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1.
Pharm Biol ; 57(1): 392-402, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31188689

RESUMO

Context: Citrus unshiu Markovich (Rutaceae) peel is known to contain high concentrations of flavonoids and exerts pharmacological effects on antioxidant, anti-inflammation, allergies, diabetes and viral infections. Objective: Very little is known about potential activity of fermented dried Citrus unshiu peel extracts (FCU) using Bacillus subtilis, as well as its mechanism of action. We investigated the effects of FCU on the anti-inflammatory activities in murine macrophages and moisturizing effects in human keratinocytes. Materials and methods: We isolated the Bacillus subtilis from Cheonggukjang and FCU using these Bacillus subtilis to prepare samples. The cells were pre-treated with various extracts for 2 h and then induced with LPS for 22 h. We determined the NO assay, TNF-α, IL-6 and PGE2 in RAW 264.7 ells. The expression of SPT and Filaggrin by FCU treatment was measured in HaCaT cells. Result: We found that two types of FCU highly suppressed LPS-induced nitric oxide (NO) without exerting cytotoxic effects on RAW 264.7 cells (21.9 and 15.4% reduction). FCU inhibited the expression of LPS-induced iNOS and COX-2 proteins and their mRNAs in a concentration-dependent manner. TNF-α (59 and 30.9% reduction), IL-6 (39.1 and 65.6% reduction), and PGE2 secretion (78.6 and 82.5% reduction) were suppressed by FCU in LPS-stimulated macrophages. Furthermore, FCU can induce the production of hyaluronic acid (38 and 38.9% induction) and expression of Filaggrin and SPT in HaCaT keratinocyte cells. Discussion and conclusion: FCU potentially inhibits inflammation, improves skin moisturizing efficacy, and it may be a therapeutic candidate for the treatment of inflammation and dry skin.

2.
J Chromatogr A ; 1602: 188-198, 2019 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-31147156

RESUMO

A liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS)-based dereplicative method was developed for identifying oplopane- and bisabolane-type sesquiterpenoids from buds of Tussilago farfara L. The analysis of these chemical analogues, sesquiterpene esters, is challenging by MS-based nontargeted metabolomic approaches because of their in-source fragmentation and structural diversity. To profile these sesquiterpenoids, four diagnostic ions (m/z 215.143, 217.158, 229.123, and 231.138) were suggested in the positive ion mode and the developed method utilized two sequential MS/MS scan modes to identify common skeletons and investigate the fragmentation patterns of their parent molecules. Precursor ion scan by triple quadrupole MS/MS provided the parent molecular ions from their diagnostic ions, and product ion scan by quadrupole time-of-flight MS/MS confirmed their fragmentation behaviors. Under the optimized UHPLC-MS/MS method, 74 sesquiterpenoids were characterized from the Farfarae Flos and 11 compounds were isolated for the method validation. Among those compounds, three sesquiterpenoids were newly separated from the Farfarae Flos. Furthermore, the diagnostic ions and the MS/MS fragment behaviors were applied to the accurate quantification of the 8 isolated sesquiterpenoids. Therefore, the developed LC-MS/MS-based method highlighted the chemical composition of the Farfarae Flos and could be extended to the screening and quantification of other sesquiterpene esters.


Assuntos
Técnicas de Química Analítica/métodos , Espectrometria de Massas , Sesquiterpenos/química , Tussilago/química , Cromatografia Líquida , Íons/química , Sesquiterpenos/isolamento & purificação
3.
BMC Complement Altern Med ; 19(1): 90, 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31036001

