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1.
Mutat Res ; 850-851: 503147, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32247562

RESUMO

Bulbus Fritillariacirrhosa D. Don (BFC) has been widely used as an herbal medicament for respiratory diseases in China for over 2000 years. The ethnomedicinal effects of BFC have been scientifically verified, nevertheless its toxicity has not been completely studied. Previously, we have reported that the aqueous extract of BFC induces mitotic aberrations and chromosomal instability (CIN) in human colon epithelial NCM460 cells via dysfunctioning the mitotic checkpoint. Here, we extend this study and specifically focus on the influence of BFC on cytokinesis, the final step of cell division. One remarkable change in NCM460 cells following BFC treatment is the high incidence of binucleated cells (BNCs). More detailed investigation of the ana-telophases reveals that furrow ingression, the first stage of cytokinesis, is inhibited by BFC. Asynchronous cultures treatment demonstrates that furrow ingression defects induced by BFCs are highly associated with the formation of BNCs in ensuing interphase, indicating the BNCs phenotype after BFC treatment was resulted from cytokinesis failure. In line with this, the expression of genes involved in the regulation of furrow ingression is significantly de-regulated by BFC (e.g., LATS-1/2 and Aurora-B are upregulated, and YB-1 is downregulated). Furthermore, long-term treatment of BFC elucidates that the BNCs phenotype is transient and the loss of BNCs is associated with increased frequency of micronuclei and nuclear buds, two biomarkers of CIN. In supporting of these findings, the Nin Jiom Pei Pa Koa and Chuanbei Pipa Gao, two commercially available Chinese traditional medicines containing BFC, are able to induce multinucleation and CIN in NCM460 cells. Altogether, these data provide the first in vitro experimental evidence linking BFC to cytokinesis failure and suggest the resultant BNCs may be intermediates to produce CIN progenies.


Assuntos
Instabilidade Cromossômica/efeitos dos fármacos , Citocinese/efeitos dos fármacos , Fritillaria/química , Extratos Vegetais/farmacologia , Aurora Quinase B/genética , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/genética , Instabilidade Cromossômica/genética , Colo/efeitos dos fármacos , Colo/patologia , Citocinese/genética , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Mitose/efeitos dos fármacos , Extratos Vegetais/química , Raízes de Plantas/química , Proteínas Serina-Treonina Quinases/genética , Proteína 1 de Ligação a Y-Box/genética
2.
Zhongguo Zhong Yao Za Zhi ; 45(6): 1393-1398, 2020 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-32281353

RESUMO

An UPLC method was established for the direct determination of six major bioactive isosteroidal alkaloids, namely peimisine, imperialine, sipeimine-3-D-glucoside, verticinone, verticine and hupehenine from the bulbus of Fritillaria(Beimu), a commonly used antitussive traditional Chinese medicinal(TCM) herb. An Acquity UPLC~(TM) CSH C_(18) column(2.1 mm×100 mm, 1.7 µm) was used for all analysis. The investigated six compounds were all separated with gradient mobile phase consisting of 0.02% diethylamine-water-methanol at a flow rate of 0.3 mL·min~(-1). The temperature of sample manager was set at 20 ℃. Drift tube temperature was 45 ℃, and spray parameter was 40% with injection volume of 1 µL. Then, the further quality assessment of Beimu was carried out by cluster analysis(CA) and principal component analysis(PCA). The investigated all had good linearity(r≥0.998 9) over the tested ranges. The method is simple, accurate and reproducible, and can be used for determining the content of six major bioactive isosteroidal alkaloids.


Assuntos
Alcaloides/análise , Medicamentos de Ervas Chinesas/química , Fritillaria/química , Alcaloides/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Compostos Fitoquímicos/análise , Compostos Fitoquímicos/isolamento & purificação , Raízes de Plantas/química
3.
PLoS One ; 15(2): e0229181, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32053689

RESUMO

Fritillariae cirrhosae bulbus is a famous type of traditional Chinese medicine used for cough relief and eliminating phlegm. The medicine originates from dried bulbs of five species and one variety of Fritillaria. Recently, immature bulbs from other congeneric species, such as F. ussuriensis, have been sold as adulterants of Fritillariae cirrhosae bulbus in medicine markets owing to the high price and limited availability of the genuine medicine. However, it is difficult to accurately identify the bulbs from different original species of Fritillariae cirrhosae bulbus and its adulterants based on traditional methods, although such medicines have different prices and treatment efficacies. The present study adopted DNA barcoding to identify these different species and compared the discriminatory power of super, universal, and specific barcodes in Fritillaria. The results revealed that the super-barcode had strong discriminatory power (87.5%). Among universal barcodes, matK provided the best species resolution (87.5%), followed by ITS (62.5%), rbcL (62.5%), and trnH-psbA (25%). The combination of these four universal barcodes provided the highest discriminatory power (87.5%), which was equivalent to that of the super-barcode. Two plastid genes, ycf1 and psbM-psbD, had much better discriminatory power (both 87.5%) than did other plastid barcodes, and were suggested as potential specific barcodes for identifying Fritillaria species. Phylogenetic analyses indicated that F. cirrhosa was not a "good" species that was composed of multiple lineages, which might have affected the evaluation of the discriminatory ability. This study revealed that the complete plastid genome, as super barcode, was an efficient and reliable tool for identifying the original species of Fritillariae cirrhosae bulbus and its adulterants.


