Design of a simple detection cell with extended optical path length for capillary electrophoresis: Application to multiresidue pesticide analysis

Absorbance detection in capillary electrophoresis (CE), offers an excellent mass sensitivity, but poor concentration detection limits owing to very small injection volumes (normally I to 10 nL). This aspect can be a limiting factor in the applicability of CE/UV to detect species at trace levels, particularly pesticide residues. In the present work, the optical path length of an on-column detection cell was increased through a proper connection of the column (75 mu m i.d.) to a capillary detection cell of 180 mu m optical path length in order to improve detectability. It is shown that the cell with an extended optical path length results in a significant gain in terms of signal to noise ratio. The effect of the increase in the optical path length has been evaluated for six pesticides, namely, carbendazim, thiabendazole, imazalil, procymidone triadimefon, and prochloraz. The resulting optical enhancement of the detection cell provided detection limits of ca. 0.3 mu g/mL for the studied compounds, thus enabling the residue analysis by CE/UV.

Determination of fluoroacetate and fluoride in blood serum by capillary zone electrophoresis using capacitively coupled contactless conductivity detection

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

Seasonal influence on biochemical profile and serum protein electrophoresis for Boa constrictor amarali in captivity

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

Viable human buccal mucosa cells do not yield typical nucleoids: Impacts on the single-cell gel electrophoresis/comet assay

Buccal mucosa (BM) cells have been used in human biomonitoring studies for detecting DNA adducts and chromosomal damage in an epithelial cell population. In the present study, we have investigated if human BM cells are suitable for use in the single-cell gel electrophoresis (SCGE)/Comet assay as an approach for estimating the exposure of epithelial cells to DNA-damaging agents. Our results indicate that only a few cells from BM cell samples yield comets that can be analyzed by current methods, and that the yield of cells with comets is independent of the percentage of viable BM cells in the sample. Data generated after enzymatic enrichment of viable cells and immunomagnetic separation of epithelial cells suggest that most of the BM cells that do form comets are probably leukocytes. Moreover, by reevaluating specific cells after running the Comet assay, we found that viable epithelial BM cells give rise to atypical comets that are not included in the analysis. Comparing DNA migration patterns between small groups of smokers and nonsmokers indicated that long-term smoking had no effect on the subpopulation of cells that yield typical comets. Our results indicate that the SCGE assay, as it is commonly performed, may not be useful for genotoxicity monitoring in human epithelial BM cells.

Disparity between Multilocus Enzyme Electrophoresis, Microsatellite Markers and Pulsed-Field Gel Electrophoresis in epidemiological tracking of Candida albicans

Various molecular systems are available for epidemiological, genetic, evolutionary, taxonomic and systematic studies of innumerable fungal infections, especially those caused by the opportunistic pathogen C. albicans. A total of 75 independent oral isolates were selected in order to compare Multilocus Enzyme Electrophoresis (MLEE), Electrophoretic Karyotyping (EK) and Microsatellite Markers (Simple Sequence Repeats - SSRs), in their abilities to differentiate and group C. albicans isolates (discriminatory power), and also, to evaluate the concordance and similarity of the groups of strains determined by cluster analysis for each fingerprinting method. Isoenzyme typing was performed using eleven enzyme systems: Adh, Sdh, M1p, Mdh, Idh, Gdh, G6pdh, Asd, Cat, Po, and Lap (data previously published). The EK method consisted of chromosomal DNA separation by pulsed-field gel electrophoresis using a CHEF system. The microsatellite markers were investigated by PCR using three polymorphic loci: EF3, CDC3, and HIS3. Dendrograms were generated by the SAHN method and UPGMA algorithm based on similarity matrices (S(SM)). The discriminatory power of the three methods was over 95%, however a paired analysis among them showed a parity of 19.7-22.4% in the identification of strains. Weak correlation was also observed among the genetic similarity matrices (S(SM)(MLEE) x S(SM)(EK) x S(SM)(SSRs)). Clustering analyses showed a mean of 9 +/- 12.4 isolates per cluster (3.8 +/- 8 isolates/taxon) for MLEE, 6.2 +/- 4.9 isolates per cluster (4 +/- 4.5 isolates/taxon) for SSRs, and 4.1 +/- 2.3 isolates per cluster (2.6 +/- 2.3 isolates/taxon) for EK. A total of 45 (13%), 39(11.2%), 5 (1.4%) and 3 (0.9%) clusters pairs from 347 showed similarity (Si) of 0.1-10%, 10.1-20%, 20.1-30% and 30.1-40%, respectively. Clinical and molecular epidemiological correlation involving the opportunistic pathogen C. albicans may be attributed dependently of each method of genotyping (i.e., MLEE, EK, and SSRs) supplemented with similarity and grouping analysis. Therefore, the use of genotyping systems that give results which offer minimum disparity, or the combination of the results of these systems, can provide greater security and consistency in the determination of strains and their genetic relationships. (C) 2010 Elsevier B.V. All rights reserved.

