A pre-column derivatization method for the sensitive determination of amines using a labeling reagent 2-(11H-benzo[a]-carbazol-11-yl) ethyl chloroformate (BCEC-Cl) followed by high-performance liquid chromatography with fluorescence detection has been developed. Identification of derivatives was carried out by LC/APCI/MS in positive-ion mode. The chromophore of 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC-Cl) reagent was replaced by 2-(11H-benzo[a]-carbazol-11-yl) ethyl functional group, which resulted in a sensitive fluorescence derivatizing reagent BCEC-Cl. BCEC-Cl could easily and quickly label amines. Derivatives were stable enough to be efficiently analyzed by HPLC and showed an intense protonated molecular ion corresponding m/z [M+H](+) under APCI/MS in positive-ion mode. The collision-induced dissociation of the protonated molecular ion formed characteristic fragment ions at m/z 261.8 and m/z 243.8 corresponding to the cleavages of CH(2)O-CO and CH(2)-OCO bonds. Studies on derivatization demonstrated excellent derivative yields over the pH 9.0-10.0. Maximal yields close to 100% were observed with three- to four-fold molar reagent excess. In addition, the detection responses for BCEC-derivatives were compared to those obtained using 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC-Cl) and 9-fluorenyl methylchloroformate (FMOC-Cl) as labeling reagents. The ratios I(BCEC)/I(BCEOC)=1.94-2.17 and I(BCEC)/I(FMOC)=1.04-2.19 for fluorescent (FL) responses (here, I was relative fluorescence intensity). Separation of the derivatized amines had been optimized on reversed-phase Eclipse XDB-C(8) column. Detection limits calculated from 0.50pmol injection, at a signal-to-noise ratio of 3, were 1.77-14.4fmol. The relative standard deviations for within-day determination (n=11) were 1.84-2.89% for the tested amines. The mean intra- and inter-assay precision for all amines levels were <3.64% and 2.52%, respectively. The mean recoveries ranged from 96.6% to 107.1% with their standard deviations in the range of 0.8-2.7. Excellent linear responses were observed with coefficients of >0.9996.

中文翻译：

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HPLC-荧光检测和APCI / MS鉴定使用2-（11H-苯并[a]咔唑-11-基）氯甲酸乙酯（BCEC-Cl）作为标记试剂测定胺。

一种使用标记试剂2-（11H-苯并[a]-咔唑-11-基）氯甲酸乙酯（BCEC-Cl）进行灵敏测定胺的柱前衍生化方法，然后采用具有荧光检测功能的高效液相色谱法已开发。通过LC / APCI / MS以阳离子模式进行衍生物鉴定。1,2-苯并-3,4-二氢咔唑-9-氯甲酸乙酯（BCEOC-Cl）试剂的发色团被2-（11H-苯并[a]-咔唑-11-基）乙基官能团取代，得到在灵敏的荧光衍生试剂BCEC-Cl中。BCEC-Cl可以轻松快速地标记胺。衍生物足够稳定，可以通过HPLC有效分析，并且在APCI / MS下在正离子模式下显示出与m / z [M + H]（+）相对应的强质子化分子离子。质子化的分子离子的碰撞诱导解离在m / z 261.8和m / z 243.8处形成特征碎片离子，对应于CH（2）O-CO和CH（2）-OCO键的裂解。衍生化研究表明，在pH 9.0-10.0范围内具有出色的衍生物产率。当摩尔试剂过量三到四倍时，观察到最大收率接近100％。此外，将BCEC衍生物的检测响应与使用1,2-苯并3,4-二氢咔唑-9-氯甲酸乙酯（BCEOC-Cl）和9-芴基甲基氯甲酸酯（FMOC-Cl）的检测响应进行了比较。 。荧光（FL）响应的比率I（BCEC）/ I（BCEOC）= 1.94-2.17和I（BCEC）/ I（FMOC）= 1.04-2.19（此处，I为相对荧光强度）。在反相Eclipse XDB-C（8）色谱柱上已优化了衍生胺的分离。从0.50pmol进样计算出的信噪比为3时，检测限为1.77-14.4fmol。对于测试的胺，日内测定的相对标准偏差（n = 11）为1.84-2.89％。所有胺水平的平均测定内和测定间精密度分别为<3.64％和2.52％。平均回收率从96.6％到107.1％，标准偏差在0.8-2.7范围内。观察到极好的线性响应，系数> 0.9996。1％，标准偏差在0.8-2.7范围内。观察到极好的线性响应，系数> 0.9996。1％，标准偏差在0.8-2.7范围内。观察到极好的线性响应，系数> 0.9996。