Kinetex biphenyl lc column

A Thermo Finnigan Surveyor Plus HPLC system was used for analyzing microdialysis samples. Neurochemical separation was achieved with a Phenomenex (Torrance, CA) Kinetex biphenyl LC column (50 × 2.1 mm, 1.7 μm particle size, 100 Å pore size). Mobile phase A was 10 mM ammonium formate and 0.15% (v/v) formic acid in HPLC water. Mobile phase B was acetonitrile. The mobile phase gradient for all of the analytes was: initial, 0% B; 0.1 min, 10% B; 0.12 min, 10% B; 2.3 min, 20% B; 3.7 min, 50% B; 4.0 min, 80% B; 4.5 min, 100% B; 5.0 min 100% B; 6.5 min, 0% B. The flow rate was 200 μL/min, and the sample injection volume was 7 μL. The autosampler and column were maintained at ambient temperature throughout the analysis. A Thermo Finnigan TSQ Quantum Ultra triple quadrupole mass spectrometer operating in positive mode was used for detection. Electrospray ionization (ESI) voltage was 3.5 kV, and heated ESI probe (HESI-I) was set at 300°C. Capillary temperature was 350°C, and sheath gas, aux gas, and ion sweep gas were maintained at 25, 15, and 0 arb, respectively. The intercycle delay was 200 ms. Automated peak integration was performed using Thermo X Calibur Quan Browser version 2.1. All peaks were visually inspected to ensure proper integration. Calibration curves were constructed based on peak area ratio (P_analyte/P_IS) versus concentrations by linear regression.

Chromatographic separation was performed using an ACQUITY® UPLC® H-Class UPLC system (Waters, USA) with a Kinetex Biphenyl LC column (100 mm × 2.1 mm, 2.6 μm, Phenomenex) preceded by a Security Guard™ ULTRA Cartridge UHPLC Biphenyl 2.1 mm ID column. The mobile phases comprised solvents A and B: 2% and 99.9% methanol in 0.1% formic acid solution, respectively. The gradient elution profile is shown in Table 1. The analytes were then quantified using a Xevo® TQ-XS tandem mass spectrometer (Waters Corporation, Milford, MA US). The mass spectrometer was operated in multiple reaction monitoring modes (MRMs). The MRM transitions and conditions for the analytes and ISs, as well as the retention times, target ions, and qualifier ions used for identification and quantification, are shown in Table 2.