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Allene oxide synthase (AOS) is the second enzyme in the biosynthesis of the plant defensive hormone jasmonic acid (JA). In rice, there are two AOSs, OsAOS1 and OsAOS2. However, the role of these two AOS genes in herbivore-induced defenses in rice remains unidentified. We cloned the two rice AOS genes and observed that the transcript level of both OsAOS1 and OsAOS2 was enhanced by mechanical wounding, the infestation of the striped stem borer (SSB) (Chilo suppressalis) or brown planthopper (BPH) (Niaparvata lugens), and treatment with JA; however, OsAOS1 responded more rapidly to SSB infestation and JA treatment than did OsAOS2. The antisense expression of OsAOS1 (as-aos1) or OsAOS2 (as-aos2) decreased levels of SSB- or BPH-induced JA, which, in turn, reduced the production of SSB-induced trypsin protease inhibitor (TrypPI) and volatiles as well as the resistance of rice to SSB. In contrast, BPH preferred to feed and oviposit on wild-type (WT) plants over as-aos1 and as-aos2 plants. Moreover, the survival of BPH nymphs on as-aos1 or as-aos2 lines was significantly lower than on WT plants. The increased resistance of as-aos1 or as-aos2 plants to BPH correlated with higher levels of BPH-induced H2O2 and SA. These results indicate that OsAOS1 and OsAOS2 are both involved in herbivore-induced JA biosynthesis and play a vital role in determining the resistance of rice to chewing and phloem-feeding herbivores.
Currently, no clinical methods reliably predict the development of castration-resistant prostate cancer (CRPC) that occurs almost universally in men undergoing androgen deprivation therapy. Hyperpolarized (HP) 13C magnetic resonance imaging (MRI) could potentially detect the incipient emergence of CRPC based on early metabolic changes. To characterize metabolic shifts occurring upon the transition from androgen-dependent to castration-resistant prostate cancer (PCa), the metabolism of [U-13C]glucose and [U-13C]glutamine was analyzed by nuclear magnetic resonance spectroscopy. Comparison of steady-state metabolite concentrations and fractional enrichment in androgen-dependent LNCaP cells and transgenic adenocarcinoma of the murine prostate (TRAMP) murine tumors versus castration-resistant PC-3 cells and treatment-driven CRPC TRAMP tumors demonstrated that CRPC was associated with upregulation of glycolysis, tricarboxylic acid metabolism of pyruvate; and glutamine, glutaminolysis, and glutathione synthesis. These findings were supported by 13C isotopomer modeling showing increased flux through pyruvate dehydrogenase (PDH) and anaplerosis; enzymatic assays showing increased lactate dehydrogenase, PDH and glutaminase activity; and oxygen consumption measurements demonstrating increased dependence on anaplerotic fuel sources for mitochondrial respiration in CRPC. Consistent with ex vivo metabolomic studies, HP [1-13C]pyruvate distinguished androgen-dependent PCa from CRPC in cell and tumor models based on significantly increased HP [1-13C]lactate.
Leuconostoc spp. are environmental microorganisms commonly associated with fermented foods. Absence of antibiotic resistance (AR) in bacteria is a critical issue for global food safety. Herein, we updated the occurrence of AR genes in the Leuconostoc genus through in silico analyses of the genomes of 17 type strains. A total of 131 putative AR traits associated with the main clinically relevant antibiotics were detected. We found, for the first time, the lsaA gene in L. fallax ATCC 700006T and L. pseudomesenteroides NCDO 768T. Their amino acid sequences displayed high similarities (59.07% and 52.21%) with LsaA of Enterococcusfaecalis V583, involved in clindamycin (CLI) and quinupristin-dalfopristin (QUD) resistance. This trait has different distribution patterns in Leuconostoc nontype strains-i.e., L. pseudomesenteroides, L. lactis and L. falkenbergense isolates from fermented vegetables, cheeses, and starters. To better explore the role of lsaA, MIC for CLI and QUD were assessed in ATCC 700006T and NCDO 768T; both strains were resistant towards CLI, potentially linking lsaA to their resistant phenotype. Contrarily, NCDO 768T was sensitive towards QUD; however, expression of lsaA increased in presence of this antibiotic, indicating an active involvement of this trait and thus suggesting a revision of the QUD thresholds for this species.
