Triple cross-linking collagen, preparation method and uses thereof

A collagen and manufacturing method technology, applied in dental preparations, pharmaceutical formulations, dental prostheses, etc., can solve the problems of unsatisfactory enzymatic decomposition resistance, low cross-linking degree, and uneven cross-linking.

Active Publication Date: 2014-09-17
SUNMAX BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, if only the cross-linking of amino groups and amino groups is carried out, it is easy to produce collagen fibers with uneven cross-linking and low cross-linking degree, so that its resistance to enzymatic decomposition cannot meet expectations

Method used

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  • Triple cross-linking collagen, preparation method and uses thereof
  • Triple cross-linking collagen, preparation method and uses thereof
  • Triple cross-linking collagen, preparation method and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] 1-1 Collagen fiber reorganization

[0055] Extract the pig-derived collagen solution from pig skin, mix this collagen solution with 0.2M, pH 11.2 phosphate buffer solution at a volume ratio of 9:1, and slowly stir for 6 hours at 25°C to Collagen fiber reorganization of collagen is performed. Next, the restructured collagen fibers were collected by centrifugation, and dispersed in 20 mM, pH 7 phosphate buffer solution to obtain a recombined collagen fiber solution with a concentration of about 2.7 mg / mL.

[0056] 1-2 The first cross-linking reaction

[0057] Glutaraldehyde, which belongs to the aldehyde crosslinking agent, is used as the primary crosslinking agent in this embodiment. Glutaraldehyde was added to the above recombinant collagen fiber solution with a concentration of about 2.7 mg / mL, and the final concentration of glutaraldehyde was 35 ppm. Next, stir slowly for 16 hours at 30°C and pH 7.2 to carry out the covalent cross-linking reaction between amine gro...

experiment example 1

[0066] 1-1 Cross-linking degree test

[0067] In this experimental example, trinitrobenzene sulfonic acid (trinitrobenzenesulphonate, TNBS) reagent was used to analyze the cross-linking degree of triple cross-linked collagen fibers.

[0068]

[0069] Add 4 mg of lyophilized triple cross-linked collagen fibers to 1 ml of 0.1 M sodium bicarbonate solution (sodium bicarbonate solution) at pH 8.5, then add 1 ml of 0.5% TNBS solution by volume. At 40°C, react for 2 hours. After the reaction, 3 ml of 6N hydrochloric acid (HCl) was added, and the reaction was carried out at 60° C. for 1.5 hours. After the reaction was completed, after the reaction solution was cooled to room temperature, 5 ml of deionized water was added and mixed evenly. Take out 5ml of the mixed solution into a spiral test tube, add 10ml of ether, shake evenly and let it stand still, wait for the liquid to separate into layers, and then take out the upper layer of ether. Then, 10 ml of diethyl ether was added,...

Embodiment 2

[0081] 2-1 Collagen fiber reorganization

[0082] Same as the collagen fiber reorganization steps in Example 1.

[0083] 2-2 The first cross-linking reaction

[0084] Same as the first heavy cross-linking reaction step of Example 1.

[0085] 2-3 The second cross-linking reaction

[0086] The second heavy cross-linking reaction in this embodiment is based on the imine cross-linking agent 1-ethyl-3-3-dimethylaminopropyl-carbodiimide (1-ethyl-3-(3-dimethylaminopropyl ) carbodiimide hydrochloride, EDC) as the second cross-linking agent in this embodiment, and also mix EDC with N-hydroxy-succinimide (N-hydroxysuccinimide, NHS) at a molar ratio of 5:1 to The second crosslinker mixture (EDC / NHS) of this example was formed. The first heavy cross-linked collagen collected by centrifugation was dispersed in 0.1M 2-morpholineethanesulfonic acid (MES) buffer solution (pH 5.5) to obtain a heavy cross-linked collagen fiber solution with a concentration of about 2.7mg / mL, Then add the s...

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Abstract

The invention relates to a triple cross-linking collagen preparation method, which comprises: providing a soluble collagen sample, mixing the collagen sample and a first cross-linking agent to form a single cross-linking collagen, mixing the single cross-linking collagen and a second cross-linking agent to form a double cross-linking collagen, and mixing the double cross-linking collagen and a third cross-linking agent to form the triple cross-linking collagen, wherein the first cross-linking agent, the second cross-linking agent and the third cross-linking agent are separately selected from a group comprising an aldehyde cross-linking agent, an imine cross-linking agent and an epoxide cross-linking agent, the first cross-linking agent is different from the second cross-linking agent, and the third cross-linking agent is different from the first cross-linking agent and the second cross-linking agent.

Description

technical field [0001] The present invention mainly relates to a method for producing triple cross-linked collagen, especially a triple cross-linked collagen suitable for manufacturing high melting point, low enzymatic degradation rate, and high stability, and the triple cross-linked Collagen is used in medical treatments that require long-term implantation of collagen. Background technique [0002] Collagen exists in the connective tissue of the human body and is the structural matrix of tissues and organs such as bones, cartilage, tendons, ligaments, blood vessels, corneas, basement membranes, and skin. Collagen is often used in medical treatment and tissue engineering to strengthen or protect human tissues due to its low immune response. [0003] However, related medical materials mainly composed of collagen are usually degraded by enzymes in the body within a short period of time after being implanted into the human body, and then metabolized and disappear. Therefore, m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C08J3/24C08L89/00C08K5/07C08K5/1515C08K5/29C08K5/3415A61K6/02A61K47/42A61L27/24A61L27/60
Inventor 林育德林建兴卢香吟林峰辉
Owner SUNMAX BIOTECH
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