Furthermore, elevated EETs triggered massive, unprecedented patterns of metastatic spread and escape from tumor dormancy [88], raising concerns about therapeutic strategies that involve up-regulation of EETs [89]. 2. Analysis of Arachidonic Acid-Derived Eicosanoids by Selleck Ceritinib targeted LC-MS. The profile of arachidonic acid metabolites is complicated by the enantioselectivity of eicosanoid formation as well as the variety of regioisomers that arise (Figure

1). In order to investigate the metabolism of arachidonic acid in vitro or in vivo, targeted chiral methods Inhibitors,research,lifescience,medical are advantageous, to help distinguish between the enantiomers that are formed by different pathways. In a 2009 review article [90], we observed Inhibitors,research,lifescience,medical that there were few reports of targeted approaches for more than one class of eicosanoids. Since that time, a number of targeted approaches have appeared [91,92,93,94,95,96,97,98,99,100] where more than one class of eicosanoid and/or other metabolites arising from the same metabolic pathway were analyzed

[93]. To efficiently conduct targeted eicosanoid analyses, the LC separations are coupled with CID and MS/MS analysis. Product ion profiles are often diagnostic for particular regioisomers. The highest sensitivity that can be achieved for the analysis of eicosanoids Inhibitors,research,lifescience,medical involves the use of LC-SRM/MS. Highest specificity is obtained when the SRM transition is between an intense parent ion which contains the intact molecule (M) and a structurally significant product ion. An example of this useful situation arises with HETEs, where product ions are formed through α-cleavage

adjacent to a double Inhibitors,research,lifescience,medical bond [101]. In some cases, fragment ions produced in the collision cell are not very specific, and isomeric eicosanoids sometimes produce very similar product ion profiles. An example of this less desirable situation arises with PGE2 and PGD2, where the isomers Inhibitors,research,lifescience,medical need to be well separated by LC for correct quantification. Most LC-SRM/MS methods that have been reported employ ESI in the negative ion mode, where the parent ion arises from de-protonation of the eicosanoid molecule (M) to give an ion corresponding to [M-H]-. LC has generally been performed using reversed stationary else phases coupled with aqueous mobile phases [91,93,94,95]. Analyses are normally conducted using stable isotope dilution methodology with deuterium-labeled eicosanoid analogs as internal standards (ISTDs), which confer much greater specificity than structural analog ISTDs. Quantification is performed by constructing calibration curves for each analyte. Standard solutions of different concentrations are prepared by serial dilution from commercially available standard eicosanoids and they are spiked with the same amount of the deuterium labeled ISTD as the samples to be determined. However, most targeted methods do not use chiral chromatography and so they cannot distinguish between pairs of eicosanoid enantiomers.