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The synthesis of several new lipoidal dicarborane compounds prepared by reaction of decaborane with several fatty mono- and bis-alkyl-1,3-diols is described.

We present a method for the quantitative ESR analysis of the antioxidant properties of drugs using the acetaldhehydelxanthine oxidase (ACIXOD) superoxide generating system and 5,5-dimethyl-l -pyrroline-N-oxide (DMPO) as spin trap. In stoichiometric conditions (ACIXOD, 60 mM l 0.01 8 U), the resulting paramagnetic DMPO adduct disappeared with superoxide dismutase and remained when catalase

Un code de simulation Monte-Carlo est developpe pour etudier les modalites d'action de la Therapie par Capture de Neutrons (TCN) sur le 10 B. L'objectif est le calcul de la probabilite de depot d'une dose letale dans les noyaux cellulaires. Les membranes cytoplasmique et nucleaire sont schematisees par des ellipsoides non concentriques dont tous les parametres sont ajustables a des configurations reelles. Les reactions considerees, 10 B(n,γα) 7 Li et 14 N(n,p) 14 C, produisent des ions dont les trajectoires peuvent etre simulees en considerant les fluctuations sur les longueurs de parcours. Les contributions respectives de chaque reaction aux doses deposees dans les divers compartiments cellulaires peuvent etre etudiees et analysees en fonction des distributions de 10 B.

International audience; The polynuclear manganese(II) complex of L-dihydroorotic acid has been synthesized and characterized by X-ray crystallography, UV-visible spectroscopy, and magnetic susceptibility measurements. Crystal data: triclinic, space group P1 bar , a = 4.7239(5) , b = 7.902(1) , c = 10.552(2) , = 105.80(1), = 98.864(7), = 104.69(1), V = 355.9(1) D 3, Z = 1, R = 0.035, Rw = 0.028. This complex consists of polymer chains of manganese atoms that are bound together via carboxylate bridges with a Mn . . .Mn distance in the chain of 4.724 D . The radical scavenging properties of the complex have been evaluated by ESR spectroscopy and cyclic voltammetry methods and compared to those of vitamin E and BHT (2,6-di-tert-butyl-4-methylphenol).Key words: orotic acid, L-dihydroorotic acid, manganese(II) complex, antioxidative properties.; Le complexe du manganèse(II) et de l'acide L-dihydroorotique a été synthétisé et caractérisé par spectroscopie UV-visible, susceptibilité magnétique et par diffraction des rayons X. Données cristallographiques : triclinique, groupe d'espace P 1 bar , a = 4.7239(5) D , b = 7.902(1) D , c = 10.552(2) D , α = 105.80(1)°, β = 98.864(7)°, γ~= 104.69(1)°, V = 355.9(1) D 3, Z = 1, R = 0.035, Rw = 0.028. Ce complexe se présente comme une chaine polymérique d'atomes de manganèse liés entre eux par des ponts carboxylate, la distance Mn . . .Mn étant de 4.724 D . Les propriétés antioxydantes de ce composé ont été évaluées par RPE et voltammétrie cyclique et comparées à celles de la vitamine E et du BHT (2,6-di-tert-butyl-4-methylphenol).Mots clés : acide orotique, acide L-dihydroorotique, complexe du manganèse(II), propriétés antioxydantes.

Deux protocoles de fixation d'un radioelement (M = 111 In, 153 Gd) sur les LDL par l'intermediaire d'un agent complexant lipophile (L) sont compares. Trois analogues lipophiles de l'EDTA ou du DTPA derives de la stearylamine ou de la phosphatidylethanolamine ont ete utilises. Lors du protocole A, le conjugue L-LDL est d'abord prepare puis marque avec le radioelement M; lors du protocole B, le complexe M-L est prepare en premier puis insere dans les LDL. Le traceur final est la particule M-L-LDL. Par precipitation du radiometal non lie (addition de tropolone) ou des particules M-L-LDL (addition d'heparine), le comptage radioactif montre que 55 a 92% de la quantite initiale de radioelement demeure fixee aux particules M-L-LDL avec le protocole A, et 55 a 85% avec le protocole B. Ces deux protocoles peuvent etre utilises pour radiomarquer les LDL.

There is an urgent need for new antimalarial drugs with novel mechanisms of action on novel targets. Indolone- N -oxides (INODs) display antimalarial properties in vitro and in vivo , but identified leads such as 6-(4-chloro-phenyl)-5-oxy-[1,3]dioxolo[4,5- f ]indol-7-one 1 , suffer from very poor aqueous solubility. In this study, structural modifications have been made by introducing various amino and bulky groups to produce sufficiently water soluble and active compounds for further pharmacological and pharmacokinetic studies. We report here the preparation of twelve novel amino derivatives and their antiplasmodial activities including those of two other structurally known compounds. The 5-methoxy-2-(4-morpholin-4-yl-phenyl)-1-oxy-indol-3-one, 9 , has the highest antiplasmodial activity in vitro (IC 50 = 6.5 nM; FcB1 strain) and selectivity index (SI (CC 50 MCF7/IC 50 FcB1) = 4538.5). The 6-amino-2-(4-chloro-phenyl)-1-oxy-indol-3-one, 14 , (IC 50 = 183 nM; SI = 60), is an excellent candidate for further mechanistic studies. Indeed, this is structurally the closest analogue to the current lead, 1 , bearing an NH 2 group at R 2 offering possibilities for functionalization and labeling.

