Bianco Giuliana, Abate Salvatore, Labella Cristiana, Cataldi Tommaso R I
Dipartimento di Chimica, Università degli Studi della Basilicata, Via N. Sauro 85, 85100 Potenza, Italy.
Rapid Commun Mass Spectrom. 2009 Apr;23(7):1065-74. doi: 10.1002/rcm.3969.
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.
采用液相色谱(LC)与正离子电喷雾电离(ESI+)联用,结合混合四极杆线性离子阱(LTQ)和傅里叶变换离子回旋共振质谱(FTICRMS),在一次色谱运行中同时测定人尿中的咖啡因及其代谢物。LC/ESI-FTICRMS能够明确测定所研究化合物的分子量,而不受其他生物分子的干扰。通过对质子化分子M+H进行源内离子阱碰撞诱导解离(CID),利用串联质谱(MS/MS)对咖啡因及其十四种代谢物的质谱行为进行了系统而全面的研究。在CID过程中观察到的主要裂解途径是逆狄尔斯-阿尔德(RDA)过程以及环收缩反应。黄嘌呤前体的基峰源于甲基异氰酸酯(CH(3)NCO,57 Da)或异氰酸(HNCO,43 Da)的损失,它们继而再损失一个CO单元。尿酸衍生物也有一个共同的裂解过程,即RDA重排以及连续损失CO(2)或CO。尿嘧啶衍生物显示质子化分子损失一个乙烯酮单元(CH(2)CO,42 Da),同时损失H(2)O或CO。为评估本方法的潜力,通过摄入咖啡因后健康人尿样中一些已鉴定的代谢物,对用于测量P450 CYP1A2、N-乙酰转移酶和黄嘌呤氧化酶活性的三种既定代谢物比率进行了评估。