The Assessment Of Folic Acid And Human Insulin By Mass Balance Method And Quantitative Nuclear Magnetic Resonance Method

Folic acid is an essential vitamin which plays an important role in the process of nucleic acid synthesis and amino acid metabolism.It is of practical significance to monitor the purity of synthetic folic acid and detect the content of impurities in the quality control of drugs but there is no literature reporting about the analysis of organic impurity.In this work,the impurities of folic acid were systematic analyzed based on the mass balance method.The triple quadruple liquid chromatography-mass spectrometry was utilized to identify organic impurities of folic acid,and it was found that there are 0.69%petroic acid with the RSD of 2.05%,and N-(4-aminobenzoyl)-glutamic acid 3.1%with the RSD of 0.61%in the folic acid.The moisture content measured by Karl Fischer titration method was 7.60%,and the content of chloride by ion chromatography method was 0.029%.Thus,the final purity of folic acid measured by this mass balance method was 91.1%with an uncertainty of 0.6%.This further analysis study of the impurities in folic acid will assure the accuracy of determination of folic acid and lay a solid foundation for the development of national first grade certified reference material of folic acid.Human insulin is the hormone with a relatively simple structure to lower blood glucose.Among the commonly detecting methods,the immunization method is lack of recognition of the inactivated insulin,and the isotope dilution mass spectrometry(IDMS)has a quite complicated pretreatment.An analytical method for the purity determination of human insulin(HI)was developed in this study.Firstly,the human insulin was prepurificated by high performance liquid chromatography(HPLC)andthe purity by HPLC was up to 99.78%.Since the insulin is easily degradable during purification,the stabilizer,the pH of mobile phase,the suitable salt concentration in the mobile phase and the temperature during freeze-drying process were investigated and optimized.Then the purified human insulin was assessed by quantitative nuclear magnetic resonance(qNMR)to act as a reference material.Finally,the human insulin bought from Sigma company was measured by HPLC as the puritymean with its uncertainty84.52%± 1.14%,which was consistent with that by IDMS method(86.6%± 3.4%).This method by qNMR with a prepurification firstly expanded the application of qNMR to proteins with molecular weight of about 5800.