CN112831529A

CN112831529A - Preparation method of chiral 3-hydroxyvaleric acid and salts/esters thereof

Info

Publication number: CN112831529A
Application number: CN202110017324.6A
Authority: CN
Inventors: 刘明明; 李世燕
Original assignee: Nanjing Jinran Biological New Material Technology Co ltd
Current assignee: Nanjing Jinran Biological New Material Technology Co ltd
Priority date: 2021-01-07
Filing date: 2021-01-07
Publication date: 2021-05-25

Abstract

The invention relates to a preparation method of 3-hydroxyvaleric acid and salts/esters thereof, belonging to the technical field of organic chemical synthesis. The invention aims to solve the problem of environmental pollution in the prior art for preparing 3-hydroxyvaleric acid. The technical scheme of the invention provides an enzymatic method for preparing 3-hydroxyvaleric acid. The method has the advantages of low raw material cost, high product yield, simple process and no pollution to the environment.

Description

Preparation method of chiral 3-hydroxyvaleric acid and salts/esters thereof

Technical Field

The invention belongs to the field of organic chemical synthesis, and particularly relates to a preparation method of chiral 3-hydroxyvaleric acid and salts/esters thereof.

Background

Polyhydroxyalkanoate (PHA) is a carbon source and energy storage substance synthesized by bacteria under unbalanced growth conditions, and it has been found that up to 150 monomers of R-configuration 3-hydroxy fatty acids, 4-hydroxy fatty acids, 5-hydroxy fatty acids, etc. can be a component of PHA. 3-hydroxyvaleric acid is one of important monomers for synthesizing PHA, but the prior art has few methods for preparing 3-hydroxyvaleric acid, mainly based on 5 carbon compounds of petroleum sources, and the 3-hydroxyvaleric acid is prepared by performing a Baeyer-Villiger reaction on cyclopentanone, and the method generates a large amount of acidic waste water due to the use of a strong oxidant, thereby causing environmental pollution.

Disclosure of Invention

In order to solve the problems in the prior art, the invention provides a method for preparing 3-hydroxyvaleric acid and salts/esters thereof, which has the characteristics of low raw material cost, higher product yield, simple process and no pollution to the environment.

The preparation method of chiral 3-hydroxypentanoic acid provided by the invention comprises the following implementation steps:

a) adding compound V, reductase, hydrogen donor, Nicotinamide Adenine Dinucleotide Phosphate (NADP) into phosphate buffer solution, and heating at 20-50 deg.C for 10-30 hr;

b) after the reaction is finished, removing enzyme and other residues in the reaction system by filtration;

c) adding ethyl acetate with the same volume as the rest solution for extraction;

d) adding anhydrous sodium sulfate into the organic phase obtained after extraction to remove water;

e) removing the organic solvent by rotary evaporation to obtain a chiral compound III or IV;

f) dissolving the chiral compound III or IV prepared in the step e) in deionized water;

g) adding inorganic base or biological enzyme for hydrolysis reaction, hydrolyzing the compound III to obtain a chiral compound I, and hydrolyzing the compound IV to obtain a chiral compound II;

in the preceding implementation step, the concentration of compound V is between 10 and 200 mg/ml.

The reductase used in step a) mainly plays a catalytic role, and can be any one of ketoreductase, hydrogenase and N5, N10-methylene tetrahydrofolate reductase, preferably Ketoreductase (KRED), and the addition amount of the ketoreductase can be 5-30%.

The NADP addition in step a) may be between 0.01% and 5%.

The hydrogen donor in step a) can be glucose, the addition amount of the glucose is 1.0-2.0 equivalent, Glucose Dehydrogenase (GDH) needs to be added, the addition ratio is 0.1-5%, and the reaction is carried out in a salt buffer solution at 30-40 ℃ for 10-48h, and the reaction equation of the method is as follows:

the hydrogen donor in the step a) can also be isopropanol, the addition amount of the isopropanol is 10-50%, and the reaction is carried out for 30 hours at a constant temperature of 30 ℃, and the reaction equation of the method is as follows:

in step g), inorganic base can be selected for hydrolysis, and the inorganic base used for reaction can be sodium hydroxide, sodium bicarbonate, potassium hydroxide, potassium bicarbonate, calcium hydroxide, magnesium hydroxide and the like, preferably sodium hydroxide. The addition amount of the inorganic base is 1.0-3.0 equivalent, and the reaction can be quickly completed at 27 ℃.

