高压热灭菌中乳化剂吐温80的应用研究

Studies on the Application of Emulsifier Tween 80 in High-pressure Thermal Sterilization Technique

使用高压热HPTS结合质量分数为0.5%,1%,2%的吐温80处理枯草杆菌芽孢。用平板计数法测定吐温80溶液处理后对芽孢菌落存活数的影响,采用流式细胞术、比色法、ATP酶试剂盒、傅里叶变换红外光谱分析吐温80溶液对芽孢内膜通透性、DPA(2,6-吡啶二羧酸)释放、芽孢Na+/K+-ATP、Ca2+/Mg2+-ATP酶活以及蛋白二级结构稳定性的影响。结果表明:吐温80能够削弱HPTS对枯草杆菌芽孢的杀灭效果,对芽孢产生保护作用。当吐温80的质量分数为0.5%时,结合低压热(200 MPa,75℃)处理,芽孢的最大存活量可达5.3 lg(CFU/mL)。这种保护作用主要体现在吐温80溶液对芽孢的内膜通透性、关键物质DPA、ATP酶活以及芽孢蛋白二级结构稳定性的保护。动力学研究发现芽孢的灭菌效果与处理时间呈正比例相关,处理时间越长,芽孢的杀灭效果越强。HPTS结合吐温80的杀菌过程符合Weibull模型,且添加吐温80后Weibull模型的参数b值、n值均减小(n<1)。随着处理时间的延长,对温度与压力敏感的芽孢很快被杀灭,而一些超级深度休眠芽孢仍然存活。

The Bacillus subtilis spores were treated with High-pressure thermal sterilization combined with mass fraction fraction of 0.5%,1%,and 2%tween 80 on Bacillus subtilis.The effect of tween 80 solution treatment on the survival of spore colonies was determined by plate counting,The flow cytometry,colorimetric method,ATP enzyme kit,Fourier transform infrared spectroscopy was used to analyse the effect of tween 80 solution on spore inner membrane permeability,DPA(2,6-pyriddiarboxylic acid)release,spore Na+/K+-ATP,Ca2+/Mg2-ATP enzyme kit and and stability of protein secondary structure.The results showed that tween 80 could weaken the inactivating effect of HPTS on Bacillus subtilis and exert a protective effect on spores.When the tween 80 mass fraction was 0.5%,combining the low pressure heat(200 MPa,75℃),the maximum survival of spores could reach 5.3 lg(CFU/mL).It was found that protection of the inner membrane permeability,DPA,ATP enzyme activity,and the secondary structural stability of the spore protein.Kinetic studies found that the spore sterilization effect was positively proportional to the treatment time,and the longer the treatment time,the stronger the spore inactivating effect.The sterilization process of HPTS combined with tween 80 complied with the Weibull model,and the parameter b and n(n<1)values of the Weibull model decreased after the addition of tween 80.As the treatment time continued,the temperature and pressure sensitive spores were quickly inactivated,while some super-deep dormant spores were survived.