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Method for extracting chorionic gonadotrophin crude product from large-volume pregnant woman urine on basis of ultrafiltration concentration

A technology for pregnant women's urine and chorionic gonadotropin, which is applied in the field of biotechnology to extract natural medicines, can solve the problems of wasting hydrochloric acid, unfavorable HCG purification operation, and low efficiency of target protein precipitation, so as to improve the activity yield, reduce production costs, Effect of increasing protein concentration

Inactive Publication Date: 2015-11-25
江西浩然生物制药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disadvantage of this method is that the original urine volume is too large, and the adsorption rate of sodium benzoate is low. To ensure sufficient adsorption, excess sodium benzoate (generally about 2-3% of the original urine amount) is often added, and the reaction time is long. Stirring and adsorption are required for at least 2 hours, HCG is prone to degradation during this process; the original urine contains a lot of water, and a large amount of hydrochloric acid needs to be wasted when adjusting the pH
[0007] The method disclosed in CN103193880A adopts the ultrafiltration concentration method to concentrate the urine to 4-5% of the original weight, and directly precipitates with acetone after adding a small amount of anhydrous calcium chloride, the precipitation efficiency of the target protein is low, and the product yield is only 40% In addition, because there are a lot of impurity proteins in urine, this method is easy to precipitate various impurity proteins in urine, and the obtained product has low unit activity, which is not conducive to the subsequent HCG purification operation

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  • Method for extracting chorionic gonadotrophin crude product from large-volume pregnant woman urine on basis of ultrafiltration concentration

Examples

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Effect test

Embodiment 1

[0016] Example 1 A method for extracting crude chorionic gonadotropin from the urine of large-volume pregnant women based on ultrafiltration concentration, the steps are as follows:

[0017] Collect 1000kg of urine from healthy pregnant women, cool it to a temperature of 2-8°C, and concentrate it with an ultrafiltration membrane with a molecular weight cut-off of 10kDa to 100kg of urine concentrate; add 6kg of sodium benzoate to it, and stir for 15 minutes until the sodium benzoate is completely dissolved; Add hydrochloric acid to adjust the pH to 3.9, and continue stirring for 30 minutes at a temperature of 2-8°C. A large amount of white insoluble matter is precipitated, and the solution becomes a white suspension; pour the suspension into a filter cloth bag, filter to collect the white insoluble matter, and shake Dry; Stir and add edible alcohol (v / w95%) pre-cooled at 2-8°C to the insoluble matter until the final concentration of alcohol is 80%, a large amount of white floccu...

Embodiment 2

[0018] Example 2 A method for extracting crude chorionic gonadotropin from the urine of large-volume pregnant women based on ultrafiltration concentration, the steps are as follows:

[0019] Collect 1200kg of urine from healthy pregnant women, cool it to a temperature of 2-8°C and concentrate it with an ultrafiltration membrane with a molecular weight cut-off of 10kDa to 120kg of urine concentrate; add 8.4kg of sodium benzoate to it, and stir for 15 minutes until the sodium benzoate is completely dissolved ; Add hydrochloric acid to adjust the pH to 3.9, and continue to stir for 45 minutes at a temperature of 2-8°C, a large amount of white insoluble matter is precipitated, and the solution becomes a white suspension; pour the suspension into a filter cloth bag, filter and collect the white insoluble matter, Shake dry; Stir and add 2-8°C pre-cooled edible alcohol (v / w95%) to the insoluble matter until the final concentration of alcohol is 80%, a large amount of white flocculent ...

Embodiment 3

[0020] Example 3 A method for extracting crude chorionic gonadotropin from the urine of large-volume pregnant women based on ultrafiltration concentration, the steps are as follows:

[0021] Collect 1500kg of urine from healthy pregnant women, cool it to a temperature of 2-8°C, and concentrate it with an ultrafiltration membrane with a molecular weight cut-off of 10kDa to 150kg of urine concentrate; add 12kg of sodium benzoate to it, and stir for 15 minutes until the sodium benzoate is completely dissolved; Add hydrochloric acid to adjust the pH to 3.9, continue stirring at 2-8°C for 60 minutes, a large amount of white insoluble matter is precipitated, and the solution becomes a white suspension; pour the suspension into a filter cloth bag, filter to collect the white insoluble matter, shake Dry; Stir and add edible alcohol (v / w95%) pre-cooled at 2-8°C to the insoluble matter until the final concentration of alcohol is 80%, a large amount of white flocculent precipitates precip...

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Abstract

The invention relates to a method for extracting a chorionic gonadotrophin crude product from large-volume pregnant woman urine on the basis of ultrafiltration concentration, which comprises the following steps: a. collecting fresh pregnant woman urine, and carrying out ultrafiltration concentration; b. adding sodium benzoate into the urine concentrated solution, and dissolving by stirring; c. acidifying with hydrochloric acid to precipitate abundant white insoluble substances, filtering to collect the insoluble substances, and carrying out centrifugal drying; d. adding precooled alcohol into the insoluble substances while stirring to precipitate abundant flocculent precipitate, and standing; and e. siphoning the solution supernate, carrying out vacuum filtration, collecting the precipitate, dewatering and drying. Compared with the traditional crude product preparation process, the method for producing the human urine chorionic gonadotrophin crude product is implemented by carrying out sodium benzoate adsorption after the ultrafiltration concentration, thereby reducing the consumption of the sodium benzoate, hydrochloric acid and ethanol and lowering the production cost; and the increase of the HCG (human chorionic gonadotrophin) concentration in the concentrated urine is beneficial to the adsorptivity of the sodium benzoate for the HCG, thereby enhancing the adsorption efficiency and enhancing the activity yield to 80%.

Description

technical field [0001] The invention belongs to the extraction of natural medicines by biotechnology, in particular to a method for extracting crude chorionic gonadotropin from the urine of large-volume pregnant women based on ultrafiltration and concentration. Background technique [0002] Chorionic Gonadotrophin (Chorionic Gonadotrophin), also known as Human Chorionic Gonadotrophin (HCG for short), was discovered by Aschhein and Zondek in the urine of pregnant women in 1927. Chorionic Gonadotropin is a sugar secreted by the trophoblast cells of the placenta Protein, mainly found in the urine and blood of pregnant women. [0003] Chorionic gonadotropin is a hormone composed of glycoproteins of α and β dimers. Its α subunit and β subunit are bound together by non-covalent hydrophobic and ionic interactions. The molecular weight of choriogonadotropin is about It is 32KD, 25%-41% of its molecular weight comes from sugar side chains (25%-30% for conventional chorionic gonadotr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/59C07K1/36C07K1/34C07K1/30
CPCC07K14/59
Inventor 万偲魏超娟黄朝晖张素芳徐富勇
Owner 江西浩然生物制药有限公司
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