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The effluent from the column was directed to a nanospray ionization source connected to a hybrid quadrupole-Orbitrap mass spectrometer (Q Exactive Plus, Thermo Fisher Scientific, USA) acquiring mass spectra in a data-dependent mode alternating between a full scan (m/z 350-1700, automated gain control (AGC) target 3 106, 50 ms maximum injection time, FWHM resolution 70,000 at m/z 200) and up to 15 MS/MS scans (quadrupole isolation of charge states 2-7, isolation window 0.7 m/z) with previously optimized fragmentation conditions (normalized collision energy of 32, dynamic exclusion of 30 s, AGC target 1 105, 100 ms maximum injection time, FWHM resolution 35,000 at m/z 200)