Uncatalyzed racemization processes in atropisomeric diphenyl-like frameworks are classically described as the result of the rotation around the pivotal single bond linking two planar frameworks. Severe constraints leading to more or less distorted transition states account for the experimental barrier to atropenantiomerization. In 1988, one of us hypothesized that, in N-aryl-2(1H)-pyrimidin-(thi)ones, a ring-opening/ring-closure process was contributing to the observed racemization process accounting for the lower barriers in the sulfur analogues than in oxygen analogues. Now, a series of six novel 6-amino-5-cyano-1,4-disubstituted-2(1H)-pyrimidinones 5a-5f and two 6-amino-5-cyano-4-p-tolyl-1-substituted-2(1H)-pyrimidinethiones 6a and 6b were synthesized and characterized through spectroscopic and X-ray diffraction studies. Semipreparative HPLC chiral separation was achieved, and enantiomerization barriers were obtained by thermal racemization. The rotational barriers of 6-amino-5-cyano-1-o-tolyl-4-p-tolyl-2(1H)-pyrimidinone (5b) and 6-amino-5-cyano-1-(naphthalen-1-yl)-4-p-tolyl-2(1H)-pyrimidinone (5e) were found to be 120.4 and 125.1 kJ·mol-1 (n-BuOH, 117 C), respectively, and those of the corresponding thiones were 116.8 and 109.6 kJ·mol -1 (EtOH, 78 C), respectively. DFT calculations of the rotational barriers clearly ruled out the classical rotation around the pivotal bond with distorted transition states in the case of the sulfur derivatives. Instead, the ranking of the experimental barriers (sulfur versus oxygen, and o-tolyl versus 1-naphthyl in both series) was nicely reproduced by calculations when the rotation occurred via a ring-opened form in N-aryl-2(1H)-pyrimidinethiones. © 2013 American Chemical Society.

International audience; Targeting the redox metabolism of Plasmodium falciparum to create a fatal overload of oxidative stress is a route to explore the discovery of new antimalarial drugs. There are three main possibilities to target the redox metabolism of P. falciparum at the erythrocytic stage: selective targeting and inhibition of a redox P. falciparum protein or enzyme; oxidant drugs targeting essential parasite components and heme by-products; and redox cycler drugs targeting the parasitized red blood cell. Oxidants and redox cycler agents, with or without specific targets, may disrupt the fragile parasitized erythrocyte redox-dependent architecture given that: redox equilibrium plays a vital role at the erythrocytic stage; P. falciparum possesses major NADPH-dependent redox systems, such as glutathione and thioredoxin ones; and the protein–NADPH-dependent phosphorylation–dephosphorylation process is involved in building new permeation pathways and channels for the nutrient–waste import–export traffic of the parasite.

International audience; Circulating blood monocytes belong to the first line of defense against pathogens and inflammation. Monocytes can be divided into three populations defined by the expression of the cell surface molecules, CD 14 and CD 16. The CD 14(++) CD 16(-) cells, called "classical" monocytes, represent 85% to 95% of the total monocytes in a healthy person whereas CD 14(-) CD 16(+), called "proinflammatory" monocytes, are found in greater numbers in the blood of patients with acute inflammation and infectious diseases. This increase in the concentration of proinflammatory monocytes can be a good indicator of an infectious state. This study presents an immunosensor based on impedance detection for specific cell trapping of classical and proinflammatory monocytes. The grafting of specific antibodies (CD 14 or CD 16) was based on the use of mixed SAM associated with protein G. Each step of the functionalization was characterized by electrochemical methods, quartz crystal microbalance and atomic force microscopy. Faradaic electrochemical impedance spectroscopy and voltametric analysis confirmed the success of the modification process with a surface coverage reaching 92% for the antibody layer. The increase in the deposited mass at each step of the modification process confirmed this results revealing that one protein G in two was bound to an antibody. The cell trapping capacity, evaluated by the variation in the film resistance using non-faradaic impedance spectroscopy revealed that the cell trapping is selective, depending on the specific antibody grafted and quantitative with the range of detection being 1000 to 30,000 infected cells. This range of detection is consistent with the application targeted.

International audience; Amyloid plaques are a hallmark in the brain of Alzheimer’s disease (AD) victims. These plaques consist mainly of an aggregated peptide dubbed amyloid‐β (Aβ), which is also present in healthy brains in a soluble form. It is thought that soluble oligomeric forms of Aβ are the most toxic species, rather than more aggregated fibrils or protofibrils. The presence of oxidative damage on neuronal lipids and proteins is evidence of a link between oxidative stress and AD. Redox‐active copper ions are highly accumulated in amyloid plaques, where they bind to Aβ. Cu‐Aβ complexes are able to catalyze the production of reactive oxygen species (ROS), such as H2O2 and HO., in vitro. These reactions might contribute to the oxidative damage of diverse biomolecules observed in AD, including the peptide Aβ itself, which has been found to be oxidized in amyloid plaques in vivo.

Ethnopharmacological relevance Crinum latifolium L. (CL) leaf extracts have been traditionally used in Vietnam and are now used all over the world for the treatment of prostate cancer. However, the precise cellular mechanisms of the action of CL extracts remain unclear. Aim of the study To examine the effects of CL samples on the anti-tumour activity of peritoneal murine macrophages. Materials and methods The properties of three extracts (aqueous, flavonoid, alkaloid), one fraction (alkaloid), and one pure compound (6-hydroxycrinamidine) obtained from CL, were studied (i) for redox capacities (DPPH and bleaching beta-carotene assays), (ii) on murine peritoneal macrophages (MTT assay) and on lymphoma EL4-luc2 cells (luciferine assay) for cytotoxicity, (iii) on macrophage polarization (production of ROS and gene expression by PCR), and (iv) on the tumoricidal functions of murine peritoneal macrophages (lymphoma cytotoxicity by co-culture with syngeneic macrophages). Results The total flavonoid extract with a high antioxidant activity (IC 50 =107.36 mg/L, DPPH assay) showed an inhibitory action on cancer cells. Alkaloid extracts inhibited the proliferation of lymphoma cells either by directly acting on tumour cells or by activating of the tumoricidal functions of syngeneic macrophages. The aqueous extract induced mRNA expression of tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin 6 (IL-6) indicating differentiation of macrophages into pro-inflammatory M1 polarized macrophages. The total flavonoid, alkaloid extracts and an alkaloid fraction induced the expression of the formyl peptide receptor (FPR) on the surface of the polarized macrophages that could lead to the activation of macrophages towards the M1 phenotype. Aqueous and flavonoid extracts enhanced NADPH quinine oxido-reductase 1 (NQO1) mRNA expression in polarized macrophages which could play an important role in cancer chemoprevention. All the samples studied were non-toxic to normal living cells and the pure alkaloid tested, 6-hydroxycrinamidine, was not active in any of the models investigated. Conclusions Our results indicate that CL extracts and alkaloid fraction (but not pure 6-hydroxycrinamidine) inhibit the proliferation of lymphoma cells in multiple pathways. Our results are in accordance with traditional usage and encourage further studies and in vivo assays.

