Enzymatic Preparation Of S-Citalopram Key Intermediate And Its Kinetics Study

Citalopram is a highly selective inhibitor of serotonin(5-HT) reuptake which contains a quaternary chiral center and almost the entire inhibitory activity resides in the S-enantiomer.S-citalporam,the single enantiomer drug with advantage of low dose and high efficacy,is one of the best-selling antidepressant and gradually replace the racemic citalopram.In this dissertation,the enzymatic preparation of citalopram chiral key intermediate was described.The main contents were as follows:1. Lipase-catalyzed remote resolution of citalopram key intermediate and kinetics study; 2.The separation of chiral diol and diol acetate after kinetic resolution;3.The utilization of unwanted R-enantiomer;4.S-citalopram synthesis and scale-up.Firstly,the lipase-catalyzed kinetic resolution of citalopram intermediate in organic media and its kinetics was studied.The lipase-catalyzed asymmetric transestereaction was superior to the asymmetric alcoholysis.The lipase-catalyzed remote kinetic resolution of diol,a key intermediate of citalopram,in organic media was studied.Through screening,the preferable system for the asymmetric transestereaction was:Novozym 435 as catalyst,acetonitrile as solvent and vinyl acetate as acyl donor.The influence of temperature,rotation speed, enzyme concentration and substrate concentration on the enzymatic resolution was studied.Reusability study of lipase was also carried out.The optimal condition was as following:60 mmol/L diol,the molar ratio of diol to vinyl acetate,1 to 5,10 mg/mL lipase,250 r/min rotation speed and 30℃.After 22h reaction,the yield of S-diol was 40.4%and the e.e._s was 99.1%.The E value of the reaction was 25.3 and the half-life of the lipase was 364.8 h.The kinetic modeling of Novozym 435 catalyzed remote resolution of diol was studied using vinyl acetate as acylating agent in solvent-free system.The substrate inhibition by each enantiomer of diol was proved.Enzyme deactivation and product inhibition were studied and could be negligible.The diffusion limitation was proved to be negligible by the calculation of Thiele module and Biot number.A kinetic model based on ping-pong bi-bi mechanism with competitive substrates using King-Altman method was proposed.The spontaneous transesterification was also considered.The reaction rate equation of both R- and S-diol acetate were derived.The model parameters were successfully simulated by Matlab program using time-concentration curves of different diol concentrations and the simulated values fitted the experimental values well with an average relative error of 12.8%.Furthermore,the developed model was used to investigate the effect of diol concentration on reaction enantioselectivity.The lipase-catalyzed asymmetric alcoholysis of diol acetate has been studied. Through screening,the preferable system for the asymmetric reaction was:Novozym 435 as catalyst,isobutyl alcohol as acyl accepter and the mixed solvent(acetonitrile: methyl tert-butyl ether=3:1).The influence of various parameters on the enzymatic resolution was studied and the reaction condition was optimized.A kinetic model based on ping-pong bi-bi mechanism with competitive substrates was proposed.The product inhibition by each enantiomer of diol and substrate inhibition by isobutyl alcohol were also considered.The reaction rate equation of both R- and S- diol were derived.The model parameters were successfully simulated by Matlab program using time-concentration curves of different diol acetate concentrations and the simulated values fitted the experimental values well with an average relative error of 11.3%.Secondly,a derivatization-extraction method was developed to separate diol from diol acetate after lipase-catalyzed kinetic resolution.It was found that succinic anhydride was the best derivative agent and water/toluene was the optimal extraction system.The effects of pH and initial diol acetate concentration on the total distribution coefficients of diol acetate were investigated.With the extraction conditions of initial diol acetate concentration of 10 mmol/L,pH 8.0 and 25℃,the total distribution coefficient achieved 157.7.The practical extraction process of the mixture of diol(ee value 99.5%) and diol acetate obtained by lipase-catalyzed reaction was carried out,within three extraction stages,the ee value and the yield of S-diol were 98.4%and 86.0%,respectively.This extraction method with anhydrides derivatization is promising in the separation of aromatic amino-alcohols for its low expense and easy scale up. Thirdly,the unwanted R-enantiomer was converted into S-citalopram by cyclic resolution and stereoinversion.The cyclic resolution of the R-diol with low optical purity obtained from the first resolution was carried out by asymmetric transestereaction.Using the kinetic model to predict the reaction process,R-diol acetate with ee value of 91.2%was obtained by controlling the conversion of cyclic resolution under 20.8%.R-diol was stereoinverted to S-citalopram by cyclization in acidic condition with 98.0%yield and ee value of 91.0%.S-citalopram with ee value of 98.0%was obtained by cyclization of S-diol with ee value of 98.2%in alkaline condition and the yield was 97.8%.Fourthly,the reaction volume of enzymatic asymmetric transestereaction of diol was scale-uped to 1 L and 14 L,gradually.There was no significant scale-up effect and the kinetic model fitted the result well.Scale-up of S-citalopram synthesis route was carried out for industrial production.