OBJECTIVES: This study presents a versatile, AI-guided workflow for the targeted isolation and characterization of prenylated flavonoids from Paulownia tomentosa (Thunb.) Steud. (Paulowniaceae). METHODS: The approach integrates established extraction and chromatography-based fractionation protocols with LC-UV-HRMS/MS analysis and supervised machine-learning (ML) custom-trained classification models, which predict prenylated flavonoids from LC-HRMS/MS spectra based on the recently developed Python package AnnoMe (v1.0). RESULTS: The workflow effectively reduced the chemical complexity of plant extracts and enabled efficient prioritization of fractions and compounds for targeted isolation. From the pre-fractionated plant extracts, 2687 features were detected, 42 were identified using reference standards, and 214 were annotated via spectra library matching (public and in-house). Furthermore, ML-trained classifiers predicted 1805 MS/MS spectra as derived from prenylated flavonoids. LC-UV-HRMS/MS data of the most abundant presumed prenyl-flavonoid candidates were manually inspected for coelution and annotated to provide dereplication. Based on this, one putative prenylated (C5) dihydroflavonol (1) and four geranylated (C10) flavanones (2-5) were selected and successfully isolated. Structural elucidation employed UV spectroscopy, HRMS, and 1D as well as 2D NMR spectroscopy. Compounds 1 and 5 were isolated from a natural source for the first time and were named 6-prenyl-4'-O-methyltaxifolin and 3',4'-O-dimethylpaulodiplacone A, respectively. CONCLUSIONS: This study highlights the combination of machine learning with analytical techniques to streamline natural product discovery via MS/MS and AI-guided pre-selection, efficient prioritization, and characterization of prenylated flavonoids, paving the way for a broader application in metabolomics and further exploration of prenylated constituents across diverse plant species.

• Klíčová slova
• bioactive compounds, geranylated flavonoids, prenylated polyphenols, specialized metabolites, untargeted metabolomics,
• Publikační typ
• časopisecké články MeSH

INTRODUCTION: Within oncology research, there is a high effort for new approaches to prevent and treat cancer as a life-threatening disease. Specific plant species that adapt to harsh conditions may possess unique properties that may be utilized in the management of cancer. HYPOTHESIS: Chokeberry fruit is rich in secondary metabolites with anti-cancer activities potentially useful in cancer prevention and treatment. AIMS OF THE STUDY AND METHODS: Based on mentioned hypothesis, the main goal of our study was to evaluate the antitumor effects of dietary administered Aronia melanocarpa L. fruit peels (in two concentrations of 0.3 and 3% [w/w]) in the therapeutic syngeneic 4T1 mouse adenocarcinoma model, the chemopreventive model of chemically induced mammary carcinogenesis in rats, a cell antioxidant assay, and robust in vitro analyses using MCF-7 and MDA-MB-231 cancer cells. RESULTS: The dominant metabolites in the A. melanocarpa fruit peel extract tested were phenolic derivatives classified as anthocyanins and procyanidins. In a therapeutic model, aronia significantly reduced the volume of 4T1 tumors at both higher and lower doses. In the same tumors, we noted a significant dose-dependent decrease in the mitotic activity index compared to the control. In the chemopreventive model, the expression of Bax was significantly increased by aronia at both doses. Additionally, aronia decreased Bcl-2 and VEGF levels, increasing the Bax/Bcl-2 ratio compared to the control group. The cytoplasmic expression of caspase-3 was significantly enhanced when aronia was administered at a higher dosage, in contrast to both the control group and the aronia group treated with a lower dosage. Furthermore, the higher dosage of aronia exhibited a significant reduction in the expression of the tumor stem cell marker CD133 compared to the control group. In addition, the examination of aronia`s epigenetic impact on tumor tissue through in vivo analyses revealed significant alterations in histone chemical modifications, specifically H3K4m3 and H3K9m3, miRNAs expression (miR155, miR210, and miR34a) and methylation status of tumor suppressor genes (PTEN and TIMP3). In vitro studies utilizing a methanolic extract of A.melanocarpa demonstrated significant anti-cancer properties in the MCF-7 and MDA-MB-231 cell lines. Various analyses, including Resazurin, cell cycle, annexin V/PI, caspase-3/7, Bcl-2, PARP, and mitochondrial membrane potential, were conducted in this regard. Additionally, the aronia extract enhanced the responsiveness to epirubicin in both cancer cell lines. CONCLUSION: This study is the first to analyze the antitumor effect of A. melanocarpa in selected models of experimental breast carcinoma in vivo and in vitro. The utilization of the antitumor effects of aronia in clinical practice is still minimal and requires precise and long-term clinical evaluations. Individualized cancer-type profiling and patient stratification are crucial for effectively implementing plant nutraceuticals within targeted anti-cancer strategies in clinical oncology.

