Micellar electrokinetic capillary chromatography for the determination of fluoxetine and its metabolite norfluoxetine in biological fluids

A micellar electrokinetic capillary chromatography (MEKC) for determining fluoxetine and its metabolite (norfluoxetine) is proposed. Optimal conditions for the quantitative separation were investigated. A background electrolyte solution consisting of 5 mM phosphate buffer adjusted to pH 12.3 and 40 mM of 1-decanesulfonic acid sodium salt (DSS), hydrodynamic injection and 25 kV of separation voltage were used. Good linearity and precision were obtained for both compounds. Detection limits of 0.2 mg/l for fluoxetine and norfluoxetine were obtained. The developed method is rapid and it has been applied to determine fluoxetine and its metabolite in human serum and urine. The samples were purified and enriched by means of extraction-preconcentration step with a preconditioned C18 cartridge and eluting the compounds with methanol.     

Organic matter transformations and soil fertility in a treed pasture in semiarid NE Brazil

Planted silvo-pastoral systems are formed by sparing selected native trees when land is cleared for pasture establishment, or by planting selected species – often known agroforestry species – into the establishing pasture. Isolated trees within pastures and savannas are often associated with `resource islands', characterized by higher fertility and organic matter levels under the tree canopies. We here examine the processes underlying the differences in fertility and organic matter in a buffel grass (Cenchrus ciliaris L.) pasture that contained two tree species (Ziziphus joazeiro Mart., Spondias tuberosa Arruda Cam.) preserved from the native thorn forest and a planted agroforestry species (Prospois juliflora Swartz D.C). The objective is to distinguish effects of soil variability from those induced by the presence of trees or the planting of pasture. The ¹³C signatures of the original (largely C3) vegetation, the preserved and planted trees, and the planted C4 grass were used to distinguish the provenance of organic matter in the top soil (0–15 cm). This allowed the conclusion that all trees maintained C3 derived C at the original thorn forest level, while lower levels under pasture were due to mineralisation of organic matter. The net rates of forest-derived C loss under pasture varied with soil type amounting to between 25 and 50% in 13 years after pasture establishment. Only on Alfisol, C inputs from the pasture compensated for the C3-C losses. Analysis of organic and inorganic P fractions indicated Z. joazeiro and P. juliflora enriched the soil under their canopy with P, whereas S. tuberosa had no positive effect on fertility. A combination of ANOVA and spatial analysis and mapping was used to show vegetation effects.     

How can malaria rapid diagnostic tests achieve their potential? A qualitative study of a trial at health facilities in Ghana

Malaria is still a major public health problem in Brazil, with approximately 306 000 registered cases in 2009, but it is estimated that in the early 1940s, around six million cases of malaria occurred each year. As a result of the fight against the disease, the number of malaria cases decreased over the years and the smallest numbers of cases to-date were recorded in the 1960s. From the mid-1960s onwards, Brazil underwent a rapid and disorganized settlement process in the Amazon and this migratory movement led to a progressive increase in the number of reported cases. Although the main mosquito vector (Anopheles darlingi) is present in about 80% of the country, currently the incidence of malaria in Brazil is almost exclusively (99,8% of the cases) restricted to the region of the Amazon Basin, where a number of combined factors favors disease transmission and impair the use of standard control procedures. Plasmodium vivax accounts for 83,7% of registered cases, while Plasmodium falciparum is responsible for 16,3% and Plasmodium malariae is seldom observed. Although vivax malaria is thought to cause little mortality, compared to falciparum malaria, it accounts for much of the morbidity and for huge burdens on the prosperity of endemic communities. However, in the last few years a pattern of unusual clinical complications with fatal cases associated with P. vivax have been reported in Brazil and this is a matter of concern for Brazilian malariologists. In addition, the emergence of P. vivax strains resistant to chloroquine in some reports needs to be further investigated. In contrast, asymptomatic infection by P. falciparum and P. vivax has been detected in epidemiological studies in the states of Rondonia and Amazonas, indicating probably a pattern of clinical immunity in both autochthonous and migrant populations. Seropidemiological studies investigating the type of immune responses elicited in naturally-exposed populations to several malaria vaccine candidates in Brazilian populations have also been providing important information on whether immune responses specific to these antigens are generated in natural infections and their immunogenic potential as vaccine candidates. The present difficulties in reducing economic and social risk factors that determine the incidence of malaria in the Amazon Region render impracticable its elimination in the region. As a result, a malaria-integrated control effort - as a joint action on the part of the government and the population - directed towards the elimination or reduction of the risks of death or illness, is the direction adopted by the Brazilian government in the fight against the disease.     

Mapping of citrullinated fibrinogen B-cell epitopes in rheumatoid arthritis by imaging surface plasmon resonance

Introduction  

Rheumatoid arthritis (RA) frequently involves the loss of tolerance to citrullinated antigens, which may play a role in pathogenicity. Citrullinated fibrinogen is commonly found in inflamed synovial tissue and is a frequent target of autoantibodies in RA patients. To obtain insight into the B-cell response to citrullinated fibrinogen in RA, its autoepitopes were systematically mapped using a new methodology.     