Myofibrillar myopathies (MFM) are heterogeneous hereditary muscle diseases with characteristic myopathological features of Z-disk dissolution and aggregates of its degradation products. The onset and progression of the disease are variable, with an elusive genetic background, and around half of the cases lacking molecular diagnosis. Here, we attempted to establish possible genetic foundations of MFM by performing whole exome sequencing (WES) in eleven unrelated families of 13 patients clinically diagnosed as MFM spectrum. A filtering strategy aimed at identification of variants related to the disease was used and included integrative analysis of WES data and human phenotype ontology (HPO) terms, analysis of muscle-expressed genes, and analysis of the disease-associated interactome. Genetic diagnosis was possible in eight out of eleven cases. Putative causative mutations were found in the DES (two cases), CRYAB, TPM3, and SELENON (four cases) genes, the latter typically presenting with a rigid spine syndrome. Moreover, a variety of additional, possibly phenotype-affecting variants were found. These findings indicate a markedly heterogeneous genetic background of MFM and show the usefulness of next generation sequencing in the identification of disease-associated mutations. Finally, we discuss the emerging concept of variant load as the basis of phenotypic heterogeneity.
Virtual reality (VR) was introduced to maximize the effect of cognitive behavioral therapy (CBT) by efficiently performing exposure therapy. The purpose of this study was to find out whether VR-based individual CBT with relatively few treatment sessions is effective in improving social anxiety disorder (SAD). This therapy was applied to 115 patients with SAD who were retrospectively classified into 43 patients who completed the nine or 10 sessions normally (normal termination group), 52 patients who finished the sessions early (early termination group), and 20 patients who had extended the sessions (session extension group). The Brief Fear of Negative Evaluation Scale (BFNE) scores tended to decrease in all groups as the session progressed, and the slope of decrease was the steepest in the early termination group and the least steep in the session extension group. Severity of social anxiety in the last session and symptom reduction rate showed no significant group difference. Our findings suggest that short-term VR-based individual CBT of nine to 10 sessions may be effective. When the therapeutic effect is insufficient during this period, the additional benefit may be minimal if the session is simply extended. The improvement in the early termination group suggests that even shorter sessions of five or six can also be effective.
Background and objectives: Ureaplasma urealyticum (UU) and Mycoplasma hominis (MH) are two commensal microorganisms that form the urogenital microbiota. Under a state of dysbiosis, both bacteria cause intrauterine infection. Material and methods: Therefore, the purpose of the present study was to analyze the prevalence of UU and MH among four hundred and eleven infertile women. Results: Women between thirty and thirty-five years old were the most affected group, followed by those that were 25 and 30 years old, respectively. Cumulatively, the prevalence of single UU and MH, and coinfection, was 28.46% (n = 117), (n = 2) 0.48%, and 2.91% (n = 12), respectively, with an overall detection rate of 31.87% (n = 131). To assess the associated drug susceptibility, endocervical samples were unequally sent to Regina Maria (n = 281) and Synevo (n = 130) laboratories for further analyses. Pristinamycin (100% vs. 100%) and Josamycin (100% vs. 98.00%) were the most efficient antibiotics in eradicating UU and MH, several others also displaying a high efficiency, among which can be mentioned Doxycycline (98.23%), Minocycline (96.00%), Tetracycline (96.48% vs. 68.00%), and Erythromycin (70.17% vs. 92.00%). Based on antibiograms, Clarithromycin (88.00%), Roxithromycin (88.00%), Levofloxacin (82.00%), and Azithromycin (78.94%) can be further used in treating such infections. On the other hand, Clindamycin (4.00%) and Ciprofloxacin (12.27% vs. 2.00%) are no longer viable because both UU and MH display an intermediate response towards gained resistance. Interestingly, the efficiency of Ofloxacin (22.79% vs. 60.00%) was conflicting, this possibly suggesting a transient stage to a gradual adaptability of these microorganisms to Ofloxacin. Conclusions: The most susceptible age groups in each case were women that were between twenty and forty years old. It can be concluded that four antibiotics can be safely used for treating UU, MH, or dual infections whose efficiency was over 95%.