• Klíčová slova
• Aronia melanocarpa L., breast carcinoma, epigenetics, in vitro models, mechanism of action, rodent models,
• Publikační typ
• časopisecké články MeSH

Within the group of higher fungi, edible medicinal mushrooms have a long history of being used as food and in folk medicine. These species contain biologically active substances with many potential beneficial effects on human health. The Pleurotus genus is representative of medicinal mushrooms because Pleurotus ostreatus is one of the most commonly cultivated culinary mushrooms. In our study, we focused on lesser-known species in the genus Pleurotus and measured their antioxidant and anti-inflammatory activity. We prepared extracts of the mushrooms and analyzed them using HPLC-HRMS, GC-MS, and 1H-NMR. Significant differences in biological activities were found among the Pleurotus spp. extracts. A MeOH extract of P. flabellatus was the most active as a radical scavenger with the highest ORAC, while a chloroform extract had significant anti-inflammatory COX-2 activity. The 80% MeOH extract of P. flabellatus contained the highest amounts of ergosterol, ergothioneine, and mannitol. The 80% MeOH extract of P. ostreatus Florida was the most active in the NF-κB inhibition assay and had the highest content of β-glucans (43.3% by dry weight). Given the antioxidant and anti-inflammatory properties of P. flabellatus, the potential therapeutic usefulness of this species is worth evaluating through in-depth investigations and confirmation by clinical trials.

• Klíčová slova
• Indian oyster mushroom, basidiomycetes, bioactivity, cyclooxygenase-2, immunomodulatory effect, inflammation, oyster mushroom, pink oyster mushroom, radical scavenging effect, secondary metabolites,
• Publikační typ
• časopisecké články MeSH

Background: Oxidative stress is a key factor in the pathophysiology of many diseases. This study aimed to verify the antioxidant activity of selected plant phenolics in cell-based assays and determine their direct or indirect effects. Methods: The cellular antioxidant assay (CAA) assay was employed for direct scavenging assays. In the indirect approach, the influence of each test substance on the gene and protein expression and activity of selected antioxidant enzymes was observed. One assay also dealt with activation of the Nrf2-ARE pathway. The overall effect of each compound was measured using a glucose oxidative stress protection assay. Results: Among the test compounds, acteoside showed the highest direct scavenging activity and no effect on the expression of antioxidant enzymes. It increased only the activity of catalase. Diplacone was less active in direct antioxidant assays but positively affected enzyme expression and catalase activity. Morusin showed no antioxidant activity in the CAA assay. Similarly, pomiferin had only mild antioxidant activity and proved rather cytotoxic. Conclusions: Of the four selected phenolics, only acteoside and diplacone demonstrated antioxidant effects in cell-based assays.

• Klíčová slova
• CAA, Nrf2-ARE, antioxidants, catalase, glucose toxicity, plant phenolics, superoxide dismutase,
• MeSH
• antioxidační responzivní elementy MeSH
• antioxidancia chemie   farmakologie   MeSH
• biologické markery MeSH
• exprese genu MeSH
• faktor 2 související s NF-E2 genetika   metabolismus   MeSH
• fenoly chemie   farmakologie   MeSH
• glukosa MeSH
• lidé MeSH
• molekulární struktura MeSH
• oxidační stres MeSH
• protinádorové látky chemie   farmakologie   MeSH
• rostlinné extrakty chemie   farmakologie   MeSH
• superoxiddismutasa 1 genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antioxidancia MeSH
• biologické markery MeSH
• faktor 2 související s NF-E2 MeSH
• fenoly MeSH
• glukosa MeSH
• NFE2L2 protein, human MeSH Prohlížeč
• protinádorové látky MeSH
• rostlinné extrakty MeSH
• SOD1 protein, human MeSH Prohlížeč
• superoxiddismutasa 1 MeSH

Multidrug resistance (MDR) is a common problem when fighting cancer with chemotherapy. P-glycoprotein (P-gp, or MDR1) is an active pump responsible for the efflux of xenobiotics out of the cell, including anti-cancer drugs. It is a validated target against MDR. No crystal structure of the human P-gp is available to date, and only recently several cryo-EM structures have been solved. In this paper, we present a comprehensive computational approach that includes constructing the full-length three-dimensional structure of the human P-gp and its refinement using molecular dynamics. We assessed its flexibility and conformational diversity, compiling a dynamical ensemble that was used to dock a set of lignan compounds, previously reported as active P-gp inhibitors, and disclose their binding modes. Based on the statistical analysis of the docking results, we selected a system for performing the structure-based virtual screening of new potential P-gp inhibitors. We tested the method on a library of 87 natural flavonoids described in the literature, and 10 of those were experimentally assayed. The results reproduced the theoretical predictions only partially due to various possible factors. However, at least two of the predicted natural flavonoids were demonstrated to be effective P-gp inhibitors. They were able to increase the accumulation of doxorubicin inside the human promyelocytic leukemia HL60/MDR cells overexpressing P-gp and potentiate the antiproliferative activity of this anti-cancer drug.

