UPLC-quadrupole time-of-flight-tandem mass spectrometry combined with chemometrics and network pharmacology to differentiate Coreopsis tinctoria Nutt.

Coreopsis tinctoria Nutt. (C. tinctoria) is a traditional medicinal plant, primarily found in plateau areas with altitudes exceeding 3000 m. The efficacy of C. tinctoria appears to be intricately tied to its quality. However, there is a scarcity of studies focused on evaluating the quality of C. tinctoria from diverse geographical locations. In this study, we used ultra-performance liquid chromatography-quadrupole time-of-flight-tandem mass spectrometry to analyze and identify the prevalent chemical components in 12 batches of C. tinctoria sourced from Xinjiang, Qinghai, Tibet, and Yunnan provinces in China. By using cluster analysis and discriminant analysis of partial least squares, we assessed the similarity and identified varying components in the 12 batches of C. tinctoria. Subsequently, their quality was further evaluated. Utilizing network pharmacology, we identified potential active components for the treatment of diabetes mellitus. The findings revealed the presence of 16 flavonoids, 3 phenylpropanes, 2 sugars, 2 amino acids, and 7 hydrocarbons in the analyzed samples. Through variable importance screening, 17 constituents were identified as quality difference markers. Marein and flavanomarein emerged as pivotal markers, crucial for distinguishing variations in C. tinctoria. In addition, network pharmacology predicted 187 targets for 9 common active components, including marein and flavanomarein. Simultaneously, 1747 targets related to diabetes mellitus were identified. The drug-component-disease target network comprised 91 nodes and 179 edges, encompassing 1 drug node, 9 component nodes, and 81 target nodes. In summary, marein and flavanomarein stand out as key biomarkers for assessing the quality of C. tinctoria, offering a scientific foundation for the quality evaluation of C. tinctoria Nutt.

Protective effects of flavonoids in Coreopsis tinctoria Nutt. in a mouse model of type 2 diabetes.

ETHNOPHARMACOLOGICAL RELEVANCE: Coreopsis tinctoria Nutt., a popular tea drink used in the Xinjiang region of China, has been traditionally used to treat diabetes and chronic metabolic diseases in China, Portugal, and North America. The bioactive extraction and potential mechanism have not been fully elucidated until now. AIM OF THE STUDY: Traditional herbal medicines usually share network targets due to multicomponent therapeutics. Therefore, we tried to explore the protective effects of C. tinctoria on diabetes and the related molecular mechanism. MATERIALS AND METHODS: A flavonoid-rich fraction of C. tinctoria (CTF) was prepared. After 15 weeks of continuous treatment with CTF, the blood glucose and blood lipid levels of experimental mice were evaluated. Tissue was collected for differentially expressed genes (DEGs), bioinformatics analysis, RT‒PCR and Western blot for target-related DEGs. RESULTS: After 15 weeks of continuous treatment with CTF, db/db mice showed reversed levels of glucose, insulin, glucagon and glycated hemoglobin A1c. CTF treatment also regulated total cholesterol, triglycerides, low density lipoprotein, nonesterified fatty acid, alanine transaminase, and aspartate transaminase. Major metabolic pathways were found to be dysregulated in the liver using a combined analysis of transcriptomics and network pharmacology. CTF treatment regulated 48.2% of 6357 dysregulated genes in db/db mice. The mitochondrial electron transport chain and tricarboxylic acid cycle were mainly affected. The sequencing data showed that fifty-nine predicted target genes for CTF were reverse regulated. Together with 1528 coexpressed genes, these genes reflected the main characteristics of the whole perturbed transcriptomic profile, i.e., dysregulated mitochondrial metabolism. The important genes of the target and coexpressed genes were further verified at the gene and protein levels. CONCLUSIONS: The results confirm that the metabolic changes induced by hyperglycemia are closely related to mitochondrial metabolism in the liver. CTF alters a core gene set that exerts regulatory effects at the biological pathway level in db/db mice. In conclusion, our data reveal that an important molecular event for CTF treatment is the regulation of mitochondrial metabolism and support the idea that herbs or natural compounds are potential therapeutic substances for mitochondrial dysfunction-related diabetes.

Aromatase inhibitors isolated from a flowering tea, snow Chrysanthemum (the capitula of Coreopsis tinctoria Nutt.).

