Towards a definitive assay of creatinine in serum and in urine: separation by high-performance liquid chromatography.

A fast and sensitive method for the separation of serum and urinary creatinine is described. For the preliminary purification of serum and urine, a cation-exchange column is used to remove protein, anions and neutral compounds prior to isolation of creatinine by high-performance liquid chromatography. A reversed-phase system with 0.01 M ammonium acetate solution as the mobile phase can separate creatinine in 7.5 min at a flow-rate of 1 ml/min. The purity of the separated creatinine is proved by derivatization using trifluoracetic anhydride, followed by gas chromatography and mass spectrometry. Although this method of purification was designed for incorporation into a definitive assay, the ease and speed of analysis makes is very attractive for routine clinical use.