Double-strand DNA cleavage induced by oxindole-Schiff base copper(II) complexes with potential antitumor activity

Some oxindole-Schiff base copper(II) complexes have already shown potential antitumor activity towards different cells, inducing apoptosis in a process modulated by the ligand, and having nuclei and mitochondria as main targets. Here, three novel copper(II) complexes with analogous ligands were isolated and characterized by spectroscopic techniques, having their reactivity compared to the so far most active complex in this class. Cytotoxicity experiments carried out toward human neuroblastoma SH-SY5Y cells confirmed its proapoptosis property. DNA cleavage studies were then performed in the presence of these complexes, in order to verify the influence of ligand structural features in its nuclease activity. All of them were able to cause double-strand DNA scissions, giving rise to nicked circular Form II and linear Form III species, in the presence of hydrogen peroxide. Additionally, DNA Form II was also detected in the absence of peroxide when the most active complex, [Cu(isaepy)(2)](2+) 1, was used. In an effort to better elucidate their interactions with DNA, solutions of the different complexes titrated with DNA had their absorption spectra monitored. An absorbance hyperchromism observed at 260 nm pointed to the intercalation of these complexes into the DNA structure. Further, investigations of 2-deoxy-D-ribose (DR) oxidation catalyzed by each of those complexes, using 2-thiobarbituric acid reactive species (TBARS) method, and detection of reactive oxygen species (ROS) formation by spin-trapping EPR, suggested that their mechanism of action in performing efficiently DNA cleavage occurs preferentially, but not only by oxidative pathways. (C) 2007 Elsevier Inc. All rights reserved.

Oxindole-Schiff base copper(II) complexes interactions with human serum albumin: Spectroscopic, oxidative damage, and computational studies

CD and EPR were used to characterize interactions of oxindole-Schiff base copper(II) complexes with human serum albumin (HSA). These imine ligands form very stable complexes with copper, and can efficiently compete for this metal ion towards the specific N-terminal binding site of the protein, consisting of the amino acid sequence Asp-Ala-His. Relative stability constants for the corresponding complexes were estimated from CD data, using the protein as competitive ligand, with values of log K(CuL) in the range 15.7-18.1, very close to that of [Cu(HSA)] itself, with log K(CuHSA) 16.2. Some of the complexes are also able to interfere in the a-helix structure of the protein, while others seem not to affect it. EPR spectra corroborate those results, indicating at least two different metal species in solution, depending on the imine ligand. Oxidative damage to the protein after incubation with these copper(II) complexes, particularly in the presence of hydrogen peroxide, was monitored by carbonyl groups formation, and was observed to be more severe when conformational features of the protein were modified. Complementary EPR spin-trapping data indicated significant formation of hydroxyl and carbon centered radicals, consistent with an oxidative mechanism. Theoretical calculations at density functional theory (DFT) level were employed to evaluate Cu(II)-L binding energies, L -> Cu(II) donation, and Cu(II) -> L back-donation, by considering the Schiff bases and the N-terminal site of HSA as ligands. These results complement previous studies on cytotoxicity, nuclease and pro-apoptotic properties of this kind of copper(II) complexes, providing additional information about their possibilities of transport and disposition in blood plasma. (C) 2009 Elsevier Inc. All rights reserved.

Spectroscopic characterization of schiff base-copper complexes immobilized in smectite clays

Herein, the immobilization of some Schiff base-copper(II) complexes in smectite clays is described as a strategy for the heterogenization of homogeneous catalysts. The obtained materials were characterized by spectroscopic techniques, mostly UV/Vis, EPR, XANES and luminescence spectroscopy. SWy-2 and synthetic Laponite clays were used for the immobilization of two different complexes that have previously shown catalytic activity in the dismutation of superoxide radicals, and disproportionation of hydrogen peroxide. The obtained results indicated the occurrence of an intriguing intramolecular redox process involving copper and the imine ligand at the surface of the clays. These studies are supported by computational calculations.

