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A method for extracting chymotrypsinogen and trypsinogen by fractional precipitation

A technology of chymotrypsinogen and trypsinogen, which is applied in the direction of biochemical equipment and methods, enzymes, peptidases, etc., can solve the problems of low activity and low yield, and achieve the effects of improving economic benefits, improving efficiency and saving costs

Active Publication Date: 2016-08-03
QINGDAO KANGYUAN PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 1. The present invention provides a method for extracting chymotrypsinogen and trypsinogen by fractional precipitation, in order to solve the shortcomings of low yield and low activity of chymotrypsinogen extraction process, improve the process of chymotrypsinogen extraction, thereby increasing the yield , to reduce the loss of protein activity, especially the use of affinity chromatography to obtain the by-product trypsinogen

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] 1) Raw material pretreatment

[0032] After washing the pancreas, remove fat, connective tissue and other debris, immediately immerse it in a pre-frozen sulfuric acid solution, cool it suddenly, and store it at about zero degrees.

[0033] 2) Protein maceration and extraction

[0034] 2.1) Put the pretreated pancreas into a meat grinder and mince it into pancreas pulp, weigh it to 100kg, put it into a reaction tank, and add 200L of ice-cold sulfuric acid. Put it in a cold room, stir once every 1-2 hours, soak and extract for 24 hours.

[0035] 2.2) The impregnation is filtered with a coarse filter bag (or two layers of gauze), and after drying, 86 kg of filter residue is obtained. Repeat the above process with 86L of ice-cold sulfuric acid, then filter, discard the filter residue, combine the filtrates from two times to a total of 270L, add 65.34kg of solid ammonium sulfate to make the concentration of ammonium sulfate reach 40% saturation, and place it in a cold room...

Embodiment 2

[0049] 1) Raw material pretreatment

[0050] After washing the pancreas, remove fat, connective tissue and other debris, immediately immerse it in a pre-frozen sulfuric acid solution, cool it suddenly, and store it at about zero degrees.

[0051] 2) Protein maceration and extraction

[0052] 2.1) Put the pretreated pancreas into a meat grinder and mince it into pancreas pulp, weigh it to 100kg, put it into a reaction tank, and add 200L of ice-cold sulfuric acid. Put it in a cold room, stir once every 1-2 hours, soak and extract for 24 hours.

[0053] 2.2) The impregnation is filtered with a coarse filter bag (or two layers of gauze), and after drying, 88 kg of filter residue is obtained. Repeat the above process with 88L of ice-cold sulfuric acid, then filter, discard the filter residue, combine the filtrates from two times to a total of 275L, add 66.55kg of solid ammonium sulfate to make the concentration of ammonium sulfate reach 40% saturation, and place it in a cold room...

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Abstract

The invention discloses a method for extracting chymotrypsinogen and trypsinogen through fractional precipitation. The principle is that high-concentration saline ions can compete with protein for water molecules in a protein solution so as to damage a hydrated sheath on the surface of the protein and reduce the solubility, and accordingly, the protein can be precipitated from the solution. Ammonium sulfate is most widely used due to large solubility, small temperature coefficient and low probability of protein denaturation. Therefore, the invention adopts an ammonium sulfate fractional precipitation method to research and improve the technology for extracting the chymotrypsinogen and the trypsinogen, the yield of the prepared chymotrypsinogen is improved to 0.5%-0.6% especially by adopting affinity chromatography, and the trypsinogen as a by-product is obtained simultaneously.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for obtaining trypsinogen while extracting chymotrypsinogen by using ammonium sulfate fractional precipitation. Background technique [0002] Chymotrypsinogen is the precursor of chymotrypsin, which belongs to serine protease with a molecular weight of 25000 Daltons. Chymotrypsin, also known as chymotrypsin, has a molecular weight of 42,000 Daltons. It is used for the enzymatic hydrolysis of peptide chains. It specifically hydrolyzes peptide bonds. It belongs to endopeptidases and has a high hydrolysis yield, but its specificity is lower than that of trypsin. . Its solid state is relatively stable, showing white rod-like crystals; while its aqueous solution is extremely unstable, when the pH is 5-8, its activity is the strongest, and it is also most likely to lose its activity. [0003] Trypsinogen is the precursor of trypsin, synthesized in the pancreas, present in the pa...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/76
CPCC12N9/6427C12Y304/21001C12Y304/21004
Inventor 刘乃山林晓磊孙延年葛翠凤
Owner QINGDAO KANGYUAN PHARMA
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