Evaluation of the antigenotoxicity of polysaccharides and beta-glucans from Agaricus blazei, a model study with the single cell gel electrophoresis/Hep G2 assay

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Molecular weight estimation of esterase isoenzymes in closely related Drosophila Species (Diptera: Drosophilidae) in non-denaturing polyacrylamide gel electrophoresis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Typing Candida albicans oral isolates from healthy Brazilian schoolchildren using multilocus enzyme electrophoresis reveals two highly polymorphic taxa

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

Agarose gel electrophoresis of cerebrospinal fluid proteins of dogs after sample concentration using a membrane microconcentrator technique

Background: Cerebrospinal fluid (CSF) is produced in the cerebral ventricles through ultrafiltration of plasma and active transport mechanisms. Evaluation of proteins in CSF may provide important information about the production of immunoglobulins within the central nervous system as well as possible disturbances in the blood-brain barrier. Objective: the objective of this study was to measure the concentration and fractions of protein in CSF samples using a membrane microconcentrator technique followed by electrophoresis, and to compare the protein fractions obtained with those in serum. Methods: CSF samples from 3 healthy dogs and 3 dogs with canine distemper virus infection were concentrated using a membrane microconcentrator having a 0.5 to 30,000 d nominal molecular weight limit (Ultrafree, Millipore, Billerica, MA, USA). Protein concentration was determined before and after concentration. Agarose gel electrophoresis was done on concentrated CSF samples, serum, and serial dilutions of one of the CSF samples. Results: Electrophoretic bands were clearly identified in densitometer tracings in CSF samples with protein concentrations as low as 1.3 g/dL. The higher CSF protein concentration in dogs with distemper was mainly the result of increased albumin concentration. Conclusion: the microconcentrating method used in this study enables characterization of the main protein fractions in CSF by routine electrophoresis and may be useful for interpreting the underlying cause of changes in CSF protein concentrations

Alternative methodologies for the determination of aldehydes by capillary electrophoresis

This paper describes 2 alternative methodologies for the determination of selected aldehydes (formaldehyde, acetaldehyde, propionaldehyde, acrolein, and benzaldehyde) by capillary electrophoresis (CE), the first approach is based on the formation of aldehyde-bisulfite adducts and employs free solution CE with reversed electroosmotic flow and indirect detection, using 10 mmol/L 3,5-dinitrobenzoic acid (pH 4.5) containing 0.2 mmol/L cetyltrimethylammonium bromide as the electrolyte. This novel methodology showed a fairly good sensitivity to concentration, with detection limits with respect to a single aldehyde on the order of 10-40 mu g/L, a reasonable analysis time (separation was achieved in <8 min), and no need for sample manipulation. A second approach was proposed in which 2,4-dinitrophenylhydrazine derivatives of the aldehydes were detected in a micellar electrolyte medium (20 mmol/L berate buffer containing 50 mmol/L sodium dodecyl sulfate and 15 mmol/L beta-cyclodextrin). This latter methodology included a laborious sample preconcentration step and showed much poorer sensitivity (0.5-2 mg/L detection limit, with respect to a single aldehyde), despite the use of sodium chloride to promote sample stacking. Both methodologies proved adequate to evaluate aldehyde levels in vehicular emissions. Samples from the tailpipe exhaust of a passenger car vehicle without a catalytic converter and operated with an ethanol-based fuel were collected and analyzed; the results showed high levels of formaldehyde and acetaldehyde (0.41-6.1 ppm, v/v). The concentrations estimated by the 2 methodologies, which were not in good agreement, suggest the possibility of striking differences in sample collection efficiency, which was not the concern of this work.

Indirect determination of chloride and sulfate ions in alcohol fuel by capillary electrophoresis

A capillary zone electrophoresis method using indirect UV detection for the analysis of chloride and sulfate in alcohol fuel samples was developed. The anions were analyzed in less than 3 min using an electrolyte containing 10 mmol 1(-1) chromate and 0.75 mmol 1(-1) hexamethonium bromide (HMB) as electroosmotic flow modifier. Coefficients of variation were better than 0.6% for migration time (n = 10) and between 2.05 and 2.82% for peak area repeatabilities. Analytical curves of peak area versus concentration in the range of 0.065-0.65 mg kg(-1) for chloride and 0.25-4.0 mg kg(-1) for sulfate were linear with coefficients of correlation higher than 0.9996. The limits of detection for sulfate and chloride were 0.033 and 0.041 mg kg(-1), respectively. Recovery values ranged from 85 to 103%. The method was successfully applied for the quantification of sulfate and chloride in five alcohol fuel samples. The concentration of sulfate varied from 0.45 to 3.12 mg kg(-1). Chloride concentrations were below the method's LOD.

Seasonal influence on hematologic values and hemoglobin electrophoresis in Brazilian Boa constrictor amarali

As ectothermic animals, snakes depend exclusively on the environment for proper temperature maintenance, which may greatly influence their activity. Twenty-five adult Boa constrictor amarali snakes maintained in captivity were used to determine the influence of seasons on their hematologic values and electrophoretic profile of hemoglobin. A complete blood cell count (CBC) and examination for hemoparasites were performed in the summer and winter of 2004. Hemoglobin was stored for later electrophoresis. Significant differences (P < 0.05) were obtained in RBC, WBC. lymphocyte, thrombocyte, and monocyte counts, demonstrating the importance of the period of the year in the interpretation of reference values in these animals. Two snakes were detected with blood parasites (Hepatozoon sp.) in the winter and four in the summer. although it appears that their presence did not cause any significant alterations in the CBC. The electrophoretic analysis of the samples demonstrated two-four hemoglobin bands in this species.