An
Agilent 1290 Infinity HPLC system coupled with an Agilent 6410 B triple
quadrupole mass spectrometer (Agilent Technologies, Santa Clara, CA)
was employed for the analysis.
Chromatographic separation was
achieved on an Thermo Scientific Syncronis HILIC HPLC column (2.1
mm × 100 mm i.d., 5 μm particle size) at 30 °C. The
flow rate was set at 0.4 mL/min. A gradient elution program was run
for chromatographic separation, with mobile phase A (0.1% formic acid,
5 mM ammonium formate in water) and mobile phase B (0.1% formic acid
in acetonitrile), as follows: the gradient elution method was used
with 75% solvent B, 75–60% over 0–1.2 min, 60–50%
over 1.2–2.5 min, 50–75% over 2.5–2.8 min, followed
by a re-equilibration time of 2.4 min. The total run time was 5.2
min for each sample.
The optimized ion source conditions were
as follows: transfer capillary
temperature, 320 °C; capillary voltage, 4000 V; nebulizer, 20
psi; and gas flow, 10 L/min. The mass spectrometer was operated in
the electrospray ionization (ESI)-positive ion selected reaction monitoring
mode using a dwell time of 50 ms per transition to detect ion pairs
at m/z 104.01–87 for GABA.
The fragmentor was 62 V and collision energy was 8 eV.
Data
were acquired and analyzed using the software Gastroplus V9.0
(Simulations Plus).
Agilent 1290 Infinity HPLC system coupled with an Agilent 6410 B triple
quadrupole mass spectrometer (Agilent Technologies, Santa Clara, CA)
was employed for the analysis.
Chromatographic separation was
achieved on an Thermo Scientific Syncronis HILIC HPLC column (2.1
mm × 100 mm i.d., 5 μm particle size) at 30 °C. The
flow rate was set at 0.4 mL/min. A gradient elution program was run
for chromatographic separation, with mobile phase A (0.1% formic acid,
5 mM ammonium formate in water) and mobile phase B (0.1% formic acid
in acetonitrile), as follows: the gradient elution method was used
with 75% solvent B, 75–60% over 0–1.2 min, 60–50%
over 1.2–2.5 min, 50–75% over 2.5–2.8 min, followed
by a re-equilibration time of 2.4 min. The total run time was 5.2
min for each sample.
The optimized ion source conditions were
as follows: transfer capillary
temperature, 320 °C; capillary voltage, 4000 V; nebulizer, 20
psi; and gas flow, 10 L/min. The mass spectrometer was operated in
the electrospray ionization (ESI)-positive ion selected reaction monitoring
mode using a dwell time of 50 ms per transition to detect ion pairs
at m/z 104.01–87 for GABA.
The fragmentor was 62 V and collision energy was 8 eV.
Data
were acquired and analyzed using the software Gastroplus V9.0
(Simulations Plus).