Thermochemical materials (TCM) are among the most promising systems to store high energy density for long-term energy storage. To be eligible as candidates, the materials have to fit many criteria such as complete reversibility of the reaction and cycling stability, high availability of the material at low cost, environmentally friendliness, and non-toxicity. Among the most promising TCM, the Mg(OH)2/MgO system appears worthy of attention for its properties in line with those required. In the last few decades, research focused its attention on the optimization of attractive hydroxide performance to achieve a better thermochemical response, however, often negatively affecting its energy density per unit of volume and therefore compromising its applicability on an industrial scale. In this study, pure Mg(OH)2 was developed using different synthesis procedures. Reverse deposition precipitation and deposition precipitation methods were used to obtain the investigated samples. By adding a cationic surfactant (cetyl trimethylammonium bromide), deposition precipitation Mg(OH)2 (CTAB-DP-MH) or changing the precipitating precursor (N-DP-MH), the structural, physical and morphological characteristics were tuned, and the results were compared with a commercial Mg(OH)2 sample. We identified a correlation between the TCM properties and the thermochemical behavior. In such a context, it was demonstrated that both CTAB-DP-MH and N-DP-MH improved the thermochemical performances of the storage medium concerning conversion (64 wt.% and 74 wt.% respectively) and stored and released heat (887 and 1041 kJ/kgMg(OH)2). In particular, using the innovative technique not yet investigated for thermal energy storage (TES) materials, with NaOH as precipitating precursor, N-DP-MH reached the highest stored and released heat capacity per volume unit, ~684 MJ/m3.
The study investigated the phenols, sugar and the antioxidant capacities of date fruit extracts obtained by organic solvents and by hydrothermal treatment from six different Algerian cultivars at two ripening stages for the first time. The analyzed cultivars exhibited potent antioxidant properties (ferric reducing antioxidant power (FRAP), 1,1-Diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) scavenging capacities) and different phenols regardless of the solvents and the maturity stages. About 18 phenols were identified and quantified, mainly in the hydrothermal extracts. The earlier stages were characterized by high amounts of o-coumaric acid, cinnamic acid and luteolin, with a noticeable absence of quercetin. The tamr stage presented the highest sugar content (78.15-86.85 mg/100 mg dry weight (DW)) with an abundance of glucose. Galactose was present only in some cultivars from the kimri stage (tamjouhert). Uronic acids were mostly detected at the tamr stage (4.02-8.82 mg gallic acid equivalent/100 mg dried weight). The obtained results highlight the potential of using date fruit extracts as natural antioxidants, especially at industrial scales that tend use hydrothermal extraction.
Viral infection-induced activation of inflammasome complexes has both positive and negative effects on the host. Proper activation of inflammasome complexes induces down-stream effector mechanisms that inhibit viral replication and promote viral clearance, whereas dysregulated activation has detrimental effects on the host. Coronaviruses, including SARS-CoV and MERS-CoV, encode viroporins that activate the NLRP3 inflammasome, and the severity of coronavirus disease is associated with the inflammasome activation. Although the NLRP3 inflammasome activation is implicated in the pathogenesis of coronaviruses, these viruses must evade inflammasome-mediated antiviral immune responses to establish primary replication. Screening of a complementary DNA (cDNA) library encoding 28 SARS-CoV-2 open reading frames (ORFs) showed that two nonstructural proteins (NSPs), NSP1 and NSP13, inhibited caspase-1-mediated IL-1β activation. NSP1 amino acid residues involved in host translation shutoff and NSP13 domains responsible for helicase activity were associated with caspase-1 inhibition. In THP-1 cells, both NSP1 and NSP13 significantly reduced NLRP3-inflammasome-induced caspase-1 activity and IL-1β secretion. These findings indicate that SARS-CoV-2 NSP1 and NSP13 are potent antagonists of the NLRP3 inflammasome.
Fluorescence in situ hybridization (FISH) and Hi-C methods are largely used to investigate the three-dimensional organization of the genome in the cell nucleus and are applied here to study the organization of genes (LMBR1, NOM1, MNX1, UBE3C, PTPRN2) localized in the human 7q36.3 band. This region contains the MNX1 gene, which is normally not expressed in human lymphocytes beyond embryonic development. However, this homeobox gene is frequently activated in leukemic cells and its expression is associated with an altered gene positioning in the leukemia cell nuclei. In this study, we used FISH on 3D-preserved nuclei to investigate the nuclear positioning of MNX1 in the leukemia-derived cell line K562. Of the five copies of the MNX1 gene present in K562, four alleles were positioned in the nuclear periphery and only one in the nuclear interior. Using the Juicebox's Hi-C dataset, we identified five chromatin loops in the 7q36.3 band, with different extensions related to the size and orientation of the genes located here, and independent from their expression levels. We identified similar loops in 11 human and three mouse cell lines, showing that these loops are highly conserved in different human cell lines and during evolution. Moreover, the chromatin loop organization is well conserved also during neuronal cell differentiation, showing consistency in genomic organization of this region in development. In this report, we show that FISH and Hi-C are two different approaches that complement one another and together give complete information on the nuclear organization of specific chromosomal regions in different conditions, including cellular differentiation and genetic diseases.
