Institute of Pharmaceutical and Medicinal Chemistry, University of Münster, Corrensstrasse 48, D-48149, Münster, Germany.
Institute of Pharmaceutical and Medicinal Chemistry, University of Münster, Corrensstrasse 48, D-48149, Münster, Germany.
Prostaglandins Other Lipid Mediat. 2021 Aug;155:106551. doi: 10.1016/j.prostaglandins.2021.106551. Epub 2021 Apr 30.
An automatic on-line dilution/on-line solid phase extraction (SPE) system has been developed for the detection of metabolites of the arachidonic acid cascade in platelets. The method allows the direct injection of larger quantities of centrifugates from cell suspensions previously treated with an equal volume of an acetonitrile/methanol mixture for protein precipitation. The method was used to study the effect of inhibitors of platelet arachidonic acid cascade enzymes (cytosolic phospholipase Aα, cyclooxygenase-1, thromboxane synthase, 12-lipoxygenase) and related targets (cyclooxygenase-2, microsomal prostaglandin E synthase-1, 5-lipoxygenase) in intact platelets after stimulation with calcium ionophore A23187. In addition to enzyme inhibition, the cell-damaging properties of the test compounds was determined by measuring the release of serotonin from the platelets into the incubation buffer.
已经开发出一种用于检测血小板中花生酸级联代谢物的自动在线稀释/在线固相萃取 (SPE) 系统。该方法允许直接注射从先前用等体积的乙腈/甲醇混合物处理以进行蛋白质沉淀的细胞悬浮液的离心沉淀物。该方法用于研究血小板花生酸级联酶（胞质磷脂酶 Aα、环加氧酶-1、血栓素合酶、12-脂氧合酶）和相关靶点（环加氧酶-2、微粒体前列腺素 E 合酶-1、5-脂氧合酶）的抑制剂对经钙离子载体 A23187 刺激后的完整血小板的影响。除了酶抑制作用外，还通过测量血小板将 5-羟色胺释放到孵育缓冲液中来确定测试化合物的细胞损伤特性。
