Highly Sensitive Fluorescent Method for the Detection of Cholesterol Aldehydes Formed by Ozone and Singlet Molecular Oxygen

Cholesterol oxidation gives rise to a mixture of oxidized products. Different types of products are generated according to the reactive species being involved. Recently, attention has been focused on two cholesterol aldehydes, 3 beta-hydroxy-5 beta-hydroxy-B-norcholestane-6 beta-carboxyaldehyde (1a) and 3 beta-hydroxy-5-oxo-5,6-secocholestan-6-al (1b). These aldehydes can be generated by ozone-, as well as by singlet molecular oxygen-mediated cholesterol oxidation. It has been suggested that 1b is preferentially formed by ozone and la is preferentially formed by singlet molecular oxygen. In this study we describe the use of 1-pyrenebutyric hydrazine (PBH) as a fluorescent probe for the detection of cholesterol aldehydes. The formation of the fluorescent adduct between la with PBH was confirmed by HPLC-MS/MS. The fluorescence spectra of PBH did not change upon binding to the aldehyde. Moreover, the derivatization was also effective in the absence of an acidified medium, which is critical to avoid the formation of cholesterol aldehydes through Hock cleavage of 5 alpha-hydroperoxycholesterol. In conclusion, PBH can be used as an efficient fluorescent probe for the detection/quantification of cholesterol aldehydes in biological samples. Its analysis by HPLC coupled to a fluorescent detector provides a sensitive and specific way to quantify cholesterol aldehydes in the low femtomol range.

Flow injection analysis of ethyl xanthate by gas diffusion and UV detection as CS(2) for process monitoring of sulfide ore flotation

A sensitive and robust analytical method for spectrophotometric determination of ethyl xanthate, CH(3)CH(2)OCS(2)(-) at trace concentrations in pulp solutions from froth flotation process is proposed. The analytical method is based on the decomposition of ethyl xanthate. EtX(-), with 2.0 mol L(-1) HCl generating ethanol and carbon disulfide. CS(2). A gas diffusion cell assures that only the volatile compounds diffuse through a PTFE membrane towards an acceptor stream of deionized water, thus avoiding the interferences of non-volatile compounds and suspended particles. The CS(2) is selectively detected by UV absorbance at 206 nm (epsilon = 65,000 L mol(-1) cm(-1)). The measured absorbance is directly proportional to EtX(-) concentration present in the sample solutions. The Beer`s law is obeyed in a 1 x 10(-6) to 2 x 10(-4) mol L(-1) concentration range of ethyl xanthate in the pulp with an excellent correlation coefficient (r = 0.999) and a detection limit of 3.1 x 10(-7) mol L(-1), corresponding to 38 mu g L. At flow rates of 200 mu L min(-1) of the donor stream and 100 mu L min(-1) of the acceptor channel a sampling rate of 15 injections per hour could be achieved with RSD < 2.3% (n = 10, 300 mu L injections of 1 x 10(-5) mol L(-1) EtX(-)). Two practical applications demonstrate the versatility of the FIA method: (i) evaluation the free EtX(-) concentration during a laboratory study of the EtX(-) adsorption capacity on pulverized sulfide ore (pyrite) and (ii) monitoring of EtX(-) at different stages (from starting load to washing effluents) of a flotation pilot plant processing a Cu-Zn sulfide ore. (C) 2010 Elsevier By. All rights reserved.

Detection of Paraquat in Oral Fluid, Plasma, and Urine by Capillary Electrophoresis for Diagnosis of Acute Poisoning

FAPESP

Environmental formaldehyde analysis by active diffusive sampling with a bundle of polypropylene porous capillaries followed by capillary zone electrophoretic separation and contactless conductivity detection

