Associations Between Flavonoid Intake and Subclinical Atherosclerosis: The MESA.

BACKGROUND: Flavonoids may play a role in mitigating atherosclerotic cardiovascular diseases, with evidence suggesting effects may differ between vascular beds. Studies examining associations with subclinical markers of atherosclerosis between subpopulations with different underlying risks of atherosclerosis are lacking. METHODS: Among 5599 participants from the MESA (Multi-Ethnic Study of Atherosclerosis), associations between dietary flavonoid intakes (estimated from a food frequency questionnaire) and subclinical measures of atherosclerosis (ankle-brachial index, carotid plaques and intima-media thickness, and coronary artery calcification) were examined using repeated measures models. Exposures and outcomes were measured at exam 1 (2000-2002) and exam 5 (2010-2011). Stratified analyses and interaction terms were used to explore effect modification by time, sex, race/ethnicity, and smoking status. RESULTS: In the analytic population, at baseline, ≈46% were males with a median age of 62 (interquartile range, 53-70) years and total flavonoid intakes of 182 (interquartile range, 98-308) mg/d. After multivariable adjustments, participants with the highest (quartile 4) versus lowest (quartile 1) total flavonoid intakes had 26% lower odds of having an ankle-brachial index <1 (odds ratio, 0.74 [95% CI, 0.60-0.92]) and 18% lower odds of having a carotid plaque (odds ratio, 0.82 [95% CI, 0.69-0.99]), averaged over exams 1 and 5. Moderate (quartile 3) to high (quartile 4) intakes of flavonols, flavanol monomers, and anthocyanins were associated with 19% to 34% lower odds of having an ankle-brachial index <1 and 18% to 20% lower odds of having carotid plaque. Participants with the highest intakes of anthocyanins (quartile 4) at baseline had a marginally slower rate of carotid plaque progression than those with moderate intakes (quartiles 2 and 3). There were no significant associations with intima-media thickness or coronary artery calcification. Observed associations did not differ by sex, race/ethnicity, or smoking status. CONCLUSIONS: In this multi-ethnic population, higher dietary flavonoid intakes were associated with lower odds of peripheral and carotid artery atherosclerosis. Increasing intakes of healthy, flavonoid-rich foods may protect against atherosclerosis in the peripheral and carotid arteries.

Flavonol derivatives containing piperazine and quinoxaline fragments: synthesis and antifungal activity.

A series of flavonol derivatives containing piperazine and quinoxaline had been designed and synthesized. The biological activity test results showed that some of the target compounds had good antifungal activity against various fungi. N5 had the best antifungal activity against Phomopsis sp (P.s.) and Phytophthora capsica (P.c.). The half maximal effective concentration (EC50) was 12.9 and 25.8 µg/mL against P.s. and P.c., respectively, which were better than azoxystrobin (Az, 25.4 and 71.1 µg/mL). In addition, the protective and curative activities of N5 against kiwifruit were 85.9 and 67.0% at 200 µg/mL in vivo, which were better than that of Az (65.9 and 57.0%). The protective and curative activities against chili leaves were 80.6 and 66.5% at 200 µg/mL, which were better than that of Az (77.6 and 60.0%). The scanning electron microscopy (SEM) experiment showed that the action of N5 caused the mycelium to bend and fold, changed its morphology and caused damaged to the mycelium. Through the measurement of relative conductivity, leakage of cytoplasmic contents and determination of malondialdehyde (MDA) content indicated that N5 could damage the integrity of pathogenic fungal cell membranes, change the permeability of cell membranes, and affect the normal growth of mycelium.

Grapevine varieties show different sensitivities to flavonoid alterations caused by high temperatures under two irrigation conditions.

This study aimed to assess the response of four red grapevine (Vitis vinifera L.) varieties to elevated temperature, drought and their combination, focusing on the concentration and profile of grape flavonoids. Fruit-bearing cuttings of Tempranillo, Cabernet Sauvignon, Merlot and Grenache grew in greenhouses under, either ambient temperature (T) or ambient temperature + 4 °C (T+4). Plants also received either full irrigation (FI, substrate field capacity) or deficit irrigation (DI, 50 % substrate field capacity). In general, T+4 decreased the concentration of anthocyanins, but DI mitigated this effect. T+4 and DI increased the abundance of methylated anthocyanins and flavonols with additive effects. Grapes under T+4 had higher abundance of acylated anthocyanins, while DI increased the proportion of tri-hydroxylated anthocyanins and flavonols. The impact of interacting elevated temperature and drought on grape composition was genotype dependent. In terms of anthocyanin concentration and profile, Tempranillo was the most affected variety, whereas Grenache was less sensitive.

