Valentine J L, Middleton R
Department of Pediatrics, University of Arkansas for Medical Sciences and Arkansas Children's Hospital, Little Rock 72202-3591, USA.
J Anal Toxicol. 2000 Apr;24(3):211-22. doi: 10.1093/jat/24.3.211.
A method was developed that permitted rapid identification in urine of the following sympathomimetic amines: amphetamine, benzphetamine, cathinone, desmethylsegiline, diethylpropion, ephedrine, fenfluramine, mazindol, methylenedioxyamphetamine, methylenedioxyethylamphetamine, methylenedioxymethamphetamine, mescaline, methamphetamine, methcathinone, methylaminorex, methylphenidate, pemoline, phendimetrazine, phenylepherine, phentermine, phenylpropanolamine, pseudoephedrine, and selegiline. In addition, two alpha-phenylethylamine-like monoamine oxidase inhibitors, phenelizine and tranylcypromine, were studied. Those sympathomimetic amines containing a primary or secondary amine, a hydrazine, and/or hydroxyl (except mazindol) functional groups were derivatized effectively using an on-column derivatization technique that used a reagent consisting of 10% fluoroanhydride in hexane, whereas the other sympathomimetic amines, including mazindol, were analyzed underivatized. Three different fluoroanhydrides, trifluoroacetic (TFAA), pentafluoropropionic (PFPA), and heptafluorobutyric (HFBA), and three different injection-port temperatures (160, 200, and 260 degrees C) were investigated. Both TFAA and PFPA gave sympathomimetic amine derivatives with essentially identical retention times, whereas HFBA gave longer retention times and better separation of individual compounds. The base fragmentation ion was noted to increase 50 amu (CF2) for each derivatized sympathomimetic amine as the length of the carbon-fluorine chain increased. Fragmentation ion abundance was maximized at an injection-port temperature of 260 degrees C, and this enhanced sensitivity coupled with the better chromatographic resolution of the individual sympathomimetic amines prompted the selection of HFBA as the derivatizing agent of choice. Assignments were made for the fragmentation ions produced by each derivatized drug. The developed method was adapted to analyze urine specimens that might be encountered in emergency toxicology testing. For identification of sympathomimetic amines requiring derivatization, 0.1 mL of the patient specimen had amphetamine-d5 and methamphetamine-d5 added as internal standard followed by adjustment of pH to 9.3 with borate buffer, extraction with 9:1 chloroform/isopropanol, centrifugation and separation of the organic phase, addition of 10% methanolic HCI and evaporation under nitrogen, reconstitution with HFBA reagent, and on-column derivatization during gas chromatographic-mass spectrometric (GC-MS) analysis. For those sympathomimetic amines not requiring derivatization, 1.0 mL of urine specimen had diazepam-d5 added as internal standard followed by the same extraction procedure and reconstitution accomplished with ethyl acetate. Because precolumn derivatization was eliminated and only 8 min was required for GC-MS analysis, complete analysis time was approximately 30 min, making the method suitable for clinical emergency toxicology purposes.
已开发出一种方法，可快速鉴定尿液中的以下拟交感神经胺：苯丙胺、苄非他明、卡西酮、去甲基司来吉兰、二乙丙胺、麻黄碱、芬氟拉明、马吲哚、亚甲二氧基苯丙胺、亚甲二氧基乙基苯丙胺、亚甲二氧基甲基苯丙胺、三甲氧苯乙胺、甲基苯丙胺、甲卡西酮、甲基氨基苯丙胺、哌醋甲酯、匹莫林、苯双甲吗啉、去氧肾上腺素、苯丁胺、苯丙醇胺、伪麻黄碱和司来吉兰。此外，还研究了两种α-苯乙胺样单胺氧化酶抑制剂，苯乙肼和反苯环丙胺。那些含有伯胺或仲胺、肼基和/或羟基（马吲哚除外）官能团的拟交感神经胺，使用柱上衍生化技术进行有效衍生化，该技术使用由10%己烷中的氟酐组成的试剂，而其他拟交感神经胺，包括马吲哚，则在未衍生化的情况下进行分析。研究了三种不同的氟酐，三氟乙酸（TFAA）、五氟丙酸（PFPA）和七氟丁酸（HFBA），以及三种不同的进样口温度（160、200和260℃）。TFAA和PFPA得到的拟交感神经胺衍生物的保留时间基本相同，而HFBA得到的保留时间更长，各化合物的分离效果更好。随着碳氟链长度的增加，每个衍生化拟交感神经胺的基峰碎裂离子增加50 amu（CF2）。在进样口温度为260℃时，碎裂离子丰度最大，这种增强的灵敏度与各拟交感神经胺更好的色谱分辨率相结合，促使选择HFBA作为首选衍生化试剂。对每种衍生化药物产生的碎裂离子进行了归属。所开发的方法适用于分析急诊毒理学检测中可能遇到的尿液标本。对于需要衍生化的拟交感神经胺的鉴定，向0.1 mL患者标本中加入苯丙胺-d5和甲基苯丙胺-d5作为内标，然后用硼酸盐缓冲液将pH值调至9.3，用9:1氯仿/异丙醇萃取，离心并分离有机相，加入10%甲醇盐酸并在氮气下蒸发，用HFBA试剂复溶，并在气相色谱-质谱（GC-MS）分析过程中进行柱上衍生化。对于那些不需要衍生化的拟交感神经胺，向1.0 mL尿液标本中加入地西泮-d5作为内标，然后进行相同的萃取程序，并用乙酸乙酯复溶。由于消除了柱前衍生化，GC-MS分析仅需8分钟，因此完整的分析时间约为30分钟，使该方法适用于临床急诊毒理学目的。