Compared to other volatile carbonylic compounds present in outdoor air, formaldehyde (CH2O) is the most toxic, deserving more attention in terms of indoor and outdoor air quality legislation and control. The analytical determination of CH2O in air still presents challenges due to the low-level concentration (in the sub-ppb range) and its variation with sampling site and time. Of the many available analytical methods for carbonylic compounds, the most widespread one is the time consuming collection in cartridges impregnated with 2,4-dinitrophenylhydrazine followed by the analysis of the formed hydrazones by HPLC. The present work proposes the use of polypropylene hollow porous capillary fibers to achieve efficient CH2O collection. The Oxyphan (R) fiber (designed for blood oxygenation) was chosen for this purpose because it presents good mechanical resistance, high density of very fine pores and high ratio of collection area to volume of the acceptor fluid in the tube, all favorable for the development of air sampling apparatus. The collector device consists of a Teflon pipe inside of which a bundle of polypropylene microporous capillary membranes was introduced. While the acceptor passes at a low flow rate through the capillaries, the sampled air circulates around the fibers, impelled by a low flow membrane pump (of the type used for aquariums ventilation). The coupling of this sampling technique with the selective and quantitative determination of CH2O, in the form of hydroxymethanesulfonate (HMS) after derivatization with HSO3-, by capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-(CD)-D-4) enabled the development of a complete analytical protocol for the CH2O evaluation in air. (C) 2008 Published by Elsevier B.V.

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

Fluoroacetate is a highly toxic species naturally found in plants and in commercial products (compound 1080) for population control of several undesirable animal species. However, it is non-selective and toxic to many other animals including humans, and thus its detection is very important for forensic purposes. This paper presents a sensitive and fast method for the determination of fluoroacetate in blood serum using capillary electrophoresis with capacitively coupled contactless conductivity detection. Serum blood samples were treated with ethanol to remove proteins. The samples were analyzed in BGE containing 15 mmol/L histidine and 30 mmol/L gluconic acid (pH 3.85). The calibration curve was linear up to 75 mu mol/L (R(2) = 0.9995 for N = 12). The detection limit in the blood serum was 0.15 mg/kg, which is smaller than the lethal dose for humans and other animals. Fluoride, a metabolite of the fluoroacetate defluorination, could also be detected for levels greater than 20 mu mol/L, when polybrene was used for reversion of the EOF. CTAB and didecyldimethylammonium bromide are not useful for this task because of the severe reduction of the fluoride level. However, no interference was observed for fluoroacetate.

Determination of ciclopirox olamine in pharmaceutical products by capillary electrophoresis with capacitively coupled contactless conductivity detection

This paper describes the determination of ciclopirox olamine in pharmaceutical formulations using capillary electrophoresis with capacitively coupled contactless conductivity detection. In an alkaline medium, ciclopirox olamine is converted into an anionic species and its detection is possible in capillary electrophoresis with capacitively coupled contactless conductivity detection without an electroosmotic flow modifier, because it is a low-mobility species. A linear working range from 2.64 to 264 mu g/mL in sodium hydroxide electrolyte as well as low detection limit (0.39 mu g/mL) and a good repeatability (RSD = 3.4% for 264 mu g/mL ciclopirox solution (n = 10)) were achieved. It was also possible to determine olamine in its cationic form when acetic acid was used as the electrolyte solution. The results obtained include a linear range from 26.4 to 184.8 mu g/mL and a detection limit of 2.6 mu g/mL olamine. The proposed methods were applied to the analysis of commercial pharmaceutical products and the results were compared with the values indicated by the manufacturer as well as those obtained using a titrimetric method recommended by American Pharmacopoeia.

Improving the Detectability of Sequential Injection Chromatography (SIC): Determination of Triazines by Exploiting Liquid Core Waveguide (LCW) Detection

Coupling a liquid core waveguide cell to a sequential injection chromatograph improved the detection limits for determination of triazine herbicides without compromising peak resolution. Separation of simazine, atrazine, and propazine was achieved in water samples by a 25mm long C18 monolithic column. Detection was made at 238nm using a type II LCW (silica capillary coated with Teflon (R) AF2400) cell with 100cm of optical path length. Detection limits for simazine, atrazine, and propazine were 2.3, 1.9, and 4.5 mu g L-1, respectively. Reduced analysis time and low solvent consumption are other remarkable features of the proposed method.