Design and Synthesis of an Azo Reductase Responsive Flavonol-Indomethacin Hybrid Used for the Diagnosis and Treatment of Colitis.

Human intestinal bacteria are the primary producers of azo reductase, and the content of azo reductase is closely associated with various intestinal diseases, including ulcerative colitis (UC). The rapid detection of changes in azo reductase levels is crucial for diagnosing and promptly intervening in UC. In this study, a therapeutic agent, FAI, specifically targeting UC, was designed and synthesized. This agent was developed by linking the anti-inflammatory drug indomethacin to flavonols with antioxidant activity via an azo bond (off-on). Breakage of the azo bond breaks results in the release of both fluorophores and drugs, achieving targeted tracing and integrated treatment effects. In vivo and in vitro fluorescence imaging experiments were used to demonstrate the potential of FAI in the diagnosis of UC, together with synergistic therapeutic effects through the release of both fluorophores and anti-inflammatory agents. Therefore, this diagnostic agent shows promise as a potential tool for diagnosing and treating UC.

Kaempferia galanga L. alleviates the expression of COX-2 and NF-kappaB-p65 in the oral mucosa ulcer of Wistar rats and exhibits no irritation toward the vascular membrane of chicken eggs and the skin of albino rabbits.

The global incidence of recurrent aphthous stomatitis in 2018 reached 5-66 % of the population, while in Indonesia 8 %. Moreover, the prevalence of oral mucosal fibrosis and recurrent aphthous stomatitis among male doctors and nurses in China was 21.24 % and 24.27 %, respectively. Our previous study has shown that the ethanol extract of Kaempferia galanga L. rhizome (EKGR) revealed an accelerated wound-healing effect in the oral mucosa ulcer of Wistar rats. This study aims to explore the effects of EKGR on the expression of NF-kappaB-p65 and COX-2 in the tongue tissue of male Wistar rats by Western blot analysis and immunohistochemistry technique, its safety towards the vascular membrane of the egg chorioallantoic membrane, and its single-dose application on the skin of male rabbits. The rats were randomly assigned into 7 groups: the normal control; the negative control; the positive control (treated with triamcinolone acetonide); and 4 treatment groups of EKGR (0.5 %; 1 %; 2 %; 4 %). Western blot and immunohistochemistry methods were used to measure the expression of NF-kappaB-p65 and COX-2. The hen's egg test-chorioallantoic membrane assay was employed to predict the safety of EKGR towards the vascular membrane. Moreover, the effect of 200 mg/kg BW EKGR application on the dorsal skin of male albino rabbits was also evaluated. EKGR inhibits the expression of NF-kappaB-p65 and COX-2 as proven by WB and IHC results. In the HET-CAM assay, all concentrations of EKGR do not induce irritation responses, which elicits the safety of EKGR. The administration of EKGR causes mild irritation to the dorsal skin of male rabbits but does not induce erythema and edema, no significant changes in BW, no toxic effects on organ macroscopic examination or histopathology, and does not induce abnormalities in the hematological profile of male albino rabbits. EKGR has confirmed its anti-inflammatory activity by suppressing the expression of COX-2 and NF-kappaB-p65 in the oral mucosa ulcer of Wistar rats. EKGR is safe as it does not exhibit irritating potential and harmful effects.

New combined Inverse-QSAR and molecular docking method for scaffold-based drug discovery.

