1. Into salt. Weigh heparin sodium crude 30g, add proper amount of pure water to dissolve, get heparin sodium solution; 75g benzalkonium chloride was weighed and dissolved in pure water to obtain benzalkonium chloride solution. Slowly add benzathonium chloride solution to heparin sodium solution, stir while adding, stir evenly and leave for 2 hours. Then centrifuge the centrifuge operation, the centrifuged solid was washed with pure water, and finally the obtained heparin benzyl chloride ammonium salt was vacuum dried to obtain heparin benzyl chloride ammonium salt.
2. The esterification. 5 times of dichloromethane was added to the obtained heparin benzoxo chloro-ammonium salt, and then diploid benzyl chloride was added to the obtained heparin benzoxo chloro-ammonium salt at 35℃ for 24 hours. At the end of the reaction, a 10% sodium acetate methanol solution containing 6 times heparin benzyl chloride ammonium salt was added, stirred for 10 minutes, left for 4 hours, and then the precipitate was filtered. The filtered precipitate was washed twice with methanol to obtain wet crude heparin benzazonium chloride. Add 8% sodium chloride aqueous solution 3 times the weight of heparin ethanoammonium salt to the wet crude product of heparin benzyl ester, stir until completely dissolved, then add methanol 4 times the volume of sodium chloride solution, stir again for 10 minutes, stand for 2 hours, filter, repeat this operation 3 times to obtain the wet product of heparin benzyl ester. Benzyl heparin was obtained by vacuum drying the wet product of benzyl heparin.
3. The depolymerization. Purified water 20 times its weight was added to benzyl heparin, stirred to dissolve and heated to 60℃. The sodium hydroxide of 10% weight of heparin benzyl ester was dissolved in pure water, and then slowly added into the heparin benzyl ester solution, kept at 60℃, stirred for 2 hours. Then, the ph of the solution was adjusted to 6.0-6.5 with appropriate hydrochloric acid, and cooled to room temperature to obtain the depolymerization solution. Add an appropriate amount of sodium chloride to the depolymerization solution, make its concentration 10%, stir until completely dissolved, and then add methanol 60 times the weight of heparin benzyl ester while stirring, stir evenly, and then leave it for 5 hours to separate the supernatant and obtain enoxaparin sodium wet crude product.
4. Oxidation alcohol precipitation. Add 3 times the volume of pure water by weight to the wet crude enoxaparin sodium product, stir until it is completely dissolved, and add pure water to bring the solution concentration to 20%. Then add 1% of the volume of sodium chloride solution, stir and heat to 35℃, adjust the ph of the solution to 9.5-10.0 with sodium hydroxide; Then, 1% of the volume of hydrogen peroxide (30% concentration) was added to the solution, the temperature and ph were maintained, and the oxidation solution was obtained for 8 hours. The oxidation solution was filtered with 0.45μm filter element, and the pH was adjusted to 6.0~6.5 with hydrochloric acid. Methanol, 5 times the volume of filtrate, was added after agitation, and stirred evenly. After standing for 5 hours, the supernatant was separated to obtain enoxaparin sodium oxidized alcohol precipitate.
5. Sterilization and filtration. Add proper amount of pure water to the alcohol precipitate after oxidation, make its concentration reach 25%, stir until completely dissolved, and then filter the solution through the filter element. Sterile filtrate was dried, crushed and mixed with freeze dryer to obtain enoxaparin sodium.