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1.
Biomed Chromatogr ; 34(4): e4788, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31899545

RESUMO

The objective of this paper was to develop a preparative method for the isolation and purification of liquiritigenin and glycyrrhetic acid from Glycyrrhiza uralensis Fisch using hydrolytic extraction combined with high-speed countercurrent chromatography (HSCCC). Liquiritigenin and glycyrrhetic acid were well hydrolyzed from liquiritin and glycyrrhizic acid by hydrochloric acid, respectively. The optimal extraction conditions were obtained by single-factor and orthogonal experiments, which were 100% ethanol, 1.5 mol/L hydrochloric acid, 1:25 ratio of solid to liquid, and extracted 2 h for one time. Using the two-phase solvent system of n-hexane-ethyl acetate-methanol-water (4:5:4:5, v/v), 2.1 mg liquiritigenin (the purity was 96.5% with a recovery of 87.6%) and 12.3 mg glycyrrhetic acid (the purity was 97.1% with a recovery of 74.4%) were obtained from 315-mg crude extraction by HSCCC. The retention ratio of stationary phase was 47.2%. Their structures were identified by HPLC, melting points, UV, Fourier-transform infrared, Electrospray ionization-MS, 1 H nuclear magnetic resonance (NMR), and 13 C NMR spectra. According to the antioxidant activity assays, liquiritigenin and glycyrrhetic acid had some scavenging abilities on 1,1-diphenyl-2-picrylhydrazyl free radicals; liquiritigenin had stronger scavenging ability on hydroxyl radicals.


Assuntos
Distribuição Contracorrente/métodos , Flavanonas/isolamento & purificação , Ácido Glicirretínico/isolamento & purificação , Glycyrrhiza uralensis/química , Extração Líquido-Líquido/métodos , Antioxidantes/química , Antioxidantes/isolamento & purificação , Flavanonas/química , Ácido Glicirretínico/química , Extratos Vegetais/química
2.
J Chromatogr A ; 1617: 460834, 2020 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-31916986

RESUMO

Sertraline is an antidepressant in a group of drugs called selective serotonin reuptake inhibitors. Four stereoisomeric compounds would be produced in its synthetic preparation due to two chiral carbons on its chemical structures. In the present work, stereoselective liquid-liquid extraction of isomeric sertraline with substituted cyclodextrins as stereoselective extractant was investigated. Factors affecting the distribution performance, including organic solvents, types of extractants, pH value, buffer solution of aqueous phase, concentration of extractant and temperature, were investigated. Under optimized conditions, a stereoselectivity of 1.404 was obtained for cis-sertraline and a stereoselectivity of 2.373 was obtained for trans-sertraline when hydroxypropyl-ß-cyclodextrin was used as the stereoselective extractant, and a stereoselectivity of 1.685 was achieved for trans-sertraline with methyl-ß-cyclodextrin as extractant. An unusual stereoselective combination was observed for trans-sertraline and cis-sertraline when sodium carbonate buffer was used. Successful stereoselective separation of trans-sertraline and cis-sertraline, (1S, 4R) and (1R, 4S)-sertraline by analytical countercurrent chromatography with methyl-ß-cyclodextrin as stereoselective selector was achieved, using a biphasic solvent system composed of n-hexane : 0.1 mol L-1 citrate buffer solution with pH7.6 (1:1, v/v).


Assuntos
Antidepressivos/química , Distribuição Contracorrente/métodos , Inibidores de Captação de Serotonina/química , Sertralina/química , 2-Hidroxipropil-beta-Ciclodextrina/química , Antidepressivos/isolamento & purificação , Ciclodextrinas/química , Extração Líquido-Líquido , Inibidores de Captação de Serotonina/isolamento & purificação , Sertralina/isolamento & purificação , Solventes , Estereoisomerismo , beta-Ciclodextrinas/química
3.
Planta Med ; 86(2): 151-159, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31777054

