Henke D C, Kouzan S, Eling T E
Anal Biochem. 1984 Jul;140(1):87-94. doi: 10.1016/0003-2697(84)90137-4.
High-performance liquid chromatography procedures were developed which separate leukotrienes (LTs), hydroxy-fatty acids (HETEs), prostaglandins (PGs), the stable metabolite of prostacyclin (6-keto-PGF1 alpha), the stable metabolite of thromboxane A2 (TXB2), 12-hydroxyheptadecatrienoic acid (HHT), and arachidonic acid (AA). Two methods employing reverse-phase columns are described. One method uses a radial compression system, the other a conventional steel column. Both systems employ methanol and buffered water as solvents. The radial compression system requires 60 min for separation of the AA metabolites, while the conventional system requires 100 min. Both methods provide good separation and recovery of 6-keto-PGF1 alpha, TXB2, PGE2, PGF2 alpha, PGD2, LTC4, LTB4, LTD4, LTE4, HHT, 15-, 12-, and 5-HETE; and AA. The 5S,12S-dihydroxy-6-trans, 8-cis, 10-trans, 14-cis-eicosatetraenoic acid (5S,12S-diHETE), a stereoisomer of LTB4, coelutes with LTB4. To determine the applicability of the methods to biologic systems, AA metabolism was studied in two models, guinea pig lung microsomes and rat alveolar macrophages. Both HPLC systems demonstrated good recovery and resolution of eicosanoids from the two biological systems. A simple evaporation technique for HPLC sample preparation, which avoids the use of chromatographic and other time-consuming methodology, is also described.
已开发出高效液相色谱法，可分离白三烯（LTs）、羟基脂肪酸（HETEs）、前列腺素（PGs）、前列环素的稳定代谢产物（6-酮-PGF1α）、血栓素A2的稳定代谢产物（TXB2）、12-羟基十七碳三烯酸（HHT）和花生四烯酸（AA）。描述了两种采用反相柱的方法。一种方法使用径向压缩系统，另一种使用传统的钢柱。两种系统均使用甲醇和缓冲水作为溶剂。径向压缩系统分离AA代谢产物需要60分钟，而传统系统需要100分钟。两种方法都能很好地分离和回收6-酮-PGF1α、TXB2、PGE2、PGF2α、PGD2、LTC4、LTB4、LTD4、LTE4、HHT、15-、12-和5-HETE以及AA。5S,12S-二羟基-6-反式、8-顺式、10-反式、14-顺式-二十碳四烯酸（5S,12S-二HETE），LTB4的立体异构体，与LTB4共洗脱。为了确定这些方法对生物系统的适用性，在豚鼠肺微粒体和大鼠肺泡巨噬细胞这两种模型中研究了AA代谢。两种高效液相色谱系统都显示出从这两种生物系统中很好地回收和分离类花生酸。还描述了一种用于高效液相色谱样品制备的简单蒸发技术，该技术避免了使用色谱法和其他耗时的方法。
