Powell W S
Anal Biochem. 1985 Jul;148(1):59-69. doi: 10.1016/0003-2697(85)90628-1.
High-pressure liquid chromatography is required to resolve the complex mixtures of arachidonic acid metabolites synthesized by many tissues. We have investigated some of the factors which affect the retention times of these substances in reversed-phase HPLC on columns of 5-micron octadecylsilyl silica. There are considerable differences in selectivity between mobile phases based on methanol and those based on acetonitrile, the latter being much better for cyclooxygenase products. The chromatographic behavior of peptidoleukotrienes (LTC4, LTD4, and LTE4) is quite different from that of other arachidonic acid metabolites which do not contain amino acids. Addition of phosphoric acid to the mobile phase results in very long retention times for peptidoleukotrienes. Very low concentrations of trifluoroacetic acid have effects similar to that of phosphoric acid, but as its concentration is raised, the retention times of peptidoleukotrienes decrease, whereas those of other arachidonic acid metabolites are unaffected. Changing the concentration of acetonitrile in the mobile phase also affects the retention times of peptidoleukotrienes differently from those of other metabolites. This information has been used to devise simple linear gradients which separate most of the major cyclooxygenase and lipoxygenase products of arachidonic acid metabolism.
需要用高压液相色谱法来分离由许多组织合成的花生四烯酸代谢物的复杂混合物。我们研究了一些影响这些物质在5微米十八烷基硅烷硅胶柱上进行反相高效液相色谱时保留时间的因素。基于甲醇的流动相和基于乙腈的流动相之间在选择性上存在相当大的差异，后者对环氧化酶产物的分离效果要好得多。肽白三烯（LTC4、LTD4和LTE4）的色谱行为与其他不含氨基酸的花生四烯酸代谢物有很大不同。向流动相中添加磷酸会导致肽白三烯的保留时间非常长。极低浓度的三氟乙酸具有与磷酸类似的效果，但随着其浓度升高，肽白三烯的保留时间会缩短，而其他花生四烯酸代谢物的保留时间则不受影响。改变流动相中乙腈的浓度对肽白三烯保留时间的影响也与其他代谢物不同。这些信息已被用于设计简单的线性梯度，以分离花生四烯酸代谢的大多数主要环氧化酶和脂氧化酶产物。
