Sephadex lh 20 gel

The NMR spectra were recorded on Bruker Advance NEO 600 spectrometer (Bruker Corporation, Zurich, Switzerland) using TMS as an internal standard. MS analysis were carried out on Agilent 1100 instrument (Agilent Technologies, Santa Clara, CA, USA) and Thermo ultimate 3000/Q EXACTIVE FOCUS mass spectrometers (Thermo Scientific™, Waltham, MA, USA), respectively. Optical rotations were determined on Rudolph Autopol1 automatic polarimeter (Rudolph Research Analytical, Hackettstown, NJ, USA). UV spectra were detected on a Cary 60-UV-Vis spectrometer (Agilent Technologies, Santa Clara, CA, USA). IR spectra were determined on an iCAN 9 infrared spectrophotometer (Tianjin Nengpu Technology Co., Ltd, Tianjin, China) with KBr disks. X-ray data were generated using a Bruker Smart-1000 CCD (Bruker Corporation, Billerica, MA, USA) area detector diffractometer with graphite monochromatic Cu-Kα radiation. Column chromatography was performed on silica gel (200–300 mesh; Qingdao Puke Parting Materials Co., Ltd., Qingdao, China), Sephadex LH-20 gel (Amersham Biosciences, Uppsala, Sweden). HPLC separation was performed on HITACHI Primaide with an ODS-A column (YMC-pack ODS-A, 10 × 250 mm, 5 μm, 4 mL/min). Melting point instrument (SGW X-4).

Optical rotations were determined on a Perkin-Elmer 341 polarimeter at 20 °C. UV spectra were performed on a Shimadzu UV-2450 spectrophotometer, and ECD spectra were performed on an Applied Photophysics Chirascan spectrometer. Infrared spectra (IR) were measured on a Bruker Tensor 37 infrared spectrophotometer. NMR experiments were carried out on a Bruker AM-400 spectrometer at the temperature thermostatically controlled at 25 °C. Exact mass measurements and molecular formulas were obtained from ESIMS and HR-ESI-MS using a Finnigan LCQ Deca and a Thermo Scientific LTQ Orbitrap XL spectrometers, respectively. Semi-preparative reversed-phase (RP) HPLC was performed with a YMC-pack ODS-A column (10 × 250 mm, S-5 μm) or a Phenomenex Lux cellulose-2 chiral column (10 × 250 mm, 5 μm) under Shimadzu LC-20 AT equipped with a SPD-M20A PDA detector. Column chromatography (CC) was performed on RP-C₁₈ silica gel (S-50 μm, 12 nm, YMC Co., Ltd.), MCI gel (CHP20P, 75–150 μm, Mitsubishi Chemical Industries Ltd.), Sephadex LH-20 gel (Amersham Biosciences), and silica gel (300–400 mesh, Qingdao Haiyang Chemical Co., Ltd.). For RP-HPLC and CC, the analytical grade solvents (Guangzhou Chemical Reagents Company, Ltd.) were employed.

¹H and ¹³C NMR spectra were acquired on a Bruker Avance 600 or a Bruker Avance 400 instrument. A LTQ-Orbitrap XL instrument was used for the determination of high-resolution ESI-MS mass spectra. Semipreparative HPLC using a Shimadzu SPD-20A instrument (Shimadzu, Japan) equipped with an YMC-Pack ODS-A column (250 × 10 mm, 5 μm) was used for sample isolation. Silica gel (200–300 mesh, Haiyang Co., Qingdao, China) and Sephadex LH-20 gel (Amersham Biosciences) were adopted for column chromatography. TLC analysis was performed on precoated silica GF254 plates (Haiyang Co., Qingdao, China).

