CN106947795B

CN106947795B - Method for synthesizing 2-thiophene ethylamine through biocatalysis

Info

Publication number: CN106947795B
Application number: CN201710336249.3A
Authority: CN
Inventors: 秦育红; 邢俊德; 范晓军; 朱国峰; 孙权威; 冯敏敏
Original assignee: Taiyuan University of Technology
Current assignee: SHANXI YITONG ENVIRONMENTAL ENERGY TECHNOLOGY GROUP CO LTD
Priority date: 2017-05-13
Filing date: 2017-05-13
Publication date: 2020-05-22
Anticipated expiration: 2037-05-13
Also published as: CN106947795A

Abstract

一种生物催化合成2‑噻吩乙胺的方法，所述方法是将葡萄糖、磷酸氢二钠、磷酸二氢钾按一定比例配成水溶液，灭菌冷却得地衣芽孢杆菌培养基；再按照一定的菌体浓度，在摇床中恒温下活化；然后将2‑硝基乙烯基噻吩溶解，加入到活化培养基中进行生物催化反应，后经离心分离，上清液用乙酸乙酯萃取、洗涤、干燥、减压蒸馏得产物2‑噻吩乙胺。本方法具有工艺简单、合成收率高、成本低廉以及环境污染少的显著特点和进步。A method for biocatalytically synthesizing 2-thiopheneethylamine, the method comprises preparing an aqueous solution of glucose, disodium hydrogen phosphate and potassium dihydrogen phosphate in a certain proportion, and sterilizing and cooling to obtain a Bacillus licheniformis culture medium; The bacterial concentration was activated at a constant temperature in a shaker; then 2-nitrovinylthiophene was dissolved, added to the activation medium for biocatalytic reaction, and then centrifuged, and the supernatant was extracted with ethyl acetate, washed, The product 2-thiopheneethylamine is obtained by drying and distillation under reduced pressure. The method has the remarkable characteristics and progress of simple process, high synthesis yield, low cost and less environmental pollution.

Description

Method for synthesizing 2-thiophene ethylamine through biocatalysis

Technical Field

The invention relates to a method for synthesizing 2-thiophene ethylamine, in particular to a method for synthesizing 2-thiophene ethylamine by biologically catalyzing and reducing 2-nitrovinyl thiophene with bacillus licheniformis, belonging to the technical field of chemical industry and medicine.

Background

The 2-thiophene ethylamine is an important intermediate for synthesizing ticlopidine hydrochloride which is a medicament for treating cardiovascular and cerebrovascular diseases, and is also an intermediate of other biological active medicaments such as a hypolipidemic medicament, a platelet aggregation inhibitor, a cardiovascular vasodilator, a 5-lipoxygenase inhibitor, a plurality of antibacterial agents and the like. The most common synthetic method reported in the literature at present is to prepare 2-thiophenecarboxaldehyde by taking thiophene as a raw material, then react with nitromethane to obtain 2-nitrovinyl thiophene, and finally reduce to obtain 2-thiopheneethylamine. In the designed synthetic route, the double bond and the nitro group are reduced in the last step of reduction reaction, and the molecules contain sulfur, so that the synthetic step has high cost and harsh reaction conditions. The main reduction methods at present are the following three: (1) reducing by using sodium borohydride as a reducing agent: the yield of the method is 75%, but the consumption of reducing agents and solvents is large, and the cost is high; (2) reducing by using lithium aluminum hydride as a reducing agent: the reducing agent is easy to cause explosion, is expensive and is generally only used in laboratories; (3) reducing by using hydrogen as a reducing agent: the method needs to carry out nitro reduction and double bond reduction step by step, the activity of the catalyst is inhibited by the thiophene sulfur, the yield is low, the process needs to be carried out at high temperature and high pressure, and the safety risk is high.

The methods do not accord with three major trends of high-efficiency reducibility, atom economy and environmental protection of organic synthesis. The invention adopts a biological catalytic reduction method, and the method has the characteristics of high efficiency, strong selectivity, low cost, simple device, less environmental pollution and the like, and has remarkable advantages.

Disclosure of Invention

The invention aims to provide a synthesis method with mild reaction conditions, simple operation, high yield and low cost aiming at the defect of high cost of the existing chemical synthesis technology, in particular to a synthesis method for reducing 2-nitrovinyl thiophene into 2-thiophene ethylamine by using microorganisms with almost no environmental pollution.