Biological aging is associated with various morphological and functional changes, yet the mechanisms of these phenomena remain unclear in many tissues and organs. Hyperlipidemia is among the factors putatively involved in the aging of the liver and heart. Here, we analyzed morphological, ultrastructural, and biochemical features in adult (7-month-old) and elderly (17-month-old) mice, and then compared age-related features between wild type (C57Bl/6 strain) and ApoE-deficient (transgenic ApoE-/-) animals. Increased numbers of damaged mitochondria, lysosomes, and lipid depositions were observed in the hepatocytes of elderly animals. Importantly, these aging-related changes were significantly stronger in hepatocytes from ApoE-deficient animals. An increased number of damaged mitochondria was observed in the cardiomyocytes of elderly animals. However, the difference between wild type and ApoE-deficient mice was expressed in the larger size of mitochondria detected in the transgenic animals. Moreover, a few aging-related differences were noted between wild type and ApoE-deficient mice at the level of plasma biochemical markers. Levels of cholesterol and HDL increased in the plasma of elderly ApoE-/- mice and were markedly higher than in the plasma of elderly wild type animals. On the other hand, the activity of alanine transaminase (ALT) decreased in the plasma of elderly ApoE-/- mice and was markedly lower than in the plasma of elderly wild type animals.
The mycotoxin ochratoxin A (OTA) is a potent nephrocarcinogen, mainly in male rats. The aim of this study was to determine the time course of gene expression (GeneChip® Rat Gene 2.0 ST Array, Affymetrix) in kidney samples from male and female F344 rats, treated daily (p.o) with 0.50 mg/kg b.w. (body weight) of OTA for 7 or 21 days, and evaluate if there were differences between both sexes. After OTA treatment, there was an evolution of gene expression in the kidney over time, with more differentially expressed genes (DEG) at 21 days. The gene expression time course was different between sexes with respect to the number of DEG and the direction of expression (up or down): the female response was progressive and consistent over time, whereas males had a different early response with more DEG, most of them up-regulated. The statistically most significant DEG corresponded to metabolism enzymes (Akr1b7, Akr1c2, Adh6 down-regulated in females; Cyp2c11, Dhrs7, Cyp2d1, Cyp2d5 down-regulated in males) or transporters (Slc17a9 down-regulated in females; Slco1a1 (OATP-1) and Slc51b and Slc22a22 (OAT) down-regulated in males). Some of these genes had also a basal sex difference and were over-expressed in males or females with respect to the other sex.
The dissolution and extraction of lignin from biomass represents a great challenge due to the complex structure of this natural phenolic biopolymer. In this work, several surfactants (i.e., non-ionic, anionic, and cationic) were used as additives to enhance the dissolution efficiency of model lignin (kraft) and to boost lignin extraction from pine sawdust residues. To the best of our knowledge, cationic surfactants have never been systematically used for lignin dissolution. It was found that ca. 20 wt.% of kraft lignin is completely solubilized using 1 mol L-1 octyltrimethylammonium bromide aqueous solution. A remarkable dissolution efficiency was also obtained using 0.5 mol L-1 polysorbate 20. Furthermore, all surfactants used increased the lignin extraction with formic acid, even at low concentrations, such as 0.01 and 0.1 mol L-1. Higher concentrations of cationic surfactants improve the extraction yield but the purity of extracted lignin decreases.
Bacteria belonging to the genus Aneurinibacillus within the family Paenibacillaceae are Gram-positive, endospore-forming, and rod-shaped bacteria inhabiting diverse environments. Currently, there are eight validly described species of Aneurinibacillus; however, several unclassified species have also been reported. Aneurinibacillus spp. have shown the potential for producing secondary metabolites (SMs) and demonstrated diverse types of enzyme activities. These features make them promising candidates with industrial implications. At present, genomes of 9 unique species from the genus Aneurinibacillus are available, which can be utilized to decipher invaluable information on their biosynthetic potential as well as enzyme activities. In this work, we performed the comparative genome analyses of nine Aneurinibacillus species representing the first such comprehensive study of this genus at the genome level. We focused on discovering the biosynthetic, biodegradation, and heavy metal resistance potential of this under-investigated genus. The results indicate that the genomes of Aneurinibacillus contain SM-producing regions with diverse bioactivities, including antimicrobial and antiviral activities. Several carbohydrate-active enzymes (CAZymes) and genes involved in heavy metal resistance were also identified. Additionally, a broad range of enzyme classes were also identified in the Aneurinibacillus pan-genomes, making this group of bacteria potential candidates for future investigations with industrial applications.
