Investigation of stationary phases performance for eicosanoids profiling in RP-HPLC.

Abstract

Eicosanoids - oxidative derivatives from arachidonic acid - represent biologically active lipid mediators in inflammatory processes. Different analytical methods treat eicosanoid analysis. Among which, reverse phase liquid chromatography figures as the appropriate method for eicosanoid profiling. RP-HPLC for eicosanoid analysis is often conducted on C18 columns. Some studies focused on profiling one family of eicosanoids; others considered all eicosanoid families. In both cases, co-elution remained a major issue and detection in mass spectrometry partially resolves this problem. In fact, the mass transitions used to monitor eicosanoid species are not specific enough and many isobars can be listed. For this, optimizing the RP-HPLC separation remains important. Based on the parameter Fs - deriving from the hydrophobic-subtraction model - and radar plots, we chose columns with different selectivities. The hydrophobic-subtraction model guided our interpretation of molecular interactions between eicosanoids and stationary phases. We founded our approach for selectivity optimization on peak capacity per minute and time needed values. Herein, we screened seven stationary phases and evaluated their chromatographic performances in RP-HPLC. Stationary phases presented different chemistry, type of silica, length, and particle size. Superficially porous particle columns registered better chromatographic profiles than classical stationary phases; and columns with embedded polar group did not serve our purpose. The stationary phase Accucore C30 - even being the least retentive - revealed the best selectivity and efficiency, and recorded the shorter duration for eicosanoid analysis.