Assuntos
Código de Barras de DNA Taxonômico , Fraude/prevenção & controle , Fritillaria/classificação , Fritillaria/genética , Variação Genética , Genomas de Plastídeos/genética , Filogenia
4.
Fitoterapia ; 140: 104434, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31760067

RESUMO

Fritillaria cirrhosa bulbus is a Chinese folk herb famous for its antitussive, expectorant, anti-asthma and anti-inflammatory properties, and is widely used to treat respiratory diseases. However, the impacts of F. cirrhosa bulbus on oxidative stress are still unkown. In the present study, we investigated the potential effect and mechanism of six isosteroid alkaloids with different chemical structures from F. cirrhosa bulbus on protection against cigarette smoke-induced oxidative stress in RAW264.7 macrophages. The results showed that six isosteroid alkaloids reduced reactive oxygen species (ROS) production, elevated glutathione (GSH) level and promoted heme oxygenase (HO-1) expression, which is in association with induction of NF-E2-related factor 2 (Nrf2) nuclear translocation and up-regulation of Nrf2 expression. Among these alkaloids, verticinone, verticine, imperialine-3-ß-D-glucoside, delavine and peimisine exhibited more potent effect against CSE-induced oxidative stress than that of imperialine. These findings for the first time demonstrated that F. cirrhosa bulbus may play a protective role in cellular oxidative stress by activating Nrf2-mediated antioxidant pathway. Furthermore, the differences in antioxidant effects of these alkaloids were compared, as well as the corresponding structure-activity relationships were preliminarily elucidated. This suggested that F. cirrhosa bulbus might be a promising therapeutic treatment for the prevent of oxidative stress-related diseases.


Assuntos
Alcaloides/farmacologia , Fritillaria/química , Estresse Oxidativo/efeitos dos fármacos , Fumaça/efeitos adversos , Alcaloides/isolamento & purificação , Animais , Glutationa/metabolismo , Heme Oxigenase-1/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Estrutura Molecular , Fator 2 Relacionado a NF-E2/metabolismo , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Raízes de Plantas/química , Plantas Medicinais/química , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade , Produtos do Tabaco
5.
J Ethnopharmacol ; 247: 112283, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31605736

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Bulbus Fritillaria cirrhosa D. Don (BFC) is a Chinese traditional herbal medicine that has long been used as an indispensable component in herbal prescriptions for bronchopulmonary diseases due to its well-established strong anti-inflammation and pulmonary harmonizing effects. Interestingly, there are few case reports in traditional Chinese medicine available where they found it to contribute in anti-tumor therapies. Imperialine is one of the most favored active substances extracted from BFC and has been widely recognized as an anti-inflammatory agent. AIM OF THE STUDY: The aim of the current work is to provide first-hand evidences both in vitro and in vivo showing that imperialine exerts anti-cancer effects against non-small cell lung cancer (NSCLC), and to explore the molecular mechanism of this anti-tumor activity. It is also necessary to examine its systemic toxicity, and to investigate how to develop strategies for feasible clinical translation of imperialine. MATERIALS AND METHODS: To investigate anti-NSCLC efficacy of imperialine using both in vitro and in vivo methods where A549 cell line were chosen as in vitro model NSCLC cells and A549 tumor-bearing mouse model was constructed for in vivo study. The detailed underlying anti-cancer mechanism has been systematically explored for the first time through a comprehensive set of molecular biology methods mainly including immunohistochemistry, western blot and enzyme-linked immunosorbent assays. The toxicity profile of imperialine treatments were evaluated using healthy nude mice by examining hemogram and histopathology. An imperialine-loaded liposomal drug delivery system was developed using thin film hydration method to evaluate target specific delivery. RESULTS: The results showed that imperialine could suppress both NSCLC tumor and associated inflammation through an inflammation-cancer feedback loop in which NF-κB activity was dramatically inhibited by imperialine. The NSCLC-targeting liposomal system was successfully developed for targeted drug delivery. The developed platform could favorably enhance imperialine cellular uptake and in vivo accumulation at tumor sites, thus improving overall anti-tumor effect. The toxicity assays revealed imperialine treatments did not significantly disturb blood cell counts in mice or exert any significant damage to the main organs. CONCLUSIONS: Imperialine exerts anti-cancer effects against NSCLC both in vitro and in vivo, and this previously unknown function is related to NF-κB centered inflammation-cancer feedback loop. Imperialine mediated anti-cancer activity is not through cytotoxicity and exhibit robust systemic safety. Furthermore, the liposome-based system we commenced would dramatically enhance therapeutic effects of imperialine while exhibiting extremely low side effects both on cellular and in NSCLC model. This work has identified imperialine as a promising novel anti-cancer compound and offered an efficient target-delivery solution that greatly facilitate practical use of imperialine.


