3.5. Cholesterol Analysis Using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) with Atmospheric Pressure Chemical Ionization (APCI)

A 1290 Infinity II HPLC system (Agilent Technologies, Santa Clara, CA, USA) equipped with a tandem mass spectrometer (QTRAP, 6500, AB SCIEX, Framingham, MA, USA), an electron multiplier detector, and a Poroshell 120 EC-C18 (4.6 × 75 mm, 2.7 µm, Agilent Technologies, Santa Clara, CA, USA) was used [44]. Multiple reaction monitoring (MRM) mode was used for cholesterol identification. The mobile phase consisted of 100% acetonitrile, and 5 µL of the sample was injected. The vacuum pump pressure was set to 2.1 torr, and nitrogen gas was used for fragmentation of precursor ions. Selected characteristic product ions generated from the precursor ions are shown along with collision energies in Table 2. An organic extract phase (12.5 mL) was evaporated to dryness under nitrogen and re-dissolved in methanol (3 mL) before injection.