title: In-source formation of N-acetyl-p-benzoquinone imine (NAPQI), the putatively toxic acetaminophen (paracetamol) metabolite, after derivatization with pentafluorobenzyl bromide and GC-ECNICI-MS analysis.
abstract: Pentafluorobenzyl (PFB) bromide (PFB-Br) is a versatile derivatization reagent for numerous classes of compounds. Under electron-capture negative-ion chemical ionization (ECNICI) conditions PFB derivatives of acidic compounds readily and abundantly ionize to produce intense anions due to [M-PFB](-). In the present article we investigated the PFB-Br derivatization of unlabelled acetaminophen (N-acetyl-p-aminophenol, NAPAP-d(0); paracetamol; MW 151) and tetradeuterated acetaminophen (NAPAP-d(4); MW 155) in anhydrous acetonitrile and their GC-ECNICI-MS behavior using methane as the buffer gas. In addition to the expected anions [M-PFB](-) at m/z 150 from NAPAP-d(0) and m/z 154 from NAPAP-d(4), we observed highly reproducibly almost equally intense anions at m/z 149 and m/z 153, respectively. Selected ion monitoring of these ions is suitable for specific and sensitive quantification of acetaminophen in human plasma and urine. Detailed investigations suggest in-source formation of N-acetyl-p-benzoquinone imine (NAPQI; MW 149), the putatively toxic acetaminophen metabolite, from the PFB ether derivative of NAPAP. GC-ECNICI-MS of non-derivatized NAPAP did not produce NAPQI. The peak area ratio of m/z 149 to m/z 150 and of m/z 153 to m/z 154 decreased with increasing ion-source temperature in the range 100-250°C. Most likely, NAPQI formed in the ion-source captures secondary electrons to become negatively charged (i.e., [NAPQI](-)) and thus detectable. Formation of NAPQI was not observed under electron ionization (EI) conditions, i.e., by GC-EI-MS, from derivatized and non-derivatized NAPAP. NAPQI was not detectable in flow injection analysis LC-MS of native NAPAP in positive electrospray ionization (ESI) mode, whereas in negative ESI mode low extent NAPQI formation was observed (<5%). Our results suggest that oxidation of drug derivatives in the ion-sources of mass spectrometers may form intermediates that are produced from activated drugs in enzyme-catalyzed reactions.
authors.affiliation: Institute of Clinical Pharmacology, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany. tsikas.dimitros@mh-hannover.de
authors.forename: Dimitrios
authors.initials: D
authors.lastname: Tsikas
authors.forename: Arne
authors.initials: A
authors.lastname: Trettin
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Zörner
authors.forename: Frank-Mathias
authors.initials: FM
authors.lastname: Gutzki
mesh_terms: D000082:Acetaminophen / Q000737:chemistry*; D000097:Acetonitriles; D018712:Analgesics, Non-Narcotic / Q000737:chemistry*; D016227:Benzoquinones / Q000737:chemistry*; D005464:Fluorobenzenes / Q000737:chemistry*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D007097:Imines / Q000737:chemistry*; D008697:Methane; D005466:Fluorocarbons
chemical_list: D000082:Acetaminophen; D000097:Acetonitriles; D018712:Analgesics, Non-Narcotic; D016227:Benzoquinones; D005464:Fluorobenzenes; D007097:Imines; D008697:Methane; C025063:pentafluorobenzyl bromide; D005466:Fluorocarbons; C028473:N-acetyl-4-benzoquinoneimine; C032159:acetonitrile