RESUMO

The biological properties of Achyrocline satureioides have been mostly ascribed to its major flavonoids quercetin (QCT), luteolin (LUT), and 3-O-methylquercetin (3OMQ). The present study aimed to optimize the extraction by dynamic maceration of the major phenolic compounds in order to obtain in a subsequent step a flavonoid-enriched fraction (FEF) using high performance countercurrent chromatography (HPCCC). A 3-level Box-Behnken design (BBD) was applied to maximize the extraction of the substances, using the plant : solvent ratio (X1 ), extraction time (X2 ), and ethanol concentration (X3 ) as factors. One-step HPCCC semipreparative separation with a solvent system composed of hexane : ethyl acetate : methanol : water (0.9 : 0.9 : 0.8 : 1.0, v/v) was employed to obtain the FEF. The second-order polynomial model was able to fit the experimental data adequately. The linear and quadratic terms of X3 were the most significant factors that affected all the responses. The positive linear term of X3 indicated a substantial increase in extraction yield, while the negative quadratic term showed a nonlinear tendency. Linear terms of X1 suggested a tendency to solvent saturation, except for QCT. The terms of X2 did not affect the responses substantially. The HPCCC method was found to be efficient and rapid for separating the FEF with 71% (w/w) flavonoid content. Overall, the developed extraction procedure coupled with HPCCC proved to be efficient for obtaining an enriched fraction with a very high content of flavonoids from A. satureioides.


Assuntos
Achyrocline/química , Distribuição Contracorrente/métodos , Flavonoides/isolamento & purificação , Extratos Vegetais/isolamento & purificação
4.
Food Chem ; 310: 125854, 2020 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-31784067

RESUMO

Zeaxanthin dipalmitate (ZDP) is a major non-saponified carotenoid in fully ripe fruits of Lycium barbarum L. In the present study, response surface methodology was used to optimize the ultrasonic-assisted extraction (UAE) conditions of carotenoids from the fruits of L. barbarum, and the optimal extraction conditions were determined as follows: ultrasonic power of 360 W, ultrasonic time of 40 min and the ratio of extraction solvent to sample of 30 mL/g. An actual value of ZDP content of 5.40 mg/g and short extraction time indicated the efficiency of UAE. Furthermore, a promising high-speed counter-current chromatography (HSCCC) method was established for the purification of ZDP from the fruits of L. barbarum. With a developed two-phase solvent system composed of n-hexane/dichloromethane/acetonitrile (10/3/7, v/v/v), ZDP with a purity of higher than 95% was successfully isolated from the crude extract. This is the first report on the purification of ZDP by using HSCCC.


Assuntos
Distribuição Contracorrente/métodos , Lycium/química , Palmitatos/isolamento & purificação , Sonicação , Xantofilas/isolamento & purificação , Cromatografia Líquida de Alta Pressão/métodos , Frutas/química , Solventes/química
5.
J Chromatogr A ; 1613: 460657, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-31685246

RESUMO

Aristolochic acids (AAs), the major components in Aristolochia manshuriensis Kom stems (AMK), may cause Chinese herb nephropathy during clinical application. Therefore, it is necessary to distinguish AMK from other herbs and Chinese medicines using AAs with high purity as standards. So, an efficient method for separation and purification of AAs is required because of their similar structures. In this study, six AAs with purities of >98% were obtained by pH-zone-refining counter-current chromatography (PZRCCC) in a single run. The optimum two-phase solvent system was petroleum ether-ethyl acetate-methanol-water (3:7:3:7, v/v). Triethylamine (10 mM) was added to the aqueous mobile phase and trifluoroacetic acid (10 mM) to the organic stationary phase. As a result, 9.7 mg aristolochic acid IIIa, 12.0 mg aristolochic acid IVa, 32.2 mg aristolochic acid II, 103.7 mg aristolochic acid I, 24.6 mg aristolic acid II, and 26.1 mg aristolic acid I were obtained from 800 mg AAs crude extract. The elution order of AAs during PZRCCC separation corresponded with the pKa values and hydrophobicities of the target compounds. PZRCCC is an efficient method for isolation of AAs with similar structures.


