naphthoquinones and chimaphilin

Epithelial-to-mesenchymal transition is a cellular process associated with cancer invasion and metastasis. However, the antimetastatic effects of chimaphilin remain elusive. In this study, we attempted to investigate the potential use of chimaphilin as an inhibitor of TGF-β1-induced epithelial-to-mesenchymal transition in U2OS cells. We found that TGF-β1 induced epithelial-to-mesenchymal transition to promote U2OS cell invasion and metastasis. Western blotting demonstrated that chimaphilin inhibited U2OS cell invasion and migration, increased the expression of the epithelial phenotype marker E-cadherin, repressed the expression of the mesenchymal phenotype marker vimentin, as well as decreased the level of epithelial-to-mesenchymal-inducing transcription factors Snail1 and Slug during the initiation of TGF-β1-induced epithelial-to-mesenchymal transition. In this study, we revealed that chimaphilin up-regulated the E-cadherin expression level and inhibited the production of vimentin, Snail1, and Slug in TGF-β1-induced U2OS cells by blocking PI-3K/Akt and ERK 1/2 signaling pathway. Additionally, the TGF-β1-mediated phosphorylated levels of Smad2/3 were inhibited by chimaphilin pretreatment. Above all, we conclude that chimaphilin represents an effective inhibitor of the metastatic potential of U2OS cells through suppression of TGF-β1-induced epithelial-to-mesenchymal transition.

Topics: Biomarkers, Tumor; Cadherins; Cell Death; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation, Neoplastic; Humans; Naphthoquinones; Neoplasm Invasiveness; Neoplasm Metastasis; Osteosarcoma; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Signal Transduction; Smad Proteins; Transcription Factors; Transforming Growth Factor beta1; Vimentin

A novel and effective extraction method, namely negative pressure cavitation-microwave assisted extraction technique (NMAE), was developed for the preparation of extracts of Pyrola incarnata Fisch., which are rich in the main constituents hyperin, 2'-O-galloylhyperin and chimaphilin. Single factor experiments and Box-Behnken design (BBD) were combined with a response surface methodology to examine factors affecting extraction. Maximum extraction yields of hyperin, 2'-O-galloylhyperin and chimaphilin (1.339±0.029, 4.831±0.117 and 0.329±0.011mg/g, respectively) were achieved under the following optimised conditions: 700W microwave power, 50°C extraction temperature, 30:1mL/g liquid-solid ratio, -0.05MPa negative pressure, 55% ethanol concentration and 12min extraction time. First-order kinetics equation demonstrated that NMAE offered significant savings in extraction time, and enhancing extraction efficiency. Furthermore, NMAE extracts yielded excellent antioxidant activity (IC50 0.121mg/mL for DPPH 2.896mmol FeSO4/g DW FRAP).

Topics: Antioxidants; Gallic Acid; Microwaves; Naphthoquinones; Plant Extracts; Pressure; Pyrola; Quercetin

Chimaphilin, an active compound separated from pyrola, possesses the highly efficient antitumor activities. Insulin-like growth factor-I receptor (IGF-IR) plays an important role in tumor cell survival. To look for effective strategies for interrupting IGF-IR signaling pathway, we found that chimaphilin can inhibit the receptor tyrosine kinase activity of IGF-IR. Chimaphilin inhibited the growth of both drug-sensitive and drug-resistant osteosarcoma cell lines in a time and dose-dependent manner; however, it showed relatively little toxicity in normal osteoblast cell lines. Chimaphilin can increase the sensitivity of doxorubicin in doxorubicin-resistant osteosarcoma cell lines. Additionally, small interfering RNA downregulation of IGF-IR expression in drug-resistant cell lines also caused resensitization to doxorubicin. Above all, we conclude that chimaphilin represents a valuable natural source and may potentially be applicable for reversing the drug-resistant phenotype in osteosarcoma therapy.

