A determination method for purity of creatinine was developed through qualitative analysis,quantitative analysis and uncertainty evaluation.The chemical composition of creatinine sample was determined by liquid chromatography-tandem mass spectrometry(LC-MS/MS) and nuclear magnetic resonance spectrometry(NMR) at first,and then the certified value was studied by mass balance method(including analysis of liquid chromatography,water content,ash content,volatile material and inorganic elements) and quantitative nuclear magnetic resonance(qNMR) method as well.The certified value of creatinine was 99.7%,with the expanded uncertainty of 0.4%.The research is important for correct diagnosis and treatment since it could assure the accuracy of determination of creatinine in Lab.The certified value of creatinine was so far as to use as the internal standard for qNMR.A certified reference material(CRM) for purity of creatinine was developed after homogeneity and stability study.