Symmetry reverse phase c 18 column

Aliquots of tomato powders (0.25 g dw) were analyzed monthly up to 180 d, except for samples incubated at aw 0.56, which were analyzed up to 106 d. Extraction was performed with 10 mL of methanol. The mixture was vortexed for 1 min, mixed continuously for 30 min with a magnetic stirrer, and then centrifuged at 12.000× g and 5 °C for 10 min. Extractions were carried out in triplicate on initial samples and in duplicate for samples stored at different a_w levels. The phenolic contents of methanolic extracts were analyzed by HPLC as described previously [13 (link)]. A 250 × 4.6 mm i.d., 5 μm particle size, Symmetry reverse phase C-18 column (Waters) equipped with a Symmetry C-18 precolumn was used. Formic acid (5%) was added to both methanol and water before the following mobile phases were prepared: (A) water/methanol (95:5, v/v); (B) water/methanol (88:12, v/v); (C) water/methanol (20:80, v/v); (D) and methanol. The following gradient elution was used: 0−5 min, 100% A; 5−10 min linear gradient to reach 100% B; 10−13 min, 100% B; 13−35 min linear gradient to reach 75% B and 25% C; 35−50 min linear gradient to reach 50% B and 50% C; 50−52 min linear gradient to reach 100% C; 52−57 min, 100% C; 57−60 min, 100% D. The flow rate was 1 mL/min. Chlorogenic acid was quantified at 330 from calibration curves using pure standard and expressed as milligrams per kilogram of dry product.

LC separation was performed with a Shimadzu Prominence UFLC XR (Shimadzu Corporation). The multiple-reaction-monitoring (MRM) spectra were obtained with a QTRAP 5500 mass spectrometer (AB SCIEX) equipped with an ESI source. A Waters Symmetry reverse-phase C18 column (2.1 mm × 150 mm, 3.5 μm) was used for the LC separation as described (Meng et al., 2016 (link)). Up to 35 eicosanoids and two internal standards [15(S)-HETE-d₈ and PGB₂] were monitored. Optimized LC-MS/MS parameters for the analysis of eicosanoids are in accordance with previous research (Meng et al., 2016 (link)).