The isatin-Schiff base copper(II) complex Cu(isaepy)(2) acts as delocalized lipophilic cation, yields widespread mitochondrial oxidative damage and induces AMP-activated protein kinase-dependent apoptosis

We previously demonstrated that Bis[(2-oxindol-3-ylimino)-2-(2-aminoethyl) pyridine-N, N`] copper(II) [Cu(isaepy)(2)] was an efficient inducer of the apoptotic mitochondrial pathway. Here, we deeply dissect the mechanisms underlying the ability of Cu(isaepy)(2) to cause mitochondriotoxicity. In particular, we demonstrate that Cu(isaepy)(2) increases NADH-dependent oxygen consumption of isolated mitochondria and that this phenomenon is associated with oxy-radical production and insensitive to adenosine diphosphate. These data indicate that Cu(isaepy)(2) behaves as an uncoupler and this property is also confirmed in cell systems. Particularly, SH-SY5Y cells show: (i) an early loss of mitochondrial transmembrane potential; (ii) a decrease in the expression levels of respiratory complex components and (iii) a significant adenosine triphosphate (ATP) decrement. The causative energetic impairment mediated by Cu(isaepy)(2) in apoptosis is confirmed by experiments carried out with rho(0) cells, or by glucose supplementation, where cell death is significantly inhibited. Moreover, gastric and cervix carcinoma AGS and HeLa cells, which rely most of their ATP production on oxidative phosphorylation, show a marked sensitivity toward Cu(isaepy)(2). Adenosine monophosphate-activated protein kinase (AMPK), which is activated by events increasing the adenosine monophosphate: ATP ratio, is deeply involved in the apoptotic process because the overexpression of its dominant/negative form completely abolishes cell death. Upon glucose supplementation, AMPK is not activated, confirming its role as fuel-sensing enzyme that positively responds to Cu(isaepy)(2)-mediated energetic impairment by committing cells to apoptosis. Overall, data obtained indicate that Cu(isaepy)(2) behaves as delocalized lipophilic cation and induces mitochondrial-sited reactive oxygen species production. This event results in mitochondrial dysfunction and ATP decrease, which in turn triggers AMPK-dependent apoptosis.

Mechanical, thermal and morphological properties of glutaraldehyde crosslinked bovine pericardium followed by glutamic acid treatment

Major problems with valve bioprostheses are associated with progressive structural deterioration and calcification, directly associated with the use of glutaraldehyde (GA). This work describes the effects of GA processing and borate/glutamic acid buffer treatment on the mechanical, thermal and morphological properties of 0.5% GA crosslinked bovine pericardium (BP). The results showed that while the treatment of 0.5% GA crosslinked BP with borate/glutamic acid significantly improves the mechanical properties, it had no visible effect on surface morphology. Better surface preservation was only achieved for BP pre-treated with a lower GA concentration followed by the conventional treatment (0.5% GA). Improvements in mechanical properties probably arises from structural changes probably involving the depolymerization of polymeric GA crosslinks and an increase electrostatic interaction due to covalent binding of glutamic acid to free carbonyl groups (Schiff base).The results indicate that the treatment GA crosslinked BP with borate/glutamic acid buffer may be an attractive procedure for the manufacture of heart valve bioprostheses.

Acidity and photolability of ruthenium salen nitrosyl and aquo complexes in aqueous solutions

The photochemical behavior of nitrosyl complexes Ru(salen)(NO)(OH(2))(+) and Ru(salen)(NO) Cl (salen = N, N`-ethylenebis-(salicylideneiminato) dianion) in aqueous solution is described. Irradiation with light in the 350-450 nm range resulted in nitric oxide (NO) release from both. For Ru(salen)(NO) Cl secondary photoreactions also resulted in chloride aquation. Thus, in both cases the final photoproduct is the diaquo cation Ru(III) (salen) (OH(2))(2)(+), for which pK(a)`s of 5.9 and 9.1 were determined for the coordinated waters. The pK(a) of the Ru(salen)(NO)(OH(2))+ cation was also determined as 4.5 +/- 0.1, and the relative acidities of these ruthenium aquo units are discussed in the context of the bonding interactions between Ru(III) and NO. (C) 2007 Elsevier B.V. All rights reserved.

Synthesis and Crystal Structures of Octahedral Sn(IV) Complexes Prepared from SnCl(2)center dot 2H(2)O and 2-Hydroxyacetophenone (S-benzydithiocarbazate) Ligand (H(2)L)

Two coordination octahedral Sn(IV) complexes [Sn(L)(2)] and cis-[SnCl(2)(L)(dmso)], where H(2)L is 2-hydroxyacetophenone (S-benzydithiocarbazate), were prepared and characterized by elemental analysis, IR, NMR ((1)H, (13)C), (119)Sn Mossbauer spectroscopies and X-ray diffraction techniques to investigate their structural properties. Both crystallize in the Monoclinic system, with parameters: a = 8.1905(3), b = 30.8811(15), c = 12.8959(7) angstrom, beta = 94.465(3)degrees and Z = 4 for [Sn(L)(2)] and a = 8.5247(2), b = 21.5445(7), c = 12.3706(3) angstrom, beta = 96.932(2)degrees and Z = 4 for cis-[SnCl(2)(L)(dmso)]. In both complexes, the Sn(IV) central atom is coordinated in a distorted octahedral geometry with the thiolate ligand (L(2-)) coordinated via O, N and S atoms. The (119)Sn Mossbauer spectroscopy of the complexes were studied and the results revealed that both complexes posses isomer shift (delta) and quadrupole splitting (Delta), which are almost the same.

