Purifying method of encephalitis B vaccine

Abstract

The invention provides a purifying method of an encephalitis B vaccine, which comprises the following steps: carrying out column chromatography on an inactivated encephalitis B virus concentrated solution through a polypropylene glucan molecular sieve, and detecting at a wavelength of 280 nm with an ultraviolet detector, wherein an eluting solvent is a phosphoric acid buffer solution with the pH of 7.5-8.5; collecting a first eluting peak, and carrying out ion exchange chromatographic film chromatography by using a cellulose acetate film with quaternary ammonium positive ions, wherein linear elution is adopted; further, the volume ratio of a buffer solution A to a buffer solution B, which is equal to 0-100% in the eluting solvent is changed at a constant speed to the volume ratio of the buffer solution A to the buffer solution B, which is equal to 100%-0; the buffer solution A is the phosphoric acid buffer solution with the pH of 7.5-8.5; the buffer solution B is the phosphoric acid buffer solution which comprises sodium chloride with the concentration of 500 mM / L and has the pH of 7.5-8.0; and an eluting speed is 200-600 cm / hour; and collecting an eluent, wherein the concentration of NaCl (sodium chloride) is 70-250 mM. By using the purifying method, impurities, particularly DNA (deoxyribonucleic acid) which significantly influences the safety of a product, can be effectivelyremoved on the basis that the immunogenicity of the vaccine is protected; simultaneously, the purifying time of the encephalitis B vaccine is shortened; and the production cost is decreased.

Description

technical field [0001] The invention relates to a purification method of Japanese encephalitis vaccine, in particular to a method for purifying inactivated virus Japanese encephalitis vaccine by molecular sieve chromatography and ion exchange membrane chromatography. Background technique [0002] Transforming traditional vaccine products with modern biotechnology, improving the safety and protective effect of traditional vaccines, and making the control of vaccine production process and product quality fully meet the requirements of modern pharmaceutical management are the development direction of high-tech supported by the state ("High-tech Industrialization "Eleventh Five-Year Plan" National Development and Reform Commission). Adopting a vaccine production process centered on large-scale animal cell culture technology, large-scale chromatographic purification technology and new preparation technology is the main way to achieve this goal. However, these new technologies ha...

AI Technical Summary

Problems solved by technology

This method only uses a single principle of different molecular weights to separate impurities, and can only separate substances with different molecular weights. For substances with similar molecular weights, they cannot be separated even after a longer chromatographic path. For impurities with similar molecular weights to antigens, especially It is DNA that cannot be effectively removed

Moreover, in order to achieve a higher protein removal rate in one step, it is necessary to pass the antigen through a longer chromatographic path, and the protein on the surface of the virus will inevitably be damaged, reducing the immunogenicity of the antigen. In order to ensure sufficient efficacy, it is necessary to add Higher levels of antigens, resulting in a higher probability of adverse reactions at the time of vaccination

[0009] Traditional ion-exchange chromatography uses ion-exchange packing as the column chromatography medium, and ion-exchange packing is connected to cations or anions and other groups on the skeleton of particles such as agarose or dextran, and 90% of the groups are located in the packing particles. Internally, when separating and purifying macromolecular substances such as viruses, the macromolecular substances cannot enter the small pores of the granular gel and cannot combine with the groups in the pores, resulting in the traditional packing capacity not meeting the requirements of the chromatography process or causing More than 90% waste

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