Assuntos
Aristolochia/química , Ácidos Aristolóquicos/isolamento & purificação , Distribuição Contracorrente/métodos , Ácidos Aristolóquicos/química , Concentração de Íons de Hidrogênio , Extratos Vegetais/química , Caules de Planta/química , Solventes/química
6.
J Chromatogr A ; 1613: 460660, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-31685247

RESUMO

Pyrethrum extract is a natural insecticide that has been used worldwide for centuries, with pyrethrins being considered active ingredients. Their purification is difficult due to their chemical structural similarity and instability to light. In the present study, a coordination complex high-speed counter-current chromatography was used to separate pyrethrins from pyrethrum extract. The two-phase solvent system composed of petroleum ether-ethyl acetate-methanol-water (10:2:10:2, v/v/v/v) with 0.30 mol/L silver nitrate was selected for separation. As a result, five pyrethrins including cinerin II (31.2 mg), pyrethrin II (43.9 mg), jasmalin II (39.1 mg), pyrethrin I (32.4 mg), and jasmalin I (16.0 mg) were obtained from 400 mg crude extract in 6 h by one-step HSCCC separation, with purities over 95%. The isolated compounds were identified by MS, 1H-NMR and 13C-NMR spectroscopy. The results demonstrated that the addition of silver nitrate in the two-phase solvent system of HSCCC significantly increased the HSCCC separation factor of pyrethrins. HSCCC is a rapid method for purification of pyrenthrins from pyrethrum extract. The established method may also be applied to separate analogous compounds from natural products in which the chemical structure differs in the position or numbers of alkenes.


Assuntos
Chrysanthemum/química , Distribuição Contracorrente/métodos , Piretrinas/isolamento & purificação , Nitrato de Prata/química , Flores/química , Extratos Vegetais/química , Piretrinas/análise
7.
J Chromatogr Sci ; 57(10): 901-909, 2020 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-31609449

RESUMO

We established an efficient method using high-speed counter-current chromatography (HSCCC) combined with preparative high-performance liquid chromatography (prep-HPLC) for isolating and purifying phenolic glycoside isomers. The method involves a rapid and sensitive ultra-performance liquid chromatography-under voltage(UPLC-UV) technique using a sub-2 µm core-shell particle column for qualitative and quantitative analysis of four phenolic glycoside isomers from Idesia polycarpa Maxim. leaves. The partially purified samples from ethyl acetate extraction of ethanol extracts of I. polycarpa Maxim. leaves were obtained by HSCCC with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water solution (3:5:3:5, v/v/v/v) to obtain fractions I and II, which contain two phenolic glycoside isomers. The two fractions were further isolated by prep-HPLC to yield compounds 1, 2, 3, and 4 with purities of 98.29%, 98.71%, 98.49% and 98.52%, and total recoveries of 93.5%, 72.2%, 75.5% and 88.3%, respectively. Compound 1 was first isolated from I. polycarpa Maxim., while compound 2 was reported to be a new phenolic glycoside, which is 1-[(6'-O-(Z)-p-coumaroyl)-ß-D-glucopyranosyl]-oxy-2-phenol. The chemical structures of the four phenolic glycoside isomers were analyzed and confirmed by UPLC, UV, electrospray ionization mass spectrometry (ESI-MS), fourier transform infra-red (FT-IR), 1H-nuclear magnetic resonance (1H NMR), 13C-nuclear magnetic resonance (13C NMR) and 2D nuclear magnetic resonance (2D NMR) spectra. This study opens prospects for broad industrial applications of HSCCC/prep-HPLC for the isolation and purification of isomers.


Assuntos
Distribuição Contracorrente/métodos , Glicosídeos/isolamento & purificação , Fenóis/isolamento & purificação , Extratos Vegetais/química , Salicaceae/química , Cromatografia Líquida de Alta Pressão/métodos , Glicosídeos/análise , Glicosídeos/química , Isomerismo , Fenóis/análise , Fenóis/química , Folhas de Planta/química
8.
J Chromatogr A ; 1609: 460503, 2020 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-31561970

RESUMO

In counter-current chromatography (CCC), the hydrodynamic motion of a two-phase solvent system in the column/coil is very important. There are some previous visualization studies on CCC using stroboscopic photography. As CCC separation is a continuous liquid-liquid extraction process, observing the distribution and movement of the solvent system on-line will be helpful to understand the hydrodynamic behavior during the whole CCC separation process. In the present study, a high-speed imaging camera was employed to take videos of a running CCC bobbin (visualized continuously on-line). The dynamic motion and phase distribution of conventional quaternary solvent system hexane/ethyl acetate/methanol/water (HEMWat) and ternary solvent system dichloromethane/methanol/water were both investigated. Wave-like mixing was observed in the area where the centrifugal force is minimum in different diameter columns. When the coil rotated, the mixing zone was "fixed" at the minimum centrifugal force position. Several photographs of the rotating coiled tube were taken which revealed that the phase dispersion hardly changes once equilibrium is established. Finally, the sample dispersion process was also recorded. These results will help us to understand the separation process in a CCC column/coil and also present some more interesting questions related to separation efficiency which are shown in discussion part of this paper.


