Preparative isolation and purification of 12 main antioxidants from the roots of Polygonum multiflorum Thunb. using high‐speed countercurrent chromatography and preparative HPLC guided by 1,1′‐diphenyl‐2‐picrylhydrazyl‐HPLC

Abstract

A hyphenated strategy by off-line coupling of 1,1 ′ -diphenyl-2-picrylhydrazyl-highperformance liquid chromatography, high-speed countercurrent chromatography, and preparative high-performance liquid chromatography was established to screen and separate antioxidants from ethyl acetate fraction of the roots of Polygonum multiflorum. Under the targeted guidance of 1,1 ′ -diphenyl-2-picrylhydrazyl-highperformance liquid chromatography experiment, 12 compounds were identified as potential antioxidants and readily isolated by high-speed counter-current chromatography and preparative high-performance liquid chromatography. Ultraviolet spectroscopy, mass spectrometry, and 1 H NMR spectroscopy were employed to identify their structures, which were assigned as gallic acid (1, 6.2 mg, 98.28%), catechin (2, 8.8 mg, 90.69%), epicatechin (3, 4.1 mg, 96.71%), polydatin (4, 5.3 mg, 94.91%), 2,3,5,4 ′ -tetrahydroxy stilbene-2-O-β-D-glucoside (5, 20.2 mg, 95.23%), piceatannol (6, 5.3 mg, 96.85%), rutin (7, 5.4 mg, 97.92%), resveratrol (8, 5.2 mg, 96.94%), isorhapontigenin (9, 11.4 mg, 94.81%), hyperoside (10, 9.7 mg, 98.52%), rhein (11, 4.9 mg, 97.46%), and emodin (12, 8.2 mg, 95.74%). Notably, compounds 6 and 9 were isolated from Polygonum multiflorum for the first time. In addition, antioxidant activity of compounds 1-12 were evaluated, and compounds 1-8 and 10 exhibited stronger antioxidant activity than ascorbic acid (positive control). These results indicated that the proposed method is a highly efficient strategy to screen and isolate antioxidants from complex natural products. K E Y W O R D Santioxidants, high-speed countercurrent chromatography, preparative high-performance liquid chromatography, traditional Chinese medicine Article Related Abbreviations: DAD, diode array detector; DPPH, 1,1 ′ -diphenyl-2-picrylhydrazyl; HSCCC, high-speed countercurrent chromatography; PEMW, petroleum ether/ethyl acetate/methanol/water; PM, Polygonum multiflorum Thunb.; prep-HPLC, preparative high-performance liquid chromatography.

Cytoprotective Effect of Liposomal Puerarin on High Glucose-Induced Injury in Rat Mesangial Cells

“…To further resolve the components of puerarin extract, high-performance liquid chromatography (Alliance HPLC system, Waters, Milford, MA, USA), equipped with an Atlantis C18 column (150 × 4.6 mm id, 5 μm) was employed. HPLC condition was adapted from previous works [ 25 , 26 ]. Briefly, 100 μL of the sample or standard (1 mg/mL) were injected into HPLC system and eluted with 0.1% formic acid (FA) and acetonitrile (AN) at a flow rate of 1.0 mL/min.…”

Preparative high‐performance liquid chromatography: Isolation of natural chemical compounds for identification and characterization

“…Also worth mentioning are: some alkaloids isolated from the plant Zanthoxylum usabarense and Zanthoxylum chalybeum using ion-pair chromatography [25]; seven caffeoylquinic acid derivatives isolated from ethyl acetate extract of Stemona japonica roots [26] (Figure 2) two new isoindolinone compounds, daldinans B (1) and C (2), two new phthalide compounds, daldinolides A (3) and B (4), and a new naphthoquinone, daldiquinone (5), were isolated together from Daldinia concentrica [27]; 12 main antioxidants from the roots of Polygonum multiflorum Thunb. using high-speed countercurrent chromatography and preparative HPLC guided by 1,1′-diphenyl-2-picrylhydrazyl-HPLC [28]; a new Prenylated Cinnamic Acid Derivative from Brazilian Green Propolis [29]; G-quadruplex ligands from the butanol extract of Zanthoxylum ailanthoides [30]; some lignans from rhizomes of Acorus gramineus [31]; four mayasinoids Maytenus rothiana [32]; novel Catechin-Tiopronin Conjugates Derived from Grape Seed Proanthocyanidin Degradation [33] phloridzin from apple leaves using macroporous [34]. Some enantiomers of the piperidine alkaloid ammodendrin type were also isolated from Lupinus formosus using preparative HPLC column in reversed-phase chromatography after derivation to generate a mixture of diastereoisomers [35].…”

Comprehensive evaluation of the antioxidant capacity of Sceptridium ternatum using multiple colorimetric methods and 1,1‐diphenyl‐2‐picrylhydrazyl–high‐performance liquid chromatography analysis

“…In this study, the antioxidants in ST were efficiently screened by DPPH-HPLC experiment. It was based on the hypothesis that upon reaction with DPPH, the antioxidants in ST would undergo structural changes during the oxidation process, leading to a significant decrease of their peak intensities in HPLC chromatogram [21][22][23]. Comparison of the HPLC chromatograms of DPPH-treated and untreated samples in Fig.…”