Rapid method for the determination. of organic acids in wine by capillary electrophoresis with indirect UV detection

A capillary electrophoresis method for organic acids in wine was developed and validated. The optimal electrolyte consisted of 10 mmol/L 3,5-dinitrobenzoic acid (DNB) at pH 3.6 containing 0.2 mmol/L cetyltrimethylammonium bromide as flow reverser. DNB was chosen because it has an effective mobility similar to the organic acids under investigation, good buffering capacity at pH 3.6, and good chromophoric characteristics for indirect UV-absorbance detection at 254 nm. Sample preparation involved dilution and filtration. The method showed good performance characteristics: Linearity at 6 to 285 mg/L (r > 0.99); detection and quantification limits of 0.64 to 1.55 and 2.12 to 5.15 mg/L, respectively; separation time of less than 5.5 min. Coefficients of variation for ten injections were less than 5% and recoveries varied from 95% to 102%. Application to 23 samples of Brazilian wine confirmed good repeatability and demonstrated wide variation in the organic acid concentrations. (C) 2008 Elsevier Ltd. All rights reserved.

Evaluation of simulated bone lesion in the head of the mandible by using multislice computed tomography

Conventional radiography has shown limitation in acquiring image of the ATM region, thus, computed tomography (CT) scanning has been the best option to the present date for diagnosis, surgical planning and treatment of bone lesions, owing to its specific properties. OBJECTIVE: The aim of the study was to evaluate images of simulated bone lesions at the head of the mandible by multislice CT. MATERIAL AND METHODS: Spherical lesions were made with dental spherical drills (sizes 1, 3, and 6) and were evaluated by using multislice CT (64 rows), by two observers in two different occasions, deploying two protocols: axial, coronal, and sagittal images, and parasagittal images for pole visualization (anterior, lateral, posterior, medial and superior). Acquired images were then compared with those lesions in the dry mandible (gold standard) to evaluate the specificity and sensibility of both protocols. Statistical methods included: Kappa statistics, validity test and chi-square test. Results demonstrated the advantage of associating axial, coronal, and sagittal slices with parasagittal slices for lesion detection at the head of the mandible. RESULTS: There was no statistically significant difference between the types of protocols regarding a particular localization of lesions at the poles. CONCLUSIONS: Protocols for the assessment of the head of the mandible were established to improve the visualization of alterations of each of the poles of the mandible's head. The anterior and posterior poles were better visualized in lateral-medial planes while lateral, medial and superior poles were better visualized in the anterior-posterior plane.

Detection of human cytomegalovirus and Epstein-Barr virus in coronary atherosclerotic tissue

Previous studies indicated that patients with atherosclerosis are predominantly infected by human cytomegalovirus (HCMV), but rarely infected by type 1 Epstein-Barr virus (EBV-1). In this study, atheromas of 30 patients who underwent aortocoronary bypass surgery with coronary endartherectomy were tested for the presence of these two viruses. HCMV occurred in 93.3% of the samples and EBV-1 was present in 50% of them. Concurrent presence of both pathogens was detected in 43.3% of the samples.

Validity and reliability of methods for the detection of secondary caries around amalgam restorations in primary teeth

Secondary caries has been reported as the main reason for restoration replacement. The aim of this in vitro study was to evaluate the performance of different methods - visual inspection, laser fluorescence (DIAGNOdent), radiography and tactile examination - for secondary caries detection in primary molars restored with amalgam. Fifty-four primary molars were photographed and 73 suspect sites adjacent to amalgam restorations were selected. Two examiners evaluated independently these sites using all methods. Agreement between examiners was assessed by the Kappa test. To validate the methods, a caries-detector dye was used after restoration removal. The best cut-off points for the sample were found by a Receiver Operator Characteristic (ROC) analysis, and the area under the ROC curve (Az), and the sensitivity, specificity and accuracy of the methods were calculated for enamel (D2) and dentine (D3) thresholds. These parameters were found for each method and then compared by the McNemar test. The tactile examination and visual inspection presented the highest inter-examiner agreement for the D2 and D3 thresholds, respectively. The visual inspection also showed better performance than the other methods for both thresholds (Az = 0.861 and Az = 0.841, respectively). In conclusion, the visual inspection presented the best performance for detecting enamel and dentin secondary caries in primary teeth restored with amalgam.

