Direct quantitation of psychosine from alkaline-treated lipid extracts with a semi-synthetic internal standard

Abstract

Psychosine is an important bioactive sphingolipid metabolite and plays an essential role in the pathogenesis of Krabbeʼs disease. Herein, we extended shotgun lipidomics for the characterization and quantitation of psychosine in alkaline-treated crude lipid extracts by using neutral loss scan of 180 amu (i.e., galactose) in the positive-ion mode. Specifically, we semi-synthesized N,N-dimethyl psychosine and used it as an internal standard for quantitation of psychosine. After characterization of the fragmentation patterns of psychosine and the selected internal standard and optimization of the experimental conditions, we demonstrated that a broad linear dynamic range for the quantitation of psychosine and a limit of detection at a concentration of low fmol/ml were achieved using this approach. The developed method is generally simpler and more efficient than other previously reported methods. Multiple factors influencing quantitation of psychosine were extensively examined and/or discussed. The levels of psychosine in diabetic mouse nerve tissue samples were determined by the developed methodology. Collectively, the developed approach, as a new addition to the shotgun lipidomics technology, will be extremely useful for understanding the pathways/ networks of sphingolipid metabolism and for exploring the important roles of psychosine in a variety of physiological and pathological conditions.-Jiang, X., K. Yang, and X. Han. Direct quantitation of psychosine from alkaline-treated lipid extracts with a semi-synthetic internal standard.

Bone Marrow Transplantation Augments the Effect of Brain- and Spinal Cord-Directed Adeno-Associated Virus 2/5 Gene Therapy by Altering Inflammation in the Murine Model of Globoid-Cell Leukodystrophy

“…Psychosine levels were quantified in the brains and spinal cords at postnatal day 36, essentially as described previously (Jiang et al, 2009). Briefly, tissues were flash-frozen, pulverized, and weighed.…”

“…Lipids were then dried under nitrogen and resuspended in 1:1 chloroform/methanol. This sample was analyzed by mass spectrometry as described previously (Jiang et al, 2009). Psychosine concentration was calculated by comparing the psychosine peak intensity to that of the internal standard.…”

Selection of internal standards for accurate quantification of complex lipid species in biological extracts by electrospray ionization mass spectrometry—What, how and why?

“…The concentration where lipids start to aggregate is estimated through determination of the linear dynamic range for quantification of lipid of interest as described previously (Jiang, Yang, & Han, ). In the linear dynamic range, the upper concentration where regression deviates from linearity represents the concentration where the lipid species begin to form aggregates.…”

A HILIC‐MS/MS method for simultaneous quantification of the lysosomal disease markers galactosylsphingosine and glucosylsphingosine in mouse serum

“…The fragmentation patterns of GalSph and GluSph in collision‐induced dissociation are nearly indistinguishable, and have been studied previously (Dekker et al, ; Jiang et al, ). The two most abundant product ions at m / z 282.3 and 264.3 resulted from the neutral loss of a galactose and a galactose plus a water molecule, respectively.…”