Analysis of N(ε) -ethyllysine in human plasma proteins by gas chromatography-negative ion chemical ionization/mass spectrometry as a biomarker for exposure to acetaldehyde and alcohol.
BACKGROUND
N(ε) -ethyllysine (NEL) is a major stable adduct formed by the reaction of acetaldehyde (AA) with lysine residues in proteins. However, its occurrence and levels in biological specimens and its relationship with AA/alcohol exposure-associated disorders have not been fully elucidated. In this study, we have developed a sensitive and specific method to quantitate NEL levels in human plasma proteins.


METHODS
The method consists of (1) purification of the protein fraction of interest by Sephadex G-15 to remove low molecular substances, (2) hydrolysis of proteins with Pronase E in the presence of stable isotope-labeled internal standards, (3) derivatization of amino acids with pentafluorobenzyl (PFB) bromide, and (4) quantification of the PFB derivatives of NEL and l-lysine using gas chromatography-negative ion chemical ionization/mass spectrometry in a selected ion monitoring mode.


RESULTS
Using the above method, the NEL levels in human plasma proteins obtained from 10 each of control subjects and alcoholic patients were measured. NEL was detected in all samples analyzed, the average level of NEL in the plasma proteins of alcoholic patients (1.17 ± 0.36 NEL/1,000 l-lysine) being significantly higher than that of control subjects (0.26 ± 0.07 NEL/1,000 l-lysine).


CONCLUSIONS
The method could be applied to molecular epidemiological studies to investigate possible associations between the NEL levels in human tissue proteins and human diseases associated with exposure to AA and alcohol.
Graduate School of Nutritional and Environmental Sciences and Global COE program, University of Shizuoka, Shizuoka, Japan.
Ryota
R
Mabuchi
Aya
A
Kurita
Noriyuki
N
Miyoshi
Akira
A
Yokoyama
Takumi
T
Furuta
Toshinao
T
Goda
Yoshihide
Y
Suwa
Toshiyuki
T
Kan
Takashi
T
Amagai
Hiroshi
H
Ohshima
D000079:Acetaldehyde / Q000097:blood*; D000328:Adult; D000368:Aged; D000437:Alcoholism / Q000097:blood* / Q000175:diagnosis; D015415:Biomarkers; D002247:Carbon Isotopes; D016022:Case-Control Studies; D002492:Central Nervous System Depressants / Q000097:blood*; D000431:Ethanol / Q000097:blood*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D007553:Isotope Labeling; D008239:Lysine / Q000031:analogs & derivatives* / Q000032:analysis; D008297:Male; D008875:Middle Aged; D009587:Nitrogen Isotopes
D015415:Biomarkers; D002247:Carbon Isotopes; D002492:Central Nervous System Depressants; D009587:Nitrogen Isotopes; C117804:ethyl lysine; D000431:Ethanol; D000079:Acetaldehyde; D008239:Lysine