Structural and functional characterization of myotoxin I, a Lys49 phospholipase A(2) homologue from Bothrops moojeni (Caissaca) snake venom

Myotoxin-I (MjTX-I) was purified to homogeneity from the venom of Bothrops moojeni by ion-exchange chromatography on CM-Sepharose. Its molecular weight, estimated by SDS-PAGE, was 13,400 (reduced) or 26,000 (unreduced). The extinction coefficient (E-1.0 cm(1.0 mg/ml)) of MjTX-I was 1.145 at lambda = 278 nm, pH 7.0, and its isoelectric point was 8.2 at ionic strength mu = 0.1. When lyophilized and stored at 4 degrees C, dimeric, trimeric, and pentameric forms of the protein were identified by SDS-PAGE. This heterogeneous sample could be separated into three fractions by gel filtration on Sephadex 6-50. The fractions were analyzed by isoelectric focusing, immunoelectrophoresis, and amino acid composition, which indicated that heterogeneity was the result of different levels of self-association. Protein sequencing indicated that MjTX-I is a Lys49 myotoxin and consists of 121 amino acids (M-r = 13,669), containing a high proportion of basic and hydrophobic residues. It shares a high degree of sequence identity with other Lys49 PLA(2)-like myotoxins, but shows a significantly lower identity with catalytically active Asp49 PLA(2)s. The three-dimensional structure of MjTX-I was modeled based on the crystal structures of three highly homologous Lys49 PLA(2)-like myotoxins. This model showed that the amino acid substitutions are conservative, and mainly the beta-wing region, and the C-terminal extended random coil. MjTX-I displays local myotoxic and edema-inducing activities in mice, and is lethal by intraperitoneal injection, with an LD50 value of 8.5 +/- 0.8 mg/kg, In addition, it is cytotoxic to myoblasts/ myotubes in culture, and disrupts negatively charged liposomes. In comparison with the freshly prepared dimeric sample, the more aggregated forms showed significantly reduced myotoxic activity. However, the edema-inducing activity of MjTX-I was independent of molecular association. Phospholipase A(2) activity on egg yolk, as well as anticoagulant activity, were undetectable both in the native and in the more associated forms. His, Tyr, and Trp residues of the toxin were chemically modified by specific reagents. Although the myotoxic and lethal activities of the modified toxins were reduced by these treatments, neither its edema-inducing or Liposome-disrupting activities were significantly altered. Rabbit antibodies to native MjTX-I cross-reacted with the chemically modified forms, and both the native and modified MjTX-I preparations were recognized by antibodies against the C-terminal region 115-129 of myotoxin II from B. asper, a highly Lys49 PLA(2)-homologue with high sequencial similarity. (C) 2000 Academic Press.

Formation and characterization of soy bean oil/surfactant/water microemulsions

Pseudoternary phase diagrams, at 25 degrees C, were constructed for the systems soy bean oil (SBO)/surfactant/water, with single anionic sodium bis(2-ethylhexyl)sulfosuccinate (AOT), nonionic monoolein (MO) and mixtures of these surfactants, showing the isotropic phase of W/O microemulsions (MEs). The area of ME formation in the phase diagrams was shown to be dependent of the relative amount of surfactants, being larger for MO:AOT equals to 2:1. Rheological and dynamic light scattering (DLS) studies indicated that the viscosity of the isotropic ME phase exhibited two different behaviors depending on composition. The viscosity of dry MEs initially decreased with increasing amount of water following a dilution line in the phase diagram, i.e., a constant surfactant:SBO percentage ratio. As the water content increased the relative viscosity attained a minimum and then increased. This minimum could be related to the transition between two ME regions, L-2 and L'(2), having different characteristics. DLS measurements confirm the existence of ordinary W/O ME droplets in the L-2 region and suggest the existence of another structure in the L'(2) region. The size of the MEs droplets in L-2 phase ranges from 3.6 to 16.5 nm, depending on composition of SBO, surfactant and water. Small angle X-ray scattering (SAXS) also indicates the existence of structures with different characteristics, for the SAXS curves exhibit a typical micelle asymmetrical peak at low scattering vector q for MEs in L-2 but a symmetrical correlation peak at higher q vector in L'(2). (c) 2006 Elsevier B.V. All rights reserved.