Computer-aided drug discovery plays a vital role in developing novel medications for various diseases. The COVID-19 pandemic has heightened the need for innovative approaches to design lead compounds with the potential to become effective drugs. Specifically, designing promising inhibitors of the SARS-CoV-2 main protease (Mpro) is crucial, as it plays a key role in viral replication. Phytochemicals, primarily flavonoids and flavonols from medicinal plants, were screened. Fifty small molecules were selected for molecular docking analysis against SARS-CoV-2 Mpro (PDB ID: 6LU7). Binding energies and interactions were analyzed and compared to those of the anti-SARS-CoV-2 inhibitor Nirmatrelvir. Using these 50 structures as a training set, a QSAR model was built employing simple, reversible topological descriptors. An inverse-QSAR analysis was then performed on 29 = 512 hydroxyl combinations at nine possible positions on the flavone and flavonol scaffold. The model predicted three novel, promising compounds exhibiting the most favorable binding energies (-8.5 kcal/mol) among the 512 possible hydroxyl combinations: 3,6,7,2',4'-pentahydroxyflavone (PF9), 6,7,2',4'-tetrahydroxyflavone (PF11), and 3,6,7,4'-tetrahydroxyflavone (PF15). Molecular dynamics (MD) simulations demonstrated the stability of the PF9/Mpro complex over 300 ns of simulation. These predicted structures, reported here for the first time, warrant synthesis and further evaluation of their biological activity through in vitro and in vivo studies.

Flavonols improve tomato pollen thermotolerance during germination and tube elongation by maintaining ROS homeostasis.

Elevated temperatures impair pollen performance and reproductive success, resulting in lower crop yields. The tomato (Solanum lycopersicum) anthocyanin reduced (are) mutant harbors a mutation in FLAVANONE 3-HYDROXYLASE (F3H), resulting in impaired flavonol antioxidant biosynthesis. The are mutant has reduced pollen performance and seed set relative to the VF36 parental line, phenotypes that are accentuated at elevated temperatures. Transformation of are with the wild-type F3H gene, or chemical complementation with flavonols, prevented temperature-dependent reactive oxygen species (ROS) accumulation in pollen and restored the reduced viability, germination, and tube elongation of are to VF36 levels. Overexpression of F3H in VF36 prevented temperature-driven ROS increases and impaired pollen performance, revealing that flavonol biosynthesis promotes thermotolerance. Although stigmas of are had reduced flavonol and elevated ROS levels, the growth of are pollen tubes was similarly impaired in both are and VF36 pistils. RNA-seq was performed at optimal and stress temperatures in are, VF36, and the F3H overexpression line at multiple timepoints across pollen tube elongation. The number of differentially expressed genes increased over time under elevated temperatures in all genotypes, with the greatest number in are. These findings suggest potential agricultural interventions to combat the negative effects of heat-induced ROS in pollen that lead to reproductive failure.

Enhancing Vascular Health and Lowering Blood Pressure in Spontaneously Hypertensive Rats through Syrah Grape (Vitis vinifera) Pomace: The Role of Phenolic Compounds.

BACKGROUND: The beneficial properties of wine by-products include actions that help prevent and treat cardiovascular conditions such as hypertension, primarily due to their antioxidant effects. Novel pharmacotherapies are being developed to treat arterial hypertension, including investigations into natural products exhibiting biological activity, necessitating rigorous evaluation of their efficacy and safety. This study aimed to identify and quantify phenolic compounds in Syrah (Vitis vinifera) grapes grown in the Brazilian Cerrado and their presence in winemaking by-products. It also examined the effects of grape pomace on blood pressure. METHODS: Fresh grapes, pomace, and lees, were subjected to spectrophotometric determination of total phenolic compounds, followed by identification and quantification using HPLC-DAD-ESI-MSn. Normotensive male rats (Wistar) and spontaneously hypertensive rats (SHR) received grape pomace-enriched (150 or 300 mg/kg/day, 14 days) or standard chow. Indirect arterial pressure was assessed, while vascular reactivity was evaluated in mesenteric resistance arteries. RESULTS: Pomace samples exhibited higher total phenolic compound concentrations than grapes or lees. Seven derivatives of hydroxycinnamic acids and twenty-one flavonols were identified. Quercetin-3-glucoside and ethyl caffeate were the most abundant phenolic compounds. Grape pomace-enriched chow demonstrated a dose-dependent hypotensive effect in rats. CONCLUSION: the abundance of flavonols and hydroxycinnamic acids, combined with their hypotensive effects, underscores the therapeutic potential of fine wine-making by-products produced in the Brazilian Cerrado.

Exploring the Anti-Inflammatory and Antioxidant Potential, Metabolite Composition and Inorganic Profile of Cistus monspeliensis L. Aerial Parts and Roots.

