Securityguard c18 guard column

Tumors and plasma samples were prepared and paclitaxel concentrations were analyzed by LC-MS/MS. Briefly, the samples were extracted with 100% acetonitrile containing carbamazepine as an internal standard, and chromatography was conducted on an Xterra C18 column (50 × 2.1 mm i.d., 5 μm, Waters, Milford, MA, USA) with a SecurityGuard C₁₈ guard column (2.0 × 4.0 mm i.d., Phenomenex, Torrance, CA, USA) maintained at room temperature. The mobile phase was 95% v/v solvent A (deionized water containing 0.1% v/v formic acid)/5% v/v solvent B (acetonitrile containing 0.1% v/v formic acid) at a flow rate of 0.4 mL/min. A linear gradient of the two solvents was used: start at 95% A and hold for 0.5 min, ramp to 5% A to 0.6 min, and hold until 4 min. The flow rate was 0.4 mL/min throughout the gradient. The retention times of paclitaxel and the internal standard (IS) were 3.0 and 2.8 min, respectively. The electrospray ionization source was operated at 5500 V and 550°C. The samples were analyzed via multiple reaction monitoring. The monitoring ions were set as m/z 876 → 308 for paclitaxel and m/z 237 → 194 for the IS. The scan dwell time was 0.1 sec for each channel. Acquisition and analysis of data were performed using the Analyst software ver. 1.5.2 (Applied Biosystems, Foster City, CA, USA).

Samples resulting from liver microsomal incubations were analyzed by LC-MS/MS. Aliquots of the supernatants were injected onto an X-Bridge C18 column (2.1 × 50 mm, 3.5-μm particles) (Waters, Milford, MA), preceded by a SecurityGuard C18 guard column (4.0 × 2.0 mm; Phenomenex Inc.). Separations were accomplished with a gradient of 0.1% acetic acid in water (v/v) versus 0.1% acetic acid in acetonitrile: methanol (75:25, v/v) at a rate of 0.8 mL/min. The eluents were further applied into an equipped API 4000 QTrap mass spectrometer (AB Sciex, Foster City, CA) with electrospray ionization (ESI) interface by two coupled LC-10 AT pumps. Peak area ratios of FTY720-C2 or FTY720-Mitoxy against the C17 sphingosine internal standard were used to determine the amount of parent remaining compared to that at 0 min. The percent of FTY720-C2 or FTY720-Mitoxy remaining was calculated by dividing the peak area ratio obtained at each time point by that obtained at 0 min. The in vitro intrinsic clearance (CL_int) was calculated by linear regression of the log percent of compound remaining versus time plots using Microsoft Excel, two independent experiments (n = 2) were performed for each compound and species. Then, the mean and standard deviation (SD), of two values for each condition, were calculated using Microsoft Excel with the “AVERAGE” and “STDEV” functions, respectively.