Time and temperature dependant changes in red blood cell analytes used for testing recombinant erythropoietin abuse in sports.

There has been a long debate since the introduction of blood analysis prior to major sports events, to find out whether blood samples should be analysed right away on the site of competition or whether they should be transported and analysed in an anti-doping laboratory. Therefore, it was necessary to measure blood samples and compare the results obtained right after the blood withdrawal with those obtained after a few hours delay. Furthermore, it was interesting to determine the effect of temperature on the possible deterioration of red blood cell analytes used for testing recombinant erythropoietin abuse. Healthy volunteers were asked to give two blood samples and one of these was kept at room temperature whereas the second one was put into a refrigerator. On a regular basis, the samples were rolled for homogenisation and temperature stabilisation and were analysed with the same haematological apparatus. The results confirmed that blood controls prior to competition should be performed as soon as possible with standardised pre-analytical conditions to avoid too many variations notably on the haematocrit and the reticulocyte count. These recommendations should ideally also be applied to the all the blood controls compulsory for the medical follow up, otherwise unexplainable values could be misinterpreted and could for instance lead to a period of incapacity.

Magnetic coupled electrochemistry: Exploring the use of superparamagnetic nanoparticles for capturing, transporting and concentrating trace amounts of analytes

We propose the use of functionalized superparamagnetic nanoparticles for capturing, and transporting analytes, in association with an external miniature magnet to deposit such nanocarrier species at the electrode surface. This approach can be employed for the electroanalytical determination of chemical species capable of interacting with the nanoparticles, or in the opposite case, to block their response at the electrode surface. The concept was successfully demonstrated by using aminofunctionalized nanoparticles to block the discharge of hexacyanoferrate(II) ions, and to enhance the signals of aquapentacyanoferrate(II) ions via coordination to the surface amino groups. Selective analysis was also performed for silver ions, surpassing the stripping methods in terms of versatility and usefulness. (C) 2010 Elsevier B.V. All rights reserved.

Sample stacking in CZE using dynamic thermal junctions I. Analytes with low dpK(a)/dT crossing a single thermally induced pH junction in a BGE with high dpH/dT

The possibility to compress analyte bands at the beginning of CE runs has many advantages. Analytes at low concentration can be analyzed with high signal-to-noise ratios by using the so-called sample stacking methods. Moreover, sample injections with very narrow initial band widths (small initial standard deviations) are sometimes useful, especially if high resolutions among the bands are required in the shortest run time. In the present work, a method of sample stacking is proposed and demonstrated. It is based on BGEs with high thermal sensitive pHs (high dpH/dT) and analytes with low dpK(a)/dT. High thermal sensitivity means that the working pK(a) of the BGE has a high dpK(a)/dT in modulus. For instance, Tris and Ethanolamine have dpH/dT = -0.028/degrees C and -0.029/degrees C, respectively, whereas carboxylic acids have low dpK(a)/dT values, i.e. in the -0.002/degrees C to+0.002/degrees C range. The action of cooling and heating sections along the capillary during the runs affects also the local viscosity, conductivity, and electric field strength. The effect of these variables on electrophoretic velocity and band compression is theoretically calculated using a simple model. Finally, this stacking method was demonstrated for amino acids derivatized with naphthalene-2,3-dicarboxaldehyde and fluorescamine using a temperature difference of 70 degrees C between two neighbor sections and Tris as separation buffer. In this case, the BGE has a high pH thermal coefficient whereas the carboxylic groups of the analytes have low pK(a) thermal coefficients. The application of these dynamic thermal gradients increased peak height by a factor of two (and decreased the standard deviations of peaks by a factor of two) of aspartic acid and glutamic acid derivatized with naphthalene-2,3-dicarboxaldehyde and serine derivatized with fluorescamine. The effect of thermal compression of bands was not observed when runs were accomplished using phosphate buffer at pH 7 (negative control). Phosphate has a low dpH/dT in this pH range, similar to the dK(a)/dT of analytes. It is shown that vertical bar dK(a)/dT-dpH/dT vertical bar >> 0 is one determinant factor to have significant stacking produced by dynamic thermal junctions.