Assuntos
Distribuição Contracorrente/métodos , Hidrodinâmica , Hexanos/química , Metanol/química , Reologia , Solventes/química , Água/química
9.
J Chromatogr A ; 1611: 460576, 2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31606158

RESUMO

Countercurrent chromatography (or counter-current chromatography, CCC) is a unique support-free liquid-liquid partition chromatography. Since it was invented by Y. Ito in 1960s, CCC has been widely accepted and applied as popular separation and purification technique for natural and synthetic complex. However, up to date there is little study to address on hydrophobic and hydrophilic interactions in CCC process, although hydrophobic interaction chromatography (HIC) and hydrophilic interaction chromatography (or hydrophilic interaction liquid chromatography, HILIC) as solid-support chromatographic techniques are widely applied at different stages of downstream processing. In fact, hydrophobic and hydrophilic interactions might be more popular in CCC separation than that in the liquid chromatography. For example, adding small solvents or additives in two-phase solvent systems may change significantly hydrophobic or hydrophilic interactions between solvents and solutes. Normally, CCC separation employs a light and hydrophobic organic phase as the stationary phase, and a heavy and hydrophilic aqueous phase as the mobile phase. Hydrophobic interactions between the solvent system and solutes (targets) will increase the partition coefficients (K values) of solutes and lengthen retention time, while hydrophilic interactions will reduce the K values and separation time. In this work, we aim to provide a general insight on the hydrophobic and hydrophilic interactions in CCC separation. We also highlight the current advances in utilizing the hydrophobic and hydrophilic interactions for K-targeted CCC separation and purification.


Assuntos
Distribuição Contracorrente/instrumentação , Cromatografia Líquida , Distribuição Contracorrente/métodos , Interações Hidrofóbicas e Hidrofílicas , Extratos Vegetais/química , Solventes/química , Água/análise
10.
Anal Bioanal Chem ; 412(3): 795-802, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31858167

RESUMO

Countercurrent chromatography (CCC) was used for the enrichment of α-tocodienol (α-T2), a rare vitamin E-related minor compound previously tentatively detected in palm oil. Hitherto, only one isomer has been mentioned to occur at traces in palm oil. However, CCC fractionation followed by GC/MS measurements of all fractions resulted in the detection of two α-T2 isomers in five different palm oil vitamin E dietary supplement capsules. Five repetitive CCC separations of ~ 1 g sample and additional purification steps by column chromatography provided ~ 2 mg of two equally abundant α-T2 isomers with a purity of ~ 85%. The positions of the double bonds in the alkyl side chain could be assigned by means of two characteristic chemical shifts in the 1H NMR spectrum. Accordingly, the structures of the α-T2 isomers were 2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridec-3,11-dienyl)chroman-6-ol (double bonds in 3',11'-position) and 2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridec-7,11-dienyl)chroman-6-ol (double bonds in 7',11'-position). Natural occurrence of both isomers was proven by GC/MS screening of crude palm oil after saponification and CCC separation. Moreover, GC/MS analysis allowed the tentative assignment of γ-tocomonoenol (γ-T1) and ß-tocomonoenol (ß-T1) as trace compounds in palm oil.


