Two Prenylated Chalcones, 4-Hydroxyderricin, and Xanthoangelol Prevent Postprandial Hyperglycemia by Promoting GLUT4 Translocation via the LKB1/AMPK Signaling Pathway in Skeletal Muscle Cells.

SCOPE: Stimulation of glucose uptake in the skeletal muscle is crucial for the prevention of postprandial hyperglycemia. Insulin and certain polyphenols enhance glucose uptake through the translocation of glucose transporter 4 (GLUT4) in the skeletal muscle. The previous study reports that prenylated chalcones, 4-hydroxyderricin (4-HD), and xanthoangelol (XAG) promote glucose uptake and GLUT4 translocation in L6 myotubes, but their underlying molecular mechanism remains unclear. This study investigates the mechanism in L6 myotubes and confirms antihyperglycemia by 4-HD and XAG. METHODS AND RESULTS: In L6 myotubes, 4-HD and XAG promote glucose uptake and GLUT4 translocation through the activation of adenosine monophosphate-activated protein kinase (AMPK) and liver kinase B1 (LKB1) signaling pathway without activating phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) and Janus kinases (JAKs)/signal transducers and activators of transcriptions (STATs) pathways. Moreover, Compound C, an AMPK-specific inhibitor, as well as siRNA targeting AMPK and LKB1 completely canceled 4-HD and XAG-increased glucose uptake. Consistently, oral administration of 4-HD and XAG to male ICR mice suppresses acute hyperglycemia in an oral glucose tolerance test. CONCLUSION: In conclusion, LKB1/AMPK pathway and subsequent GLUT4 translocation in skeletal muscle cells are involved in Ashitaba chalcone-suppressed acute hyperglycemia.

Synthesis, spectroscopic characterization, and antibacterial activity of chalcone (2E)-1-(3'-aminophenyl)-3-(4-dimethylaminophenyl)-prop-2-en-1-one against multiresistant Staphylococcus aureus carrier of efflux pump mechanisms and ß-lactamase.

BACKGROUND: The bacterium Staphylococcus aureus has stood out for presenting a high adaptability, acquiring resistance to multiple drugs. The search for natural or synthetic compounds with antibacterial properties capable of reversing the resistance of S. aureus is the main challenge to be overcome today. Natural products such as chalcones are substances present in the secondary metabolism of plants, presenting important biological activities such as antitumor, antidiabetic, and antimicrobial activity. OBJECTIVES: In this context, the aim of this work was to synthesize the chalcone (2E)-1-(3'-aminophenyl)-3-(4-dimethylaminophenyl)-prop-2-en-1-one with nomenclature CMADMA, confirm its structure by nuclear magnetic resonance (NMR), and evaluate its antibacterial properties. METHODS: The synthesis methodology used was that of Claisen-Schmidt, and spectroscopic characterization was performed by NMR. For microbiological assays, the broth microdilution methodology was adopted in order to analyze the antibacterial potential of chalcones and to analyze their ability to act as a possible inhibitor of ß-lactamase and efflux pump resistance mechanisms, present in S. aureus strain K4100. RESULTS: The results obtained show that CMADMA does not show direct antibacterial activity, expressing a MIC of ≥1024 µg/mL, or on the enzymatic mechanism of ß-lactamase; however, when associated with ethidium bromide in efflux pump inhibition assays, CMADMA showed promising activity by reducing the MIC of the bromide from 64 to 32 µg/mL. CONCLUSION: We conclude that the chalcone synthesized in this study is a promising substance to combat bacterial resistance, possibly acting in the inhibition of the QacC efflux pump present in S. aureus strain K4100, as evidenced by the reduction in the MIC of ethidium bromide.

Chalcone T4 Inhibits RANKL-Induced Osteoclastogenesis and Stimulates Osteogenesis In Vitro.