Sample stacking in CZE using dynamic thermal junctions II: Analytes with high dpK(a)/dT crossing a single thermal junction in a BGE with low dpH/dT

In a previous work [M. Mandaji, et al., this issue] a sample stacking method was theoretically modeled and experimentally demonstrated for analytes with low dpK(a)/dT (analytes carrying carboxylic groups) and BGEs with high dpH/dT (high pH-temperature-coefficients). In that work, buffer pH was modulated with temperature, inducing electrophoretic mobility changes in the analytes. In the present work, the opposite conditions are studied and tested, i.e. analytes with high dpK(a)/dT and BGEs that exhibit low dpH/dT. It is well known that organic bases such as amines, imidazoles, and benzimidazoles exhibit high dpK(a)/dT. Temperature variations induce instantaneous changes on the basicity of these and other basic groups. Therefore, the electrophoretic velocity of some analytes changes abruptly when temperature variations are applied along the capillary. This is true only if BGE pH remains constant or if it changes in the opposite direction of pK(a) of the analyte. The presence of hot and cold sections along the capillary also affects local viscosity, conductivity, and electric field strength. The effect of these variables on electrophoretic velocity and band stacking efficacy was also taken into account in the theoretical model presented. Finally, this stacking method is demonstrated for lysine partially derivatized with naphthalene-2,3-dicarboxaldehyde. In this case, the amino group of the lateral chain was left underivatized and only the alpha amino group was derivatized. Therefore, the basicity of the lateral amino group, and consequently the electrophoretic mobility, was modulated with temperature while the pH of the buffer used remained unchanged.

Temporal response pattern of biochemical analytes in experimental diabetes

The activities of the enzymes aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LD), creatine kinase (CK), amylase (AMS) and angiotensin converting enzyme (ACE) have been used to assess the toxic effects of xenobiotics that have hypoglycaemic action in hepatic, pancreatic, renal and muscle tissue. Using a validated experimental model of diabetes mellitus in rats, we ascertained whether this syndrome itself affected the serum activities of these enzymes over a 53-day period. Levels of hepatic enzymes AST, ALT and ALP were higher in the streptozotocin (STZ)diabetic rats (group D), but were controlled by insulin therapy (group DI). AMS was reduced in group D and unchanged in group DI rats. Proteinuria was detected 1 day after STZ administation and partially controlled by insulin (group DI); its early presence in group D rats, and the lack of any change in serum ACE in this group, indicates that proteinuria is the better marker for microangiopathy. Microscopic examination of liver, kidney, heart and skeletal muscles (soleus and extensor digitorum longus) revealed various alterations in group D rat tissues, which were less pronounced in group DI. The liver, pancreas and kidney tissue-damage was consistent with the altered serum levels of AST, ALT, ALP and AMS and proteinuria. We conclude that: (i) rigorous control is required when these serum-enzyme levels are used as indicators of tissue toxicity in experimental diabetes, and (ii) LD, CK and bilirubin serum levels, which are unaffected by diabetes, can be used when testing effects of xenobiotics on tissues.

Analysis of N-nitrosames in water using micellar electrokinetic chromatography (MEKC) and development of online and offline methods of extraction and enrichment of neutral polar analytes

Evaluation of the quality of the environment is essential for human wellness as pollutants in trace amounts can cause serious health problem. Nitrosamines are a group of compounds that are considered potential carcinogens and can be found in drinking water (as disinfection byproducts), foods, beverages and cosmetics. To monitor the level of these compounds to minimize daily intakes, fast and reliable analytical techniques are required. As these compounds are relatively highly polar, extraction and enrichment from environmental samples (aqueous) are challenging. Also, the trend of analytical techniques toward the reduction of sample size and minimization of organic solvent use demands new methods of analysis. In light of fulfilling these requirements, a new method of online preconcentration tailored to an electrokinetic chromatography is introduced. In this method, electroosmotic flow (EOF) was suppressed to increase the interaction time between analyte and micellar phase, therefore the only force to mobilize the neutral analytes is the interaction of analyte with moving micelles. In absence of EOF, polarity of applied potential was switched (negative or positive) to force (anionic or cationic) micelles to move toward the detector. To avoid the excessive band broadening due to longer analysis time caused by slow moving micelles, auxiliary pressure was introduced to boost the micelle movement toward the detector using an in house designed and built apparatus. Applying the external auxiliary pressure significantly reduced the analysis times without compromising separation efficiency. Parameters, such as type of surfactants, composition of background electrolyte (BGE), type of capillary, matrix effect, organic modifiers, etc., were evaluated in optimization of the method. The enrichment factors for targeted analytes were impressive, particularly; cationic surfactants were shown to be suitable for analysis of nitrosamines due to their ability to act as hydrogen bond donors. Ammonium perfluorooctanoate (APFO) also showed remarkable results in term of peak shapes and number of theoretical plates. It was shown that the separation results were best when a high conductivity sample was paired with a BGE of lower conductivity. Using higher surfactant concentrations (up to 200 mM SDS) than usual (50 mM SDS) for micellar electrokinetic chromatography (MEKC) improved the sweeping. A new method for micro-extraction and enrichment of highly polar neutral analytes (N-Nitrosamines in particular) based on three-phase drop micro-extraction was introduced and its performance studied. In this method, a new device using some easy-to-find components was fabricated and its operation and application demonstrated. Compared to conventional extraction methods (liquid-liquid extraction), consumption of organic solvents and operation times were significantly lower.

