Development and validation of an LC-MS/MS method for quantitative analysis of mirtazapine in human plasma

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

AN EFFICIENT HPLC-UV METHOD FOR THE QUANTITATIVE DETERMINATION OF CEFADROXIL IN HUMAN PLASMA AND ITS APPLICATION IN PHARMACOKINETIC STUDIES

Cefadroxil is a semi-synthetic first-generation oral cephalosporin used in the treatment of mild to moderate infections of the respiratory and urinary tracts, skin and soft tissue infections. In this work a simple, rapid, economic and sensitive HPLC-UV method is described for the quantitative determination of cefadroxil in human plasma samples using lamivudine as internal standard. Sample pre-treatment was accomplished through protein precipitation with acetonitrile and chromatographic separation was performed with a mobile phase consisting of a mixture of sodium dihydrogen phosphate monohydrate solution, methanol and acetonitrile in the ratio of 90:8:2 (v/v/v) at a flow rate of 1.0mL/min. The proposed method is linear between 0.4 to 40.0 mu g/mL and its average recovery is 102.21% for cefadroxil and 97.94% for lamivudine. The method is simple, sensitive, reproducible, less time consuming for determination of cefadroxil in human plasma. The method can therefore be recommended for pharmacokinetics studies, including bioavailability and bioequivalence studies.

A new method to monitor visual field defects caused by photoreceptor degeneration by quantitative optical coherence tomography

PURPOSE: To correlate the dimension of the visual field (VF) tested by Goldman kinetic perimetry with the extent of visibility of the highly reflective layer between inner and outer segments of photoreceptors (IOS) seen in optical coherence tomography (OCT) images in patients with retinitis pigmentosa (RP). METHODS: In a retrospectively designed cross-sectional study, 18 eyes of 18 patients with RP were examined with OCT and Goldmann perimetry using test target I4e and compared with 18 eyes of 18 control subjects. A-scans of raw scan data of Stratus OCT images (Carl Zeiss Meditec, AG, Oberkochen, Germany) were quantitatively analyzed for the presence of the signal generated by the highly reflective layer between the IOS in OCT images. Starting in the fovea, the distance to which this signal was detectable was measured. Visual fields were analyzed by measuring the distance from the center point to isopter I4e. OCT and visual field data were analyzed in a clockwise fashion every 30 degrees , and corresponding measures were correlated. RESULTS: In corresponding alignments, the distance from the center point to isopter I4e and the distance to which the highly reflective signal from the IOS can be detected correlate significantly (r = 0.75, P < 0.0001). The greater the distance in VF, the greater the distance measured in OCT. CONCLUSIONS: The authors hypothesize that the retinal structure from which the highly reflective layer between the IOS emanates is of critical importance for visual and photoreceptor function. Further research is warranted to determine whether this may be useful as an objective marker of progression of retinal degeneration in patients with RP.

Quantitative myocardial contrast echocardiography: a new method for the non-invasive detection of chronic heart transplant rejection

Chronic heart transplant rejection, i.e. cardiac allograft vasculopathy (CAV) is a major adverse prognostic factor after heart transplantation (HTx). This study tested the hypothesis that the relative myocardial blood volume (rBV) as quantified by myocardial contrast echocardiography accurately detects severe CAV as defined by coronary intravascular ultrasound (IVUS).

A quantitative phenytoin GC-MS method and it's validation for samples from human ex situ brain microdialysis, blood and saliva using solid-phase extraction

