Yun Wei

Separation of epigallocatechin and flavonoids from Hypericum perforatum L. by high-speed counter-current chromatography and preparative high-performance liquid chromatography

High-speed counter-current chromatography (HSCCC) and preparative high-performance liquid chromatography (prep-HPLC) were successively used for the separation of epigallocatechin and flavonoids from Hypericum perforatum L. The two-phase solvent system composed of ethyl acetate-methanol-water (10:1:10, v/v) was used for HSCCC. About 900mg of the crude extract was separated by HSCCC, yielding 7.8mg of quercitrin at a purity of over 97%, 12.6mg of quercetin at a purity of over 93%, and 38.9mg of a mixture of hyperoside, isoquercitrin and miquelianin constituting over 97% of the fraction. A mixture of epigallocatechin and avicularin pooled from three HSCCC runs, a total amount of 54.3mg, was further separated by prep-HPLC yielding 23.4mg of epigallocatechin and 15.3mg of avicularin each at a purity of over 97%.

Online isolation and purification of four phthalide compounds from Chuanxiong rhizoma using high-speed counter-current chromatography coupled with semi-preparative liquid chromatography ☆

The phthalide compounds of Chuanxiong rhizoma including senkyunolide A, levistolide A, Z-ligustilide and 3-butylidenephthalide, have been reported as the biologically active compounds because of their therapeutic effects. In this work, online high-speed counter-current chromatography coupled with semi-preparative liquid chromatography instrument was set up, and online separation of the four compounds has been simultaneously achieved using this instrument. In this study, using all the selected solvent system, Z-ligustilide and 3-butylidenephthalide were eluted in one peak by high-speed counter-current chromatography. Using high-speed counter-current chromatography with a solvent system of n-hexane-ethyl acetate-methanol-water-acetonitrile (8:2:5:5:5, v/v), 3.6 mg of senkyunolide A (94.4%) and 3.0mg of levistolide A (95.3%) were obtained from 100mg of the crude extract. Coeluted Z-ligustilide and 3-butylidenephthalide peak fraction (8 mL) from high-speed counter-current chromatography was directly transferred and injected to the semi-preparative liquid chromatography for further separation. Finally, 5.6 mg of Z-ligustilide (97.5%) and 4.8 mg of 3-butylidenephthalide (99.3%) were obtained. The identification of these four compounds was performed by quadrupole time-of-flight mass spectrometer, (1)H and (13)C nuclear magnetic resonance spectrometer.

Enantioseparation of Lomefloxacin Hydrochloride by High-speed Counter-current Chromatography Using Sulfated-β-cyclodextrin as a Chiral Selector

Enantiomers of lomefloxacin hydrochloride were separated by high-speed counter-current chromatography (HSCCC) using sulfated-β-cyclodextrin as a chiral selector (CS). The separation was performed with a two-phase solvent system composed of ethyl acetate-methanol-water (10:1:10, v/v) containing CS at 0-60mmol/l in a head-to-tail elution mode, while obtained fractions were identified by polarimeter and spectropolarimeter. The results show that the concentration of the CS in the system strongly affects the peak resolution (Rs). As the concentration of CS increases, the Rs first increases reaching the maximum at 50mmol/l and then decreases. When the CS concentration is kept constant in the solvent systems, the Rs decreases as the concentration of the lomefloxacin hydrochloride increases. The overall results of our studies indicated that sulfated-β-cyclodextrin is very useful for the chiral separation by HSCCC.

PREPARATIVE ISOLATION OF ISORHAMNETIN FROM STIGMA MAYDIS USING HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY

Abstract Preparative high speed countercurrent chromatography (HSCCC) has been successfully used for the isolation and purification of isorhamnetin from Stigma maydis. This was achieved in two stages: the first separation was performed with a two phase solvent system composed of n-hexane-ethyl acetate-methanol-water (HEMW) at a volume ratio of 5:5:5:5, yielding isorhamnetin at 65.6%, which is followed by the second run using a two phase solvent system composed of HEMW 5:5:6:4, v/v. From 700 mg of the crude extract, 11.8 mg of isorhamnetin was obtained at a high purity of 98%. The final identification was performed by MS, 1H-NMR and 13C-NMR spectra.

ISOLATION OF CAFFEIC ACID FROM EUPATORIUM ADENOPHORUM SPRENG BY HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY AND SYNTHESIS OF CAFFEIC ACID-INTERCALATED LAYERED DOUBLE HYDROXIDE

Preparative high-speed countercurrent chromatography (HSCCC) was successfully used for isolation and purification of caffeic acid from Eupatorium adenophorum Spreng with a solvent system composed of ethyl acetate-methanol-water at a volume ratio of 10:1:10, v/v. Using a preparative unit of the HSCCC centrifuge, about a 938 mg amount of the crude extract was separated, yielding 63.2 mg of caffeic acid at purity of 96.0%. Then, the antimicrobial and antivirus drug caffeic acid (C9H8O4) was intercalated into layered double hydroxides for the first time by anion exchange under a nitrogen atmosphere. The product caffeic acid–LDH has been characterized by powder X-ray diffraction (XRD), Fourier transform infrared (FT-IR), and Scanning electron micrographs (SEM), indicating that the drug has been successfully intercalated into LDH.