Development of wearable electrochemical sensors for the determination of biological analytes

Wearable biosensors are attracting interest due to their potential to provide continuous, real-time physiological information via dynamic, non-invasive measurements of biochemical markers in biofluids, such as interstitial fluid (ISF). One notable example of their applications is for glycemic monitoring in diabetic patients, which is typically carried out either by direct measurement of blood glucose via finger pricking or by wearable sensors that can continuously monitor glucose in ISF by sampling it from below the skin with a microneedle. In this context, the development of a new and minimally invasive multisensing tattoo-based platform for the monitoring of glucose and other analytes in ISF extracted through reverse iontophoresis in proposed by the GLUCOMFORT project. This elaborate describes the in-vitro development of flexible electrochemical sensors based on inkjet-printed PEDOT:PSS and metal inks that are capable of determining glucose and chloride at biologically relevant concentrations, making them good candidates for application in the GLUCOMFORT platform. In order to make PEDOT:PSS sensitive to glucose at micromolar concentrations, a biocompatible functionalization based on immobilized glucose oxidase and electrodeposited platinum was developed. This functionalization was successfully applied to bulk and flexible amperometric devices, the design of which was also optimized. Using the same strategy, flexible organic electrochemical transistors (OECTs) for glucose sensing were also made and successfully tested. For the sensing of chloride ions, an organic charge-modulated field-effect transistor (OCMFET) featuring a silver/silver chloride modified floating gate electrode was developed and tested.

Assessment Of Robustness On Analysis Using Headspace Solid-phase Microextraction And Comprehensive Two-dimensional Gas Chromatography Through Experimental Designs.

Plackett-Burman experimental design was applied for the robustness assessment of GC×GC-qMS (Comprehensive Two-Dimensional Gas Chromatography with Fast Quadrupolar Mass Spectrometric Detection) in quantitative and qualitative analysis of volatiles compounds from chocolate samples isolated by headspace solid-phase microextraction (HS-SPME). The influence of small changes around the nominal level of six factors deemed as important on peak areas (carrier gas flow rate, modulation period, temperature of ionic source, MS photomultiplier power, injector temperature and interface temperature) and of four factors considered as potentially influential on spectral quality (minimum and maximum limits of the scanned mass ranges, ions source temperature and photomultiplier power). The analytes selected for the study were 2,3,5-trimethylpyrazine, 2-octanone, octanal, 2-pentyl-furan, 2,3,5,6-tetramethylpyrazine, and 2-nonanone e nonanal. The factors pointed out as important on the robustness of the system were photomultiplier power for quantitative analysis and lower limit of mass scanning range for qualitative analysis.

Chemical Immobilization Of Crosslinked Polymeric Ionic Liquids On Nitinol Wires Produces Highly Robust Sorbent Coatings For Solid-phase Microextraction.

Super elastic nitinol (NiTi) wires were exploited as highly robust supports for three distinct crosslinked polymeric ionic liquid (PIL)-based coatings in solid-phase microextraction (SPME). The oxidation of NiTi wires in a boiling (30%w/w) H2O2 solution and subsequent derivatization in vinyltrimethoxysilane (VTMS) allowed for vinyl moieties to be appended to the surface of the support. UV-initiated on-fiber copolymerization of the vinyl-substituted NiTi support with monocationic ionic liquid (IL) monomers and dicationic IL crosslinkers produced a crosslinked PIL-based network that was covalently attached to the NiTi wire. This alteration alleviated receding of the coating from the support, which was observed for an analogous crosslinked PIL applied on unmodified NiTi wires. A series of demanding extraction conditions, including extreme pH, pre-exposure to pure organic solvents, and high temperatures, were applied to investigate the versatility and robustness of the fibers. Acceptable precision of the model analytes was obtained for all fibers under these conditions. Method validation by examining the relative recovery of a homologous group of phthalate esters (PAEs) was performed in drip-brewed coffee (maintained at 60 °C) by direct immersion SPME. Acceptable recoveries were obtained for most PAEs in the part-per-billion level, even in this exceedingly harsh and complex matrix.

Comparison Of Techniques For The Isolation Of Volatiles From Cashew Apple Juice.