This study describes the development and validation of a gas chromatography-mass spectrometry (GC-MS) method to identify and quantitate phenytoin in brain microdialysate, saliva and blood from human samples. A solid-phase extraction (SPE) was performed with a nonpolar C8-SCX column. The eluate was evaporated with nitrogen (50°C) and derivatized with trimethylsulfonium hydroxide before GC-MS analysis. As the internal standard, 5-(p-methylphenyl)-5-phenylhydantoin was used. The MS was run in scan mode and the identification was made with three ion fragment masses. All peaks were identified with MassLib. Spiked phenytoin samples showed recovery after SPE of ≥94%. The calibration curve (phenytoin 50 to 1,200 ng/mL, n = 6, at six concentration levels) showed good linearity and correlation (r² > 0.998). The limit of detection was 15 ng/mL; the limit of quantification was 50 ng/mL. Dried extracted samples were stable within a 15% deviation range for ≥4 weeks at room temperature. The method met International Organization for Standardization standards and was able to detect and quantify phenytoin in different biological matrices and patient samples. The GC-MS method with SPE is specific, sensitive, robust and well reproducible, and is therefore an appropriate candidate for the pharmacokinetic assessment of phenytoin concentrations in different human biological samples.

Transconvolution and the virtual positron emission tomograph--a new method for cross calibration in quantitative PET∕CT imaging

PURPOSE Positron emission tomography (PET)∕computed tomography (CT) measurements on small lesions are impaired by the partial volume effect, which is intrinsically tied to the point spread function of the actual imaging system, including the reconstruction algorithms. The variability resulting from different point spread functions hinders the assessment of quantitative measurements in clinical routine and especially degrades comparability within multicenter trials. To improve quantitative comparability there is a need for methods to match different PET∕CT systems through elimination of this systemic variability. Consequently, a new method was developed and tested that transforms the image of an object as produced by one tomograph to another image of the same object as it would have been seen by a different tomograph. The proposed new method, termed Transconvolution, compensates for differing imaging properties of different tomographs and particularly aims at quantitative comparability of PET∕CT in the context of multicenter trials. METHODS To solve the problem of image normalization, the theory of Transconvolution was mathematically established together with new methods to handle point spread functions of different PET∕CT systems. Knowing the point spread functions of two different imaging systems allows determining a Transconvolution function to convert one image into the other. This function is calculated by convolving one point spread function with the inverse of the other point spread function which, when adhering to certain boundary conditions such as the use of linear acquisition and image reconstruction methods, is a numerically accessible operation. For reliable measurement of such point spread functions characterizing different PET∕CT systems, a dedicated solid-state phantom incorporating (68)Ge∕(68)Ga filled spheres was developed. To iteratively determine and represent such point spread functions, exponential density functions in combination with a Gaussian distribution were introduced. Furthermore, simulation of a virtual PET system provided a standard imaging system with clearly defined properties to which the real PET systems were to be matched. A Hann window served as the modulation transfer function for the virtual PET. The Hann's apodization properties suppressed high spatial frequencies above a certain critical frequency, thereby fulfilling the above-mentioned boundary conditions. The determined point spread functions were subsequently used by the novel Transconvolution algorithm to match different PET∕CT systems onto the virtual PET system. Finally, the theoretically elaborated Transconvolution method was validated transforming phantom images acquired on two different PET systems to nearly identical data sets, as they would be imaged by the virtual PET system. RESULTS The proposed Transconvolution method matched different PET∕CT-systems for an improved and reproducible determination of a normalized activity concentration. The highest difference in measured activity concentration between the two different PET systems of 18.2% was found in spheres of 2 ml volume. Transconvolution reduced this difference down to 1.6%. In addition to reestablishing comparability the new method with its parameterization of point spread functions allowed a full characterization of imaging properties of the examined tomographs. CONCLUSIONS By matching different tomographs to a virtual standardized imaging system, Transconvolution opens a new comprehensive method for cross calibration in quantitative PET imaging. The use of a virtual PET system restores comparability between data sets from different PET systems by exerting a common, reproducible, and defined partial volume effect.

