Dionex ultimate 3000 series uhplc

Chemical structures of anthocyanins were determined by UHPLC–ESI–HRMS performed with a Dionex Ultimate 3000 series UHPLC (Thermo Scientific, Schwerte, Germany) and a Bruker timsToF mass spectrometer (Bruker Daltonics, Bremen, Germany). UHPLC was used applying a Zorbax Eclipse XDB‐C18 column (100 mm × 2.1 mm, 1.8 µm; Agilent Technologies) with a solvent system of 0.1% (v/v) formic acid (A) and acetonitrile (B) at a flow rate of 0.3 mL·min⁻¹. The elution profile was the following: 0–0.5 min, 5% B; 0.5–11.0 min, 5–60% B; 11.0–11.1 min, 60–100% B, 11.1–12.0 min, 100% B and 12.1–15.0 min 5% B. ESI in positive ionization mode was used for the coupling of LC to MS. The mass spectrometer parameters were set as follows: capillary voltage 4.5 KV, end plate offset of 500 V, nebulizer pressure 2.8 bar, nitrogen at 280 °C at a flow rate of 8 L·min⁻¹ as drying gas. Acquisition was achieved at 12 Hz with a mass range from m/z 50 to 1500 with data‐dependent MS². Fragmentation was triggered at the two most intense peaks applying a target intensity of 20 000 counts, with MS² spectra acquisition at 2 Hz, and a limited total cycle time of 2 s. Collision energy was alternated between 20 and 50 to achieve mixed MS² spectra.

LC separations
were performed
using Dionex Ultimate 3000 series UHPLC (Thermo Fisher Scientific)
coupled to an Orbitrap Q-Exactive Plus Mass Spectrometer (Thermo Fisher
Scientific). The used column was Acclaim 120 (C18, 5 μm, 120
Å, 4.6 mm × 100 mm) (Thermo Fisher Scientific). The mobile
phases were A, H₂O, and B, AcN, both with 0.1% formic acid.
The flow rate used in all LC–MS experiments was 1 mL·min^–1, and sample elution was performed by using the gradient
from 5 to 95% of B over 6.5 min. 10 μL of sample injection was
set. Ionization was performed using an electrospray ion source operating
in positive ion mode with a capillary voltage of 3.80 kV and a capillary
temperature of 400 °C. The sheath gas, auxiliary gas, and sweep
gas flow rates were set at 75, 20, and 1 (arbitrary units), respectively.
The auxiliary gas temperature was set at 500 °C. For each studied
mass, a set of MS/MS spectra were acquired using an isolation width
of 1.0 and 5.0 m/z, and a screening of collision
energies (from 10 to 40 HCD) was carried out. All MSⁿ data
were analyzed using the Qual Browser embedded in the Thermo Fisher
Scientific Xcalibur program and the FreeStyle software.