This work focuses on Cistus monspeliensis L. aerial parts (AP) and roots (R) extracts, investigating the anti-inflammatory and antioxidant potential of the two organs in comparison. At dosages between 1.56 and 6.25 µg/mL, both extracts showed a protective effect against LPS inflammatory stimulus on a macrophage cell line (RAW 264.7). Interestingly, only R was able to significantly reduce both IL-1ß and IL-6 mRNA gene expression in the presence of LPS. Moreover, the treatment of a neuroblastoma cell line (SH-SY5Y) with AP and R at 6.25 µg/mL increased the cell survival rate by nearly 20% after H2O2 insult. However, only R promoted mitochondria survival, exhibited a significantly higher production of ATP and a higher activity of the enzyme catalase than the control. Both AP and R had similar primary metabolites; in particular, they both contained 1-O-methyl-epi-inositol. Labdane and methoxylated flavonoids were the most characteristic compounds of AP, while R contained mainly catechins, gallic acid, and pyrogallol derivatives. Considering the importance of elemental composition in plants, the inorganic profile of AP and R was also investigated and compared. No potentially toxic elements, such as Pb, were detected in any sample.

Attenuation of Quorum Sensing Mediated Virulence Factors and Biofilm Formation in Pseudomonas Aeruginosa PAO1 by Substituted Chalcones and Flavonols.

Flavonoids epitomize structural scaffolds in many biologically active synthetic and natural compounds. They showcase a diverse spectrum of biological activities including anticancer, antidiabetic, antituberculosis, antimalarial, and antibiofilm activities. The antibiofilm activity of a series of new chalcones and flavonols against clinically significant Pseudomonas aeruginosa PAO1 strain was studied. Antivirulence activities were screened by analysing the effect of compounds on the production of virulence factors like pyocyanin, LasA protease, cell surface hydrophobicity, and rhamnolipid. The best ligands towards the quorum sensing proteins LasR, RhlR, and PqsR were recognised using a molecular docking study. The gene expression in P. aeruginosa after treatment with test compounds was evaluated on quorum sensing genes including rhlA, lasB, and pqsE. The antibiofilm potential of chalcones and flavonols was confirmed by the efficient reduction in the production of virulence factors and downregulation of gene expression.

A Study of the UV Spectral Features in Wine and Their Correlation with Phenolic Constituents.

BACKGROUND: This study investigated the ultraviolet (UV) absorption spectra of various types and ages of grape wines and the correlation these spectra presented with their phenolic constituents. Firstly, the differences in UV spectra were characterized for different wine samples, depending on their type and age. METHODS: The following methods were used in this study: ultraviolet visible spectrophotometry, Folin-Ciocalteu spectrophotometric method, high-performance liquid chromatography. RESULTS: Then, it was demonstrated that for identically aged wines, the 280 nm absorbance is proportional to the concentration of phenolic compounds, as determined by the Folin-Ciocalteu method. Next, an investigation was conducted into the absorption coefficients of different phenolic classes commonly found in grapes and wine. Finally, the range in variation of phenolic compounds in various types of grape wines was established. CONCLUSIONS: This work provides a methodological approach to rapidly determine the concentration of phenolic compounds in wines using UV spectroscopy, provided that their age is known. As UV spectrophotometers are available in nearly all laboratories, this may provide a cheaper and faster alternative to current methods, including high-performance liquid chromatography (HPLC).

Association of enzymatic and optimized ultrasound-assisted aqueous extraction of flavonoid glycosides from dried Hippophae rhamnoides L. (Sea Buckthorn) berries.

The main purpose of the present study was to determine the effect of associating an optimized ultrasound-assisted extraction (UAE) protocol with enzyme-assisted extraction (EAE) in aqueous media, using the dried berries of Hippophae rhamnoides L. (sea buckthorn) as plant material. A specialized software was used for the determination of potential optimal extraction parameters, leading to the development of four optimized extracts with different characteristics (UAE ± EAE). For these extracts, buffered or non-buffered solutions have been used, with the aim to determine the influence of adjustable pH on extractability. As enzymatic solution, a pectinase, cellulase, and hemicellulase mix (2:1:1) has been applied, acting as pre-treatment for the optimized protocol. The highest extractive yields have been identified for non-buffered extracts, and the E-UAE combination obtained extracts with the highest overall in vitro antioxidant activity. The HPLC-MSn analysis demonstrated a rich composition in different types of isorhamnetin-O-glycosides, as well as some quercetin-O-glycosides, showing a high recovery of specific flavonol-type polyphenolic species. Moreover, we have tentatively identified two flavanols (i.e., catechin and epigallocatechin) and one flavone derivative (i.e., luteolin).

