Analysis of carbohydrates in Fusarium verticillioides using size-exclusion HPLC – DRI and direct analysis in real time ionization – time-of-flight – mass spectrometry (DART-MS)

Abstract

Direct analysis in real time ionization – time-of-flight – mass spectrometry (DART-MS) and size-exclusion HPLC – DRI are used, respectively, to qualitatively and quantitatively determine the carbohydrates extracted from the corn rot fungus Fusarium verticillioides. In situ permethylation in the DART beam forms tri-methylammonium adducts of the carbohydrates, which enables positive ion MS detection and analysis of these compounds in either whole fungal extracts or size-exclusion separated HPLC fractions of the extracts. This method detects the disaccharide trehalose at concentrations greater than 3.0 ppm. Additionally, this procedure allows for analysis of glucose as well as the sugar alcohols mannitol, arabitol, and glycerol. The reliability of the DART-MS method is confirmed by the absence of trehalose in a mutant strain of F. verticillioides lacking the gene for trehalose-6-phosphate synthase. The capabilities demonstrated here suggest that the DART-MS method, in conjunction with size-exclusion HPLC, is an effective approach for the analysis of saccharide biomarkers in fungi.