| The relative contents of four kinds of glycan forms in the inovator and biosimilar products of recombinant human thromboplastin(rhTPO) were accurately determined using matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF MS) with quasi-internal calibration,the consistencies between products and among batches were evaluated.The N-glycan,sialic acid and O-glycan of rhTPO were removed off with different endoglycosidase,respectively.bovine serum albumin(BSA)used as the standard was pointed target separate with the sample.The molecular weights of the intact glycoproteins and the different forms of deglycosylated proteins were determined by MALDI-TOF MS with quasi-internal calibration.The glycan contents in samples were calculated by the molecular weight differences among different deglycosylated samples.Finally,the consistency of the inovator and biosimilar products were evaluated.The multi-charge peaks of BSA were used for the quasi-internal calibration of MALDI-TOF MS.And their calibration relative standard deviations(RSDs) were all no more than 0.075%,and the RSDs among different calibrations were among 0.001%-0.004%.The method deviations away from the sequence molecular weight were less than 0.01%.For the biosimilar drug,the relative contents of N-glycan,sialic acid,O-glycan,and glycation were 24.6%,2.9%，9.0%and 5.1%,respectively,and the batch-to-batch consistency was quite good.The total glycan contents were consistent with those reported in the literature.For the innovator drug,the relative contents of N-glycan,sialic acid,O-glycan and glycation were 25.6%,2.9%,7.9%and 3.5%,respectively.The total glycan content was lower than that reported in the literature.The relative contents of glycosylation between the innoventor products and the biosimilar products were almost the same but the relative contents of glycation showed some differences.Glycation of rhTPO has not been reported up-to-now,the differences of the relative contents of glycation might be the important factors for the different rhTPO-manufacturers,the glycation differences were mainly caused by the differences of fermentation process. |