Fourier transform-infrared (FT-IR) spectroscopy is being developed as a stand-alone method for glycosyl composition analysis and as an adjunct to capillary electrophoretic (CE) separation of the monosaccharide components of glycoconjugates. Early results indicated that FT-IR spectroscopy can be used to differentiate among a wide spectrum of monosaccharide isomers, and that buffers typically used for CE separations do not interfere with IR detection. More recently, a new experimental method has been developed for FT-IR quantitative analysis with the use of FT-IR microscopy. Film spectra of sample solutions deposited and dried on a ZnSe plate were collected with an FT-IR microscope and a high-sensitivity mercury cadmium telluride detector. A fairly extensive library of monosaccharide IR spectra has been collected at high resolution under different conditions. The library is being extended to include di- and higher oligosaccharides. Recent work has focused on the pote ntial of performing glycosyl composition analysis by FT-IR spectroscopy of per-N,O-acetylated derivatives of the monosaccharide methylglycoside mixtures obtained after acidic methanolysis of mono- and oligosaccharides. The peracetylation step was introduced after methanolysis because the monosaccharide methylglycosides were observed to have poor deposition characteristics. Since succesful application of the method will depend on prior acquisition of training spectra from a wide distribution of known samples, including pure monosaccharides as well as standard mixtures, a large number of these samples were generated from combinations of sugars known to comprise N- and O-linked oligosaccharides of mammalian glycoproteins. Spectra from 145 standards of known composition have been acquired, and the method is now being tested on unknown oligosaccharides and monosaccharide mixtures.
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