The above object of the present invention is achieved by the following means.

A method for biocatalytic synthesis of 2-thienylethylamine, the method comprising the steps of:

1) preparation of culture medium for bacillus licheniformis

Adding water, glucose, disodium hydrogen phosphate and potassium dihydrogen phosphate into a vessel according to the mass ratio of 1000: 30: 7: 6 to prepare an aqueous solution, adjusting the pH to be =8, sterilizing for 30 min, and cooling to obtain a bacillus licheniformis culture medium;

2) activation of Bacillus licheniformis

Adding the bacillus licheniformis according to the mass ratio of the solution of the bacillus licheniformis culture medium to the bacillus licheniformis of 10: 1, and activating for 1 hour at the constant temperature of 37 ℃ and the rotation speed of 180 r/min in a shaking table;

3) synthesis of 2-thiopheneethylamine

Dissolving 2-nitrovinyl thiophene in tetrahydrofuran, adding 5 g/L of reactant into an activated culture medium, carrying out biocatalytic reaction for 96 hours in a shaking table at 37 ℃ and the rotation speed of 180 r/min, keeping the pH =8 during the reaction, centrifugally separating reaction liquid after the reaction, extracting supernatant with ethyl acetate, washing, drying, and distilling under reduced pressure to obtain the product 2-thiophene ethylamine.

Wherein, the concentration of the bacillus licheniformis liquid is as follows: 5.0*10⁹cfu/mL。

Compared with the existing chemical method for synthesizing 2-thiophene ethylamine, the method for synthesizing 2-thiophene ethylamine by biological catalysis has the following three substantial characteristics and remarkable progress.

Firstly, the method adopts the bacillus licheniformis to reduce double bonds and nitro groups in the 2-nitroethylene thiophene in one step, thereby greatly reducing the cost, and having mild reaction conditions and simple and convenient operation.

Secondly, the synthetic route of the method is environment-friendly and pollution-free. The reduction preparation of the 2-thiophene ethylamine by a chemical method can cause a certain degree of environmental pollution, and the bacillus licheniformis is nontoxic and harmless and has high yield.

Thirdly, the used bacillus licheniformis is activated and then reused, and the production cost is further reduced.

Detailed Description

In order to clearly illustrate the embodiments of the present invention, the following further describes the embodiments of the present invention.

Example 1

Dissolving 3 g of glucose, 0.7 g of disodium hydrogen phosphate and 0.6 g of potassium dihydrogen phosphate in 100 mL of water, adjusting the pH to be =8, sterilizing for 30 min, and cooling to obtain a bacillus licheniformis culture medium;

10 g of Bacillus licheniformis was added to the culture medium and activated for 1 hour in a shaker at a constant temperature of 37 ℃ at a rotation speed of 180 rpm.

Dissolving 0.5g of 2-nitrovinyl thiophene in 5 mL of tetrahydrofuran, adding the solution into an activated culture medium solution, carrying out biocatalytic reaction for 96 hours in a shaking table at 37 ℃ and a rotation speed of 180 revolutions per minute, keeping pH =8 during the reaction, carrying out centrifugal separation on a reaction solution after the reaction, extracting a supernatant with an ethyl acetate solvent, washing with saturated saline, drying with anhydrous sodium sulfate, and carrying out reduced pressure distillation to obtain a product 2-thiophene ethylamine with the yield of 75.5%.

Example 2

Activation and reutilization of bacillus licheniformis

15 g of Bacillus licheniformis obtained by centrifugal separation in example 1 was added into the culture medium, and a solution consisting of 0.5g of 2-nitrovinylthiophene and 5 mL of tetrahydrofuran was added, biocatalytic reaction was carried out in a shaker at 37 ℃ and a rotation speed of 180 rpm for 96 hours, the reaction was carried out while maintaining pH =8, after the reaction, the reaction solution was centrifugally separated, the supernatant was extracted with an ethyl acetate solvent, washed with saturated saline, dried with anhydrous sodium sulfate, and distilled under reduced pressure to obtain 2-thienylethylamine with a yield of 73.0%.

Comparative example 1

This example differs from example 1 in that yeast was added to the medium solution instead of Bacillus licheniformis, and the other experimental steps and reaction conditions were the same as in example 1, yielding a yield of 65.8% 2-thienylethylamine. The yield was lower than in example 1.

