ONE-POT CHEMOENZYMATIC SYNTHESIS OF 3'-FUNCTIONALIZED CEPHALOSPORINES(CEFAZOLIN) BY 3 CONSECUTIVE BIOTRANSFORMATIONS IN FULLY AQUEOUS-MEDIUM

We illustrate a new chemoenzymatic synthesis of cefazolin from cephalo sporin C, involving three consecutive biotransformations in full aqueo us medium. This one-pot three-step synthesis includes the D-amino acid oxidase catalyzed oxidative deamination of the cephalosporin C side c hain, hydrolysis of the resulting glutaryl derivative catalyzed by glu taryl acylase, and the final penicillin G acylase (PGA)-catalyzed acyl ation of 7-aminocephalosporanic acid (1, 7-ACA). The product, do]-3-(a cetoxymethyl)-Delta(3)-cephem-4-carboxylic acid (5), was used as an in termediate for cefazolin synthesis by 3'-acetoxy group displacement wi th 2-mercapto-5-methyl-1,3,4-thiadiazole. Very high yields have been a chieved with all the enzymatic reactions performed; high product conce ntrations were obtained in short reaction times. This synthetic approa ch presents several advantages when compared with the conventional che mical processes. The use of the toxic reagents and chlorinated solvent s is avoided, while the substrate specificity and chemoselectivity of the enzymes makes reactive group protection and intermediate purificat ion unnecessary. The enzymatic deacylation of cephalosporin C was perf ormed by the simultaneous use of D-amino acid oxidase and glutaryl acy lase. The substrate specificity of PGA allowed the acylation of 7-ACA (1) to be performed without purification from the glutaric acid produc ed during the enzymatic deacylation. These results were achieved by op timization and correct assembly of the different biotransformations in volved. Special attention has been applied to the kinetically controll ed acylation reaction. High yields were obtained through a careful sel ection of the enzyme catalyst, experimental conditions, and synthetic strategy.