Patterned library analysis: A method for the quantitative assessment of hypotheses concerning the determinants of protein structure

Site-directed mutagenesis and combinatorial libraries are powerful tools for providing information about the relationship between protein sequence and structure. Here we report two extensions that expand the utility of combinatorial mutagenesis for the quantitative assessment of hypotheses about the determinants of protein structure. First, we show that resin-splitting technology, which allows the construction of arbitrarily complex libraries of degenerate oligonucleotides, can be used to construct more complex protein libraries for hypothesis testing than can be constructed from oligonucleotides limited to degenerate codons. Second, using eglin c as a model protein, we show that regression analysis of activity scores from library data can be used to assess the relative contributions to the specific activity of the amino acids that were varied in the library. The regression parameters derived from the analysis of a 455-member sample from a library wherein four solvent-exposed sites in an α-helix can contain any of nine different amino acids are highly correlated (P < 0.0001, R2 = 0.97) to the relative helix propensities for those amino acids, as estimated by a variety of biophysical and computational techniques.

Laser-induced breakdown spectroscopy quantitative analysis method via adaptive analytical line selection and relevance vector machine regression model

Abstract A new LIBS quantitative analysis method based on analytical line adaptive selection and Relevance Vector Machine (RVM) regression model is proposed. First, a scheme of adaptively selecting analytical line is put forward in order to overcome the drawback of high dependency on a priori knowledge. The candidate analytical lines are automatically selected based on the built-in characteristics of spectral lines, such as spectral intensity, wavelength and width at half height. The analytical lines which will be used as input variables of regression model are determined adaptively according to the samples for both training and testing. Second, an LIBS quantitative analysis method based on RVM is presented. The intensities of analytical lines and the elemental concentrations of certified standard samples are used to train the RVM regression model. The predicted elemental concentration analysis results will be given with a form of confidence interval of probabilistic distribution, which is helpful for evaluating the uncertainness contained in the measured spectra. Chromium concentration analysis experiments of 23 certified standard high-alloy steel samples have been carried out. The multiple correlation coefficient of the prediction was up to 98.85%, and the average relative error of the prediction was 4.01%. The experiment results showed that the proposed LIBS quantitative analysis method achieved better prediction accuracy and better modeling robustness compared with the methods based on partial least squares regression, artificial neural network and standard support vector machine.

A laser induced breakdown spectroscopy quantitative analysis method based on the robust least squares support vector machine regression model

Data fluctuation in multiple measurements of Laser Induced Breakdown Spectroscopy (LIBS) greatly affects the accuracy of quantitative analysis. A new LIBS quantitative analysis method based on the Robust Least Squares Support Vector Machine (RLS-SVM) regression model is proposed. The usual way to enhance the analysis accuracy is to improve the quality and consistency of the emission signal, such as by averaging the spectral signals or spectrum standardization over a number of laser shots. The proposed method focuses more on how to enhance the robustness of the quantitative analysis regression model. The proposed RLS-SVM regression model originates from the Weighted Least Squares Support Vector Machine (WLS-SVM) but has an improved segmented weighting function and residual error calculation according to the statistical distribution of measured spectral data. Through the improved segmented weighting function, the information on the spectral data in the normal distribution will be retained in the regression model while the information on the outliers will be restrained or removed. Copper elemental concentration analysis experiments of 16 certified standard brass samples were carried out. The average value of relative standard deviation obtained from the RLS-SVM model was 3.06% and the root mean square error was 1.537%. The experimental results showed that the proposed method achieved better prediction accuracy and better modeling robustness compared with the quantitative analysis methods based on Partial Least Squares (PLS) regression, standard Support Vector Machine (SVM) and WLS-SVM. It was also demonstrated that the improved weighting function had better comprehensive performance in model robustness and convergence speed, compared with the four known weighting functions.