Topics: Apoptosis; Bone Neoplasms; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Humans; Naphthoquinones; Osteosarcoma; Receptor, IGF Type 1

A novel extraction method, homogenate-assisted negative pressure cavitation extraction (HNPCE), was designed for the extraction and determination of the main phenolic compounds of Pyrola incarnata Fisch. by LC-MS/MS. The particle sizes and extraction yields in the process of homogenization were compared with conventional pulverization. The results showed that homogenization for less than 120 s could produce more suitable particle size powders for analyte extraction. The following NPCE parameters were optimized by a BBD test and under the optimal conditions, the maximum extraction yields of arbutin, epicatechin, hyperin, 2'-O-galloylhyperin and chimaphilin increased by 68.7%, 72.0%, 43.3%, 62.5% and 34.5% with respect to normal NPCE. The LC-MS/MS method was successfully applied for the quantification of five target compounds in pyrola, and the results of the precision test indicated a high accuracy of the present method for the quantification of the target compounds in pyrola. Furthermore, the antioxidant activities of the pyrola extracts were also determined. The results showed that pyrola had good antioxidant activities and it was a valuable antioxidant natural source.

Topics: Antioxidants; Arbutin; Catechin; Chromatography, Liquid; Gallic Acid; Naphthoquinones; Phenols; Plant Extracts; Pyrola; Quercetin; Reproducibility of Results; Tandem Mass Spectrometry

Chimaphilin, 2,7-dimethyl-1,4-naphthoquinone, is extracted from pyrola [Passiflora incarnata Fisch.]. In this study, the anticancer activity and underlying mechanisms of chimaphilin toward human breast cancer MCF-7 cells are firstly investigated. Chimaphilin could inhibit the viability of MCF-7 cells in a concentration-dependent manner, and the IC50 value was 43.30μM for 24h. Chimaphilin markedly induced apoptosis through the investigation of characteristic apoptotic morphological changes, nuclear DNA fragmentation, annexin V-FITC/propidium iodide (PI) double staining. Flow cytometry assay revealed that chimaphilin triggered a significant generation of ROS and disruption of mitochondrial membrane potential. Additionally, western blotting assay showed that chimaphilin suppressed Bcl-2 level and enhanced Bad level, then activated caspase-9 and caspase-3, and further activated the poly ADP-ribose polymerase (PARP), finally induced cell apoptosis involving the mitochondrial pathway. Furthermore, free radical scavengers N-acetyl-L-cysteine (NAC) pretreatment test testified that chimaphilin could increase the generation of ROS, then induce cell apoptosis. In general, the present results demonstrated that chimaphilin induced apoptosis in human breast cancer MCF-7 cells via a ROS-mediated mitochondrial pathway.

Topics: Acetylcysteine; Annexin A5; Apoptosis; bcl-2-Associated X Protein; bcl-Associated Death Protein; Caspase 3; Caspase 9; Cell Proliferation; DNA Fragmentation; Fluorescein-5-isothiocyanate; Humans; Inhibitory Concentration 50; MCF-7 Cells; Membrane Potential, Mitochondrial; Mitochondria; Naphthoquinones; Passiflora; Propidium; Reactive Oxygen Species

Bioassay-guided fractionation of Chimaphila umbellata (L.) W. Bart (Pyrolaceae) ethanol extracts led to the identification of 2,7-dimethyl-1,4-naphthoquinone (chimaphilin) as the principal antifungal component. The structure of chimaphilin was confirmed by 1H and 13C NMR spectroscopy. The antifungal activity of chimaphilin was evaluated using the microdilution method with Saccharomyces cerevisiae (0.05mg/mL) and the dandruff-associated fungi Malassezia globosa (0.39mg/mL) and Malassezia restricta (0.55mg/mL). Pronounced antioxidant activity of C. umbellata crude extract was also identified using the DPPH (2,2-diphenyl-1-picrylhydrazyl) assay, suggesting this phytomedicine has an antioxidant function in wound healing. A chemical-genetic profile was completed with chimaphilin using approximately 4700 S. cerevisiae gene deletion mutants. Cellular roles of deleted genes in the most susceptible mutants and secondary assays indicate that the targets for chimaphilin include pathways involved in cell wall biogenesis and transcription.

Topics: Antifungal Agents; Antioxidants; Biphenyl Compounds; Dose-Response Relationship, Drug; Hydrazines; Magnetic Resonance Spectroscopy; Malassezia; Microbial Sensitivity Tests; Molecular Structure; Naphthoquinones; Picrates; Pyrolaceae; Saccharomyces cerevisiae; Structure-Activity Relationship

To investigate the chemical constituents of Pyrola calliatha.. The chemical constituents were isolated by various column chromatographic methods. The structures were identified by spectral data.. Ten compounds were isolated and identified as chimaphilin (1), uvaol(2), ursolic acid (3), 2beta,3beta,23-trihydroxy-12-ene-28-ursolic acid (4), daucosterol (5), 2alpha,3beta,23,24-tetrahydroxy-12-ene-28-ursolic acid (6), emodin (7), gallic acid (8), monotropein (9), adenosine (10).. Compounds 2,4,6,7,10 were obtained from this genus for the first time, compounds 5, 9 were obtained from this species for the first time. Antifungal activity of compounds 1-4, 6-9 were evaluated. Compound 1 showed the strong activity.