In the step g), biological enzyme can be selected for hydrolysis, wherein the biological enzyme can be hydrolytic enzyme such as lipase, esterase, protease and the like for hydrolysis, the addition ratio is 1% -50%, and the reaction is carried out for 24 hours at the temperature of 30-50 ℃.

Adding the chiral compound I or II obtained in the step g) into a 500ml three-neck flask, then adding 200ml deionized water, stirring to dissolve, adding inorganic base, and reacting to obtain the 3-hydroxy valerate, wherein the inorganic base used for the reaction can be sodium hydroxide, sodium bicarbonate, potassium hydroxide, potassium bicarbonate, calcium hydroxide, magnesium hydroxide and the like, and for example, the chiral compounds I and II are respectively reacted with sodium hydroxide to obtain sodium salts such as VI and VII.

Dissolving the chiral compound I or II obtained in the step g) in an organic alcohol solvent, wherein the organic alcohol is preferably ethanol, reacting for 12-24 hours at the temperature of 20-80 ℃, and removing the organic alcohol solvent in a distillation mode after the reaction is finished to obtain the 3-hydroxyvalerate.

The 3-hydroxyvaleric acid and its salt/ester prepared by the process of the present invention may be used in the fields of food, medicine and nutritional supplement.

Compared with the prior art, the method has the beneficial effects that: the substrate raw material used by the method is easy to obtain, and compared with the prior art, the method has the characteristics of higher product yield, simple process and no pollution to the environment.

Detailed Description

The technical solutions in the embodiments of the present invention are clearly and completely described below, the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and all other embodiments obtained by a person of ordinary skill in the art without creative efforts based on the embodiments of the present invention belong to the protection scope of the present invention.

The embodiment of the invention relates to the addition amount, content and concentration of various substances, wherein the percentage content refers to the mass percentage content except for special description.

In the example of the present invention, if no specific description is made on the operation temperature, the temperature is usually room temperature and the temperature range is 15 to 30 ℃.

Example 1

100ml of 0.1M phosphate buffer was measured and added to a 250ml 4-necked reaction flask, and 30g of glucose and 100mg of NADP were added in this order, and the mixture was stirred at a rate of 100/min and slowly warmed to 30 ℃. Subsequently, 10g of Compound V, 100mg of GDH and 5g of ketoreductase were added and the reaction was carried out at 30 ℃ for 24 hours.

After the reaction is finished, enzyme and other residues in the system are removed through filtration, the residual solution is extracted by ethyl acetate with the same volume, and the extraction frequency is preferably three times for achieving a better extraction effect.

Anhydrous sodium sulfate was added to the extracted organic phase to remove water.

And (3) removing the organic solvent by water pump rotary evaporation at 50 ℃ to obtain a product IV, wherein the enantiomeric excess percentage of the product IV is 100%.

The following is the reaction equation for example 1:

example 2

90ml of 0.1M phosphate buffer was measured and added to a 250ml 4-necked reaction flask, and 100mg of NADP was added thereto, and the mixture was stirred at a rate of 100/min and slowly warmed to 30 ℃. 10g of the compound V is weighed and dissolved in 10ml of isopropanol, the dissolved solution is added into a reaction system, 5g of ketoreductase is added, and the reaction is carried out for 30 hours at the constant temperature of 30 ℃.

The following is the reaction equation for example 2:

example 3

100ml of 0.1M phosphate buffer was measured and added to a 250ml 4-necked reaction flask, and 30g of glucose and 100mg of NADP were added in this order, and the mixture was stirred at a rate of 100/min and slowly warmed to 35 ℃. 10g of Compound V, 200mg of GDH and 5g of ketoreductase were then added and the reaction was incubated at 35 ℃ for 24 hours.

And (3) removing the organic solvent by water pump rotary evaporation at 50 ℃ to obtain a product III, wherein the enantiomeric excess percentage of the product III is 98.6%.

