Tsq endura triple quadrupole ms

The analysis of probe metabolites from CYP‐specific marker reactions was conducted with a LC/MS/MS method modified from earlier works (Turpeinen et al., 2005; Tolonen et al., 2007) and also reported by (Showande et al., 2013). Briefly, a Waters Acquity UPLC system (Waters Corp., Milford, MA) was used together with a Waters HSS C18 column (2.1 mm × 50 mm; 1.8 μm particle size) and an online filter at 35°C. The injection volume was 4 μl, and UPLC eluents were aqueous 0.1% acetic acid (pH 3.2, A) and acetonitrile (B). The gradient elution from 2%–65%–95% B was applied in 0–2.5–3.5 min, followed by column equilibration, giving a total time of 4.5 min/injection. The eluent flow rate was 0.5 ml/min. Data were acquired using a Thermo TSQ Endura triple quadrupole MS. Multiple reaction monitoring (MRM) mode using positive ion mode. For all compounds, the spray voltage was 4500 V, vaporizer temperature and transfer tube temperature were 400°C and 350°C, respectively. The CID argon pressure was set to 2.0 mTorr. The MRM transitions were as previously described (Tolonen et al., 2007; Turpeinen et al., 2005). For acetaminophen, hydroxyrepaglinide and 4‐hydroxydiclofenac MRM's were m/z 152 >  m/z 110, m/z 469 >  m/z 246, m/z 312 >  m/z 231, respectively. The instruments were controlled using Thermo Xcalibur 3.0.63 software.

Saponin extracts were analyzed using an Ultimate 3000 HPLC coupled with a TSQ Endura triple-quadrupole MS (Thermo Fisher Scientific, San Jose, CA, USA) equipped with a Thermo Syncronis C₁₈ column (100 mm × 2.1 mm, 1.7 μm). The mobile phase consisted of ultrapure water with 0.1% (v/v) formic acid (A) and acetonitrile (B). The gradient elution program was carried out as follows: 35–90% A for 0–20 min, 90–35% A for 20–20.5 min, and 35% A for 20.5–25 min at a flow rate of 0.2 mL/min. The column temperature was set at 25 °C and the injection volume was 2 µL. The effluent was delivered to the MS via the ESI ion source with the sheath gas, auxiliary gas, and sweep gas set to 38, 11, and 1 arbitrary unit, respectively. The spray voltage, ion transfer tube temperature, and vaporizer temperature were set at −2500 V, 329 °C, and 296 °C, respectively. Ultrapure argon was introduced as the collision gas, and the collision energy was between 15 and 45 V. MS was operated in full-scan, product ion scan, and MRM modes for the differential, qualitative, and quantitative analysis of the saponins, respectively.