酶-化学级联催化制备(S)-烟碱

Production of(S)-nicotine by enzymatic-chemical cascade catalysis

传统(S)-烟碱合成方法步骤复杂且依赖多种化学催化剂。为了实现(S)-烟碱的绿色合成,设计了酶-化学级联途径:首先,构建了亚胺还原酶(IRED)与甲酸脱氢酶(FDH)的偶联体系,以麦斯明为底物合成(S)-降烟碱;然后,将(S)-降烟碱经Eschweiler-Clarke反应制备(S)-烟碱。通过大肠杆菌宿主分别对IRED和FDH进行表达,得到含酶细胞及酶液,考察了pH和温度对细胞中IRED和FDH酶活力的影响。优化酶催化制备(S)-降烟碱的反应条件,在30℃、pH 7.5条件下,添加质量浓度为30 g/L的麦斯明、600 U/L FDH细胞、900 U/L IRED细胞、0.6 mmol/L NADP^(+)和质量浓度为90 g/L的甲酸钠,(S)-降烟碱的产率>98%,对映体过量(e.e.)值>99%。最后,通过化学法将(S)-降烟碱甲基化得到(S)-烟碱,此步骤产率和e.e.值均达99%以上。

The traditional synthesis method of (S)-nicotine involves complex steps and relies on multiple chemical catalysts.Herein,a enzymatic-chemical cascade pathway was designed to achieve the green synthesis of (S)-nicotine.Firstly,a coupling system of imine reductase (IRED) and formate dehydrogenase (FDH) was constructed to catalyzed the synthesis of (S)-nornicotine from myosmine as substrate,then (S)-nicotine was prepared via Eschweiler-Clarke reaction of (S)-nornicotine.The IRED and FDH enzyme-containing cells and enzyme solution were obtained from E.coli expression system and the effects of pH and temperature on the enzyme activity of IRED and FDH were investigated.The yield of (S)-nornicotine reached>98%with enantiomeric excess (e.e.) value of>99%under the optimal reaction conditions of temperature 30℃,pH 7.5,myosmine mass concentration 30 g/L,FDH cells 600 U/L,IRED cells 900 U/L,NADP~+0.6 mmol/L and sodium formate mass concentration 90 g/L Finally,(S)-nicotine with a yield and e.e.value of above 99%was obtained by chemical methylation of(S)-nornicotine.