纳豆芽孢杆菌固态发酵罗非鱼下脚料过程相关参数变化及产脂肽动力学

摘要

为了探明发酵参数与脂肽生成间的关系,实验以罗非鱼下脚料为主要基质接种纳豆芽孢杆菌,控制一定条件在三角瓶中进行固态发酵,测定发酵过程中菌体量、基质的温度、pH、水分含量、生物酶活性、蛋白含量及水解度和抗菌脂肽生成量;通过数学拟合建立底物消耗、菌体生长、脂肽生成的动力学模型,并分析各参数与脂肽生成和蛋白水解间的关系及其变化规律.结果显示,在下脚料固态基质中生长曲线为典型的S型,最大生长速率μm为0.2355×108 cfu/h,48 h菌体处于最大生长量,通过动力学模型拟合,符合Logistic模型.脂肽的生成趋势和菌体生长基本一致.同样符合Logistic模型,最大生成量为7.30 g/kg,最大生成速率为0.1112 g/h.在菌体生长过程中,基质中的蛋白酶活性在前期呈直线上升的趋势,48 h达到最高30304.56 U/g,随后迅速下降.在菌体生长前60 h,基质中的粗蛋白含量呈直线下降,最终水解度达26.26%,蛋白消耗动力学模型符合物料衡算理论模型.发酵过程中,基质的温度、pH和水分会发生小幅波动.研究表明,脂肽生成和菌体生长是偶联型;蛋白酶是同步合成型,蛋白酶产量可以为后续二步酶解制备小肽提供充足酶源;建立的动力学模型拟合度好,可以用于描述发酵过程和脂肽生成规律.本研究为利用罗非鱼下脚料固态发酵生产抗菌脂肽和营养小肽复合型活性肽工业化生产调控提供了重要的理论依据.%A 60 h solid state fermentation experiment was conducted to investigate the production technology of tilapia sp. byproduct as an antimicrobial product and source of nutritious feed. Crushed tilapia sp. byproduct and water was put in a triangular flask and then inoculated with 5% Bacillus Natto NT-6 with fermentation temperature at 30 ℃. Studies on changes in pH, temperature, ratio of water and enzymatic activity during the fermentation process was conducted. In addition to this, modelling of the antimicrobial concentration of the biomass and protein concentration was done. Analysis of the relationship between these parameters and lipopeptide yield was also made. The results showed that during fermentation the biomass increased for 48 h, then stabilized, the maximum growth rate was 0.2355×108 cfu/h. The production trend of lipopeptide is same as the biomass, the maximum yield was 7.3 g/kg and the maximum generation rate was 0.1112 g/h. The bacteria (B. Natto NT-6) secreted a high amount of enzymes, extraordinarily,the protease enzymatic activity was as highly as 30304.56 U/g. During the process, the temperature, pH and water content of the substrate was fluctuating slightly. On the bases of Logistic and material balance equation, the model of bacteria growth kinetics, comprehensive product generation and substrate consumption were set up and the non-linear fitting was employed to model the kinetics of fermentation by the software Origin 8.5. At the time,the models and the experimental data were well fitted and the relative error non-significant. The model developed in this study could predict the concentration of surfactin in biomass, substrate consumption and also help to determine whether the fermentation process was normal or not. This would enable a sound scientific control of surfactin fermentation process in real-time. This is of great significance to antimicrobial lipopeptides at large-scale production and application.