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Method for extracting and purifying pepsin-soluble collagens of black sea cucumber in East China Sea

A technology of collagen and purification method, which is applied in the field of extraction and purification of enzymatically soluble collagen of Donghai black ginseng, which can solve problems such as pollution and inability to eat directly, and achieve the effect of high-yield extraction

Active Publication Date: 2013-07-03
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Other patents or documents related to sea cucumbers do not involve the extraction or production of collagen of the East China Sea black cucumber variety. Because the East China Sea continental shelf contains a huge amount of East China Sea black cucumber, the pollution of metal elements in the seawater makes it impossible to eat directly, and it is a low-value marine resource. , needs to be removed from the chemical elements before it can be used, so it is necessary to develop it into a high value-added collagen product through deep processing

Method used

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  • Method for extracting and purifying pepsin-soluble collagens of black sea cucumber in East China Sea
  • Method for extracting and purifying pepsin-soluble collagens of black sea cucumber in East China Sea
  • Method for extracting and purifying pepsin-soluble collagens of black sea cucumber in East China Sea

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] 1. Fresh Donghai black ginseng (commercially purchased, produced in Xiangshan, Zhejiang) samples were washed with tap water at 4°C to remove sediment and internal organs, cut into small pieces of 5mm*5mm, and 200g of block samples were washed with 1000mL0.2M EDTA (pH8 .0) Soak and stir for 5 days, replace the solution every day to remove calcium, iron, manganese, magnesium, lead and other pollution elements and residual impurities.

[0028] 2. After washing with water, add 100g of ice cubes (to reduce the temperature of the solution to below 4°C), homogenize with a stainless steel homogenizer (IKA T25Homogenizer), 18000rpm, 20min, and take the precipitate A after centrifugation.

[0029] 3. Add 0.1M Tris-HCl (pH 8.0) with 20 times volume (solution 20mL / 1g precipitate) to precipitate A, stir for 48 hours to fully dissolve tissue protein, and take precipitate B after centrifugation.

[0030] 4. Add 20 times the volume of 0.1M NaOH to the precipitate B and stir for 48 hour...

Embodiment 2

[0035] 1. Fresh black ginseng samples from the East China Sea were frozen and cut into granules (5mm*5mm in size), washed with ice water and stirred three times to remove impurities such as sediment and viscera, then took 200g block samples and washed them with 2000mL0.2M EDTA (pH8. 0) Soak and stir for 5 days, replace the solution every day to remove calcium, iron, manganese, magnesium, lead and other polluting elements.

[0036] 2. After washing with water, add 200g of ice cubes (to lower the temperature of the solution to below 4°C), use a stainless steel homogenizer (IKA T25Homogenizer) to homogenize at 18000rpm for 20min, and take the precipitate A after centrifugation.

[0037] 3. Add 20 times the volume of 0.1M, pH8.0 Tris-HCl+0.5M NaCl+50mM EDTA+0.2M β-mercaptoethanol to the precipitate A, stir and extract for 72 hours, and take the precipitate B after centrifugation.

[0038] 4. Add 20 times the volume of 0.1M NaOH to the precipitate B and stir for 72 hours. Change th...

Embodiment 3

[0043] 1. The tissue water content of fresh black ginseng is 83.61±0.2%, the ash content is 3.9±0.05%, the protein is 13.71±0.1%, and the fat is 0.9±0.05%. The content of heavy metal elements seriously exceeds the standard, as shown in the table below:

[0044]

[0045] It can be seen from the table that the contents of As, Pb and Cr far exceed the standard limit of 0.5mg / Kg.

[0046] 2. The epidermis of fresh Donghai black ginseng is seriously polluted, yellow in color, and partly has a serious rust red color. In 0.2M EDTA with pH 8.0, the solution is changed every day. After soaking for 5 days, the removal rate of Ca, Fe, Mn, Mg, Zn, Sn and other metal elements is above 90%, and the tissue is obviously softened and swollen, and it is easy to homogenize. Add ice water, homogenize at 18000rpm for 20min, centrifuge at 12000G and take precipitate A.

[0047] 3. Precipitate A was dissolved with 20 times the volume of 0.1M Tris-HCl (pH 8.0) and stirred for 72 hours. This step...

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Abstract

The invention provides a method for extracting and purifying pepsin-soluble collagens of black sea cucumbers in East China Sea. The method comprises the following steps: soaking a clean, massive black sea cucumber sample in East China Sea with an EDTA (Ethylene Diamine Tetraacetic Acid) solution; performing homogenized homogeneity; performing extraction in sequence by using a Tris-HCl solution, a NaOH solution and a pepsase acetic acid solution; and performing salting out and dialysis to obtain the pepsin-soluble collagens. According to the invention, the EDTA chelating agent is used to remove chemical element with high pollution concentration from sea cucumber tissues, and the removal efficiency can reach more than 90%, a crosslinking effect in chemical bonds and among the chemical bonds of a collagen albumen product can be effectively removed after pepsase is used for performing hydrolysis on collagen fibers, so that maximally extracting the solvation collagen albumen can be realized; after the purification steps of sodium chloride salting out and dialysis, and the like are carried out, and the purity of the obtained collagen albumen product by extraction can reach more than 90%, the yield can reach 56.99-61.52% (dry weight), which is 97.1-102.5% of the total collagen albumen content of the black sea cucumber in East China Sea, so that the extraction of collagen albumen content with high productivity of the black sea cucumber in East China Sea can be realized.

Description

(1) Technical field [0001] The invention relates to a method for extracting and purifying enzymatically soluble collagen from black ginseng. (2) Background technology [0002] Collagen is rich in amino acids such as glycine, alanine, arginine, and proline that the human body needs. Collagen is the most important part of the extracellular matrix. It is an extracellular protein. Collagen is a fibrous protein with three peptide chains twisted into a helical shape. The peptide chain is composed of a large number of repeated Gly-X-Y-sequences (X : proline, Y: hydroxyproline), 15-20 amino acid residues of the telopeptide (C- and N-terminal) do not participate in the triple helix structure, most of the residues are lysine and hydroxylysine It is composed of its aldehyde derivatives, forming covalent crosslinks between molecules and within minutes, and the triple helix structure is maintained by hydrogen bonds within and between peptide chains. [0003] Collagen is the most abunda...

Claims

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Application Information

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IPC IPC(8): C12P21/06C07K1/14
Inventor 郑裕国林赛君薛亚平单恩莉沈寅初
Owner ZHEJIANG UNIV OF TECH
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