Synthesis and Characterization of Homoleptic and Heteroleptic Cobalt, Nickel, Copper, Zinc and Cadmium Compounds with the 2-(Tosylamino)-N-[2-(tosylamino)benzylidene]aniline Ligand

The electrochemical oxidation of anodic metal (cobalt, nickel, copper, zinc and cadmium) in an acetonitrile solution of the Schiff-base ligand 2-(tosylamino)-N-[2-(tosylamino)-benzylidene] aniline (H(2)L) afforded the homoleptic compounds [ML]. The addition of 1,1-diphenylphosphanylmethane (dppm), 2,2`-bipyridine (bipy) or 1,10-phenanthroline (phen) to the electrolytic phase gave the heteroleptic complexes [NiL(dppm)], [ML(bipy)] and [ML(phen)]. The crystal structures of H(2)L (1), [NiL] (2), [CuL] (3), [NiL(dppm)] (4), [CoL(phen)] (5), [CuL(bipy)] (6) and [Zn(Lphen)] (7) were determined by X-ray diffraction. The homoleptic compounds [NiL] and [CuL] are mononuclear with a distorted square planar [MN(3)O] geometry with the Schiff base acting as a dianionic (N(amide)N(amide)N(imine)O(tosyl)) tetradentate ligand. Both compounds exhibit an unusual pi-pi stacking interaction be-tween a six-membered chelate ring containing the metal and a phenylic ring of the ligand. In the heteroleptic complex [NiL(dppm)], the nickel atom is in a distorted tetrahedral [NiN(3)P] environment defined by the imine, two amide nitrogen atoms of the L(2-) dianionic tridentate ligand and one of the phosphorus atoms of the dppm molecule. In the other heteroleptic complexes, [CoL(phen)], [CuL(bipy)] and [ZnL(phen)], the metal atom is in a five-coordinate environment defined by the imine, two amide nitrogen atoms of the dianionic tridentate ligand and the two bipyridine or phenanthroline nitrogen atoms. The compounds were characterized by microanalysis, IR and UV/Vis (Co, Ni and Cu complexes) spectroscopy, FAB mass spectrometry and (1)H NMR ([NiL] and Zn and Cd complexes) and EPR spectroscopy (Cu complexes).

Dinuclear Azide-Bridged Copper(II) Complex as Building Block for the Assembly of a 2D-Supramolecular Array

A novel Schiff base-copper(II) complex [Cu(2)L(2)(N(3))(2)](ClO(4))(2) 1, where L = (4-imidazolyl)ethylene-2-amino-1-ethylpyridine (apyhist), containing azide-bridges between adjacent copper ions in a dinuclear arrangement was isolated and characterized both in the solid state and in solution by X-ray crystallography and different spectroscopic techniques. Azide binding constants were estimated from titrations of the precursor [CuL(H(2)O)(2)](2+) solutions with sodium azide, giving rise to the azido-bridged species, [Cu(2)L(2)(N(3))(2)](2+). Raman spectra showed asymmetric stretching band at 2060 cm(-1), indicating the presence of azido ligands with a symmetric mu(1,) (1) binding geometry. EPA spectra, in frozen methanol/water solutions at 77 K, exhibited characteristic features of copper centers in tetragonal pyramidal coordination geometry, exhibiting magnetic interactions between them. Further, in solid state, two different values for magnetic coupling in this species were obtained, J/k = -(5.14 +/- 0.02) cm(-1) attributed to the mu(1, 1) azide-bridge mode, and J`z`/k = -(2.94 +/- 0.11) cm(-1) for the interaction between dinuclear moieties via water/perchorate bridges. Finally, an attempt was made to correlate structure and magnetic data for this dinuclear asymmetric end-on azido bridged-copper(II) 1 complex with those of another correlated dinuclear system, complex [Cu(2)L(2)Cl(2)](ClO(4))(2) 2, containing the same tridentate diimine ligand, but with chloro-bridged groups between the copper centres.

