2.4. Ceramide Quantification
dryness
Plasma samples (700 µL) were frozen over liquid nitrogen and lyophilized to dryness. HPLC-grade methanol (1.4 mL) was added to the dried material and the mixture was sonicated for 3 min (on/off pulse cycles of 3 s on, 2 s off, at power 100%) using a Qsonica Ultrasonic Processor (Model Q700; Qsonica, Newtown, CT, USA) with a water bath cup horn adaptor (Model 431C2) and the water bath maintained at room temperature. Samples were placed on an end-over-end rotator, and extraction continued for 24 h, then centrifuged at 18,000× Ceramide analyses were performed using a Thermo Scientific Vanquish Horizon UHPLC System coupled with a Thermo Scientific TSQ Quantis Triple Quadrupole mass spectrometer equipped with a HESI ion source. Mobile phase A was 78.6% water, 20% acetonitrile, and 0.4% formic acid (