Identification and fragmentation pathways of caffeine metabolites in urine samples via liquid chromatography with positive electrospray ionization coupled to a hybrid quadrupole linear ion trap (LTQ) and Fourier transform ion cyclotron resonance mass spectrometry and tandem mass spectrometry

Abstract

Liquid chromatography (LC) with positive ion electrospray ionization (ESI+) coupled to a hybrid quadrupole linear ion trap (LTQ) and Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) was employed for the simultaneous determination of caffeine and its metabolites in human urine within a single chromatographic run. LC/ESI-FTICRMS led to the unambiguous determination of the molecular masses of the studied compounds without interference from other biomolecules. A systematic and comprehensive study of the mass spectral behaviour of caffeine and its fourteen metabolites by tandem mass spectrometry (MS/MS) was performed, through in-source ion trap collision-induced dissociation (CID) of the protonated molecules, [M+H](+). A retro-Diels-Alder (RDA) process along with ring-contraction reactions were the major fragmentation pathways observed during CID. The base peak of xanthine precursors originates from the loss of methyl isocyanate (CH(3)NCO, 57 Da) or isocyanic acid (HNCO, 43 Da), which in turn lose a CO unit. Also uric acid derivatives shared a RDA rearrangement as a common fragmentation process and a successive loss of CO(2) or CO. The uracil derivatives showed a loss of a ketene unit (CH(2)CO, 42 Da) from the protonated molecule along with the loss of H(2)O or CO. To assess the potential of the present method three established metabolite ratios to measure P450 CYP1A2, N-acetyltransferase and xanthine oxidase activities were evaluated by a number of identified metabolites from healthy human urine samples after caffeine intake.

Metabolite Profiling of Bendamustine in Urine of Cancer Patients after Administration of [14C]Bendamustine

“…The proposed elemental composition of the conjugate was C 5 H 4 N 4 O 3 (168.0278 Da; ⌬ppm ϭ Ϫ.57), which corresponds with the endogenous compound uric acid. Neutral loss of 43 Da (HCON), followed by loss of 28 Da (CO), is frequently observed for uric acid derivatives (Bianco et al, 2009) and also occurred for Met11, supporting the proposed identification of this compound as a uric acid conjugate of didechlorinated bendamustine.…”

Collision‐induced dissociation studies of caffeine in positive electrospray ionisation mass spectrometry using six deuterated isotopomers and one N1‐ethylated homologue

“…This fragment arises directly from the parent substance a by a neutral loss of methyl isocyanate (CH 3 NCO). In the literature, the formation of this fragment is explained by a retro‐Diels‐Alder reaction and the resulting molecule is consequently presented as an open‐chain structure . Taking into account the calculated empirical formula C 6 H 7 N 3 O (Table ) and the calculated number of ‘double‐bond equivalents’ a cyclic β‐lactam structure for fragment c seems to be feasible .…”

Investigation on fragmentation pathways of rutaecarpine and its two derivatives using electrospray ionization ion‐trap time‐of‐flight tandem mass spectrometry

“…The MS 2 spectrum from protonated molecule 1, m/z 288, consists of major fragment ions at m/z 273, 243, 167 and 142. Most interesting is the main product ion at m/z 273 corresponding to the loss of methyl radical, which has been reported for the fragmentation of some other similar structures 27,28. The loss of this methyl radical could be explained by the initial cleavage of the nonconjugated covalent bond at the C-7-N-6 position and transfer of a hydrogen radical to give an intermediate ion with a new double bond and a methyl radical.…”