Methanol extract from the capitula of Coreopsis tinctoria Nutt. (Asteraceae), which is also known as a flowering tea or blooming tea "Snow Chrysanthemum," was found to inhibit the enzymatic activity of aromatase. A total of 24 known isolates (1-24) were identified from the extract, including three chalcones (1-3), an aurone (4), five flavanones (5-9), four flavanols (10-13), a flavonol (14), and two biflavanones (15, 16). Among them, okanin (1, Ki = 1.6 µM), (2S)-naringenin (5, 0.90 µM), isookanin (6, 0.81 µM), (2S)-7,3',5'-trihydroxyflavaone (7, 0.13 µM), and (2S)-5,7,3',5'-tetrahydroxyflavanone (8, 0.32 µM) exhibited relatively potent competitive inhibition. Specifically, the isolates 7 and 8, having a common 3',5'-resorcinol moiety at the B ring in their flavanone skeleton, exhibited potent inhibitory activities compared to those of a clinically applied aminoglutethimide (0.84 µM) and naturally occurring flavone, chrysin (0.23 µM), which is a common non-steroidal aromatase inhibitor. Importantly, the active flavonoid constituents (1 and 5-8) did not inhibit the activity of 5α-reductase enzyme, which normally reacts with the same substrate "testosterone," thus, these compounds were suggested to be specific to aromatase.

Lanceolanone A, a new biflavanone, and a chalcone glucoside from the flower heads of Coreopsis lanceolata and their aldose reductase inhibitory activity and AMPK activation.

The MeOH extract of the flower heads of Coreopsis lanceolata L. (Asteraceae) exhibited aldose reductase (AR) inhibitory activity (IC50 8.36 µg/mL). Bioassay-guided fractionation of the extract resulted in the isolation of a new biflavanone-named Lanceolanone A (1) and a chalcone glucoside (6), along with 12 known compounds (2-5 and 7-14), of which 4, 7, 9, 10, and 12 were isolated from C. lanceolata for the first time. The structures of the new compounds (1 and 6) were determined by extensive spectroscopic analysis, including two-dimensional (2D) NMR, and ECD calculation method. Compounds 2, 4, 11, 13, and 14 exhibited AR inhibitory activities with IC50 values between 2.40 and 9.99 µM. Furthermore, 8-13 at 1.0 mM activated AMPK expression in HepG2 human hepatoma cells compared to the control.

Development of an LC/MS/MS Method for Simultaneous Detection of 11 Polyphenols in Rat Plasma and Its Pharmacokinetic Application after Oral Administration of Coreopsis tinctoria Extract.

Eleven polyphenols, classified as flavonoid glycosides, flavonoid aglycones, and phenolic acids, are important bioactive components in the capitula of Coreopsis tinctoria (CCT). Nevertheless, their full pharmacokinetic profiles have not been demonstrated simultaneously. Therefore, a liquid chromatography - tandem mass spectrometry (LC/MS/MS) method was developed in the present work and used it to study the pharmacokinetics of these 11 compounds. We performed LC/MS/MS with a gradient mobile phase composed of water containing 0.1 % formic acid and acetonitrile containing 0.1 % formic acid on a Proshell 120 SB C18 column (2.1 mm×100 mm, 2.7 µm). We achieved a good chromatographic peak shape, resolution, and mass signal response, and multiple reaction monitoring facilitated the simultaneous detection of 11 analytes. In addition, we validated the selectivity, correlation coefficient, precision, extraction recovery, matrix effects, and stability of the LC/MS/MS method to be acceptable for 11 analytes in rat plasma. Subsequently, rats were orally administered with 50 % ethanol eluent of CCT (ECCT). Nine of 11 polyphenols were absorbed quickly (except for QCD and TCA), and their plasma levels peaked within 40 min. The exposure and Cmax values of flavonoid glycosides and phenolic acids were lower than those of flavonoid aglycones. This is the first report to demonstrate the pharmacokinetics of 11 polyphenols in ECCT, which may play an important role in future studies of the bioactive components of ECCT and their bioactive mechanisms.

Okanin from Coreopsis tinctoria Nutt. alleviates cognitive impairment in bilateral common carotid artery occlusion mice by regulating the miR-7/NLRP3 axis in microglia.