Assuntos
Distribuição Contracorrente/métodos , Óleo de Palmeira/química , Vitamina E/análogos & derivados , Cromatografia Gasosa-Espectrometria de Massas , Isomerismo , Estrutura Molecular , Espectroscopia de Prótons por Ressonância Magnética , Vitamina E/química
11.
Talanta ; 206: 120195, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514831

RESUMO

The ability to separate bioactive compounds from herbal medicines, which contain abundant components, is crucial for drug discovery. Conventional Countercurrent chromatography (CCC) methods for separating bioactive compounds are labor intensive and show low efficiency. Here, we present a novel integrative CCC method for separating lysine-specific demethylase 1 (LSD1) inhibitors from the roots of Salvia miltiorrhiza (RSM). The methanol extracts of RSM were separated into hydrosoluble and liposoluble fractions, which were online stored in coils. Subsequently, the targeting LSD1 constituents were isolated using isocratic, gradient, or recycling elution mode. All separation processes could be accomplished using one CCC apparatus. Using our separation strategy, two phenylpropanoids and four tanshinones were isolated, which were determined to be new classes of natural LSD1 inhibitors. Salvianolic acid B, which showed the most potent inhibitory activity with an IC50 of 0.11 µM, exhibiting a considerable potential as an anticancer agent. Promisingly, the integrative CCC could be a crucial tool for the target separation of enzyme inhibitors from herbal medicines.


Assuntos
Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Raízes de Plantas/química , Salvia miltiorrhiza/química , Benzofuranos/isolamento & purificação , Benzofuranos/metabolismo , Benzofuranos/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Cinamatos/isolamento & purificação , Cinamatos/metabolismo , Cinamatos/farmacologia , Distribuição Contracorrente/métodos , Depsídeos/isolamento & purificação , Depsídeos/metabolismo , Depsídeos/farmacologia , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/metabolismo , Histona Desmetilases/metabolismo , Humanos , Simulação de Acoplamento Molecular , Fenantrenos/isolamento & purificação , Fenantrenos/metabolismo , Fenantrenos/farmacologia , Ligação Proteica
12.
Mar Drugs ; 17(12)2019 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-31818004

RESUMO

Sargassum horneri, a sargassaceae brown alga, is one of the main species in the subtidal seaweeds flora extensively distributed in the Yellow and East China Sea. It has been proven that the phytosterols are an important class of bioactive substances in S. horneri. In this work, a counter-current chromatography approach is proposed for preparative separation of phytol and two analogue sterols from a crude extract of S. horneri. A two-phase solvent system composed of n-hexane-acetonitrile-methanol (5:5:6, v/v) was selected and optimized. The effects of rotary speed and flow rate on the retention of the stationary phase were carefully studied. Under the optimum conditions, phytol and two analogue sterols, fucosterol and saringosterol, were baseline separated, producing 19.8 mg phytol, 23.7 mg fucosterol, and 3.1 mg saringosterol from 300 mg of crude S. horneri extract in one-step separation. The purities of three target compounds were all above 85%. The structures of phytol and two sterols were identified by nuclear magnetic resonance spectroscopy.


Assuntos
Distribuição Contracorrente/métodos , Fitosteróis/isolamento & purificação , Sargassum/química , Espectroscopia de Ressonância Magnética , Fitol/química , Fitol/isolamento & purificação , Fitosteróis/química , Solventes/química , Estigmasterol/análogos & derivados , Estigmasterol/química , Estigmasterol/isolamento & purificação
13.
J Agric Food Chem ; 67(41): 11508-11517, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31538478

RESUMO

Tiopronin, as a novel thiol-containing nucleophile, was introduced for depolymerizing polymeric proanthocyanidins from grape seed into catechins and three new proanthocyanidin-tiopronin degradation products: (+)-catechin-4ß-S-tiopronin methyl ester (CT), (-)-epicatechin-4ß-S-tiopronin methyl ester (ECT), and (-)-epicatechin gallate-4ß-S-tiopronin methyl ester (ECGT). A Box-Behnken design was employed to optimize degradation conditions based on single-factor experiments to obtain target products. Each of the new degradation compounds was isolated by the high-speed counter-current chromatography combined with semipreparative high performance liquid chromatography in large amounts, and then, their structures were identified by 1H NMR, 13C NMR, 2D-NMR, as well as mass spectrometry analysis. The absolute configurations were further confirmed by comparison between the calculated electronic circular dichroism and experimental spectra. Further evaluation of antibacterial activities of these compounds showed that CT and ECT possessed more inhibiting capacity against Staphylococcus aureus and Escherichia coli than parent compound catechin and epicatechin. However, ECGT has no bacteriostatic capacity against these two bacteria.