International audience; Indolone-N-oxides (INODs) are bioreducible and possess remarkable anti-malarial activities in the low nanomolar range in vitro against different Plasmodium falciparum (P. falciparum) strains and in vivo. INODs have an original mechanism of action: they damage the host cell membrane without affecting non-parasitized erythrocytes. These molecules produce a redox signal which activates SYK tyrosine kinases and induces a hyperphosphorylation of AE1 (band 3, erythrocyte membrane protein). The present work aimed to understand the early stages of the biochemical interactions of these compounds with some erythrocyte components from which the redox signal could originate. The interactions were studied in a biomimetic model and compared with those of chloroquine and artemisinin. The results showed that INODs i) do not enter the coordination sphere of the metal in the heme iron complex as does chloroquine; ii) do not generate iron-dependent radicals as does artemisinin; iii) generate stable free radical adducts after reduction at one electron; iv) cannot trap free radicals after reduction. These results confirm that the bioactivity of INODs does not lie in their spin-trapping properties but rather in their pro-oxidant character. This property may be the initiator of the redox signal which activates SYK tyrosine kinases.

Two lipophilic chelating agents (L) prepared by reaction of dipalmitoylphosphatidylethanolamine with the bis(anhydride) form of ethylenediaminetetraacetic acid (EDTA) or diethylenetriaminepentaacetic acid (DTPA) were characterized. L, indium-L or gadolinium-L complexes are soluble in buffered solutions and may be used for metal labeling of LDL.

L'anion superoxyde est implique dans la genese de plusieurs pathologies comme l'ischemie, l'atherosclerose et l'inflammation des lors que sa formation n'est plus regulee par les systemes moleculaires ou enzymatiques antioxydants. Dans le but de rechercher des composes mimetiques de la superoxyde dismutase (SOD), une methode utilisant la Resonance de Spin Electronique (RSE) a ete mise au point pour quantifier l'activite antioxydante de composes du manganese(Il). Le systeme acetaldehyde/xanthine oxydase genere l'anion superoxyde et le 5,5-dimethyl-1-pyroline-N-oxyde (DMPO) est utilise comme piegeur de spin. Le pouvoir complexant du tampon phosphate, habituellement utilise, vis-a-vis de l'ion Mn 2+ ne pouvant etre neglige comparativement a celui des ligands etudies, les activites des ligands libres et des complexes sont comparees en fonction de la nature du systeme tampon, phosphate ou organique.

In order to use the LDL receptor pathway to target radionuclides to cancer sites for imaging and diagnostic purposes, a labeling procedure of LDL with 111In using the DTPA-bis(stearylamide) (L) has been developed. This bifunctional ligand is intended to be incorporated into the phospholipid monolayer of LDL and to specifically chelate the In3+ cation at the surface. The ligand was incorporated into LDL in buffered medium with a 65-80% yield. The L-LDL samples are stable over a 24 h period when examined by dialysis, allowing their storage before indium-111 radiolabeling. In vitro studies of In-L-LDL particles show that indium labeling is rapidly achieved (1 h). More than 85% of the indium atoms are bound to the chelating functions of the incorporated DTPA derivatives and less than 10% to the nonspecific complexation sites of LDL (e.g., protein residues). After incubation in human serum, the indium activity recovered in the LDL fraction of In-L-LDL samples (95%) is much higher than in In-LDL samples (35%), pointing out the strong stabilizing chelating effect of the ligand. Competitive binding studies show that In-L-LDL are recognized by LDL receptors of A549 cells like native LDL when the In-L/LDL ratio varies from 5 to 30. All these in vitro experiments demonstrate that the In-L-LDL conjugates possess properties suitable for further work with in vivo experiments.

Afin de cibler des tumeurs par des radiotraceurs pour des applications de diagnostic humain en utilisant la voie du recepteur aux lipoproteines de basse densite (LDL), les LDL ont ete marquees a l'indium-111 par l'intermediaire du bis(stearylamide) de l'acide diethylene-triaminepentaacetique (DTPA), L. Les etudes in vitro ayant demontre la stabilite des particules 111 In-L-LDL en milieu plasmatique et leur reconnais-sance par le recepteur aux LDL, des experiences ont ete menees in vivo. Lors de premiers essais chez la souris saine, les images scintigraphiques enregistrees 24 h apres injection intra-veineuse d' 111 In-citrate ou de 111 In-L-LDL montrent une difference de biodistribution suggerant que les particules 111 In-L-LDL suivent le metabolisme des LDL. Lors d'experiences chez la souris porteuse du Melanome B16 depuis 22 jours, la visualisation des tumeurs 24 h apres l'injection de 111 In-L-LDL a ete obtenue pour une quantite de proteines injectees de 400 μg et un rapport In-L/LDL variant entre 3/10 et 3/100. Ces premieres experiences in vivo mettent en evidence l'importance de ces deux parametres pour visualiser la tumeur.

A series of bis -indolone- N -oxides, 1a–f , was prepared from bis (ethynyl)benzenes and o -halonitroaryls and studied for their in vitro antiplasmodial activities against Plasmodium falciparum and representative strains of bacteria and candida as well as for their cytotoxicity against a human tumor cell line (MCF7). They did not cause any haemolysis (300 μg mL −1 ). Of the synthesized bis -indolones, compound 1a had the most potent antiplasmodial activity (IC 50 = 0.763 μmol L −1 on the FcB1 strain) with a selectivity index (CC 50 MCF7/IC 50 FcB1) of 35.6. No potency against the tested microbial strains was observed.

