4000 qtrap hybrid triple quadrupole linear ion trap lc ms ms system

Adduct analysis was performed using a 4000 QTRAP hybrid triple quadrupole/linear ion trap LC-MS/MS system (AB Sciex, Redwood City, CA) interfaced with a Flexar UHPLC system (Perkin-Elmer, Waltham, MA) using an Acuity UPLC 1.7 µm BEH C₁₈, 100 × 2.1 mm column (Waters, Milford, MA). The UHPLC solvent method utilized a gradient of solvent A (5 mM NH₄OAc + 0.1% formic acid in HPLC grade water) and solvent B (acetonitrile) at a flow rate of 500 µl/min. The 7.5 minute gradient was as follows: 1) 0.5 minute 80% A and 20% B; 2) 6 minutes to 10% A and 90% B; 3) 0.1 min 10% A and 90% B; 4) 0.9 minute wash out 10% A and 90% B. The MS parameters were set as follows: electrospray ionization – positive mode; electrospray source temperature – 600°C; declustering potential – 60 eV; collision energy – 30 eV; entrance potential – 10 eV; cell exit potential – 10 eV; collision activated dissociation gas – high; curtain gas – 20 psi. Adducts were analyzed in multiple reaction monitoring (MRM) mode with the following transitions 604.2/335.0, 604.2/317.0, 604.2/289.1 (targeted adducts) and 609.2/335.0, 609.2/317.0, 609.2/289.1 (internal standards). Peak integration was performed using Analyst 1.5.2 Software (AB Sciex, Redwood City, CA).