Cognitive impairment is the main clinical feature following stroke, and microglia-mediated inflammatory response is a major contributor to it. Coreopsis tinctoria Nutt., an edible chrysanthemum, is commonly used as a functional ingredient in healthcare beverages and food. Okanin, the main active ingredient of Coreopsis tinctoria Nutt. flower, inhibits microglial activation. However, the role of okanin in cognitive impairment following ischemic stroke is still unknown. In this study, we investigated the effect of okanin on ischemic stroke and its underlying mechanism both in vivo and in vitro. Okanin was found to attenuate cognitive impairment in bilateral common carotid artery occlusion (BCCAO) mice, inhibit neuronal loss and microglial activation, decrease NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation, and increase miR-7 expression. Okanin suppressed NLRP3 inflammasome activation in oxygen-glucose deprivation (OGD) and lipopolysaccharide (LPS)-stimulated microglia by increasing miR-7 expression and inhibited microglia-induced neuronal injury. This study provides new insights into the role of okanin in ischemic stroke and shows that the miR-7/NLRP3 axis plays an important role in mediating the beneficial effects of okanin on cerebral ischemia. These findings suggest that okanin has great potential as a functional food for stroke recovery.

Impact of Coreopsis tinctoria Nutt. Essential oil microcapsules on the formation of biogenic amines and quality of smoked horsemeat sausage during ripening.

The present study evaluated the effect of Coreopsis tinctoria Nutt. essential oil (CEO) and its microcapsules (CEOM) on the accumulation of biogenic amines (BAs) and the quality of smoked horsemeat sausage during fermentation. The results showed that CEO could effectively inhibit Enterobacteriaceae growth and the formation of BAs (cadaverine, putrescine, tyrosine, histamine and tryptamine) in smoked horsemeat sausages, and the inhibition of CEO was enhanced after embedding (P < 0.05). Compared with other groups, thiobarbituric acid reactive substances, total volatile basic nitrogen and pH were lower in the microcapsule group. Furthermore, the sensory evaluation indicated that the addition of CEOM was a more effective way to maintain color and delay the deterioration of the sensory quality of sausages. Therefore, it is suggested that the CEOM can be used as a natural preservative in traditional fermented meat products to inhibit BAs accumulation and improve quality.

Metabolomics study of flavonoids in Coreopsis tinctoria of different origins by UPLC-MS/MS.

To analyze the flavonoids in Coreopsis tinctoria and compare the differences in flavonoids among C. tinctoria of different origins, the chemical composition of C. tinctoria capitulum was analyzed by ultra-high-performance liquid chromatography-tandem quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS), and the flavonoid metabolites were analyzed and identified based on their retention time, mass-to-charge ratio and fragment ions in the UPLC-QTOF-MS matrix. Capitulum samples of C. tinctoria were collected from three locations in the Xinjiang region at different altitudes. A total of 204 flavonoid compounds were identified, and 31 different flavonoid metabolites were then identified from flowers of C. tinctoria of different origins. Further analysis of these 31 significantly accumulated metabolites identified seven flavonoid metabolites, namely, homoplantaginin, kaempferol, quercetin, isorhamnetin, avicularin, quercetin 3-O-(6'-galloyl)-ß-D-galactopyranoside and isorhamnetin 3-O-glucoside, with high accumulation only in sample collected from Tashkurgan Tajik (TX) and low expression in sample collected from Yutian County (YT) and Shaya County (SY). Moreover, 7,4'-dihydroxyflavone and 4,4'-dimethoxychalcone showed high accumulation only in SY, and afzelin was specifically highly accumulated in YT. In addition, the identified flavonoid metabolites were annotated using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, and key pathways that might regulate the biosynthesis of these flavonoid compounds were analyzed. These findings provide key information for research on flavonoids and their biosynthesis in C. tinctoria and will provide a theoretical basis for studying the herbal quality and origin of C. tinctoria.

Antioxidant properties of polyphenols from snow chrysanthemum (Coreopsis tinctoria) and the modulation on intestinal microflora in vitro.

