Evaluation of comprehensive two-dimensional gas chromatography coupled to rapid scanning quadrupole mass spectrometry for quantitative analysis

Comprehensive two-dimensional gas chromatography (GC x GC) is a powerful technique that provides excellent separation and identification of analytes in highly complex samples with considerable increase in GC peak capacities. However, since second dimension analyses are very fast, detectors with a rapid acquisition rate are required. Over the last years, quite a number of studies have discussed the potential and limitations of the combination GC x GC with a variety of quadrupole mass spectrometers. The present research focuses on the evaluation of qMS effectiveness at a 10,000-amu/s scan speed and 20-Hz scan frequency for the identification (full scan mode acquisition-TIC) and quantification (extracted ion chromatogram) of target pesticide residues in tomato samples. The following MS parameters have been evaluated: number of data points per peak, mass spectrum quality, peak skewing, and sensitivity. The validated proposed GC x GC/qMS method presented satisfactory results in terms of repeatability (coefficient of variation lower than 15%), accuracy (84-117%), and linearity (ranging from 25 to 500 ng/g), while significant enhancement in sensitivity was observed (a factor of around 10) under scan conditions. (C) 2012 Elsevier B.V. All rights reserved.

A graphical tool for an analytical approach of scattering photons by the Compton effect

The photons scattered by the Compton effect can be used to characterize the physical properties of a given sample due to the influence that the electron density exerts on the number of scattered photons. However, scattering measurements involve experimental and physical factors that must be carefully analyzed to predict uncertainty in the detection of Compton photons. This paper presents a method for the optimization of the geometrical parameters of an experimental arrangement for Compton scattering analysis, based on its relations with the energy and incident flux of the X-ray photons. In addition, the tool enables the statistical analysis of the information displayed and includes the coefficient of variation (CV) measurement for a comparative evaluation of the physical parameters of the model established for the simulation. (C) 2012 Elsevier B.V. All rights reserved.

Evaluation of Solid-Phase Microextraction using a Polythiophene Film and Liquid Chromatography with Spectrophotometric Detection for the Determination of Antidepressants in Plasma Samples

Polythiophene (PTh) phase electropolymerized on the stainless steel wire was evaluated as solid-phase microextraction (SPME), and analysis by liquid chromatography with spectrophotometric detection (LC-UV) for determination of new-generation antidepressants, selective serotonin reuptake inhibitors (SSRIs) (citalopram, paroxetine, fluoxetine and sertraline), in plasma samples. The influence of electropolymerization variables (scan rate, potential range and scan cycles) was evaluated on SPME performance. The SPME variables (extraction time, temperature, matrix pH, ionic strength and desorption procedure), as well as the influence of plasma proteins on sorption mechanisms were also evaluated. The SPME/LC-UV method developed for determination of antidepressants in plasma sample presented a linear range between the limit of quantification (LOQ, 200-250 ng mL(-1)) to 4000 ng mL(-1), and interday precision with coefficient of variation (CV) ranged from 11 to 15%. The proposed method can be a useful tool for the determination of antidepressants in human plasma samples in urgent toxicological analysis after the accidental or suicidal intake of higher doses of medications.

Isokinetic analysis of the elbow joint in children with cerebral palsy

The joint torque is an important variable related to children with cerebral palsy. The present study analyzed kinetic parameters during elbow flexion and extension movements in healthy and cerebral palsy children. Ten healthy and 10 cerebral palsy children participated of the study. An isokinetic dynamometer was used to measure the elbow mean peak torque, mean angle peak torque, coefficient of variation and acceleration during flexion and extension movements at different angular speeds. The mean peak torque on extension movement in healthy children group was significant higher compared to the cerebral palsy group. The coefficient of variation on both flexion and extension movements was significantly higher in cerebral palsy group. However there were significantly difference on both groups compared the lowest and highest velocities. Although the results showed no difference in flexor peak torque, the acceleration is significantly lower in lowest and highest angular velocity.

Influence of fast and slow distillation on ethyl carbamate content and on coefficient of non-alcohol components in Brazilian sugarcane spirits

Ethyl carbamate is an impurity present in distilled beverages. Given the risk of it being a carcinogenic substance, Brazilian legislation has determined that its presence in distilled beverages, such as 'aguardente' and 'cachaca' (two types of sugarcane spirits), should be limited to a maximum of 150 mu g/L. Ordinary spirits usually contain variable amounts of ethyl carbamate, although in lower concentrations than the maximum determined by law. The finding that commercial spirits had a much lower concentration of this impurity (around 50 mu g/L) led the authors to research the reasons for the differences, and these are explored in this paper, with a focus on the speed of the distillation process and its influence on the spirit's composition. The team conducted research in a sugarcane distillery producing 'aguardente' using a simple pot still and measured the influence of fast and slow distillation on the presence of ethyl carbamate and non-alcohol components in the process. The results demonstrated that the speed of distillation was proportionally related to the concentration of ethyl carbamate and secondary components in the beverage's composition. Copyright (c) 2012 The Institute of Brewing & Distilling

