Determination of neutral lipid hydroperoxides by size exclusion HPLC with fluorometric detection. Application to fish oil enriched mayonnaises during storage

A fast (12 min) high-performance size exclusion chromatography (HPSEC) method for the separation of neutral lipid class hydroperoxides in the oil phases from fish oil enriched mayonnaises was developed. Detection and quantification were performed using the postcolumn fluorometric (FL) diphenyl-1-pyrenylphosphine oxidation principle. The reproducibilities judged by intra- and inter-assay variations were 0.64 and 7.2%, respectively. The HPSEC-FL method was applied to assess the effect of supplementations with emulsifier, gallic acid, and EDTA on the oxidative processes in the mayonnaises during storage. Substantial amounts of hydroperoxy triacylglycerols (TAGOOH) and cholesterol esters (CEOOH), together with traces of TAGOOH-dimers, were detected. All supplementations significantly decreased the levels of TAGOOH and to a lesser degree CEOOH. Supplementations with EDTA and gallic acid resulted in constant and slightly increasing levels of TAGOOH, respectively, thus affecting the oxidation mechanisms seen in reference mayonnaise. The emulsifier Panodan TR DATEM reduced the levels of TAGOOH as compared to the appropriate controls.