Comparative example 2

This example differs from example 1 in that the pH of the culture broth was adjusted by dropping 10% NaOH solution or 2 mol/L HCl solution during the media preparation and biocatalysis to pH =4, pH =5, pH =6, pH =7 and pH =9, respectively, and other experimental steps and reaction conditions were the same as example 1, yielding yields of 2-thiopheneethylamine of 25.8%, 34.5%, 58.9%, 72.3% and 53.2%, respectively. The yields are all lower than in example 1.

Comparative example 3

This example is different from example 1 in that the reaction temperature was changed to 30 ℃, 32 ℃, 35 ℃, 40 ℃ and 42 ℃ during the biocatalytic reaction, and other experimental steps and reaction conditions were the same as example 1, to obtain yields of 2-thienylethylamine of 53.2%, 58.3%, 65.9%, 67.8% and 40.2%, respectively. The yields are all lower than in example 1.

Comparative example 4

This example is different from example 1 in that the reaction time was changed to 24 hours, 48 hours, 72 hours, 120 hours, 144 hours and 168 hours during the biocatalytic reaction, and the other experimental procedures and reaction conditions were the same as example 1, to obtain yields of 2-thienylethylamine of 20.7%, 45.9%, 65.8%, 71.3%, 66.6% and 52.0%, respectively. The yields are all lower than in example 1.

Comparative example 5

This example differs from example 1 in that the starting 2-nitrovinylthiophene, having different masses of 0.2 g, 0.4 g, 0.6 g, 0.8 g and 1.0 g, respectively, was dissolved in 5 mL of tetrahydrofuran. The other experimental procedures and reaction conditions were the same as in example 1, and the yields of 2-thienylethylamine were 45.9%, 74.3%, 66.5%, 52.0% and 35.7%, respectively. The yields are all lower than in example 1.

Comparative example 6

This example differs from example 1 in that different masses of Bacillus licheniformis were added to the medium solution at 5g, 8 g, 12g, 15 g and 20 g, respectively, and activated in a shaker at a rotation speed of 180 rpm at a constant temperature of 37 ℃ for 1 hour to obtain an activated Bacillus licheniformis culture solution. The other experimental procedures and reaction conditions were the same as in example 1, and the yields of 2-thienylethylamine were 45.5%, 68.3%, 74.6%, 62.0% and 55.7%, respectively. The yields are all lower than in example 1.

Comparative example 7

This example differs from example 1 in that 2-nitrovinylthiophene was dissolved in different solvents, methanol, ethanol and dioxane, respectively. The other experimental procedures and reaction conditions were the same as in example 1, and the yields of 2-thienylethylamine were 56.2%, 65.4% and 68.1%, respectively. The yields are all lower than in example 1.

From example 1 to comparative example 7, it can be seen that the reaction process of the present invention is the optimum process for preparing 2-thienylethylamine by 2-nitrovinylthiophene biocatalytic reaction, and changes in reaction conditions all result in a decrease in product yield.

It should be understood that the above-mentioned embodiments of the present invention are only examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention, and it will be obvious to those skilled in the art that other variations or modifications may be made on the basis of the above description, and all embodiments may not be exhaustive, and all obvious variations or modifications may be included within the scope of the present invention.

Claims

1. a method for biocatalytic synthesis of 2-thiopheneethylamine, the method is carried out according to the following steps:

1) Preparation of Bacillus licheniformis medium

Water, glucose, disodium hydrogen phosphate and potassium dihydrogen phosphate are added to the vessel in a mass ratio of 1000:30:7:6 to prepare an aqueous solution, adjust pH=8, sterilize for 30 min, and cool to obtain Bacillus licheniformis culture medium;

2) Activation of Bacillus licheniformis

According to the mass ratio of the solution of Bacillus licheniformis medium: Bacillus licheniformis is 10:1, add Bacillus licheniformis, in a shaker at a constant temperature of 37 ° C, and activate for 1 hour at a rotating speed of 180 rev/min;

3) Synthesis of 2-thiopheneethylamine

Dissolve 2-nitrovinylthiophene in tetrahydrofuran, add 5 g/L of reactant to the activated culture medium, and conduct biocatalytic reaction at 37°C with a rotating speed of 180 rpm for 96 hours in a shaker. During the period, pH=8 was maintained, the reaction solution was centrifuged after the reaction, and the supernatant was extracted with ethyl acetate, washed, dried, and distilled under reduced pressure to obtain the product 2-thiopheneethylamine.