奶中一株具有抗氧化活性乳酸菌的分离及鉴定

本研究旨在从传统发酵牦牛酸奶中筛选获得具有抗氧化活性的乳酸菌。采用平板计数法对牦牛酸奶样品进行总菌与乳酸菌的菌落计数,采用添加2.5% CaCO₃的M17固体培养基进行乳酸菌的分离筛选,对筛选出的菌株采用16S rDNA基因克隆测序进行分子鉴定以及产酸、糖醇发酵试验,通过测定菌株培养上清液、菌体裂解液和菌体PBS重悬液的总抗氧化能力、DPPH自由基清除能力和羟自由基清除能力,评价乳酸菌的抗氧化活性。结果表明,分离出的革兰氏阳性球形菌株X1具备产酸特性,糖醇发酵实验表现为屎肠球菌生化反应特征,16S rDNA克隆测序鉴定菌株X1为屎肠球菌。体外抗氧化活性检测发现,菌株X1培养上清液的总抗氧化能力与DPPH自由基清除率显著高于对应的菌体裂解液和菌体PBS重悬液(P<0.05);而菌体PBS重悬液对羟自由基清除率显著高于培养上清液和菌体裂解液(P<0.05)。本试验成功从牦牛发酵酸奶中分离出一株具备抗氧化活性的屎肠球菌X1,其抗氧化活性物质主要存在于菌体培养上清液中,该结果为将牦牛酸奶来源乳酸菌开发为新型抗氧化微生态制剂、缓解畜禽养殖过程中的氧化应激提供菌种来源。

Screening and Identification of a Lactic Acid Bacterium Strain with Antioxidant Activity from Yak Yogurt

The study was conducted to screen and identify lactic acid bacterium with antioxidant activities from yak yogurt. The number of total bacterium and lactic acid bacterium strain was enumerated by plate counting method. The lactic acid bacterium was isolated by M17 agar supplemented with 2.5% CaCO₃.The isolated strain was molecularly identified by 16S rRNA gene cloning and sequencing,and acid producing test and biochemical reaction test was also conducted to determine the biological characteristics of the isolated strain. Total antioxidative activity,DPPH free radical scavenging ability and hydroxyl radical scavenging ability was analyzed to determine the in vitro antioxidant activities of the isolated strain. The results showed that the isolated gram-positive lactic acid bacterial strain X1 identified as an acid-producing bacterium,with the biochemical characteristics of Enterococcus faecium. Furthermore,16S rRNA gene sequencing result confirmed that the isolated strain X1 belongs to the genus Enterococcus faecium. In vitro anti-oxidative activity analysis showed that the total anti-oxidative ability and the DPPH free radical scavenging ability of the cultured supernatant was significantly greater compared with the bacterial lysates and the bacterial resuspension,respectively(P<0.05). However,the hydroxyl radical scavenging ability of the bacterial resuspension was significantly greater compared with the cultured supernatant and the lysates(P<0.05). In conclusion,in the current study, Enterococcus faecium X1 with antioxidant activities was isolated and identified successfully from yak yogurt,and the antioxidant substances mainly existed in the cultured supernatant of this strain. These results could provide references for developing yak yogurt derived lactic acid bacterium into a novel antioxidative probiotic feed additive,and would be valuable for reducing oxidative stress during animal production.