Quantification of Plasma Glycosphingolipids

Plasma globotriaosylsphingosine (lyso-GB3) was measured by liquid chromatography tandem mass spectrometry (LC-MS/MS) using a published method with minor modifications [20 (link)]. Dimethyl psychosine was used as internal standard. Acquity UPLC BEH C18 column (2.1 mm X 50 mm with 1.7 μm particle size) was used for chromatography. Plasma glucosylsphingosine (lyso-GL1) was measured by a published LC-MS/MS method with minor modifications [21 (link)]. D5-glucosylsphingosine was used as internal standard. Lyso-GL1 is separated from its isomer galactosylsphingosine (psychosine) using Acquity UPLC BEH amide column (2.1 mm X 100 mm with 1.7 μm particle size). Agilent 6490 triple quadruple mass spectrometer coupled with Agilent 1290 liquid chromatography system was used for both analyses. Measurements were performed in cell pellets and culture supernatants as indicated in each figure.