VALIDATION OF A GC-FID METHODOLOGY FOR THE DETERMINATION OF 2,5-HEXANEDIONE IN URINE

Abstract

2,5-hexanedione is the main metabolite of hexane and methyl butyl ketone excreted into urine and is currently used to estimate the human exposure levels to these solvents in professional environments. In Brazil, occupational exposure is regulated by a national control program (PCMSO) and applied to public and private organizations that hire your employees according to the actual Brazilian legislation. In the present work, we validated a methodology based on a gas chromatography−flame ionization detector (GC-FID) for the quantification of 2,5-hexanedione in urine to attend the new occupational limit of urinary 2,5-hexanedione, established by a recent revision of the Brazilian legislation, which reduced this limit from 5.0 to 0.5 mg/L. Before GC-FID analysis, sample treatment was based on a simple liquid-liquid extraction with dichloromethane. Considering that there is no specific validation guide for occupational purposes, the adopted validation process was performed using a “fit-for-purpose” approach based on different guides, including FDA bioanalytical method validation, European Commission Decision 2002-657-EC, and ANVISA Guidelines RDC 166/2017. After method optimization, the performance characteristics determined by the present validation study were considered satisfactory and demonstrate the method suitability for the routine analysis in the Brazilian PCMSO, including CCα and CCβ values, which are used to avoid the presentation of false-positive and false-negative results. Intra and interday reproducibility were below 2.66, and 4.08 % and analyte recoveries were above 95.4% for three evaluated levels. The limits of detection and quantification were 0.05 and 0.17, respectively. To our knowledge, this is the first application of CCα and CCβ approaches in an analytical method intended for occupational purposes, which are important to know when a sample is out or following the legislation. Then real samples were analyzed herein, and none presented 2,5-hexanedione above the required limits.