Mass Spectrometric Analysis of Triacylglycerol Hydroperoxide Isomers

Q1 mass and product ion mass spectra in the TG 18:1_18:1_18:1;OOH isomer mixture were obtained using a 4000 QTRAP mass spectrometer (SCIEX, Tokyo, Japan). Standard TG 18:1_18:1_18:1;OOH was diluted in methanol (0.5 µM) and directly infused into the MS at a flow rate of 10 µL/min. Positive ESI was used as the ion source. MS spectra were obtained in a range of m/z 100–1000. Analytical parameters were optimized using the Analyst software (ver. 1.6.2, SCIEX, Tokyo, Japan) (Supplementary Materials).
TG 18:1_18:1_18:2;OOH and TG 18:1_18:1_18:1;OOH isomers were analyzed in multiple-reaction-monitoring (MRM) mode. The MRM transitions described in the Supplementary Materials were used. LC-MS/MS analysis was conducted using an ExionLC HPLC system (SCIEX, Tokyo, Japan) equipped with a 4000 QTRAP mass spectrometer. An Inertsil SIL-100A column (5 µm, 2.1 × 250 mm, GL Sciences Inc., Tokyo, Japan) was eluted with hexane/2-propanol/acetic acid (100:0.6:0.5, v/v/v) at 0.2 mL/min (40 °C). A post-column solvent consisting of methanol/2-propanol (1:1, v/v) containing 0.2 mM sodium acetate was mixed with the eluent at 0.2 mL/min to promote ionization [29 (link)]. TG 18:1_18:1_18:2;OOH and TG 18:1_18:1_18:1;OOH isomers were quantitated with external standard curves.