Therefore, each compound was mixed with KBr and compressed under reduced pressure to form a pellet for IR absorbance measurement. However, spectroscopic interference derived from water absorption by the pellet required the use of an alternative method, in which the saponin was dissolved in MeOH, cast onto CaF2 or LiF plates, and allowed to evaporate. Ginsenosides Re (1), Rf (2), Rg2 (3), and 20-gluco-Rf (4) exhibited check details absorption peaks corresponding to the O–H stretching of each hydroxyl group (3359, 3360, 3391, and 3360, respectively), C–H stretching (2929, 2924, 2930, and 2930), C=C stretching (1642, 1637, 1635, and

1635), C–H bending (1072, 1071, 1070, and 1074), and C–O bending (1045, 1031, 1048, and 1032). The multiple hydroxyl groups of ginsenosides also result in very low volatility.

Thus, mass spectra are usually obtained with FAB/MS instead of EI/MS. The soft ionization method FAB/MS distinguishes between molecular ions and fragment ions of relatively smaller proportions. The negative ionization method showed better spectrums for the ginsenosides than a positive ionization method. Ginsenoside Re (1) showed a molecular ion at m/z 945 ([M-H]–) and fragment ions peaks at m/z 765, 475, and 265. The molecular ion of ginsenoside Rf (2) was observed at m/z 799 ([M-H]–) with fragment peaks at m/z 475 and 325. Ginsenoside Rg2 (3) showed m/z 765 ([M-H2O-H]–) as a pseudomolecular ion peak and m/z 281 and 255 as fragment ion peaks. 20-Gluco-ginsenoside Rf (4) revealed a molecular ion peak at m/z 961 ([M-H]–) with a fragment ion peak at m/z 799. NMR spectra were obtained at 40°C from 0.08 M solutions of compounds dissolved in pyridine-d5. Each Alisertib nmr spectrum was the accumulation of eight scans for 1H-NMR and > 1024 scans for 13C-NMR. TMS was used as an internal standard adjusted to 0 ppm. Because ginsenoside Re (1) contains two attached

sugar moieties, it dissolved easily in methanol, pyridine, and DMSO. Pyridine-d5 has few double bonds and many oxygen-linked carbon atoms so it was a better solvent for NMR measurements because it resulted in less overlap between the ginsenoside- and solvent-derived signals than deuterated methanol or DMSO-d6. The methyl carbon atoms C-18, C-19, C-29, and C-30 of ginsenoside Re (1) in pyridine-d5 corresponded to peaks at δC 17.386, 17.628, 17.780, and 17.325, respectively. However, the crotamiton order of the chemical shifts differed from those in the literature [7], [8] and [11]. The carbon signals were confirmed based on cross peaks with corresponding proton signals for C-18, C-19, C-29, and C-30 at δH 1.14, 0.93, 1.33, and 0.92, respectively, in the HSQC spectrum ( Fig. 2A). Cross peaks were also seen in the HMBC spectrum, with H-26 at δH 1.58 showing cross peaks with the carbon signal at δC 17.886 (C-27), and H-28 at δH 2.04 with the carbon signal at δC 17.780 (C-29; Fig. 3A). Methylene proton signals H-15 (δH 0.82, 1.48) and H-22 (δH 1.75, 2.34) differed from the chemical shifts in the literature [5] and [8].