Extraction and Purification of R

To begin extraction, dried Sophora japonica was added into a boiling aqueous solution of 0.4% sodium tetraborate. To reach a pH between 8 and 9, calcium hydroxide was added into the solution. After 0.5 h, the solution was filtered with gauze and filter paper, respectively. We observed that the temperature of the solution needed to remain above 60°C in order to avoid unwanted precipitation of R. Next, an appropriate amount of hydrochloric acid solution was added to the filtrate to render the solution at a pH between 2 and 3. After this adjustment, a mass of crude R precipitated from solution. Crude R was then re-dissolved in boiling water, and the hot solution was then quickly filtered again. From this filtrate, the R recrystallized while it cooled. The crystals (purified R) were then collected via vacuum filtration and allowed to dry.

The ¹H NMR spectra of purified R was shown in Supplementary Figure 1, and the data was as follows: ¹H NMR (DMSO-d₆ 600 MHz): 12.594 (1H, s, 5-OH), 10.833 (1H, s,7-OH), 9.670 (1H, s, 3′-OH), 9.176 (1H, s, 4′-OH), 7.548 (2H, m, 2′-H, 6′-H), 6.842 (1H, d, J = 8.16 Hz, 5′-H), 6.386 (1H, d, J = 2.04 Hz, 8-H), 6.195 (1H, d, J = 2.04 Hz, 6-H), 5.344 (1H, d, J = 7.32 Hz, 1′′-H), 4.385 (1H, s, 1′′′-H), 4.346–4.384 and 4.527–5.283 (6H, m, surgar moieties-OH), 3.058–3.714 (10H, m, surgar moieties-H), 0.991 (3H, d, J = 6.18 Hz, surgar moieties-CH₃). The results confirmed R was successfully extracted from Sophora japonica (Nam et al., 2015; Yingyuen et al., 2020).