CONTEXT: Coreopsis tinctoria Nutt (Asteraceae), named snow chrysanthemum, is known to have a high level of polyphenols. However, the potential prebiotic effect on modulating intestinal microflora is still unclear. OBJECTIVE: The chemical composition, antioxidant properties of snow chrysanthemum polyphenols (SCPs) and their effects on human intestinal microbiota were investigated. MATERIALS AND METHODS: SCPs were extracted using ultrasonic-assisted extraction, and further determined using UPLC-QE Orbitrap/MS. Five assays were used to investigate the antioxidant activities of SCPs. Subsequently, the effects of SCPs on intestinal microbiota in vitro were determined by high throughput sequencing and bioinformatics analysis. RESULTS: Marein, isookanin and cymaroside were the major phenolic compounds, which accounted for 42.17%, 19.53% and 12.25%, respectively. Marein exhibited higher scavenging capacities in DPPH (EC50 = 8.84 µg/mL) and super anion radical assay (EC50 = 282.1 µg/mL) compared to cymaroside and isookanin. The antioxidant capacity of cymaroside was weakest among the three phenolic compounds due to the highest EC50 values, especially for superoxide anion radical assay, EC50 > 800 µg/mL. The result of in vitro fermentation showed that the three phenolic compounds increased the relative abundances of Escherichia/Shigella, Enterococcus, Klebsiella, etc., and isookanin notably increased the relative abundance of Bifidobacterium and Lactobacillus. DISCUSSION AND CONCLUSIONS: SCPs exhibited antioxidant properties and potential prebiotic effects on modulating the gut microbiota composition. The findings indicated that SCPs consumption could exert prebiotic activity that is beneficial for human health.

Study on the metabolism profile of flavanomarein in Coreopsis tinctoria Nutt.

Coreopsis tinctoria Nutt. (family Asteraceae) is a popular medicine-food plant, which improves chronic diseases such as hyperlipemia, hypertension, and diabetes. Flavanomarein is the main active component of Coreopsis tinctoria Nutt, in which the blood concentration of volunteers is low and bioavailability is poor. Thus, the understanding of flavanomarein metabolites and metabolic pathways is significant to clarify its effectiveness. This study systematically studied the metabolites of flavanomarein by oral and injection. The biological samples (feces, urine, and plasma) were analyzed by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry in negative ion mode. The metabolic law of flavanomarein in the liver was further verified by a liver microsomal incubation experiment in vitro. A total of 12 metabolites were identified by oral administration while 15 metabolites were detected by injection. It was shown that metabolic pathways include acetylation, hydroxylation, glucuronidation, methylation, dehydrogenation, and so forth. The liver extraction rate of flavanomarein was 0.08, which means the metabolic stability of flavanomarein is well in rats' liver microsomes. It is a systematic study on the metabolism of flavanomarein and provides a metabolic rationale for further in-depth in vivo biotransformation.

Effect of Different Roasting Conditions and Coreopsis Extract on Heterocyclic Amine Formation in Roast Lamb Products.

ABSTRACT: Heterocyclic amines (HCAs), which are known carcinogens in thermally processed foods, were investigated in roast lamb patties under various time and temperature conditions. HCAs in lamb products roasted at some temperatures increased with roasting time. An exponential model with a time factor fit well for the production of HCAs. The mean pH and cooking loss at various temperatures were also determined. The mean pH decreased as the temperature increased. Coreopsis extract was added to lamb patties roasted at 230°C for 15 min per side. The amount of coreopsis extract added had a significant effect on HCA development. A weak positive relationship was observed between the antioxidant activity of the lamb patty with the coreopsis extract and the inhibitory effect of coreopsis extract on various HCAs, with a correlation coefficient of 0.14 to 0.44 (P > 0.05). Coreopsis extract containing flavonoids can be a beneficial additive for production of barbecue meat.

Physiological and rhizospheric response characteristics to cadmium of a newly identified cadmium accumulator Coreopsis grandiflora Hogg. (Asteraceae).

