Thermospray liquid chromatography-mass spectrometry of nucleosides and of enzymatic hydrolysates of nucleic acids.

Nucleosides dissolved in aqueous buffered solutions undergo ionization during direct introduction of the solution into a mass spectrometer using a thermospray interface. The principal ions formed represent the protonated molecule, the corresponding protonated free base, and sugar. In addition to potential utility for characterization of new nucleosides, the technique can be used to monitor nucleosides separated from enzymatic hydrolysates by liquid chromatography. The selectivity of chromatographic detection is significantly greater than with UV absorbance alone so that independent detection of components of unresolved chromatographic peaks is usually possible. Detection limits, with signal/noise greater than 10 for most nucleosides, are approximately 0.1-1 ng per component for selected ion monitoring and 10-50 ng for full-scan mass spectra. Examples are given from the detection of modified nucleosides in enzymatic hydrolysates of 0.05 A260 units (2.5 micrograms) of rabbit liver tRNAVal and of unfractionated H. volcanii tRNA.