Senescence marker protein 30 (SMP30) is a cell survival factor playing an important role in vitamin C synthesis and antiapoptosis. Moreover, its cytoprotective role suggests a possibility to be related to cancer cell survival. Mammary carcinoma is a common cancer in both humans and animals. Because of its histopathological diversity, especially in the early stage, histopathological diagnosis may be complicated; therefore, a diagnostic marker is helpful for confirmation. The present study analyzed the expression pattern of SMP30 in mammary carcinoma in humans, dogs, and cats. Immunohistochemistry, immunofluorescence, and western blot analysis were used to investigate SMP30 expression patterns. The expression was specifically observed in neoplastic glandular epithelial cells. The expression increased with the malignancy of glandular epithelial cells with a highly proliferative status. However, SMP30 expression was low in normal mammary gland tissues or well-differentiated adenoma tissues. The patterns were consistently reproduced in canine primary mammary carcinoma cells and MCF-7 and MDA-MB-231 human carcinoma cell lines. This study provides useful information to understand SMP30 expression in various stages of mammary carcinoma and to suggest its utility as a pan-species diagnostic marker, thereby helping to establish strategies for diagnosing mammary carcinoma in several species.
Indocyanine green (ICG) is a clinically approved near-infrared dye that has shown promise as a photosensitizer for the phototherapy of cancer. However, its chemical instability in an aqueous solution has limited its clinical application. Encapsulating ICG in liposomes, phosphatidylcholine nanoparticles (PC-NP), has shown partial effectiveness in stabilizing it. Prompted by our recent finding that the zein-phosphatidylcholine hybrid nanoparticles (Z/PC-NP) provide an advanced drug carrier compared to PC-NP, we herein investigated the potential of Z/PC-NP as an improved ICG formulation. Dynamic light scattering analysis, transmission electron microscopy, and Fourier-transform infrared spectroscopy studies showed that ICG was encapsulated in Z/PC-NP without hampering the high colloidal stability of the Z/PC-NP. During storage, the Z/PC-NP almost completely inhibited the ICG aggregation, whereas the PC-NP did so partially. The Z/PC-NP also more effectively blocked the ICG degradation compared to the PC-NP. The phototoxicity of ICG encapsulated in Z/PC-NP on cancer cells was twofold higher than that in the PC-NP. The ICG encapsulated in Z/PC-NP, but not in PC-NP, maintained its photocytotoxicity after four-day storage. These findings highlight the promising potential of Z/PC-NP as an ICG formulation that provides a higher stabilization effect than PC-NP.
Forward osmosis (FO) modules currently suffer from performance efficiency limitations due to concentration polarisation (CP), as well as pressure drops during operation. There are incentives to further reduce CP effects, as well as optimise spacer design for pressure drop improvements and mechanical support. In this study, the effects of applying transmembrane pressure (TMP) on FO membrane deformation and the subsequent impact on module performance was investigated by comparing experimental data to 3D computational fluid dynamics (CFD) simulations for three commercial FO modules. At a TMP of 1.5 bar the occlusion of the draw-channel induced by longitudinal pressure hydraulic drop was comparable for the Toray (16%) and HTI modules (12%); however, the hydraulic perimeter of the Profiera module was reduced by 46%. CFD simulation of the occluded channels indicated that a change in hydraulic perimeter due to a 62% increase in shear strain resulted in a 31% increase in the Reynolds number. This reduction in channel dimensions enhanced osmotic efficiency by reducing CP via improved draw-channel hydrodynamics, which significantly disrupted the external concentration polarization (ECP) layer. Furthermore, simulations indicated that the Reynolds number experienced only modest increases with applied TMP and that shear strain at the membrane surface was found to be the most important factor when predicting flux performance enhancement, which varied between the different modules. This work suggests that a numerical approach to assess the effects of draw-spacers on pressure drop and CP can optimize and reduce investment in the design and validation of FO module designs.