Chalcones are phenolic compounds produced during the biosynthesis of flavonoids that have numerous biological activities, including anti-inflammatory, antioxidant and anticancer. In this in vitro study, we investigate a newly synthesized chalcone (Chalcone T4) in the context of bone turnover, specifically on the modulation of osteoclast differentiation and activity and osteoblast differentiation. Murine macrophages (RAW 264.7) and pre-osteoblasts (MC3T3-E1) were used as models of osteoclasts and osteoblasts, respectively. Differentiation and activity osteoclasts were induced by RANKL in the presence and absence of non-cytotoxic concentrations of Chalcone T4, added in different periods during osteoclastogenesis. Osteoclast differentiation and activity were assessed by actin ring formation and resorption pit assay, respectively. Expression of osteoclast-specific markers (Nfatc1, Oscar, Acp5, Mmp-9 and Ctsk) was determined by RT-qPCR, and the activation status of relevant intracellular signaling pathways (MAPK, AKT and NF-kB) by Western blot. Osteoblast differentiation and activity was induced by osteogenic culture medium in the presence and absence of the same concentrations of Chalcone T4. Outcomes assessed were the formation of mineralization nodules via alizarin red staining and the expression of osteoblast-related genes (Alp e Runx2) by RT-qPCR. Chalcone T4 reduced RANKL-induced osteoclast differentiation and activity, suppressed Oscar, Acp5 and Mmp-9 expression, and decreased ERK and AKT activation in a dose-dependent manner. Nfact1 expression and NF-kB phosphorylation were not modulated by the compound. Mineralized matrix formation and the expression of Alp and Runx2 by MC3T3-E1 cells were markedly stimulated by Chalcone T4. Collectively, these results demonstrate that Chalcone T4 inhibits in osteoclast differentiation and activity and stimulates osteogenesis, which indicates a promising therapeutic potential in osteolytic diseases.

Chalcone-inspired rA1 /A2A adenosine receptor ligands: Ring closure as an alternative to a reactive substructure.

Over the past few years, great progress has been made in the development of high-affinity adenosine A1 and/or A2A receptor antagonists-promising agents for the potential treatment of Parkinson's disease. Unfortunately, many of these compounds raise structure-related concerns. The present study investigated the effect of ring closures on the rA1 /A2A affinity of compounds containing a highly reactive α,ß-unsaturated carbonyl system, hence providing insight into the potential of heterocycles to address these concerns. A total of 12 heterocyclic compounds were synthesised and evaluated in silico and in vitro. The test compounds performed well upon qualitative assessment of drug-likeness and were generally found to be free from potentially problematic fragments. Most also showed low/weak cytotoxicity. Results from radioligand binding experiments confirm that heterocycles (particularly 2-substituted 3-cyanopyridines) can replace the promiscuous α,ß-unsaturated ketone functional group without compromising A1 /A2A affinity. Structure-activity relationships highlighted the importance of hydrogen bonds in binding to the receptors of interest. Compounds 3c (rA1 Ki  = 16 nM; rA2A Ki  = 65 nM) and 8a (rA1 Ki  = 102 nM; rA2A Ki  = 37 nM), which both act as A1 antagonists, showed significant dual A1 /A2A affinity and may, therefore, inspire further investigation into heterocycles as potentially safe and potent adenosine receptor antagonists.

Chalcone Scaffolds, Bioprecursors of Flavonoids: Chemistry, Bioactivities, and Pharmacokinetics.

Chalcones are secondary metabolites belonging to the flavonoid (C6-C3-C6 system) family that are ubiquitous in edible and medicinal plants, and they are bioprecursors of plant flavonoids. Chalcones and their natural derivatives are important intermediates of the flavonoid biosynthetic pathway. Plants containing chalcones have been used in traditional medicines since antiquity. Chalcones are basically α,ß-unsaturated ketones that exert great diversity in pharmacological activities such as antioxidant, anticancer, antimicrobial, antiviral, antitubercular, antiplasmodial, antileishmanial, immunosuppressive, anti-inflammatory, and so on. This review provides an insight into the chemistry, biosynthesis, and occurrence of chalcones from natural sources, particularly dietary and medicinal plants. Furthermore, the pharmacological, pharmacokinetics, and toxicological aspects of naturally occurring chalcone derivatives are also discussed herein. In view of having tremendous pharmacological potential, chalcone scaffolds/chalcone derivatives and bioflavonoids after subtle chemical modification could serve as a reliable platform for natural products-based drug discovery toward promising drug lead molecules/drug candidates.

Chalcones identify cTXNPx as a potential antileishmanial drug target.