Assuntos
Alcaloides/administração & dosagem , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Cevanas/administração & dosagem , Medicamentos de Ervas Chinesas/administração & dosagem , Fritillaria/química , Neoplasias Pulmonares/tratamento farmacológico , Células A549 , Alcaloides/efeitos adversos , Alcaloides/química , Alcaloides/isolamento & purificação , Animais , Contagem de Células Sanguíneas , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Cevanas/efeitos adversos , Cevanas/química , Cevanas/isolamento & purificação , Medicamentos de Ervas Chinesas/efeitos adversos , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/isolamento & purificação , Retroalimentação Fisiológica/efeitos dos fármacos , Humanos , Lipossomos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , NF-kappa B/antagonistas & inibidores , NF-kappa B/imunologia , Testes de Toxicidade , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Molecules ; 24(18)2019 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-31500338

RESUMO

Fritillariae Bulbus is a precious Chinese herbal medicine that is grown at high elevation and used to relieve coughs, remove phlegm, and nourish the lungs. Historically, Fritillariae Bulbus has been divided into two odourless crude drugs: Fritillariae Cirrhosae Bulbus and Fritillariae Thunbergii Bulbus. However, now the Chinese Pharmacopoeia has described five Fritillariae Bulbus-the new additions include Fritillariae Pallidiflorae Bulbus, Fritillariae Ussuriensis Bulbus, and Fritillariae Hupehensis Bulbus. Because the morphology of dried Fritillariae Bulbus is similar, it is difficult to accurately identify the different types of Fritillariae Bulbus. In the current study, we develop a method combining DNA barcoding and high-performance liquid chromatography (HPLC) to help distinguish Fritillariae Cirrhosae Bulbus from other Fritillariae Bulbus and guarantee species traceability of the five types of Fritillariae Bulbus. We report on the validation of an integrated analysis method for plant species identification using DNA barcoding that is based on genetic distance, identification efficiency, inter- and intra-specific variation, calculated nearest distance, neighbour-joining tree and barcoding gap. Our results show that the DNA barcoding data successfully identified the five Fritillariae Bulbus by internal transcribed spacer region (ITS) and ITS2, with the ability to distinguish the species origin of these Fritillariae Bulbus. ITS2 can serve as a potentially useful DNA barcode for the Fritillaria species. Additionally, the effective chemical constituents are identified by HPLC combined with a chemical identification method to classify Fritillaria. The HPLC fingerprint data and HCA (hierarchical clustering analysis) show that Fritillariae Cirrhosae Bulbus is clearly different from Fritillariae Thunbergii Bulbus and Fritillariae Hupehensis Bulbus, but there is no difference between Fritillariae Cirrhosae Bulbus, Fritillariae Ussuriensis Bulbus, and Fritillariae Pallidiflorae Bulbus. These results show that DNA barcoding and HPLC fingerprinting can discriminate between the five Fritillariae Bulbus types and trace species to identify related species that are genetically similar.


Assuntos
Código de Barras de DNA Taxonômico/métodos , Medicamentos de Ervas Chinesas/química , Fritillaria/classificação , Cromatografia Líquida de Alta Pressão , Análise por Conglomerados , DNA Mitocondrial/genética , DNA de Plantas/genética , Fritillaria/química , Fritillaria/genética , Filogenia , Análise de Sequência de DNA
7.
Zhongguo Zhong Yao Za Zhi ; 44(11): 2231-2235, 2019 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-31359647

RESUMO

In order to reveal the relationship between the amount of soil microorganisms and the quality of Fritillaria taipaiensis, both cultivated and wild F. taipaiensis were collected from Chongqing, Wuxi at different stages of their growth as objects of the research. The mycorrhizal infection rate and colonization intensity, peimisine and total alkaloid content in bulbs, the amount of microorganisms and biomass carbon content in rhizospheric soil were all determined using common methods. The results showed that the typical arbuscular-vesicle roots were formed after the AM fungi infected the F. taipaiensis roots which were collected from different origins. The mycorrhizal infection rates were ranged from 78.74% to 98.68% and the colonization intensities were ranged from 13.29% to 37.06%. The rhizospheric microorganisms of F. taipaiensis showed abundant resources. The distribution rule of them in the rhizospheric soil was as follows: the amount of bacteria>the amount of actinomycetes>the amount of fungi. The rhizospheric bacteria, decomposition inorganic phosphorus bacteria, decomposition organic phosphorus bacteria, actinomycetes amount and the total number of microbes increased first and then decreased with the increase of years, while decomposition potassium bacteria showed decreasing trend and fungi showed gradual increasing trend. The soil microbial flora content in the soil changed from "bacterial type" with a high fertility to "fungal type" with a low fertility. The mass fraction of peimisine and total alkaloid content increased first and then decreased with the increase of over the years, the same trend of culturable rhizosphere soil bacteria and actinomycetes indicated that the growth years affected the quality of soil and medicinal materials on different levels. Therefore, the diversity of microbial communities in rhizosphere soil reduced with the increase of years leading to the continuous cropping obstacles and the destruction of medicinal quality of F. taipaiensis.


