Development, validation and clinical evaluation of a dried urine spot method for determination of hippuric acid and creatinine

Abstract

ObjectivesThe purpose of this study was to develop and validate a HPLC–DAD method for determination of creatinine and hippuric acid in dried urine spots (DUS), evaluating its clinical applicability. Design and methodsSample preparation was based on a simple one step extraction with water. Analysis was performed in a reversed phase column Hypersil Gold® C18 (150×4.6mm, 3μm) with isocratic elution. Mobile phase was a mixture of triethylammonium phosphate buffer 5mM, pH3.3 and acetonitrile (90:10, v/v). ResultsTotal analytical run time was 6min. Precision assays presented CV% lower than 11.8%. Accuracy was 96.0–103%. The lower limit of quantitation was 50mgL−1. Mean extraction yields were 99% for CR and 79.3% for HA. Analytes were stable in DUS up to 11days at 40°C. The method was tested in 49 paired DUS and urine samples, with acceptable agreement. HA–CR concentration ratios in DUS were in the range of 0.06 to 2.74, being in average 100.7% from those values found in urine. ConclusionsDUS samples are a useful alternative for biological monitoring of toluene occupational exposure, especially in Developing Countries where sample logistics could be an important concern.