With current drug treatments failing due to toxicity, low efficacy and resistance; leishmaniasis is a major global health challenge that desperately needs new validated drug targets. Inspired by activity of the natural chalcone 2',6'-dihydroxy-4'-methoxychalcone (DMC), the nitro-analogue, 3-nitro-2',4',6'- trimethoxychalcone (NAT22, 1c) was identified as potent broad spectrum antileishmanial drug lead. Structural modification provided an alkyne containing chemical probe that labelled a protein within the parasite that was confirmed as cytosolic tryparedoxin peroxidase (cTXNPx). Crucially, labelling is observed in both promastigote and intramacrophage amastigote life forms, with no evidence of host macrophage toxicity. Incubation of the chalcone in the parasite leads to ROS accumulation and parasite death. Deletion of cTXNPx, by CRISPR-Cas9, dramatically impacts upon the parasite phenotype and reduces the antileishmanial activity of the chalcone analogue. Molecular docking studies with a homology model of in-silico cTXNPx suggest that the chalcone is able to bind in the putative active site hindering access to the crucial cysteine residue. Collectively, this work identifies cTXNPx as an important target for antileishmanial chalcones.

Docking Studies and Antiproliferative Activities of 6-(3-aryl-2-propenoyl)-2(3H)- benzoxazolone Derivatives as Novel Inhibitors of Phosphatidylinositol 3-Kinase (PI3Kα).

BACKGROUND: Cancer is a life-threatening group of diseases and universally, the second main cause of death. The design and development of new scaffolds targeting selective cancer cells are considered a promising goal for cancer treatment. AIMS AND OBJECTIVE: Chalcone derivatives; 6-(3-aryl-2-propenoyl)-2(3H)-benzoxazolone, were previously prepared and evaluated against the oral cavity squamous cell carcinoma cell line, HSC-2, and were reported to have remarkably high tumor selectivity. The aim of this study was to further investigate the anticancer activities of the chalcone derivatives against human colon cancer cells with a possible elucidation of their mechanism of action. METHODS: Computational studies were conducted to explore the potential interaction of the synthesized molecules with the phosphatidylinositol-4,5-bisphosphate 3-kinaseα (PI3Kα). Biological evaluation of the antiproliferative activities associated with compounds 1-23 was carried out against the colon cancer cell line, HCT116. Lactate Dehydrogenase (LDH) activity was measured to study necrosis, while the caspase-3 activation and DNA measurements were used to evaluate apoptosis in the treated cells. RESULTS: Glide studies against PI3Kα kinase domain demonstrated that the 6-(3-aryl-2-propenoyl)-2(3H)- benzoxazolone scaffold forms H-bond with K802, Y836, E849, V851, N853, Q859, and D933, and it fits the fingerprint of PI3Kα active inhibitors. Biological evaluation of the reported compounds in HCT116 cell line confirmed that the series inhibited PI3Kα activity and induced apoptosis via activation of caspase-3 and reduction of DNA content. CONCLUSION: The recently developed compounds might be employed as lead structures for the design of new antitumor drugs targeting PI3Kα.

Adenosine Receptor Ligands: Coumarin-Chalcone Hybrids as Modulating Agents on the Activity of hARs.

Adenosine receptors (ARs) play an important role in neurological and psychiatric disorders such as Alzheimer's disease, Parkinson's disease, epilepsy and schizophrenia. The different subtypes of ARs and the knowledge on their densities and status are important for understanding the mechanisms underlying the pathogenesis of diseases and for developing new therapeutics. Looking for new scaffolds for selective AR ligands, coumarin-chalcone hybrids were synthesized (compounds 1-8) and screened in radioligand binding (hA1, hA2A and hA3) and adenylyl cyclase (hA2B) assays in order to evaluate their affinity for the four human AR subtypes (hARs). Coumarin-chalcone hybrid has been established as a new scaffold suitable for the development of potent and selective ligands for hA1 or hA3 subtypes. In general, hydroxy-substituted hybrids showed some affinity for the hA1, while the methoxy counterparts were selective for the hA3. The most potent hA1 ligand was compound 7 (Ki = 17.7 µM), whereas compound 4 was the most potent ligand for hA3 (Ki = 2.49 µM). In addition, docking studies with hA1 and hA3 homology models were established to analyze the structure-function relationships. Results showed that the different residues located on the protein binding pocket could play an important role in ligand selectivity.

Antiproliferative Effect of Acridine Chalcone Is Mediated by Induction of Oxidative Stress.

