Compound 1, a phenolic acid, is commonly found in plants and is a known antioxidant, metal chelator, and AR inhibitor in vitro 
Compound 1, a phenolic acid, is commonly found in plants and is a known antioxidant, metal chelator, and AR inhibitor in vitro . widely in the northeast of China as well as in the southern part of the Russian Far East and North Korea. The dried stem bark of this plant has been used as folk medicine for the treatment of cancer, cholecystitis, and arthritis . In previous reports, MA has been found to contain various flavones, isoflavones, stilbenes, pterocarpans, dimeric stilbenes, and so forth [8, 9]. To identify and isolate bioactive components from natural products, bioassay-guided fractionation is commonly used; however, this old-fashioned approach is time-consuming and labor intensive . Since the advent of high-throughput screening in the early 1990s, offline HPLC-based activity profiling has been proposed and implemented for the effective tracking of bioactive compounds in natural product extracts. Consequently, in recent times, many offline HPLC-based assays have been developed, and many studies have reported successful application of target isolation, that JUN is, offline HPLC-ABTS assay and microfractionation bioassays . To date, however, no data are available on the inhibitory effects of the MA and its constituents on AR. Therefore, as CD-161 part of our continuing search for new AR inhibitors and antioxidants from natural products, we were to investigate the antioxidant and AR inhibitory effects of the dried stem bark of MA and isolate its active components using high-speed countercurrent chromatography (HSCCC) and Sephadex LH-20 column chromatography guided by an offline HPLC-ABTS assay and HPLC microfractionation AR assay. 2. Materials and Methods 2.1. General Points 1H and 13C NMR spectra and correlation 2D NMR spectra were obtained from a Bruker Avance DPX 400 (or 600) spectrometer. These spectra were obtained at operating frequencies of 400?MHz (1H) and 100 (or 150)?MHz (13C) with CD3OD, (CD3)2SO, (CD3)2CO, or D2O and TMS used as an internal standard; chemical shifts were reported in values. The molecular mass was measured using the Voyager DE STR matrix assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometer (MS, Applied Biosystems, Foster City, CA, USA), the low resolution-electronic impact (EI) MS equipped JMS-700 (Tokyo, Japan). Fast atom bombardment (FAB) MS was recorded in the negative form usingmnnnnvalue of the target compound. The value was calculated as the peak area in the upper phase divided by the lower layer and the upper CD-161 stage was utilized as the fixed phase, and the low phase was utilized CD-161 as the cellular stage. 2.5.3. High-Speed Countercurrent Chromatography (HSCCC) The HSCCC device was a model TBE-1000A HSCCC (Tauto Biotechnique Firm, Shanghai, China) with three multilayer coil columns (from the tubes: 1.8?mm, column quantity: 260?mL) connected in series and a 50?mL test loop. The worthiness (= may be the distance in the coil towards the holder shaft and may be the distance between your holder axis and central axis from the centrifuge) from the multilayer coil varies from 0.60 (internal terminal) to 0.80 (exterior terminal). The trend speed from the equipment was controlled at 0C1000?rpm with an electric quickness controller. The HSCCC program was built with a Model Hitachi L-6200 smart pump (Hitachi, Tokyo, Japan) and an Isolera Display purification CD-161 program (Biotage, Uppsala, Sweden) as UV monitor. The multilayer coil column was initially entirely filled up with top of the organic stage at a stream price of 20?mL?min?1. The low aqueous stage was pumped in to the inlet column as the cellular stage at 5?mL?min?1, as the apparatus was rotated in 400?rpm. The setting for HSCCC parting was check out tail. Following the hydrodynamic equilibrium was set up, the EtOAc small percentage of the MA remove (2?g in 40?mL of every stage) was injected in to the parting column through the shot valve, and each top fraction was collected in 25 then?mL pipes while monitored using a UV detector in 254?nm. 2.5.4. Sephadex LH-20 Column Chromatography A cup column (90?cm 3?cm we.d.) was filled with Sephadex LH-20 gel in 60% methanol at area temperature. After that, 0.49?g from the EtOAc small percentage of the MA remove in 1.5?mL of 60% methanol was loaded towards the column and eluted. 2.6. Evaluation of Trolox Similar Antioxidant Capability (TEAC) ABTS radical scavenging activity was examined by changing a previously defined process [12, 13]. A 2?mM.