Analysis of urinary porphyrins by high performance liquid chromatography-electrospray ionization mass spectrometry

A high-performance liquid chromatography coupled with electrospray ionization mass spectrometry (HPLC-ESI-MS) method was developed for the simultaneous separation and detection of six porphyrins, namely uroporphyrin (UP), heptacarboxylic acid porphyrin (HEPTAP), hexacarboxylic acid porphyrin (HEXAP), pentacarboxylic acid porphyrin (PENTAP), coproporphyrin (CP), and mesoporphyrin (MP), in human urine samples. The recoveries of the porphyrins in urine ranged from 84 to 108%. The intra- and inter-day precisions at spiked porphyrin concentrations were 4.0-9.7% and 5.5-15%, respectively. The limits of detection (LOD) ranged from 0.2 to 3nM. The developed method was successfully applied to the determination of endogenous porphyrins in human urine. UP, HEPTAP, and PENTAP were detected at levels of 42.4, 21.3, and 18.0nM, respectively. CP was detected as the predominant species at 135.6nM, wheres MP and HEXAP were not detected in the urine samples from healthy females. Significant increases of CP excretion in urine from liver disease patients were observed. This method proved potentially applicable for the routine diagnosis and treatment of porphyrin-related diseases.