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Cattle-sourced hyaluronidase affinity medium and adsorption method thereof

A technology of hyaluronidase and hyaluronic acid, applied in chemical instruments and methods, separation methods, solid adsorbent liquid separation, etc., can solve problems such as inability to carry out industrial production, low recovery rate, and complicated operation, and achieve large-scale industrial production Application potential, great economic benefit, effect of increased purification efficiency

Active Publication Date: 2018-08-03
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The defects of the above studies are that the recovery rate is too low, the cost is high, the operation is complicated, and industrial production cannot be carried out.

Method used

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  • Cattle-sourced hyaluronidase affinity medium and adsorption method thereof
  • Cattle-sourced hyaluronidase affinity medium and adsorption method thereof
  • Cattle-sourced hyaluronidase affinity medium and adsorption method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: Synthesis and application of Sepharose-6-Chloromethyluracil-ECH

[0030] Sepharose CL 4B (100g) was washed with 10 times the volume of deionized water, and dried into a wet cake; suspended in 50ml activation buffer (0.8M NaOH, 20% DMSO, 10% Epichlorohydrin, 0.5mg / ml sodium borohydride) Shake at 40°C for 2.5 hours, then pour into a glass frosted funnel, and wash with 10 times the volume of distilled water each time under suction filtration until the pH of the washing solution is neutral, and then drained to form a wet cake. Suspend the activated Sepharose CL 4B medium in 500ml 0.1M NaOH solution, add 20mL ammonia water, keep the gel at 30°C for 12h under stirring (200rpm), and wash it with deionized water. Weigh 2g of 6-Chloromethyluracil, mix with 20g of the washed medium at 60°C for 12h, maintain pH 8.0, and wash with deionized water. The medium Sepharose-6-Chloromethyluracil-ECH is obtained, the structure of which is shown in the general formula (1), re...

Embodiment 2

[0039] Example 2: Synthesis and application of Sepharose-6-Chloromethyluracil-1,3-propanediamine-ECH

[0040]Sepharose CL 4B (100g) was washed with 10 times the volume of deionized water, and dried into a wet cake; suspended in 50ml activation buffer (0.8M NaOH, 20% DMSO, 10% Epichlorohydrin, 0.5mg / ml sodium borohydride) Shake at 40°C for 2.5 hours, then pour into a glass frosted funnel, and wash with 10 times the volume of distilled water each time under suction filtration until the pH of the washing solution is neutral, and then drained to form a wet cake. Suspend the activated Sepharose CL 4B medium in 500ml 0.1M NaOH solution, add 20ml 1,3-propanediamine, keep the gel at 30°C for 12h under stirring (200rpm), and wash it with deionized water. Weigh 2g of 6-Chloromethyluracil, mix with 20g of the washed medium at 60°C for 12h, maintain pH 8.0, and wash with deionized water.

[0041] The medium Sepharose-6-Chloromethyluracil-1,3-propanediamine-ECH is obtained, the structure ...

Embodiment 3

[0048] Embodiment 3: Synthesis and application of Sepharose-2-Aminobenzonzimidazole-ECH

[0049] Sepharose CL 4B (100g) was washed with 10 times the volume of deionized water, and dried into a wet cake; suspended in 50ml activation buffer (0.8M NaOH, 20% DMSO, 10% Epichlorohydrin, 0.5mg / ml sodium borohydride) Shake at 40°C for 2.5 hours, then pour into a glass frosted funnel, and wash with 10 times the volume of distilled water each time under suction filtration until the pH of the washing solution is neutral, and then drained to form a wet cake. Suspend 20g of activated Sepharose CL 4B medium in 50ml of 0.1M NaOH solution, weigh and add 2g of 2-Aminobenzimidazole, keep the temperature at 60°C for 12h, and wash with deionized water.

[0050] The medium Sepharose-6-Chloromethyluracil-ECH was obtained, the structure of which was shown in the general formula (3), referred to as structure 3.

[0051] (1) Determination of the maximum adsorption capacity of the medium

[0052] Det...

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Abstract

The invention provides a cattle-sourced hyaluronidase affinity medium, formed by connecting affinity ligand and a chromatography medium. Preferably, the affinity ligand is a modified molecule of 6-chloromethyluracil and 2-aminobenzimi-dazole, and the chromatography medium is sepharose. The invention also provides a synthesis method of the cattle-sourced hyaluronidase affinity matrix, and a methodof using the affinity matrix to adsorb cattle-sourced hyaluronidase. By using the affinity matrix, the cattle-sourced hyaluronidase affinity can be specifically adsorbed and purified, and the application and popularization potential is great.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to the technical field of medical enzyme production, in particular to a group of bovine-derived hyaluronidase affinity media and an adsorption process thereof. Background technique [0002] Hyaluronidase (HAase; EC 4.2.99.1) is an enzyme that can hydrolyze hyaluronic acid (hyaluronic acid is a component in the tissue matrix that restricts the diffusion of water and other extracellular substances), used for human body energy Temporarily reducing the viscosity of the intercellular substance can promote the diffusion of subcutaneous infusion, locally accumulated exudate or blood to facilitate absorption. It is an important drug diffusing agent. It is clinically used as a drug penetration agent to promote the absorption of drugs and promote the dissipation of local edema or hematoma after surgery and trauma. In the plastic surgery industry, hyaluronidase is used to digest the ag...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/24B01J20/28B01D15/38B01J20/30
CPCB01D15/3814B01J20/22B01J20/24B01J20/28002
Inventor 辛瑜张梁
Owner JIANGNAN UNIV
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