Multiwavelength analyses of derivative spectra: An application for the determination of urinary porphyrins

Abstract

Quantification of excreted poiphyrins is of value in the diagnosis of the various disorders of po~hy~n metabolism. However, by far the greatest use of po~hy~n testing is to exclude porphyria as the cause of an unexplained illness. For the purpose of excluding porphyria, the determination of urinary porphyrins can be considered a ‘screening’ procedure since this type of testing is most often done on urine samples that are essentially normal. For tests done on normal samples, the more sophisticated chromatographic methods [l-3] for porphyrin analyses are not warranted. Jones and Sweeney used first f4] and second-derivative [5] spectra obtained from acidified urine samples for the se~quantitative dete~nation of urinary po~hy~ns. They found that the wavelength where the second derivative maxima occurred correlated to the relative proportion of coproporphyrin and uroporphyrin in a mixture. They used the height of the peak to estimate the concentration of total porphyrin present. We have improved this method of analysis by employing curve-fitting of multiwavelength derivative data for the quantitation of coproporphyrin and uroporphyrin in the mixture. Sternberg et al [6] described a method for the cute-fitting of multiwaveleng~ absorbance data to quantitate indi~dual components of a complex mixture. Weiser and Pardue [7] extended this method of multiwavelength analysis to quantitate data from derivatives of absorbance spectra. They found that the precision for measuring methemoglobin progressively improved as the computations were applied first to absorbance, then to first-derivative, and finally to second-derivative data. The objective of our study was to evaluate the method of multiwavelength analysis for the quantitative determination of coproporphyrin and uroporphyrin in a complex mixture, such as acidified urine.