Application of LC–TOF MS to analysis of hemoglobin acetaldehyde adducts in alcohol detoxification patients

Abstract

Acetaldehyde adducts of hemoglobin have been regarded as potential biochemical markers of alcohol exposure. In this study a novel sensitive method using liquid chromatography coupled to time-of-flight mass spectrometry (LC-TOF MS) has been used to investigate changes in adduct levels in alcohol detoxification patients. Hemoglobin and authentic blood samples from 66 adults with an alcohol-dependence syndrome and from 12 children were analyzed for acetaldehyde modifications with and without trypsin digestion using LC-TOF MS. After in-vitro incubation of hemoglobin with increasing concentrations of acetaldehyde, followed by tryptic digestion, 21 modified peptide fragments could be identified from their accurate mass and retention time shift. Eight of these could also be detected in authentic human blood samples. Trace amounts in children's blood were indicative of an endogenous source. Modified peptide levels in patients' samples with and without ethanol were significantly different, as also were levels in samples from admission and from five days later. Samples obtained 5, 10, or 15 days after admission did not differ in adduct levels. The LC-TOF MS method was sensitive enough to detect acetaldehyde-modified hemoglobin peptides in blood samples from patients with an alcohol-dependence syndrome. However, elevated levels were only observed after recent ethanol consumption and decreased during five days of abstinence, suggesting that acetaldehyde-modified tryptic peptides of hemoglobin are potential biomarkers only for short-term ethanol ingestion.