Rapid biomonitoring of heterocyclic aromatic amines in human urine by tandem solvent solid phase extraction liquid chromatography electrospray ionization mass spectrometry.
A rapid and facile tandem solvent solid phase extraction method was established to isolate the heterocyclic aromatic amines (HAAs) 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, and 2-amino-9H-pyrido[2,3-b]indole from urine. The HAAs were separated by reversed phase liquid chromatography and quantified by electrospray ionization tandem mass spectrometry (ESI/MS/MS) using selected reaction monitoring. The limits of detection and quantitation of these HAAs approached 1-3 and 2-8 pg/mL, respectively, using only 0.3 mL of urine for analysis. Full product ion spectra were acquired to corroborate analyte identities. The pretreatment of urine from human volunteers that had consumed a grilled beef meal with acid or base at 70 degrees C increased the concentration of HAAs by as much as 6-fold, indicating the presence of phase II conjugates of the parent compounds. HAAs containing an N-methylimidazole moiety undergo facile cleavage of the N-methyl group under collision-induced dissociation conditions, and MS/MS analysis in the constant neutral loss scan mode monitoring the transition [M + H](+) --> [M + H - CH(3)(*)](+) revealed the presence of two other HAAs. 2-Amino-3-methylimidazo[4,5-f]quinoxaline (IQx) was identified by coelution of the analyte with synthetic IQx and by acquisition of the product ion spectrum. The second HAA was present in a relatively high abundance in urine. The molecule had the same nominal mass as 8-MeIQx (MH(+) at m/z 214), and the product ion spectrum was similar to that of 8-MeIQx. This novel HAA was also found in the grilled meat consumed by the volunteers at a concentration of 8 parts per billion. The accurate mass measurement and product ion spectrum of this molecule by ESI quadrupole time-of-flight mass spectrometry revealed that it was an isomer of 8-MeIQx. This tandem solvent solid phase extraction LC/ESI/MS/MS procedure may be used to rapidly assess the daily exposure to a variety of HAAs in urine.
Holland
Ricky D
RD
Division of Chemistry, National Center for Toxicological Research, 3900 NCTR Road, Jefferson, Arkansas 72079, USA.
Taylor
Jason
J
Schoenbachler
Laura
L
Jones
Richard C
RC
Freeman
James P
JP
Miller
Dwight W
DW
Lake
Brian G
BG
Gooderham
Nigel J
NJ
Turesky
Robert J
RJ
D000588:Amines / Q000652:urine*; D000818:Animals; D002243:Carbolines / Q000652:urine; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods*; D005502:Food / Q000633:toxicity; D006571:Heterocyclic Compounds / Q000009:adverse effects / Q000652:urine; D006801:Humans; D008297:Male; D011084:Polycyclic Aromatic Hydrocarbons / Q000009:adverse effects / Q000652:urine*; D011810:Quinoxalines / Q000652:urine; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D000588:Amines; D002243:Carbolines; D006571:Heterocyclic Compounds; D011084:Polycyclic Aromatic Hydrocarbons; D011810:Quinoxalines; C036990:2-amino-3,8-dimethylimidazo(4,5-f)quinoxaline; C017822:2-amino-9H-pyrido(2,3-b)indole