Synthesis, spectral studies and X-ray crystal structure of N,N `-(+/-)-trans-1,2-cyclohexylenebis(3-ethoxysalicylideneamine) H-2(t-3-EtOsalchxn)

The synthesis, spectra and X-ray crystal structure of N,N`-(+/-)-trans-1,2-cyclohexylenebis(3-ethoxysalicylideneamine) H-2(t-3-EtOsalchxn), a salen-type ligand, are reported. The Schiff base was characterized by elemental analysis, m.p., IR, electronic spectra, H-1 and C-13 NMR spectra. The spectra are discussed and compared with those of N,N`-(+/-)-trans-1,2-cyclohexylenebis(salicylideneamine), H-2(t-salchxn). The electronic and IR spectra were also resolved by deconvolution. The influence of the ethoxy group on the IR, electronic spectrum, H-1 and C-13 NMR spectra is discussed. Strong intramolecular forces are present as supported by the IR and H-1 NMR spectra and the X-ray crystal structure. An intermolecular hydrogen bond is observed and appears twice in a pair of molecules in the unit cell. (c) 2007 Elsevier B.V. All rights reserved.

Ionic mobility of the solvated proton and acid-base titration in a four-compartment capillary electrophoresis system

Although H(+) and OH(-) are the most common ions in aqueous media, they are not usually observable in capillary electrophoresis (CE) experiments, because of the extensive use of buffer solutions as the background electrolyte. In the present work, we introduce CE equipment designed to allow the determination of such ions in a similar fashion as any other ion. Basically, it consists of a four-compartment piece of equipment for electrolysis-separated experiments (D. P. de Jesus et at, Anal. Chem., 2005, 77, 607). In such a system, the ends of the capillary are placed in two reservoirs, which are connected to two other reservoirs through electrolyte-filled tubes. The electrodes of the high-voltage power source are positioned in these reservoirs. Thus, the electrolysis products are kept away from the inputs of the capillary. The detection was provided by two capacitively coupled contactless conductivity detectors (CD), each one positioned about 11 cm from the end of the capillary. Two applications were demonstrated: titration-like procedures for nanolitre samples and mobility measurements. Strong and weak acids (pK(a) < 5), pure or mixtures, could be titrated. The analytical curve is linear from 50 mu M up to 10 mM of total dissociable hydrogen (r = 0.99899 for n =10) in 10-nL samples. By including D(2)O in the running electrolyte, we could demonstrate how to measure the mixed proton/deuteron mobility. When H(2)O/D(2)O (9 : 1 v/v) was used as the solvent, the mobility was 289.6 +/- 0.5 x 10(-5) cm(2) V(-1) s(-1). Due to the fast conversion of the species, this value is related to the overall behaviour of all isotopologues and isotopomers of the Zundel and Eigen structures, as well as the Stokesian mobility of proton and deuteron. The effect of neutral (o-phenanthroline) and negatively charged (chloroacetate) bases and aprotic solvent (DMSO) over the H(+) mobility was also demonstrated.

Reproductive biology and pollination of Govenia utriculata: A syrphid fly orchid pollinated through a pollen-deceptive mechanism

The reproductive biology and pollination mechanisms of Govenia utriculata (Sw.) Lindl. were studied in a mesophytic semideciduous forest at Serra do Japi, south-eastern Brazil. The floral visitors and pollination mechanisms were recorded, and experimental pollinations were carried out to determine the breeding system of this species. Populations of G. utriculata growing at Serra do Japi are exclusively visited and pollinated by two species of hoverflies in the genus Salpingogaster (Diptera: Syrphidae) that are attracted by deceit to the flowers of this orchid species. The lip apex and the column base present small brownish and yellow to orange spots that mimic pollen clusters. Govenia utriculata is self-compatible, but pollinator dependent. Natural fruit set was low (10%), but similar to that of other non-obligatorily autogamous sympatric orchid species that occur at Serra do Japi and of other fly-pollinated orchid species pollinated through deceptive mechanisms.

