Rapid reversed-phase high-performance liquid chromatographic method with double derivatization for the assay of urinary hydroxyproline.

An HPLC method with two derivatizations, the first with o-phthaldehyde in order to eliminate interferences due to some primary amino acids eluting with retention times similar to those of hydroxyproline and the second with dabsyl chloride, was developed and evaluated. Calibration graph linearity, influence of agitation and temperature on the preparation of the first derivative and the influence of the detection wavelength were assessed. The analysis time is shorter in comparison with other available methods, and therefore this method is suitable for laboratories that analyse both small and large series of samples.