Assuntos
Alcaloides/análise , Fritillaria/química , Micorrizas , Rizosfera , Microbiologia do Solo , Fritillaria/microbiologia , Raízes de Plantas
8.
J Ethnopharmacol ; 238: 111853, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-30954613

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Fritillariae Bulbus ("Beimu" in Chinese) is a famous traditional Chinese medicine used to treat cough, expectoration and asthma for more than 2000 years, which belongs to the Fritillaria genus in Liliaceae family. Bulbs of Fritillaria cirrhosa D.Don (BFC) and bulbs of Fritillaria pallidiflora Schrenk (BFP) are two important drugs of Beimu. Due to the significant similarities in their outward appearance characters and chemical profiles, BFC has often been adulterated with BFP in Chinese Traditional Medicine markets. AIM OF THE STUDY: This study aims to compare the oral acute toxicity and the traditional pharmacological activities including antitussive, expectorant and anti-inflammatory effects between the extract of BFC and BFP, to clear and definite if the BFP can be used as a substitute of the BFC in the application of traditional medicine. MATERIALS AND METHODS: The extracts were prepared through refluxing with 80% ethanol solvent. For the acute toxicity tests, graded doses of BFP extracts and the maximum dose of BFC extracts were administered orally to mice. The animals were observed for toxic symptoms and mortality daily for 14 days. For the pharmacological activities tests, graded doses of BFP and BFC extracts were administered orally to mice. To observe the effects relieving cough, expelling phlegm and lessening the ear swelling of BFC extracts and BFP extracts through ammonia liquor inducing cough, phenol red apophlegmating in mice and the xylene-induced auricular swelling of mouse, respectively. RESULTS: In the acute toxicity study, the LD50 value of BFP in mice was calculated to be 213.57 g/kg body weight, and the maximum feasible dose (MFD) value of BFC in mice was 452.14 g/kg. Histopathological analysis has shown inflammatory cells infiltration and cells edema in liver, multinucleated giant cell proliferation in spleen, perivascular exudate and hemorrhage in lung, glomerulus atrophy in kidney of mice after oral administrations of BFP extracts. But only liver cells edema was observed in BFC group. Both BFC extract and BFP extract significantly increased latent period of cough and inhibited cough frequency in mice induced by ammonia. Besides, the two extracts also obviously enhanced mice's tracheal phenol red output in expectorant assessment and inhibited the development of ear edema in anti-inflammatory evaluation assay. CONCLUSION: To summarize, the BFP has the significant similarities in morphological characteristics, chemical profiles and traditional pharmacological activities compared with the BFC. The result of this study provide some valid scientific support for using BFP as a plant substitute of the BFC, but considering the toxicity of BFP is much higher than BFC, we don't recommend long-term oral administration of BFP or exceeding recommended dosage of Chinese Pharmacopoeia 2015.


Assuntos
Anti-Inflamatórios , Antitussígenos , Tosse/tratamento farmacológico , Edema/tratamento farmacológico , Expectorantes , Fritillaria , Extratos Vegetais , Administração Oral , Animais , Anti-Inflamatórios/uso terapêutico , Anti-Inflamatórios/toxicidade , Antitussígenos/uso terapêutico , Antitussígenos/toxicidade , Expectorantes/uso terapêutico , Expectorantes/toxicidade , Feminino , Rim/efeitos dos fármacos , Rim/patologia , Fígado/efeitos dos fármacos , Fígado/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Medicina Tradicional Chinesa , Camundongos , Extratos Vegetais/uso terapêutico , Extratos Vegetais/toxicidade , Raízes de Plantas , Baço/efeitos dos fármacos , Baço/patologia , Testes de Toxicidade Aguda
9.
Zhongguo Zhong Yao Za Zhi ; 44(3): 448-453, 2019 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-30989906

RESUMO

A new variety "Zhebei 3(Zhejiao Pharmaceutical 2018002)" was selected and bred from multi seeded Fritillaria thunbergii mutants by systematic breeding method. From 2012 to 2016, the traits assessment, disease resistance appraisal, plot ratios and regional trials of the variety were continuously carried out. The results showed that "Zhebei 3" emerged early and had late seedlings. The average growth period was about 100 days, which was 6 days and 12 days higher than the "Zhebei 1" and "Zhebei 2". The average yield was 5 095.5 kg·hm~(-2), which was 14.42% and 17.71% higher than of the control respectively. The average proliferation rate of bulbs was 261.2%, which was 37.46% and 31.58% higher than that of the control, respectively. The propagation coefficient of bulbs was about 1∶2.6, and the total amount of peimine and peiminine was 0.172 2%, which was 4.49% and 29.47% higher than the control, respectively. The identification of disease resistance showed that it was resistance to bulb stem(soft) rot, better than the control. "Zhebei 3" has stable characters, high yield, good quality, strong disease resistance, and moderate propagation coefficient which is suitable for planting in Zhejiang province.