The aim of this study was to compare the performance of the following techniques on the isolation of volatiles of importance for the aroma/flavor of fresh cashew apple juice: dynamic headspace analysis using PorapakQ(®) as trap, solvent extraction with and without further concentration of the isolate, and solid-phase microextraction (fiber DVB/CAR/PDMS). A total of 181 compounds were identified, from which 44 were esters, 20 terpenes, 19 alcohols, 17 hydrocarbons, 15 ketones, 14 aldehydes, among others. Sensory evaluation of the gas chromatography effluents revealed esters (n = 24) and terpenes (n = 10) as the most important aroma compounds. The four techniques were efficient in isolating esters, a chemical class of high impact in the cashew aroma/flavor. However, the dynamic headspace methodology produced an isolate in which the analytes were in greater concentration, which facilitates their identification (gas chromatography-mass spectrometry) and sensory evaluation in the chromatographic effluents. Solvent extraction (dichloromethane) without further concentration of the isolate was the most efficient methodology for the isolation of terpenes. Because these two techniques also isolated in greater concentration the volatiles from other chemical classes important to the cashew aroma, such as aldehydes and alcohols, they were considered the most advantageous for the study of cashew aroma/flavor.

Classificação de água de coco processada e natural por meio de HCA, PCA e teores de íons metálicos determinados por ICP OES

Inductively Coupled Plasma Optical Emission Spectrometry was used to determine Ca, Mg, Mn, Fe, Zn and Cu in samples of processed and natural coconut water. The sample preparation consisted in a filtration step followed by a dilution. The analysis was made employing optimized instrumental parameters and the results were evaluated using methods of Pattern Recognition. The data showed common concentration values for the analytes present in processed and natural samples. Principal Component Analysis (PCA) and Hierarchical Cluster Analysis (HCA) indicated that the samples of different kinds were statistically different when the concentrations of all the analytes were considered simultaneously.

Aplicação do ultra-som em sistemas eletroquímicos: considerações teóricas e experimentais

The aim of this review is to present and discuss the applications of ultrasound in electrochemical systems such as in sonoelectroanalysis and sonoelectrolysis for the electrochemical combustion of organic compounds. Initially, theoretical and experimental aspects are discussed, particularly those related to the enhancement of mass transport and the surface cleaning effects. Some results are included to illustrate alternative geometries for the experimental measurements and the working electrodes used in these systems. In the sequence, the available publications are presented and discussed to demonstrate that ultrasound combined with electrochemical techniques is a powerful set-up for the detection of analytes such as metals and/or organic compounds in hostile media and for the effective destruction of toxic organic substances. At the end, a table summarizes the results already published in the literature.

Estratégias de pré-concentração em eletroforese capilar: parte 2. Manipulação da velocidade da fase dispersa/secundária

This work describes CE preconcentration strategies based on the effect of manipulation of the disperse/secondary velocity. Introduced by Terabe et al. in 1984, micellar electrokinetic chromatography is a powerful separation approach that increases the usage of electrokinetic phenomena for the separation of nonionic compounds. The main disadvantage of MEKC is the low concentration sensitivity associated with the limited optical path length for on-capillary photometric detection and the limited volume of sample solution that can be injected. This paper compiles on-line concentration strategies for neutral analytes by sample stacking and sweeping in micellar electrokinetic chromatography.

Optimized architecture for Tyrosinase-containing Langmuir-Blodgett films to detect pyrogallol

The control of molecular architectures has been a key factor for the use of Langmuir-Blodgett (LB) films in biosensors, especially because biomolecules can be immobilized with preserved activity. In this paper we investigated the incorporation of tyrosinase (Tyr) in mixed Langmuir films of arachidic acid (AA) and a lutetium bisphthalocyanine (LuPc(2)), which is confirmed by a large expansion in the surface pressure isotherm. These mixed films of AA-LuPc(2) + Tyr could be transferred onto ITO and Pt electrodes as indicated by FTIR and electrochemical measurements, and there was no need for crosslinking of the enzyme molecules to preserve their activity. Significantly, the activity of the immobilised Tyr was considerably higher than in previous work in the literature, which allowed Tyr-containing LB films to be used as highly sensitive voltammetric sensors to detect pyrogallol. Linear responses have been found up to 400 mu M, with a detection limit of 4.87 x 10(-2) mu M (n = 4) and a sensitivity of 1.54 mu A mu M(-1) cm(-2). In addition, the Hill coefficient (h = 1.27) indicates cooperation with LuPc(2) that also acts as a catalyst. The enhanced performance of the LB-based biosensor resulted therefore from a preserved activity of Tyr combined with the catalytic activity of LuPc(2), in a strategy that can be extended to other enzymes and analytes upon varying the LB film architecture.