Electro-assisted extraction of ionic drugs from biological fluids through a supported liquid membrane (SLM) and into an aqueous acceptor solution was recently introduced as a new sample preparation technique termed electromembrane extraction (EME). The applied electrical potential across the SLM has typically been in the range of 1 – 300 V. Successful extractions have been demonstrated even with common batteries (9 V) instead of a power supply. The chemical composition of the SLM has been crucial for the selectivity and for the recoveries of the extraction. Compared to other liquid-phase microextraction techniques (LPME), extraction times have been reduced by a factor of 6 – 17, and successful extractions have been obtained at extraction times of 1 – 5 min, and even down to a few seconds with online microfluidic EME devices. The technique has provided very efficient sample clean-up and has been found well suited for the extraction of sample sizes in the low μL range. Extractions have been performed with both rod-shaped hydrophobic porous fibers and with flat hydrophobic porous sheets as SLM support. The technique has been successfully downscaled into the micro-chip format. The nature of the SLM has been tuned for extraction of drugs with different polarity allowing extractions to be tailored for specific applications depending on the analyte of interest. The technique has been found to be compatible with a wide range of biological fluids and extraction of drugs directly from untreated human plasma and whole blood has been demonstrated. EME selectively extracts the compounds from the complex biological sample matrix as well as allowing concentration of the drugs. With home-built equipment fully acceptable validation results have been obtained.
A precise understanding of individual cellular processes is essential to meet the expectations of most advanced cell biology. Therefore single-cell analysis is considered to be one of possible approach to overcome any misleading of cell characteristics by averaging large groups of cells in bulk conditions. In the present work, we modified a newly designed microchip for single-cell analysis and regulated the cell-adhesive area inside a cell-chamber of the microfluidic system. By using surface-modification techniques involving a silanization compound, a photo-labile linker and the 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer were covalently bonded on the surface of a microchannel. The MPC polymer was utilized as a non-biofouling compound for inhibiting non-specific binding of the biological samples inside the microchannel, and was selectively removed by a photochemical reaction that controlled the cell attachment. To achieve the desired single-macrophage patterning and culture in the cell-chamber of the microchannel, the cell density and flow rate of the culture medium were optimized. We found that a cell density of 2.0 × 106 cells/ml was the appropriate condition to introduce a single cell in each cell chamber. Furthermore, the macrophage was cultured in a small size of the cell chamber in a safe way for 5 h at a flow rate of 0.2 μl/min under the medium condition. This strategy can be a powerful tool for broadening new possibilities in studies of individual cellular processes in a dynamic microfluidic device.
The thermal inactivation of glucose oxidase (GOD) in aqueous solution has been studied by the electrochemical method to follow the bioelectrocatalytic current due to the oxidation of glucose by GOD. Exponential time-dependent decrease in bioelectrocatalytic current, that is, the decrease in the enzymatic activity of GOD, was observed at given temperatures to determine the rate constant (k) of a simple inactivation process: GOD (active) → GOD (inactive). The ln[k] vs. T−1 plots gave straight lines with all solution conditions tested, so that the resulting Arrhenius activation parameters including ΔH‡ and ΔS‡ can be compared with each other. In the 50 mmol/L phosphate buffer at 70°C, k was determined to be (6.6 ± 1.6)× 10−4 s−1, and ΔH‡ and ΔS‡ were calculated to be 202 ± 13 kJ mol−1 and 282 ± 39 J K−1 mol−1, respectively. By addition of 3 mol/L guanidine hydrochloride, the k was increased to (4.7 ± 0.6)× 10−3 s−1, indicating that the denaturant accelerates the thermal inactivation. In this case, ΔH‡ was significantly reduced. By addition of 1 g/L ε-poly-L-lysine, which may adsorb onto the GOD surface to reduce the local disorder, k was decreased to (1.8 ± 0.6)× 10−4 s−1. In this case, ΔS‡ was reduced but ΔH‡ was not decreased much. This can be used as an important indication for selection of the enzyme stabilizer in solution.