Indolone-N-oxide derivatives possess interesting biological properties. The analysis of these compounds using mass spectrometry (MS) may lead to interference or under-estimation due to the tendency of the N-oxides to lose oxygen. All the previous works focused only on the temperature of the heated parts (vaporizer and ion-transfer tube) of the mass spectrometer without investigating other parameters. This work is extended to the investigation of other parameters.The behavior of N-oxides during atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI) has been investigated using MS(n) ion trap mass spectrometry. Different parameters were investigated to clarify the factors implicated in the deoxygenation process. The investigated parameters were vaporizer temperature (APCI), ion-transfer tube temperature, solvent type, and the flow rates of the sheath gas, auxiliary gas, sweep gas and mobile phase.The deoxygenation increased when the vaporizer temperature increased. The extent of the 'thermally' induced deoxygenation was inversely proportional to the ion-transfer tube temperature and auxiliary gas flow rate and in direct proportion to the mobile phase flow rate. Deoxygenation was not detected under MS/MS fragmentation and hence it is a non-collision-induced dissociation. N-Oxides have the tendency to form abundant 'non-classical' dimers under ESI, which fragment via dehydration rather than giving their corresponding monomer.Deoxygenation is not solely a 'classical' thermal process but it is a thermal process that is solvent-mediated in the source. Deoxygenation was maximal with an APCI source while dimerization was predominant with an ESI source. Therefore, attention should be paid to these molecular changes in the mass spectrometer as well as to the choice of the ionization mode for N-oxides.

International audience; An alarming increase in microbial resistance to traditional drugs and classical pharmacophores has spurred the search for new antimicrobial compounds. Indolone-N-oxides (INODs) possess a redox pharmacophore with promising, recently established, antimalarial activities. In this study, the anti-infectious properties of a series of INODs were investigated. The antibacterial activity was evaluated against five bacterial strains Gram-positive (Staphylococcus aureus, Enterococcus hirae), Gram-negative (Pseudomonas aeruginosa, Escherichia coli) and acid-fast (Mycobacterium tuberculosis). The antifungal activity was assessed using two fungal strains (Aspergillus niger, Candida albicans). The antileishmanial activity was tested against two leishmanial strains, axenically-cultured amastigote (Leishmania infantum, Leishmania amazonensis). The pharmacological activities are discussed as a function of structural and lipophilic characteristics. The Gram-positive bacterial strain E. hirae was found to be the most sensitive strain, whereas the Gram-negative E. coli was resistant to this family of compounds. One compound (64) was more potent than nalidixic acid against E. hirae, whereas another one (52) was equipotent as clotrimazole against C. albicans. INODs were microbe -cidal rather than -static. INODs showed good antitubercular activity in the low micromolar range (similar to ciprofloxacin). In addition, INOD-antiprotozoal potencies were confirmed against the leishmania parasite. INODs showed a broad spectrum of antimicrobial activity and offer a promising anti-infectious prototype worthy of being developed.

Dans de nombreuses conditions pathologiques les systemes de defense de l'organisme vis-a-vis d'un stress oxydant ne sont pas suffisants pour lutter contre les lesions induites par la formation des radicaux oxygenes. Il est donc necessaire de disposer de molecules interceptant les radicaux pour developper des therapies antioxidantes. Afin d'evaluer le pouvoir antioxydant de differentes substances, la Resonance Paramagnetique Electronique associee a la technique de piegeage de spin a ete utilisee pour developper des tests in vitro

NRH:quinone oxidoreductase 2 (QR2) is a cytosolic enzyme that catalyzes the reduction of quinones, such as menadione and co-enzymes Q. With the aim of understanding better the mechanisms of action of QR2, we approached this enzyme catalysis via electron paramagnetic resonance (EPR) measurements of the by-products of the QR2 redox cycle. The variation in the production of oxidative species such as H(2)O(2), and subsequent hydroxyl radical generation, was measured during the course of QR2 activity under aerobic conditions and using pure human enzyme. The effects on the activity of the following were compared: (i) synthetic (N-benzyldihydronicotinamide, BNAH) or natural (nicotinamide riboside, NRH) co-substrates; (ii) synthetic (menadione) or natural (co-enzyme Q0, Q2) substrates; (iii) QR2 modulators and inhibitors (melatonin, resveratrol and S29434); (iv) a pro-drug activated via a redox cycle [CB1954, 5-(aziridin-1-yl)-2,4-dinitrobenzamide]. The results were also compared with those obtained with human QR1. The production of hydroxyl radicals is: (i) observed whatever the substrate/co-substrate used; ii) quenched by adding catalase; (iii) not observed with the specific QR2 inhibitor S29434; (iv) observed with the pro-drug CB1954. While QR2 produced free radicals with this pro-drug, QR1 gave no EPR signal showing the strong reducing capacity of QR2. In conclusion, EPR analysis of QR2 enzyme activity through free radical production enables modulators and effective inhibitors to be distinguished.

The binding affinity of human serum albumin (HSA) to three antimalarial indolone-N-oxide derivatives, INODs, was investigated under simulated physiological conditions using fluorescence spectroscopy in combination with UV-vis absorption and circular dichroism (CD) spectroscopy. Analysis of fluorescence quenching data of HSA by these compounds at different temperatures using Stern-Volmer and Lineweaver-Burk methods revealed the formation of a ground state indolone-HSA complex with binding affinities of the order 10(4) M(-1). The thermodynamic parameters ΔG, ΔH, and ΔS, calculated at different temperatures, indicated that the binding reaction was endothermic and hydrophobic interactions play a major role in this association. The conformational changes of HSA were investigated qualitatively using synchronous fluorescence and quantitatively using CD. Site marker competitive experiments showed that the binding process took place primarily at site I (subdomain IIA) of HSA. The number of binding sites and the apparent binding constants were also studied in the presence of different ions.