RESUMO

BACKGROUND: Papaver nudicaule belongs to the Papaveraceae family, which is planted as an annual herbaceous species generally for ornamental purpose. Papaver rhoeas in the same family has been reported to have various pharmacological activities such as antioxidant and analgesic effects. In contrast, little is known about the pharmacological activity of Papaver nudicaule. In this study, the anti-inflammatory activity of Papaver nudicaule extracts and the action mechanisms were investigated in RAW264.7 macrophage cells. METHODS: To investigate the anti-inflammatory activity of five cultivars of Papaver nudicaule with different flower color, samples were collected from their aerial parts at two growth stages (60 and 90 days) and their ethanol extracts were evaluated in the lipopolysaccharide (LPS)-treated RAW264.7 cells by measuring nitric oxide (NO) and prostaglandin E2 (PGE2) levels. Interleukin 1-beta (IL-1ß), Interleukin-6 (IL-6) and Tumor necrosis factor alpha (TNF-α) production were also analyzed by RT-PCR and multiplex assays. Nuclear Factor-kappa-light-chain-enhancer of activated B cells (NF-κB) and Signal transducer and activator of transcription 3 (STAT3) signaling pathways were examined using western blotting and luciferase reporter assays to reveal the action mechanism of Papaver nudicaule extracts in their anti-inflammatory activity. RESULTS: All of the Papaver nudicaule extracts were effective in reducing the LPS-induced NO, which is an important inflammatory mediator, and the extract of Papaver nudicaule with white flower collected at 90 days (NW90) was selected for further experiments because of the best effect on reducing the LPS-induced NO as well as no toxicity. NW90 lowered the LPS-induced PGE2 level and decreased the LPS-induced Nitric oxide synthase 2 (NOS2) and Cyclooxygenase 2 (COX2). In addition, NW90 reduced the LPS-induced inflammatory cytokines, IL-1ß and IL-6. Furthermore, NW90 inhibited the LPS-induced activation of NF-κB and STAT3. CONCLUSIONS: These results indicate that NW90 may restrain inflammation by inhibiting NF-κB and STAT3, suggesting the potential therapeutic properties of Papaver nudicaule against inflammatory disease.


Assuntos
Anti-Inflamatórios/farmacologia , NF-kappa B/metabolismo , Papaver/química , Extratos Vegetais/farmacologia , Fator de Transcrição STAT3/metabolismo , Animais , Anti-Inflamatórios/química , Sobrevivência Celular/efeitos dos fármacos , Inflamação/induzido quimicamente , Lipopolissacarídeos/efeitos adversos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Óxido Nítrico/metabolismo , Extratos Vegetais/química , Células RAW 264.7 , Transdução de Sinais/efeitos dos fármacos
4.
Int J Mol Sci ; 19(10)2018 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-30332811

RESUMO

The Papaver spp. (Papaver rhoeas (Corn poppy) and Papaver nudicaule (Iceland poppy)) genera are ornamental and medicinal plants that are used for the isolation of alkaloid drugs. In this study, we generated 700 Mb of transcriptome sequences with the PacBio platform. They were assembled into 120,926 contigs, and 1185 (82.2%) of the benchmarking universal single-copy orthologs (BUSCO) core genes were completely present in our assembled transcriptome. Furthermore, using 128 Gb of Illumina sequences, the transcript expression was assessed at three stages of Papaver plant development (30, 60, and 90 days), from which we identified 137 differentially expressed transcripts. Furthermore, three co-occurrence heat maps are generated from 51 different plant genomes along with the Papaver transcriptome, i.e., secondary metabolite biosynthesis, isoquinoline alkaloid biosynthesis (BIA) pathway, and cytochrome. Sixty-nine transcripts in the BIA pathway along with 22 different alkaloids (quantified with LC-QTOF-MS/MS) were mapped into the BIA KEGG map (map00950). Finally, we identified 39 full-length cytochrome transcripts and compared them with other genomes. Collectively, this transcriptome data, along with the expression and quantitative metabolite profiles, provides an initial recording of secondary metabolites and their expression related to Papaver plant development. Moreover, these profiles could help to further detail the functional characterization of the various secondary metabolite biosynthesis and Papaver plant development associated problems.