Flavonoids from the leaves and stems of Sedum japonicum var. senanense and their antioxidant activity.

Twenty flavonoids (1-20) were isolated from the leaves and stems of Sedum japonicum var. senanense endemic to Japan. Among them, nine compounds were reported in nature for the first time, and identified as herbacetin 3-O-neohesperidoside-8-O-(2‴-acetylxyloside) (2), gossypetin 8-O-(2″-acetylxyloside) (4), gossypetin 8-O-(3″-acetylxyloside) (5), gossypetin 3-O-glucoside-8-O-(3‴-acetylxyloside) (9), gossypetin 3-O-glucoside-8-O-(2‴,3‴-diacetylxyloside) (10), gossypetin 3-O-neohesperidoside-8-O-xyloside (11), gossypetin 3-O-neohesperidoside-8-O-(2⁗-acetylxyloside) (12), gossypetin 3-O-neohesperidoside-8-O-(3⁗-acetylxyloside) (13) and gossypetin 3-O-glucoside-8-O-xylofuranoside (14) by UV spectral survey, HR-MS, LC-MS, acid hydrolysis, NMR including 1H and 13C NMR, COSY, NOESY, HSQC and HMBC. Moreover, nine major flavonoids were surveyed for antioxidant activity by H-ORAC method. As the results, gossypetin 3-O-glucoside-8-O-(2‴-acetylxyoside) (8) showed the highest antioxidant activity. Conversely, gossypetin 3-O-neohesperidoside-8-O-xyloside (11) and gossypetin 3-O-neohesperidoside-8-O-(2⁗-acetylxyloside) (12) which attach neohesperidose showed the lowest values.

Structural and Interactional Analysis of the Flavonoid Pathway Proteins: Chalcone Synthase, Chalcone Isomerase and Chalcone Isomerase-like Protein.

Chalcone synthase (CHS) and chalcone isomerase (CHI) catalyze the first two committed steps of the flavonoid pathway that plays a pivotal role in the growth and reproduction of land plants, including UV protection, pigmentation, symbiotic nitrogen fixation, and pathogen resistance. Based on the obtained X-ray crystal structures of CHS, CHI, and chalcone isomerase-like protein (CHIL) from the same monocotyledon, Panicum virgatum, along with the results of the steady-state kinetics, spectroscopic/thermodynamic analyses, intermolecular interactions, and their effect on each catalytic step are proposed. In addition, PvCHI's unique activity for both naringenin chalcone and isoliquiritigenin was analyzed, and the observed hierarchical activity for those type-I and -II substrates was explained with the intrinsic characteristics of the enzyme and two substrates. The structure of PvCHS complexed with naringenin supports uncompetitive inhibition. PvCHS displays intrinsic catalytic promiscuity, evident from the formation of p-coumaroyltriacetic acid lactone (CTAL) in addition to naringenin chalcone. In the presence of PvCHIL, conversion of p-coumaroyl-CoA to naringenin through PvCHS and PvCHI displayed ~400-fold increased Vmax with reduced formation of CTAL by 70%. Supporting this model, molecular docking, ITC (Isothermal Titration Calorimetry), and FRET (Fluorescence Resonance Energy Transfer) indicated that both PvCHI and PvCHIL interact with PvCHS in a non-competitive manner, indicating the plausible allosteric effect of naringenin on CHS. Significantly, the presence of naringenin increased the affinity between PvCHS and PvCHIL, whereas naringenin chalcone decreased the affinity, indicating a plausible feedback mechanism to minimize spontaneous incorrect stereoisomers. These are the first findings from a three-body system from the same species, indicating the importance of the macromolecular assembly of CHS-CHI-CHIL in determining the amount and type of flavonoids produced in plant cells.

A functional study reveals CsNAC086 regulated the biosynthesis of flavonols in Camellia sinensis.

