Enhancement of the Yu and Ledeen gas-liquid chromatographic method for sialic acid estimation: Use of methane chemical ionization mass fragmentography

J. Ashraf, University of Kentucky
D. A. Butterfield, University of Kentucky
J. Jarnefelt, University of Kentucky
R. A. Laine, University of Kentucky


The sialic acid present in erythrocyte ghosts was estimated by methane chemical ionization mass fragmentography of the trimethylsilyl methyl glycosides. The internal standard was 3,4,6-tris-trimethylsilyl-α-phenyl-2-deoxy-2-acetamide-D-glucosaminide, as proposed by Yu and Ledeen. The [MH-16]+ions (m/e 610 for the TMS-methylglycoside of N-acetylneuraminic acid and m/e 498 for the internal standard) were used for quantifying nanogram levels of sialic acid in the presence of other contaminating substances. This effectively raises the signal-to-noise ratio for the Yu and Ledeen method by at least two orders of magnitude. The sensitivity and linearity of the method, without use of isotopic carriers, were tested using known quantities of N-acetylneuraminic acid. The limit of detection was below 0.4 nanograms (approximately one picomole). The useful range of detection was 10 ng-1 μg, showing a large dynamic range.