Quantifying Hepatic Long-Chain Acyl-CoA Esters

The level of hepatic LCACoAs was analyzed using UHPLC/MS/MS as described by Blachnio-Zabielska et al. [26 (link)]. LCACoA extraction was performed according to Minkler et al. [27 (link)] with the use of internal standards (C15:0-, 16:0(d4)-, C17-, C19:0-, C21:0-, C23:0- and 24:0(d4)-CoA) spiked into each sample. Samples were resolved on 2.1 mm × 150 mm Agilent ZORBAX Extend-C18, using a reverse-phase gradient of ammonium hydroxide (NH₄OH) in water and NH₄OH in ACN. The quantitative analysis of LCACOAs was performed in positive electrospray ionization mode (ESI+) on Sciex QTRAP 6500 + LC/MS/MS system (AB Sciex, Darmstadt, Germany).