Journal of Chromatography B, 947–948 (2014) 1–7 Contents lists available at ScienceDirect Journal of Chromatography B j ourna l h om epage: www.elsevier.com/locate/chromb Rapid and sensitive LC-ESI-MS of gangliosides Aldo D. Garcia, Jorge L. Chavez, Yehia Mechref ∗ Department of Chemistry and Biochemistry, Texas Tech University, Lubbock 79409, TX, USA a r t i c l e i n f o Article history: Received 25 May 2013 Received in revised form 12 November 2013 Accepted 17 November 2013 Available online 25 November 2013 Keywords: Glycolipids Gangliosides LC-ESI-MS Hydrophilic interaction chromatography a b s t r a c t Gangliosides are a class of sphingolipids characterized by a ceramide lipid chain attached to an anionic oligosaccharide moiety that varies in complexity based on the level of sialylation. Heterogeneity in the oligosaccharide chain of gangliosides is a direct result of the monosaccharide structure, content, sequence, and connections. Gangliosides are highly concentrated in the central nervous system, and are cell type- specific as well as development-dependent and their quantities and species can undergo drastic changes during cell differentiation. Specific localization of gangliosides also allows for interaction with a variety of bioeffectors, including glycoproteins, antibodies, peptide hormones, and growth factors. There are currently no rapid analytical assays capable of identifying and quantifying gangliosides. The aim of this study is to establish a reliable chromatographic mass spectrometry based assay capable of profiling ganglioside levels in complex biological samples at high sensitivity. We describe here a chromatographic method using an amino column on which the separation is based on hydrophilic interaction with the sugar moiety of gangliosides. Several gangliosides, including GM1–3, GD1 a,b , GD2–3, and GT1 a,b , were efficiently separated in less than 10 min at a limit of detection ranging between 10–50 pg on column with a concentration dynamic range extending over 4 orders of magnitude. The developed method allowed the sensitive quantitation of gangliosides derived from the blood serum of patients with different esophagus diseases, including, adenocarcinoma, high-grade dysplasia, and Barrett’s. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Gangliosides are a class of sphingolipids characterized by a ceramide lipid chain attached to an anionic oligosaccharide moiety that varies in complexity based on the level of sialyla- tion (N-acetylneuraminic acid or Neu5Ac). Heterogeneity in the oligosaccharide chain of gangliosides is a direct result of the monosaccharide structure, content, sequence, and connections [1]. When taking the variability of the lipid moiety into account, this makes gangliosides a very large family of compounds. Gangliosides are integral constituents of cells and are especially abundant in neuronal membranes. Most importantly, gangliosides are known to serve a wide array of biological functions, including cellular recog- nition, adhesion, and cellular signaling [2]. They have even been reported to play critical structural roles as well. Recently, theoreti- cal considerations and experimental data suggest that gangliosides actively participate in the organization and maintenance of mem- brane lipid domains/zones [3]. Moreover, ganglioside expression is developmentally regulated and is not only cell type specific, but also goes hand in hand with the differentiation state of the cell [4,5]. Multiple investigations have ∗ Corresponding author at: Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409-1061, 806-834-8246806 742 1289. E-mail address: yehia.mechref@ttu.edu (Y. Mechref). also postulated that changes of ganglioside profiles during cellular differentiation is closely related to their metabolism, and in par- ticular their biosynthesis [6]. Interestingly, gangliosides may even have neuroprotective effects to the cell [7–9]. Many studies have also highlighted the neuroprotective effects of GM1 in diseases such as Alzheimer’s disease [10], Parkinson’s disease [11], stroke, and Guillain–Barré syndrome [12]. Recently, gangliosides have also become an attractive target of biomarker discovery because of their biological significance. Even though gangliosides have neuroprotective effects, unusual gan- glioside proliferation can also signify diseases, such as Tay-Sachs, Sandhoff disease, and the AB variant of GM2 gangliosidosis [13–17]. Glycoconjugates have also been associated with tumor develop- ment. In the mid 1980s, it was discovered that glycoconjugates are often expressed as onco-developmental antigens [18]. Today, there is growing evidence to suggest that glycosphingolipids (GSLs) play an important role in tumor biology. For instance, multiple studies have discovered increased levels of GSL-bound sialic acid found in the serum of patients with mammary carcinoma [19], as well as colorectal cancer [20] and melanoma [21]. Abnormally high levels of GM3 and GD3 in the presence of melanoma and elevated levels of GD2 in retinoblastoma patients have also been found through targeted monitoring studies [22]. Establishment of reliable cancer biomarkers is still severely lacking as there is yet no streamlined, facile, rapid, and sensitive assay capable of being applied in a clinical setting. 1570-0232/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jchromb.2013.11.025