Photoreactivation and dark repair in UV-treated microorganisms: effect of temperature

Because of the lack of readily available information about the influence of temperature on microorganism reactivation processes subsequent to inactivation with UV radiation, a series of batch reactivation studies were performed at 5, 10, 15, 20, 25, and 30 degrees C. A special effort was made to model the reactivation process to enable the effect of the temperature variable to be quantified. Because an earlier-proposed kinetic model (K. Kashimada, N. Kamiko, K. Yamamoto, and S. Ohgaki, Water Sci. Technol. 33:261-269, 1996), a first-order saturation type, does not adequately fit the data obtained in experiments of reactivation in conditions of light and darkness, a modification of that model is proposed. The new model, which actually coincides with the classical logistic equation, incorporates two kinetic parameters: the maximum survival ratio (Sm) and the second-order reactivation rate constant (k2). In order to interpret correctly the reactivation occurring in conditions of darkness, a new term for the decay is added to the logistic equation. The new model accurately fits the data obtained in reactivation experiments, permitting the interpretation of the kinetic parameters Sm, k2, and M (for only repair in darkness), where M is mortality, a zero-order decay rate constant, and their relationship with various environmental conditions, such as microbial type, light, and temperature. The parameters Sm and k2 (and M for reactivation in conditions of darkness) show exponential dependence on the reactivating temperature, and it is possible to predict their values and hence the reactivation curve from the equations proposed in this work.     

Assessment of algorithms for high throughput detection of genomic copy number variation in oligonucleotide microarray data

Background

Genomic deletions and duplications are important in the pathogenesis of diseases, such as cancer and mental retardation, and have recently been shown to occur frequently in unaffected individuals as polymorphisms. Affymetrix GeneChip whole genome sampling analysis (WGSA) combined with 100 K single nucleotide polymorphism (SNP) genotyping arrays is one of several microarray-based approaches that are now being used to detect such structural genomic changes. The popularity of this technology and its associated open source data format have resulted in the development of an increasing number of software packages for the analysis of copy number changes using these SNP arrays.

Results

We evaluated four publicly available software packages for high throughput copy number analysis using synthetic and empirical 100 K SNP array data sets, the latter obtained from 107 mental retardation (MR) patients and their unaffected parents and siblings. We evaluated the software with regards to overall suitability for high-throughput 100 K SNP array data analysis, as well as effectiveness of normalization, scaling with various reference sets and feature extraction, as well as true and false positive rates of genomic copy number variant (CNV) detection.

Conclusion

We observed considerable variation among the numbers and types of candidate CNVs detected by different analysis approaches, and found that multiple programs were needed to find all real aberrations in our test set. The frequency of false positive deletions was substantial, but could be greatly reduced by using the SNP genotype information to confirm loss of heterozygosity.     

Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis

Recent studies have implicated the dying cell as a potential reservoir of modified autoantigens that might initiate and drive systemic autoimmunity in susceptible hosts. A number of subunits of the exosome, a complex of 3'→5' exoribonucleases that functions in a variety of cellular processes, are recognized by the so-called anti-PM/Scl autoantibodies, found predominantly in patients suffering from an overlap syndrome of myositis and scleroderma. Here we show that one of these subunits, PM/Scl-75, is cleaved during apoptosis. PM/Scl-75 cleavage is inhibited by several different caspase inhibitors. The analysis of PM/Scl-75 cleavage by recombinant caspase proteins shows that PM/Scl-75 is efficiently cleaved by caspase-1, to a smaller extent by caspase-8, and relatively inefficiently by caspase-3 and caspase-7. Cleavage of the PM/Scl-75 protein occurs in the C-terminal part of the protein at Asp369 (IILD³⁶⁹↓G), and at least a fraction of the resulting N-terminal fragments of PM/Scl-75 remains associated with the exosome. Finally, the implications of PM/Scl-75 cleavage for exosome function and the generation of anti-PM/Scl-75 autoantibodies are discussed.     

Similar hypotensive responses to resistance exercise with and without blood flow restriction

Low intensity resistance exercise (RE) with blood flow restriction (BFR) has gained attention in the literature due to the beneficial effects on functional and morphological variables, similar to those observed during traditional RE without BFR, while the effects of BFR on post-exercise hypotension remain unclear. The aim of the present study was to compare the blood pressure (BP) response of trained normotensive individuals to RE with and without BFR. In this cross-over randomized trial, eight male subjects (23.8 ± 4 years, 74 ± 3 kg, 174 ± 4 cm) completed two exercise protocols: traditional RE (3 x 10 repetitions at 70% one-repetition maximum [1-RM]) and low intensity RE (3 x 15 repetitions at 20% 1-RM) with BFR. Blood pressure measurements were performed after 15 min of seated rest (0), immediately after and 10 min, 20 min, 30 min, 40 min, 50 min and 60 min after the experimental sessions. Similar hypotensive effects for systolic BP (SBP) were observed for both protocols (P < 0.05) after exercise, with no differences between groups (P > 0.05) and no statistically significant difference for diastolic BP (P > 0.05). These results suggest that in normotensive trained individuals, both traditional RE and RE with BFR induce hypotension for SBP, which is important to prevent cardiovascular disturbances.