• Klíčová slova
• P-glycoprotein, flavonoids, molecular docking, molecular dynamics, multidrug resistance, natural compounds, structure-based virtual screening,
• Publikační typ
• časopisecké články MeSH

Morusin is a prenylated flavonoid isolated from the root bark of Morus alba. Many studies have shown the ability of flavonoids to act as anti-inflammatory agents. The aim of this study was to evaluate the effect of morusin on experimentally colitis induced by 2,4,6-trinitrobenzensulfonic acid in Wistar rats and to compare it with sulfasalazine, a drug conventionally used in the treatment of inflammatory bowel disease. Morusin was administered by gavage at doses of 12.5, 25, or 50 mg/kg/day for five days. The colonic tissue was evaluated macroscopically, histologically, and by performing immunodetection and zymographic analysis to determine the levels of antioxidant enzymes [superoxide dismutase (SOD) and catalase (CAT)], interleukin (IL)-1β, and transforming growth factor (TGF)-β1 and the activities of matrix metalloproteinases (MMP) 2 and 9. The tissue damage scores were significantly reduced with increasing dose of morusin, however efficacy was not demonstrated at the highest dose. At the dose of 12.5 mg/kg, morusin exerted therapeutic effectivity similar to that of sulfasalazine (50 mg/kg). This was associated with significant reduction of TGF-β1 levels and MMP2 and MMP9 activities, and slight reduction of IL-1β. Our results suggest that morusin possesses therapeutic potential for the treatment of chronic inflammatory diseases.

• MeSH
• flavonoidy farmakologie   MeSH
• kolitida chemicky indukované   enzymologie   prevence a kontrola   MeSH
• kolon účinky léků   enzymologie   patologie   MeSH
• kyselina trinitrobenzensulfonová MeSH
• matrixová metaloproteinasa 2 metabolismus   MeSH
• potkani Wistar MeSH
• prenylace MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• flavonoidy MeSH
• kyselina trinitrobenzensulfonová MeSH
• matrixová metaloproteinasa 2 MeSH
• Mmp2 protein, rat MeSH Prohlížeč
• morusin MeSH Prohlížeč

Aims. In this work we studied cytodifferentiation effects of newly characterized prenyl flavonoid 4'-O-methylkuwanon E (4ME) isolated from white mulberry (Morus alba L.). Main Methods. Cell growth and viability were measured by dye exclusion assay; cell cycle and surface antigen CD11b were monitored by flow cytometry. For the cytodifferentiation of cells the NBT reduction assay was employed. Regulatory proteins were assessed by western blotting. Key Findings. 4ME induced dose-dependent growth inhibition of THP-1 cells, which was not accompanied by toxic effect. Inhibition of cells proliferation caused by 4ME was associated with the accumulation in G1 phase and with downregulation of hyperphosphorylated pRb. Treatment with 4ME led to significant induction of NBT-reducing activity of PMA stimulated THP-1 cells and upregulation expression of differentiation-associated surface antigen CD11b. Our results suggest that monocytic differentiation induced by 4ME is connected with up-regulation of p38 kinase activity. Significance. Our study provides the first evidence that 4ME induces the differentiation of THP-1 human monocytic leukemia cells and thus is a potential cytodifferentiating anticancer agent.

• Publikační typ
• časopisecké články MeSH

Morus alba L. (MA) is a natural source of many compounds with different biological effects. It has been described to possess anti-inflammatory, antioxidant, and hepatoprotective activities. The aim of this study was to evaluate cytotoxicity of three flavonoids isolated from MA (kuwanon E, cudraflavone B, and 4'-O-methylkuwanon E) and to determine their effects on proliferation of THP-1 cells, and on cell cycle progression of cancer cells. Anti-inflammatory effects were also determined for all three given flavonoids. Methods used in the study included quantification of cells by hemocytometer and WST-1 assays, flow cytometry, western blotting, ELISA, and zymography. From the three compounds tested, cudraflavone B showed the strongest effects on cell cycle progression and viability of tumor and/or immortalized cells and also on inflammatory response of macrophage-like cells. Kuwanon E and 4'-O-methylkuwanon E exerted more sophisticated rather than direct toxic effect on used cell types. Our data indicate that mechanisms different from stress-related or apoptotic signaling pathways are involved in the action of these compounds. Although further studies are required to precisely define the mechanisms of MA flavonoid action in human cancer and macrophage-like cells, here we demonstrate their effects combining antiproliferative and anti-inflammatory activities, respectively.

• Publikační typ
• časopisecké články MeSH