植物乳杆菌B02012对酸面团小麦蛋白结构和免疫特性的影响

本实验以植物乳杆菌（Lactobacillus plantarum）B02012和消化乳杆菌（Lactobacillus alimentarius）20998两株具有相近产酸速率和酸化性能的乳酸菌制备长时发酵酸面团（分别简称为SLB02012和SL20998），并以传统酵母菌长时发酵酸面团（SY）和可食性有机酸化学酸化面团（SCA）为对照，通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（sodium dodecyl sulfate-polyacrylamide gel electrophoresis，SDS-PAGE）、傅里叶变换红外光谱（Fourier transform infrared spectrophotometer，FTIR）、内源性荧光扫描（intrinsic fluorescence scanning，IFS）、竞争性酶联免疫吸附测定（enzyme linked immunosorbent assay，ELISA）并结合层次聚类分析（agglomerative hierarchical clustering，AHC）等方法研究不同发酵面团中小麦蛋白分子二、三级结构及麦醇溶蛋白免疫性的变化和相关性。SDS-PAGE结果显示，乳杆菌的添加对降解面团蛋白质起主导作用；与面团SL20998相比，面团SLB02012的高分子质量麦谷蛋白及部分麦醇溶蛋白发生了更明显降解。FTIR和IFS进一步验证，面团SLB02012的蛋白分子柔韧性增加，α-螺旋含量下降最明显，α-螺旋含量/β-折叠含量的比值最小；面团SY、SLB02012和SYLB02012的λmax发生蓝移，而SL20998的λmax发生红移，蛋白质三级结构得到伸展。ELISA结果表明，对比空白组，面团SLB02012麦醇溶蛋白的免疫性下降了35%，且与面团SCA差异不显著，而SL20998麦醇溶蛋白的免疫性增加了29.5%。AHC结果表明，不同发酵条件获得的发酵面团中麦醇溶蛋白的二、三级结构及其免疫性呈现良好的聚类关系。本研究结果表明，植物乳杆菌B02012可作为制备低敏发酵制品的优势菌，为进一步深入研究乳酸菌降敏机理提供理论支撑。

Impacts of Lactobacillus plantarum B02012 on Gluten Structure and Immunoreactivity in Sourdough

Two different sourdoughs, named SLB02012 and SL20998, were prepared respectively with Lactobacillus plantarum B02012 and Lactobacillus alimentarius 20998, having similar acid production rate and acidification performance in this study. Also, sourdoughs SYLB02012 and SYL20998 were made with each of the two strains in combined with yeast, respectively. Traditional yeast-fermented sourdough (SY) and non-fermented sourdough made with edible organic acids (SCA) as well as non-inoculated dough with erythromycin (CK) were used as controls. Gliadin was extracted from each sourdough and was evaluated for secondary and tertiary structures and immunoreactivity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Fourier transform infrared spectroscopy (FTIR), intrinsic fluorescence spectroscopy (IFS), enzyme linked immunosorbent assay (ELISA), and their correlation were analyzed by agglomerative hierarchical clustering (AHC). SDS-PAGE results indicated that protein hydrolysis in dough was mainly governed by lactic acid bacteria. High-molecular-mass glutenin (HMM-GS) and some gliadins in SLB02012 were more effectively degraded than in SL20998. Further, FTIR and IFS analyses indicated that the molecular flexibility of glutens in SLB02012 increased. Moreover, SLB02012 exhibited the most marked decrease in α-helix content and the smallest α-helix/β-sheet content ratio among the 7 doughs. In addition, SY, SLB02012 and SYLB02012 showed blue shifts of λmax, while SL20998 showed a red shift of λmax, implying that the tertiary structure of the proteins was extended. In terms of the immunoreactivity of gliadins as measured by ELISA , SLB02012 showed a decrease of 35% compared with CK, while no significant difference was noted for SCA and SL20998 increased by 29.5%. AHC showed a good correlation between the secondary and tertiary structures and the immunoreactiviity of gliadins for each of these sourdoughs. Taken together, Lactobacillus plantarum B02012 can be used as a dominant bacterium in low allergenic fermented foods, and this study provides theoretical support for understanding the mechanism of action of lactic acid bacteria in reducing allergenicity.