Fusarium genus comprises important saprophytic and phytopathogenic fungi and is widespread in nature. The present study reports the occurrence of Fusarium spp. in soils from two mangrove forests in northern Peninsular Malaysia and analyzed physico-chemical properties of the mangrove soil. Based on TEF-1α sequences, nine Fusarium species were identified: Fusarium solani species complex (FSSC) (n = 77), Fusarium verticillioides (n = 20), Fusarium incarnatum (n = 10), Fusarium proliferatum (n = 7), Fusarium lateritium (n = 4), Fusarium oxysporum (n = 3), Fusarium rigidiuscula (n = 2), Fusarium chlamydosporum (n = 1), and Fusarium camptoceras (n = 1); FSSC isolates were the most prevalent. Phylogenetic analysis of the combined TEF-1α and ITS sequences revealed diverse phylogenetic affinities among the FSSC isolates and potentially new phylogenetic clades of FSSC. Soil analysis showed varied carbon content, pH, soil moisture, and salinity, but not nitrogen content, between sampling locations. Regardless of the physico-chemical properties, various Fusarium species were recovered from the mangrove soils. These were likely saprophytes; however, some were well-known plant pathogens and opportunistic human pathogens. Thus, mangrove soils might serve as inoculum sources for plant and human pathogenic Fusarium species. The present study demonstrates the occurrence of various Fusarium species in the extreme environment of mangrove soil, thereby contributing to the knowledge on species diversity in Fusarium.
Using glycosaminoglycan Chemical Exchange Saturation Transfer (gagCEST) magnetic resonance imaging (MRI), this study comparatively evaluated the GAG contents of lumbar intervertebral disks (IVDs) of patients with non-specific low back pain (nsLBP), radiculopathy, and asymptomatic volunteers to elucidate the association of clinical manifestation and compositional correlate. A total of 18 patients (mean age 57.5 ± 22.5 years) with radiculopathy, 16 age-matched patients with chronic nsLBP and 20 age-matched volunteers underwent standard morphologic and compositional gagCEST MRI on a 3T scanner. In all cohorts, GAG contents of lumbar IVDs were determined using gagCEST MRI. An assessment of morphologic IVD degeneration based on the Pfirrmann classification and T2-weighted sequences served as a reference. A linear mixed model adjusted for multiple confounders was used for statistical evaluation. IVDs of patients with nsLBP showed lower gagCEST values than those of volunteers (nsLBP: 1.3% [99% confidence intervals (CI): 1.0; 1.6] vs. volunteers: 1.9% [99% CI: 1.6; 2.2]). Yet, IVDs of patients with radiculopathy (1.8% [99% CI: 1.4; 2.1]) were not different from patients with nsLBP or volunteers. In patients with radiculopathy, IVDs directly adjacent to IVD extrusions demonstrated lower gagCEST values than distant IVDs (adjacent: 0.9% [99% CI: 0.3; 1.5], distant: 2.1% [99% CI: 1.7; 2.5]). Advanced GAG depletion in nsLBP and directly adjacent to IVD extrusions in radiculopathy indicates close interrelatedness of clinical pathology and compositional degeneration.
Cytotoxic flavonoids of Murraya tetramera were investigated in this study. A novel flavonoid and twelve known flavonoids, including seven flavones (1-7), three flavanones (8-10), and three chalcones (11-13) were isolated from the leaves and twigs of Murraya tetramera. Chemical structures were elucidated by NMR combined with MS spectral analysis, and the new compound (6) was confirmed as 3',5'-dihydroxy-5,6,7,4'-tetramethoxyflavone. Furthermore, all the isolated flavonoids were evaluated for their cytotoxicities against murine melanoma cells (B16), and human breast cancer cells (MDA-MB-231) by CCK-8 assay. Among them, compounds 7, 13, and 5 exhibited potent cytotoxic activities against B16 cell lines (IC50 = 3.87, 7.00 and 8.66 μg/mL, respectively). Compounds 5, 13, and 12 displayed potent cytotoxicities against MDA-MB-231 cell lines (IC50 = 3.80, 5.95 and 7.89 μg/mL, respectively). According to the correlation of the structure and activity analysis, 5-hydroxyl and 8-methoxyl substituents of the flavone, 8-methoxyl substituent of the flavanone, and 3',5'-methoxyl substituents of the chalcone could be critical factors of the high cytotoxicity. The results indicated that the active flavonoids have potential to be developed as leading compounds for treating cancers.
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