Selekt flash chromatography system

The PBE components were isolated using the methods described in our previous study (S. Lee et al., 2021 ). Briefly, 50.0 g of the aqueous extract was separated on a Diaion HP‐20 gel (Supelco) in a Selekt flash chromatography system (Biotage) using two 340 g Biotage empty cartridges (water:MeOH, 100:0–0:100) to obtain nine fractions (F01 − F09). Of these, F05 (1.55 g) was further fractionated using a flash chromatography system using a 400 g Biotage Sfär C18 D cartridge (water:MeOH, 60:40–0:100), to obtain inosine (122.0 mg) along with 14 subfractions (F0501 − F0514). Furthermore, F0506 (56.1 mg) and F0512 (23.3 mg) were purified using a Waters preparative high performance liquid chromatography (HPLC) system consisting of a 2545Q gradient module and a 2998 photodiode array detector (Waters) equipped with a YMC‐Pack ODS‐A semi‐preparative column (21.2 × 250 mm; 5 μm; 120 Å; water:CH3CN, 98:2–90:10; 6 ml/min; YMC), yielding hypoxanthine (3.4 mg) and uridine (2.9 mg) from F0506, and adenine (2.2 mg) and adenosine (3.1 mg) from F0512. Benzoic acid (7.4 mg) was obtained from F08 (40.1 mg) after purification by preparative HPLC using a YMC‐Pack ODS‐A semi‐preparative column (water:CH3CN, 75:25–60:40; 6 ml/min). Nuclear magnetic resonance (NMR) was performed using a spectraMHz Cryo‐NMR system (Bruker,).

The PBWE (50.0 g) was separated using a Diaion HP-20 gel (Supelco, Bellefonte, PA, USA) in two 340 g scale Biotage empty cartridges (water/MeOH, 100:0 to 0:100) to afford nine fractions (F01−F09) using a Selekt flash chromatography system (Biotage, Uppsala, Sweden). Among the fractions obtained, F05 (1.55 g) was further fractionated employing a flash chromatography system using a 400 g Biotage Sfär C₁₈ D cartridge (water/MeOH, 60:40 to 0:100) to obtain compound 5 (122.0 mg) along with 14 subfractions (F0501−F0514). F0506 (56.1 mg) and F0512 (23.3 mg) were purified using a Waters preparative high-performance liquid chromatography (HPLC) system consisting of a 2545Q gradient module and a 2998 Photodiode array (PDA) detector (Waters, Milford, MA, USA) with a YMC-Pack ODS-A semi-preparative column (21.2 × 250 mm, 5 μm, 120 Å, water/CH₃CN, 98:2 to 90:10, 6 mL/min, YMC, Kyoto, Japan) to yield compounds 2 (3.4 mg) and 3 (2.9 mg) from F0506 and compounds 1 (2.2 mg) and 4 (3.1 mg) from F0512. Compound 6 (7.4 mg) was also obtained after preparative HPLC using a YMC-Pack ODS-A semi-preparative column (water/CH₃CN, 75:25 to 60:40, 6 mL/min) from F08 (40.1 mg). NMR spectra of isolated compounds were acquired using the AVANCE III 600 MHz Cryo-NMR equipment (Bruker, Rheinstetten, Germany).