Simultaneous Quantification of Lycopene, beta-Carotene, Retinol and alpha-Tocopherol in Plasma after a Simple Extraction Procedure: Stability Study and Application to Human Volunteers

A method for the simultaneous quantification of lycopene, beta-carotene, retinol and alpha-tocopherol by high-performance liquid chromatography (HPLC) with Vis/fluorescence detection with isocratic elution was optimized and validated. The method consists of a rapid and simple liquid-liquid extraction procedure and a posterior quantification of extracted supernatants by HPLC. Aliquots of plasma were stored at -20 degrees C for three months for stability study. The methodology was applied to samples from painters and individuals not exposed to paints (n = 75). The assay was linear for all vitamins (r > 0.99). Intra-and inter-run precisions were obtained with coefficient of variation smaller than 5%. The accuracies ranged from 0.29 to -5.80% and recoveries between 92.73 and 101.97%. Plasma samples and extracted supernatants were stable for 60 days at -20 degrees C. A significant decrease of lycopene, beta-carotene and retinol concentrations in plasma from exposed individuals compared to non-exposed individuals (p < 0.05) was observed. The method is simple, reproducible, precise, accurate and sensitive, and can be routinely utilized in clinical laboratories.

Application of the Z-scan technique to determine the optical Kerr coefficient and two-photon absorption coefficient of magnetite nanoparticles colloidal suspension

We investigate the occurrence of the optical Kerr effect and two-photon absorption when an oil-based magnetic Fe3O4 nanoparticles colloidal suspension is illuminated with high intensity femtosecond laser pulses. The frequency of the pulses is controlled and the Z-scan technique is employed in our measurements of the nonlinear optical Kerr coefficient (n(2)) and two-photon absorption coefficient (beta). From these values it was possible to calculate the real and imaginary parts of the third-order susceptibility. We observed that increasing the pulse frequency, additional physical processes take place, increasing artificially the absolute values of n(2) and beta. The experimental conditions are discussed to assure the obtention of reliable values of these nonlinear optical parameters, which may be useful in all-optical switching and optical power limiting applications. (C) 2012 American Institute of Physics. [http://dx.doi.org/10.1063/1.4723829]

Evaluation of reference-based two-color methods for measurement of gene expression ratios using spotted cDNA microarrays

Abstract Background Spotted cDNA microarrays generally employ co-hybridization of fluorescently-labeled RNA targets to produce gene expression ratios for subsequent analysis. Direct comparison of two RNA samples in the same microarray provides the highest level of accuracy; however, due to the number of combinatorial pair-wise comparisons, the direct method is impractical for studies including large number of individual samples (e.g., tumor classification studies). For such studies, indirect comparisons using a common reference standard have been the preferred method. Here we evaluated the precision and accuracy of reconstructed ratios from three indirect methods relative to ratios obtained from direct hybridizations, herein considered as the gold-standard. Results We performed hybridizations using a fixed amount of Cy3-labeled reference oligonucleotide (RefOligo) against distinct Cy5-labeled targets from prostate, breast and kidney tumor samples. Reconstructed ratios between all tissue pairs were derived from ratios between each tissue sample and RefOligo. Reconstructed ratios were compared to (i) ratios obtained in parallel from direct pair-wise hybridizations of tissue samples, and to (ii) reconstructed ratios derived from hybridization of each tissue against a reference RNA pool (RefPool). To evaluate the effect of the external references, reconstructed ratios were also calculated directly from intensity values of single-channel (One-Color) measurements derived from tissue sample data collected in the RefOligo experiments. We show that the average coefficient of variation of ratios between intra- and inter-slide replicates derived from RefOligo, RefPool and One-Color were similar and 2 to 4-fold higher than ratios obtained in direct hybridizations. Correlation coefficients calculated for all three tissue comparisons were also similar. In addition, the performance of all indirect methods in terms of their robustness to identify genes deemed as differentially expressed based on direct hybridizations, as well as false-positive and false-negative rates, were found to be comparable. Conclusion RefOligo produces ratios as precise and accurate as ratios reconstructed from a RNA pool, thus representing a reliable alternative in reference-based hybridization experiments. In addition, One-Color measurements alone can reconstruct expression ratios without loss in precision or accuracy. We conclude that both methods are adequate options in large-scale projects where the amount of a common reference RNA pool is usually restrictive.