Screening for superior cadmium (Cd) phytoremediation resources and uncovering the mechanisms of plant response to Cd are important for effective phytoremediation of Cd-polluted soils. In this study, the characteristics of Coreopsis grandiflora related to Cd tolerance and accumulation were analyzed to evaluate its Cd phytoremediation potential. The results revealed that C. grandiflora can tolerate up to 20 mg kg-1 of Cd in the soil. This species showed relatively high shoot bioconcentration factors (1.09-1.85) and translocation factors (0.46-0.97) when grown in soils spiked with 5-45 mg kg-1 Cd, suggesting that C. grandiflora is a Cd accumulator and can potentially be used for Cd phytoextraction. Physiological analysis indicated that antioxidant enzymes (i.e., superoxide dismutase, peroxidase, and catalase) and various free amino acids (e.g., proline, histidine, and methionine) participate in Cd detoxification in C. grandiflora grown in soil spiked with 20 mg kg-1 of Cd (Cd20). The overall microbial richness and diversity remained similar between the control (Cd0) and Cd20 soils. However, the abundance of multiple rhizospheric microbial taxa was altered in the Cd20 soil compared with that in the Cd0 soil. Interestingly, many plant growth-promoting microorganisms (e.g., Nocardioides, Flavisolibacter, Rhizobium, Achromobacter, and Penicillium) enriched in the Cd20 soil likely contributed to the growth and vitality of C. grandiflora under Cd stress. Among these, some microorganisms (e.g., Rhizobium, Achromobacter, and Penicillium) likely affected Cd uptake by C. grandiflora. These abundant plant growth-promoting microorganisms potentially interacted with soil pH and the concentrations of Cd and AK in soil. Notably, potassium-solubilizing microbes (e.g., Rhizobium and Penicillium) may effectively solubilize potassium to assist Cd uptake by C. grandiflora. This study provides a new plant resource for Cd phytoextraction and improves our understanding of rhizosphere-associated mechanisms of plant adaptation to Cd-contaminated soil.

Coreopsis tinctoria and Its Flavonoids Ameliorate Hyperglycemia in Obese Mice Induced by High-Fat Diet.

With the prevalence of obesity all over the world, human health has been seriously affected. In particular, the number of diabetic and cardiovascular diseases has increased dramatically. The herb Coreopsis tinctoria (C. tinctoria) shows diverse biological and pharmacological activities, which are mainly attributed to its flavonoids. However, the specific functional substances that play an active role in C. tinctoria remain unclear, and its mechanism has not been deeply explored. In this study, we established a diet-induced obesity (DIO) mice model and treated mice with C. tinctoria or kaempferol for 8 weeks. The results showed that both C. tinctoria and kaempferol lowered body weight, reduced fasting blood glucose, and improved glucose tolerance and insulin resistance to alleviate obesity in DIO mice. The level of hemoglobin A1c also decreased significantly after treatment with C. tinctoria and kaempferol. Moreover, the administration of C. tinctoria and kaempferol also restored gut microbiota imbalance and significantly increased Desulfovibrio and Butyricimonas levels, which have been reported to improve glucose metabolism and intestinal health. In general, our study shows that C. tinctoria is a potential hypoglycemic substance for obesity and may reduce blood glucose by regulating gut microbiota, and that kaempferol is one of the effective substances of C. tinctoria.

Simultaneous Determination and Comparison of Phenolic Bioactives among Three Main Kinds of Edible Chrysanthemums.

In this study, we report a simple and reliable high-performance liquid chromatography coupled with diode array detection method for simultaneous and quantitative analysis and comparison of major phenolic compounds dominant phytochemicals in Chrysanthemum morifolium, Florists chrysanthemum and snow chrysanthemum (Coreopsis tinctoria or C. tinctoria). The chromatographic separation was achieved using a reversed phase C18 column with a mobile phase of water [containing 0.1% trifluoroacetic acid (TFA)] and acetonitrile. The major phenolic compounds were completely separated within 16 min at a flow rate of 1.0 mL/min. Flavonoid and phenolic acid profiles of the ethanol extracts of the three flowers were analyzed. The results revealed that C. tinctoria possessed the highest amount of flavonoids (flavanomarein, flavanokanin, marein and okanin) and relative lower content of phenolic acid (chlorogenic acid and 3,5-dicafeoylquinic acid). The total content of the four flavonoids in C. tinctoria reached 53.99 ± 1.32 mg/g. In particular, the marein content in C. tinctoria was as high as 36.50 mg/g. Flavanomarein was only detected in C. tinctoria, whereas chlorogenic acid and 3,5-dicafeoylquinic acid were abundant in Chrysanthemum morifolium and Florists chrysanthemum. The content of marein in Chrysanthemum morifolium was slightly higher than that in Florists chrysanthemum, whereas no okanin was detected in Florists chrysanthemum under these high-performance liquid chromatography conditions. The results indicated phenolic components differ significantly depending on the cultivar, especially between C. tinctoria and common commercially available chrysanthemums. The method adopted in this study is helpful for quality control of different chrysanthemum species as well as their products, which is essential for usage and functionality clarification.

Rapid prediction method of α-Glycosidase inhibitory activity of Coreopsis tinctoria extract from different habitats by near infrared spectroscopy.