Chalcones are naturally occurring phytochemicals with diverse biological activities including antioxidant, antiproliferative, and anticancer effects. Some studies indicate that the antiproliferative effect of chalcones may be associated with their pro-oxidant effect. In the present study, we evaluated contribution of oxidative stress in the antiproliferative effect of acridine chalcone 1C ((2 E)-3-(acridin-9-yl)-1-(2,6-dimethoxyphenyl)prop-2-en-1-one) in human colorectal HCT116 cells. We demonstrated that chalcone 1C induced oxidative stress via increased reactive oxygen/nitrogen species (ROS/RNS) and superoxide production with a simultaneous weak adaptive activation of the cellular antioxidant defence mechanism. Furthermore, we also showed chalcone-induced mitochondrial dysfunction, DNA damage, and apoptosis induction. Moreover, activation of mitogen activated phosphokinase (MAPK) signalling pathway in 1C-treated cancer cells was also observed. On the other hand, co-treatment of cells with strong antioxidant, N-acetyl cysteine (NAC), significantly attenuated all of the above-mentioned effects of chalcone 1C, that is, decreased oxidant production, prevent mitochondrial dysfunction, DNA damage, and induction of apoptosis, as well as partially preventing the activation of MAPK signalling. Taken together, we documented the role of ROS in the antiproliferative/pro-apoptotic effects of acridine chalcone 1C. Moreover, these data suggest that this chalcone may be useful as a promising anti-cancer agent for treating colon cancer.

Chalcone Based Homodimeric PET Agent, 11C-(Chal)2DEA-Me, for Beta Amyloid Imaging: Synthesis and Bioevaluation.

Homodimeric chalcone based 11C-PET radiotracer, 11C-(Chal)2DEA-Me, was synthesized, and binding affinity toward beta amyloid (Aß) was evaluated. The computational studies revealed multiple binding of the tracer at the recognition sites of Aß fibrils. The bivalent ligand 11C-(Chal)2DEA-Me displayed higher binding affinity compared to the corresponding monomer, 11C-Chal-Me, and classical Aß agents. The radiolabeling yield with carbon-11 was 40-55% (decay corrected) with specific activity of 65-90 GBq/µmol. A significant ( p < 0.0001) improvement in the binding affinity of 11C-(Chal)2DEA-Me with synthetic Aß42 aggregates over the monomer, 11C-Chal-Me, demonstrates the utility of the bivalent approach. The PET imaging and biodistribution data displayed suitable brain pharmacokinetics of both ligands with higher brain uptake in the case of the bivalent ligand. Metabolite analysis of healthy ddY mouse brain homogenates exhibited high stability of the radiotracers in the brain with >93% intact tracer at 30 min post injection. Both chalcone derivatives were fluorescent in nature and demonstrated significant changes in the emission properties after binding with Aß42. The preliminary analysis indicates high potential of 11C-(Chal)2DEA-Me as in vivo Aß42 imaging tracer and highlights the significance of the bivalent approach to achieve a higher biological response for detection of early stages of amyloidosis.

Chalcone-templated Hsp90 inhibitors and their effects on gefitinib resistance in non-small cell lung cancer (NSCLC).

The molecular chaperone Hsp90 has emerged as an attractive cancer therapeutic target due to its role in cellular homeostasis by modulating the stabilization and maturation of many oncogenic proteins. In this study, we designed and synthesized a series of Hsp90 inhibitors that hybridized NVP-AUY992 (2) and PU3 (3) in the chalcone scaffold using a structure-based approach. Our results indicate that compound 1g inhibited the proliferation of gefitinib-resistant non-small cell lung cancer (H1975) cells, downregulated the expression of client proteins of Hsp90 including EGFR, MET, Her2 and Akt, and up-regulated the expression of Hsp70. The compound 1g represents a new class of Hsp90 inhibitors with a chalcone structure. The design, synthesis, and evaluation of 1g are described herein.

Neuroprotective effect of synthetic chalcone derivatives as competitive dual inhibitors against µ-calpain and cathepsin B through the downregulation of tau phosphorylation and insoluble Aß peptide formation.

