ISSN print edition: 0366-6352 ISSN electronic edition: 1336-9075 Registr. No.: MK SR 9/7 Published monthly

Ketoreductase-assisted synthesis of chiral selective tert-butyl{5-[(4-cyanophenyl)(hydroxy)methyl]-2-fluorophenyl}carbamate: process minutiae, optimization and characterization

Amol Gulab Khatik, Abhijeet Bhimrao Muley, and Arvind Kumar Jain

School of Basic and Applied Sciences, Galgotias University, Gautam Buddh Nagar, Greater Noida, India

E-mail: amol.khatik@gmail.com

Received: 5 August 2022  Accepted: 19 January 2023

Abstract:

Ketoreductases capable of performing chiral selective reduction in tert-butyl[5-(4-cyanobenzoyl)-2-fluorophenyl]carbamate to tert-butyl{5-[(4-cyanophenyl)(hydroxy)methyl]-2-fluorophenyl}carbamate were screened, and ES-KRED-213 and KRED-P1-H01 were the best ones for targeted biotransformation. Dimethylsulfoxide (10% V/V) was the best co-solvent, and the optimum parameters were 40 °C, pH 7.0, 10% enzyme loading, and 100 g/L substrate loading for maximum conversion and > 99% chiral selectivity. Gram scale batches were performed at different temperature (30 °C, 40 °C, and 50 °C), enzyme loading (5%, 10%, 15%, and 20%), and substrate loading (50 g/L, 100 g/L, and 150 g/L) and assessed for timely progression of reaction, and then, product formation and yield were calculated. ES-KRED-213 showed 99.4% conversion, 4.9 g/50 mL product formation, with an actual product recovery of 4.2 g corresponding to a product yield 85% with respect to the product formed after reaction. The purity of tert-butyl{5-[(4-cyanophenyl)(hydroxy)methyl]-2-fluorophenyl}carbamate formed was ≥ 99% (RP-HPLC) and chiral purity ≥ 99% (NP-HPLC). The recovered product was confirmed and characterized with instrumental analysis using HPLC analysis, specific optical rotation, melting point and boiling point, LC–MS, ATR-FTIR, ¹H NMR, and ¹³C NMR.

Keywords: Ketoreductase; Reduction; Tert-butyl[5-(4-cyanobenzoyl)-2-fluorophenyl]carbamate; Tert-butyl{5-[(4-cyanophenyl)(hydroxy)methyl]-2-fluorophenyl}carbamate; Chiral selective

Full paper is available at www.springerlink.com.

DOI: 10.1007/s11696-023-02698-3

Chemical Papers 77 (6) 3213–3227 (2023)