Fig. 2. Conserved and diversification independent feature space for nonulosonic acids (NulOs). (A and C) outline the empirical fragmentation space systematically spanning the oxidation and saturation space of ulosonic acids with different carbon chain lengths. More specifically, (A) outlines (from left to right) one universal fragmentation route for the three main sialic acid compositions (Kdn, Neu and Pse/Leg/Aci). (B) shows the frequency (>50 counts, see ESI-Table sheet 4‡) of binned fragments from ulosonic acid related compounds observed in the large-scale study. This confirmed the theoretically established fragment feature space for different ulosonic acid classes. The high frequency for 283 and 301 (8 C, black bars) and 295 and 297 (9 C, red bars) correspond to fragments from KDO, Neu/Kdn and P.L.A respectively. Furthermore, this provided mass spectrometric evidence for potential higher carbon chain ulosonic acids (blue). (C) The alpha-keto acid specific chemical labelling introduced a significantly altered mass defect and fragment features such as the (quinoxaline-based) ulosonic acid unique core fragments and carbon chain length features. Deviations in the degree of saturation and oxidation allow further differentiation between types of NulOs and the identification of completely new structures.