5.
J Sep Sci ; 41(12): 2517-2527, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29607619

RESUMO

Papaver plants can produce diverse bioactive alkaloids. Papaver rhoeas Linnaeus (common poppy or corn poppy) is an annual flowering medicinal plant used for treating cough, sleep disorder, and as a sedative, pain reliever, and food. It contains various powerful alkaloids like rhoeadine, benzylisoquinoline, and proaporphine. To investigate and identify alkaloids in the aerial parts of P. rhoeas, samples were collected at different growth stages and analyzed using liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry. A liquid chromatography with mass spectrometry method was developed for the identification and metabolite profiling of alkaloids for P. rhoeas by comparing with Papaver somniferum. Eighteen alkaloids involved in benzylisoquinoline alkaloid biosynthesis were used to optimize the liquid chromatography gradient and mass spectrometry conditions. Fifty-five alkaloids, including protoberberine, benzylisoquinoline, aporphine, benzophenanthridine, and rhoeadine-type alkaloids, were identified authentically or tentatively by liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry in samples taken during various growth stages. Rhoeadine alkaloids were observed only in P. rhoeas samples, and codeine and morphine were tentatively identified in P. somniferum. The liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry method can be a powerful tool for the identification of diverse metabolites in the genus Papaver. These results may help understand the biosynthesis of alkaloids in P. rhoeas and evaluate the quality of this plant for possible medicinal applications.


Assuntos
Alcaloides/química , Cromatografia Líquida/métodos , Papaver/química , Extratos Vegetais/química , Espectrometria de Massas em Tandem/métodos , Componentes Aéreos da Planta/química , Plantas Medicinais/química
6.
J Nat Prod ; 79(2): 317-23, 2016 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-26829656

RESUMO

Glioblastoma is one of the most malignant primary tumors, and the prognosis for glioblastoma patients remains poor. Tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL) is considered a promising anticancer agent due to its remarkable ability to selectively kill tumor cells. However, since many cancers are resistant to TRAIL, strategies to overcome resistance are required for the successful use of TRAIL in the clinic. In the present study, the potential of morusin as a TRAIL sensitizer in human glioblastoma cells was evaluated. Treatment with TRAIL or morusin alone showed weak cytotoxicity in human glioblastoma cells. However, combination treatment of TRAIL with morusin synergistically decreased cell viability and increased apoptosis compared with single treatment. Morusin induced expression of death receptor 5 (DR5), but not DR4 or decoy receptors (DcR1 and DcR2). Furthermore, morusin significantly decreased anti-apoptotic molecules survivin and XIAP. In addition, morusin reduced expression of EGFR and PDFGR as well as phosphorylation of STAT3, possibly mediating down-regulation of survivin and XIAP. Together these results suggest that morusin enhances TRAIL sensitivity in human glioblastoma cells through regulating expression of DR5 and EGFR. Therefore, the combination treatment of TRAIL and morusin may be a new therapeutic strategy for malignant glioma patients.


Assuntos
Receptores ErbB/metabolismo , Flavonoides/farmacologia , Glioblastoma/genética , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/genética , Apoptose/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Flavonoides/química , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Humanos , Masculino , Estrutura Molecular , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/efeitos dos fármacos , Fator de Transcrição STAT3/efeitos dos fármacos , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo
7.
J Exp Bot ; 67(5): 1519-33, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26733692

RESUMO

To gain insights into the regulatory networks related to anthocyanin biosynthesis and identify key regulatory genes, we performed an integrated analysis of the transcriptome and metabolome in sprouts germinated from three colored potato cultivars: light-red Hongyoung, dark-purple Jayoung, and white Atlantic. We investigated transcriptional and metabolic changes using statistical analyses and gene-metabolite correlation networks. Transcript and metabolite profiles were generated through high-throughput RNA-sequencing data analysis and ultraperformance liquid chromatography quadrupole time-of-flight tandem mass spectrometry, respectively. The identification and quantification of changes in anthocyanin were performed using molecular formula-based mass accuracy and specific features of their MS(2) spectra. Correlation tests of anthocyanin contents and transcriptional changes showed 823 strong correlations (correlation coefficient, R (2)>0.9) between 22 compounds and 119 transcripts categorized into flavonoid metabolism, hormones, transcriptional regulation, and signaling. The connection network of anthocyanins and genes showed a regulatory system involved in the pigmentation of light-red Hongyoung and dark-purple Jayoung potatoes, suggesting that this systemic approach is powerful for investigations into novel genes that are potential targets for the breeding of new valuable potato cultivars.