International audience; The physiological changes caused by external stimuli can be employed as parameters to study pathogen infection in cells and the effect of drugs. Among analytical methods, impedance is potentially useful to give insight into cellular behavior by studying morphological changes, alterations in the physiological state, production of charged or redox species without interfering with in vitro cellular metabolism and labeling. The present work describes the use of electrochemical impedances spectroscopy to simply monitor by modeling impedance plots (Nyquist diagram) in appropriate equivalent circuit, the changes affecting murine macrophage cell line (RAW 264.7) in response to parasite infection by Leishmania amazonensis or to lipopolysaccharide (LPS) treatment. These results demonstrate the ability of electrochemical impedance spectroscopy to discriminate between two opposite cell responses associated to two different stimuli, one caused by the internalization of a parasite, and the other by activation by a bacterium component. Indeed, the study has allowed the characterization, from an electrical point of view, of the extra-cellular NO radical produced endogenously and in great quantities by the inducible form of NO-synthase in the case of LPS-stimulatedmacrophages. This production was not observed in the case of Leishmania-infectedmacrophages for which to survive and multiply, the parasite itself possesses mechanisms which may interfere with NO production. In this latest case, only the intracellular production of ROS was observed. To confirm these interpretations confocal microscopy analysis using the ROS (reactive oxygen species) fluorescent probe 2′,7′-dichlorodihydrofluorescein diacetate and electron paramagnetic resonance experiments using Fe(DETC)2 as NO radical spin trap were carried out.

The synthesis of several new lipoidal dicarborane compounds prepared by reaction of decaborane with several fatty mono- and bis-alkyl-1,3-diols is described.

Two amphiphilic cheating agents have been synthesized from diethylenetriaminepentaacetic acid (DTPA). The dialkylamine conjugates of DTPA present a symmetrical structure with two lipophilic long chains.

International audience; The malaria parasite, Plasmodium falciparum, invades human erythrocytes and induces dramatic changes in the host cell. The idea of this work was to use RBC modified electrode to perform electrochemical impedance spectroscopy (EIS) with the aim of monitoring physiological changes affecting the erythrocyte after invasion by the malaria parasite. Impedance cell-based devices are potentially useful to give insight into cellular behavior and to detect morphological changes. The modelling of impedance plots (Nyquist diagram) in equivalent circuit taking into account the presence of the cellular layer, allowed us pointing out specific events associated with the development of the parasite such as (i) strong changes in the host cell cytoplasm illustrated by changes in the film capacity, (ii) perturbation of the ionic composition of the host cell illustrated by changes in the film resistance, (iii) releasing of reducer (lactic acid or heme) and an enhanced oxygen consumption characterized by changes in the charge transfer resistance and in the Warburg coefficient characteristic of the redox species diffusion. These results show that the RBC-based device may help to analyze strategic events in the malaria parasite development constituting a new tool in antimalarial research.

Soy germ is a remarkable source of bioactive phytochemicals offering an interesting alternative as starting ingredient for fermented food. This work aimed to determine whether lactic acid bacteria fermentation of soy germ induces changes on its phytochemical composition. The antioxidant properties of fermented soy germ samples periodically taken during the fermentation process were evaluated and correlated with the concentration and structural modifications of isoflavones, saponins, phytosterols and tocopherols. Fermented soy germ extracts exhibited a higher inhibition effect against the superoxide anion radical, and lesser but significant ferric-reducing and DPPH radical scavenging effects compared with raw soy germ. By comparison to the traditional whole seed-based products, soy germ exhibits higher levels of isoflavones, saponins, phytosterols and tocopherols. All these phytochemicals contributed to the antioxidant capacity of soy germ and were conserved under lactic acid bacteria fermentation.

Background Parasite concentration methods facilitate molecular, biochemical and immunological research on the erythrocytic stages of Plasmodium. In this paper, an adaptation of magnetic MACS® columns for the purification of human Plasmodium species is presented. This method was useful for the concentration/purification of either schizonts or gametocytes. Results and conclusions The magnetic removal of non-parasitized red blood cells (in vivo and in vitro) using magnetic columns (MACS) was evaluated. This easy-to-use technique enriched schizonts and gametocytes from Plasmodium falciparum in vitro cultures with a very high degree of purity. In addition, all haemozoin-containing stages (schizonts and/or gametocytes) from the peripheral blood of infected patients could be concentrated using this method. This method is particularly useful for the concentration of non-falciparum species, which do not grow in culture and are otherwise difficult to obtain in large amounts.

International audience; Intraerythrocytic Plasmodium produces large amounts of toxic heme during the digestion of hemoglobin, a parasite specific pathway. Heme is then partially biocristallized into hemozoin and mostly detoxified by reduced glutathione. We proposed an in vitro micro assay to test the ability of drugs to inhibit heme-glutathione dependent degradation. As glutathione and o-phthalaldehyde form a fluorescent adduct, we followed the extinction of the fluorescent signal when heme was added with or without antimalarial compounds. In this assay, 50 microM of amodiaquine, arthemether, chloroquine, methylene blue, mefloquine and quinine inhibited the interaction between glutathione (50 microM) and heme (50 microM), while atovaquone did not. Consequently, this test could detect drugs that can inhibit heme-GSH degradation in a fast, simple and specific way, making it suitable for high throughput screening of potential antimalarials.

Svrha je ovog istraživanja bila proučiti odnos antioksidativnog kapaciteta i udjela izoflavona u ekstraktu soje s obzirom na geografsko podrijetlo i kultivar. Uzorci soje uzeti su iz dva dijela zrna soje, klice i kotiledona, s dvaju zemljopisnih lokacija (L1 i L2) i iz dva kultivara (Queen i Imari), ukupno 8 različitih uzoraka. HPLC metoda potvrdila je veći udio izoflavona u klicama nego u kotiledonima, i to u uzorcima s lokacije L2 i u kultivaru Queen. Antioksidativni kapacitet uzoraka soje određen je dvjema metodama, uklanjanjem 2,2'- difenil-1-pikrilhidrazil radikala i određivanjem sposobnosti apsorpcije kisikovih radikala. Rezultati obiju metoda pokazali su veću antioksidativnu aktivnost ekstrakta klice od ekstrakta kotiledona., The aim was to study the relationship between the antioxidant capacity and the isoflavone content of soybean extracts depending on both geographic origin and cultivar. Soybean samples were obtained from two soybean seed parts, germ and cotyledon, from two geographical locations (L1, L2) and two cultivars (Queen, Imari), which gave 8 different samples. HPLC determination confirmed higher isoflavone content in germs than in cotyledons, with higher contents in site L2, and in the Queen cultivar. The antioxidant capacity of soybean samples was determined with two methods, the 2,2-diphenyl-1-picrylhydrazyl scavenging assay and the oxygen radical absorbance capacity assay. The results obtained with both assays showed differences in antioxidant capacity between germ and cotyledon extracts, with a higher antioxidant activity of germ extracts.

