Ultimate 3000 uhplc pump

Reversed phase UHPLC/MS was conducted using a Dionex Ultimate 3000 UHPLC pump interfaced with a Thermo Fisher Q-Exactive Orbitrap mass spectrometer. An ACE Excel PFP-C18 2.1 × 100 mm with 2.0 μm particle size column (ACE #EXL-1010-1002U) was used, with 2 μL injections of chemical extracts. UHPLC was conducted using a flow rate of 350 μL/min with water and acetonitrile gradient, where each contained 0.1% formic acid. An initial mobile phase of 100% water was held for 3 min, followed by a linear gradient from 0–80% acetonitrile over 10 min, then held at 80% acetonitrile for 3 min. MS was conducted in ESI⁺ and ESI⁻ modes with a probe temperature of 350 °C, spray voltage of 3500 V, capillary temperature of 320 °C, sheath gas flow rate of 40 arbitrary units, and auxiliary gas flow rate of 10 arbitrary units. Tandem mass spectra for DKPs 1–6 and 10 were collected using a Dionex Ultimate 3000 UHPLC pump interfaced with a Thermo Fisher LTQ-XL mass spectrometer in ESI⁺ mode with parameters analogous to those above and a normalized collision energy of 40 for MS².