All ly ophilized samples were stored at space temperature. Column chromatography was performed applying Diaion HP 20P, Sephadex LH 20, MCI gel CHP 20P, and octadecyl silane columns, TLC was performed on pre coated Si gel 60 F254 plates, The 1H and 13C NMR spectra were recorded on an Avance DRX 500 instrument, Electrospray ionization mass spectrometry spectra were obtained on a VG platform electrospray mass spectrom eter, Extraction and isolation The leaves from the regenerated H. pogonocalyx had been macerated with 95% EtOH at area temperature for five days, then filtered to provide the residue and filtrate. The residue was treated inside a similar manner as above three occasions. The combined filtrates have been concentrated below reduced pressure to give the EtOH extract, which was divided into fractions soluble in n hexane, ethyl acetate and H2O by liquid liquid partitioning.
The EtOAc extract was re suspended in H2O, subjected to chromatography on a Diaion HP 20 column, eluted with MeOH H2O and analyzed by thin layer chromatography to obtain seven respective fractions, Fractions E 3 and E four have been passed through a Sephadex LH 20 column to acquire 13 and 11 subfractions, respectively. Re crystallization of fraction selelck kinase inhibitor E three 11 with MeOH yielded compound, Fraction E four 3 was separated by semi preparative HPLC to give compounds and, Fraction E 4 five was separated by semi preparative HPLC to acquire compounds and, Fraction E four six was separated by semi preparative HPLC to provide compound, Compounds and were obtained from fraction E four 7 by semi preparative HPLC, Compound was obtained from fraction E 4 9 by semi preparative HPLC, Fraction E 6 was subjected to an ODS column and eluted with 20 100% MeOH to receive compound, The n butanol extract was eluted on a Sephadex LH 20 column with 100% MeOH to obtain nine frac tions, Right after monitoring by HPLC evaluation, B 6 was subjected to MCI gel CHP 20P column chroma tography.
Fraction B 6 was eluted using a stepwise gradi ent of aqueous methanol, yielding 14 fractions, A precipitate was evi dent inside the B 6 five fraction, Re crystallizing the precipitate with MeOH and H2O yielded pure com pound, Compound was ob tained from B six 14, A precipitate from B 9 was re crystallized irreversible JAK inhibitor with MeOH and H2O to yield pure compound, The spectral data and physical constants for isolated compounds have been in cluded in Supporting details, Antioxidant activities 1, 1 Diphenyl two picrylhydrazyl radical scavenging activity DPPH radical scavenging effect was measured based on the method of Hou et al. Each tested sample was mixed with 160 uM DPPH in an MeOH option. Following a 20 min incubation at room temperature inside the dark, the absorbance was read at 517 nm. The inhibitory percentage of DPPH was calcu lated in accordance with the following equation.