The possible selves method in evaluating youth development interventions: The use of Integrated Qualitative /Quantitative Data Analytic Strategies (IQ -DAS)

This study investigated the feasibility of using qualitative methods to provide empirical documentation of the long-term qualitative change in the life course trajectories of “at risk” youth in a school based positive youth development program (the Changing Lives Program—CLP). This work draws from life course theory for a developmental framework and from recent advances in the use of qualitative methods in general and a grounded theory approach in particular. Grounded theory provided a methodological framework for conceptualizing the use of qualitative methods for assessing qualitative life change. The study investigated the feasibility of using the Possible Selves Questionnaire-Qualitative Extension (PSQ-QE) for evaluating the impact of the program on qualitative change in participants' life trajectory relative to a non-intervention control group. Integrated Qualitative/Quantitative Data Analytic Strategies (IQ-DAS) that we have been developing a part of our program of research provided the data analytic framework for the study. ^ Change was evaluated in 85 at risk high school students in CLP high school counseling groups over three assessment periods (pre, post, and follow-up), and a non-intervention control group of 23 students over two assessment periods (pre and post). Intervention gains and maintenance and the extent to which these patterns of change were moderated by gender and ethnicity were evaluated using a mixed design Repeated Measures Multivariate Analysis of Variance (RMANOVA) in which Time (pre, post) was the within (repeated) factor and Condition, Gender, and Ethnicity the between group factors. The trends for the direction of qualitative change were positive from pre to post and maintained at the year-end follow-up. More important, the 3-way interaction for Time x Gender x Ethnicity was significant, Roy's Θ =. 205, F(2, 37) = 3.80, p <.032, indicating that the overall pattern of positive change was significantly moderated by gender and ethnicity. Thus, the findings also provided preliminary evidence for a positive impact of the youth development program on long-term change in life course trajectory, and were suggestive with respect to the issue of amenability to treatment, i.e., the identification of subgroups of individuals in a target population who are likely to be the most amenable or responsive to a treatment. ^

The use of quantitative polymerase chain reaction amplification method to determine sex ratio of human spermatozoa in semen and to determine maternal contamination of amniotic fluid samples

We have modified a technique which uses a single pair of primer sets directed against homologous but distinct genes on the X and Y chromosomes, all of which are coamplified in the same reaction tube with trace amounts of radioactivity. The resulting bands are equal in length, yet distinguishable by restriction enzyme sites generating two independent bands, a 364 bp X-specific band and a 280 bp Y-specific band. A standard curve was generated to show the linear relationship between X/Y ratio average vs. %Y or %X chromosomal content. Of the 51 purified amniocyte DNA samples analyzed, 16 samples showed evidence of high % X contamination while 2 samples demonstrated higher % Y than the expected 50% X and 50% Y chromosomal content. With regards to the 25 processed sperm samples analyzed, X-sperm enrichment was evident when compared to the primary sex ratio whereas Y-sperm was enriched when we compared before and after selection samples.

[factors That Influence Bone Mass Of Healthy Children And Adolescents Measured By Quantitative Ultrasound At The Hand Phalanges: A Systematic Review].

To analyze the main factors that influence bone mass in children and teenagers assessed by quantitative ultrasound (QUS) of the phalanges. A systematic literature review was performed according to the PRISMA method with searches in databases Pubmed/Medline, SciELO and Bireme for the period 2001-2012, in English and Portuguese languages, using the keywords: children, teenagers, adolescent, ultrasound finger phalanges, quantitative ultrasound of phalanges, phalangeal quantitative ultrasound. 21 articles were included. Girls had, in QUS, Amplitude Dependent Speed of Sound (AD-SoS) values higher than boys during pubertal development. The values of the parameters of QUS of the phalanges and dual-energy X-ray Absorptiometry (DXA) increased with the increase of the maturational stage. Anthropometric variables such as age, weight, height, body mass index (BMI), lean mass showed positive correlations with the values of QUS of the phalanges. Physical activity has also been shown to be positively associated with increased bone mass. Factors such as ethnicity, genetics, caloric intake and socioeconomic profile have not yet shown a conclusive relationship and need a larger number of studies. QUS of the phalanges is a method used to evaluate the progressive acquisition of bone mass during growth and maturation of individuals in school phase, by monitoring changes that occur with increasing age and pubertal stage. There were mainly positive influences in variables of sex, maturity, height, weight and BMI, with similar data when compared to the gold standard method, the DXA.