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Chemical characterization of the major glycolipids from bovine and starfish spermatozoa Development of new methodologies

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Description: The development of new methodologies for glycolipid analysis including, (1) separation of acidic and neutral glycolipids in a diphasic high-performance thin-layer chromatographic system consisting of an amino-bonded silica gel layer with anion exchange properties interfaced to a neutral silica gel layer (HPTLC); (2) separation and purification of gram quantities of glycosphingolipids by centrifugal partition chromatography (CPC); (3) analysis of sphingolipid sphingoid bases by HPTLC-fluorescence spectrodensitometry following in situ reaction with the Fluorescamine reagent; (4) analysis of free fatty acids by diphasic two-dimensional HPTLC-fluorescence spectrodensitometry following reaction with the fluorescence probe, bromomethyl dimethoxy coumarin (Br-Mdmc). This diphasic HPTLC system consisted of a C18 layer interfaced to a AgNO$\sb3$-modified silica gel layer. The former allowed separation of the Br-Mdmc-derivatized fatty acid esters based on the number of carbons. The latter allowed separation based on the number of double bonds; and (5) analysis of picogram amounts of oligosaccharides by HPTLC-radioscanning, is presented. The oligosaccharides were peracetylated with a mixture of pyridine/ ($\sp3$H) -acetic anhydride (2:1, v/v), separated by normal phase HPTLC, and quantified using HPTLC-radioscanning. All these methodologies in conjunction with one- and two-dimensional (COSY) proton nuclear magnetic resonance spectrometry, fast atom bombardment mass spectrometry, and gas chromatography-mass spectrometric (GC-MS) analysis of the partially methylated alditol acetates and fatty acid methyl esters, were applied to the analysis of the major glycolipids from bovine and starfish spermatozoa. The major glycolipids characterized from bovine spermatozoa were 1-O-hexadecyl-2-O-hexadecanoyl-3-O-$\beta$-D-(3$\sp \prime$-sulfo)-galactopyranosyl-sn-glycerol (seminolipid) and Gal$\alpha$1-4Gal$\beta$1-4Glc$\beta$1-1Cer (globotriaosyl ceramide). The major glycolipid from starfish spermatozoa was Glc$\alpha$1-1Cer containing saturated hydroxylated fatty acids and an unusual sphingoid base with a cis double bond at the 12 position.
Language: English
Format: Degree Work