We describe here the development, validation, quantification and application of a method for determination of heme porphyrin precursors in the urine of porphyric patients. The isomers coproporphyrinogen I and III (COPRO I and III), uroporphyrinogen I (URO I), heptacarboxylporphyrinogen I (HEPTA I), pentacarboxylporphyrinogen (PENTA I), and hexacarboxylporphyrinogen I (HEXA I) were analyzed. These six urinary heme precursors were determined in urine samples collected from 24 patients by high-performance liquid chromatography (HPLC) equipped with a fluorescence detector. The inter- and intra-day precision (coefficient of variation < 5%) and accuracy (95-99%) were evaluated. The limits of detection and of quantification of the porphyrins, expressed in nmol L-1, were as follows: URO I, 0.62 and 2.05; HEPTA I, 0.59 and 1.96; HEXA I, 0.54 and 1.81; PENTA I, 0.52 and 1.73; COPRO I, 2.03 and 6.77; and COPRO III, 0.43 and 1.44. The method described here satisfactorily results in an acceptable cost-benefit ratio, precision and speed for determining the concentrations of heme precursors in the urine of latent or symptomatic acute intermittent porphyria individuals or porphyria cutanea tarda carriers. Since it was analytically validated, this method may be used for accurate and reliable diagnostic reports to follow-up the onset of acute crisis in porphyria carriers to adopt preventive pharmacological treatment.
porphyrins; heme biosynthesis; acute intermittent porphyria; porphyria cutanea tarda; porphyria diagnosis; HPLC analysis of porphyrinogens