α-Glucosidase is one of the main enzymes causing elevated blood glucose, and Coreopsis tinctoria extract can be used as a natural inhibitor of α-Glucosidase. Therefore, a new method was proposed for predicting the inhibitory activity on α-Glucosidase of Coreopsis tinctoria extract based on near infrared spectroscopy. The absorbance of the inhibitory system was measured by ultraviolet spectroscopy, which was used to study the inhibitory activity on a-glucosidase of Coreopsis tinctoria extract. The near infrared spectra of the solid samples were collected. By selecting spectral preprocessing and optimizing spectral bands, a rapid prediction model of the inhibitory activity was established by partial least squares regression. The root mean square error of cross-validation (RMSECV), correlation coefficient (R) value and the ratio of prediction to deviation (RPD) value were used as indicators of the evaluation model. The near infrared spectrum model was established by combining the best spectral preprocessing of the continuous wavelet transform (CWT) and the best spectral band. The root mean square error of cross-validation (RMSECV) of this model was 0.815%, the correlation coefficient (R) value was 0.942, and the ratio of prediction to deviation (RPD) was 3.0. The root mean square error of prediction (RMSEP) of the model by prediction set was 0.819%, the correlation coefficient (R) value was 0.950, and the RPD was 3.2. The model shows that the fitting relationship between the predicted inhibition value and the reference inhibition value of the near infrared spectral model is good. The results showed that there was a good correlation between near infrared spectroscopy and the inhibitory activity of Coreopsis tinctoria extract. Thus, the established model was robust and effective and could be used for rapid quantification of α-Glucosidase inhibitory activity. The prediction method is simple and rapid, and can be extended to study the inhibition of other medicinal plants.

Mechanisms of Coreopsis tinctoria Nutt in the treatment of diabetic nephropathy based on network pharmacyology analysis of its active ingredients / Mecanismos de Coreopsis tinctoria Nutt. En el tratamiento de la nefropatía diabética basado en el análisis farmacológico de red de sus principios activos

SUMMARY: Coreopsis tinctoria Nutt. (C. tinctoria Nutt.) can protect diabetic kidneys, but the mechanisms are unclear. This work is to investigate the potential mechanisms of C. tinctoria Nutt. in the treatment of diabetic nephropathy based on network pharmacology analysis of its active ingredients. Twelve small molecular compounds of C. tinctoria Nutt. and targets related to diabetic nephropathy were docked by Discovery Studio 3.0. DAVID database was used for GO enrichment and KEGG pathway analysis. Cytoscape 3.6.1 was used to construct active ingredient-target network. Cell viability was detected with MTT. Glucose consumption was analyzed with glucose oxidase method. Protein expression was measured with Western blot and immunofluorescence. Electron microscopy observed autophagosomes. The core active ingredients of C. tinctoria Nutt. included heriguard, flavanomarein, maritimein, and marein. Twenty-one core targets of the 43 potential targets were PYGM, TLR2, RAF1, PRKAA2, GPR119, INS, CSF2, TNF, IAPP, AKR1B1, GSK3B, SYK, NFKB2, ESR2, CDK2, FGFR1, HTRA1, AMY2A, CAMK4, GCK, and ABL2. These 21 core targets were significantly enriched in 50 signaling pathways. Thirty- four signaling pathways were closely related to diabetic nephropathy, of which the top pathways were PI3K/AKT, insulin, and mTOR, and insulin resistance. The enriched GO terms included biological processes of protein phosphorylation, and the positive regulation of PI3K signaling and cytokine secretion; cellular components of cytosol, extracellular region, and extracellular space; and molecular function of protein kinase activity, ATP binding, and non-membrane spanning protein tyrosine kinase activity. In vitro experiments found that marein increased the expression of phosphorylated AKT/AKT in human renal glomerular endothelial cells of an insulin resistance model induced by high glucose, as well as increased and decreased, respectively, the levels of the microtubule-associated proteins, LC3 and P62. C. tinctoria Nutt. has many active ingredients, with main ingredients of heriguard, flavanomarein, maritimein, and marein, and may exert anti-diabetic nephropathy effect through various signaling pathways and targets.

