Effect of hydrolysis conditions on the determination of urinary 2,5-hexanedione in workers exposed or not exposed to N-hexane

Abstract

In order to determine the optimal conditions of acid hydrolysis for urinary 2,5-hexanedione (HD) measurement, the effects of urine pH or volume of HCl added to urine and hydrolysis period were studied using gas chromatography-mass spectrometry (GC-MS). When 0.3 ml of concentrated HCl was added to 3 ml of urine, complete hydrolysis to liberate HD was attained 2 h after the start of heating at 100 degrees C for urine from non-exposed subjects. On the contrary, in urine from exposed subjects most of HD was liberated in 30 min at 100 degrees C, suggesting that the urinary substrates converted to HD by acid hydrolysis were different between exposed and non-exposed subjects. It was confirmed that hydrolysis for 2 h at 100 degrees C with 0.3 ml of concentrated HCl added to 3 ml of urine gave the most reliable levels and the greatest amounts of HD in both urine from exposed and non-exposed subjects. Reference values were determined by the hydrolysis conditions presented here. In 84 males, 52 females, and 136 with sexes combined the urinary HD levels were 0.35, 0.49, and 0.39 mg/l (geometric mean), respectively. Urinary HD levels determined without hydrolysis (free HD) were less than 0.006 mg/l in 31 control subjects examined. In urine from exposed subjects the amount of free HD was about one-fifth of the total HD (free plus conjugated HD) as determined with acid hydrolysates, although the percentage of free to total HD varied from 0.4 to 28%.(ABSTRACT TRUNCATED AT 250 WORDS)