Assuntos
Redes Reguladoras de Genes , Metaboloma/genética , Pigmentação/genética , Solanum tuberosum/genética , Transcriptoma/genética , Antocianinas/metabolismo , Vias Biossintéticas , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Análise de Componente Principal , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
8.
Am J Chin Med ; 43(3): 425-41, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25967662

RESUMO

As a treatment for allergic asthma, inhaled treatments such as bronchodilators that contain ß2-agonists have an immediate effect, which attenuates airway obstructions and decreases airway hypersensitivity. However, bronchodilators only perform on a one off basis, but not consistently. Asthma is defined as a chronic inflammatory disease of the airways accompanying the overproduction of mucus, airway wall remodeling, bronchial hyperreactivity and airway obstruction. Liriope platyphylla radix extract (LPP), a traditional Korean medicine, has been thoroughly studied and found to be an effective anti-inflammatory medicine. Here, we demonstrate that an inhaled treatment of LPP can attenuate airway hyperresponsiveness (AHR) in an ovalbumin-induced asthmatic mouse model, compared to the saline-treated group (p < 0.01). Moreover, LPP decreases inflammatory cytokine levels, such as eotaxin (p < 0.05), IL-5 (p < 0.05), IL-13 (p < 0.001), RANTES (p < 0.01), and TNF-α (p < 0.05) in the bronchoalveolar lavage (BAL) fluid of asthmatic mice. A histopathological study was carried out to determine the effects of LPP inhalation on mice lung tissue. We performed UPLC/ESI-QTOF-MS, LC/MS, and GC/MS analyses to analyze the chemical constituents of LPP, finding that these are ophiopogonin D, spicatoside A, spicatoside B, benzyl alcohol, and 5-hydroxymethylfurfural. This study demonstrates the effect of an inhaled LPP treatment both on airway AHR and on the inflammatory response in an asthmatic mouse model. Hence, LPP holds significant promise as a nasal inhalant for the treatment of asthmatic airway disease.


Assuntos
Asma/tratamento farmacológico , Liriope (Planta) , Medicina Tradicional Coreana , Fitoterapia , Extratos Vegetais/administração & dosagem , Hipersensibilidade Respiratória/tratamento farmacológico , Administração por Inalação , Animais , Asma/induzido quimicamente , Asma/metabolismo , Líquido da Lavagem Broncoalveolar/química , Citocinas/metabolismo , Depressão Química , Modelos Animais de Doenças , Feminino , Mediadores da Inflamação/metabolismo , Camundongos Endogâmicos BALB C , Ovalbumina , Extratos Vegetais/farmacologia , Hipersensibilidade Respiratória/induzido quimicamente
9.
J Sep Sci ; 38(3): 502-10, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25413645

RESUMO

This study was designed to classify and identify closely related thistle species in the genus Cirsium, as well as Carduus and Cephalonoplos species, which are also thistles. The comprehensive and untargeted metabolite profiles of nine Korean thistles were determined using ultra high performance liquid chromatography combined with hybrid quadrupole time-of-flight mass spectrometry. The difference in metabolite profiles among species was explored using principal component analysis and hierarchical clustering analysis. The significantly different metabolites (Bonferroni-corrected P-value < 0.001) were used to construct a partial least squares discriminant analysis model to predict the species of thistle. Nine species were successfully classified using a partial least squares discriminant analysis model and confirmed using a cross-validation method. Species with similar features were grouped based on unique patterns in variable clusters. The present study suggests that liquid chromatography with quadrupole time-of-flight mass spectrometry untargeted metabolomic profiling with chemometric analysis is an efficient and powerful tool for discriminating between different species of medicinal herbs.


Assuntos
Cirsium/química , Cromatografia Líquida de Alta Pressão , Cirsium/metabolismo , Análise por Conglomerados , Análise Discriminante , Espectrometria de Massas , Análise de Componente Principal , República da Coreia , Fatores de Tempo
10.
PLoS One ; 9(11): e113573, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25419661

RESUMO

Radiation exposure is a threat to public health because it causes many diseases, such as cancers and birth defects, due to genetic modification of cells. Compared with the past, a greater number of people are more frequently exposed to higher levels of radioactivity today, not least due to the increased use of diagnostic and therapeutic radiation-emitting devices. In this study, ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS)-based metabolic profiling was used to investigate radiation- induced metabolic changes in human fibroblasts. After exposure to 1 and 5 Gy of γ-radiation, the irradiated fibroblasts were harvested at 24, 48, and 72 h and subjected to global metabolite profiling analysis. Mass spectral peaks of cell extracts were analyzed by pattern recognition using principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA). The results showed that the cells irradiated with 1 Gy returned to control levels at 72 h post radiation, whereas cells irradiated with 5 Gy were quite unlike the controls; therefore, cells irradiated with 1 Gy had recovered, whereas those irradiated with 5 Gy had not. Lipid and amino acid levels increased after the higher-level radiation, indicating degradation of membranes and proteins. These results suggest that MS-based metabolite profiling of γ-radiation-exposed human cells provides insight into the global metabolic alterations in these cells.


