Purificação e caracterização parcial de duas n-acetil-B-hexosaminidases do equinoderma marinho echinometra lucunter

Two b-N-acetylhexosaminidases (F11 e F15) were purified from Echinometra lucunter gonads extracts. The purified enzymes were obtained using ammonium sulfate fractionation, followed by gel filtration chromatographies (Sephacryl S-200, Sephadex G-75 and Sephacryl S-200). The F11 fraction was purified 192.47 -fold with a 28.5% yield, and F15 fraction 85.41 -fold with a 32.3% yield. The molecular weights of the fractions were 116 kDa for F11 and 42 kDa for F15 using SDS-PAGE. In Sephacryl S-200, F15 was 84 kDa, indicating that it is a dimeric protein. When p-nitrophenyl-β-D-glycosaminide was used as substrate, we determined an apparent Km of 0.257 mM and Vmax of 0.704 for F11 and for F15 the Km was 0.235 mM and Vmax of 0.9 mM of product liberated by hour. Both enzymes have optimum pH and temperature respectively at 5.0 and 45 °C. The enzymes showed inhibition by silver nitrate, while the glucuronic acid was a potent activator. The high inhibition of F15 by N-etylmaleimide indicates that sulphydril groups are involved in the catalysis of synthetic substrate

Análise in vitro da atividade antifúngica e de toxicidade da anfotericina B pré-aquecida

The aim of this work was to evaluate how an aqueous micellar system containing Amphotericin B (AmB) and sodium deoxycholate (DOC) can be rebuilt after heating treatment. Also a review of the literature about the new physicochemical and biological properties of this new system was carried out. Afterwards, heated (AmB-DOC-H) and unheated (AmB-DOC) micelles were subsequently diluted at four different concentrations (50mg.L-1, 5mg.L-1, 0.5mg.L-1 and 0.05mg.L-1) to perform the physicochemical study and, then, the pharmacotoxicity assay, in which two cell models were used for the in vitro experiments, Red Blood Cells (RBC) from human donors and Candida parapisilosis (Cp). While potassium (K+) and hemoglobin leakage from RBC were the used parameters to evaluate the acute and chronic toxicity, respectively, the efficacy of AmB-DOC and AmB-DOC-H were assessed by K+ leakage and cell survival rate from Cp. The spectral study revealed a slight change on the aggregate peak from 327nm to 323nm for AmB-DOC-H compared to AmB-DOC. Concerning the toxicity, although AmB-DOC and AmB-DOC-H presented different behavior for hemoglobin leakage, AmB-DOC produced higher leakage than AmB-DOC-H at high concentrations (from 5mg.L-1) with values tending to zero. However, concerning K+ leakage, both AmB-DOC and AmB-DOC-H, showed similar profile for both cell models, RBC and Cp (p<0,05). AmB-DOC-H and AmB-DOC also revealed similar profile of activity against Cp with equivalent survival rate. In short, the AmB-DOC-H showed much less toxicity than AmB-DOC, but remained as active as the late one against fungal cell. Therefore, the results highlight the importance of this new procedure as a simple, inexpensive and safe alternative to produce a new kind of micelle system for treatment of systemic fungal infections

A dialética do amor em pigmaleão, de G. B. Shaw.

Pygmalion (1913), by George Bernard Shaw (1856-1950), has many studies in literary criticism. However, this study brings a new interpretation to Shaw s play based on Harold Bloom s theory and methodology, that is, the anxiety of influence and the dialectic of revisionism. Through the analysis of poetic influence and the dialectic of love, we can see that Pygmalion represents an apophrades in relation to William Shakespeare s The Taming of the Shrew (1593) and Ovid s myth of Pygmalion and Galatea in Metamorphosis (c. 14), which creates a family romance between the three stories. Shaw s play surpasses The Taming of the Shrew when it shows the possibility of the relation between this parent poem and Ovid s myth, which it is also its parent poem, and because it represents a strong misreading of Shakespeare s play as well as of Ovid s myth.

BSmart: desenvolvimento rigoroso de aplicações Java Card com base no método formal B

Java Card technology allows the development and execution of small applications embedded in smart cards. A Java Card application is composed of an external card client and of an application in the card that implements the services available to the client by means of an Application Programming Interface (API). Usually, these applications manipulate and store important information, such as cash and confidential data of their owners. Thus, it is necessary to adopt rigor on developing a smart card application to improve its quality and trustworthiness. The use of formal methods on the development of these applications is a way to reach these quality requirements. The B method is one of the many formal methods for system specification. The development in B starts with the functional specification of the system, continues with the application of some optional refinements to the specification and, from the last level of refinement, it is possible to generate code for some programming language. The B formalism has a good tool support and its application to Java Card is adequate since the specification and development of APIs is one of the major applications of B. The BSmart method proposed here aims to promote the rigorous development of Java Card applications up to the generation of its code, based on the refinement of its formal specification described in the B notation. This development is supported by the BSmart tool, that is composed of some programs that automate each stage of the method; and by a library of B modules and Java Card classes that model primitive types, essential Java Card API classes and reusable data structures

Formal verification of PLC programs using the B Method

PLCs (acronym for Programmable Logic Controllers) perform control operations, receiving information from the environment, processing it and modifying this same environment according to the results produced. They are commonly used in industry in several applications, from mass transport to petroleum industry. As the complexity of these applications increase, and as various are safety critical, a necessity for ensuring that they are reliable arouses. Testing and simulation are the de-facto methods used in the industry to do so, but they can leave flaws undiscovered. Formal methods can provide more confidence in an application s safety, once they permit their mathematical verification. We make use of the B Method, which has been successfully applied in the formal verification of industrial systems, is supported by several tools and can handle decomposition, refinement, and verification of correctness according to the specification. The method we developed and present in this work automatically generates B models from PLC programs and verify them in terms of safety constraints, manually derived from the system requirements. The scope of our method is the PLC programming languages presented in the IEC 61131-3 standard, although we are also able to verify programs not fully compliant with the standard. Our approach aims to ease the integration of formal methods in the industry through the abbreviation of the effort to perform formal verification in PLCs