Real-time PCR quantitation of glucocorticoid receptor alpha isoform

Abstract Background The expression of glucocorticoid-receptor (GR) seems to be a key mechanism in the regulation of glucocorticoid (GC) sensitivity and is potentially involved in cases of GC resistance or hypersensitivity. The aim of this study is to describe a method for quantitation of GR alpha isoform (GRα) expression using real-time PCR (qrt-PCR) with analytical capabilities to monitor patients, offering standard-curve reproducibility as well as intra- and inter-assay precision. Results Standard-curves were constructed by employing standardized Jurkat cell culture procedures, both for GRα and BCR (breakpoint cluster region), as a normalizing gene. We evaluated standard-curves using five different sets of cell culture passages, RNA extraction, reverse transcription, and qrt-PCR quantification. Intra-assay precision was evaluated using 12 replicates of each gene, for 2 patients, in a single experiment. Inter-assay precision was evaluated on 8 experiments, using duplicate tests of each gene for two patients. Standard-curves were reproducible, with CV (coefficient of variation) of less than 11%, and Pearson correlation coefficients above 0,990 for most comparisons. Intra-assay and inter-assay were 2% and 7%, respectively. Conclusion This is the first method for quantitation of GRα expression with technical characteristics that permit patient monitoring, in a fast, simple and robust way.

Residual stresses in Y-TZP crowns due to changes in the thermal contraction coefficient of veneers

Objective. To test the hypothesis that the difference in the coefficient of thermal contraction of the veneering porcelain above (˛liquid) and below (˛solid) its Tg plays an important role in stress development during a fast cooling protocol of Y-TZP crowns. Methods. Three-dimensional finite element models of veneered Y-TZP crowns were developed. Heat transfer analyses were conducted with two cooling protocols: slow (group A) and fast (groups B–F). Calculated temperatures as a function of time were used to determine the thermal stresses. Porcelain ˛solid was kept constant while its ˛liquid was varied, creating different ˛/˛solid conditions: 0, 1, 1.5, 2 and 3 (groups B–F, respectively). Maximum ( 1) and minimum ( 3) residual principal stress distributions in the porcelain layer were compared. Results. For the slowly cooled crown, positive 1 were observed in the porcelain, orientated perpendicular to the core–veneer interface (“radial” orientation). Simultaneously, negative 3 were observed within the porcelain, mostly in a hoop orientation (“hoop–arch”). For rapidly cooled crowns, stress patterns varied depending on ˛/˛solid ratios. For groups B and C, the patterns were similar to those found in group A for 1 (“radial”) and 3 (“hoop–arch”). For groups D–F, stress distribution changed significantly, with 1 forming a “hoop-arch” pattern while 3 developed a “radial” pattern. Significance. Hoop tensile stresses generated in the veneering layer during fast cooling protocols due to porcelain high ˛/˛solid ratio will facilitate flaw propagation from the surface toward the core, which negatively affects the potential clinical longevity of a crown.

Evaluation of solid-phase microextraction using a polythiophene film and liquid chromatography with spectrophotometric detection for the determination of antidepressants in plasma samples

Polythiophene (PTh) phase electropolymerized on the stainless steel wire was evaluated as solid-phase microextraction (SPME), and analysis by liquid chromatography with spectrophotometric detection (LC-UV) for determination of new-generation antidepressants, selective serotonin reuptake inhibitors (SSRIs) (citalopram, paroxetine, fluoxetine and sertraline), in plasma samples. The influence of electropolymerization variables (scan rate, potential range and scan cycles) was evaluated on SPME performance. The SPME variables (extraction time, temperature, matrix pH, ionic strength and desorption procedure), as well as the influence of plasma proteins on sorption mechanisms were also evaluated. The SPME/LC-UV method developed for determination of antidepressants in plasma sample presented a linear range between the limit of quantification (LOQ, 200-250 ng mL-1) to 4000 ng mL-1, and interday precision with coefficient of variation (CV) ranged from 11 to 15%. The proposed method can be a useful tool for the determination of antidepressants in human plasma samples in urgent toxicological analysis after the accidental or suicidal intake of higher doses of medications.