Assuntos
Fibroblastos/efeitos da radiação , Raios gama , Metaboloma/efeitos da radiação , Metabolômica , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Cromatografia Líquida de Alta Pressão , Derme/citologia , Análise Discriminante , Relação Dose-Resposta à Radiação , Fibroblastos/metabolismo , Humanos , Análise dos Mínimos Quadrados , Espectrometria de Massas/métodos , Análise Multivariada , Análise de Componente Principal , Fatores de Tempo
11.
J Sep Sci ; 37(1-2): 61-8, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24327461

RESUMO

In this study, enzymatic hydrolysis and chemometric methods were utilized to discriminate glycosylated platycosides in the extract of Platycodi Radix by LC-MS. Laminarinase, whose enzymatic activity was evaluated using gentiobiose and laminaritriose, was a suitable enzyme to identify the glycosylated platycosides. The laminarinase produced deapi-platycodin D and platycodin D from the isolated deapi-platycoside E and platycoside E through the loss of two glucose units by enzymatic reaction, respectively. After hydrolyzing a crude extract by laminarinase, the reconstructed total ion chromatogram generated by a chemometric technique sorted peaks of deglycosylated platycosides easily. Structural information of the glycosylated isomers was revealed through fragment ions generated by the sodiated C0ß ion corresponding to reduced disaccharides in the positive MS(4) spectra. Characteristic fragment ions of Glc-(1→6)-Glc moieties were observed through ring cleavages of (0,2)A0ß, (0,3)A0ß, and (0,4)A0ß, whereas Glc-(1→3)-Glc moieties produced only (0,3)A0ß ions. Lithium-adducted platycosides allowed more detailed structural analysis of glycosidic bond cleavage corresponding to Y1ß and B1ß in addition to ring cleavage.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Fabaceae/química , Ácido Oleanólico/análogos & derivados , Extratos Vegetais/química , Saponinas/química , Espectrometria de Massas em Tandem/métodos , Biocatálise , Celulases/química , Proteínas Fúngicas/química , Estrutura Molecular , Ácido Oleanólico/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Trichoderma/enzimologia
12.
J Chromatogr A ; 1310: 66-73, 2013 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-23992840

RESUMO

The targeted purification of compounds with a broad polarity range from traditional medicinal plants is a big challenge for counter-current chromatography (CCC). Gradient elution was introduced in CCC to address this problem. However, once a suitable solvent system is selected, the separation process requires optimization of operational parameters. The present study was conducted to optimize various operational parameters to integrate the flow rate and solvent gradients for the rapid isolation of eight coumarins from Seseli resinosum in a single run. An increase in the system temperature from 15°C to 35°C increased the stationary phase retention and solubility of the sample, whereas the operation time and viscosity of the system were decreased. The high purity of each compound was ensured by collecting the fractions from the main peaks while all the shoulder peaks were mixed and separated under the same conditions with semi-preparative CCC. GC-FID was used to analyze the components of each phase, which was prepared without presaturation to save the time and solvent consumption. Finally, eight coumarins were purified, including (1) d-laserpitin, (2) (3'S,4'S)-3'-angeloyloxy-4'-hydroxy-3',4'-dihydroseselin, (3) (+)-samidin, (4) (3'S,4'S)-3'-acetoxy-4'-angeloyloxy-3',4'-dihydroseselin, (5) deltoin (6), calipteryxin, (7) (3'S,4'S)-3',4'-disenecioyloxy-3',4'-dihydroseselin, and (8) (-)-anomalin. The present technique has successfully accomplished the goal of one-step separation of these compounds with high purity and recovery in an economic and time efficient manner.


