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Method for separating albumin nonconjugate, albumin conjugate and micro-molecular compound

A technology of small molecular compounds and conjugates, which is applied in the preparation methods of peptides, chemical instruments and methods, animal/human proteins, etc., can solve the problems of small batch processing and small sample loading, and achieve large processing batches and separation effects. Good, solve the effect of not removing free small molecule compounds

Inactive Publication Date: 2018-11-20
HEBEI CHANGSHAN BIOCHEM PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the albumin solution containing small molecular compounds, the molecular weight of albumin is more than ten times the molecular weight of small molecular compounds, so professionals usually use gel filtration chromatography according to the large difference in molecular weight when separating small molecular compounds. For example, GE’s superdex75 or superdex200 filler, although effective separation can be achieved by gel filtration chromatography, the disadvantage of this technology is that the sample volume is small and the processing batch is small

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  • Method for separating albumin nonconjugate, albumin conjugate and micro-molecular compound
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  • Method for separating albumin nonconjugate, albumin conjugate and micro-molecular compound

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Embodiment 1

[0029] a. After cleaning and disinfecting the phenyl sepharose column, equilibrate the phenyl sepharose with 2 column volumes of pH 6.0 low-salt loading eluent (20mM sodium phosphate, 1mM ammonium sulfate, 5mM sodium caprylate) Gel chromatographic column, then the mixed solution containing albumin binder and unbound GLP-1 analog after the reaction of human serum albumin and GLP-1 analog is loaded onto the phenyl sepharose chromatographic column;

[0030] b. After loading the sample, continue to wash the phenyl sepharose column with pH 6.0 low-salt loading eluent (composed of 20mM sodium phosphate, 1mM ammonium sulfate, 5mM sodium caprylate), when the liquid flows out of the column When the UV absorbance of the chromatographic column reaches 10mAU, the washing solution containing only the albumin conjugate is collected, and the collection is stopped when the UV absorbance of the effluent liquid in the chromatographic column drops to the baseline level;

[0031] c. After stoppin...

Embodiment 2

[0032] Embodiment 2: The difference between this embodiment and embodiment 1 is that the concentration of ammonium sulfate is adjusted from 1 mM to 5 mM.

[0033] a. After cleaning and disinfecting the phenyl sepharose column, equilibrate the phenyl sepharose with 2 column volumes of pH 6.0 low-salt loading eluent (composed of 20mM sodium phosphate, 5mM ammonium sulfate, 5mM sodium caprylate) Gel chromatographic column, and then the mixed solution containing albumin binder and non-binding GLP-1 analog after the reaction of albumin and GLP-1 analog is loaded onto the phenyl sepharose chromatographic column;

[0034] b. After loading the sample, continue to wash the phenyl sepharose column continuously with pH 6.0 low-salt loading eluent (composed of 20mM sodium phosphate, 5mM ammonium sulfate, 5mM sodium octanoate). When the UV absorbance of the chromatographic column reaches 10mAU, the washing solution containing only the albumin conjugate is collected, and the collection is s...

Embodiment 3

[0036] Embodiment 3: The difference between this embodiment and embodiment 1 is that the concentration of ammonium sulfate is adjusted from 1 mM to 10 mM.

[0037] a. After cleaning and disinfecting the phenyl sepharose column, equilibrate the phenyl sepharose with 2 column volumes of pH 6.0 low-salt loading eluent (20mM sodium phosphate, 10mM ammonium sulfate, 5mM sodium caprylate) Gel chromatographic column, and then the mixed solution containing albumin binder and non-binding GLP-1 analog after the reaction of albumin and GLP-1 analog is loaded onto the phenyl sepharose chromatographic column;

[0038] b. After loading the sample, continue to wash the phenyl sepharose column continuously with pH 6.0 low-salt loading eluent (composed of 20mM sodium phosphate, 10mM ammonium sulfate, 5mM sodium caprylate), when the liquid flows out of the column When the UV absorbance of the chromatographic column reaches 10mAU, the washing solution containing only the albumin conjugate is col...

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Abstract

The invention discloses a method for separating an albumin nonconjugate, an albumin conjugate and a micro-molecular compound. The method comprises the following steps: balancing a chromatographic column by using eluent, and loading a mixed solution containing the albumin conjugate and the unconjugated micro-molecular compound to a chromatographic column; and washing the chromatographic column by using the eluent, starting to collect washing liquid containing the albumin conjugate when the UV absorbance of the liquid flowing out of the chromatographic column reaches to 10 mAU, and stopping collecting when the UV absorbance is reduced to a baseline. The free micro-molecular compound is separated from the solution containing the albumin conjugate and the albumin nonconjugate, and the advantages of good separation effect and large treatment batch are achieved. The defect that the free micro-molecular compound cannot be removed is overcome and the defect that the treatment batch by a gel filtration chromatography method is small is overcome.

Description

technical field [0001] The present invention relates to a method for separating albumin non-conjugates and albumin conjugates and free small molecule compounds from a solution containing albumin non-conjugates and albumin conjugates. Background technique [0002] Albumin itself has the characteristics of biodegradability, non-toxicity, non-antigenicity, patient tolerance and high bioavailability, so albumin has attracted extensive attention in the field of biomedicine. Among them, albumin drug-loading technology is widely used and mature. Albumin drug-loaded system can improve the pharmacokinetic characteristics of drugs, such as prolonging the half-life of drugs and enhancing drug activity. At present, a number of drugs have been marketed using related technologies, such as albumin nano-paclitaxel and semaglutide. [0003] Conjuchem's drug affinity construction technology platform technology is also a technology platform that uses albumin-coupled drugs. The platform is to ...

Claims

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Application Information

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IPC IPC(8): C07K14/605C07K14/62C07K1/20
CPCC07K14/605C07K14/62
Inventor 杜旭召庞田白茜茜李云峰
Owner HEBEI CHANGSHAN BIOCHEM PHARMA