Optical rotations were measured using an Anton Paar MCP-500 instrument (Anton Paar, Graz, Austria) and the circular dichroism (ECD) as well as the UV spectra were collected on a Jasco 820 spectropolarimeter (JASCO, Tokyo, Japan) in the 200–400 nm range (under N₂ protection). Infrared (IR) spectra were recorded on an IR Affinity-1 spectrophotometer (Shimadzu, Kyoto, Japan). All the 1D and 2D NMR data were recorded on a Avance-III 600 MHz HD spectrometer (Bruker, Bremen, Germany) with tetramethylsilane as an internal standard. HR-ESI-MS were collected on Bruker maXis high resolution mass spectrometer. A Shimadzu LC-20 AT equipped with an SPD-M20A PDA detector was used for HPLC analysis and preparative separations. An ACE 5 PFP-C₁₈ column (250 × 10.0 mm, 5 μm, 12 nm) was used for semipreparative HPLC separation, meanwhile, a CHIRAL-MD (2)-RH column (250 × 10.0 mm, 5 μm) was used for chiral-phase chromatography (Guangzhou FLM Scientific Instrument Co., Ltd., Guangzhou, China). Column chromatography material: commercial silica gel (200–300 mesh) was purchased from Qingdao Marine Chemical Plant (Qingdao, China); Sephadex LH-20 gel was purchased from Amersham Biosciences, Shanghai, China). All analytical grade solvents were purchased from Guangzhou Chemical Regents Company (Guangzhou, China). The natural sea salt was produced by Guangdong Yueyan saltern (Guangdong, China).

Optical rotation was measured on a Perkin-Elmer 341 polarimeter (Perkin-Elmer, Boston, MA, USA). UV spectra were recorded in MeOH on a Perkin-Elmer Lambda 35 UV–vis spectrophotometer (Perkin-Elmer, Boston, MA, USA). 1D and 2D NMR spectra were recorded on a Bruker Advance-500 spectrometer with TMS as internal standard (Bruker BioSpin AG, Fällanden, Switzerland). HRESIMS data were obtained on a Bruker Bio TOF IIIQ mass spectrometer (Bruker Daltonics, Billerica, MA, USA). All solvents were analytical grade (Shanghai Chemical Plant, Shanghai, China). Silica gel (200–300 mesh) was used for column chromatography, and precoated silica gel GF₂₅₄ plates (Qingdao Haiyang Chemical Plant, Qingdao, China) were used for TLC analysis. C₁₈ reversed-phase silica gel (150–200 mesh, Merck), MCI gel (CHP20P, 75–150 μm, Mitsubishi Chemical Industries Ltd.), and Sephadex LH-20 gel (Amersham Biosciences) were also used for column chromatography. TLC spots were visualized under UV light and by dipping into 5% H₂SO₄ in alcohol, followed by heating. In the biological and biophysical experiments, the solvent used to dissolve 1 were DMSO.

All of the chemicals were obtained from commercial sources. Circular dichroism (CD) spectra were measured on a JASCO J-810 instrument (JASCO, Tokyo, Japan). Optical rotations were measured on Autopol VI Polarimeter (Rudolph, NJ, USA). NMR spectra were measured on a Bruker DRX-400 or Bruker DRX-500 spectrometer (BrukerBiospin AG, Fällanden, Germany). Chemical shifts (δ) are reported in ppm with reference to the solvent signals, and coupling constants (J) are in Hz. MS spectra were recorded on a Finnigan-MAT-95 mass spectrometer (FinniganMAT, San Jose, CA, USA). Commercial silica gel (Qingdao Haiyang Chemical Group Co., Ltd., Qingdao, China, 200–300 and 400–600 mesh), and Sephadex LH-20 gel (Amersham Biosciences, Little Chalfont, UK) were used for column chromatography, and precoated silica gel plates (Yan Tai Zi Fu Chemical Group Co., Yantai, China, G60 F-254) were used for analytical TLC (thin layer chromatography). Reversed-phase (RP) HPLC was performed on an Agilent 1260 series liquid chromatography equipped with a DAD G1315D detector at 210 and 254 nm. A semi-preparative ODS-HG-5 column (5 µm, 250 × 9.4 mm) was employed for the purifications. All solvents used for CC and HPLC were of analytical grade (Shanghai Chemical Reagents Co., Ltd., Shanghai, China) and chromatographic grade (Dikma Technologies Inc., Lake Forest, CA, USA), respectively.