源于奶酪的产β-半乳糖苷酶乳酸片球菌的筛选及其合成低聚半乳糖特性  

Screening of Aβ-Galactosidase-producing Pediococcus acidilactici Strain from Cheese and Its Characteristics for Galactooligosaccharides Synthesis

作　　者：李欣 张秋涵 马子尧 关波[1,2] 胡有贞[1,2] 李谞 倪永清[1,2] LI Xin;ZHANG Qiuhan;MA Ziyao;GUAN Bo;HU Youzhen;LI Xu;NI Yongqing(Key Laboratory of Agricultural Product Processing and Quality Control of Specialty(Co-construction by Ministry and Province),School of Food Science and Technology,Shihezi University,Shihezi 832000,China;Key Laboratory for Food Nutrition and Safety Control of Xinjiang Production and Construction Corps,School of Food Science and Technology,Shihezi University,Shihezi 832000,China)

机构地区：[1]农业农村部特色农产品加工与质量安全控制重点实验室(部省共建),石河子大学食品学院,新疆石河子832000 [2]兵团食品营养与安全控制重点实验室,石河子大学食品学院,新疆石河子832000

出　　处：《食品工业科技》2025年第6期170-178,共9页Science and Technology of Food Industry

基　　金：国家自然科学基金(32060522);兵团科技创新人才计划青年拔尖人才项目(2024DB028);石河子大学成果转化与技术推广计划(CGZH202305)。

摘　　要：为了给益生性低聚半乳糖(GOS)的合成提供新的乳酸菌酶源,本研究从新疆传统奶酪中筛选具有高转糖苷活性的β-半乳糖苷酶产酶菌株。在以乳糖为单一碳源的MRS固体培养基中加入显色剂5-溴-4-氯-3-吲哚-β-D-半乳糖苷(X-gal),以菌落生长颜色进行初筛,以粗酶液酶活性以及转糖苷反应混合物的薄层色谱(TLC)进行复筛,结合其形态学、生理生化特征及16S rRNA序列同源性分析对产转糖苷活性的β-半乳糖苷酶菌株进行鉴定。单因素实验确定菌株的产酶条件和粗酶液的转糖苷反应条件,高效液相色谱分析转糖苷反应产物的组分及含量。筛选获得产转糖苷活性β-半乳糖苷酶的菌株6株,其中Pediococcus acidilactici Y1的产酶水平和转糖苷活性最高。单因素实验结果表明,P.acidilactici Y1在20 mg/mL乳糖,37℃培养18 h时,产酶水平最高可达(15.52±0.34)U/g。在初始乳糖浓度300 mg/mL,加酶量4.24 U/mL,50℃反应28 h时,GOS得率最高为38.4%±0.56%(w/w),乳糖残留量为30.9%±0.44%(w/w)。其中转移二糖、转移三糖以及转移四糖的质量分数分别为15.6%±0.21%(w/w)、18.3%±0.15%(w/w)和4.5%±0.01%(w/w)。以上结果表明,P.acidilactici Y1是一株产转糖苷活性β-半乳糖苷酶的新菌株,在益生性GOS的合成领域具有应用前景。This study aimed to provide a new lactic acid bacteria(LAB)enzyme source for synthesizing probiotic galactooligosaccharides(GOS).It screenedβ-galactosidase producing strains with high transglycosidase activity from traditional cheese in Xinjiang,China.A modified MRS medium with lactose as the single carbon source supplemented with 5-bromo-4-chloro-3-indole-β-D-galactoside was used for the first-round screening of LAB strains producingβ-galactosidases.Thin layer chromatography(TLC)analysis of the reaction products catalyzed by the crude enzyme from selected strains was used for the second-round screening.The isolate with the highest transglycosylation activity was identified based on the physiological and biochemical characteristics,along with the sequence analysis of the 16S rRNA gene.The conditions for enzyme production and transglycosylation reaction were determined by one-way experiment.TLC combined with high performance liquid chromatography was conducted to analyze the content of each GOS component.Six strains producingβ-galactosidase with transglycosidase activity were screened,among which Pediococcus acidilactici Y1 exhibited the highest enzyme production level and transglycosidase activity.After optimization,P.acidilactici Y1 producedβ-galactosidase at a level of up to 15.52±0.34 U/g(with 20 mg/mL of initial lactose after 18 h at 37℃).The highest GOS yield of 38.4%±0.56%(w/w)was achieved with an initial lactose concentration of 300 mg/mL,an enzyme amount of 4.24 U/mL,and a reaction temperature of 50℃ for 28 h.Also,the residual rate of lactose decreased to 30.9%±0.44%(w/w).The content of transfer disaccharide,transfer trisaccharide,and transfer trisaccharide was 15.6%±0.21%(w/w),18.3%±0.15%(w/w),and 4.5%±0.01%(w/w),respectively.The results of this study showed that P.acidilactici Y1 was a newβ-galactosidase-producing strain,with application prospects in the synthesis of probiotic GOS.

关 键 词：Β-半乳糖苷酶 转糖苷活性 低聚半乳糖 乳酸片球菌 奶酪 

分 类 号：Q815[生物学—生物工程]