CN-121972137-A - Modified chitosan magnetic adsorbent, preparation method thereof and method for removing high molecular substances in posterior pituitary injection

CN121972137ACN 121972137 ACN121972137 ACN 121972137ACN-121972137-A

Abstract

The application discloses a modified chitosan magnetic adsorbent, a preparation method thereof and a method for removing high molecular substances in posterior pituitary injection, belonging to the technical field of biological medicine purification. The method for removing high molecular substances in the posterior pituitary injection comprises heating glacial acetic acid solution, adding posterior pituitary powder and chitosan, maintaining the temperature, stirring, filtering to remove residue, adjusting pH, heating, adding modified chitosan magnetic adsorbent, stirring, cooling, filtering to remove adsorbent, and filtering with multi-stage microporous membrane to obtain the final product. The application can efficiently and targetedly remove high molecular protein polymer, endotoxin and other impurities in the posterior pituitary injection, the removal rate is more than 100 percent, active ingredients are retained to the greatest extent, the process is simplified, the efficiency and the controllability are improved, the preparation purity and the medication safety are obviously improved, and the application is suitable for purifying polypeptide biological preparations and has wide industrial application value.

Inventors

JIANG DABO
Shi Lingyang
SHI SHANWEN
ZHANG ANLING
XIE LEI
LI PAN

Assignees

成都市海通药业有限公司
成都通德药业有限公司

Dates

Publication Date: 20260505
Application Date: 20260209

Claims (10)

1. The preparation method of the modified chitosan magnetic adsorbent is characterized by comprising the following steps of: The amphoteric carboxymethyl chitosan-based flocculant is prepared by taking carboxymethyl chitosan as a main chain, acrylamide and methacryloyloxyethyl trimethyl ammonium chloride as grafted side chains and performing ultraviolet light initiation reaction; Introducing magnetic nano particles into the amphoteric carboxymethyl chitosan-based flocculant to prepare composite magnetic particles; and chemically modifying the composite magnetic particles by using citric acid and diethylenetriamine to prepare the modified chitosan magnetic adsorbent.
2. The method of claim 1, wherein the ultraviolet light initiated reaction comprises the steps of: adding an acrylamide monomer and a methacryloyloxyethyl trimethyl ammonium chloride monomer into a carboxymethyl chitosan aqueous solution, and continuously introducing high-purity nitrogen for 30-50 min; Adding a photoinitiator, carrying out a reaction for 1-3 hours at room temperature under the irradiation of an ultraviolet lamp, and controlling the pH of the solution to 7.5-8.5; Precipitating, washing, vacuum drying and pulverizing the colloidal product after the reaction is finished to obtain amphoteric carboxymethyl chitosan-based flocculant powder, wherein, The mass of the photoinitiator is 0.03% -0.05% of the total mass of the monomers.
3. The preparation method of claim 1 or 2, wherein the mass ratio of carboxymethyl chitosan, acrylamide and methacryloyloxyethyl trimethyl ammonium chloride is (0.8-1.2): 8-12): 6-10.
4. The preparation method of claim 1, wherein the magnetic nanoparticles are introduced into the amphoteric carboxymethyl chitosan-based flocculant to prepare composite magnetic particles, and the preparation method comprises the following steps: Adding FeCl 3 ·6H 2 O and FeCl 2 ·4H 2 O into an amphoteric carboxymethyl chitosan-based flocculant aqueous solution, and stirring in a water bath at 38-42 ℃ for 25-35 min under the protection of nitrogen to obtain a mixed solution; dropwise adding ammonia water into the mixed solution, and controlling the dropwise adding time to be 25-35 min to obtain a black solution; Heating the black solution to 55-65 ℃, continuously reacting for 1-3 hours, separating the magnetic material after the reaction is finished, and washing and drying the product to obtain composite magnetic particles; the volume ratio of the ammonia water to the amphoteric carboxymethyl chitosan-based flocculant is 12-18:200, the concentration of the amphoteric carboxymethyl chitosan-based flocculant is 2-3 wt%, the concentration of the ammonia water is 24-26 wt%, the molar ratio of FeCl 3 ·6H 2 O to FeCl 2 ·4H 2 O is 1.8-2.2:1, and the mass ratio of FeCl 3 ·6H 2 O to the amphoteric carboxymethyl chitosan-based flocculant is 1:2-3.
5. The method of claim 1, wherein the chemically modifying the composite magnetic particles with citric acid and diethylenetriamine comprises the steps of: dispersing the composite magnetic particles in a citric acid aqueous solution, and stirring at a constant temperature of 75-85 ℃ for 3-5 hours to obtain a first product; Dispersing the first product in dimethylformamide, adding diethylenetriamine and a catalyst, and carrying out reflux reaction for 5-7.5 hours under the protection of nitrogen at 82-88 ℃; magnetically separating the obtained product after the reaction is finished, and washing and drying to obtain the modified chitosan magnetic adsorbent; Wherein the proportion relation among the composite magnetic particles, the diethylenetriamine and the catalyst is 5g (4 mL-6 mL): 0.5 g-1.5 g, and the catalyst is 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride.
6. A modified chitosan magnetic adsorbent, characterized by being obtained according to the preparation method of any one of claims 1 to 5.
7. A method for removing high molecular substances in posterior pituitary injection is characterized in that the modified chitosan magnetic adsorbent of claim 6 is used for adsorption, and the removal rate of the high molecular substances in posterior pituitary injection reaches 100%.
8. The method of claim 7, wherein the adsorption process comprises the steps of: Preparing an initial product of posterior pituitary injection, and performing primary impurity removal through a filter membrane to obtain a liquid medicine; adjusting the pH of the liquid medicine to 7.0-7.2, and heating the liquid medicine to above 90 ℃ to obtain a pretreatment liquid medicine; And adding the modified chitosan magnetic adsorbent into the pretreatment liquid medicine, stirring for 30-40 min, cooling to 20-30 ℃, and performing secondary impurity removal by using a filter membrane to remove high molecular substances in the posterior pituitary injection.
9. The method of claim 8, wherein the pituitary posterior leaflet injecta is formulated by: And heating the glacial acetic acid solution with the concentration of 0.24-0.28 wt% to 48-52 ℃, and adding pituitary powder and chitosan according to the feed liquid ratio of 50mL (1.5-2.0 g), and carrying out heat preservation and stirring for 20-40 min, wherein the rotation speed of heat preservation and stirring is 100-500 rpm.
10. The method of claim 8, wherein the primary removal of impurities uses a filter of 0.45 μm and the secondary removal of impurities comprises filtering using filters of 0.45 μm, 0.22 μm and 0.22 μm in sequence.

