Analysis of 12/15-lipoxygenase metabolism of EPA and DHA with special attention to authentication of docosatrienes.
Analysis of 12/15-lipoxygenase metabolism of EPA and DHA with special attention to authentication of docosatrienes.
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DOI：
10.1016/j.jlr.2021.100088
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发表时间：
2021
影响因子：
6.5
通讯作者：
Brash AR
中科院分区：
文献类型：
--
作者：
Jin J;Boeglin WE;Brash AR
高度不饱和的“鱼油”脂肪酸的一个拟议的有益影响是它们被脂氧合酶(LOX)酶转化为专门的促分解脂质介体，包括来自EPA和DHA的12/15-LOX产物。DHA的转化包括形成二十二碳三烯，因双键的独特共轭三烯而得名。为了加深对生物合成途径和机制的了解，我们将重组人15-LOX-1形成的不稳定的白三烯A(LTA)相关环氧化物中间体的生物合成和核磁共振表征以及稳定的酶产物鉴定结合在一起，并将研究结果扩展到常驻小鼠腹膜巨噬细胞中12/15-LOX的代谢。15-LOX-1氧化EPA可将最初的15S-过氧化氢转化为14S，15S-反式-环氧5Z，8Z，10E，12E，17Z-EPA(以其8，15-二醇的水解物形式出现)和二氢过氧基脂肪酸的混合物，而在小鼠细胞中主要是环氧化物的水解物。DHA还被15-LOX-1转化为环氧化物和二氢过氧物，生成10，17-二氢(过氧基)XY衍生物(二十二碳三烯)和少量7S，17S-和14，17S-二氢过氧物的混合物。在小鼠巨噬细胞中，与LTA相关的环氧化物中间体的10，17S-二羟基水解产物占主导地位，而与白三烯B4相关的带有反式、反式、顺式共轭三烯的(Neuro)Protectin D1则未被检测到。在这项研究中，我们强调了UV光谱特征在产品鉴定中的作用，它可以诊断不同的双键结构和羟基脂肪酸官能度与过氧化氢的对比。LC-MS不能确定构型异构体。安全的鉴定是基于色谱保留时间、与正品标准的比较以及高度独特的UV光谱。
A proposed beneficial impact of highly unsaturated “fish oil” fatty acids is their conversion by lipoxygenase (LOX) enzymes to specialized proresolving lipid mediators, including 12/15-LOX products from EPA and DHA. The transformations of DHA include formation of docosatrienes, named for the distinctive conjugated triene of the double bonds. To further the understanding of biosynthetic pathways and mechanisms, herein we meld together biosynthesis and NMR characterization of the unstable leukotriene A (LTA)-related epoxide intermediates formed by recombinant human 15-LOX-1, along with identification of the stable enzymatic products, and extend the findings into the 12/15-LOX metabolism in resident murine peritoneal macrophages. Oxygenation of EPA by 15-LOX-1 converts the initial 15S-hydroperoxide to 14S,15S-trans-epoxy-5Z,8Z,10E,12E,17Z-EPA (appearing as its 8,15-diol hydrolysis products) and mixtures of dihydroperoxy fatty acids, while mainly the epoxide hydrolysis products are evident in the murine cells. DHA also undergoes transformations to epoxides and dihydroperoxides by 15-LOX-1, resulting in a mixture of 10,17-dihydro(pero)xy derivatives (docosatrienes) and minor 7S,17S- and 14,17S-dihydroperoxides. The 10,17S-dihydroxy hydrolysis products of the LTA-related epoxide intermediate dominate the product profile in mouse macrophages, whereas (neuro)protectin D1, the leukotriene B4-related derivative with trans,trans,cis conjugated triene, was undetectable. In this study, we emphasize the utility of UV spectral characteristics for product identification, being diagnostic of the different double bond configurations and hydroxy fatty acid functionality versus hydroperoxide. LC-MS is not definitive for configurational isomers. Secure identification is based on chromatographic retention times, comparison with authentic standards, and the highly distinctive UV spectra.