CN103804525A

CN103804525A - Method for removing organic residual in heparin sodium

Info

Publication number: CN103804525A
Application number: CN201310631226.7A
Authority: CN
Inventors: 刘乃山; 陆玉梅; 迟培升; 夏衬来
Original assignee: QINGDAO JIULONG BIO-PHARMACEUTICAL Co Ltd
Current assignee: QINGDAO JIULONG BIO-PHARMACEUTICAL Co Ltd
Priority date: 2013-11-26
Filing date: 2013-11-26
Publication date: 2014-05-21

Abstract

The invention discloses a method for removing organic residual in heparin sodium, wherein the product is mainly prepared through the steps of enzymolysis, protein removal, precipitation, drying and the like. The method mainly aims at solving the problems of complex flow, high dosage of alkaline and acid or salt, long preparation period, low yield and the like of the existing production process of the products; the method is capable of extracting cartilage in chondroitin sulfate through only one step of enzymolysis without alkaline hydrolysis or soaking in salt; the prepared product is pure white and fluffy in texture, and the yield of the product can be 43% and above; the purity of the product is high and up to the domestic and foreign oral administration; and the method is simple in production steps and short in period, the reaction time only takes 3-8 hours, the alkaline dosage is saved by more than 80% in contrast with a diluted alkali enzymolysis method, and in the meantime, the difficulty of downstream purification is reduced and pollution on the environment is reduced, and therefore, the method is environment-friendly.

Description

A kind of method of removing organic residue in heparin sodium

Technical field

The present invention relates to biochemical pharmacy field, be specifically related to a kind of method of removing organic residue in heparin sodium.

Background technology

Sodium chondroitin sulfate A, English name Chondroitin sulfate sodium, it is the acidic mucopolysaccharide extracting in animal cartilage, be present in cartilage, larynx bone, nasal bone (41%), ox, Malaysia and China's diaphragm and the tracheae of animal the polysaccharide chain that the repetition disaccharide unit that its molecular structure forms by β 1 → 3 glycosidic link for β-D-Glucose aldehydic acid and N-acetylamino galactosamine forms more.

Sodium chondroitin sulfate A is mainly used in treating the medicine of rheumatism and rheumatism, also has the effects such as anti-freezing, anticancer, antithrombotic, reducing blood-fat, can be used for treating the symptoms such as headache, migraine, coronary heart diseases and angina pectoris.Abroad, the research of Sodium chondroitin sulfate A pharmacology has been reached to molecular level.

The method of current domestic extraction Sodium chondroitin sulfate A mainly contains three kinds: salt solution, alkaline hydrolysis and enzymolysis process.The main separating and purifying technology adopting has: ethanol precipitation, ion exchange chromatography, Mierocrystalline cellulose partition method, quaternary ammonium salt companion method, absorption method, zone electrophoresis partition method etc.In above concentrated extracting method, although yield can reach very high level, but all there is very large problem in production method, removing acidic protein as existed in salt solution technique regulates pH value 1-2 consuming time very long in the time that product volume is large, amino in chondroitin sulfate is easily dissociated, thereby has affected the yield of chondroitin sulfate.

Summary of the invention

The object of the invention is, a kind of preparation method who improves chondroitin sulfate yield is provided, this method mainly, from energy-conservation, saving reagent, shortening production time, simplification production technique equal angles, provides a kind of preparation method who improves chondroitin sulfate yield, and the method is beneficial to suitability for industrialized production.

Because chondroitin sulfate and protein in cartilage are with together with glycopeptide covalent bonds, under alkaline condition, take trypsinase as catalyzer, glycosaminoglycan can be separated in the collagen protein from cartilaginous tissue.Around this principle, this technique directly adds trypsinase, regulates the PH of reaction, temperature condition, direct production chondroitin sulfate.Like this, shortened technical process, the reaction times only needs 4-8 hour, and alkali charge is saved more than 80% than diluted alkaline enzymolysis process, and yield does not only decline, on the contrary than original high 3-5 percentage point.

Step of the present invention is as follows:

1, pure white clean animal cartilage is put into retort, add appropriate water soaking, and add the trypsinase of the 0.1%-1.6% of cartilage weight, regulate PH between 8.5-9.5, control temperature between 40-55 ℃, stir 2-9 hour;

2, after animal cartilage reacts completely, with hydrochloric acid regulate enzymolysis solution make its PH between 5.5-7.5, be heated to 65-80 ℃, insulation 10-40 minute;

3, after enzymolysis solution is cooling, filter or centrifugal slagging-off, filtrate enzymolysis solution that must be limpider after filtration, adds the sodium-chlor of enzymolysis solution weight 0.2%-2.0%, makes PH between 3.5-5.5, stirs 20-50 minute, filters or centrifugal slagging-off.

4, upper step gained enzymolysis solution is added to the ethanol that its volume 2-4 concentration is doubly 90%, make alcohol concn reach 50%-80%, precipitation;

5, the ethanol that is 95% by upper step throw out by concentration heavy 2 to 3 times again, obtains described chondroitin sulfate finished product after being dried.

Advantage of the present invention is: this preparation method is by the improvement innovation to current technique, adopt further enzymolysis process to produce chondroitin sulfate, pass through orthogonal experiment, find the optimized production process condition of production technique to be: hydrolysis time is 6 hours, hydrolysis temperature is 48 ℃, trypsinase consumption is 0.8% cartilage weight, in three kinds of factors, Hydrolysis degree is mainly to hydrolysis temperature, is secondly enzyme dosage and hydrolysis time.Whole reaction process pH value is controlled between 8.5-9.5.This best of breed can make chondroitin sulfate yield reach 43% and more than.After adopting new technology, can simplify Production Flow Chart, greatly shorten the production time, reduce alkali consumption, reduce the difficulty of downstream purification simultaneously, alleviate the pollution to environment.

Claims

1. remove a method for organic residue in heparin sodium, it is characterized in that comprising the following steps:

(1) pure white clean animal cartilage is put into retort, add appropriate water soaking, and add the trypsinase of the 0.1%-1.6% of cartilage weight, regulate PH between 8.5-9.5, control temperature between 40-55 ℃, stir 2-9 hour;

(2) after animal cartilage reacts completely, with hydrochloric acid regulate enzymolysis solution make its PH between 5.5-7.5, be heated to 65-80 ℃, insulation 10-40 minute;

(3) after enzymolysis solution is cooling, filter or centrifugal slagging-off, filtrate enzymolysis solution that must be limpider after filtration, adds the sodium-chlor of enzymolysis solution weight 0.2%-2.0%, makes PH between 3.5-5.5, stirs 20-50 minute, filters or centrifugal slagging-off;

(4) upper step gained enzymolysis solution is added to the ethanol that its volume 2-4 concentration is doubly 90%, make alcohol concn reach 50%-80%, precipitation;

(5) ethanol that is 95% by upper step throw out by concentration heavy 2 to 3 times again, obtains described chondroitin sulfate finished product after being dried.

2. method according to claim 1, is characterized in that: the sodium-chlor of the heavy 0.2%-2.0% of enzymolysis solution adding in (3) step is saltoutd, to strengthen the sedimentation effect of chondroitin sulfate.

3. method according to claim 1, is characterized in that: hydrolysis temperature is 48 ℃.

4. method according to claim 1, is characterized in that: hydrolysis time is 6h, and pancreatin consumption is 0.8% times of cartilage weight.