Background and aims: Development of a candidate reference method based on bidimensional liquid chromatography coupled to ESI-MS/MS and double spike isotope dilution for serum creatinine quantification capable of correcting for creatinine-creatine interconversion during sample pretreatment. Study of the impact of the creatine-creatinine interconversion during the analysis of human serum samples.
Materials and methods: 13C1-creatinine and 13C2-creatine are added to the serum sample. Separation carried out by bidimensional liquid chromatography combining reversed phase and a strong cation exchange chromatography. The heart cut, containing creatine and creatinine, is automatically transferred to the second dimension. Quantification carried out by double spike isotope dilution tandem MS/MS.
Results: Minimization of spectral interferences and ion suppression due to matrix effects while increasing sample throughput compared to the direct coupling of cation exchange chromatography to the ESI source. Trueness of the method studied with the satisfactory analysis of two certified reference materials. Satisfactory intra- and inter-day precisions obtained analysing a serum pool and control sera. Analysis of 93 serum samples revealed negligible interconversions with no correlation with creatine levels.
Conclusions: The method provides adequate analytical figures of merit for serum creatinine determination according to CSLI guidelines. Negligible creatine-creatinine interconversion is promoted with the applied sample preparation procedure.
Keywords: Analyte interconversion; Bidimensional chromatography; Double spike isotope dilution; Serum creatinine; Tandem mass spectrometry.
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