Swiss Laboratory for Doping Analyses, University Center of Legal Medecine, Geneva and Lausanne, Centre Hospitalier Universitaire Vaudois and University of Lausanne, Ch. des Croisettes 22, 1066 Epalinges, Switzerland.
J Chromatogr A. 2010 Nov 26;1217(48):7528-38. doi: 10.1016/j.chroma.2010.10.005. Epub 2010 Oct 25.
Consumption of nicotine in the form of smokeless tobacco (snus, snuff, chewing tobacco) or nicotine-containing medication (gum, patch) may benefit sport practice. Indeed, use of snus seems to be a growing trend and investigating nicotine consumption amongst professional athletes is of major interest to sport authorities. Thus, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the detection and quantification of nicotine and its principal metabolites cotinine, trans-3-hydroxycotinine, nicotine-N'-oxide and cotinine-N-oxide in urine was developed. Sample preparation was performed by liquid-liquid extraction followed by hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-MS/MS) operated in electrospray positive ionization (ESI) mode with selective reaction monitoring (SRM) data acquisition. The method was validated and calibration curves were linear over the selected concentration ranges of 10-10,000 ng/mL for nicotine, cotinine, trans-3-hydroxycotinine and 10-5000 ng/mL for nicotine-N'-oxide and cotinine-N-oxide, with calculated coefficients of determination (R(2)) greater than 0.95. The total extraction efficiency (%) was concentration dependent and ranged between 70.4 and 100.4%. The lower limit of quantification (LLOQ) for all analytes was 10 ng/mL. Repeatability and intermediate precision were ≤9.4 and ≤9.9%, respectively. In order to measure the prevalence of nicotine exposure during the 2009 Ice Hockey World Championships, 72 samples were collected and analyzed after the minimum of 3 months storage period and complete removal of identification means as required by the 2009 International Standards for Laboratories (ISL). Nicotine and/or metabolites were detected in every urine sample, while concentration measurements indicated an exposure within the last 3 days for eight specimens out of ten. Concentrations of nicotine, cotinine, trans-3-hydroxycotinine, nicotine-N'-oxide and cotinine-N-oxide were found to range between 11 and 19,750, 13 and 10,475, 10 and 8217, 11 and 3396, and 13 and 1640 ng/mL, respectively. When proposing conservative concentration limits for nicotine consumption prior and/or during the games (50 ng/mL for nicotine, cotinine and trans-3-hydroxycotinine and 25 ng/mL for nicotine-N'-oxide and cotinine-N-oxide), about half of the hockey players were qualified as consumers. These findings significantly support the likelihood of extensive smokeless nicotine consumption. However, since such conclusions can only be hypothesized, the potential use of smokeless tobacco as a doping agent in ice hockey requires further investigation.
以无烟烟草（鼻烟、鼻烟、咀嚼烟草）或含尼古丁药物（口香糖、贴剂）形式摄入尼古丁可能有益于体育实践。事实上，鼻烟的使用似乎是一种正在增长的趋势，调查职业运动员中尼古丁的使用是体育当局的主要关注点。因此，开发了一种用于检测和定量尿液中尼古丁及其主要代谢物可替宁、反式-3-羟基可替宁、尼古丁-N'-氧化物和可替宁-N-氧化物的液相色谱-串联质谱（LC-MS/MS）方法。样品制备通过液液萃取进行，然后通过亲水相互作用色谱-串联质谱（HILIC-MS/MS）进行分析，采用电喷雾正电离（ESI）模式进行操作，并采用选择性反应监测（SRM）数据采集。该方法经过验证，在选定的浓度范围内，尼古丁、可替宁、反式-3-羟基可替宁的校准曲线呈线性，范围为 10-10,000ng/mL，尼古丁-N'-氧化物和可替宁-N-氧化物的浓度为 10-5,000ng/mL，计算得出的决定系数（R(2)）大于 0.95。总提取效率（%）与浓度有关，范围在 70.4%至 100.4%之间。所有分析物的定量下限（LLOQ）均为 10ng/mL。重复性和中间精密度分别为≤9.4%和≤9.9%。为了测量 2009 年冰球世界锦标赛期间尼古丁暴露的流行程度，在经过 3 个月的储存期和完全去除 2009 年国际实验室标准（ISL）要求的识别手段后，收集并分析了 72 个样本。在十个样本中，有八个样本的尼古丁和/或代谢物被检测到，而浓度测量表明在过去 3 天内有暴露。尼古丁、可替宁、反式-3-羟基可替宁、尼古丁-N'-氧化物和可替宁-N-氧化物的浓度范围分别为 11 至 19,750、13 至 10,475、10 至 8,217、11 至 3,396 和 13 至 1,640ng/mL。当提出比赛前和/或期间尼古丁消费的保守浓度限制（50ng/mL 用于尼古丁、可替宁和反式-3-羟基可替宁，25ng/mL 用于尼古丁-N'-氧化物和可替宁-N-氧化物）时，大约一半的冰球运动员被认定为消费者。这些发现显著支持广泛使用无烟尼古丁的可能性。然而，由于这些结论只能假设，因此需要进一步调查无烟烟草作为冰球兴奋剂的潜在用途。
