Analytical Methods for Lipid: Ceramide
Determination of ceramides by liquid chromatography tandem mass spectrometry
A rapid five minute liquid chromatography tandem mass spectrometry (LC-MS/MS) method operating in multiple reaction ion monitoring mode (MRM) was developed for identification and simultaneous quantification of six ceramides with mass transitions m/z 370/264 (C4), 398/264(C6), 426/264 (C8), 552/264 (C17, IS), 566/264 (C18), 594/264(C20), and 650/264 (C24) (Fig. 1). The method was demonstrated to be specific and sensitive, and a linear response for each of these was observed over a range of 0.01-100 ng/mL with correlation coefficients 91-117, except for C8, C24 and C20 whose lower limit of quantification is 0.1 and 1 ng/mL, respectively. The separation was carried out on a Zorbax C8 3.5u 2x50 mm column at 500C with water containing [A] and acetonitrile/isopropanol (50/50) (both containing 0.2% formic acid, pH 4) as the mobile phase under gradient conditions at a flow rate of 0.75 mL/min. This LC-MS/MS method is very useful for high throughput analysis of ceramides and proved to be simple, sensitive, reproducible, and reliable.