A new liquid chromatography/tandem mass spectrometry method for quantification of gangliosides in human plasma

Abstract

Gangliosides are a family of glycosphingolipids characterized by mono- or polysialic acid-containing oligosaccharides linked through 1,3- and 1,4-β glycosidic bonds with subtle differences in structure that are abundantly present in the central nervous systems of many living organisms. Their cellular surface expression and physiological malfunction are believed to be pathologically implicated in considerable neurological disorders, including Alzheimer and Parkinson diseases. Recently, studies have tentatively elucidated that mental retardation or physical stagnation deteriorates as the physiological profile of gangliosides becomes progressively and distinctively abnormal during the development of these typical neurodegenerative syndromes. In this work, a reverse-phase liquid chromatography/tandem mass spectrometry (LC/MS/MS) assay using standard addition calibration for determination of GM2, GM3, GD2, and GD3 in human plasma has been developed and validated. The analytes and internal standard were extracted from human plasma using a simple protein precipitation procedure. Then the samples were analyzed by reverse-phase ultra-performance liquid chromatography (UPLC)/MS/MS interfaced to mass spectrometry with electrospray ionization using a multiple reaction monitoring mode to obtain superior sensitivity and specificity. This assay was validated for extraction recovery, calibration linearity, precision, and accuracy. Our quick and sensitive method can be applied to monitor ganglioside levels in plasma from normal people and neurodegenerative patients.

Quantitative GSL-glycome analysis of human whole serum based on an EGCase digestion and glycoblotting method

“…Kundu et al ( 15 ) observed 25 GSLs consisting of 15 acidic gangliosides and 10 neutral GSLs in human plasma by TLC separation and gas-liquid chromatography analysis. Recent advances in MS analysis allowed quantitation of the intact forms of targeted GSLs using the multiple reaction monitoring mode (16)(17)(18). However, efforts to clarify the comprehensive GSL-glycomics of human serum have been limited since the characterization of serum GSLs using TLC and MS.…”

Profiling gangliosides from milk products and other biological membranes using LC/MS

“…The third quadrupole is set to allow sialic acid to pass and to collect fragments. Triple quadrupole MS with MRM has been recently used due to the high specificity for simultaneous detection of multiple components in complex samples (Zhang et al, 2012;Lee et al, 2013;Giuffrida et al, 2014;Huang et al, 2014).…”

Apprehending ganglioside diversity: a comprehensive methodological approach

“…On the contrary, other authors reported a suppression of GG precursor ion signal to ‫ف‬ 14% of the original intensity by addition of dioleoylphosphatidylcholine in the analyte ( 35 ). This result was corroborated by Huang et al ( 36 ) who also showed important suppression of the GG signals (from 22% to 51% extinction for GM2, GM3, GD2, and GD3) due to contaminants present in the extracts prepared by simple protein precipitation procedure by comparing MS signals of spiked GG standards in analyte-free extracts and in pure solvents. To avoid any potential suppression, we preferred a sample extraction aiming at depleting phospholipids and enriching the extract with GGs.…”

“…Most of the studies published in the literature do not actually use any internal standard (44)(45)(46). Some have used GM3d3 for various GG classes (GM3, GM2, GD3, and GD2) in human plasma ( 36 ) or GM1d3 for GM2 in cells and plasma ( 47 ), while others have used phospholipids ( 30,31 ). Recently, authors reported the synthesis of GG-bearing nonnatural FAs for the major classes of mammalian brain and also some tumorassociated classes, which will be valuable tools to use as internal standards for MS analyses ( 48 ).…”