RESUMEN: Coreopsis tinctoria Nutt. (C. tinctoria Nutt.) puede proteger riñones diabéticos, sin embargo los mecanismos son desconocidos. Este trabajo se realizó para investigar los potenciales mecanismos de C. tinctoria Nutt. en el tratamiento de la nefropatía diabética basado en el análisis de farmacología en red de sus principios activos. Doce compuestos moleculares pequeños de C. tinctoria Nutt. y los objetivos relacionados con la nefropatía diabética fueron acoplados por Discovery Studio 3.0. La base de datos DAVID se utilizó para el enriquecimiento GO y el análisis de la vía KEGG. Se usó Cytoscape 3.6.1 para construir una red de ingrediente-objetivo activa. La viabili- dad celular se detectó mediante MTT. El consumo de glucosa se analizó con el método de glucosa oxidasa. La expresión proteica fue determinada mediante Western blot e inmunofluorescencia. En la microscopía electrónica se observó autofagosomas. Los principales ingredientes activos de C. tinctoria Nutt. incluyeron heriguard, flavanomarein, maritimin y marein. Veintiún de los 43 objetivos potenciales fueron PYGM, TLR2, RAF1, PRKAA2, GPR119, INS, CSF2, TNF, IAPP, AKR1B1, GSK3B, SYK, NFKB2, ESR2, CDK2, FGFR1, HTRA1, AMY2A, CAMK4, GCK y ABL2. Estos 21 objetivos principales se enriquecieron significativamente en 50 vías de señalización. Treinta y cuatro vías de señalización estuvieron estrechamente relacionadas con la nefropatía diabética, de las cuales las principales vías fueron PI3K/ AKT, insulina y mTOR, y resistencia a la insulina. Los términos GO enriquecidos incluyeron procesos biológicos de fosforilación proteica, la regulación positiva de la señalización de PI3K y la secreción de citoquinas; componentes celulares del citosol, región extracelular y espacio extracelular; y la función molecular de la actividad de la proteína quinasa, la unión de ATP y la actividad de la proteína tirosina quinasa que no se extiende por la membrana. Los experimentos in vitro encontraron que la mareína aumentaba la expresión de AKT/AKT fosforilada en células endoteliales glomerulares renales humanas en un modelo de resistencia a la insulina inducida por niveles elevados de glucosa, así como aumentaron y disminuyeron respectivamente, los niveles de las proteínas asociadas a los microtúbulos, LC3 y P62. C. tinctoria Nutt. tiene muchos principios activos, con ingredientes principales de heriguard, flavanomarein, maritimain y marein, y puede ejercer un efecto de nefropatía antidiabética a través de distintass vías de señalización y objetivos.

Aurones and Flavonols from Coreopsis lanceolata L. Flowers and Their Anti-Oxidant, Pro-Inflammatory Inhibition Effects, and Recovery Effects on Alloxan-Induced Pancreatic Islets in Zebrafish.

(1) Background: Many flavonoids have been reported to exhibit pharmacological activity; a preparatory study confirmed that Coreopsis lanceolata flowers (CLFs) contained high flavonoid structure content; (2) Methods: CLFs were extracted in aqueous methanol (MeOH:H2O = 4:1) and fractionated into acetic ester (EtOAc), normal butanol (n-BuOH), and H2O fractions. Repeated column chromatographies for two fractions led to the isolation of two aurones and two flavonols; (3) Results: Four flavonoids were identified based on a variety of spectroscopic data analyses to be leptosidin (1), leptosin (2), isoquercetin (3), and astragalin (4), respectively. This is the first report for isolation of 2-4 from CLFs. High-performance liquid chromatography (HPLC) analysis determined the content levels of compounds 1-4 in the MeOH extract to be 2.8 ± 0.3 mg/g (1), 17.9 ± 0.9 mg/g (2), 3.0 ± 0.2 mg/g (3), and 10.9 ± 0.9 mg/g (4), respectively. All isolated compounds showed radical scavenging activities and recovery activities in Caco-2, RAW264.7, PC-12, and HepG2 cells against reactive oxygen species. MeOH extract, EtOAc fraction, and 1-3 suppressed NO formation in LPS-stimulated RAW 264.7 cells and decreased iNOS and COX-2 expression. Furthermore, all compounds recovered the pancreatic islets damaged by alloxan treatment in zebrafish; (4) Conclusions: The outcome proposes 1-4 to serve as components of CLFs in standardizing anti-oxidant, pro-inflammatory inhibition, and potential anti-diabetic agents.

Anti-Neuroinflammatory and Anti-Inflammatory Activities of Phenylheptatriyne Isolated from the Flowers of Coreopsis lanceolata L. via NF-κB Inhibition and HO-1 Expression in BV2 and RAW264.7 Cells.

