New technique for diagnosis and monitoring of alcaptonuria: quantification of homogentisic acid in urine with mid-infrared spectrometry Original Research Article

For the first time we used Fourier transform-infra-red (FT-IR) spectrometry in the investigation of an inborn error of metabolism, i.e. alcaptonuria, which expresses itself by the excretion of high concentrations of homogentisic acid in urine. The 200 homogentisic acid-spiked urine samples, ranging from 0 to 60 mmol l−1, were analysed in the mid-IR region using an attenuated total reflection accessory. Quantification is performed with partial least squares regression analysis. The standard errors of calibration, cross validation and prediction in the IR-predicted homogentisic acid concentration are 0.64, 0.78 and 0.79 mmol l−1, respectively, with R2>0.998. The 2σ detection limit is 2.0 mmol l−1 and the intra- and inter-assay coefficients of variation are below 5% for three concentration levels. The clear difference in the concentrations of homogentisic acid in the urine of four alcaptonuria patients compared to seven control samples demonstrates the potential suitability of IR analysis for the diagnosis and monitoring of alcaptonuria patients.