Xevo triple quadrupole ms

StromaNKtert cells were treated with vehicle, 20 mM R-2HG or 1 mM Octyl-R-2HG for 48 h and the metabolites were extracted from cells as previously described35 (link). R-2HG was measured using UPLC coupled with a Xevo-TQ mass spectrometer. Liquid chromatography was performed on Acquity UPLC system (Waters) using a BEH C18 column (1.7 μm, 2.1 mm × 100 mm, Waters Corporation). UPLC linear gradient conditions were: 0–3 min, 1% B; 3–5 min, 30% B; 6–8.5 min; 99% B; and 9.5–12 min 99% A [solvent system A: water/formic acid (100:0.1, vol/vol); B: acetonitrile/formic acid (100:0.1, vol/vol)]. The injection volume was 2 μL, and the column temperature was maintained at 35 °C. Mass spectrometry detection was performed by using a Xevo™ Triple Quadrupole MS (Waters Corporation) equipped with an electrospray ionisation (ESI) source operating in negative ionization mode. The online MS analysis was at the Multiple Reaction Monitoring (MRM) mode. Parameters for the cone energy and collision energy for R-2HG are: parent ion 147.02 m/z, daughter ion 129.02 (m/z), cone energy 14 V, collision 12 V. Quantification of R-2HG was done using Target Lynx software (Waters Corporation).

BSA224S Precision electronic balance was purchased from Beijing Sartorius Scientific Instrument Co., Ltd. (Beijing, China). KQ-500VDE double frequency digital ultrasonic cleaning instrument was purchased from Kunshan Ultrasonic Instrument Co., Ltd. (Kunshan, China). Chromatographic analysis was performed on a Waters Acquity UHPLC system (Waters, Corp., Milford, MA, USA), consisting of a binary pump solvent management system, an online degasser, and an autosampler. Mass spectrometry detection was performed using a Xevo Triple Quadrupole MS (Waters Corp., Milford, MA, USA) equipped with an electrospray ionization source (ESI). The ESI-MS spectra were acquired by using multiple reaction monitoring (MRM). Inorganic elements analysis was performed on Agilent 7800 ICP-MS system (Agilent Technologies Inc., USA). CEM MARS6 microwave digestion apparatus was purchased from BERGHOF Co., Ltd. (CEM MARS6, Berghof Co., Germany).