Assuntos
Apiaceae/química , Cumarínicos/análise , Distribuição Contracorrente/métodos , Cromatografia Líquida de Alta Pressão/métodos , Cumarínicos/química , Extratos Vegetais/química , Raízes de Plantas/química , Solventes , Temperatura Ambiente
13.
Planta Med ; 79(8): 639-45, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23576176

RESUMO

The present study was undertaken to investigate the antiproliferative and apoptotic activities of Platycodon saponins, including platycodin D, 2''-O-acetylplatycodin D, 3''-O-acetylplatycodin D, polygalacin D, 2''-O-acetylpolygalacin D, and 3''-O-acetylpolygalacin D, isolated from Platycodon grandiflorum, and prosapogenins which lack the C-3 or C-28 sugar residues, obtained from hydrolysis of platycodin D. We also clarified the structure-activity relationships of these molecules to define structural features that are crucial for the biological activity of Platycodon saponins and prosapogenins. The results showed that all Platycodon saponins had antiproliferative effects on the seven types of cancer cell lines tested. In particular, O-acetylation at the C-2 or C-3 position of rhamnose and dehydroxylation at C-24 increase the compound's cytotoxicity, while the loss of sugar residues linked to C-3 or C-28 dramatically reduced cytotoxicity. This cytotoxicity was associated with apoptosis, which was indicated by DNA fragmentation, phosphatidylserine externalization, and the activation of caspases in AGS cells. Furthermore, Platycodon saponins suppressed the phosphorylation of Akt, which resulted in the inhibition of mTOR and NF-κB signaling following the inhibition of their downstream proteins. In conclusion, six Platycodon saponins have antiproliferative activity, and the presence of sugar residues, an O-acetyl group on the rhamnose, and a methyl group at C-4 contributes to their cytotoxicity and apoptotic activity. These findings may be useful in evaluating the structure-activity relationships of Platycodon saponins and modifying them as a potent apoptosis-inducing agent.


Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Extratos Vegetais/farmacologia , Raízes de Plantas/química , Platycodon/química , Saponinas/farmacologia , Triterpenos/farmacologia , Linhagem Celular Tumoral , Humanos , Extratos Vegetais/química , Saponinas/química , Relação Estrutura-Atividade , Triterpenos/química
14.
Phytochem Anal ; 24(2): 148-54, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22930642

RESUMO

INTRODUCTION: The roots of Adenophorae species have been reported to exhibit anti-obese, anti-oxidant, anti-cancer, and anti-bacterial activities. However, there has been no single report regarding the preparative isolation and biological activities of the chemical components from Adenophora triphylla. OBJECTIVE: To develop an efficient method for the determination of the active fraction from the methanol extract from the roots of Adenophora triphylla and for the preparative isolation and purification of target compounds having cytotoxicity on carcinoma cells from the active fraction by high-speed counter-current chromatography (HSCCC). METHODS: The Plant (5 kg, dry weight) was extracted with methanol. Three hundred grams of the dried methanol extract (885 g) were fractionated by open-column chromatography with a stepwise gradient of water-methanol. Preparative isolation of bioactive components was performed by HSCCC with a two-phase solvent system composed of ethyl acetate-n-butanol-0.2% trifluoroacetic acid in water (5:5:10, v/v). The cytotoxicity of column fractions and isolated compounds was evaluated by 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay. RESULTS: The 70% MeOH column fraction showed inhibitory effects against three human carcinoma cells A549, AGS and HepG2. Two saponins were separated from 400 mg of the active fraction by HSCCC. After further purification with solid phase extraction column, 25 mg of peak fraction 1 and 20 mg of peak fraction 2 were obtained. Their structures were identified by ¹H-NMR, ¹³C-NMR, Fourier transform infrared, fast atom bombardment-MS and electrospray ionisation-MS/MS. They exhibited strong cytotoxic effects against three cancer cells. CONCLUSION: Two cytotoxic saponins were isolated for the first time from the roots of Adenophora triphylla by HSCCC.


