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Method utilizing microsomal enzyme of animal liver to biosynthesize 7-dehydrocholesterol

A technology for dehydrocholesterol and biosynthesis, which is applied in the fields of biochemical engineering and enzyme engineering, can solve the problems of low efficiency, difficult control of conditions, affecting the production cost of vitamin D, and marketing promotion of sales price, so as to reduce environmental pollution and reduce production costs. Effect

Inactive Publication Date: 2009-11-11
GUANGXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, no matter what kind of chemical synthesis method is used to prepare the raw material 7-dehydrocholesterol, there are disadvantages such as long reaction steps, low overall yield, many side reactions, difficult conditions to control, low efficiency, and high cost, which will affect vitamin D. 3 production cost, sales price, marketing

Method used

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  • Method utilizing microsomal enzyme of animal liver to biosynthesize 7-dehydrocholesterol
  • Method utilizing microsomal enzyme of animal liver to biosynthesize 7-dehydrocholesterol
  • Method utilizing microsomal enzyme of animal liver to biosynthesize 7-dehydrocholesterol

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Experimental program
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Effect test

Embodiment 1

[0025] (1) Preparation of microsomal enzyme solution

[0026] Put 50g of fresh pig liver cut into pieces into pH 5, 0.01mol / L glutathione phosphate buffer solution, homogenize with a tissue homogenizer at 4°C, filter with two layers of gauze, and place the filtrate in In the ultrasonic cell disruptor, the processing conditions are: power 400W, temperature 4°C, time 5min. Then centrifuge (4°C, 12000r min -1 , 10 min) separation, and the supernatant was taken to obtain the microsomal enzyme solution.

[0027] (2) Enzymatic conversion reaction

[0028] Add 20ml 1ug / ml squalene, 2ml 2mmol / L reduced coenzyme II, 1ml 5mmol / L adenosine triphosphate, 2ml 20mmol / L nicotinamide to the obtained 20ml microsomal enzyme solution, and react for 5 hours to obtain an enzymatic conversion reaction solution .

[0029] (3) Saponification reaction

[0030] The enzymatic conversion reaction solution was added to 5 ml of 15% KOH-ethanol solution, and the reaction solution was shaken at 30° C. f...

Embodiment 2

[0034] (1) Preparation of microsomal enzyme solution

[0035] Put 50 g of fresh pig liver cut into pieces into pH 6.5, 0.05 mol / L glutathione phosphate buffer solution, homogenize it with a tissue homogenizer at 4°C, filter it with two layers of gauze, and place the filtrate in In the ultrasonic cell disruptor, the processing conditions are: power 150W, temperature 35°C, time 40min. Then centrifuge (4°C, 12000r min -1 , 10 min) separation, and the supernatant was taken to obtain the microsomal enzyme solution.

[0036] (2) Enzymatic conversion reaction

[0037] Add 20ml of 40ug / ml squalene, 2ml of 1mmol / L reduced coenzyme II, 1ml of 2.5mmol / L adenosine triphosphate, and 2ml of 30mmol / L nicotinamide to the obtained 20ml microsomal enzyme solution, and react for 5 hours to obtain an enzymatic conversion reaction liquid.

[0038] (3) Saponification reaction

[0039] The enzymatic conversion reaction solution was added to 5 ml of 15% KOH-ethanol solution, and the reaction sol...

Embodiment 3

[0043] (1) Preparation of microsomal enzyme solution

[0044] Put 50 g of fresh sheep liver cut into pieces into pH 9, 0.05 mol / L glutathione phosphate buffer solution, homogenize it with a tissue homogenizer at 4°C, filter it with two layers of gauze, and place the filtrate in In the ultrasonic cell disruptor, the processing conditions are: power 300W, temperature 4°C, time 20min. Then centrifuge (4°C, 12000r min -1 , 10 min) separation, and the supernatant was taken to obtain the microsomal enzyme solution.

[0045] (2) Enzymatic conversion reaction

[0046] Add 20ml 40ug / ml squalene, 2ml 2mmol / L reduced coenzyme II, 1ml 5mmol / L adenosine triphosphate, 2ml 40mmol / L nicotinamide, 1ml 1mmol / L MgCL to the obtained 20ml microsomal enzyme solution 2 , and reacted for 5h to obtain an enzymatic conversion reaction liquid.

[0047] (3) Saponification reaction

[0048] The enzymatic conversion reaction liquid was added to 5 ml of 15% KOH-ethanol solution, and the reaction was sh...

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Abstract

The invention belongs to the technical field of biochemical engineering and enzyme engineering, and in particular relates to a method utilizing the microsomal enzyme of an animal liver to biosynthesize 7-dehydrocholesterol. The method includes the following steps: (1) preparation of microsomal enzyme solution; (2) enzymatic conversion reaction; (3) saponification reaction; and (4) extraction. The method can prepare the 7-dehydrocholesterol which is completely identical with that prepared by chemosynthesis; and meanwhile, the technology is simple and easy, and not only the production cost of vitamin D3 can be reduced, but also the environment pollution can be reduced.

Description

technical field [0001] The invention belongs to the technical fields of biochemical engineering and enzyme engineering. It specifically relates to a method for synthesizing 7-dehydrocholesterol by using animal liver microsomal enzymes Background technique [0002] Vitamin D 3 It is an essential fat-soluble vitamin for the growth, development, reproduction, maintenance of life and health of humans and animals. Its main function is to regulate calcium and phosphorus metabolism, promote intestinal calcium and phosphorus absorption and bone calcification, maintain the balance of blood calcium and blood phosphorus, and can be used as pharmaceutical preparations, food and feed additives, etc., and can also be used clinically to treat rickets , senile osteoporosis, hypothyroidism, etc. current vitamin D 3 It is produced by photochemical method, that is, 7-dehydrocholesterol (7-DHC) is used as raw material, which is converted into vitamin D after ultraviolet irradiation 3 Precu...

Claims

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Application Information

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IPC IPC(8): C12P33/00C12P33/02
Inventor 黄时海李湘萍夏小斌吴孔阳陈桂光梁智群张云开李灿明汪晟康超何鑫平
Owner GUANGXI UNIV
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