Diaion hp 20

Optical rotations, CD, UV and FT-IR spectra were measured with a Rudolph Research analytical AutoPol automatic polarimeter (Rudolph Research Analytical, NJ, United States), JASCO J-815 CD (Jasco Corporation, Japan), Shimadzu UV spectrophotometer UV-1800 and Thermo Scientific Nicolet iS10 IR spectrometer (Thermo Fisher Scientific, WI, United States), respectively. The structure characterizations of all the compounds were based on 1D NMR (¹H, ¹³C) and 2D NMR (COSY, HSQC, HMBC, 1D-NOE and ROESY) data, recorded on a Bruker AM-400 spectrometer (Bruker BioSpin AG, Switzerland). An Agilent 6,530 Accurate-Mass Q-TOF LC-MS spectrometer (Agilent Technologies, Germany) was used to record high-resolution mass spectra. Preparative RP-HPLC was carried out on an Ultimate 3000 chromatographic system (Agilent Technologies, Germany) with a Phenomenex preparative column (Phenyl-hexyl, 5 μ, 100 × 21.2 mm) and semipreparative RP-HPLC on an Ultimate 3000 chromatographic system (Agilent Technologies, Germany) with a Phenomenex semipreparative column (C₁₈, 5 μ, 250 × 10 mm), connected to a Dionex Ultimate 3000 DAD detector (Agilent Technologies, Germany) (detected at 210, 254, 320, and 365 nm) and a Dionex Ultimate 3000 automated fraction collector. All solvents were HPLC grade. Diaion HP-20 (Alfa Aesar, Japan) was used to run the open-column chromatography.

Optical rotations, CD and IR spectra were measured with a Rudolph Research analytical AutoPol automatic polarimeter (Rudolph Research Analytical, Hackettstown, NJ, USA), JASCO J-815 CD (Jasco Corporation, Tokyo, Japan) and Thermo Scientific Nicolet iS10 IR spectrometer (Thermo Fisher Scientific, Madison, WI, USA), respectively. The structure characterizations of all the compounds were based on 1D NMR (¹H, ¹³C) and 2D NMR (COSY, HSQC, HMBC, 1D-NOE and ROESY) data, recorded on a Bruker AM-400 spectrometer (Bruker BioSpin AG, Faellanden, Switzerland). An Agilent 6530 Accurate-Mass Q-TOF LC-MS spectrometer (Agilent Technologies, Waldbronn, Germany) was used to record high-resolution mass spectra. Preparative RP-HPLC was carried out on an Ultimate 3000 chromatographic system (Agilent Technologies, Waldbronn, Germany) with a Phenomenex preparative column (Phenyl-hexyl, 5 μ, 100 × 21.2 mm) and semipreparative RP-HPLC on an Ultimate 3000 chromatographic system (Agilent Technologies, Waldbronn, Germany) with a Phenomenex semipreparative column (C₁₈, 5 μ, 250 × 10 mm), connected to a Dionex Ultimate 3000 DAD detector (Agilent Technologies, Waldbronn, Germany) (detected at 210, 254, 320 and 365 nm) and a Dionex Ultimate 3000 automated fraction collector. All solvents were HPLC grade. Diaion HP-20 (Alfa Aesar, Japan) was used to run the open-column chromatography.

Teaghrelin was extracted from Shy-jih-Chuen oolong tea according to the method described previously [23 (link)]. A tea sample of 400 g was immersed in 4 L of 50% ethanol aqueous solution for 1 h. After extraction three times, the combined filtrates were concentrated. The crude extract was purified on a Diaion HP20 (Alfa Aesar, Ward Hill, MA, USA) open column and eluted with a stepwise gradient of water and methanol (25, 50, 75, 100%) to generate four fractions. The third fraction (75%) was further purified on a Sephadex LH–20 (GE Healthcare, Chicago, IL, USA) open column eluted with water and a stepwise gradient of methanol (from 10% to 100%) to afford ten subfractions. The sixth and seventh fractions (60 and 70%) were combined and subjected to a LiChroprep^® RP–18 (Millipore, Merck Sigma, MA, USA) open column eluted with water and a step gradient of methanol (30, 70, and 100%) to generate three subfractions. Finally, the second fraction (70%) was collected and analyzed using high-performance liquid chromatography (Waters Corporation, Milford, MA).