Optimization of Production Process for Diethylene Insulin Active Pharmaceutical Ingredient
OBJECTIVE To construct and express Detemir proinsulin,develop raw material production processes,optimize key processes to improve and stabilize recovery rates,and manafacture the raw materials with the same quality as the original drug.METHODS By establishing DesB30 engineering bacteria for fermentation and expression of Detemir proinsulin.Capturing Detemir proinsulin through ion exchange chromatography.Cut off the linked short peptides through enzyme reaction.Reverse phase chromatography after targeted modification of fatty acid chains to obtain Levemir.Finally,through two steps of reverse phase chromatography and freeze-drying,the raw material powder was obtained.RESULTS Optimization exchange chromatography to increase the elution concentration of proinsulin,and convenient for production operations;Optimize enzyme reaction and modification reaction,improve and stabilize recovery rates and target protein purity,to ensure the quality of the final product.The overall yield of this process reaches 25％,and the purity of the target protein＞99％.CONCLUSION The molecular weight,isoelectric point,amino acid sequence,disulfide bond,and insulin receptor affinity of the samples prepared by this process are consistent with the quality of the original drug.The process is robustness and can be used as a reference for large-scale production.
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