An optimized method for the measurement of acetaldehyde by high-performance liquid chromatography.
An optimized method for the measurement of acetaldehyde by high-performance liquid chromatography.
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DOI：
10.1111/j.1530-0277.2011.01612.x
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发表时间：
2012-03
期刊：
影响因子：
--
通讯作者：
Anni H
中科院分区：
文献类型：
--
作者：
Guan X;Rubin E;Anni H
乙醛主要在肝脏中通过乙醇脱氢酶在乙醇代谢过程中产生，并通过醛脱氢酶氧化为乙酸盐而迅速消除。通过顶空气相色谱法和反相高效液相色谱法（RP-HPLC）评估生物基质中的循环乙醛水平。我们建立了一个优化的反相高效液相色谱法测定肝癌细胞培养基，血液和血浆中乙醛的方法。样品脱蛋白后，乙醛用2，4-二硝基苯肼（DNPH）衍生。对反应的pH值、衍生化试剂用量、反应时间和反应温度进行了优化。对乙酰丙酮-腙（AcH-DPN）稳定衍生物的提取方法和产品稳定性进行了研究。使用新的色谱梯度程序，通过与AcH-DPN标准品相比的保留时间鉴别乙醛，并在存在和不存在乙醇的情况下基于外部参比标准品和标准品加入校准曲线进行定量。在pH 4.0下用80倍摩尔过量的DNPH进行乙醛的衍生化。在环境温度下，反应在40分钟内完成，并且产物稳定2天。用一种新的11分钟层析程序可使AcH-DNP与DNPH完全分离。乙醛检测线性高达80 μM。乙醛的回收率在培养基中>88%，在血浆中>78%。我们定量测定了肝癌细胞、大鼠血液和血浆中乙醇衍生的乙醛，检测限约为3 μM。在小体积（70 μl）血浆采样中，该方法的日内测量准确度<9%，日间测量准确度<15%。采用DNPH衍生化，反相高效液相色谱法（RP-HPLC）快速分离AcH-DNP，建立了生物体系中乙醛的定量分析方法。该方法具有扩展的线性范围，重现性好，适用于培养基和生物液体的小体积取样。
Acetaldehyde is produced during ethanol metabolism predominantly in the liver by alcohol dehydrogenase, and rapidly eliminated by oxidation to acetate via aldehyde dehydrogenase. Assessment of circulating acetaldehyde levels in biological matrices is performed by headspace gas chromatography and reverse phase high-performance liquid chromatography (RP-HPLC). We have developed an optimized method for the measurement of acetaldehyde by RP-HPLC in hepatoma cell culture medium, blood and plasma. After sample deproteinization, acetaldehyde was derivatized with 2,4-dinitrophenylhydrazine (DNPH). The reaction was optimized for pH, amount of derivatization reagent,, time and temperature. Extraction methods of the acetaldehyde-hydrazone (AcH-DPN) stable derivative and product stability studies were carried out. Acetaldehyde was identified by its retention time in comparison to AcH-DPN standard, using a new chromatography gradient program, and quantitated based on external reference standards and standard addition calibration curves in the presence and absence of ethanol. Derivatization of acetaldehyde was performed at pH 4.0 with a 80-fold molar excess of DNPH. The reaction was completed in 40 min at ambient temperature, and the product was stable for 2 days. A clear separation of AcH-DNP from DNPH was obtained with a new 11-min chromatography program. Acetaldehyde detection was linear up to 80 μM. The recovery of acetaldehyde was >88% in culture media, and >78% in plasma. We quantitatively determined the ethanol-derived acetaldehyde in hepatoma cells, rat blood and plasma with a detection limit around 3 μM. The accuracy of the method was <9% for intraday and <15% for interday measurements, in small volume (70 μl) plasma sampling. An optimized method for the quantitative determination of acetaldehyde in biological systems was developed using derivatization with DNPH, followed by a short RP-HPLC separation of AcH-DNP. The method has an extended linear range, is reproducible and applicable to small volume sampling of culture media and biological fluids.
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DOI：
10.1111/j.1530-0277.2009.01031.x
发表时间：
2009-11
期刊：
Alcoholism, clinical and experimental research
影响因子：
--
作者：
Arolfo MP;Overstreet DH;Yao L;Fan P;Lawrence AJ;Tao G;Keung WM;Vallee BL;Olive MF;Gass JT;Rubin E;Anni H;Hodge CW;Besheer J;Zablocki J;Leung K;Blackburn BK;Lange LG;Diamond I
通讯作者：
Diamond I
影响因子：
3.9
作者：
CLEMENS, DL;HALGARD, CM;TUMA, DJ
通讯作者：
TUMA, DJ
DOI：
10.1097/00000374-199709000-00020
发表时间：
1997-09-01
影响因子：
3.2
作者：
HambyMason, R;Chen, JJ;Henderson, GI
通讯作者：
Henderson, GI
DOI：
10.1111/j.1530-0277.1999.tb04020.x
发表时间：
1999-01-01
影响因子：
3.2
作者：
Anni, H;Israel, Y
通讯作者：
Israel, Y
影响因子：
--
作者：
Kempter, C;Zurek, G;Karst, U
通讯作者：
Karst, U