Assuntos
Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Catequina/química , Distribuição Contracorrente/métodos , Extrato de Sementes de Uva/isolamento & purificação , Extrato de Sementes de Uva/farmacologia , Proantocianidinas/isolamento & purificação , Proantocianidinas/farmacologia , Tiopronina/química , Antibacterianos/química , Cromatografia Líquida de Alta Pressão , Escherichia coli/efeitos dos fármacos , Extrato de Sementes de Uva/química , Proantocianidinas/química , Sementes/química , Espectrometria de Massas por Ionização por Electrospray , Staphylococcus aureus/efeitos dos fármacos , Vitis/química
14.
J Chromatogr A ; 1608: 460422, 2019 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-31500882

RESUMO

Nectandra leucantha (Lauraceae) is a tree indigenous to the tropical Atlantic forests of Brazil, one of the most biodiverse flora hotspots worldwide. This plant species contains high concentrations of neolignan and dehydrodieugenol derivatives that express significant in-vitro activities against various parasite strains. These activities are however responsible for severe tropical human infections, such as Leishmaniasis (Leishmania spp.) and Chagas disease (Trypanosoma cruzi), which have been classified by the World Health Organization (WHO) as Neglected Tropical Diseases (NTDs). In order to optimize the isolation process for these target metabolites, n-hexane extract of the leaves was separated by means of semi-preparative high performance countercurrent chromatography (HPCCC) and scale-up spiral-coil countercurrent chromatography (sp-CCC) systems. Several biphasic solvent mixtures were evaluated for their partitioning effects on neolignans, resulting in the selection of an optimized system n-hexane - ethylacetate - methanol - water (7:3:7:3, v/v/v/v). The chromatographic experiments on the HPCCC and sp-CCC were run in the head-to-tail mode with 500 mg and 16 g injections, respectively. For specific and multiple metabolite detection, the recovered CCC-fractions were off-line injected, in the sequence of recovery, to an electrospray mass-spectrometry (ESI-MS/MS) device. A projection of the single ion traces of the target compounds, in the positive ionization mode at a scan range of m/z 100-1500, located chromatographic areas where the co-elution effects occurred and pure target metabolites were present. Five major target neolignans were specifically detected, which enabled the accurate pooling of CCC-fractions for an optimum recovery of the metabolites. The direct comparison of the performance characteristics of the two CCC-devices, with very different mechanical designs was achieved by the conversion of the time axis into a partition ratio (KD) separation scale. As a result, the compound specific KD-elution values of the target neolignan were determined in high precision, while the comparison of the calculated separation factor (α) and resolution factor (RS) values revealed a superior separation performance for the HPCCC system. Also, the reproducibility of detected metabolites in the two CCC experiments was confirmed by small variations (ΔKD ±0.1). Neolignan target compounds with anti-parasite activities were successfully isolated in the 100 mg to 4 g range in a single lab-scale countercurrent chromatographic process step.


Assuntos
Distribuição Contracorrente/métodos , Lauraceae/química , Lignanas/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Espectrometria de Massas em Tandem/métodos , Brasil , Cromatografia Líquida de Alta Pressão/métodos , Eugenol/análogos & derivados , Eugenol/análise , Eugenol/isolamento & purificação , Lignanas/análise , Extratos Vegetais/análise , Folhas de Planta/química
15.
J Chromatogr A ; 1603: 433-437, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31288929

RESUMO

An efficient strategy for the selection of active components based on counter-current fractionation and bioassay-guided separation was established in the present work. Blaps rynchopetera Fairmaire was an edible medicinal insect. Its extract showed the potential RAW264.7 macrophage cell inhibitory activity. After extraction with different solvents, the active components were enriched in ethyl acetate. In order to further track the active compounds, the ethyl acetate extraction was divided into 14 fractions by means of HSCCC. The results showed that the activities of F6 and F7 were significant higher than the others. Two compounds, hydroxytyrosol and 4-ethylbenzene-1,3-diol, were separated from the mixture of F6 and F7 by column chromatography and their chemical structures were confirmed by MS, 1H NMR and 13C NMR. The IC50 of hydroxytyrosol and 4-ethylbenzene-1,3-diol against RAW264.7 macrophage cell were 38.24 ±â€¯0.26 µg/mL and 103.26 ±â€¯0.29 µg/mL, respectively, indicating that hydroxytyrosol was the major active ingredient responsible for the RAW264.7 inhibitory activity of Blaps rynchopetera Fairmaire.


