DPP-4抑制剂高产菌株的筛选、鉴定及发酵特性

Screening,Identification and Fermentation Characteristics of High-yield DPP-4 Inhibitors

目的:高产DPP-4抑制剂发酵菌株的筛选,并对菌株进行鉴定以及发酵特性的研究。方法:采用逐级稀释法从江南传统发酵食品中,利用DPP-4抑制剂体外筛选模型,筛选具有DPP-4抑制活性的菌株,以及通过形态学、生理生化和16S rRNA基因,对菌株进行观察、分析和序列比对,确定其种属关系。在以DPP-4抑制活性为指标下,通过与其他乳酸菌的比较、不同培养介质的选择、在脱脂乳(SKM)培养基中发酵条件的优化和发酵特性等。结果:从江南传统酸豆角汁中,在分离纯化得到11株具有乳蛋白水解能力的菌株中,经形态学、生理生化特征和16S rRNA基因序列分析,筛选得到一株多粘类芽孢杆菌(Paenibacillus polymyxa) BD3736。P.polymyxa BD3736以10%(W/V)脱脂乳(SKM)为培养基,厌氧培养2 d所得的发酵乳,对DPP-4抑制率为66.4%±2.9%,其抑制活性远大于乳杆菌属、双歧杆菌属和链球菌属等。P.polymyxa BD3736在10%(W/V) SKM液体培养基中,在30℃、180 r/min培养条件下,培养12 h时活菌数达到峰值为1.05×109CFU/mL,培养36 h时发酵体系的pH为5.58。在同等发酵条件下,P. polymyxa BD3736发酵2 d的发酵液上清,对DPP-4的抑制率为90.6%±2.6%,IC50值为(14.2±2.0) mg/m L。本研究为后续进一步开发新型、副作用低的口服降糖药物或功能性食品配料提供理论指导和技术支持。

Objectives: Screening of high-yield DPP-4 inhibitors fermentation strains,identification of strains and study of fermentation characteristics. Methods: Using the screening model for DPP-4 inhibitors,the strains with DPP-4 inhibitory activity were screened by the stepwise dilution method from the traditional fermented food in Jiangnan.The strains were observed and analyzed by morphological,physiological and biochemical and 16 S rRNA genes,to alignment to determine its relationship between species.Based on the DPP-4 inhibitory activity,the main research contents were compared with other lactic acid bacteria,selection of different culture media,optimization of fermentation conditions,and fermentation characteristics in skim milk( SKM) medium. Results: A strain of Paenibacillus polymyxa BD3736 was screened from the 11 strains with milk protein hydrolysis ability from Jiangnan traditional sour bean juice.The inhibition rate of DPP-4 of P.polymyxa BD3736 was fermented in 10%( W/V) skim milk( SKM) medium for 2 days under anaerobic condition was 66.4% ± 2.9%. Its inhibitory activity was much larger than Lactobacillus,Bifidobacterium and Streptococcus.The strain BD 3736 reached a peak of 1.05 × 109 CFU/mL in 10%( W/V) SKM liquid medium at 30 ℃ and 180 r/min for 12 h,and the pH of system was 5.58.Under the same fermentation conditions,the inhibition rate of DPP-4 of the supernatant of the strain BD3736 for 2 days was 90.6% ± 2.6%,and the IC50 value was( 14.2 ± 2.0) mg/m L. This study provides theoretical guidance and technical support for the further development of new oral hypoglycemic drugs or functional food ingredients with low side effects.