Abstract
Trihydroxyoctadecenoic acids (TriHOMEs) are linoleic acid-derived oxylipins with potential physiological relevance in inflammatory processes as well as in maintaining an intact skin barrier. Due to the high number of possible TriHOME isomers with only subtle differences in their physicochemical properties, the stereochemical analysis is challenging and usually involves a series of laborious analytical procedures. We herein report a straightforward analytical workflow that includes reversed-phase ultra-HPLC-MS/MS for rapid quantification of 9,10,13- and 9,12,13-TriHOME diastereomers and a chiral LC-MS method capable of resolving all sixteen 9,10,13-TriHOME and 9,12,13-TriHOME regio- and stereoisomers. We characterized the workflow (accuracy, 98-120%; precision, coefficient of variation ≤6.1%; limit of detection, 90-98 fg on column; linearity, R2 = 0.998) and used it for stereochemical profiling of TriHOMEs in bronchoalveolar lavage fluid (BALF) of individuals with chronic obstructive pulmonary disease (COPD). All TriHOME isomers were increased in the BALF of COPD patients relative to that of smokers (P ≤ 0.06). In both COPD patients and smokers with normal lung function, TriHOMEs with the 13(S) configuration were enantiomerically enriched relative to the corresponding 13(R) isomers, suggesting at least partial enzymatic control of TriHOME synthesis. This method will be useful for understanding the synthetic sources of these compounds and for elucidating disease mechanisms.
Highlights
Trihydroxyoctadecenoic acids (TriHOMEs) are linoleic acid-derived oxylipins with potential physiological relevance in inflammatory processes as well as in maintaining an intact skin barrier
We describe a straightforward chromatographic workflow that enables characterization and quantification of the eight 9,10,13-trihydroxyoctadecenoic acid (TriHOME) and the eight 9,12,13-TriHOMEs based on synthesized racemic TriHOME standard mixes and standards with defined regio- and stereochemistry
The TriHOMEs in particular have experienced an increasing degree of attention due to their potential significance in human physiology. They have been hypothesized to play an important part in maintaining the human skin water barrier [9(R),10(S),13(R)-TriHOME, compound 7b [19, 20]] and were found to be dysregulated in several recent studies in respiratory diseases [21,22,23]
Summary
Trihydroxyoctadecenoic acids (TriHOMEs) are linoleic acid-derived oxylipins with potential physiological relevance in inflammatory processes as well as in maintaining an intact skin barrier. The concentrations of individual TriHOME diastereomers were determined by preparing adequate dilutions of the standard mixtures in 84% methanol and measuring the concentration in quadruplicates, as described, Quantification of TriHOME diastereomers by reversed-phase UHPLC-MS/MS analysis.
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