Assuntos
Fritillaria/crescimento & desenvolvimento , Melhoramento Vegetal , Resistência à Doença , Doenças das Plantas , Raízes de Plantas
10.
Zhongguo Zhong Yao Za Zhi ; 44(3): 495-499, 2019 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-30989914

RESUMO

Twelve alkaloids were isolated from the bulbs of Fritillaria yuminensis by column chromatography over silica gel, ODS, and Sephadex LH-20, as well as RP-HPLC. Their structures were identified mainly by NMR and MS analyses as yubeinine(1), imperialine(2), delavinone(3), tortifoline(4), hupehenizioiside(5), imperialine-ß-D-glucoside(6), kuroyurinidine(7), pengbeisine A(8), walujewine A(9), peimisine-3-O-ß-D-glucopyranoside(10), solanidine-3-O-α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranoside(11), and solanidine-3-O-α-L-rhamnopyranosyl-(1→2)-[ß-D-glucopyranosyl-(1→4)]-ß-D-glucopyranoside(12). Compounds 4-12 were obtained from F. yuminensis for the first time.


Assuntos
Alcaloides/análise , Fritillaria/química , Raízes de Plantas/química , Cromatografia Líquida de Alta Pressão , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Compostos Fitoquímicos/análise
11.
Biomed Res Int ; 2019: 3163218, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31016188

RESUMO

Thirty-one compounds, including delavinone, were isolated from the methanol extract of F. cirrhosa by modern chromatographic techniques. The pharmacological action of Fritillaria is widely used in clinical practice. However, the pharmacokinetic studies on delavinone have not been reported. Therefore, the chemical constituents of this species were investigated. Therefore, it is necessary to establish an analytical method to monitor the concentration of delavinone. An UPLC-MS/MS method was established to determine delavinone in the mouse blood, and the pharmacokinetics of delavinone after intravenous (1.0 mg/kg) and intragastric (2.5, 10.0 mg/kg) administration were studied. The lower limit of quantification was 1.0 ng/mL. The intraday and interday precision RSD were less than 13%, the accuracy ranged from 96.8% to 104.9%, the average recovery was better than 80.6%, and the matrix effect was between 88.8% and 103.4%. The UPLC-MS/MS method has been successfully applied to the pharmacokinetics of delavinone in mice. The noncompartment model was used to fit the main pharmacokinetic parameters. It was found that AUC in mice was higher than that in mice given orally, and the bioavailability of delavinone was 12.4%.


Assuntos
Fritillaria/química , Extratos Vegetais/química , Extratos Vegetais/farmacocinética , Administração Intravenosa/métodos , Administração Oral , Animais , Disponibilidade Biológica , Cromatografia Líquida de Alta Pressão/métodos , Camundongos , Camundongos Endogâmicos ICR , Espectrometria de Massas em Tandem/métodos
12.
Int J Biol Macromol ; 131: 97-106, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30844453

RESUMO

In this paper, Fritillaria pallidiflora schrenk polysaccharides were extracted with different methods and isolated by anion-exchange and gel-permeation chromatography. Physicochemical properties, structural characteristics and antioxidant activities were investigated for the first time. The polysaccharides composed of glucose, galactose, arabinose, xylose, mannose and rhamnose with different molar ratio and molecular weight. Fourier transform infrared spectroscopy and nuclear magnetic resonance analysis exhibited the presence of key functional groups of polysaccharides whereas scanning electron microscopy analysis revealed the characteristic morphology of different fractions. The thermogravimetric analysis manifested the thermal stability of polysaccharides. The reducing power and antioxidant activities of polysaccharides were evaluated in vitro. Among them, the acidic fraction FPSP-H2-1 has a strong antioxidant effect against DPPH, hydroxyl and ABTS free radicals. The results revealed the polysaccharide's important role as potential natural antioxidant agents during the investigation and application of bioactive polysaccharide.


Assuntos
Antioxidantes/química , Antioxidantes/farmacologia , Fritillaria/química , Polissacarídeos/química , Polissacarídeos/farmacologia , Antioxidantes/isolamento & purificação , Fracionamento Químico , Relação Dose-Resposta a Droga , Depuradores de Radicais Livres/química , Depuradores de Radicais Livres/isolamento & purificação , Depuradores de Radicais Livres/farmacologia , Peso Molecular , Monossacarídeos/química , Polissacarídeos/isolamento & purificação , Análise Espectral , Termogravimetria
13.
J Biosci Bioeng ; 127(2): 231-240, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30177486

RESUMO

Exopolysaccharides (EPSs) are high-molecular-weight carbohydrates with a wide range of biophysiological activities, such as antioxidant activity, immunostimulatory activity, antitumor activity, hepatoprotective activity, and antifatigue effects. In the present work, two water-soluble EPSs, namely, A14EPS-1 and A14EPS-2, were isolated and purified from the fungal endophytic strain A14 using ethanol precipitation, DEAE-cellulose ion exchange chromatography and Sepharose G-150 gel filtration chromatography. A14EPS-1 (∼2.4 × 104 Da, the major fraction) was mainly composed of mannose, rhamnose, glucose, galactose, xylose and arabinose with a molar ratio of 0.31:0.55:10.00:0.34:0.03:0.06. The major monosaccharide of A14EPS-1 was pyranose, which was connected by α-glycosidic linkages. And the side chains of A14EPS-1 may be composed of rhamnose, arabinose, glucose and galactose; moreover, the backbone of A14EPS-1 may be composed of rhamnose, xylose, arabinose and glucose. A14EPS-2 (∼0.5 × 104 Da) was mainly composed of mannose, rhamnose, glucose, galactose, xylose and arabinose in a ratio of 0.16:0.88:10.00:0.39:0.06:0.06. Pyranose was observed in both the α- and ß-configurations in A14EPS-2, and the α configuration was dominant. In addition, the results of the bioactivity assays indicated that both A14EPS-1 and A14EPS-2 had moderate antioxidant activity in vitro, and A14EPS-2 showed a moderate antiproliferation effect on human hepatocellular carcinoma HepG2 cells.


