Method for resolving Lisinopril hydride through biocatalysis

A technology of biocatalysis and hydride, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problem of low selectivity and achieve the effects of good stereoselectivity, low environmental pollution and increased stability

Active Publication Date: 2017-08-18
CHANGXING PHARMA
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0006] At present, the synthetic route of lisinopril is mainly based on 3-ethyl benzoyl acrylate as a starting material, which undergoes an asymmetric addition reaction with trifluoroacetyl lysine under the action of lithium hydroxide, and then undergoes an asymmetric addition reaction with Pd / C hydrogenation reduction, cyclization, condensation, hydrolysis and other reaction processes are prepared. This process has low selectivity through asymmetric addition reaction, and the content ratio of (S,S)-type: (R,S)-type is usually 75 : 25

Method used

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  • Method for resolving Lisinopril hydride through biocatalysis
  • Method for resolving Lisinopril hydride through biocatalysis
  • Method for resolving Lisinopril hydride through biocatalysis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Embodiment 1: the preparation of biocatalyst

[0033](1) Inoculate Acinetobacter sp.Cxzy-L119 into LB medium and culture at 30°C for 1-2 days to obtain slant cells; the concentration of the slant medium consists of: NaCl 10g / L, yeast extract powder 5.0 g / L, peptone 10g / L, pH adjusted to 7.0, sterilized at 121°C for 20min;

[0034] (2) Inoculate the slant bacteria into the seed medium, cultivate at 30°C for 24h, and obtain the seed liquid; the concentration of the seed medium consists of: NaCl 0.5g / L, MgSO 4 ·7H 2 O 1.0g / L, K 2 HPO 4 1.0g / L, NH 4 NO 3 1.0g / L, peptone 10g / L, yeast extract powder 5.0g / L, olive oil 2.0g / L, sterilized at 121°C for 20min;

[0035] (3) Inoculate the seed solution into the fermentation medium with an inoculum volume concentration of 1% to 10%, cultivate it at 30°C and 180r / min for 48h, centrifuge the fermentation solution, discard the supernatant, and obtain wet bacteria ; The concentration of the fermentation medium consists of: NaCl ...

Embodiment 2

[0037] Embodiment 2: reaction solution monitoring method

[0038] The reaction solution was washed with 5% Na 2 CO 3 The solution was adjusted to a pH of about 7.5, extracted with ethyl acetate, and the upper organic phase was retained. Take an appropriate amount of the upper organic phase, dry it with nitrogen, add the mobile phase to dissolve, mix and filter with a 0.45 μm microporous membrane, and wait for sample injection. HPLC analysis condition: mobile phase: acetonitrile: water: trifluoroacetic acid=50: 50: 0.1; Flow rate: 1mL / min; Detection ultraviolet wavelength: 210nm; Column temperature: 30 ℃; Injection volume: 10 μ L; , 4.6×250nm).

[0039] (S,S)-lisinopril hydride enantiomeric excess e.e. and substrate conversion rate are calculated according to the following formula:

[0040] Formula 1:

[0041]

[0042] Formula 2:

[0043]

[0044] where [A] SS and [A] RS Respectively, liquid chromatography records the peak area of ​​substrate SS and RS type in the ...

Embodiment 3

[0045] Embodiment 3: lisinopril hydride small test catalytic process

[0046] 300mL catalytic reaction system: 50g / L concentration of lisinopril hydride in swirling; 30mL methanol as a co-solvent: 10g of the biocatalyst prepared in Example 1.

[0047] Add 15g of vortexed lisinopril hydride into a 500mL transformation bottle, add 30mL of methanol and start stirring to dissolve, then add 270mL of water, and use Na 2 CO 3 The solution (15%, V / V) adjusted the pH to around 7.2. The transformation temperature was 37° C., and 10 g of the immobilized cells prepared in Example 1 were added to start the transformation. use Na 2 CO 3 The solution (15%, V / V) maintained the pH at 7.0-7.3. Utilize HPLC to detect reaction transformation result, see table and accompanying drawing.

[0048] Table 1 Bioresolution of lisinopril hydride results

[0049] Catalytic time Conversion rate C enantiomeric excess e.e The main configuration of the remaining substrate 0h 0％ 33....

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Abstract

The invention belongs to the technical field of biocatalysis, and relates to a method for resolving Lisinopril hydride through biocatalysis. The method for resolving Lisinopril hydride through biocatalysis comprises the following steps: by taking Lisinopril hydride as a substrate raw material, adding a proper volume of water, generating selective esterolysis (R,S)- Lisinopril hydride under the condition of existence of lipase, and retaining (S,S)- Lisinopril hydride; after the reaction is completed, separating a mixed solution to obtain the (S,S)- Lisinopril hydride. The method has the advantages of being high in catalysis efficiency, good in stereoselectivity, easy to separate enzyme from substrate, capable of recycling enzyme, low in cost, small in environment pollution, and the like.

Description

technical field [0001] The invention belongs to the technical field of biocatalysis, and relates to a biocatalytic method for splitting lisinopril hydride. Background technique [0002] Lisinopril (Lisinopril), the chemical name is N-{N-[(S)-1-carboxy-3-phenylpropyl]-L-lysyl}-L-proline, is enalapril Lysine derivatives of La, a third-generation angiotensin-converting enzyme inhibitor (ACEI) developed and marketed by Merck in 1987, is mainly used for the treatment of hypertension, inhibiting ACE to reduce blood vessel The conversion of tensin I (AngI) to angiotensin II (AngII) leads to a decrease in the concentration of AngII, thereby reducing the pressor effect of AngII and the secretion of aldosterone. It also has the effects of reducing left ventricular mass index (LVMI), improving left ventricular function, reducing mortality in patients with acute myocardial infarction, and significantly reducing urinary albumin excretion rate. [0003] Lisinopril hydride is a key chira...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P41/00C12P13/02C12N1/20C12N9/20C12N11/14C12N11/08C12R1/01
CPCC12N9/20C12N11/08C12N11/14C12P13/02C12P41/001C12Y301/01003C12N1/205C12R2001/01Y02P20/50
Inventor 张利坤肖延铭钱敏帆杨卫华谈聪
Owner CHANGXING PHARMA
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