A series of chalcone derivatives were synthesized and evaluated for their µ-calpain and cathepsin B inhibitory activities. Among the tested chalcone derivatives, two compounds, 7 and 11, showed potent inhibitory activities against µ-calpain and cathepsin B and were selected for further evaluation. Compounds 7 and 11 showed enzyme inhibitory activities at the cellular level and displayed neuroprotective effects against oxidative stress-induced apoptosis in SH-SY5Y cells, a human neuroblastoma cell line. Moreover, compounds 7 and 11 reduced p25 formation, tau phosphorylation and insoluble Aß peptide formation. Enzyme kinetic experiments and docking studies revealed that compounds 7 and 11 competitively inhibited both µ-calpain and cathepsin B enzymes.

Flavokawain A inhibits Cytochrome P450 in in vitro metabolic and inhibitory investigations.

ETHNOPHARMACOLOGICAL RELEVANCE: Flavokawain A, the major chalcone in kava extracts, was served as beverages for informal social occasions and traditional ceremonials in most South Pacific islands. It exhibited strong antiproliferative and apoptotic effects against human prostate and urinary bladder cancer cells. AIM OF THE STUDY: The current study was purposed to investigate the interaction between Flavokawain A and Cytochrome P450, including the inhibitory effects of Flavokawain A on predominant CYP450 isotypes and further clarified the inhibitory mechanism of FKA on CYP450 enzymes. Besides, study about identifying the key CYP450 isotypes responsible for the metabolism of FKA was also performed. MATERIALS AND METHODS: In this study, probe-based assays with rat liver microsome system were used to characterize the inhibitory effects of FKA. Molecular docking study was performed to further explore the binding site of FKA on CYP450 isoforms. In addition, chemical inhibition experiments using specific inhibitors (a-naphthoflavone, quinidine, sulfamethoxazde, ketoconazole, omeprazole) were performed to clarify the individual CYP450 isoform that are responsible for the metabolism of FKA. RESULTS: FKA showed significant inhibition on CYP1A2, CYP2D1, CYP2C6 and CYP3A2 activities with IC50 values of 102.23, 20.39, 69.95, 60.22µmol/L, respectively. The inhibition model was competitive, mixed-inhibition, uncompetitive, and noncompetitive for CYP1A2, CYP2D1, CYP2C6 and CYP3A2 enzymes. Molecular docking study indicated the ligand-binding conformation of FKA in the active site of CYP450 isoforms. The chemical inhibition experiments showed that the metabolic clearance rate of Flavokawain A decreased to 19.84%, 50.38%, and 67.02% of the control in the presence of ketoconazole, sulfamethoxazde and a-naphthoflavone. CONCLUSION: The study showed that Flavokawain A has varying inhibitory effect on CYP450 enzymes and CYP3A2 was the principal CYP isoform contributing to the metabolism of Flavokawain A. Besides, CYP2C6 and CYP1A2 isoforms also play important roles in the metabolism of FKA. Our results provided a basis for better understanding the biotransformation of FKA and prediction of drug-drug interaction of FKA.

In vitro characterization of 4'-(p-toluenesulfonylamide)-4-hydroxychalcone using human liver microsomes and recombinant cytochrome P450s.

1. 4'-(p-Toluenesulfonylamide)-4-hydroxychalcone (TSAHC) is a synthetic sulfonylamino chalcone compound possessing anti-cancer properties. The aim of this study was to elucidate the metabolism of TSAHC in human liver microsomes (HLMs) and to characterize the cytochrome P450 (P450) enzymes that are involved in the metabolism of TSAHC. 2. TSAHC was incubated with HLMs or recombinant P450 isoforms (rP450) in the presence of an nicotinamide adenine dinucleotide phosphate, reduced form (NADPH)-regenerating system. The metabolites were identified and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). P450 isoforms, responsible for TSAHC metabolite formation, were characterized by chemical inhibition and correlation studies in HLMs and enzyme kinetic studies with a panel of rP450 isoforms. 3. Two hydroxyl metabolites, that is M1 and M2, were produced from the human liver microsomal incubations (K(m) and V(max) values were 2.46 µM and 85.1 pmol/min/mg protein for M1 and 9.98 µM and 32.1 pmol/min/mg protein for M2, respectively). The specific P450 isoforms responsible for two hydroxy-TSAHC formations were identified using a combination of chemical inhibition, correlation analysis and metabolism by expressed recombinant P450 isoforms. The known P450 enzyme activities and the rate of TSAHC metabolite formation in the 15 HLMs showed that TSAHC metabolism is correlated with CYP2C and CYP3A activity. The P450 isoform-selective inhibition study in HLMs and the incubation study of cDNA-expressed enzymes also showed that two hydroxyl metabolites M1 and M2 biotransformed from TSAHC are mainly mediated by CYP2C and CYP3A, respectively. These findings suggest that CYP2C8, CYP2C9, CYP2C19, CYP3A4 and CYP3A5 isoforms are major enzymes contributing to TSAHC metabolism.

