Lv Chunxiao, Li Qing, Liu Xujia, He Bosai, Sui Zhenyu, Xu Huarong, Yin Yidi, Liu Ran, Bi Kaishun
School of Pharmacy, Shenyang Pharmaceutical University, Shenyang, 110016, China; National and Local United Engineering Laboratory for Key Technology of Chinese Material Medica Quality Control, Shenyang Pharmaceutical University, Shenyang, 110016, China.
J Mass Spectrom. 2015 Feb;50(2):354-63. doi: 10.1002/jms.3536.
In order to investigate the potential links between catecholamines (CAs) and Alzheimer's disease (AD), rapid and sensitive ultra-performance liquid chromatography (UPLC)-tandem mass spectrometry (MS/MS) methods in different ionization modes for the quantification of 14 CAs and their metabolites in rat urine without derivatization or complex sample pre-treatments were developed. After addition of the internal standard, isoproterenol, the urine samples were extracted by protein precipitation and separated on an Inertsil ODS-EP column (Shimadzu, Japan) at a flow of 1.0 ml min(-1). Tandem mass spectrometric detection was performed on a 4000Q UPLC-MS/MS in the multiple reaction monitoring mode with turbo ion spray source. Tyrosine, dopamine, noradrenaline, epinephrine, 3-methoxytyramine, normetanephrine and metanephrine were determined in positive mode, while 3,4-dihyroxy-L-phenylalanine (DOPA), 3,4-dihydroxyphenylacetic acid, DL-3,4-dihydroxymandelic acid, DL-3,4-dihydroxyphenyl glycol, homovanillic acid, DL-4-hydroxy-3-methoxymandelic acid and 4-hydroxy-3-methoxy-phenylglycol were determined in negative mode. The methods were examined and were found to be precise and accurate within the linearity range of the assays. The intra-day and inter-day precision and accuracy of the analytes were well within acceptance criteria (±15%). The mean extraction recoveries of analytes and internal standard were all more than 60%. The validated methods have been successfully applied to compare CAs profiles in normal and AD rats. The results indicated the urine levels of DL-3,4-dihydroxyphenyl glycol and 4-hydroxy-3-methoxy-phenylglycol in AD rats were significantly higher than those in the normal group, and the other CAs have an opposite performance. These may attribute to the difference of some enzyme activity between rats with AD and normal. Furthermore, this may be helpful in clinical diagnostics and monitor the efficacy of AD treatment.
为了研究儿茶酚胺（CAs）与阿尔茨海默病（AD）之间的潜在联系，开发了无需衍生化或复杂样品预处理即可在不同电离模式下快速灵敏地定量大鼠尿液中14种儿茶酚胺及其代谢物的超高效液相色谱（UPLC）-串联质谱（MS/MS）方法。加入内标异丙肾上腺素后，通过蛋白沉淀法提取尿液样品，并在Inertsil ODS-EP柱（日本岛津）上以1.0 ml min⁻¹的流速进行分离。在配备涡轮离子喷雾源的4000Q UPLC-MS/MS上以多反应监测模式进行串联质谱检测。酪氨酸、多巴胺、去甲肾上腺素、肾上腺素、3-甲氧基酪胺、去甲变肾上腺素和变肾上腺素以正模式测定，而3,4-二羟基-L-苯丙氨酸（DOPA）、3,4-二羟基苯乙酸、DL-3,4-二羟基扁桃酸、DL-3,4-二羟基苯乙二醇、高香草酸、DL-4-羟基-3-甲氧基扁桃酸和4-羟基-3-甲氧基苯乙二醇以负模式测定。对这些方法进行了检测，发现在测定的线性范围内精确且准确。分析物的日内和日间精密度和准确度均在可接受标准（±15%）范围内。分析物和内标的平均提取回收率均超过60%。经过验证的方法已成功应用于比较正常大鼠和AD大鼠的儿茶酚胺谱。结果表明，AD大鼠尿液中DL-3,4-二羟基苯乙二醇和4-羟基-3-甲氧基苯乙二醇的水平显著高于正常组，而其他儿茶酚胺表现相反。这些可能归因于AD大鼠和正常大鼠之间某些酶活性的差异。此外，这可能有助于临床诊断和监测AD治疗的疗效。