Catalytic mechanism of inulinase from Arthrobacter sp S37

Detailed catalytic roles of the conserved Glu323, Asp460, and Glu519 of Arthrobacter sp. S37 inulinase (EnIA), a member of the glycoside hydrolase family 32, were investigated by site-directed mutagenesis and pH-dependence studies of the enzyme efficiency and homology modeling were carried out for EnIA and for D460E mutant. The enzyme efficiency (k(cat)/K-m) of the E323A and E519A mutants was significantly lower than that of the wild-type due to a substantial decrease in k(cat), but not due to variations in K-m, consistent with their putative roles as nucleophile and acid/base catalyst, respectively. The D460A mutant was totally inactive, whereas the D460E and D460N mutants were active to some extent, revealing Asp460 as a catalytic residue and demonstrating that the presence of a carboxylate group in this position is a prerequisite for catalysis. The pH-dependence studies indicated that the pK(a) of the acid/base catalyst decreased from 9.2 for the wild-type enzyme to 7.0 for the D460E mutant, implicating Asp460 as the residue that interacts with the acid/base catalyst Glu519 and elevates its pK(a). Homology modeling and molecular dynamics simulation of the wild-type enzyme and the D460E mutant shed light on the structural roles of Glu323, Asp460, and Glu519 in the catalytic activity of the enzyme. (C) 2008 Elsevier Inc. All rights reserved.

T-wave alternans: desempenho clínico, limitações, metodologias de análise

Reconhecer com precisão indivíduos com maior risco imediato de morte súbita cardíaca (MSC) ainda é uma questão em aberto. A natureza fortuita dos eventos cardiovasculares agudos não parece se adequar ao conhecido modelo de indução de taquicardia/fibrilação ventricular por um gatilho em sincronia a um substrato arritmogênico estático. Quanto ao mecanismo da MSC, uma instabilidade elétrica dinâmica explicaria melhor a raridade da associação simultânea de um gatilho certo a um substrato cardíaco apropriado. Diversos estudos tentaram medir essa instabilidade elétrica cardíaca (ou um equivalente válido) em uma sequência de batimentos cardíacos no ECG. Dentre os mecanismos possíveis podemos citar o prolongamento do QT, dispersão do QT, potenciais tardios, alternância de onda T ou T-wave alternans (TWA), e turbulência da frequência cardíaca. Este artigo se atém em particular ao papel da TWA no panorama atual da estratificação de risco cardíaco. Os achados sobre TWA ainda são heterogêneos, variando de um desempenho prognóstico muito bom até um quase nulo, dependendo da população clínica observada e protocolo clínico usado. Para preencher as atuais lacunas no conhecimento sobre TWA, profissionais médicos e pesquisadores devem explorar melhor as características técnicas das diversas tecnologias disponíveis para a avaliação de TWA e atentar ao fato de que os valores de TWA respondem a diversos outros fatores, além de medicamentos. Informações sobre mecanismos celulares e subcelulares da TWA estão fora do escopo deste artigo, mas são referenciados alguns dos principais trabalhos sobre este tópico, com o intuito de auxiliar no entendimento dos conceitos e fatos cobertos neste artigo.

Influence of incorporation of fluoroalkyl methacrylates on roughness and flexural strength of a denture base acrylic resin

Fluorinated denture base acrylic resins can present more stable physical properties when compared with conventional polymers. This study evaluated the incorporation of a fluoroalkyl methacrylate (FMA) mixture in a denture base material and its effect on roughness and flexural strength. A swelling behavior assessment of acrylic resin specimens (n=3, per substance) after 12 h of FMA or methyl methacrylate (MMA) immersion was conducted to determine the solvent properties. Rectangular specimens (n=30) were allocated to three groups, according to the concentration of FMA substituted into the monomer component of a heat-polymerized acrylic resin (Lucitone 550), as follows: 0% (control), 10% and 20% (v/v). Acrylic resin mixed with concentrations of 25% or more did not reach the dough stage and was not viable. The surface roughness and flexural strength of the specimens were tested. Variables were analyzed by ANOVA and Tukey's test (a=0.05). Immersion in FMA produced negligible swelling, and MMA produced obvious swelling and dissolution of the specimens. Surface roughness at concentrations of 0%, 10% and 20% were: 0.25 ± 0.04, 0.24 ± 0.04, 0.22 ± 0.03 mm (F=1.78; p=0.189, not significant). Significant differences were found for flexural strength (F=15.92; p<0.001) and modulus of elasticity (F=7.67; p=0.002), with the following results: 96 ± 6, 82 ± 5, 84 ± 6 MPa, and 2,717 ± 79, 2,558 ± 128, 2574 ± 87 MPa, respectively. The solvent properties of FMA against acrylic resin are weak, which would explain why concentrations over 20% were not viable. Surface changes were not detected after the incorporation of FMA in the denture base acrylic resin tested. The addition of FMA into denture base resin may lower the flexural strength and modulus of elasticity, regardless of the tested concentration.