Quantification of o,o'-dityrosine, o-nitrotyrosine, and o-tyrosine in cat urine samples by LC/ electrospray ionization-MS/MS using isotope dilution.
Quantification of o-tyrosine, o-nitrotyrosine, and o,o'dityrosine from cat urine samples was achieved by LC/ electrospray ionization-MS/MS (LC/ESI-MS/MS) using an isotope dilution technique in multiple reaction monitoring mode before butylation of o,o'-dityrosine and after butylation of o-tyrosine and o-nitrotyrosine. This novel approach of amino acids butylation enhanced the MS response by a factor of 7-fold for o-tyrosine and 6-fold for o-nitrotyrosine and decreased the overall chemical background noise. Butylation of o,o'-dityrosine resulted in a lower MS response as a result of the formation of both mono- and doubly butylated species. The mean recovery for the oxidized amino acids was estimated at 73 +/- 2%. The limits of quantitation of NO2-Tyr butyl ester, o-Tyr butyl ester, and di-Tyr in cat urine samples were calculated at 14.5, 28.2 and 140.9 nM, respectively. The oxidized amino acids levels in cat urine extracts ranged from 157 to 250 ng/day for o-Tyr and from 3,289 to 11,803 ng/day for di-Tyr. NO2-Tyr was found in only two urine extracts at levels below 58 ng/day. A certain trend of correlation was observed between o,o'-dityrosine and o-tyrosine when comparing these values against their respective creatinine amounts. A comparison of the data gathered from the ThermoFinnigan TSQ 7000 and Micromass Q-TOF instruments revealed several advantages of using the Q-TOF regarding the exact mass measurement, a lower ion suppression effect and the possibility to perform analyses in full scan product ion mode. These results demonstrate that a Q-TOF instrument can be a good alternative to classical triple quadrupole for quantitative purposes on a relatively small linear dynamic range (4 orders of magnitude for the Q-TOF, as compared to 6 for the triple quadrupole).
Marvin
Laure Féline
LF
Nestlé Research Center, Nestec Ltd., Vers-Chez-Les-Blanc, 1000 Lausanne 26, Switzerland.
Delatour
Thierry
T
Tavazzi
Isabelle
I
Fay
Laurent Bernard
LB
Cupp
Carolyn
C
Guy
Philippe Alexandre
PA
D000818:Animals; D002415:Cats; D002851:Chromatography, High Pressure Liquid; D007554:Isotopes; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation* / Q000592:standards; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000652:urine*
D007554:Isotopes; C002744:3-nitrotyrosine; D014443:Tyrosine; C007543:dityrosine