Authors:Yifeng Zhou, Liling Wang, Tiebing Liu, Zedong Mao, Qing Ge and Jianwei Mao
Two pairs of homoisoflavonoid analogues, 6-aldehydo-isoophiopogonanone A (1) / 6-aldehydo-isoophiopogonanone B (2) and methylophiopogonanone A (3) / methylophiopogonanone B (4), were obtained from the fibrous roots of Ophiopogon japonicus (L. f.) Ker-Gawl. (FROJ) by silica gel column chromatography (SGCC) and recycling high-speed counter-current chromatography (rHSCCC). First, the ethyl acetate fraction from the 70% ethanol extract was pre-separated by SGCC with a petroleum ether–ethyl acetate gradient (50:1–2:1, v/v). Then, the two sub-fractions containing homoisoflavonoid analogues were further separated by rHSCCC with n-hexane–ethyl acetate–methanol–acetonitrile–water (3:2:3.5:1:0.5, v/v) and n-hexane–ethyl acetate–methanol–acetonitrile–water (3:2:2.5:1:1.5, v/v). Finally, 6-aldehydo-isoophiopogonone A (16 mg), 6-aldehydo-isoophiopogonanone B (26 mg), methylophiopogonanone A (46 mg), and methylophiopogonanone B (148 mg) were obtained with purities of 97.82%, 96.70%, 97.76%, and 94.62%. Their structures were identified by high-resolution quadrupole-time-of-flight mass spectrometry (HR-QTOF-MS), ultraviolet (UV), and nuclear magnetic resonance spectroscopy (1H-NMR and 13C-NMR). These results demonstrated that rHSCCC could be used for the large scale preparation of homoisoflavonoid analogues from FROJ, which provides scientific support for utilization of untraditional medicinal part of O. japonicus and also for reduction in waste of plant resources. Additionally, an online antioxidant activity assay was investigated with hyphenated HSCCC-DPPH (1-diphenyl-2-picrylhydrazyl) radical scavenging detection.