Assuntos
Antioxidantes/farmacologia , Citostáticos/farmacologia , Fritillaria/microbiologia , Fusarium/isolamento & purificação , Fusarium/metabolismo , Polissacarídeos/farmacologia , Antioxidantes/metabolismo , Carboidratos/farmacologia , Proliferação de Células/efeitos dos fármacos , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Citostáticos/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Fusarium/química , Glicosídeos/metabolismo , Glicosídeos/farmacologia , Células Hep G2 , Humanos , Polissacarídeos/química , Polissacarídeos/metabolismo
14.
Cell Physiol Biochem ; 51(4): 1566-1583, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30497066

RESUMO

BACKGROUND/AIMS: Glioblastoma multiforme (GBM) is the most devastating and widespread primary central nervous system tumour in adults, with poor survival rate and high mortality rates. Existing treatments do not provide substantial benefits to patients; therefore, novel treatment strategies are required. Peiminine, a natural bioactive compound extracted from the traditional Chinese medicine Fritillaria thunbergii, has many pharmacological effects, especially anticancer activities. However, its anticancer effects on GBM and the underlying mechanism have not been demonstrated. This study was conducted to investigate the potential antitumour effects of peiminine in human GBM cells and to explore the related molecular signalling mechanisms in vitro and in vivo Methods: Cell viability and proliferation were detected with MTT and colony formation assays. Morphological changes associated with autophagy were assessed by transmission electron microscopy (TEM). The cell cycle rate was measured by flow cytometry. To detect changes in related genes and signalling pathways in vitro and in vivo, RNA-seq, Western blotting and immunohistochemical analyses were employed. RESULTS: Peiminine significantly inhibited the proliferation and colony formation of GBM cells and resulted in changes in many tumour-related genes and transcriptional products. The potential anti-GBM role of peiminine might involve cell cycle arrest and autophagic flux blocking via changes in expression of the cyclin D1/CDK network, p62 and LC3. Changes in Changes in flow cytometry results and TEM findings were also observed. Molecular alterations included downregulation of the expression of not only phospho-Akt and phospho-GSK3ß but also phospho-AMPK and phospho-ULK1. Furthermore, overexpression of AKT and inhibition of AKT reversed and augmented peiminine-induced cell cycle arrest in GBM cells, respectively. The cellular activation of AMPK reversed the changes in the levels of protein markers of autophagic flux. These results demonstrated that peiminine mediates cell cycle arrest by suppressing AktGSk3ß signalling and blocks autophagic flux by depressing AMPK-ULK1 signalling in GBM cells. Finally, peiminine inhibited the growth of U251 gliomas in vivo. CONCLUSION: Peiminine inhibits glioblastoma in vitro and in vivo via arresting the cell cycle and blocking autophagic flux, suggesting new avenues for GBM therapy.


Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Autofagia/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Cevanas/uso terapêutico , Glioblastoma/tratamento farmacológico , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Antineoplásicos Fitogênicos/farmacologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Cevanas/farmacologia , Feminino , Fritillaria/química , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transdução de Sinais/efeitos dos fármacos
15.
Fitoterapia ; 131: 112-118, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30336193

RESUMO

Three new isosteroidal alkaloids, frititorines A-C (1-3), were isolated from the bulbs of Fritillaria tortifolia, together with ten known ones (4-13). Their structures were elucidated by extensive spectroscopic analyses, chemical methods, and single-crystal X-ray crystallographic analysis. Compound 1 is the first 5ß-cevanine alkaloid with a cis A/B ring junction from the Fritillaria genus. Compound 2 is the first example of glycosylated isosteroidal alkaloid N-oxide. Compound 1 showed significant relaxant effect on Ach-induced tracheal contraction with pA2 and EC50 values equivalent to those of aminophylline.


Assuntos
Alcaloides/isolamento & purificação , Fritillaria/química , Raízes de Plantas/química , Traqueia/efeitos dos fármacos , Animais , China , Técnicas In Vitro , Estrutura Molecular , Compostos Fitoquímicos/isolamento & purificação , Ratos Sprague-Dawley
16.
Molecules ; 23(7)2018 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-29958456

