质谱能量分辨曲线法区分氧化脂类前列腺素异构体

目的：利用能量分辨曲线建立有效区分氧化脂类的质谱方法，为氧化脂类在细胞信号传导过程中机理的研究提供高灵敏、高效的异构体区分手段，特别是在没有对照品的情况下可以迅速地区分出氧化脂类位置异构体。 方法：对照品经甲醇溶解，0.22 μm滤膜过滤后上机分析。采用电喷雾电离源负离子电离模式，喷雾电压2.6 kV，鞘气（N₂）流量400 L·h^-1，辅助气（N₂）流量1 Arbs，离子传输毛细管温度250℃，扫描采用二级质谱子离子扫描模式，碰撞能量5~25 eV。 结果：4对氧化脂类异构体PGB2和PGA2，PGE1和PGD1，5（R）-HETE、12（S）-HETE和15（S）-HETE，9（S）-HODE和13（S）-HODE不同碰撞能量下母离子与碎片离子绘制出能量分辨曲线。其中3对异构体PGE1和PGD1，9（S）-HODE和13（S）-HODE，5（R）-HETE、12（S）-HETE和15（S）-HETE）结构上的主要区别在于羟基位点不同，结果表明羟基位点离末端羧基位点较远，则在能量分辨曲线中表现为离子能量曲线变化趋势较小，化合物较稳定；而异构体（PGA2和PGB2）在结构上的主要区别是双键位置不同，在能量分辨曲线上可以观察到共轭体系的存在使得化合物相对稳定。并通过海带中存在的4种氧化脂类进行了验证，经稳定性试验分析，这4对氧化脂类日内、日间RSD小于7%。 结论：本研究有效区分了前列腺素B2（PGB2）、前列腺素A2（PGA2）、前列腺素E1（PGE1）、前列腺素D1（PGD1）等9个氧化脂类成分，质谱能量分辨曲线法适用于氧化脂类位置异构体快速区分。

Objective: To establish a mass spectrometry method to rapidly distinguish the oxylipins in the process of lipid oxidation during cell signal transduction,especially to quickly distinguish oxylipins isomers without standards. Methods: The standards were dissolved with methanol,and then filtrated by 0.45 μm micromembrane.Mass spectrometry was performed on the high-performance liquid chromatography coupled with electrospray ionization/triple quadrupole mass spectrometry (HPLC/QqQ-MS) in negative modes with the spray voltage set at 2.6 kV.The gas flow of desolvation was set at 1 Arbs with 400 L·h^-1 of cone gas,and the capillary temperature was set at 250℃.The sampling collision energy was set at a ramp of 5-25 eV. Results: Breakdown curve is mapped by parent and diagnostic ion to distinguish four pairs of oxylipin isomers,including classes of PGB2 and PGA2,PGE1 and PGD1,5(R)-HETE,12(S)-HETE and 15(S)-HETE.According to experimental data,when the position of hydroxyl group is far from the position of the terminal carboxyl group,the compound is more stable according to the breakdown curve.The main difference among the structures of three pairs of isomer PGE1 and PGD1,9(S)-HODE and 13(S)-HODE,5(R)-HETE,12(S)-HETE and 15(S)-HETE is the position of hydroxyl group.The difference of structures between PGA2 and PGB2 is the double bond position.The results showed that the conjugate system makes the compound more stable according to the breakdown curve. Conclusion: This study effectively distinguishes prostaglandin B2 (PGB2),prostaglandin A2 (PGA2),prostaglandin E1 (PGE1) and prostaglandin D1 (PGD1);the mass spectrum energy resolution curve method is suitable for rapid identification of oxylipins isomers.