Rp 18f254 plates

Infrared spectra (IR) were obtained with a Thermo Fisher Scientific (Waltham, MA, USA) NICOLET iS5 FT-TR spectrometer from samples in CHCl₃ and MeOH. NMR spectra were measured on JEOL (Akishima, Tokyo, Japan) JNM-ECA600 and JNM-ECS400 spectrometers with tetramethylsilane as an internal standard, and chemical shifts are stated as δ values. HRMS data were recorded on a JEOL JMS-700 Mstation (FAB or EI) mass spectrometer. Analytical and preparative TLC were carried out on precoated silica gel 60F254 and RP-18F254 plates (0.25 or 0.50 mm thickness; Merck, Darmstadt, Germany). MPLC was performed on a Combiflash R_f (Teledyne Isco, Lincoln, NE, USA) with silica gel and C18 cartridges (Biotage, Uppsala Sweden). Preparative HPLC was carried out with a GL Science (Shinjuku, Tokyo, Japan) recycling system (PU714 pump and UV702 UV-Vis detector) using an InertSustain C18 column (5 µM, 20 × 250 mm).

One- and two-dimensional spectra of CC were obtained using Bruker Avance DRX Spectrometer (Billerica, MA, USA), with chemical shifts reported in ppm (δ). Mass spectrometry analyses were performed using electrospray ionization. Purification of the extract was achieved via open-column chromatography on silica gel (63–200 µm) and subsequent thin-layer chromatography using Merck Silica Gel 60 F254 and RP-18 F254 plates (Merck, Rahway, NJ, USA). Visualization of isolated compounds employed a standard protocol with 10% aqueous H₂SO₄ and heating for approximately 3 min.

Melting points were determined on a Yanaco Micro MP apparatus. Optical rotations were measured on a JASCO P-2100 polarimeter (Hachioji, Tokyo, Japan). Infrared spectra were recorded on JASCO FT/IR-460 Plus spectrometer. 1D and 2D NMR were carried out using Bruker Avance 500 spectrometer (Bruker, Mass., Billerica, MA) with tetramethylsilane as an internal standard. ESI-MS data including high-resolution mass spectrum were measured on Shimadzu LCMS-IT-TOF spectrometer (Kyoto, Japan) HREIMS was recorded on a JEOL MStation JMS-700 spectrometer (JEOL Ltd., Tokyo, Japan). Column chromatography was performed using silica gel (Kanto, 40–50 μm, Tokyo, Japan), Cosmosil 75C18-OPN (Nacalai Tesque Inc., Kyoto, Japan) and Sephadex LH-20 (Dowex® 50WX2-100, Sigma-Aldrich, Tokyo, Japan). Analytical TLC was performed on pre-coated silica gel 60F₂₅₄ and RP-18 F₂₅₄ plates (0.25 or 0.50 mm thickness, Merck KGaA, Darmstadt, Germany). Cosmosil 5C18-AR-II (Nacalai Tesque Inc., Kyoto, Japan) was used for analytical and semi-preparative HPLC (250 × 4.6 mm for analytical HPLC, and 250 × 10.0 mm for semi-preparative HPLC).

Silica gel 60 GF254 (Qingdao Haiyang) was used for vacuum liquid chromatography. Thin-layer Chromatography (TLC) was carried out using silica gel 60 F254 and RP-18 F254 plates (Merck). High-performance liquid chromatography (HPLC) was conducted using a Shimadzu system equipped with a LC-6 AD pump and a Diode Array Detector (SPD-M20A), as well as a Zorbax Eclipse XDB-C18 column (9.4 × 250 mm, 5 μm particle size, Agilent); mobile phase acetonitrile – water (9:1 v/v); flowrate 1 mL/min, injection volume 500 μL, wavelength 254 nm and 365 nm. HPLC solvents were purchased from Merck.