Isolation of Imperatorin, Oxypeucedanin, and Isoimperatorin from Angelica dahurica (Fisch. ex Hoffm) Benth. et Hook by Stepwise Flow Rate High‐Speed Countercurrent Chromatography

Abstract Stepwise flow rate preparative high‐speed countercurrent chromatography (HSCCC) was successfully used for isolation and purification of imperatorin, oxypeucedanin, and isoimperatorin from a crude root extract of Angelica dahurica (Fisch. ex Hoffm) Benth. et Hook. The separation was performed with a two‐phase solvent system composed of n‐hexane‐ethyl acetate‐methanol‐water at volume ratio of 5:5:4:6, v/v/v/v, which had been selected by analytical HSCCC. About a 600 mg amount of the crude extract was separated in a one step operation, yielding 35.6 mg of imperatorin, 16.4 mg of oxypeucedanin, and 22.7 mg of isoimperatorin, all at a high purity of over 98%.

ISOLATION OF FIVE BIOACTIVE COMPONENTS FROM EUPATORIUM ADENOPHORUM SPRENG USING STEPWISE ELUTION BY HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY

Semi-preparative high-speed countercurrent chromatography (HSCCC) was successfully performed for isolation and purification of caffeic acid and 4 bioactive flavonoids from Eupatorium adenophorum Spreng using stepwise elution with a pair of two-phase solvent systems composed of ethyl acetate–methanol–water at the volume ratios of 10:1:10 and 5:1:5 (v/v). From 378.5 mg of crude extract 24.1 mg of caffeic acid, 6.7 mg of 4′-methyl quercetagetin 7-O-(6″-O-E-caffeoylglucopyranoside), 6.5 mg of quercetagetin 7-O-(6″-O-acetyl-β-D-glucopyranoside), 31.8 mg of eupalitin 3-O-β-D-galactopyranoside, and 36.7 mg of eupalitin were obtained with purities of 96.0%, 91.2%, 82.3%, 95.1%, and 85.6%, respectively. The structures of the separated compounds were identified by EI-MS, FAB-MS, 1HNMR, and 13CNMR.

Isolation of Hyperoside and Luteolin‐Glucoside from Agrimonia pilosa Ledeb Using Stepwise Elution by High-Speed Countercurrent Chromatography

Abstract Preparative high-speed countercurrent chromatography was successfully used for isolation and purification of hyperoside and luteolin‐glucoside from Agrimonia pilosa Ledeb. The separation was performed by stepwise elution with a pair of two‐phase solvent systems composed of ethyl acetate‐methanol‐water at volume ratios of 50:1:50 and 5:1:5, which had been selected by analytical high-speed countercurrent chromatography (HSCCC). Using a preparative unit of the HSCCC centrifuge, about a 300 mg amount of the crude extract was separated, yielding 7.3 mg of hyperoside and 10.4 mg of luteolin‐glucoside at a high purity of over 97%.

PREPARATIVE SEPARATION OF AXIFOLIN-3-GLUCOSIDE, HYPEROSIDE AND AMYGDALIN FROM PLANT EXTRACTS BY HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY.

Abstract High speed countercurrent chromatography (HSCCC) was successfully used to isolate three bioactive compounds, i.e., amygdalin from bitter almond and taxifolin-3-glucoside and quercetin-3-galactoside (hyperoside) from water extract of A. pilosa Ledeb, respectively. From 1 g of the crude extract 65 mg of amygdalin was isolated at 97% purity using a two phase solvent system composed of ethyl acetate-n-butanol-water (5:2:5, v/v) by preparative HSCCC. From a 400 mg amount of crude extract of A. pilosa Ledeb, 11 mg of taxifolin-3-glucoside and 8 mg of hyperoside were isolated at 96% purity, using a two phase solvent system composed of ethyl acetate-methanol-water (25:1:25, v/v) similarly by preparative HSCCC. The final structural identification was performed by MS, 1H-NMR, and 13C-NMR spectra.

Modelling counter-current chromatography using a temperature dependence plate model

So far, research workers have built several math models of counter-current chromatography (CCC), such as cell model, eluting counter-current distribution model, continuous-stirred tank reactors model, and probability model. Based on plate theory and Vant Hoff equation, a new temperature dependence plate model has been established in this paper. When temperature was taken into consideration, the error of retention time was significantly reduced. The relationship between temperature and partition coefficient K can be summarized into linear equations (logK=A-B/T), which can be introduced to the plate model where A and B are constant and T is the temperature in Kelvin. K values decreased with the increasing temperature. Since resolution values (R) are related to K values, the change of temperature finally leads to the change of resolutions between different compounds. K values at different temperatures of a certain compound in a given system were calculated according to the equation above. New program with a temperature parameter was applied in prediction of experimental results, which diminished the error in modelling prediction. Besides, it is difficult for the CCC work when two compounds have similar K values. However, separation can be achieved by changing temperature, which can be directed by this new model.