Assuntos
Bioensaio/métodos , Distribuição Contracorrente/métodos , Insetos/química , Animais , Fracionamento Químico , Cromatografia Líquida de Alta Pressão , Camundongos , Extratos Vegetais/farmacologia , Células RAW 264.7 , Solventes/química
16.
J Chromatogr A ; 1603: 251-261, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31266644

RESUMO

Counter-current chromatography (CCC) is a unique, liquid-liquid partition chromatography process. Both the mobile and stationary phases are liquids, so no solid support matrix is used. CCC has gained wide acceptance as a preparative technique in a variety of fields. Because the mobile and stationary phases are both liquids, gradient elution is difficult to perform with CCC. Phase equilibrium must be maintained, so any change in the composition of one phase may induce a compositional change in the other. In this work, a new linear gradient elution method was developed for CCC. Biphasic solvent systems containing heptane, ethyl acetate, methanol, and water (HepEMWat) in various ratios were prepared and used to optimize both isocratic and linear gradient CCC separation with methanol. We first separated a test mixture of four standard compounds with partition coefficients ranging from 0.8 to 7.8. The separation resembled a reversed-phase process, and elution was performed while progressively decreasing the polarity of the mobile phase. Target molecules with small partition coefficients eluted first in the lower phase of the optimized HepEMWat solvent system. Elution of constituents with large partition coefficients was quite slow under isocratic conditions. Separation time was significantly reduced when elution was performed with a linear gradient using methanol and the optimal HepEMWat system. Elution with a 3:7:4:6 (v/v/v/v) HepEMWat system took approximately 200 min. This included an 80-min isocratic step, followed by gradient elution with methanol from 0% to 30%. The optimized methanol linear gradient CCC method was then used to separate a complex mixture of natural products isolated from Sinopodophyllum hexandrum (Royle) Ying roots. Twelve compounds with a wide range of polarities were well-resolved in a single separation. We have developed a convenient and cost-effective strategy for the separation of complex mixtures. No tedious mobile phase preparation step is required. The volume of unused mobile phase is minimal, so little solvent is wasted. The method is an important advance for the separation of mixtures that contain many compounds with a large range of polarities and partition coefficients, which are common features of natural products.


Assuntos
Berberidaceae/química , Produtos Biológicos/isolamento & purificação , Distribuição Contracorrente/métodos , Metanol/química , Cromatografia Líquida de Alta Pressão/métodos , Extratos Vegetais/isolamento & purificação , Solventes/química , Água/química
17.
J Chromatogr Sci ; 57(8): 738-744, 2019 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-31318427

RESUMO

Morus alba L. is a medicinal plant that contains a high amount of caffeoylquinic acids such as 3-caffeoylquinic acid (3-CQA), 5-caffeoylquinic acid (5-CQA) and 4-caffeoylquinic acid (4-CQA). This study aimed to establish a fast and efficient method for separating caffeoylquinic acids from mulberry leaves by using high-speed countercurrent chromatography coupled with macroporous resin. D101 resin showed better adsorption and desorption capacity for three caffeoylquinic acids among six macroporous resin adsorbents. The contents of 3-CQA, 5-CQA and 4-CQA reached for 4.77%, 18.95% and 9.84% through one cycle of D101 resin, which were 3.13-fold, 4.57-fold and 4.78-fold more than those in crude extracts, respectively. With a two-phase solvent system of ethyl acetate-water (1:1, V/V), >93% purity of target compounds were obtained in one cycle during 150 min with the recovery yields of 80.59%, 99.56% and 94.21% for 3-CQA, 5-CQA and 4-CQA, respectively. The structural identification of target compounds was carried out by ESI-MS, 1H NMR and 13C NMR spectra. The present result represented an easy and efficient separation strategy for the utilization of mulberry resource.


