A method for adjusting the isoelectric point of type Ⅱ collagen

An isoelectric point and collagen technology, applied in the field of electric point, can solve the problems of increasing the difficulty of dispersion, difficult to control the structure of composite materials, and aggregation of isoelectric points, and achieve the effect of improving the reaction rate

Active Publication Date: 2019-08-20
HEBEI COLLAGEN BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since the isoelectric point of type II collagen is close to neutral, the solution environment during the preparation of organic / inorganic composites is often in a neutral environment, which leads to the formation of isoelectric point aggregation of type II collagen, which increases the difficulty of dispersion and makes the prepared composite Material structure is difficult to control

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] (1) Collagen pretreatment: Weigh 100g of bovine type II collagen with a triple helix structure and remove the telopeptide, put it in a 50L stirred reaction kettle, add 10kg of acetic acid solution with a concentration of 0.1mol / L, and stir for 24 hours to make it all dissolve. The ultrafiltration membrane with a molecular weight cut-off of 1kDa is used for ultrafiltration. During the ultrafiltration process, a NaOH solution with a pH of 7.5 is added to the stirred reactor, and the ultrafiltration is continued until the pH of the permeate reaches 7.5.

[0020] (2) Reaction process: import the type II collagen alkaline solution prepared in step (1) into a constant temperature reactor, control the temperature of the solution in the reactor at 10°C, and continuously add an acid anhydride aqueous solution into the reactor, the total amount of acid anhydride added is 10g , during the reaction process, the pH of the reaction solution was controlled to be maintained at pH 7.5 b...

Embodiment 2

[0025] (1) Collagen pretreatment: Weigh 100g bovine type II collagen with a triple helix structure and remove telopeptides, put it in a 50L stirred reaction kettle, add 50kg of acetic acid solution with a concentration of 0.3mol / L, and stir for 24 hours to make it completely dissolve. The ultrafiltration membrane with a molecular weight cut-off of 20kDa is used for ultrafiltration. During the ultrafiltration process, a KOH solution with a pH of 7.8 is added to the stirred reactor, and the ultrafiltration is continued until the pH of the permeate reaches 7.8.

[0026] (2) Reaction process: import the type II collagen alkaline solution prepared in step (1) into a constant temperature reactor, control the temperature of the solution in the reactor at 25°C, and continuously add an aqueous solution of acid anhydride to the reactor, the total amount of acid anhydride added is 6g , during the reaction process, the pH of the reaction solution was controlled at pH 9.5 by using a potass...

Embodiment 3

[0031] (1) Collagen pretreatment: Weigh 80g of bovine type II collagen with a triple helix structure and remove the telopeptide, put it in a 50L stirred reaction kettle, add 80kg of acetic acid solution with a concentration of 0.5mol / L, and stir for 24 hours to make it all dissolve. The ultrafiltration membrane with a molecular weight cut-off of 30kDa is used for ultrafiltration. During the ultrafiltration process, a NaOH solution with a pH of 11.0 is added to the stirred reactor, and the ultrafiltration is continued until the pH of the permeate reaches 11.0.

[0032] (2) Reaction process: import the type II collagen alkaline solution prepared in step (1) into a constant temperature reactor, control the temperature of the solution in the reactor at 40°C, and continuously add an acid anhydride aqueous solution into the reactor, the total amount of acid anhydride added is 80g , during the reaction process, the pH of the reaction solution was controlled to be maintained at 11.0 b...

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Abstract

The invention discloses a method for adjusting isoelectric points of collagen II. The method includes the steps of 1), weighing a certain amount of the collagen II with a three-screws structure and with telopeptide removed, adding 100-1000 times the weight of 0.05-0.5mol / L of an organic acid solution, allowing to dissolve completely by means of stirring, performing displacement of solution with an ultrafiltration method, adding an alkaline solution with pH value being 7.5-11 during ultrafiltration, and continuously performing ultrafiltration till difference value between a permeate solution pH and the added alkaline solution pH is lower than 0.2; 2), feeding the alkaline solution of the collagen II to a constant-temperature reaction kettle, controlling the temperature of the solution in the reaction kettle to be 10-40DEG C, continuously adding an acid aqueous solution to the reaction kettle, controlling total adding amount of acid anhydride to be 0.01-1 times the weight of the collagen II, and adopting the alkaline solution with the pH value being 12-14 to control pH of the reaction solution to be maintained in pH 7.5-11; 3), adding an acid diluting solution to the solution prepared in the step 2) till flocky precipitate appears in the solution, recording the pH value of the solution during settling, subjecting the solution to centrifugal treatment and collecting the precipitate; 4), subjecting the precipitate to freeze drying.

Description

technical field [0001] The invention belongs to the field of biomedical materials, in particular, the invention relates to a method for adjusting the isoelectric point of type II collagen. [0002] technical background [0003] Collagen is a natural biopolymer expressed by animal cells with a triple helical structure. It consists of three chains wound in a right-handed helical form to form a superhelical structure, and each peptide chain has about 1,000 amino acid residues. Collagen, as the main protein component of the extracellular matrix (ECM), exists in almost all tissues, accounting for about 33% of the total protein in mammals. In mammals, collagen interacts with biological macromolecules such as proteins and polysaccharides to form a complex network structure, which acts as a scaffold for cells and tissue structures, and plays an important role in the morphology, metabolism, growth and differentiation of cells and tissues. Impact. Collagen is divided into various typ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/78
Inventor 王云松赵磊磊冯俊超
Owner HEBEI COLLAGEN BIOTECH
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