Lipid neutral glycosphingolipids

Ullman, M.D., McCluer, R.H. Quantitative analysis of plasma neutral glycosphingolipids by high performance liquid chromatography of their perbenzoyl derivatives. J. Lipid Res., 1977, 18, 371-377. 

Isolation of the neutra-L glycosphingolipids. In a typical extraction procedure, 10purified human neutrophils yielded 100-150 mg of total glycosphingolipids. As shown in Table I, glycosphingolipids account for approximately 10% of the total cellular dry weight of the neutrophil. Separation of the total neutrophil lipids by DEAE-sephadex and silicic acid column chromatography yielded 70-100 mg of neutral glycosphingolipids from lO cells. 

The neutral lipid fraction from the DEAE-Sephadex A-50 column was combined with the lower phase obtained after Folch partition of the total lipid extract and the combined lipids dried. To the same flask, 10Q ml of 0.6 M NaOH in methanol was added. The mixture was incubated at 37°C for 5 hours. Five volumes of acetone were then added and stored overnight at 4°C. The precipitate was collected by centrifugation at 4°C and dissolved in C M (4 1, v/v). After application to the column (2.0 x 25 cm), the column was washed with chloroform. Neutral glycolipids were then eluted with tetrahydrofuran H2O (10 1). Fractions containing neutral glycosphingolipids were pooled and their glycolipid content examined by thin-layer chromatography. 

R. S. Duarte, C. R. Polycarpo, R. Wait, R. Hartmann, and E. B. Bergter, Structural characterization of neutral glycosphingolipids from Fusarium species, Biochim. Biophys. Acta-Lip-ids and Lipid Metabolism, 1390 (1998) 186-196. 

The membrane constituents are lipids (phospholipids, glycosphingolipids, and cholesterol Figure 10-5), carbohydrates, and proteins. The ratio of protein lipid carbohydrate on a weight basis varies considerably from membrane to membrane. For example, the human erythrocyte membrane has a ratio of about 49 43 8, whereas myelin has a ratio of 18 79 3. The composition of the normal human erythrocyte membrane is shown in Table 10-2. All membrane lipids are amphipathic (i.e., polar lipids). The polar heads of the phospholipids may be neutral, anionic, or dipolar. The surface of the membrane bears a net negative charge. The distribution of lipid constituents in the bilayer is asymmetrical. For example, in the erythrocyte membrane, phosphatidylethanolamine and phosphatidylserine are located primarily in the internal monolayer, whereas phosphatidylcholine and sphingomyelin are located in the external monolayer. 

For preparation of CMHs, a lipid extract from fungal cells is obtained by successive extractions with chloroform/methanol (2 1 and 1 2 v/v). These extracts are usually combined and dried, yielding a crude lipid mixture. The crude extract is subsequently partitioned according to Folch et al. [27], in which the lower phase containing neutral glycosphingolipids is taken for further analysis. 

Quantitative Determination of Neutral Glycosphingolipids in Urine Sediment J. Lipid Res. 11(1) 31-37 (1970) CA 72 63463k... 

The five major neutral glycosphingolipids of human plasma have been characterized and each has a structure which has been identified previously. One of these glycolipids, D-glucosylceramide, and possibly a second, lactosyl ceramide, exchange between plasma and erythrocyte pools. The conclusion is drawn in terms of the relative roles of carbohydrate and lipid moieties of the glycosphingolipids in maintaining that association with erythrocyte membranes. 

Snyder, P. D., Jr., Krivit, W., and Sweeley, C. C., 1972, Generalized accumulation of neutral glycosphingolipids with GM-ganglioside accumulation in the brain, J. Lipid Res. 13 128-136. 

Glycosphingolipids (GSLs) (neutral GSLs, gangliosides, and complex species, including the ABO blood group substances) constitute about 5-10% of the total lipid. 

Fig. 1 (A) Procedure to obtain acidic and neutral fractions. (B) General structures of lipids analyzed in this study. Glycosphingolipid is...

In the case of acidic glycolipids the relative proton affinity of chemicals can shift the balance for negative ionisation in favor of co-eluted compounds. Pre-analytical separation under acidic conditions serves also to reduce as much as possible the dispersion in the MS spectrum of the metabolite into multiple m/ z representing the various adducts of counterions Na, K, NH4, organic amines, ... which improves sensitivity of the test. Sulfatides are lost during the partition between the hexane and the methanol/water phase. The analysis of sulfatides involves the isolation of the glycosphingolipid fraction and the subsequent separation of sulfatides from neutral lipids by chromatography on DEAE-sephadex or DEAE-cellulose column (the variety of methods are referenced in the website CyberLipid (http //www.cyberlipid.org/). 

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