Ab 5500 qtrap

Quantification of PTX was conducted on an LC-MS platform consisting of a Shimadzu High-performance liquid chromatography system and an AB-5500 QTrap (Sciex, Ontario, Canada) tandem mass spectrometer equipped with an electrospray ionization source. The sample injection volume was 5 µL, and chromatographic separation was performed on an Xbridge C18 column (3.5 µm, 50 mm × 2.1 mm internal diameter; Waters, Milford, MA, USA) at a flow rate of 400 µL/min. Mobile phase A consisted of 0.1% formic acid in water, and mobile phase B consisted of 0.1% formic acid in acetonitrile. Gradient elution started with 25% B for 0.5 minute, linearly increased to 65% at 2 minutes, to 95% B at 2.5 minute, kept at 95% B for 2 minutes, decreased to 25% B at 5 minutes, and kept at 25% for 2 minutes. The MS system was operated in positive-ion mode while a multiple reaction monitoring method was used for PTX (m/z 854.40 → 286.10) and IS (m/z 808.00 →226.00) detection. The gas temperature was 300°C with an ion-spray voltage of 5500 V, nitrogen and argon of 60 psi, declustering potential of 190.00 V for PTX and 173.00 V for IS, curtain gas of 30 psi, and collision energy of 21.00 V for PTX and 18.80 V for IS. LC-MS system control, data acquisition, peak integration, and quantitation were performed using the Analyst software package MDS SCIEX (version 1.6; Applied Biosystems, Toronto, Canada).