RESUMO

A simple and high sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the simultaneous determination of peimine and peiminine in beagle dog plasma after the oral administration of Fritillariae ussuriensis Maxim and Fritillariae thunbergii Miq powder. Chromatographic separation was achieved on an ACQUIT UPLC® BEH C18 column (1.7 µm, 2.1 × 100 mm) in a gradient elution way with a mobile phase consisting of acetonitrile and water containing 0.1% formic acid at a flow rate of 0.4 mL/min. The plasma samples were prepared by a liquid⁻liquid extraction (LLE) method with ethyl acetate. The analytes were detected with a triple quadrupole tandem mass spectrometry (MS) in multiple reaction monitoring (MRM) mode and a positive ion electrospray ionization (ESI) of the transitions at m/z 432.4→414.4 for peimine and m/z 430.3→412.3 for peiminine. The method was linear for two analytes over the investigated range with all determined correlation coefficients exceeding 0.9900. The lower limit of quantification (LLOQ) was 0.988 ng/mL for peimine and 0.980 ng/mL for peiminine. The mean extraction recoveries of peimine and peiminine at three quality control samples (QC) levels were ranged from 82.56 to 88.71%, and matrix effects ranged from 92.06 to 101.2%. The intra-day and inter-day precision and accuracy were within the acceptable limits at LLOQ and QC levels. The method was effectively and successfully applied to the pharmacokinetics of peimine and peiminine after oral administration of powder to beagle dogs. The obtained results may be help to guide the clinical application of Fritillaria ussuriensis Maxim and Fritillaria thunbergii Miq.


Assuntos
Cevanas/sangue , Cevanas/farmacocinética , Cromatografia Líquida/métodos , Medicamentos de Ervas Chinesas/química , Fritillaria/química , Espectrometria de Massas em Tandem/métodos , Administração Oral , Animais , Cães , Medicamentos de Ervas Chinesas/administração & dosagem , Masculino
17.
J Ethnopharmacol ; 223: 142-151, 2018 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-29751123

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Fritillaria cirrhosa D. Don bulbs contain alkaloids and are one of the most intensively exploited alpine Himalayan medicinal species. In terms of proprietary medicines, our study shows that 210 F. cirrhosa products are offered by 46 suppliers, most of which (44) are situated in China and two in Nepal. A widespread commercial use is as one of the main ingredients in cough syrups. A well known example is "Nin Jiom Pei Pa Koa Herbal Cough & Throat Syrup", which typically contains more F. cirrhosa than any other herbal ingredient in the formulation. The biggest market for F. cirrhosa bulbs is China, where demand exceeds supply of this wild harvested species for use in traditional Chinese medicine (TCM). Cross-border trade from Nepal to China occurs in significant quantities. Bhutan also imports F. cirrhosa bulbs from Nepal. In addition, F. cirrhosa is registered as an active ingredient in traditional herbal medicinal preparations in Australia, Canada, Hong Kong SAR, Malaysia, Republic of Korea, Singapore and Taiwan. There is also an export trade in F. cirrhosa to Europe. Assessing how much F. cirrhosa is traded is complex, however, due to a "look-alike" challenge, as nine Chinese Fritillaria species are traded in Europe (Fritillaria cirrhosa, F. delavayi, F. hupehensis, F. pallidiflora, F. przewalskii, F. thunbergii, F. unibracteata, F. ussuriensis and F. walujewii). AIMS OF THE STUDY: The aims of this review were to assess the scale of the global trade in F. cirrhosa, and to synthesise studies of the impacts of wild harvest on F. cirrhosa populations and on the extent of emerging cultivation initiatives as an alternative to wild harvest. METHODS: Firstly, we reviewed published information on studies on impacts of wild F. cirrhosa harvest from across the geographic range of this species. Secondly, global trade data for F. cirrhosa were analysed. RESULTS: The principal demand for F. cirrhosa bulbs is in China, where hundreds of different companies produce Fritillaria preparations. Trade data also show that in 2013, China exported over 44 tonnes of F. cirrhosa bulbs to Taiwan and 26.7 tonnes to the Republic of Korea. Extensive commercial use and limited wild stocks result in a high price (2000 - 3800 CNY per kg (around US$ 303 -560 per kg in 2017)) for F. cirrhosa bulbs. Prices of cultivated Fritillaria bulbs are much lower (600-680 CNY per kg in 2017) than wild harvested bulbs. But due to very specific growth requirements of F. cirrhosa, cultivation is not yet able to meet total demand. The consequence is continued exploitation of wild stocks. At the same time, however, an increasing proportion of the demand is met by cultivation of alternative Fritillaria species that are easier to grow than F. cirrhosa. The air-dry mass of F. cirrhosa bulbs varies between 0.0917 and 0.1116 g per bulb. This represents 8960 - 10,900 bulbs/kg or 8.9 - 10.9 million bulbs per tonne. Current demand therefore represents billions of bulbs per year. CONCLUSIONS: Demand for F. cirrhosa bulbs, particularly from China, makes this species one of the most intensively harvested alpine Himalayan medicinal bulbs. Although F. cirrhosa is listed as a Class III protected species in China, billions of these tiny, wild harvested bulbs are sold per year. Due to demand exceeding supply, the price of F. cirrhosa bulbs has increased dramatically. Between 2002 and 2017, for example, the price of wild harvested F. cirrhosa bulbs increased over nine-fold, from the equivalent of US$60 in 2002 to US$560 per kg in 2017. To date, cultivation has been unable to meet the entire market demand for F. cirrhosa bulbs, although other Fritillaria species are successfully cultivated on a larger scale.