A series of 2-alkyl and 2-aryl substituted-3H-indol-3-one-1-oxides was prepared and evaluated for its radical trapping properties. Spin trapping and electron paramagnetic resonance experiments demonstrate the ability of these indolone-1-oxides to trap hetero- and carbon-centered radicals. The most stable spin adducts (lifetime of several hours) are obtained with 2-alkyl substituted nitrones, the 2-ethyl-5,6-dioxolo-3H-indolone-1-oxide, 5e and the 2-secbutyl-3H-indolone-1-oxide, 5f. These two nitrones are also sensitive to redox reactions in solution. Therefore this indolone-1-oxide series lacking a beta-hydrogen atom gives rise to highly stable adducts with free radicals.

International audience; The capacity of estrogen and stilbene derivatives to modulate the activity of superoxide dismutases in relation with their estrogenic properties has been studied. The properties of trans-resveratrol (3,5,4′-trihydroxystilbene) and its analogues, 4-hydroxystilbene, 4,4′-dihydroxystilbene, 3,5-dihydroxystilbene, 3,5,4′-trimethoxystilbene and 4,4′-dihydroxy-3,5,3′,5′-tetramethylstilbene were compared to 17β-estradiol and its analogues (2-methoxyestradiol, estrone, 2-hydroxyestradiol and 2-methoxyestrone). Measurement of estrogen receptor-β (ER-β) binding capacity was carried out by a receptor competitor assay associated with fluorescence polarisation detection. The superoxide dismutase (SOD) modulation activity was followed with a spectrophotometric assay using the sequence xanthine/xanthine oxidase-2,3-bis[2-methoxy-4-nitro-sulfo-phenyl]-2H-tetrazolium-5-carboxanilide (X/XO-XTT). The structure–activity relationship was different for the two series tested. In the estrogenic series, a compound which does not inhibit SOD, is recognized by the ER-β. In contrast for the stilbenic series both properties are parallel each other.

International audience; Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzaldehyde, is a toxin produced by Eutypa lata, the causal agent of eutypa dieback of grapevine. The tolerance of some grapevine cultivars to the disease has been ascribed to the potential reduction of eutypine into its corresponding non-toxic alcohol, eutypinol. In the present study, eutypine biotransformation in different tissues of grapevine was investigated by HPLC and LC–MS. Grape callus tissues were able to biotransform eutypine into eutypinol within the first 3 h of culture. The grape plantlets cultured in vitro can also transform eutypine into eutypinol. Grape plantlet leaves do not have any effect on the uptake of eutypine, which goes through the tissues following a concentration gradient. Results revealed that the toxicity of eutypine in grape tissues is an active process showing that eutypinol is rapidly metabolised into other compounds. The use of micro-cuttings and in vitro plants showed that eutypine strongly accumulates in the bottom part of the diseased plant stems.

1,2-Dicarbonyl compounds employed as key-intermediates in indolone-N-oxide synthesis were prepared by direct oxidation of aryl, aryl- and aryl, alkyl-substituted alkenes assisted by benzeneseleninic anhydride.

International audience; To solve problems of sensitivity, repeatability and multi-step extraction related to 3-isobutyl-2-methoxypyrazine (IBMP) determination in wines, a simple method based on the novel combination of solid-phase microextraction and stable isotope dilution assay is presented. Among the parameters that affect this type of extraction, five of them have been optimised since the other parameters have common values or do not require optimisation (e.g. addition of sodium chloride at saturated concentration) and so were fixed. Vial volume, sample volume/vial volume ratio, pH, adsorption time and temperature have been optimised by means of two experimental designs. After extraction, quantification was performed by stable isotope dilution with gas chromatography-tandem mass spectrometry ([]-IBMP as internal standard). The final procedure allowed quantification far below IBMP’s sensory threshold (1 ng l−1 versus 15 ng l−1) with a 4% standard deviation. This method has been applied to experimental Fer servadou wines. Comparison of IBMP contents confirmed the efficiency of some viticultural and enological techniques on the herbaceous flavour decrease, such as prior fermentation maceration at high temperature (70 °C) and the use of a reflective carpet on viticultural soil.

Various oxime derivatives were evaluated as nitric oxide (NO) donors in arteries. Relaxation of rat aortic rings was used for bioassay of NO production, and electron paramagnetic resonance spectroscopy for demonstration of NO elevation. In rings with or without endothelium or adventitia, hydroxyguanidine and hydroxyurea were almost inactive, whereas formamidoxime, acetaldoxime, acetone oxime, acetohydroxamic acid and formaldoxime elicited relaxation. Active compounds increased NO levels in endothelium-denuded rings. Formaldoxime was the most potent agent for both relaxation and NO elevation in aortic rings, and it also increased NO in human aortic smooth muscle cells. In endothelium-denuded rings, relaxation was inhibited by a NO scavenger (2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide) and by inhibitors of soluble guanylyl-cyclase (1H[1,2,4,]oxadiazolo[4,3-a]quinoxalin-1-one) or cyclic GMP-dependent protein kinases (Rp-8-bromo cyclic GMP monophosphorothioate). Neither Nω-nitro- l -arginine methylester (a NO synthases inhibitor) nor proadifen (a cytochrome P450 inhibitor) decreased the effect of oxime derivatives. However, 7-ethoxyresorufin (7-ER, an inhibitor of P4501A1 which can also inhibit various NADPH-dependent reductases) abolished the relaxant effect of these compounds, without affecting the one of glyceryl trinitrate (GTN) or 2-(N,N-diethylamino)-diazenolate-2-oxide. 7-ER also abolished formaldoxime-induced NO increase in aortic rings. In rings tolerant to GTN, formaldoxime-induced relaxation and NO elevation were not different from those obtained in control rings. In conclusion, some oxime derivatives release NO by 7-ER-sensitive pathways in aortic smooth muscle, thus eliciting vasorelaxation. Pathways of NO formation are likely distinct from NO synthases and from those responsible for GTN biotransformation. Oxime derivatives could be useful for NO delivery in arteries in which endothelial NO synthase activity is impaired.