SYNTHESIS AND CRYSTAL-STRUCTURE OF THE NOVEL CYCLOPALLADATED COMPLEX DI(MU,N-S-ETA(2)-QUINOLINE-2-THIOLATE)-BIS[(N,N-DIMETHYLBENZYLAMINE-C(2),N) PALLADIUM(II)]

The compound di(mu,N-Seta2-2-quinoline-2-thiolate)-bis[(N,N-dimethylbenzylamine-C2,N)palladium(II)] was synthesized and studied by IR, NMR and X-ray diffraction: monoclinic, a = 20.138(3), b = 10.831(1), c = 14.973(2) angstrom, beta = 98.04(1)-degrees, Z = 4, space group P2(1)/c, R = 0.032. The compound is dimeric with the two [Pd(N,N-dimethylbenzylamine)]moieties being connected by the two vicinal bridging eta2-N,S-quinoline-2-thiolate anions in a square-planar coordination geometry for the palladium atoms.

beta-ZnMoO4 microcrystals synthesized by the surfactant-assisted hydrothermal method: Growth process and photoluminescence properties

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Highly swollen liquid crystals as new reactors for the synthesis of nanomaterials

Synthesis and self-assembly of nanomaterials can be controlled by the properties of soft matter. on one hand, dedicated nanoreactors such as reverse microemulsions or miniemulsions can be designed. on the other hand, direct shape control can be provided by the topology of liquid crystals that confine the reacting medium within a specific geometry. In the first case, the preparation of micro- or miniemulsions generally requires energetic mechanical stirring. The second approach uses thermodynamically stable systems, but it remains usually limited to binary (water + surfactant) systems. We report the preparation of different families of materials in highly ordered quaternary mediums that exhibit a liquid crystal structure with a high cell parameter. They were prepared with the proper ratios of salted water, nonpolar solvent, surfactant. and cosurfactants that form spontaneously swollen hexagonal phases. These swollen liquid crystals can be prepared from all classes of surfactants (cationic, anionic, and nonionic). They contain a regular network of parallel cylinders, whose diameters can be swollen with a nonpolar solvent, that are regularly spaced in a continuous aqueous salt solution. We demonstrate in the present report that both aqueous and organic phases can be used as nanoreactors for the preparation of materials. This property is illustrated by various examples such as the synthesis of platinum nanorods prepared in the aqueous phase or zirconia needles or the photo- or gamma-ray-induced polymerization of polydiacetylene in the organic phase. In all cases, materials can be easily extracted and their final shapes are directed by the structure-directing effect imposed by the liquid crystal.

Combined (13)C NMR and DFT/GIAO studies of the polyketides Aurasperone A and Fonsecinone A

Structural characterization by NMR spectroscopy and DFT calculations was performed for two dimeric naptho-gamma-pyrones, the polyketides Aurasperone A and Fonsecinone A. Experimental data ((13)C NMR chemical shifts and interatomic geometries) were found to be in reasonable agreement with theoretical ones, obtained at B3LYP level for three different basis sets (6-31G/6-31G(d)/6-31G(d,p)). Additionally, the dipolar moments calculation allowed explaining the different solubility for these molecules. The (13)C NMR theoretical chemical shifts were calculated with the GIAO method and the solvent effects were taken into account by means of the PCM approximation. In this work, the DFT/GIAO methodology shows to be a reliable tool in the assignment of experimental NMR chemical shifts of similar molecules. (C) 2008 Wiley Periodicals, Inc. Int J Quantum Chem 108: 2408-2416, 2008.

