Optimization of a Crude Deoxyribose-5-phosphate Aldolase Lyzate-Catalyzed Process in Synthesis of Statin Intermediates

Abstract

A process for providing intermediate compounds as building blocks for effectively producing statins is described. The presented process is based on acetoxyacetaldehyde and acetaldehyde as substrates, which are presented in an aldol reaction catalyzed by a crude deoxyribose-5-phosphate aldolase (DERA) expressing culture lysate. Different addition regimes of both reactants into a reaction mixture were applied. For the highest concentration of product ((2S,4R)-4,6-dihydroxytetrahydro-2Hpyran-2-yl)methyl acetate, in the presented crude DERA expressing culture lysate-catalyzed reaction used further in the production of statins, the best addition time of reactants is described. Improved process conditions and reactants' feeding regime were achieved by converting a batch reaction to a fed-batch process, reaching the highest concentration of product ((2S,4R)-4,6dihydroxytetrahydro-2H-pyran-2-yl)methyl acetate near 77 g/L. The complete process was designed in a practical and economical manner and could be used further on an industrial scale.

2‐Deoxyribose‐5‐phosphate aldolase from Thermotoga maritima in the synthesis of a statin side‐chain precursor: characterization, modeling and optimization

“…) . Spontaneous cyclization of 4 after the second addition step of 1 to 3 drives the overall equilibrium favorably because the high stability of the cyclized lactol form of the final product removes the open‐chain 4 from the aldol equilibrium . For the same reason, a further, third addition step of 1 to 4 is prevented .…”

“…By using achiral substrates, such as chloroacetaldehyde and acetaldehyde, DERA catalyzes a sequential aldol addition that results in an enantiomerically pure lactol, (3 R ,5 R )‐6‐chloro‐2,4,6‐trideoxyhexose . This 2,4,6‐trideoxyhexose is a valuable chiral synthon for HMG‐CoA reductase inhibitors, collectively known as statins, and used for the production of cholesterol‐lowering drugs, such as atorvastatin and rosuvastatin .…”

“…Therefore, the preparation of the 2,4,6‐trideoxyhexose has attracted immense effort because of its extremely high market price, and the requirement for high chemical and stereochemical purity at its two chiral centers . Because the chemical synthesis of statins is time‐consuming and consists of a number of synthetic steps that are carried out under very harsh conditions, the use of DERA for the synthesis of the statin side‐chains presents one of the most attractive and promising routes, due to its ability to introduce both stereocenters in a single step …”

“…Thus, large quantities of the enzyme are necessary to obtain industrially useful product yields . This issue can be circumvented by improving its resistance and catalytic activity using a molecular evolution approach, or by applying a cell‐free lysate or a whole‐cell biocatalyst instead of an isolated enzyme or by immobilization on appropriate supports …”

Recent Progress in the Synthesis of Super-Statins

“…Ručigaj and Krajnc [99] investigated the process of the crude DERA-lyzatecatalyzed reaction between 78 and 79a. Several parameters had relevant influence on the reaction outcome in terms of the ((2S,4R)-4,6-dihydroxytetrahydro-2H-pyran-2-yl)methyl acetate 82a yield.…”