Analysis of aflatoxin B1-lysine adduct in serum using isotope-dilution liquid chromatography/tandem mass spectrometry.
A method for quantitative analysis of aflatoxin B1-lysine adduct (B1-Lys) in serum by liquid chromatography using tandem mass spectrometry (LC/MS/MS) is presented. The protein in a 250-microL sample was digested in the presence of a stable-isotope internal standard during a 4-h incubation at 37 degrees C with Pronasetrade mark. B1-Lys and the internal standard were extracted using mixed-mode solid-phase extraction cartridges and eluted with 2% formic acid in methanol. Following evaporation and reconstitution, extracts were injected onto a Luna C-18(2) column and eluted with a step gradient of acetonitrile and 0.06% formic acid. The B1-Lys and the internal standard were detected in a positive ionization selective reaction monitoring mode with a ThermoFinnigan TSQ Quantum triple quadrupole mass spectrometer. Calibration curves were linear for concentrations from 0.05-8.0 ng/mL. The method was validated with aflatoxin B1 dosed rat serum diluted to anticipated high and low concentrations. Total imprecision determined from 30 measurements over 15 days was 5.6% and 9.1%, respectively. Recoveries of 78.8 +/- 6.4% for B1-Lys and 85.4 +/- 12.4% for the internal standard were based on the full extraction and reconstitution processes. The method can be used to quantitate B1-Lys at the 0.5 pg/mg albumin level and is suitable for routine analysis.
McCoy
Leslie F
LF
Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA 30341, USA.
Scholl
Peter F
PF
Schleicher
Rosemary L
RL
Groopman
John D
JD
Powers
Carissa D
CD
Pfeiffer
Christine M
CM
D016604:Aflatoxin B1 / Q000097:blood* / Q000302:isolation & purification; D000818:Animals; D002138:Calibration; D002853:Chromatography, Liquid; D006801:Humans; D007554:Isotopes; D007700:Kinetics; D008239:Lysine / Q000097:blood* / Q000302:isolation & purification; D011402:Pronase / Q000378:metabolism; D051381:Rats; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization
D007554:Isotopes; C066451:aflatoxin B1-lysine adduct; D016604:Aflatoxin B1; D011402:Pronase; D008239:Lysine