苯丙氨酸解氨酶合成L-2-氯苯丙氨酸的研究

Synthesis of 2-chloro-L-phenylalanine by Phenylalanine Ammonia Lyase

L-2-氯苯丙氨酸要求旋光对映体过量(ee)≥98%,导致使用化学拆分法合成L-2-氯苯丙氨酸的收率不足50%。为提高收率,文章采用具有立体专一性的苯丙氨酸解氨酶(PAL),催化邻氯肉桂酸与氨合成L-2-氯苯丙氨酸,旋光对映体过量(ee)>99%。通过考察不同形态PAL酶的稳定性,研究得出全细胞冻融法制作的PAL酶稳定性较好,结合氮气保护,反应80 h依然能保留62%的酶活。同时,优化PAL酶催化反应的pH、温度、酶用量,确定最佳条件后,结合分段补料的方式进行酶催化反应,最终使L-2-氯苯丙氨酸的累积量达到了68.5 g/L,转化率达到了97.8%。该技术合成L-2-氯苯丙氨酸的产量和收率较高,有效降低了成本,提升了L-2-氯苯丙氨酸的市场价值。

The synthesis yield of 2-chloro-L-phenylalanine by chemical resolution is less than 50%,due to the requirement of optically active enantiomer excess(ee)逸98%.In order to improve the yield,this article uses a stereospecific phenylalanine ammonia lyase(PAL)to catalyze the synthesis of 2-chloro-L-phenylalanine from 2-chlorocinnamic acid and ammonia,the product with an optically active enantiomer excess(ee)>99%.By investigating the enzyme stability of different forms of PAL,it finds that the stability of PAL produced by whole cell freeze-thaw method is better.Combined with nitrogen protection,the enzyme remains 62%active after 80 hours of reaction.The antioxidant activity of the enzyme is Successfully improved and the effective reaction time is prolonged.At the same time,this study optimizes the pH value,temperature,enzyme dosage for PAL catalysis,and after determining the best conditions,carries out the enzyme catalyze reaction in combination with the method of fractional feeding.Finally,the high yield of 2-chloro-L-phenylalanine achieves 68.5 g/L,with the conversion rate of 97.8%.The synthetic output and yield of 2-chloro-L-phenylalanine by using this technology are relatively high,which effectively reduces the cost and enhances the market value of 2-chloro-L-phenylalanine.