Modulation of the Pharmacological Activities of Secretory Phospholipase A2 from Crotalus durissus cascavella Induced by Naringin

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Efeitos tóxicos de surfactantes fitossanitários para o peixe mato grosso (Hyphessobrycon eques)

Toxic effects of phytosanitary surfactants for jewel tetra (Hyphessobrycon eques). Surfactants are amphipatic molecules that reduce the surface tension of water and make up the inert components of pesticide formulations. Thus, the objectives of this study were: to estimate the lethal concentration (LC(I)50;96h); classify and evaluate water quality during testing of the following surfactants: Agral[registered trademark], Aterbane[registered trademark]BR, Ag-bem[registered trademark], Energic[registered trademark], Fixade[registered trademark] and Gotafix[registered trademark] for jewel tetra (Hyphessobrycon eques); and the signs of intoxication in the animals. For this, the fish were acclimated for ten days in the bioassay room. The animals were exposed to the surfactants in an entirely randomized design with three replications. The LC(I)(50;96h) of surfactant Agral[registered trademark] was 3.29 mg L-1; Aterbane[registered trademark] BR 8.21 mg L-1; Energic[registered trademark] 2.34 mg L-1; Gotafix[registered trademark] 4.37 mg L-1; Fixade[registered trademark] 3.38 mg L-1; and Ag-bem[registered trademark] 34.95 mg L-1. The variables of water quality were unchanged. The fish showed an increase in the opercular beating after exposure; 4 and 24 hours, loss of gasping ability; 48 hours, slow opercular beating; and 72 and 96 hours later, recovery. The surfactants Energic[registered trademark], Agral[registered trademark], Gotafix[registered trademark], Aterbane[registered trademark] BR and Fixade[registered trademark] can be classified as moderately toxics, and Ag-Bem[registered trademark] as slightly toxic for H. eques; this organism shows similar intoxication signs for all surfactants.

Synthesis and X-ray studies of di-μ-cyanato-bis[{cyanato (N,N- dimethylethylenediamine)}copper(II)]

The compound di-μ-cyanato-bis[{cyanato(N,N-dimethylethylenediamine)} copper(II)] was synthesized, and studied by IR spectroscopy and X-ray diffraction. It is dimeric with bridging and terminal cyanate groups, and the copper atoms show a square-based pyramid coordination geometry. © 1990.

EFEITO DE SURFACTANTES NA ESTABILIDADE DE COMPOSTOS DE USO TERAPEUTICO

A critical revision of literature as regards to the drug stability in the presence of surfactants were realized. The functional groups envolved in the drug decomposition were used to the development of the discussion. The analysis indicated that the detergent effect can be used to control the rates and mechanisms of drug decomposition and to obtain specific information about the drug reactivity in the environment of pharmacological action.

Polybitoxins: A group of phospholipases A2 from the venom of the neotropical social wasp paulistinha (Polybia paulista)

The neotropical wasp Polybia paulista is very aggressive and endemic in south-east Brazil, where it frequently causes stinging accidents. By using gel filtration on Sephadex G-200, followed by ion-exchange chromatography on DEAE-Cellulose under a pH gradient, a group of four toxins (designated as polybitoxins-I, II, lII and IV) presenting phospholipase A2 (PLA2) activities was purified. These toxins are dimeric with mol. wts ranging from 115,000 to 132,000 and formed by different subunits. The four toxins contain very high sugar contents attached to their molecules (22-43% w/w) and presented different values of pH optimum from 7.8 to 9.0; when dissociated, only residual catalytic activities were maintained. The catalytic activities of polybitoxins (from 18 to 771 μmoles/mg per minute) are lower than that of PLA2 from Apis mellifera venom and hornetin from Vespa basalis. The polybitoxins presented a non-linear steady-state kinetic behavior for the hydrolysis of phosphatidylcholine at pH 7.9, compatible with the negative co- operativity phenomena. All of the polybitoxins were very potent direct hemolysins, especially the polybitoxins-III and IV, which are as potent as the lethal toxin from V. basalis and hornetin from Vespa flavitarsus, respectively; polybitoxin-IV presented hemolytic action 20 times higher than that of PLA2 from A. mellifera, 17 times higher than that of neutral PLA2 from Naja nigricolis and about 37 times higher than that of cardiotoxin from Naja naja atra venom.

Effect of micelles, microemulsions and other supramolecular aggregates on the hydrolysis of β-lactam antibiotics

A critical revision of the literature was made regarding the stability of β-lactam antibiotics in the presence of surfactants. The factors involved in the drug decomposition were analyzed in the development of the discussion. The analysis has indicated that some organized systems obtained from surfactants can be used to control rates and mechanisms of antibiotic decomposition. These organized systems can also be used to obtain specific information about the drug reactivity in a microenvironment similar to the site of pharmacological effect.