Novel carbapenem chalcone derivatives: synthesis, cytotoxicity and molecular docking studies.

A one-pot efficient synthetic protocol is described for the synthesis of carbapenem chalcone derivatives using AAPTMS@MCM-41 heterogeneous catalyst. Various substituted aromatic aldehydes were attached to highly chiral and reactive carbapenem using this approach. The cytotoxic activity evaluation of all synthesized compounds was performed against lung cancer cell lines (A-549) and breast cancer cell lines (MCF-7) using the MTT assay. Among the tested compounds, compound CPC-2 showed better activity against MCF-7 cell lines with an IC50 value 2.52 µM mL(-1); whereas compound CPC-4 showed good activity against A-549 cell lines with an IC50 value 1.59 µM mL(-1). In order to support the observed activity profiles, the representative compounds were flexibly docked into the active sites of the Anaplastic Lymphoma Kinase (ALK) enzyme and the Estrogen receptor (ERß). The most active anticancer compounds exhibited stronger binding affinities for proteins.

Synthesis of phenstatin/isocombretastatin-chalcone conjugates as potent tubulin polymerization inhibitors and mitochondrial apoptotic inducers.

A series of phenstatin/isocombretastatin­chalcones were synthesized and screened for their cytotoxic activity against various human cancer cell lines. Some representative compounds exhibited significant antiproliferative activity against a panel of sixty human cancer cell lines of the NCI, with GI50 values in the range of 0.11 to 19.0 µM. Three compounds (3b, 3c and 3e) showed a broad spectrum of antiproliferative efficacy on most of the cell lines in the sub-micromolar range. In addition, all the synthesized compounds (3a­l and 4a­l) displayed moderate to excellent cytotoxicity against breast cancer cells such as MCF-7 and MDA-MB-231 with IC50 values in the range of 0.5 to 19.9 µM. Moreover, the tubulin polymerization assay and immunofluorescence analysis results suggest that some of these compounds like 3c and 3e exhibited significant inhibitory effect on the tubulin assembly with an IC50 value of 0.8 µM and 0.6 µM respectively. A competitive binding assay suggested that these compounds bind at the colchicine-binding site of tubulin. A cell cycle assay revealed that these compounds arrest at the G2/M phase and lead to apoptotic cell death. Furthermore, this was confirmed by Hoechst 33258 staining, activation of caspase 9, DNA fragmentation, Annexin V-FITC and mitochondrial membrane depolarization. Molecular docking studies indicated that compounds like 3e occupy the colchicine binding site of tubulin.

Synthesis, biological evaluation and molecular docking of novel chalcone-coumarin hybrids as anticancer and antimalarial agents.

A new series of chalcone-coumarin derivatives (9-19) linked by the 1,2,3-triazole ring were synthesized through the azide/alkyne dipolar cycloaddition. Hybrid molecules were evaluated for their cytotoxic activity against four cancer cell lines (e.g., HuCCA-1, HepG2, A549 and MOLT-3) and antimalarial activity toward Plasmodium falciparum. Most of the synthesized hybrids, except for analogs 10 and 16, displayed cytotoxicity against MOLT-3 cell line without affecting normal cells. Analogs (10, 11, 16 and 18) exhibited higher inhibitory efficacy than the control drug, etoposide, in HepG2 cells. Significantly, the high cytotoxic potency of the hybrid 11 was shown to be non-toxic to normal cells. Interestingly, the chalcone-coumarin 18 was the most potent antimalarial compound affording IC50 value of 1.60 µM. Molecular docking suggested that the cytotoxicity of reported hybrids could be possibly due to their dual inhibition of α- and ß-tubulins at GTP and colchicine binding sites, respectively. Furthermore, falcipain-2 was identified to be a plausible target site of the hybrids given their antimalarial potency.

