Utility of solution electrochemistry mass spectrometry for investigating the formation and detection of biologically important conjugates of acetaminophen.
On-line formation and detection of glutathione and cysteine conjugates of acetaminophen were accomplished by the interfacing of a coulometric electrochemical cell with a thermospray mass spectrometer in a flow-injection experiment using a liquid chromatographic pump. Formation of the conjugates occurred only after acetaminophen was oxidized electrochemically by a two-electron transfer to N-acetyl-p-benzoquinoneimine and reacted in a mixing tee with either glutathione or cysteine. The newly formed conjugate was detected by thermospray mass spectrometry by observing the [M + H]+ ion for the acetaminophen-glutathione conjugate at m/z 457, or the [M + H]+ ion for the acetaminophen cysteine conjugate at m/z 271. Both the glutathione and cysteine conjugates produced a common fragment ion at m/z 184. The on-line reaction of glutathione and electrochemically generated N-acetyl-p-benzoquinoneimine was monitored at varying pH. At pH 8.5 the ion intensity for the acetaminophen-glutathione conjugate was greater than at lower pH, indicating that lower proton concentration enhanced the reaction of glutathione with N-acetyl-p-benzoquinoneimine. This on-line electrochemical-thermospray mass spectrometric method demonstrated that acetaminophen conjugates may be formed and detected in the time frame of 1 s.
National Center for Toxicological Research, Jefferson, AR 72079.
T A
TA
Getek
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WA
Korfmacher
T A
TA
McRae
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JA
Hinson
D000082:Acetaminophen / Q000378:metabolism*; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004563:Electrochemistry; D005978:Glutathione / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods*; D010084:Oxidation-Reduction
D000082:Acetaminophen; D005978:Glutathione