Yuanyuan Ji , Yulemni Morel , Anh Q. Tran , Marta M. Lipinski , Chinmoy Sarkar , Jace W. Jones
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Method development involved comparing the sensitivity of detection between widely used derivatization reagents: 2,4-dinitrophenylhydrazine and 3-nitrophenylhydrazine. Our data showed that 3-nitrophenylhydrazine resulted in greater sensitivity. Additional method development included evaluation of hydrolysis sample pretreatment, selection of protein precipitation reagent, and optimization of derivatization conditions. The optimized conditions included no hydrolysis and use of 20 % trichloroacetic acid as the protein precipitation reagent. The optimized derivatization condition was 25 mM 3-nitrophenylhydrazine reacted at 20 °C for 30 min. The chromatographic conditions were optimized to reduce matrix effects, ion suppression, and efficient analysis time (<7-minute analytical run). The four aldehyde species were accurately quantified in brain tissue using stable-labeled internal standards. 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引用次数: 0
摘要
反应性醛是一类亲电性低分子量化合物,在生理功能和脂质过氧化过程中发挥着重要作用。这些分子与许多疾病(尤其是心血管和神经退行性疾病)有关,是潜在的脂质过氧化内源性标志物。然而,目前能准确量化脑组织中多种活性醛的方法还很有限。本研究开发并验证了一种基于 3-硝基苯肼衍生化的 LC-MS/MS 方法来定量检测四种活性醛:丙二醛、丙烯醛、4-羟基-2-己烯醛和 4-羟基-2-壬烯醛。方法开发包括比较广泛使用的衍生试剂的检测灵敏度:2,4-二硝基苯肼和 3-硝基苯肼。数据显示,3-硝基苯肼的灵敏度更高。其他方法开发包括水解样品预处理评估、蛋白质沉淀试剂的选择和衍生条件的优化。优化条件包括不水解和使用 20% 的三氯乙酸作为蛋白质沉淀试剂。优化的衍生条件为 25 mM 3-硝基苯肼在 20 °C 下反应 30 分钟。对色谱条件进行了优化,以减少基质效应、离子抑制和有效的分析时间 (
Development and evaluation of a liquid chromatography-tandem mass spectrometry method for simultaneous measurement of toxic aldehydes from brain tissue
Reactive aldehydes are a class of electrophilic low molecular weight compounds that play an essential role in physiological function and lipid peroxidation. These molecules are implicated in many diseases, especially cardiovascular and neurodegenerative diseases, and are potential endogenous markers of lipid peroxidation. However, there are limited options to accurately quantify multiple reactive aldehydes in brain tissue. This study developed and validated a 3-nitrophenylhydrazine derivatization-based LC-MS/MS method to quantify four reactive aldehydes: malondialdehyde, acrolein, 4-hydroxy-2-hexenal and 4-hydroxy-2-nonenal. Method development involved comparing the sensitivity of detection between widely used derivatization reagents: 2,4-dinitrophenylhydrazine and 3-nitrophenylhydrazine. Our data showed that 3-nitrophenylhydrazine resulted in greater sensitivity. Additional method development included evaluation of hydrolysis sample pretreatment, selection of protein precipitation reagent, and optimization of derivatization conditions. The optimized conditions included no hydrolysis and use of 20 % trichloroacetic acid as the protein precipitation reagent. The optimized derivatization condition was 25 mM 3-nitrophenylhydrazine reacted at 20 °C for 30 min. The chromatographic conditions were optimized to reduce matrix effects, ion suppression, and efficient analysis time (<7-minute analytical run). The four aldehyde species were accurately quantified in brain tissue using stable-labeled internal standards. Application of this assay to a traumatic brain injury mouse model revealed significant accumulation of acrolein, 4-hydroxy-2-hexenal, and 4-hydroxy-2-nonenal at 28 days post injury. Overall, a validated method was developed to rapidly quantify the most prominent reactive aldehydes associated with lipid peroxidation during injury progression following acute brain trauma.
期刊介绍:
The Journal of Chromatography B publishes papers on developments in separation science relevant to biology and biomedical research including both fundamental advances and applications. Analytical techniques which may be considered include the various facets of chromatography, electrophoresis and related methods, affinity and immunoaffinity-based methodologies, hyphenated and other multi-dimensional techniques, and microanalytical approaches. The journal also considers articles reporting developments in sample preparation, detection techniques including mass spectrometry, and data handling and analysis.
Developments related to preparative separations for the isolation and purification of components of biological systems may be published, including chromatographic and electrophoretic methods, affinity separations, field flow fractionation and other preparative approaches.
Applications to the analysis of biological systems and samples will be considered when the analytical science contains a significant element of novelty, e.g. a new approach to the separation of a compound, novel combination of analytical techniques, or significantly improved analytical performance.