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CC BY 4.0 · Pharmaceutical Fronts 2022; 04(04): e267-e274
DOI: 10.1055/s-0042-1759851
Original Article

Process Study on the Enzyme-Catalyzed Preparation of Key Chiral Intermediates for Saxagliptin

Shan-Shan Li
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry Co,. Ltd., China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
,
Zong-Qing Huang
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
,
Hao-Ju Hua
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
,
Jian-Guang Lu
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
,
Wen-Jie Zhao
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry Co,. Ltd., China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
,
Jun Feng
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry Co,. Ltd., China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
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Abstract

Saxagliptin is a therapeutic drug for diabetes. The key synthesis process of the drug involves catalyzing 2-(3-hydroxy-1-adamantyl)-2-oxoacetic acid (A) into (S)-3-hydroxyadamantane glycine (B), during which enzymes phenylalanine dehydrogenase mutant from Thermoactinomyces intermedius (TiPDHm) and formate dehydrogenase (FDH) were most often used for biocatalysis. However, the process was limited due to difficulty in enzyme preparation and a low conversion rate. This study focuses on co-expression of TiPDHm and FDH in recombinant Escherichia coli, cell homogenate clarification, enzyme concentration as well as the optimized conditions of enzyme-catalyzed reaction. Our data showed that the wet weight density of bacteria reached 300 g/L, and the yields of TiPDHm and FDH were 7674.24 and 2042.52 U/L, respectively. The combination of ammonium formate and polyethyleneimine favors the clarification of the bacteria homogenate. The clarified enzyme solution obtained can be concentrated by ultrafiltration and directly used in a reductive amination reaction in a high concentration of keto acid A. The reaction time was only 12 hours and the conversion rate reached 95%. Therefore, this process could provide a reference for enzyme-catalyzed preparation of saxagliptin on an industrial scale.

Keywords

saxagliptin - phenylalanine dehydrogenase - formate dehydrogenase - clarification - biocatalysis - reductive amination

Supplementary Information

The results of liquid chromatography-tandem mass spectrometry (LC-MS) analysis and chiral analysis of product B are shown in the Supporting Information ([Supplementary Figs. S1] and [S2] [online only]).


Supplementary Material



Publication History

Received: 22 May 2022

Accepted: 15 November 2022

Article published online:
26 December 2022

© 2022. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/)

Georg Thieme Verlag KG
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