高效液相色谱法检测枯草芽孢杆菌发酵液中乙偶姻和2,3-丁二醇含量

枯草芽孢杆菌可发酵代谢产生乙偶姻和2,3-丁二醇(两者可逆转化),本研究建立了用高效液相色谱法同时检测发酵液中乙偶姻和2,3-丁二醇的方法。对比了不同色谱柱和流动相配比对乙偶姻和2,3-丁二醇含量的测定结果,进一步检测枯草芽孢杆菌发酵液中乙偶姻和2,3-丁二醇含量。结果表明:采用Bio-Rad Aminex HPX-87H Column分析;流动相:0.005 mol/L的硫酸溶液;流速:0.5 mL/min;柱温:28℃;示差检测器,在30 min内完成样品检测,乙偶姻和2,3-丁二醇标准曲线相关系数分别为0.9997和0.9998,精密度标准偏差分别为1.76%和1.74%,稳定性标准偏差分别为1.2%和1.6%,回收率为98.93%~102.10%,相对标准偏差≤ 2%。此方法简单准确,检测目标物浓度为50 g/L,检测结果重现性好,发酵液中乙偶姻和2,3-丁二醇含量呈负相关,乙偶姻的含量随着2,3-丁二醇的增加而降低,该方法适合枯草芽孢杆菌发酵液中乙偶姻和2,3-丁二醇的发酵检测。

Detection of Acetoin and 2,3-Butanediol in Fermentation Broth of Bacillus subtilis by High Performance Liquid Chromatography

The metabolism Bacillus subtilis could produce acetoin and 2,3-butanediol,and a new method was established in this paper to detect the method of acetoin and 2,3-butanediol in fermentation broth by high performance liquid chromatography. The results of determination of acetoin and 2,3-butanediol by different chromatographic columns and flow matching ratio were compared and the detection conditions were optimized. The content of acetoin and 2,3-butanediol in the fermentation broth of Bacillus subtilis was further determined.The results showed that the Bio-Rad Aminex HPX-87H column was used to complete the sample detection,and the mobile phase was 0.005 mol/L of sulfuric acid solution,flow rate was 0.5 mL/min,column temperature was 28℃,refractive index detector was used,and the sample test was completed within 30 min.The correlation coefficients of the standard curve of acetoin and 2,3-butanediol were 0.9997 and 0.9998,respectively,the precision standard deviation was 1.76% and 1.74%,the stability standard deviation was 1.2% and 1.6%,the recovery rate was 98.93%~102.10%,and the relative standard deviation was ≤ 2%. This method was simple and accurate,the detection target concentration was 50 g/L,the test results were good on reproduction. The contents of acetoin and 2,3-butanediol in fermentation solution was negatively correlated,and the content of acetoin was decreased with the increasing of 2,3-butanediol,which was suitable for the detection of acetoin and 2,3-butanediol in the fermentation broth of Bacillus subtilis.