Assuntos
Campanulaceae/química , Distribuição Contracorrente/métodos , Raízes de Plantas/química , Saponinas/isolamento & purificação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células Hep G2 , Humanos , Estrutura Molecular , Saponinas/química , Saponinas/farmacologia , Solventes/química
15.
J Sep Sci ; 35(12): 1462-9, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22740255

RESUMO

A new model of solvent gradients selection was rationally developed for the preparative separation of target compounds. The solvent gradients were selected based on a three-stage screening process where stationary phase retention was ensured by introducing a new parameter termed as the phase ratio. The phase ratio was calculated after mixing the upper phase of a solvent system with the lower phase of a different solvent system (1:1, v/v). The developed model was applied to the one-step separation of eight ginsenosides from Panax ginseng. Three gradients were selected on the basis of new model and eight ginsenosides, Rb(1), Rb(2), Rc, Rd, Re, Rg(1), Rf, and Rh(1), were efficiently separated by high-speed counter-current chromatography coupled with evaporative light scattering detector. The structures of all compounds were characterized by electrospray-ionization mass spectrometry and nuclear magnetic resonance spectroscopy.


Assuntos
Distribuição Contracorrente/métodos , Ginsenosídeos/isolamento & purificação , Panax/química , Extratos Vegetais/isolamento & purificação , Distribuição Contracorrente/instrumentação , Solventes/química
16.
J Sep Sci ; 34(10): 1116-22, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21491596

RESUMO

Ginsenosides exhibit diverse biological activities and are major well-known components isolated from the radix of Panax ginseng C.A. Meyer. In the present work, a rapid and facile method for the separation and purification of eight ginsenosides from P. ginseng by high-speed counter-current chromatography coupled with evaporative light scattering detector (HSCCC-ELSD) was successfully developed. The crude samples for HSCCC separation were first purified from ginseng extract using a macroporous resin; the extract was loaded onto a Diaion-HP20 column and fractionated by methanol and water gradient elution. The ginsenosides-protopanaxadiol (PPD) and protopanaxatriol (PPT) fractions were subsequently eluted with 65 and 80% methanol and water gradient elution, respectively. Furthermore, these two fractions were separated by HSCCC-ELSD. The two-phase solvent system used for separation was composed of chloroform/methanol/water/isopropanol at a volume ratio of 4:3:2:1. Each fraction obtained was collected and dried, yielding the following eight ginsenosides: Rg(1), Re, Rf, Rh(1), Rb(1), Rc Rb(2) and Rd. The purity of these ginsenosides was greater than 97% as assessed by HPLC-ELSD, and their structures were characterized by electrospray-ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance spectroscopy. This is the first report regarding the separation of the ginsenosides Rh(1), Rb(2) and Rc from P. ginseng by HSCCC.


Assuntos
Distribuição Contracorrente/métodos , Medicamentos de Ervas Chinesas/isolamento & purificação , Ginsenosídeos/isolamento & purificação , Panax/química , Distribuição Contracorrente/instrumentação , Medicamentos de Ervas Chinesas/química , Ginsenosídeos/química , Estrutura Molecular , Espectrometria de Massas por Ionização por Electrospray
17.
J Sep Sci ; 33(13): 1916-22, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20533341

RESUMO

Platycosides, the saponins found in the roots of Platycodon grandiflorum (Platycodi Radix), are typically composed of oleanane triterpenes with two side chains. In platycosides, platycodin D, a glucose unit at C-3, is a major component, which has several pharmacological activities. Because of the high demand for this compound, we attempted to enzymatically convert platycodin D(3) and platycoside E, having two and three glucose units at C-3, respectively, into platycodin D. In this study, we tested the ability of several glycosidases to transform platycosides, or more specifically, the ability to transform platycoside E and platycodin D(3) into platycodin D. To obtain pure platycodin D on a preparative scale, high-speed countercurrent chromatography with a solvent system of ethyl acetate/n-butanol/water (1.2:1:2, v/v/v) was used for the separation of the enzymatically transformed product. Approximately 39.4 mg of platycodin D (99.8% purity) was obtained from 200 mg of the product in a one-step separation. The results strongly support the advantage of enzymatic transformation of the platycosides for the efficient enrichment of platycodin D in the complicated extract of the medicinal plant.


