Quantification of Inhaled Anesthetics via UPLC/MS

The concentrations of dissolved isoflurane and sevoflurane were determined by liquid chromatography with mass spectrometry detection (UPLC/MS). Culture medium was collected immediately following exposure to isoflurane and sevoflurane in air-tight glass containers without air space as we previously performed^{20 (link)}. Samples were diluted in mobile phase comprised of Part A pure methanol MS grade and Part B 0.1% formic acid in MilliQ water. All samples were assayed with an Acquity H-Class UPLC with a XEVO TQ triple quadrupole mass spectrometry detector (Waters CO., Milford, MA). We used an Acquity UPLC BEH 18 1.7 μm 2.1 x 100 mm column, with a VanGuard Pre-Column 2.1 x 5 mm guard. The mobile phase was run on a gradient from 95% B to 90% A for 5.5 minutes. The flow rate was 0.3 mL/min and the injection volume was 3 μL. The detection was carried out by Multiple Reaction Monitoring (MRM) under positive mode. For isoflurane we used the transitions 185.067 > 99.867 and 185.067 > 143.927, while for sevoflurane we used 201.039 > 153.867 and 201.039 > 168.896. Data acquisition and instrument control was done by Masslynx 4.1 software (Waters Co.). The accuracy and precision (% coefficient of variation) were 97.2-99.5% and 1.8-4.7%, respectively.