Assuntos
Fritillaria , Altitude , China , Comércio , Internacionalidade , Medicina Tradicional , Nepal
18.
PLoS One ; 13(3): e0194613, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29543905

RESUMO

Fritillaria spp. constitute important traditional Chinese medicinal plants. Xinjiang is one of two diversity hotspots in China in which eight Fritillaria species occur, two of which are endemic to the region. Furthermore, the phylogenetic relationships of Xinjiang Fritillaria species (including F. yuminensis) within the genus are unclear. In the present study, we sequenced the chloroplast (cp) genomes of seven Fritillaria species in Xinjiang using the Illumina HiSeq platform, with the aim of assessing the global structural patterns of the seven cp genomes and identifying highly variable cp DNA sequences. These were compared to previously sequenced Fritillaria cp genomes. Phylogenetic analysis was then used to evaluate the relationships of the Xinjiang species and assess the evolution of an undivided stigma. The seven cp genomes ranged from 151,764 to 152,112 bp, presenting a traditional quadripartite structure. The gene order and gene content of the seven cp genomes were identical. A comparison of the 13 cp genomes indicated that the structure is highly conserved. Ten highly divergent regions were identified that could be valuable in phylogenetic and population genetic studies. The phylogenetic relationships of the 13 Fritillaria species inferred from the protein-coding genes, large single-copy, small single-copy, and inverted repeat regions were identical and highly resolved. The phylogenetic relationships of the species corresponded with their geographic distribution patterns, in that the north group (consisting of eight species from Xinjiang and Heilongjiang in North China) and the south group (including six species from South China) were basically divided at 40°N. Species with an undivided stigma were not monophyletic, suggesting that this trait might have evolved several times in the genus.


Assuntos
Fritillaria/classificação , Fritillaria/genética , Marcadores Genéticos , Genoma de Cloroplastos , DNA de Cloroplastos/análise , DNA de Cloroplastos/genética , Marcadores Genéticos/genética , Especiação Genética , Filogenia , Plantas Medicinais/classificação , Plantas Medicinais/genética , Análise de Sequência de DNA
19.
Int J Biol Macromol ; 112: 1073-1083, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29447973

RESUMO

Rich polysaccharides were directly observed in the bulbs of Fritillaria unibracteata var. wabuensis (FUW) using the periodic acid-Schiff (PAS) method and microexamination. An acidic water-soluble heteropolysaccharide (FWPS1-1) was isolated from FUW through ethanol precipitation, decoloration, deproteinization, dialysis and separation using a DE-52 anion-exchange column and a Sepharose G-150 gel filtration column. FWPS1-1 (average molecular weight: ~7.44 kDa) has many branches and long side chains; holds the triple-helix conformation; was composed of mannose (Man), galacturonic acid (GalA), galactose (Gal), xylose (Xyl) and arabinose (Ara) with a molar ratio of 2.62:5.59:10.00:0.76:9.38; and features side chains that may be composed of Ara, Man, Gal and GalA, while the backbone may be composed of Xyl, Ara and Gal. In addition, the backbone of FWPS1-1 mainly consists of α-type glycosidic bonds. Bioactivity tests in vitro showed that the polysaccharide exhibited weak 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity and low ferric reducing antioxidant power (FRAP) but high 2,2-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)l (ABTS) radical scavenging activity, good Fe(II)-chelating ability and remarkable DNA damage protective activity. FWPS1-1 was the first heteropolysaccharide purified from FUW and showed good antioxidant activity and DNA protective effect. The results confirmed that macromolecule is also bioactive ingredient that requires attention like the small-molecule active compounds in FUW.


Assuntos
Antioxidantes/farmacologia , Fritillaria/química , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Benzotiazóis/química , Compostos de Bifenilo/química , Configuração de Carboidratos , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Delgada , Misturas Complexas/farmacologia , Dano ao DNA , Depuradores de Radicais Livres/química , Fritillaria/citologia , Iodo , Quelantes de Ferro/farmacologia , Monossacarídeos/análise , Oxirredução , Picratos/química , Polissacarídeos/química , Espectroscopia de Prótons por Ressonância Magnética , Padrões de Referência , Espectroscopia de Infravermelho com Transformada de Fourier , Ácidos Sulfônicos/química
20.
J Food Drug Anal ; 26(1): 337-347, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29389572

RESUMO

Thunberg fritillary bulb (the dry bulbs of Fritillaria thunbergii Miq.), a traditional Chinese Medicine, is widely applied as an expectorant and antitussive. In this investigation, the primary metabolites of bulbs, flowers, leaves, and stems of F. thunbergii were analyzed by gas chromatography-mass spectrometry. Principal component analysis, partial least squares-discriminate analysis, orthogonal projection to latent structures-discriminate analysis, and heat map analysis showed that there were dissimilar metabolites, and a negative correlation between amino acids and saccharides in different analytes. Furthermore, carbodiimide, tryptophan, glucose-6-phosphate, xylose, 2-piperidinecarboxylic acid, monoamidomalonic acid, phenylalanine, and histidine were found to play an important role in the plant metabolism net of F. thunbergii.


Assuntos
Fritillaria/química , Cromatografia Gasosa-Espectrometria de Massas , Metaboloma , Metabolômica , Biologia Computacional/métodos , Análise de Dados , Fritillaria/metabolismo , Redes e Vias Metabólicas , Metabolômica/métodos
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