Lin Yu-Hong, Chen Cheng, Wei Shoupeng, Adrien Guillot, Mackowiak Bryan, Pan Hongna, Fu Yaojie, Maccioni Luca, Ren Tianyi, Zhang Li, Hibbeln Joseph, Pawlosky Robert, Gao Bin
Laboratory of Liver Diseases, NIAAA/NIH, Bethesda, Maryland, USA.
Laboratory of Integrative Neuroscience, NIAAA/NIH, Bethesda, Maryland, USA.
Alcohol Clin Exp Res (Hoboken). 2025 Sep;49(9):1897-1911. doi: 10.1111/acer.70126. Epub 2025 Aug 4.
Acetaldehyde, an immediate ethanol metabolite, mediates many ethanol-induced behavioral effects and is both psychoactive and toxic to animals and humans. Monitoring the kinetics of acetaldehyde using rodent models of alcohol misuse is essential for understanding and managing ethanol-associated diseases. However, quantitation of acetaldehyde in biological specimens after alcohol consumption has been challenging due to its high volatility, relatively low concentrations, and strong reactivity toward biochemical molecules. It was necessary to develop and establish an accurate and high-throughput method to quantitate acetaldehyde and ethanol.
Gas chromatography/mass spectrometry in positive chemical ionization mode coupled with a 111-vial headspace autosampler was employed to quantitate acetaldehyde and ethanol using H-acetaldehyde and H-ethanol as internal standards. A multidimensional approach was used to develop the method, including sample collection and processing, instrumental data analysis, optimization, and validation. Blood and tissues collected from genetically modified mouse models and their wild-type counterparts were studied.
The method was validated and applied to quantitate acetaldehyde and ethanol in blood and tissues from multiple mouse studies on ethanol metabolism. Acetaldehyde and ethanol were well-resolved from chromatographic interferences with linear ranges of 6.25-800 μM for acetaldehyde and 1.25-160 mM for ethanol. Both regression coefficients for calibration curves were >0.999. The within- and between-run precisions for ethanol in plasma, whole blood, and serum were all <5.0%, and for acetaldehyde in plasma and serum were <9.0%, while in whole blood it was 19.2%. Sample throughput was on the order of 60 samples per 15 h daily, with a maximum of 111 per batch.
Despite some limitations, this validated method proved to be specific, accurate, and reproducible for high-throughput quantitation of acetaldehyde and ethanol in rodent plasma, whole blood, serum, and visceral organs.
乙醛是乙醇的直接代谢产物，介导许多乙醇诱导的行为效应，对动物和人类具有精神活性和毒性。利用酒精滥用的啮齿动物模型监测乙醛的动力学对于理解和管理乙醇相关疾病至关重要。然而，由于乙醛具有高挥发性、相对低浓度以及对生物化学分子的强反应性，在饮酒后对生物样本中的乙醛进行定量分析一直具有挑战性。因此，有必要开发并建立一种准确且高通量的方法来定量乙醛和乙醇。
采用正化学电离模式的气相色谱/质谱联用111瓶顶空自动进样器，以氘代乙醛和氘代乙醇作为内标物来定量乙醛和乙醇。采用多维方法开发该方法，包括样本采集与处理、仪器数据分析、优化和验证。对从转基因小鼠模型及其野生型对照采集的血液和组织进行了研究。
该方法经过验证，并应用于多项关于乙醇代谢的小鼠研究中血液和组织中乙醛和乙醇的定量分析。乙醛和乙醇与色谱干扰物得到了很好的分离，乙醛的线性范围为6.25 - 800 μM，乙醇的线性范围为1.25 - 160 mM。校准曲线的两个回归系数均>0.999。血浆、全血和血清中乙醇的批内和批间精密度均<5.0%，血浆和血清中乙醛的精密度<9.0%，而全血中乙醛的精密度为19.2%。样本通量约为每天每15小时60个样本，每批最多111个样本。
尽管存在一些局限性，但该经过验证的方法被证明对于啮齿动物血浆、全血、血清和内脏器官中乙醛和乙醇的高通量定量分析具有特异性、准确性和可重复性。