International audience; Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzaldehyde, is a toxin produced by Eutypa lata, the causal agent of Eutypa dieback in grapevine. The effect of the toxin on anthocyanin synthesis has been investigated in Vitis vinifera cv. Gamay cell cultures. At concentrations higher than 200 μmol/L, eutypine reduced anthocyanin accumulation in cells. The reduction in anthocyanin accumulation was proportional to the eutypine concentrations and HPLC analysis showed that eutypine affected the levels of all anthocyanins. The effect of eutypine application on the expression of five genes of the anthocyanin biosynthesis pathway, including chalcone synthase (CHS), flavonone-3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), leucoanthocyanidin dioxygenase (LDOX), and UDP glucose-flavonoid 3-O-glucosyl transferase (UFGT) was determined. Expression of CHS, F3H, DFR and LDOX was not affected by the addition of eutypine to grapevine cell cultures. In contrast, expression of the UFGT gene was dramatically inhibited by the toxin. These results suggest that in grapevine cell cultures, eutypine strongly affects anthocyanin accumulation by inhibiting UFGT gene expression. The mechanism of action of eutypine is discussed.

International audience; The biogenic amine tyramine has been reported to stimulatein vitro glucose transport in adipocytes, cardiomyocytes and skeletal muscle, and to improvein vivo glucose utilization in rats. These effects were dependent on amine oxidation, since they were blocked by inhibitors of monoamine oxidase (MAO) and semicarbazide-sensitive amine oxidase (SSAO). We thus tested in this work whether a prolonged treatment with tyramine could improve glucose tolerance in streptozotocin-induced diabetic rats. First, tyramine content of standard rodent chow was determined by HPLC and daily tyramine intake of control rats was estimated to be around 26 μmol/kg body weight. Then, tyramine was administred during 3 weeks in streptozotocin-induced diabetic rats at 29 μmol/kg by daily i.p. injection alone or together with vanadate 0.02 μmol/kg. In another group of diabetic rats, tyramine was subcutaneously delivered at 116 μmol/kg/day by osmotic minipumps. All tyramine treatments resulted in a decrease of the hyperglycemic responses to an i.p. glucose load. Adipocytes isolated from either untreated or treated diabetic rats were sensitive to the stimulation of glucose uptake by tyramine. However, diabetic animals receiving tyramine for three weeks did not recover from their hyperglycemia, hypoinsulinemia and glucosuria. These results show that the improvement of glucose tolerance induced by prolonged tyramine administration occurs in an insulin-depleted model and probably results from peripheral insulin-like actions of the oxidation of MAO/SSAO substrates, such as the stimulation of glucose uptake into adipocytes.

The present experiments were designed to analyze the influence of copper and iron ions on the process of decomposition of S-nitrosocysteine (cysNO), the most labile species among S-nitrosothiols (RSNO). CysNO fate in buffer solution was evaluated by optical and electron paramagnetic resonance (EPR) spectroscopy, and the consequences on its vasorelaxant effect were studied on noradrenaline-precontracted rat aortic rings. The main results are the following: (i) copper or iron ions, especially in the presence of the reducing agent ascorbate, accelerated the decomposition of cysNO and markedly attenuated the amplitude and duration of the relaxant effect of cysNO; (ii) by contrast, the iron and copper chelators bathophenantroline disulfonic acid (BPDS) and bathocuproine disulfonic acid (BCS) exerted a stabilizing effect on cysNO, prolonged its vasorelaxant effect, and abolished the influence of ascorbate; (iii) in the presence of ascorbate, BPDS displayed a selective inhibitory effect toward the influence of iron ions (but not toward copper ions) on cysNO decomposition and vasorelaxant effect, while BCS prevented the effects of both copper and iron ions; (iv) l -cysteine enhanced stability and prolonged the relaxant effect of cysNO; (v) the process of iron-induced decomposition of cysNO was associated with the formation of EPR-detectable dinitrosyl–iron complexes (DNIC) either with non-thiol- or thiol-containing ligands (depending on the presence of l -cysteine), both of which exhibiting vasorelaxant properties. From these data, it is concluded that the amount of intrinsic copper was probably too low to produce a destabilizing effect even on the most labile RSNO, cysNO, and that only intrinsic iron, through the formation of DNIC, was responsible for the process of cysNO decomposition and thus influenced its vasorelaxant properties.

Headspace solid phase microextraction (HS-SPME) was investigated as a solvent-free alternative method for the extraction and determination of 4-ethylphenol (EP) and 4-ethylguaiacol (EG) in red wine by capillary gas chromatography with flame ionization detection (FID) and compared to liquid–liquid extraction. For HS-SPME, better results were obtained with saturated sodium chloride samples, at 55 °C, using a 85 μm polyacrylate fiber. An absorption time of 40 min was needed to reach the absorption equilibrium for EG. This 40-min duration corresponds to the beginning of EP equilibrium and was selected for the experiments. In these conditions, the calibration graphs were linear in the range 5–5000 μg l −1 and the sensitivity was nearly the same for the two compounds. The detection limits were in the low μg l −1 range. In model wine solutions, result obtained with the liquid–liquid extraction method exhibit a linear calibration between 25 and 10,000 μg l −1 with a detection limit of 1 μg l −1 , but, the relative standard deviations of the EP and EG result in the low concentration range ( −1 ) are higher than those obtained by HS-SPME (15% compared to 2% for EP and 12% compared to 5% for EG). Taking into account the numerous volatile compounds in wine, HS-SPME is a rapid and valid alternative technique for use in the determination of ethylphenols at trace levels.

