Chromatography medium taking 4-aminopyrrolo[3,2-D]pyrimidine as functional ligand

A technology of aminopyrrole and chromatographic medium, applied in the field of protein chromatographic separation technology, can solve the problems of difficult to meet the requirements of the separation process, poor specificity and selectivity, and low number of repeated uses, and achieve significant hydrophobic charge-induced chromatographic separation Features, high coupling efficiency, and the effect of reducing operation steps

Active Publication Date: 2018-03-16
SUZHOU BOJIN BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Antibody products often require high purity and must maintain biological activity, so traditional separation processes are often difficult to meet the requirements
Protein A or protein G affinity chromatography media can specifically bind antibodies, but protein affinity ligands are easily degraded by proteases in the feed solution, contaminating the product, and the number of reuses is low. The media is expensive and the operating cost is extremely high. limit its large-scale application
Although traditional methods such as ion exchange chromatography and hydrophobic interaction chromatography can bind antibodies, they have poor specificity and selectivity, many separation steps, and limited purification effects.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] A preparation method of a chromatographic medium using 4-aminopyrrolo[3,2-D]pyrimidine as a functional ligand, comprising the following steps: taking 10 g of hydroxylated methacrylic acid hydrophilic polymer microspheres, adding 3g of 30% (v / v) dimethyl sulfoxide, 5g of 3-chlorophenyl oxirane and 2g of sodium hydroxide were activated in a 200rpm shaker at 25°C for 15 hours, suction filtered, and washed with deionized water to obtain Activate the chromatography matrix; mix the activated chromatography matrix with 2g of 6-aminoacetic acid for reaction, react in a 200rpm shaker at 30°C for 3-10 hours, filter with suction, and wash with deionized water to obtain an amino-activated matrix; activate the amino The substrate was mixed with 3g of 4-aminopyrrolo[3,2-D]pyrimidine and 0.5M sodium carbonate buffer solution, the pH of the sodium carbonate buffer solution was 10, and the reaction was carried out in a shaker at 200rpm at 30°C for 15 hours to obtain 4-amino The medium a...

Embodiment 2

[0020] A preparation method of a chromatographic medium using 4-aminopyrrolo[3,2-D]pyrimidine as a functional ligand, comprising the following steps: taking 10 g of hydroxylated methacrylic acid hydrophilic polymer microspheres, adding 2g of 30% (v / v) dimethyl sulfoxide, 6g of 3-chlorophenyl oxirane and 4g of sodium hydroxide were activated in a shaker at 200rpm at 25°C for 25 hours, suction filtered, and deionized Wash with water to obtain an activated chromatography matrix; mix the activated chromatography matrix with 1 g of 6-aminoacetic acid for reaction, react in a shaker at 200 rpm at 30°C for 6 hours, filter with suction, and wash with deionized water to obtain an amino-activated matrix; Amino-activated matrix was mixed with 5 g of 4-aminopyrrolo[3,2-D]pyrimidine and 0.5M sodium carbonate buffer, the pH of the sodium carbonate buffer was 10, and reacted in a shaker at 200 rpm at 30°C for 20 hours to obtain 4-aminopyrrolo[3,2-D]pyrimidine-coupled medium; filter the 4-ami...

Embodiment 3

[0022] A preparation method of a chromatographic medium using 4-aminopyrrolo[3,2-D]pyrimidine as a functional ligand, comprising the following steps: taking 10 g of hydroxylated methacrylic acid hydrophilic polymer microspheres, adding 5g of 30% (v / v) dimethyl sulfoxide, 8g of 3-chlorophenyl oxirane and 4g of sodium hydroxide were activated for 40 hours in a 200rpm shaker at 25°C, suction filtered, and deionized Wash with water to obtain an activated chromatographic matrix; mix the activated chromatographic matrix with 3 g of 6-aminoacetic acid for reaction, react in a shaker at 200 rpm at 30°C for 8 hours, filter with suction, and wash with deionized water to obtain an amino-activated matrix; Amino-activated matrix was mixed with 6g of 4-aminopyrrolo[3,2-D]pyrimidine and 0.5M sodium carbonate buffer, the pH of the sodium carbonate buffer was 10, and reacted in a shaker at 200rpm at 30°C for 20 hours to obtain 4-aminopyrrolo[3,2-D]pyrimidine-coupled medium; the 4-aminopyrrolo[...

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Abstract

The invention relates to a chromatography medium taking 4-aminopyrrolo[3,2-D]pyrimidine as a functional ligand. The chromatography medium taking 4-aminopyrrolo[3,2-D]pyrimidine as the functional ligand is characterized by comprising a chromatography substrate and a ligand, wherein the chromatography substrate is a hydrophilic porous microsphere with hydroxide radical; the ligand is 4-aminopyrrolo[3,2-D]pyrimidine activated and coupled by 3-chlorphenyl ethylene oxide. The novel chromatography medium developed in the invention takes the 4-aminopyrrolo[3,2-D]pyrimidine as the functional ligand, has obvious hydrophobic charge-induced chromatographic separation characteristics, has extremely high adsorption capacity on antibodies and can be applied to large-scale antibody preparation.

Description

technical field [0001] The invention relates to a chromatographic medium using 4-aminopyrrolo[3,2-D]pyrimidine as a functional ligand, which belongs to protein chromatographic separation technology in the field of biochemical industry. Background technique [0002] Immunoglobulin (Ig), also known as antibody (antibody, Ab), is a type of glycoprotein in blood and interstitial fluid. It is produced by the proliferation and differentiation of B cells into plasma cells after being stimulated by antigens. It is an important effector molecule of humoral immunity. . [0003] There are various antibody preparation technologies, but no matter what technology is used to prepare antibodies, different separation and purification methods need to be used in the later stage to obtain antibody products that meet the requirements. The separation and purification of antibodies should follow the principle of reducing the Purification steps, improve the separation efficiency of each step, ther...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/26B01J20/30C07K1/16
CPCB01J20/26C07K1/16
Inventor 瞿欢欢朱至放
Owner SUZHOU BOJIN BIOLOGICAL TECH
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