Extraction and Purification of UDP-Sugars

Samples were applied to Supelclean™ ENVI™-Carb SPE Tubes (3 ml, 0.25 g, particle size: 120–400 mesh) (Supelco) equilibrated with 3 ml equilibration-elution buffer (60% acetonitrile in water containing 0.3% formic acid adjusted to pH 9 with ammonia) and flushing with 3 ml of water. After sample application, washing steps were performed, first with 3 ml of water and afterwards with 1 ml wash buffer (60% acetonitrile in water). Sample elution was done by flushing the column with 2 ml equilibration-elution buffer [29] (link). The aqueous phase containing the UDP-sugars was vaporized by under vacuum in a concentrator 5301 (Eppendorf) at 30°C and reconstituted in 100 µl water.