Endothelium produces oxygen-derived free radicals (nitric oxide, NOz.rad;; superoxide anion, O(2)(*-)) which play a major role in physiology and pathology of the vessel wall. However, little is known about endothelium-derived O(2)(*-) production, particularly due to the difficulty in assessing O(2)(*-) when its production is low and to controversies recently raised about the use of lucigenin-enhanced chemiluminescence. We compared four techniques of O(2)(*-) assessment when its production is low. In the present study, we have compared ferricytochrome c reduction, electron spin resonance (ESR) spectroscopy using DMPO as spin trap, hydroethidine fluorescence, and lucigenin-enhanced chemiluminescence to assess O(2)(*-) production in cultured bovine aortic endothelial cells (BAEC). We focused our study on extracellular O(2)(*-) production because the specificity of the signal is provided by the use of superoxide dismutase, and this control cannot be obtained intracellularly. We found that the calcium ionophore A23187 dose-dependently stimulated O(2)(*-) production, with a good correlation between all four techniques. The signals evoked by postconfluent BAEC were increased 2- to 7-fold in comparison to just-confluent BAEC, according to the technique used. Ferricytochrome c 20 microm rather than at 100 microm appears more suitable to detect O(2)(*-). However, in the presence of electron donors such as NADH or NADPH, lucigenin-enhanced chemiluminescence generated high amounts of O(2)(*-). Thus, ferricytochrome c reduction, electron spin resonance (ESR), and hydroethidine fluorescence appear as adequate tools for the detection of extracellular endothelium-derived O(2)(*-) production, whereas lucigenin may be artifactual, even when a low concentration of lucigenin is employed.

International audience; To evaluate the antioxidant activity of different phenolic acids and their esters, three types of experiments have been used. Electron paramagnetic resonance (EPR) quantitative analysis was carried out using the acetaldehyde/xanthine oxidase system and Fenton's reaction to generate superoxide and hydroxyl radicals, respectively. In a second test, hydroperoxides generated by Cu2+-catalysed oxidation of low density lipoproteins (LDL) were quantified by a modified iodometric method. In a third assay, LDL were oxidized with Esterbauer's method and modified LDL species were quantified by HPLC. The results show that the esterified phenolic derivatives present a better antioxidant activity, on the lipoperoxidation of LDL, than the corresponding phenolic acids.

Low-density lipoproteins (LDL) labeled with indium via a lipid-chelating agent, the bis(stearylamide) of diethylenetri-aminepentaacetic acid (L), were evaluated as a potential radiopharmaceutical (111In-L-LDL) for tumor localization by studying their internalization in human pancreatic cancer cells (Capan-1). Using Dil-LDL (1,1'-dioctadecyl-3,3,3',3'-tetramethylindodicarbocyanine perchlorate-LDL), this cell line was shown to bind human LDL with a high-affinity saturable component and a low-affinity non-saturable (40%) component. The single saturable high-affinity binding site had a KD of 27.5 +/- 2.1 micrograms/ml and a maximal binding of 610 +/- 7.5 ng/ml protein. Electron-microscopic examination of the In-L-LDL particles revealed the peripheral distribution of the electron-dense indium atoms at the outer surface of LDL. The modified LDL were then shown to be internalized by the cells. After conjugation of In-L-LDL to colloidal gold to follow the different stages of internalization, electron-microscopic examination showed that the In-L-LDL gold conjugates were stuck to the external sheet of the plasma apical and microvilli membrane, into earlier and later endosomes and into multivesicular bodies, suggesting the penetration of the In-L-LDL particles into lysosomal vacuoles. The observation of In-L-LDL-gold conjugates in deep-seated cytoplasm suggests that LDL could be employed as a drug-transport vehicle for targeting cytotoxics or radionuclides close to the cell nucleus.

The Superoxide anion radical, O2 −, plays a key role in the initiation stage of oxidative damage in biological systems (1). The main line of defense in mammalian organisms for controlling intracellular, and to a lesser extent extracellular, O2 − radicals are the Cu/Zn and Mn-containing Superoxide dismutase (SOD) enzymes and, recently, the application of SOD as a drug has attracted much attention (2). Since various problems are associated with using an enzyme as a drug (cost, bioavailability, stability, immunogenicity), non-toxic and low-mass metal complexes that catalyze the dismutation of O2 − might be able to substitute for SOD in such applications.

1,2-Dicarbonyl compounds employed as key-intermediates in indolone-N-oxide synthesis were prepared by direct oxidation of aryl, aryl- and aryl, alkyl-substituted alkenes assisted by benzeneseleninic anhydride.

Indolone-N-oxides, long known for their biological activities, possess remarkable anti-infectious properties. With the aim of improving the pharmacological and antimalarial properties of indolone-N-oxide derivatives (INODs), 6-(4-chlorophenyl)-7H-[1,3]dioxolo[4,5-f]indol-7-one-5-oxide, compound 1, was selected to study its penetration and biotransformation in red blood cells (RBC) in vitro. Compound 1 accumulated inside RBCs and was rapidly bio-transformed giving a major fluorescent metabolite, the dihydroanalogue, 1-HH, identified after extraction, through LC-MS and NMR analyses. This bioreductive transformation was (i) observed with other INOD derivatives (ii: 1–7); (ii) observed in normal, β-thalassemic and Plasmodium falciparum infected RBCs; (iii) temperature and thiol-dependent; (iv) not observed with heat-denatured RBCs, suggesting an enzyme-dependent biotransformation. The dihydro form, 1-HH, has antiplasmodial activity but lower than the parent compound. Since the RBCs represent 99% of the total cellular space of blood in humans, this leads to extensive metabolism of indolone-N-oxide type compounds. Given the redox events occurring in Plasmodium infected RBCs, this bioreductive transformation may be pivotal for parasite redox balance and antiplasmodial activity. However, it may be a drawback when other pharmacological properties of INODs are investigated. These results show the importance of RBCs as an in vitro model to study the biotransformation of drugs, especially antimalarial drugs in the early discovery stages.