Simultaneous quantification of lycopene, β-carotene, retinol and α -tocopherol in plasma after a simple extraction procedure: stability study and application to human volunteers

A method for the simultaneous quantification of lycopene, β-carotene, retinol and α-tocopherol by high-performance liquid chromatography (HPLC) with Vis/fluorescence detection with isocratic elution was optimized and validated. The method consists of a rapid and simple liquid-liquid extraction procedure and a posterior quantification of extracted supernatants by HPLC. Aliquots of plasma were stored at -20°C for three months for stability study. The methodology was applied to samples from painters and individuals not exposed to paints (n = 75). The assay was linear for all vitamins (r > 0.99). Intra- and inter-run precisions were obtained with coefficient of variation smaller than 5%. The accuracies ranged from 0.29 to -5.80% and recoveries between 92.73 and 101.97%. Plasma samples and extracted supernatants were stable for 60 days at -20°C. A significant decrease of lycopene, β-carotene and retinol concentrations in plasma from exposed individuals compared to non-exposed individuals (p < 0.05) was observed. The method is simple, reproducible, precise, accurate and sensitive, and can be routinely utilized in clinical laboratories.

Variability of neurally-adjusted ventilatory assist (nava) compared to pressure support ventilation (psv) during the weaning phase of mechanical ventilation.

Rationale: NAVA is an assisted ventilatory mode that uses the electrical activity of the diaphragm (Edi) to trigger and cycle the ventilator, and to offer inspiratory assistance in proportion to patient effort. Since Edi varies from breath to breath, airway pressure and tidal volume also vary according to the patient's breathing pattern. Our objective was to compare the variability of NAVA with PSV in mechanically ventilated patients during the weaning phase. Methods: We analyzed the data collected for a clinical trial that compares PSV and NAVA during spontaneous breathing trials using PSV, with PS of 5 cmH2O, and NAVA, with Nava level titrated to generate a peak airway pressure equivalent to PSV of 5 cmH2O (NCT01137271). We captured flow, airway pressure and Edi at 100Hz from the ventilator using a dedicated software (Servo Tracker v2, Maquet, Sweden), and processed the cycles using a MatLab (Mathworks, USA) code. The code automatically detects the tidal volume (Vt), respiratory rate (RR), Edi and Airway pressure (Paw) on a breath-by-breath basis for each ventilatory mode. We also calculated the coefficient of variation (standard deviation, SD, divided by the mean). Results: We analyzed data from eleven patients. The mean Vt was similar on both modes (370 ±70 for Nava and 347± 77 for PSV), the RR was 26±6 for Nava and 26±7 or PSV. Paw was higher for Nava than for PSV (14±1 vs 11±0.4, p=0.0033), and Edi was similar for both modes (12±8 for Nava and 11±6 for PSV). The variability of the respiratory pattern, assessed with the coefficient of variation, was larger for Nava than for PSV for the Vt ( 23%±1% vs 15%±1%, p=0.03) and Paw (17%±1% vs 1% ±0.1%, p=0.0033), but not for RR (21% ±1% vs 16% ±8%, p=0.050) or Edi (33%±14% vs 39% ±16%,p=0.07). Conclusion: The variability of the breathing pattern is high during spontaneous breathing trials independent of the ventilatory mode. This variability results in variability of airway pressure and tidal volume, which are higher on Nava than on PSV. Our results suggest that Nava better reflects the normal variability of the breathing pattern during assisted mechanical ventilation.

Competing regression models for longitudinal data

The choice of an appropriate family of linear models for the analysis of longitudinal data is often a matter of concern for practitioners. To attenuate such difficulties, we discuss some issues that emerge when analyzing this type of data via a practical example involving pretestposttest longitudinal data. In particular, we consider log-normal linear mixed models (LNLMM), generalized linear mixed models (GLMM), and models based on generalized estimating equations (GEE). We show how some special features of the data, like a nonconstant coefficient of variation, may be handled in the three approaches and evaluate their performance with respect to the magnitude of standard errors of interpretable and comparable parameters. We also show how different diagnostic tools may be employed to identify outliers and comment on available software. We conclude by noting that the results are similar, but that GEE-based models may be preferable when the goal is to compare the marginal expected responses.

Análise isocinética da articulação do cotovelo em crianças com paralisia cerebral

A quantidade de torque aplicado na articulação é uma medida de aptidão física importante para crianças com paralisia cerebral. O presente estudo analisou parâmetros cinéticos na articulação do cotovelo em crianças saudáveis e com paralisia cerebral. Participaram 10 crianças com paralisia cerebral e 10 crianças sem comprometimento neurológico. Avaliou-se a média do pico de torque, média do ângulo do pico de torque, coeficiente de variação do torque e aceleração angular do movimento de flexo-extensão do cotovelo nas velocidades com um dinamômetro isocinético. A média de pico de torque (extensão), aceleração (flexão) e coeficiente de variação (flexão e extensão) são diferentes entre grupos. Conclui-se que o torque e aceleração sofreram interferências no movimento de flexo-extensão; as principais diferenças encontradas foram entre os extremos das velocidades; não houve diferenças no ângulo do pico de torque. A espasticidade não interferiu na força dos músculos agonistas do movimento de flexão da articulação do cotovelo.