Aging is associated with immune disregulation and oxidative stress which lead to inflammation and neurodegenerative diseases. We have tried to identify the anti-neuroinflammatory and anti-inflammatory components of Coreopsis lanceolata L. The dried flowers of C. lanceolata were extracted with 70% EtOH, and the obtained extract was divided into CH2Cl2, EtOAc, n-BuOH, and H2O fractions. The CH2Cl2 fraction was separated using silica gel and C-18 column chromatography to yield phenylheptatriyne (1), 2'-hydroxy-3,4,4'-trimethoxychalcone (2), and 4',7-dimethoxyflavanone (3). Additionally, the EtOAc fraction was subjected to silica gel, C-18, and Sephadex LH-20 column chromatography to yield 8-methoxybutin (4) and leptosidin (5). All the compounds isolated from C. lanceolata inhibited the production of nitric oxide (NO) in LPS-induced BV2 and RAW264.7 cells. In addition, phenylheptatriyne and 4',7-dimethoxyflavanone reduced the secretion of inflammatory cytokines, tumor necrosis factor alpha (TNF-α), and interleukin (IL)-6. Among them, phenylheptatriyne was significantly downregulated in the expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). Subsequently, phenylheptatriyne also effectively inhibited nuclear factor-kappa B (NF-κB) activation in LPS-stimulated BV2 and RAW264.7 cells. Based on these results, the anti-neuroinflammatory effect of phenylheptatriyne isolated from C. lanceolata was confirmed, which may exert a therapeutic effect in treatment of neuroinflammation-related diseases.

Nitrogen deficiency maintains the yield and improves the antioxidant activity of Coreopsis tinctoria Nutt.

Nitrogen (N) deficiency levels were investigated for their potential to maintain the yield and improve antioxidant activity of Coreopsis tinctoria. Inflorescences and leaves at 0, 10, 20, 30, 40, and 50 d after flowering were frozen at -80 °C and plant growth, antioxidant activity, bioactive substance, enzyme activity, and gene expression were evaluated. N deficiency maintained the total number of flowers, promoted phenol and flavonoid accumulation, and enhanced antioxidant activity. Moreover, N deficiency stimulated activities of phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H) and 4-coumarate:coenzyme A ligase (4CL), and induced CtPAL, CtC4H and Ct4CL gene expression. The data also suggest that N-deficiency-induced phenolic and flavonoid accumulation occurs due to the activation of biosynthetic pathways in C. tinctoria. We characterize the unique features of C. tinctoria under N-deficiency conditions and provide valuable information for the cultivation of high-N use efficiency varieties with low input and high output.

[Study on rat intestinal absorption characteristics of total flavonoids from Coreopsis tinctoria].

The rat everted intestinal sac model was adopted to investigate the absorption of total flavonoids from Coreopsis tinctoria in different intestinal segments. Cyaniding-3-O-ß-D-glucoside, chlorogenic acid, flavanomarein, quercetagetin-7-O-ß-D-glucoside, iso-okanin, marein and 3,5-dicaffeoylquinic acid which as the major chemical components of total flavonoids from C. tinctoria were selec-ted as the study objects to evaluate the absorption characteristics of each component in different intestinal segments. The results showed that the absorption of seven components of total flavonoids at different intestinal segments was in consistent with zero order absorption rate. The K_a of chlorogenic acid, flavanomarein, quercetagetin-7-O-ß-D-glucoside, isookanin and 3,5-dicaffeoylquinic acid increased with increasing of concentration of total flavonoids(P<0.05), indicating that the intestinal absorption of these five components was passive transport. The K_a of cyaniding-3-O-ß-D-glucoside and marein showed a weak concentration dependence, suggesting that the absorption of them may be an positive and passive co-existing mode. The result of absorption in different intestinal segments showed that cyaniding-3-O-ß-D-glucoside, chlorogenic acid, flavanomarein, quercetagetin-7-O-ß-D-glucoside, marein and 3,5-dicaffeoylquinic acid were mainly absorbed in ileum, while isookanin was mainly absorbed in jejunum. The total flavonoids of C. tinctoria are selectively absorbed in intestinal tract, the rat everted intestinal sac model can be used to evaluate the multi-component intestinal absorption characteristics of total flavonoids from C. tinctoria.