MAIN CONCLUSION: CsNAC086 was found to promote the expression of CsFLS, thus promoting the accumulation of flavonols in Camellia sinensis. Flavonols, the main flavonoids in tea plants, play an important role in the taste and quality of tea. In this study, a NAC TF gene CsNAC086 was isolated from tea plants and confirmed its regulatory role in the expression of flavonol synthase which is a key gene involved in the biosynthesis of flavonols in tea plant. Yeast transcription-activity assays showed that CsNAC086 has self-activation activity. The transcriptional activator domain of CsNAC086 is located in the non-conserved C-terminal region (positions 171-550), while the conserved NAC domain (positions 1-170) does not have self-activation activity. Silencing the CsNAC086 gene using antisense oligonucleotides significantly decreased the expression of CsFLS. As a result, the concentration of flavonols decreased significantly. In overexpressing CsNAC086 tobacco leaves, the expression of NtFLS was significantly increased. Compared with wild-type tobacco, the flavonols concentration increased. Yeast one-hybrid assays showed CsNAC086 did not directly regulate the gene expression of CsFLS. These findings indicate that CsNAC086 plays a role in regulating flavonols biosynthesis in tea plants, which has important implications for selecting and breeding of high-flavonols-concentration containing tea-plant cultivars.

Effect of Flavonols of Aronia melanocarpa Fruits on Morphofunctional State of Immunocompetent Organs of Rats under Cyclophosphamide-Induced Immunosuppression.

Aronia melanocarpa berries contain many compounds with potential benefits for human health. The food flavonoids quercetin and rutin, found in significant amounts in the fruits of A. melanocarpa, are known to have favourable effects on animal and human organisms. However, data on the effect of flavonols isolated from black chokeberry on immune functions during immunosuppression are not available in the literature. Thus, the aim of this study was to evaluate the effect of flavonol fraction isolated from A. melanocarpa fruits, in comparison with pure quercetin and rutin substances, on the dysfunctional state of rat thymus and spleen in immunodeficiency. The study was performed on Wistar rats. The animals were orally administered solutions of the investigated substances for 7 days: water, a mixture of quercetin and rutin and flavonol fraction of A. melanocarpa. For induction of immunosuppression, the animals were injected once intraperitoneally with cyclophosphamide. Substance administration was then continued for another 7 days. The results showed that under the influence of flavonols, there was a decrease in cyclophosphamide-mediated reaction of lipid peroxidation enhancement and stimulation of proliferation of lymphocytes of thymus and spleen in rats. At that, the effect of the flavonol fraction of aronia was more pronounced.

Variability in the Qualitative and Quantitative Composition of Phenolic Compounds and the In Vitro Antioxidant Activity of Sour Cherry (Prunus cerasus L.) Leaves.

Sour cherry (Prunus cerasus L.) is a deciduous tree belonging to the Rosaceae Juss. family. Cherry leaves are an underutilized source of biologically active compounds. The aim of this study was to determine the composition of the phenolic compounds, as well as the total antioxidant activity, in leaf samples of P. cerasus cultivars and to elucidate the cultivars with particular phytochemical compositions. The phytochemical profiles of P. cerasus leaves vary significantly in a cultivar-dependent manner. The total content of identified phenolic compounds varied from 8.254 to 16.199 mg/g in the cherry leaves. Chlorogenic acid ranged between 1413.3 µg/g ('North Star') and 8028.0 µg/g ('Note'). The total content of flavonols varied from 4172.5 µg/g ('Vytenu zvaigzde') to 9030.7 µg/g ('Tikhonovskaya'). The total content of identified proanthocyanidins varied from 122.3 µg/g ('Note') to 684.8 µg/g ('Kelleris'). The highest levels of phloridzin (38.1 ± 0.9 µg/g) were found in samples of 'Molodezhnaya', while the lowest level of this compound was determined in the leaf samples of 'Turgenevka' (6.7 ± 0.2). The strongest antiradical (138.0 ± 4.0 µmol TE/g, p < 0.05) and reducing (364.9 ± 10.5 µmol TE/g, p < 0.05) activity in vitro was exhibited by the cultivar 'Vytenu zvaigzde' cherry leaf sample extracts. 'Kelleris', 'Note', and 'Tikhonovskaya' distinguish themselves with peculiar phytochemical compositions.

The interaction of plant flavones with amphotericin B: Consequences for its pore-forming ability.

