川西高原牧区牦牛酸乳中的几株乳杆菌发酵特性及系统发育研究

Fermentation Characteristics and Phylogeny of Several Lactobacilli Strains Isolated from Spontaneously Fermented Yak Yoghurt from Western Sichuan Plateau

通过感官评定和酸度测定,从川西高原自然发酵牦牛奶酸乳中分离的109株乳酸菌中筛选出6株发酵酸乳感官品质良好的乳杆菌,进一步研究这6株乳杆菌的生长曲线、产酸特性、产黏特性、产乙醛特性,并采用16S rDNA基因序列分析,初步研究其系统发育关系。结果表明:在42℃条件下,6株乳杆菌在MRS培养基中培养14h、在10%的脱脂乳中培养8h时,达到最高生长量;6株乳杆菌发酵10%脱脂乳凝乳时酸度都达到GB2746—1999的要求,凝乳黏度为1050～4900mPa·s,乙醛产量为15.11～26.64μg/mL。冷藏中菌株E-1、E-5产酸量小,后酸化能力弱。6株乳杆菌单株凝乳冷藏中黏度在第5天达最高,其中菌株E-1、E-5的凝乳黏度变化小;菌株J-2、E-5发酵乳冷藏乙醛含量在第3天达到最高,其余菌株在第2天达到最高。16S rDNA基因序列的相似性分析表明,菌株4-7、E-1、E-5之间的相似度达到99%以上,这些菌株在系统发育树上与植物乳杆菌(L.plantarum)处于发育树的同一分支;菌株J-2与副干酪乳杆菌(L.paracasei)处于发育树的同一分支,与代表菌株L.paracaseiSM63的相似度为97.6%;菌株4-1和B-2的相似度达到99%,与发酵乳杆菌(L.fermentum)处于发育树的同一分支。综合研究结果表明菌株E-1和E-5可用于开发酸乳发酵剂菌种。

According to sensory evaluation and acidity determination,6 Lactobacilli strains（4-1,4-7,B-2,E-1,E-5,J-2） were selected from 109 isolated from spontaneously fermented yak yoghurt samples from western Sichuan plateau.The fermentation characteristics of these strains,including growth curve,acid production,fermented yoghurt viscosity,acetaldehyde synthesis were compared.Along with this,the phylogenic analysis based on 16S rDNA sequence was performed.Results showed that maximum growth was observed in the strains cultured for 14 h in MRS medium or 8 h in 10% skim milk at 42 ℃,and the acidity of yoghurt fermented with each of them reached the Chinese national standard of GB2746—1999 for yoghurt,and the viscosity and acetaldehyde amount were in the ranges of 1050-4900 mPa·s and 15.11-26.64μg/mL,respectively.Strains E-1 and E-5 revealed weaker acid-producing ability during cold storage when compared with other four strains.The highest values of viscosity and acetaldehyde amount of yoghurt fermented with each strain appeared on the 5th and 2nd day of cold storage.Furthermore,only slight change was observed in the viscosity of yoghurt fermented with strains E-1 or E-5.The highest acetaldehyde content in yoghurt fermented with strains J-2 or E-5 was observed on the 3rd day of cold storage,while that in yoghurt fermented with other four strains alone on the 2nd day.Homology analysis of 16S rDNA gene sequence indicated that there was a homology above 99% among strains 4-7,E-1 and E-5 and these strains and L.plantarum strains were included in the same branch of phylogenetic tree based on 16S rDNA gene sequence.Strain J-2 belonged to the L.paracasei family,and exhibited a homology of 97.6% to the representative stain of this family,L.paracasei SM63（AJ441107 ）.There was a homology of 99% between strains 4-1 and B-2,which were included in the L.fermentum branch.All these results showed that strains E-1 and E-5 had a good potential as starters for yoghurt production.