Topics: Antifungal Agents; Cryptococcus neoformans; Iridoids; Naphthoquinones; Plants, Medicinal; Pyrola; Triterpenes

Lu xian cao (Hebra pyrolae) is a traditional Chinese medicine used for the treatment of several major diseases. Chimaphilin has been demonstrated one of the major active components in Lu xian cao by modern pharmacological studies. In this study, a liquid chromatography-mass spectrometry method for the determination of chimaphilin in rat plasma was developed and validated. The separation was carried out on a C18 column using methanol and water as mobile phase after the plasma sample was extracted with diethyl ether. Atmospheric pressure chemical ionization in negative ion mode and selected ion monitoring method were developed to determine [M]- at 186 and 210 for chimaphilin and benzil (internal standard), respectively. The lower limit of quantitation was 10 ng/ml and the calibration curve was linear (r>0.9964) over the concentration range 10-1000 ng/ml. The method was demonstrated reproducible and reliable with intra-day precision<11.5%, inter-day precision<7.6%, accuracy in the range of 88.4-113.0%, and mean extraction recovery excess of 83.0%, which were all calculated from the quality control samples at concentrations of 20, 100, and 500 ng/ml. The method was successfully applied to pharmacokinetic study of chimaphilin in rat plasma following oral administration of a 30-mg/kg dose of chimaphilin in Lu xian cao decoction to male Wistar rats.

Topics: Animals; Male; Medicine, Chinese Traditional; Naphthoquinones; Rats; Rats, Wistar; Reference Standards; Reproducibility of Results; Sensitivity and Specificity

From electron paramagnetic resonance (EPR) spectra of numerous semiquinone anion radicals we propose that the spin density distributions in benzo-, naphtho- and anthrasemiquinones are changed under the influence of various substituent in a comparable fashion, electron donating substituents having the opposite effect of withdrawing ones. These findings facilitate the work in obtaining correct assignments, further leading to safe identification of new radicals. Semiquinone anion radicals of para- and ortho-hydroquinones are demonstrated to exist at physiological pH in buffered solutions under continuous access to molecular oxygen. Reaction schemes are proposed for the formation of primary and tertiary radicals and hydroxylated products of ten quinols. From studies on crude plant extracts are three quinols, known as precursors in the biogenesis of chimaphilin, observed and identified in Chimaphila umbellata. Other studies demonstrate the power of using semiquinone radicals as chemotaxonomic markers.

Topics: Chemistry; Electron Spin Resonance Spectroscopy; Electrons; Hydrogen-Ion Concentration; Hydroquinones; Models, Chemical; Naphthoquinones; Oxygen; Plant Extracts; Quinones; Time Factors

A new naphthoquinone, 5,8-dihydro-2,7-dimethyl-[1,4]naphthoquinone (1), which was named 5,8-dihydrochimaphilin, isolated from an ethyl acetate soluble fraction from the root of Pyrola japonica, together with chimaphilin (2). Compound 1 was transformed rapidly to 2 upon exposure to air by HPLC analysis. This fact supported that chimaphilin (2) may be an artifact from 1.

Topics: Antineoplastic Agents, Phytogenic; Naphthoquinones; Plants, Medicinal

Chimaphilin is a yellow naphthoquinone which occurs naturally in various chimaphila and Pyrola species. In Chimaphila umbellata (winter green) and C. maculata, it is a major constituent. Folk medicine recommends the leaves of Chimaphila species as a topical application to treat skin diseases. Since 1887, winter green is claimed to have caused dermatitis and to have been responsible for "idiosyncrasy". Experimental sensitization using the open epicutaneous as well as Freund's complete adjuvant technique has now revealed that chimaphilin is a moderate contact sensitizer.

Topics: Animals; Dermatitis, Contact; Female; Freund's Adjuvant; Guinea Pigs; Immunization; Irritants; Naphthoquinones; Skin Tests

Topics: Asteraceae; Glycosides; Hydroquinones; Naphthoquinones; Phenols; Plants, Medicinal; Pyrolaceae