Micro-UHPLC-MS/MS method for analysis of oxylipins in plasma and platelets

Oxylipins play an important role in cell signaling and they act as auto- and paracrine factors. There are numerous reports on the analysis of oxylipins in biofluids, especially in plasma. Only a limited number of studies addressed the analysis of oxylipins in platelets using modern, sensitive LCMS methods, even though these compounds have a huge impact on platelet functions and thrombo-inflammation. In this work, a new method based on superficially porous particle (2.7 μm) capillary column (0.5 mm ID) and micro-liquid chromatography coupled to tandem mass spectrometry (μUHPLC-ESI-QqQ-MS/MS) method has been developed, optimized and validated. It has finally been successfully applied for human plasma and platelet analysis. The method allows the precise and accurate simultaneous quantification of 42 oxylipins with 13 deuterated internal standards. Solid phase extraction with Bond Elut Certify II provides good extraction recoveries (on average around 75 %). The μUHPLC-MS/MS method is selective, sensitive (LOQs between 30 and 150 pg/mL) and shows good linearity. Limits of detections for most of the compounds are between 2 and 250 fmol on column. Twenty-three oxylipins have been detected in plasma and 19 in non-activated (resting) platelets (all samples were from healthy donors). The μUHPLC-MS/MS method uses very low volume of mobile phase (less than 250 μL of organic solvents in mobile phase per analysis), and therefore is considered environmentally friendly. It also turned out to be robust enough for routine analysis.