*2.4. Elucidation of the Saponin Structures by ESI-MS<sup>2</sup>*

ESI-MS<sup>2</sup> was carried out using CID, creating ion fragments from the precursor ions (Figure 4), and was applied to differentiate the structure of isomeric saponins as described by Song *et al.* [27]. The schematic fragmentation of Lessonioside A, as a representative, and the stepwise structure elucidation of the ion at *m/z* 1477.7 is shown in Figure 4 corroborates results from the MALDI-MS2.

**Figure 4.** *Cont.*

**Figure 4.** Positive ion mode ESI-MS2 spectrum of acetylated saponins detected at *m/z* 1477.7 from Fraction 18. The schematic fragmentation of Lessonioside A as a representative (**A**), and the complete ESI-MS2 fragmentation profile of the ion at *m/z* 1477.7 (**B**). Spectrum (**B**) shows the presence of two different aglycones in the isomeric saponins. Full and dotted arrows illustrate the three main feasible fragmentation pathways. The blue arrows show the decomposition of the isomeric congeners Lessoniosides A, B and D where the green arrows indicate the fragmentation patterns of Lessoniosides C, E, F and G. The ion at *m/z* 451.1 corresponds to the hydrated three sugar units [Xyl-Xyl-MeXyl + H2O + Na].