Assuntos
Distribuição Contracorrente/métodos , Morus/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Ácido Quínico/análogos & derivados , Extração Líquido-Líquido , Espectrometria de Massas , Folhas de Planta/química , Ácido Quínico/química , Ácido Quínico/isolamento & purificação
18.
J Chromatogr A ; 1603: 240-250, 2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31221429

RESUMO

Closed-loop recycling dual-mode counter-current chromatography (CLR DM CCC) includes two separation stages: 1 - closed-loop recycling separation of solutes with mobile x-phase (CLR CCC); 2 - separation of solutes with the mobile y-phase in the opposite flow direction. Previous analysis of CLR DM CCC separations has been limited to the ideal recycling model, which neglects extra-column dispersion. In this study, the analysis of CLR CCC separations is based on the non-ideal recycling model, which takes into account the extra-column dispersion caused by the recycling system. This is of great practical importance, since by selecting the optimal parameters of the recycling system the separation can be significantly improved. Comparative analysis of CLR CCC and CLR DM CCC separations has shown that at low separations factors compounds with low partition coefficients can be separated by CLR CCC using recycling systems with a long recycling line; the separation of compounds with high partition coefficients and the separation of complex mixtures can be performed by CLR DM CCC. Simple equations for simulation and design of CLR DM CCC separations are developed. Several variants of the implementation of this separation method are discussed; examples of simulation are presented in "Mathcad" program.


Assuntos
Distribuição Contracorrente/métodos , Modelos Teóricos , Misturas Complexas/isolamento & purificação , Reciclagem , Reprodutibilidade dos Testes , Soluções/química
19.
Artigo em Inglês | MEDLINE | ID: mdl-31254782

RESUMO

Polymethoxyflavones (PMFs) are widely found in Citri Reticulatae Pericarpium (CRP) and have been investigated with a broad spectrum of biological activities as well as health promoting properties. However, separation of the PMFs from a complex sample, especially preparative separation of these PMFs with high purity, remains challenging. In the present study, an efficient method based on supercritical fluid extraction (SFE) and continuous high-speed counter-current chromatography (HSCCC) has been developed for extracting and preparative purification PMFs from CRP. Various experimental conditions were investigated to optimize the SFE and HSCCC processes. Under these optimized conditions, crude extract of CRP (extract I) was obtained with a maximum contents of nobiletin, 3,5,6,7,8,3',4'-heptamethoxyflavone and tangeretin. Further extraction of crude extract I was carried out to obtain crude extract II, which was further isolated and purified by HSCCC. It was worth mentioned that continuous injection HSCCC process were realized without lost of separation efficiency, which allowed for multiple purification cycles and therefore saved a lot of labor and time. Furthermore, high-performance liquid chromatography (HPLC) was employed to analyze the fractions separated by HSCCC, which revealed that the purities of the three PMFs were all above 98%. The structures of the three PMFs were identified by LC-MS and 1H NMR spectroscopy.


Assuntos
Cromatografia com Fluido Supercrítico/métodos , Citrus/química , Distribuição Contracorrente/métodos , Flavonas/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Dióxido de Carbono/química , Flavonas/química , Espectroscopia de Ressonância Magnética , Extratos Vegetais/química
20.
Biomed Chromatogr ; 33(9): e4574, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31062384

RESUMO

To develop an efficient method for large preparation of javanicin from Fusarium solani, a rapid and simple method by high-speed countercurrent chromatography was established based on average polarity (P' values) and partition coefficients (K values) of crude samples. A suitable solvent system for high-speed countercurrent chromatography was selected from many possible biphasic solvent systems. HSCCC was successfully applied to separate and purify javanicin, the main bioactive component of solid cultures of the fungus F. solani isolated from the fruiting body of Trametes trogii, with petroleum ether-ethyl acetate-methanol-water (4:3:2:1, v/v) as solvent system. A total amount of 40.6 mg of javanicin was obtained from 100 mg crude sample. The purity of javanicin was 92.2% with a recovery of 95.1%, as determined by high-performance liquid chromatrography. The molecular structure was identified primarily by NMR and MS methods. The results indicated that high-speed countercurrent chromatography could be a powerful technology for separating naphthoquinones from the solid cultures of the fungus F. solani. It is also of significance that the separation of javanicin from natural source was carried out for the first time utilizing high-speed countercurrent chromatography.


Assuntos
Distribuição Contracorrente/métodos , Carpóforos/química , Fusarium/química , Quassinas/isolamento & purificação , Solventes
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