The following is the reaction equation for example 3:

example 4

90ml of 0.1M phosphate buffer was measured and added to a 250ml 4-necked reaction flask, and 100mg of NADP was added thereto, and the mixture was stirred at a rate of 100/min and slowly warmed to 35 ℃. 10g of the compound V is weighed and dissolved in 10ml of isopropanol, the dissolved solution is added into a reaction system, 5g of ketoreductase is added, and the reaction is carried out for 30 hours at the constant temperature of 35 ℃.

And (3) removing the organic solvent by water pump rotary evaporation at 50 ℃ to obtain a product III, wherein the enantiomeric excess percentage of the product III is 98.9%.

The following is the reaction equation for example 4:

example 5

50ml of deionized water is weighed and added into a 100ml 4-mouth reaction bottle, stirring is carried out at the speed of 100rpm/min, 1.5g of sodium hydroxide solid is slowly added, stirring is carried out until all the sodium hydroxide solvent is dissolved, 5g of compound III is added, and reaction is carried out for 3h at the temperature of 27 ℃.

And after the reaction is finished, removing water by rotary evaporation, and drying to obtain a product I.

The following is the reaction equation for example 5:

example 6

50ml of deionized water was weighed into a 100ml 4-neck reaction flask, stirred at 100rpm/min, 5g of Compound IV was added, 1g of hydrolase was added, and the reaction was carried out at 27 ℃ for 15 hours.

And after the reaction is finished, filtering to remove hydrolase, removing water by rotary evaporation, and drying to obtain a product II.

The following is the reaction equation for example 6:

the chiral compounds I or II prepared in examples 5 and 6 are added into a 500ml three-neck flask, then 200ml deionized water is added, after stirring and dissolving, inorganic base is added, 3-hydroxy valerate can be obtained through reaction, the inorganic base used for reaction can be sodium hydroxide, sodium bicarbonate, potassium hydroxide, potassium bicarbonate, calcium hydroxide, magnesium hydroxide and the like, for example, the chiral compounds I and II are respectively reacted with sodium hydroxide, and sodium salts such as VI and VII can be obtained.

Dissolving the chiral compounds I or II prepared in examples 5 and 6 in an organic alcohol solvent, preferably ethanol, reacting at 20-80 ℃ for 12-24 hours, and removing the organic alcohol solvent by distillation after the reaction to obtain the 3-hydroxyvalerate.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.

Claims

1. A preparation method of chiral 3-hydroxy valeric acid comprises the following steps:

a) adding the compound V, reductase, hydrogen donor and nicotinamide adenine dinucleotide phosphate into phosphate buffer solution, and reacting for 10-30 hours under the heating condition of 20-50 ℃;

c) adding ethyl acetate with the same volume as the residual solution after filtration for extraction;

2. the method of claim 1, wherein: the reductase may be any one of ketoreductase, hydrogenase, and N5, N10-methylenetetrahydrofolate reductase.

3. The method of claim 2, wherein: the addition amount of the ketoreductase is 5-30%.

4. The method of claim 3, wherein: the hydrogen donor is glucose, and glucose dehydrogenase is also added in the reaction system.

5. The method of claim 3, wherein: the hydrogen donor is isopropanol.

6. A preparation method of chiral 3-hydroxy valerate is characterized in that: adding the compound I or II prepared by the preparation method of claims 1-5 into a container, adding a proper amount of deionized water, stirring for dissolving, adding an inorganic base, and reacting to obtain chiral 3-hydroxy valerate.

7. The method of claim 6, wherein: the inorganic base can be any one of sodium hydroxide, sodium bicarbonate, potassium hydroxide, potassium bicarbonate, calcium hydroxide and magnesium hydroxide.

8. A preparation method of chiral 3-hydroxy valerate is characterized by comprising the following steps: dissolving the chiral compound I or II prepared by the preparation method of claims 1-5 in an organic alcohol solvent, reacting at 20-80 ℃ for 12-24 hours, and removing the organic alcohol solvent by distillation after the reaction is finished to obtain the 3-hydroxyvalerate.

9. The method of claim 8, wherein: the organic alcohol is ethanol.

10. Use of chiral 3-hydroxyvaleric acid prepared by the process according to claims 1 to 5, chiral 3-hydroxyvaleric acid salts prepared by the process according to claims 6 to 7 and chiral 3-hydroxyvaleric acid esters prepared by the process according to claims 8 to 9 in the field of foodstuffs, pharmaceuticals and nutritional supplements.