5,6,7,4′-四羟基黄酮和5,6,7,4′-四甲氧基黄酮高分辨电喷雾串联质谱裂解规律对比研究

以5,6,7,4′-四羟基黄酮和5,6,7,4′-四甲氧基黄酮为例,运用高分辨电喷雾串联质谱在正离子模式下(HR-ESI-MS/MS)获得的碎片离子,根据一、二级质谱离子的精确质荷比,分别推导出两种黄酮可能的裂解途径,并比较多甲氧基、多羟基黄酮质谱裂解规律的异同。结果表明:在正离子模式下,5,6,7,4′-四羟基黄酮和5,6,7,4′-四甲氧基黄酮的一级质谱均可获得稳定的准分子离子。5,6,7,4′-四羟基黄酮的二级质谱碎片离子由准分子离子M+H]+经RDA裂解及进一步脱水和(或)脱一氧化碳中性分子产生,直接由准分子离子脱水及一氧化碳中性分子获得的碎片离子丰度比较低;5,6,7,4′-四甲氧基黄酮二级质谱碎片即可由准分子离子M+H]+直接经RDA裂解,还可先脱去乙烷或甲烷中性分子再经RDA裂解(1,3断裂)获得。此外,该化合物还有大量的碎片离子是由准分子离子M+H]+分别经脱水、甲烷、乙烷或进一步脱甲基、一氧化碳而获得。两个化合物质谱碎片的比较结果表明,多甲氧基黄酮的二级质谱碎片裂解机制更复杂,碎片信息更丰富。该方法可为黄酮类化合物成分的快速定性分析提供支持,也可为黄酮类化合物体内代谢产物的LC-MS/MS鉴定提供依据。

Fragmentation Pathway Comparison of 5,6,7,4′-Tetrahydroxy-Flavone and 5,6,7,4′-Tetramethoxy-Flavone by High Resolution Electrospray Ionization Tandem Mass Spectroscopy

5,6,7,4′-Tetrahydroxy-flavone and 5,6,7,4′-tetramethoxy-flavone were subjected to high resolution electrospray ionization tandem mass spectrometry (HR-ESI-MS/MS) in positive ion mode to obtain their protonated molecules and subsequent product ions, which were further analyzed to give proposed fragmentation pathways for the two flavones of polyhydroxy and polymethoxy substitution according to the high resolution mass to charge of product ions. The results show that the two compounds give stable pseudo-molecular ions (protonated molecules) in positive ion mode. Product ions for 5,6,7,4′-tetrahydroxy-flavone are mainly produced by Retro Diels-Alder (RDA) reaction from its protonated molecule ［M+H］⁺ and subsequent neutral loss of H₂O and (or) CO, while direct neutral loss of H₂O and (or) CO from ［M+H］⁺ give product ions with low relative abundance. Product ions for 5,6,7,4′-tetramethoxy-flavone are obtained both by direct RDA cleavage from protonated molecule ［M+H］⁺ and neutral loss of C₂H₆ and CH₄ with subsequent RDA cleavage. More product ions for 5,6,7,4′-tetramethoxy-flavone are produced by loss of CH₄, C₂H₆, CO and H₂O, even CH₃. HR-ESI-MS/MS comparison of the two flavones shows very different MS fragmentation pathways, which leads to more special ion peaks for polymethoxy substitution than for polyhydroxy substitution.