The growth of antibiotic resistance to antifungal drugs contributes to the search for new ways to enhance their effectiveness and reduce toxicity. The undeniable advantage of polyene macrolide antibiotic amphotericin B (AmB) which ensures low pathogen resistance is its mechanism of action related to the formation of transmembrane pores in target lipid membranes. Here, we investigated the effects of plant flavones, chrysin, wogonin, baicalein, apigenin, scutellarein, luteolin, morin and fisetin on the pore-forming activity of AmB in the sterol-enriched membranes by electrophysiological assays. Сhrysin, wogonin, baicalein, apigenin, scutellarein, and luteolin were shown to decrease the AmB pore-forming activity in the bilayers composed of palmitoyloleylphosphocholine independently of their sterol composition. Morin and fisetin led to the increase and decrease in the AmB pore-forming activity in the ergosterol- and cholesterol-containing bilayers respectively. Differential scanning microcalorimetry of the gel-to-liquid crystalline phase transition of membrane forming lipids, molecular dynamics simulations, and absorbance spectroscopy revealed the possibility of direct interactions between AmB and some flavones in the water and/or in the lipid bilayer. The influence of these interactions on the antibiotic partitioning between aqueous solution and membrane and/or its transition between different states in the bilayer was discussed.

R2R3-MYB transcription factor CsMYB60 controls mature fruit skin color by regulating flavonoid accumulation in cucumber.

Skin color is an important trait that determines the cosmetic appearance and quality of fruits. In cucumber, the skin color ranges from white to brown in mature fruits. However, the genetic basis for this important trait remains unclear. We conducted a genome-wide association study of natural cucumber populations, along with map-based cloning techniques, on an F2 population resulting from a cross between Pepino (with yellow-brown fruit skin) and Zaoer-N (with creamy fruit skin). We identified CsMYB60 as a candidate gene responsible for skin coloration in mature cucumber fruits. In cucumber accessions with white to pale yellow skin color, a premature stop mutation (C to T) was found in the second exon region of CsMYB60, whereas light yellow cucumber accessions exhibited splicing premature termination caused by an intronic mutator-like element insertion in CsMYB60. Transgenic CsMYB60c cucumber plants displayed a yellow-brown skin color by promoting accumulation of flavonoids, especially hyperoside, a yellow-colored flavonol. CsMYB60c encodes a nuclear protein that primarily acts as a transcriptional activator through its C-terminal activation motif. RNA sequencing and DNA affinity purification sequencing assays revealed that CsMYB60c promotes skin coloration by directly binding to the YYTACCTAMYT motif in the promoter regions of flavonoid biosynthetic genes, including CsF3'H, which encodes flavonoid 3'-hydroxylase. The findings of our study not only offer insight into the function of CsMYB60 as dominantly controlling fruit coloration, but also highlight that intronic DNA mutations can have a similar phenotypic impact as exonic mutations, which may be valuable in future cucumber breeding programs.

A maize enzyme from the 2-oxoglutarate-dependent oxygenase family with unique kinetic properties, mediates resistance against pathogens and regulates senescence.

In plants, salicylic acid (SA) hydroxylation regulates SA homoeostasis, playing an essential role during plant development and response to pathogens. This reaction is catalysed by SA hydroxylase enzymes, which hydroxylate SA producing 2,3-dihydroxybenzoic acid (2,3-DHBA) and/or 2,5-dihydroxybenzoic acid (2,5-DHBA). Several SA hydroxylases have recently been identified and characterised from different plant species, but no such activity has yet been reported in maize. In this work, we describe the identification and characterisation of a new SA hydroxylase in maize plants. This enzyme, with high sequence similarity to previously described SA hydroxylases from Arabidopsis and rice, converts SA into 2,5-DHBA; however, it has different kinetic properties to those of previously characterised enzymes, and it also catalysers the conversion of the flavonoid dihydroquercetin into quercetin in in vitro activity assays, suggesting that the maize enzyme may have different roles in vivo to those previously reported from other species. Despite this, ZmS5H can complement the pathogen resistance and the early senescence phenotypes of Arabidopsis s3h mutant plants. Finally, we characterised a maize mutant in the S5H gene (s5hMu) that has altered growth, senescence and increased resistance against Colletotrichum graminicola infection, showing not only alterations in SA and 2,5-DHBA but also in flavonol levels. Together, the results presented here provide evidence that SA hydroxylases in different plant species have evolved to show differences in catalytic properties that may be important to fine tune SA levels and other phenolic compounds such as flavonols, to regulate different aspects of plant development and pathogen defence.