Inhibin B and the ovarian follicular reserve

Inhibins are dimeric glycoproteins composed of an α-subunit and a βA-subunit (inhibin A) or βB-subunit (inhibin B), which inhibits pituitary gonadotropin secretion of FSH. The inhibin B is a product of the cohort of antral follicles. The ovarian follicle number decrease steadily as a function of increasing age, with consequent falls in the levels of inhibin B and increase of FSH levels. It is sufficient to maintain ovulatory function and continued secretion of estradiol. Elevated FSH levels seem to occur late in the sequence of events associated with ovarian failure. The inhibin B, produced by granulosa cells, is the earliest marker of the decline in ovarian follicular reserve across reproductive aging.

Different doses of exogenous surfactant for treatment of meconium aspiration syndrome in newborn rabbits.

OBJECTIVE: To evaluate the effects of 2 different doses of exogenous surfactant on pulmonary mechanics and on the regularity of pulmonary parenchyma inflation in newborn rabbits. METHOD: Newborn rabbits were submitted to tracheostomy and randomized into 4 study groups: the Control group did not receive any material inside the trachea; the MEC group was instilled with meconium, without surfactant treatment; the S100 and S200 groups were instilled with meconium and were treated with 100 and 200 mg/kg of exogenous surfactant (produced by Instituto Butantan) respectively. Animals from the 4 groups were mechanically ventilated during a 25-minute period. Dynamic compliance, ventilatory pressure, tidal volume, and maximum lung volume (P-V curve) were evaluated. Histological analysis was conducted using the mean linear intercept (Lm), and the lung tissue distortion index (SDI) was derived from the standard deviation of the means of the Lm. One-way analysis of variance was used with a = 0.05. RESULTS: After 25 minutes of ventilation, dynamic compliance (mL/cm H2O.kg) was 0.87 +/- 0.07 (Control); 0.49 +/- 0.04 (MEC*); 0.67 +/- 0.06 (S100); and 0.67 +/- 0.08 (S200), and ventilatory pressure (cm H2O) was 9.0 +/- 0.9 (Control); 16.5 +/- 1.7 (MEC*); 12.4 +/- 1.1 (S100); and 12.1 +/- 1.5 (S200). Both treated groups had lower Lm values and more homogeneity in the lung parenchyma compared to the MEC group: SDI = 7.5 +/- 1.9 (Control); 11.3 +/- 2.5 (MEC*), 5.8 +/- 1.9 (S100); and 6.7 +/- 1.7 (S200) (*P < 0.05 versus all the other groups). CONCLUSIONS: Animals treated with surfactant showed significant improvement in pulmonary mechanics and more regularity of the lung parenchyma in comparison to untreated animals. There was no difference in results after treatment with either of the doses used.

On the specific filtration mechanism of a mesoporous silica membrane, prepared with non-connecting parallel pores

We report the singular filtration properties of an ultrafiltration membrane made with mesoporous silica that exhibits cylindrical pores aligned mostly normal to the support. This membrane supported on tubular commercial macroporous alumina supports was prepared by the interfacial growth mechanism between stable silica-surfactant hybrid micelles made of the association of silica oligomers with polyethyleneoxide-based (PEO) surfactants and sodium fluoride, a well-known silica condensation catalyst [Boissière et al., An ultrafiltration membrane made with mesoporous MSU-X silica, Chem. Mater. 15 (2003) 460-463]. It appears that the combined effect of the silica nature of the membrane, whose surface charge can be easily adjusted by changing the pH and the non-connected cylindrical shape of the pores provides a new behavior in the retention properties, as proved by the filtration of polyoxyethylene polymers (PEO) with different molecular weights. Depending on the filtration conditions, a rejection rate of 80% and a steep cut-off at 2000 Da can be obtained or, on the reverse, polymers three times bigger than the pore diameter can diffuse through the membrane. This new filtration mechanism, which opens up new modes of separation modes, is explained in the light of both topology of the porous network and pH-dependent interactions between PEO polymers and silica porous media. © 2004 Elsevier B.V. All rights reserved.