A method to determine volatile contaminants in polyethylene terephthalate (PET) packages by HDC-GC-FID and its application to post-consumer materials

A simple and low cost method to determine volatile contaminants in post-consumer recycled PET flakes was developed and validated by Headspace Dynamic Concentration and Gas Chromatography-Flame Ionization Detection (HDC-GC-FID). The analytical parameters evaluated by using surrogates include: correlation coefficient, detection limit, quantification limit, accuracy, intra-assay precision, and inter-assay precision. In order to compare the efficiency of the proposed method to recognized automated techniques, post-consumer PET packaging samples collected in Brazil were used. GC-MS was used to confirm the identity of the substances identified in the PET packaging. Some of the identified contaminants were estimated in the post-consumer material at concentrations higher than 220 ng.g-1. The findings in this work corroborate data available in the scientific literature pointing out the suitability of the proposed analytical method.

Spatial pattern detection modeling of thrips (Thrips tabaci) on onion fields

Onion (Allium cepa) is one of the most cultivated and consumed vegetables in Brazil and its importance is due to the large laborforce involved. One of the main pests that affect this crop is the Onion Thrips (Thrips tabaci), but the spatial distribution of this insect, although important, has not been considered in crop management recommendations, experimental planning or sampling procedures. Our purpose here is to consider statistical tools to detect and model spatial patterns of the occurrence of the onion thrips. In order to characterize the spatial distribution pattern of the Onion Thrips a survey was carried out to record the number of insects in each development phase on onion plant leaves, on different dates and sample locations, in four rural properties with neighboring farms under different infestation levels and planting methods. The Mantel randomization test proved to be a useful tool to test for spatial correlation which, when detected, was described by a mixed spatial Poisson model with a geostatistical random component and parameters allowing for a characterization of the spatial pattern, as well as the production of prediction maps of susceptibility to levels of infestation throughout the area.

Detection of Brucella ovis in ovine from Paraíba State, in the Northeast region of Brazil

To determine the presence of Brucella ovis in ovine from Paraíba State, in the Northeast region of Brazil, 80 animals slaughtered in the public slaughterhouse of Patos city were used. Before slaughter, blood samples were collected by jugular venopuncture from each animal, and after slaughter, testicles, epidydimus and uterus were aseptically collected. For the serological diagnosis of B. ovis and B. abortus infections, the agar gel immunodiffusion (AGID) and Rose Bengal (RBT) tests were carried out, respectively. In addition, microbiological culture and polymerase chain reaction (PCR) were performed on testicle, epidydimus and uterus samples. Six animals (7.5%) tested positive for the presence of B. ovis antibodies and all animals tested negative for the presence of B. abortus antibodies. One AGID-positive animal tested positive at uterine swab culture. PCR was able to amplify DNA of Brucella spp. from the pool of testicle, epidydimus and uterus samples from AGID-positive animals. This is the first report of isolation and detection of B. ovis DNA by PCR in ovine from the Northeast region of Brazil.

Evaluation of DNA extraction protocols for Brucella abortus pcr detection in aborted fetuses or calves born from cows experimentally infected with strain 2308

The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives), 26 spleens (11 positives), 23 livers (8 positives) and 22 bronchial lymph nodes (7 positives). All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the primers B4 and B5. From the accumulated results for organ, the proportion of positives for the lungs was higher than the livers (p=0.04) or bronchial lymph nodes (p=0.004) and equal to the spleens (p=0.18). From the accumulated results for DNA extraction protocol, the proportion of positives for the Boom protocol was bigger than the PK (p<0.0001) and GT (p=0.0004). There was no difference between the PK and GT protocols (p=0.5). Some positive samples from the classical bacteriology were negative to the PCR and viceversa. Therefore, the best strategy for B. abortus detection in the organs of aborted fetuses or calves born from infected cows is the use, in parallel, of isolation by classical bacteriology and the PCR, with the DNA extraction performed by the Boom protocol.