Department of Pharmaceutical Sciences, College of Pharmacy, Western University of Health Sciences, Pomona, California (K.K., S.M.A., C.S.S., A.H., K.L.) and Lab Launch, Monrovia, California (K.K.)
Department of Pharmaceutical Sciences, College of Pharmacy, Western University of Health Sciences, Pomona, California (K.K., S.M.A., C.S.S., A.H., K.L.) and Lab Launch, Monrovia, California (K.K.).
Drug Metab Dispos. 2022 Dec;50(12):1454-1463. doi: 10.1124/dmd.122.001059. Epub 2022 Oct 2.
Two oxidation products of cotinine, 5-hydroxycotinine (5-HC) and cotinine N-oxide (CNO), were identified for the first time in vivo in the plasma of C57BL/6 mice after injection of nicotine (1 mg/kg) or exposure to an e-cigarette (e-cig) containing 2.4% nicotine. Liquid chromatography-mass spectrometry was used to separate 3-hydroxycotinine (3-HC), 5-HC, and CNO and to quantify each by the sensitive direct detection of their parent ion with m/z of 193.097. In nicotine-injected mice, 5-HC was as abundant as 3-HC 15 minutes postinjection, and CNO was readily detectable. In e-cig-exposed mice with plasma nicotine levels resembling that of human smokers, plasma 5-HC and CNO, as well as 3-HC, were readily quantifiable at the end of the 4-hour exposure time. In nicotine-injected mice, the combined concentration of 3-HC plus 5-HC plus CNO, all formed from cotinine by CYP2A5, was higher ( < 0.01) in females than in males, although the male-female difference in cotinine plasma level did not reach statistical significance. The result highlights the importance of considering these three oxidation products of cotinine in examining cotinine metabolism and disposition. Coumarin 7-hydroxylase activity, a specific marker of CYP2A5, measured in the hepatic microsomes of untreated mice showed that females have higher activity ( < 0.001) than males ( = 8 per sex). The abundance of plasma 5-hydroxycotinine in nicotine-treated mice raises intriguing questions about the site of its origin (hepatic or possibly kidney CYP2A5) and the routes of its disposition because urinary excretion of 5-HC has not been detected by liquid chromatography with tandem mass spectrometry in mice and is controversial in human smokers. SIGNIFICANCE STATEMENT: Nicotine is the active ingredient of tobacco, but its elimination route through its biomarker cotinine is not fully understood. By liquid chromatography-mass spectrometry, this study has identified and quantified for the first time 5-hydroxycotinine and cotinine N-oxide, which are oxidation products of cotinine, in the plasma of mice treated with nicotine or exposed to e-cigarettes. The results raise intriguing questions about nicotine disposition in vivo in this well established preclinical model of human smokers.
两种可替宁的氧化产物，5-羟基可替宁（5-HC）和可替宁 N-氧化物（CNO），在给予尼古丁（1 毫克/千克）或暴露于含有 2.4%尼古丁的电子烟后，首次在 C57BL/6 小鼠的血浆中被鉴定为体内产物。使用液相色谱-质谱法分离 3-羟基可替宁（3-HC）、5-HC 和 CNO，并通过对其母离子 m/z 为 193.097 的灵敏直接检测来定量。在尼古丁注射的小鼠中，5-HC 在注射后 15 分钟与 3-HC 一样丰富，并且可以容易地检测到 CNO。在暴露于电子烟的小鼠中，其血浆尼古丁水平与人类吸烟者相似，可在 4 小时暴露结束时定量检测到血浆 5-HC 和 CNO 以及 3-HC。在尼古丁注射的小鼠中，由 CYP2A5 从可替宁形成的 3-HC 加 5-HC 加 CNO 的总浓度（<0.01）在雌性中高于雄性，尽管雄性和雌性的可替宁血浆水平差异没有达到统计学意义。该结果突出表明，在检查可替宁代谢和处置时，需要考虑这三种可替宁的氧化产物。在未处理的小鼠肝微粒体中测量的香豆素 7-羟化酶活性，一种 CYP2A5 的特异性标志物，表明雌性（<0.001）高于雄性（每组 8 只）。尼古丁处理的小鼠中血浆 5-羟基可替宁的丰富度提出了一个有趣的问题，即其起源部位（肝脏或可能是肾脏 CYP2A5）及其处置途径，因为在小鼠中尚未通过液相色谱-串联质谱法检测到尿液中 5-HC 的排泄，并且在人类吸烟者中存在争议。意义陈述：尼古丁是烟草的有效成分，但通过其生物标志物可替宁的消除途径尚未完全了解。通过液相色谱-质谱法，本研究首次在给予尼古丁或暴露于电子烟的小鼠的血浆中鉴定和定量了可替宁的两种氧化产物，5-羟基可替宁和可替宁 N-氧化物。这些结果提出了一个有趣的问题，即在这个成熟的人类吸烟者临床前模型中，体内尼古丁的处置情况。