Description

Modified chitosan magnetic adsorbent, preparation method thereof and method for removing high molecular substances in posterior pituitary injection Technical Field The application relates to the technical field of biological medicine purification, in particular to a modified chitosan magnetic adsorbent, a preparation method thereof and a method for removing high molecular substances in posterior pituitary injection. Background The posterior pituitary injection is a polypeptide preparation extracted from posterior pituitary, and the core active ingredients are vasopressin, oxytocin and the like, thus having important application value in clinic. However, due to the limitation of the extraction process, the injection produced by the prior art is easy to remain high molecular impurities, including macromolecular protein polymers, endotoxin and the like, and the impurities not only can increase the immunogenicity of the medicine, reduce the stability of the preparation and increase the visible foreign matters, but also can obviously increase the risk of anaphylactic reaction in clinical use and influence the medication safety. Currently, the purification technology of posterior pituitary injection mainly relies on ultrafiltration membrane fractionation filtration (e.g. 0.45 μm- > 0.22 μm filtration), organic acid precipitation (e.g. acetone/acetic acid extraction), etc. However, the prior art has limited effect of removing high molecular impurities, is difficult to thoroughly solve the problem of impurity residue, and cannot fully meet the production requirements of high purity and high safety of the preparation. Therefore, developing a technology for efficiently and accurately removing high molecular impurities in posterior pituitary injection has important significance in optimizing biological agent purification process and improving product quality. Disclosure of Invention The application aims to provide a modified chitosan magnetic adsorbent, a preparation method thereof and a method for removing high molecular substances in posterior pituitary injection, and the magnetic adsorbent can be used for efficiently removing high molecular protein impurities, endotoxin and other pollutants in posterior pituitary injection by combining the advantages of a magnetic separation technology and selective adsorption, so that the purity of a medicine and the safety of the medicine are obviously improved. In order to achieve the above purpose, the application provides a preparation method of a modified chitosan magnetic adsorbent, which comprises the following steps: The amphoteric carboxymethyl chitosan-based flocculant is prepared by taking carboxymethyl chitosan as a main chain, acrylamide and methacryloyloxyethyl trimethyl ammonium chloride as grafted side chains and performing ultraviolet light initiation reaction; Introducing magnetic nano particles into an amphoteric carboxymethyl chitosan-based flocculant to prepare composite magnetic particles; And chemically modifying the composite magnetic particles by using citric acid and diethylenetriamine to prepare the modified chitosan magnetic adsorbent. Further, the ultraviolet light initiated reaction comprises the following steps: adding an acrylamide monomer and a methacryloyloxyethyl trimethyl ammonium chloride monomer into a carboxymethyl chitosan aqueous solution, and continuously introducing high-purity nitrogen for 30-50 min; Adding a photoinitiator, carrying out a reaction for 1-3 hours at room temperature under the irradiation of an ultraviolet lamp, and controlling the pH of the solution to 7.5-8.5; Precipitating, washing, vacuum drying and pulverizing the colloidal product after the reaction is finished to obtain amphoteric carboxymethyl chitosan-based flocculant powder, wherein, The mass of the photoinitiator is 0.03% -0.05% of the total mass of the monomers. Further, the mass ratio of the carboxymethyl chitosan to the acrylamide to the methacryloyloxyethyl trimethyl ammonium chloride is (0.8-1.2): 8-12): 6-10. Further, magnetic nano particles are introduced into an amphoteric carboxymethyl chitosan-based flocculant, and the composite magnetic particles are prepared and obtained, and the method comprises the following steps: Adding FeCl 3·6H2 O and FeCl 2·4H2 O into an amphoteric carboxymethyl chitosan-based flocculant aqueous solution, and stirring in a water bath at 38-42 ℃ for 25-35 min under the protection of nitrogen to obtain a mixed solution; dropwise adding ammonia water into the mixed solution, and controlling the dropwise adding time to be 25-35 min to obtain a black solution; heating the black solution to 55-65 ℃, continuously reacting for 1-3 hours, separating the magnetic material after the reaction is finished, and washing and drying the product to obtain composite magnetic particles; The volume ratio of the ammonia water to the amphoteric carboxymethyl chitosan-based flocculant aqueous solution is 12-18:200, the concentration of the amphoteric