An electrochemical biosensor was developed using boron-doped diamond (BDD) as an electrode material. To enhance the electrical performance of the electrode, the BDD electrode was decorated with Pt-nanoparticles (Pt-NPs) by electrochemical deposition. Their morphology according to the applied potentials for the synthesis of Pt-NPs was characterized by SEM. To identify the performance of the electrode modified with Pt-NPs, glucose detection was used as a sample sensing process, and the results were compared with those of a gold electrode and a bare BDD electrode. The electrochemical characteristics of the modified electrode were examined by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The BDD electrode with the Pt-NPs showed higher sensitivity and a lower detection limit than the Au electrode and BDD electrode. The proposed biosensor based on the Pt-NPs decorated BDD electrode showed high sensitivity, a low detection limit, fast direct electron transfer and good stability.
The present work describes the preparation and characterization of a carbon nanotube paste electrode modified with 2,7-bis(ferrocenyl ethyl)fluoren-9-one (2,7-BF). This electrode showed an efficient catalytic activity for the electro-oxidation of 6-thioguanine (6-TG), which leads to lowering 6-TG overpotential by more than 610 mV. Also, the values of catalytic rate constant (k = 2.7 × 103 mol−1 L s−1), and diffusion coefficient (D = 2.7 × 10−5 cm2 s) were calculated. In 0.1 M phosphate buffer solution of pH 7.0, the oxidation current increased linearly with two concentration intervals of 6-TG, one is 0.06 to 10.0 μmol L−1 and the other is 10.0 to 160.0 μmol L−1. The detection limit (3σ) obtained by differential pulse voltammetry (DPV) was 22.0 nmol L−1. DPV was used for simultaneous determination of 6-TG, uric acid (UA) and folic acid (FA) at the modified electrode, and for quantification of 6-TG, UA and FA in some real samples by the standard addition method.
A rapid, specific, and sensitive method for the simultaneous quantitation of organophosphates (fenitrothion (MEP), malathion, and phenthoate (PAP)), glufosinate (GLUF), and glyphosate (GLYP) in human serum and urine by gas chromatography–mass spectrometry (GC-MS) has been validated. All of the targeted compounds together with the internal standard were extracted from the serum and urine using a mix-mode TiO-C18 monolithic spin column. The recovery of organophosphates from serum and urine ranged from 12.7 to 49.5%. The recovery of GLUF and GLYP from serum and urine ranged from 1.9 to 7.9%. The intra- and inter-accuracy and precision (expressed as relative standard deviation, %RSD) were within 96.7 – 107.7% and 4.0 – 13.8%, respectively. The detection and quantitation limits for serum and urine were 0.1 and 0.1 μg/ml, respectively, for organophosphates, 0.1 and 0.5 μg/ml, respectively for GLUF and GLYP. The method had linear calibration curves ranging from 0.1 to 25.0 μg/ml for organophosphates and 0.5 – 100.0 μg/ml for GLUF, and GLYP. The validated method was successfully applied to a clinical GLYP poisoning case.
New moment equations were developed for chromatography using superficially porous (shell-type) spherical particles, which have recently attracted much attention as one of separation media for fast separation with high efficiency. At first, the moment equations of the first absolute and second central moments in the real time domain were derived from the analytical solution in the Laplace domain of a set of basic equations of the general rate model of chromatography, which represent the mass balance, mass-transfer rate, and reaction kinetics in the column packed with shell-type particles. Then, the moment equations were used for analyzing the experimental data of chromatography of kallidin in a Halo column, which were published in a previous paper written by other researchers. It was tried to predict the chromatographic behavior of shell-type particles having different shell thicknesses. The new moment equations are useful for a detailed analysis of the chromatographic behavior of shell-type spherical particles. It is also concluded that they can be used for the preliminarily optimization of their structural characteristics.
Inorganic anions were separated on a reversed-phase stationary phase dynamically modified with crown ether as a selector in capillary ion chromatography. The eluent contained crown ether, acetonitrile and a salt. Free and cation-trapped crown ether molecules in the eluent were adsorbed on a hydrophobic stationary phase such as triacontyl-functionalized silica (C30). The eluent cations trapped on crown ether worked as the ion-exchange sites, where the eluent anions and the analyte anions were competing for electrostatic interaction. The sizes of crown ether and the salt cation affected the retention of analyte anions. The concentrations of acetonitrile and crown ether as well as the eluent anion also affected the retention of analyte anions. An aqueous solution containing 18-crown-6-ether, potassium salt and acetonitrile achieved larger retention for analyte anions. Effects of the eluent conditions on the retention of analyte anions were examined in detail.
