Method for preparing narrow-distribution low average molecular weight polymerized hemoglobin

Abstract

A preparation method of narrow-distribution polymerized hemoglobin with low average molecular weight is characterized in that the method includes the following steps: {1} the hemoglobin is added into the gel retardation spectrum for balance through a pump and then distributively added with double-function crosslinker for polyreaction; {2} buffer solution is used to elute and collect the polymer produced in the step {1}, amide acid is used in the collected solution for stopped reaction and then hyperfiltration and concentration are carried out; {3} the obtained polymerized hemoglobin is treated by reduction by sodium borohydride and then is separated and purified through hyperfiltration and chromatography. The advantages of the present invention are that 1. the polymerized hemoglobin prepared by the present invention is narrow-distribution polymerized hemoglobin with low average molecular weight less than 200kD; 2. the present invention overcomes the poor homogeneity of the polymer in the traditional glutaraldehyde preparation method; 3. the polymerized hemoglobin is easily separated from the unpolymerized single hemoglobin and the reduced number of protein colloid decreases the osmotic pressure without any obvious change in viscosity.

Description

(1) Technical field: [0001] The invention relates to a preparation method of polymerized hemoglobin, in particular to a preparation method of narrowly distributed low average molecular weight polymerized hemoglobin. (two) background technology: [0002] Although the traditional blood transfusion method has played an important role in medical rescue, there are also various problems, such as blood shortage, virus transmission, immune response, and the disposal of expired blood. In various clinical medical treatment and first aid, the quality and quantity requirements for blood transfusion are becoming more and more stringent, so the research and development and use of blood substitutes have generally been paid attention to. [0003] Hemoglobin consists of four subunits (α 1 alpha 2 beta 1 beta 2 ), a protein in red blood cells responsible for oxygen transport. Because of its unique oxygen binding characteristics, normal physiological metabolic pathways, etc., hemoglobin-b...

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Problems solved by technology

However, unmodified polymerized hemoglobin cannot be used directly as a blood substitute, mainly because (1) unmodified hemoglobin will lose the regulation of 2,3-bisphosphoglycerate, which will increase the oxygen affinity of hemoglobin and cannot Effective supply of oxygen to tissues

(2) Unpolymerized hemoglobin will increase the colloid osmotic pressure of plasma

But this method also has some problems such as: the utilization rate of anion exchange medium is low (0.5-4mg hemoglobin / ml solid phase), the use of high molar ratio of glutaraldehyde will cause excessive modification of hemoglobin, and the use of HEPES buffer system without amino groups is expensive. Expensive etc.

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