Assuntos
Celulase/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Ácido Oleanólico/análogos & derivados , Saponinas/isolamento & purificação , Saponinas/metabolismo , Celulase/química , Conformação Molecular , Ácido Oleanólico/isolamento & purificação , Ácido Oleanólico/metabolismo , Platycodon/química
18.
J Chromatogr A ; 1217(26): 4375-82, 2010 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-20462588

RESUMO

Comprehensive two-dimensional chromatography (LCxLC) using combinations of two columns (C(18) x CN and C(18) x NH(2)) was employed with electrospray (ESI) mass spectrometry to analyze platycosides from root extract. Based on the capability of the C(18), CN and NH(2) columns to separate the platycosides, the orthogonality in two-dimensional space according to each combination of columns was predicted from the correlation coefficients between the retention times of the 17 compounds separated by the independent CN and C(18) columns, and NH(2) and C(18) columns. The expected distribution of the peaks was also compared with the two-dimensional plots obtained by practical separation in an LCxLC system. The increased peak capacities using C(18) x NH(2) allowed three minor components and five isomers of the platycosides to be newly separated, which were not identified with 1D-LC using the individual C(18) column, whereas the combination of C(18) x CN did not result in any improvement of the separation performance.


Assuntos
Campanulaceae/química , Glicosídeos/análise , Extratos Vegetais/análise , Espectrometria de Massas por Ionização por Electrospray/métodos , Cromatografia Líquida/métodos
19.
J Pharm Biomed Anal ; 51(1): 202-9, 2010 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-19726152

RESUMO

Platycosides, the main active constituents of Platycodi Radix, have been thoroughly studied for the characterization of their potent biological activities. However, metabolism of platycosides has not yet been characterized. A HPLC electrospray ionization-tandem mass spectrometry (LC/ESI-MS(n)) approach was applied to new complex platycoside metabolites transformed by human intestinal bacteria to identify their structures and determine metabolic pathway. The molecular weights of metabolites were identified by LC/ESI-MS analysis in both positive and negative modes. Structures for the platycoside metabolites were proposed by the molecular weights and the expected enzymatic activity of intestinal microbes on platycoside. In the second step, successive LC-MS(n) analysis was used to demonstrate the proposed structures. Under ESI tandem mass conditions, the sequential fragmentation patterns of [M+Na](+) ions exclusively showed signals, consistent with the cleavage of glycoside bonds, rearrangement and some cross-ring cleavage, thus allowing the rapid identification of platycoside metabolites. The metabolites identified in the time-dependent metabolism experiments enable us to propose several microbial pathways for platycosides. Even though the metabolites of some platycosides may have unknown structures and low levels, the analytical tools presented in this study made it possible to obtain a rapid and complete characterization of new metabolites and their metabolism pathway in human intestinal bacteria.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Platycodon/química , Saponinas/metabolismo , Espectrometria de Massas por Ionização por Electrospray/métodos , Bactérias/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Intestinos/microbiologia , Peso Molecular , Extratos Vegetais/análise , Extratos Vegetais/química , Raízes de Plantas , Saponinas/análise , Fatores de Tempo
20.
J Chromatogr A ; 1151(1-2): 37-44, 2007 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-17270197

RESUMO

Ginseng (Panax ginseng C. A. Meyer) has been well known to have a variety of ginsenosides that show diverse biological activities. Especially, the components of ginsenosides are quite different depending on the processing method. Recently, there have been several reports showing that less polar ginsenosides from Korean red ginseng (steam-treated Panax ginseng) have potent biological activities such as radical scavenging, vasodilating and anti-tumor activities. In this study, we have isolated four known ginsenosides Rg3, Rk1, Rg5 and F4 from Korean red ginseng by high-speed counter-current chromatography (HSCCC) coupled with evaporative light scattering detection (ELSD). The enriched saponin fraction (350 mg) was separated by using methylene chloride-methanol-water-isopropanol (6:6:4:1, v/v) as the two-phase solvent system and yielded 28.6 mg of Rg5, 26.6 mg of Rk1, 32.2 mg of Rg3 and 8.1 mg of F4. The purity of these ginsenosides was assessed by HPLC-ELSD to be over 95%, and their structures were characterized by electrospray ionization mass spectrometry (ESI-MS), (1)H NMR and (13)C NMR.


Assuntos
Distribuição Contracorrente/métodos , Ginsenosídeos/isolamento & purificação , Panax/química , Cromatografia Líquida de Alta Pressão , Ginsenosídeos/química , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Espectrometria de Massas por Ionização por Electrospray
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