[C-ring cleavage of liquiritigenin extracted from licorice roots by an oxygen-tolerant bovine rumen bacterium strain Aeroto-Niu-O16].

Aeroto-Niu-O16, an oxygen-tolerant bovine rumen bacterium, is capable of aerobically reducing isoflavones daidzein and genistein to dihydrodaidzein and dihydrogenistein through catalytic hydrogenation. In this study, it was found that bacterium strain Aeroto-Niu-O16 was able to cleavage the C-ring of liquiritigenin (LG), which is one of the main biologically active components of licorice roots, in the presence of atmospheric oxygen. LG was prepared by acid hydrolysis of the crude extract of licorice roots. The metabolite of LG obtained in strain Aeroto-Niu-O16 was identified as davidigenin (DG) based on the data of UV, MS, 1H and 13C NMR. The maximal concentration of LG that the strain Aeroto-Niu-O16 was able to transform effectively was 0.8 mmol x L(-1) and the average productivity of the metabolite DG was 71.7%. Furthermore, when 0.1% (m/v) of L-cysteine or sodium thiosulfate was added in the cultural medium, the average bioconversion rate of LG was increased from 71.7% to 78.3% and 77.2%, respectively. The in vitro antioxidant investigation showed that 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity of DG was significantly or extremely significantly higher than that of LG at the concentrations from 0.2 mmol x L(-1) to 1.6 mmol x L(-1). We discoverd for the first time that LG can be converted to DG, which has stronger and wider biological activities, through microbial biotransformation method.

Effects of novel chalcone derivatives on α-glucosidase, dipeptidyl peptidase-4, and adipocyte differentiation in vitro.

Chana series are new chalcone derivatives. To evaluate the possibility of Chana series as therapeutic agents of type 2 diabetes, the inhibitory effects of Chana series on the activities of α-glucosidase and DPP-4 were investigated using in vitro enzyme assays, and their effects on adipocyte differentiation were investigated in C3H10T1/2 cells. Chana 1 and Chana 7 among the Chana series showed significant inhibition of α-glucosidase activity. In DPP-4 enzyme assay, Chana 1 exhibited the highest inhibitory activity while Chana 7 did not. In MTT assay, Chana 1 did not show significant cytotoxicity up to a concentration of 250 µM, whereas cytotoxicity was observed with Chana 7 at a concentration of 300 µM. In addition, Chana 1 induced adipocyte differentiation. Therefore, Chana 1 showed inhibitory effects on α-glucosidase and DPP-4 as well as a stimulatory effect on adipocyte differentiation, suggesting that Chana 1 may be a potential beneficial agent for the treatment of type 2 diabetes.

Simultaneous post-transcriptional gene silencing of two different chalcone synthase genes resulting in pure white flowers in the octoploid dahlia.

Garden dahlias (Dahlia variabilis) are autoallooctoploids with redundant genes producing wide color variations in flowers. There are no pure white dahlia cultivars, despite its long breeding history. However, the white areas of bicolor flower petals appear to be pure white. The objective of this experiment was to elucidate the mechanism by which the pure white color is expressed in the petals of some bicolor cultivars. A pigment analysis showed that no flavonoid derivatives were detected in the white areas of petals in a star-type cultivar 'Yuino' and the two seedling cultivars 'OriW1' and 'OriW2' borne from a red-white bicolor cultivar, 'Orihime', indicating that their white areas are pure white. Semi-quantitative RT-PCR showed that in the pure white areas, transcripts of two chalcone synthases (CHS), DvCHS1 and DvCHS2 which share 69% nucleotide similarity with each other, were barely detected. Premature mRNA of DvCHS1 and DvCHS2 were detected, indicating that these two CHS genes are silenced post-transcriptionally. RNA gel blot analysis revealed that small interfering RNAs (siRNAs) derived from CHSs were produced in these pure white areas. By high-throughput sequence analysis of small RNAs in the pure white areas with no mismatch acceptance, small RNAs were mapped to two alleles of DvCHS1 and two alleles of DvCHS2 expressed in 'Yuino' petals. Therefore, we concluded that simultaneous siRNA-mediated post-transcriptional gene silencing of redundant CHS genes results in the appearance of pure white color in dahlias.