Two-dimensional LC
× LC–HRMS experiments were performed using an Agilent
1290 Infinity 2D- LC system (Agilent Technologies, Waldbronn, Germany)
coupled with a Waters Synapt-G2 high-resolution mass spectrometer.
The system comprised two binary pumps (G4220A) for solvent delivery,
an autosampler (G4226A), column thermostat (G1316C) equipped with
a 2D-LC 8-port 2-postion modulation valve (G4236A) with 40 μL
loops, and a diode-array detector (G4212A) equipped with an Agilent
Max-Light cartridge flow cell (G4212–6008, 10 mm, Vdet = 1.0 μL). The 1D column was a Phenomenex
Luna HILIC (150 × 2.0 mm i.d., 3.0 μm particles, 200 Å
pore size) column used for gradient-NPLC, while two Waters Acquity
APC XT columns (75 × 4.6 mm i.d., 1.7 μm particles, 45
Å pore size and 75 × 4.6 mm i.d., 2.5 μm particles,
125 Å pore size, respectively) were coupled in series for SEC
experiments.
For parallel UV/HRMS detection, the analytical
effluent was split after the second-dimension SEC column set using
a tee piece and in-house-made restriction capillaries (450 ×
0.075 mm i.d. and 900 × 0.050 mm i.d. capillaries), ensuring
a split ratio of 9:1 to the diode array and mass spectrometer, respectively.
Using the diverter valve of the Synapt-G2 system, the smallest split
flow was combined with a make-up flow (1:1 ratio) consisting of 1
mM NaI with 0.5% (v/v) 3-nitrobenzyl alcohol in deionized water.
× LC–HRMS experiments were performed using an Agilent
1290 Infinity 2D- LC system (Agilent Technologies, Waldbronn, Germany)
coupled with a Waters Synapt-G2 high-resolution mass spectrometer.
The system comprised two binary pumps (G4220A) for solvent delivery,
an autosampler (G4226A), column thermostat (G1316C) equipped with
a 2D-LC 8-port 2-postion modulation valve (G4236A) with 40 μL
loops, and a diode-array detector (G4212A) equipped with an Agilent
Max-Light cartridge flow cell (G4212–6008, 10 mm, Vdet = 1.0 μL). The 1D column was a Phenomenex
Luna HILIC (150 × 2.0 mm i.d., 3.0 μm particles, 200 Å
pore size) column used for gradient-NPLC, while two Waters Acquity
APC XT columns (75 × 4.6 mm i.d., 1.7 μm particles, 45
Å pore size and 75 × 4.6 mm i.d., 2.5 μm particles,
125 Å pore size, respectively) were coupled in series for SEC
experiments.
For parallel UV/HRMS detection, the analytical
effluent was split after the second-dimension SEC column set using
a tee piece and in-house-made restriction capillaries (450 ×
0.075 mm i.d. and 900 × 0.050 mm i.d. capillaries), ensuring
a split ratio of 9:1 to the diode array and mass spectrometer, respectively.
Using the diverter valve of the Synapt-G2 system, the smallest split
flow was combined with a make-up flow (1:1 ratio) consisting of 1
mM NaI with 0.5% (v/v) 3-nitrobenzyl alcohol in deionized water.