Method for preparing ursodeoxycholic acid through cyclic enzyme catalysis

Abstract

The invention discloses an application of acetone / isopropanol circulation in preparation of ursodeoxycholic acid (UDCA) by an enzyme method, and provides an environment-friendly, economic and efficient new idea for preparation of UDCA by a two-step enzyme method. The method comprises the following steps of 1, reducing acetone into isopropanol by using ketoreductase, and converting NADPH / NADH intoNADP+ / NAD+ to provide NADP+ / NAD+ for catalyzing chenodeoxycholic acid (CDCA) to generate 7KLCA by using 7alpha steroid dehydrogenase (7alpha HSDH); in the second step, isopropanol being oxidized intoacetone through isopropanol dehydrogenase, NADP+ / NAD+ being converted into NADPH / NADH, and NADPH / NADH being provided for catalyzing 7KLCA to generate UDCA through 7beta steroid dehydrogenase (7beta HSDH). According to the two-step enzyme method, immobilized enzyme is utilized, the process is simple, second-step reaction can be carried out through simple treatment after first-step reaction is finished, and intermediate purification is not needed; the addition amount of NADP+ / NAD+ in the reaction is small, and acetone and isopropanol can be recycled, so production cost and the environmental protection pressure are greatly reduced.

Description

technical field [0001] The invention relates to the field of biocatalysis, in particular to a method for preparing ursodeoxycholic acid catalyzed by circulating enzymes. Background technique [0002] Ursodeoxycholic acid (UDCA), whose chemical name is 3α,7β-dihydroxy-5β-cholestane-24-acid, is an organic compound with an odorless, bitter taste. In medicine, it is used to increase the secretion of bile acid, change the composition of bile, reduce cholesterol and cholesterol lipid in bile, and help the cholesterol in gallstones dissolve gradually. It is used for the treatment of gallstones, cholestatic liver disease, fatty liver, and various types of hepatitis. , toxic liver disorders, cholecystitis, cholangitis and biliary dyspepsia, bile reflux gastritis, eye diseases, etc. [0003] In recent years, the domestic UDCA market has experienced explosive growth. The current production process is generally a chemical process, which extracts CDCA from animal bile and then uses chem...

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Problems solved by technology

Patent CN106086149A discloses a method for preparing UDCA by chemical-enzymatic method. CDCA is oxidized to 7-KLCA with sodium hypochlorite, and 7-KLCA is catalyzed by enzymatic method to UDCA. The problem with chemical oxidation is that the residual oxidant will affect the next step of enzymatic catalysis, although Treated with sodium sulfite, but there are still unstable factors, which is not conducive to large-scale stable production. The glucose used for the regeneration of the coenzyme catalyzed by the second step generates a large amount of sodium gluconate after the reaction. Although it has no effect on the reaction itself, the resulting sewage However, the processing cost is very high, which does not meet the current increasingly stringent environmental protection policies. Patents such as CN107995929A, CN109055473A, and CN110643584A all use glucose coenzyme regeneration systems, while some patents use coenzyme regeneration systems such as lactic acid and sodium malate. There is a certain pressure of sewage treatment

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