An on-line microdialysis/high-performance liquid chromatography method was developed for the simultaneous determination of melamine and cyanuric acid in non-dairy coffee creamer. To collect these analytes from aqueous samples, the microdialysis system featured a microdialysis probe incorporating a polyarylethersulfone membrane and employed 0.05 M HCl in 0.1% (v/v) MeOH as the perfusate, with optimal efficiency obtained at a flow rate of 1 μL min−1. The chromatographic conditions were optimized when using a reverse-phase phenyl column and a mobile phase of phosphate buffer solution in 10% (v/v) MeOH, buffered at pH 3.0. Good linearity relationship (r2 > 0.9987), intra- and inter-day precisions (RSDs < 6.6%), recoveries (96.9 – 105.0%), and limits of detection (melamine, 3 ppb; cyanuric acid, 150 ppb) were observed for the two analytes. This method has been successfully applied to simultaneous determination of melamine and cyanuric acid in commercial creamers with the recoveries in the range of 97.5 to 102.6%.
A simple screening method was developed for the determination of glyphosate in water samples using a multi-pumping flow system. The proposed method is based on the reaction between glyphosate and p-dimethylaminocinnamaldehyde (p-DAC), in an acid medium where the reaction product can be measured spectrophotometrically at λmax = 495 nm. An experimental design methodology was used to optimize the measurement conditions. The proposed method was applied to the determination of glyphosate in water samples in a concentration range from 0.5 to 10 μg mL−1. The limit of detection and quantification were 0.17 and 0.53 μg mL−1, respectively. The results obtained (88.5 to 104.5%) in recovery studies for the determination of glyphosate in different water samples indicated good accuracy and no matrix effect for the developed method. Samples were also analyzed by a confirmatory HPLC method, and agreement within the two set of results was found.
In this work, a combined discrete and continuous wavelet transform analysis was developed for simultaneous spectrophotometric determinations of metformin hydrochloride and glibenclamide, two antidiabetic drugs, in binary mixtures without any chemical pretreatment. Absorption spectra were subjected to the 4-level db4 discrete wavelet transform (DWT) for signal de-noising. Selected continuous wavelet transform (CWT) families (rbio3.1 with scaling factor, a = 80, and gaus2, a = 60) were applied on these de-noised signals. Finally, a zero-crossing technique was used for the construction of calibration curves for both drugs. The proposed method was validated by analyzing synthetic mixtures of the investigated drugs with various concentrations. The amount of metformin hydrochloride and glibenclamide were determined by using CWT amplitudes in zero-crossing points. The mean recovery values of metformin hydrochloride and glibenclamide were found between 98.6 – 102.0 and 97.9 – 102.4% for rbio3 and 98.3 – 101.2 and 97.1 – 101.4% for gaus2 families, respectively. The obtained results showed that the developed method is a simple, rapid and precise procedure for the simultaneous determination of metformin hydrochloride and glibenclamide in binary mixtures.
New appropriate reaction system was found for spectrophotometric determination of phosphate anion. This spectrophotometric method is based on the color development due to the formation of yellow molybdophosphate anion in acidic ethylene glycol–water (EG-W) mixed solution containing Mo(VI) species. The solution containing e.g. 20 mM Na2MoO4, 0.1 M HCl, and 40% (v/v) EG is colorless, and becomes immediately yellow by addition of phosphate anion. Thus the method is simple, rapid, and easy to carry out. Although Si(IV) species is well known to interfere with the determination of phosphate anion in many cases, the EG-W Mo(VI) solution remains colorless after addition of silicate anion at 1 mM level, indicating that no yellow molybdosilicate anion was formed in the EG-W solution. Under an optimized condition, the absorbance at e.g. 400 nm of the EG-W P(V)-Mo(VI) solution was proportional to the concentration of phosphate anion with good reproducibility, and the detection limit was 1 μM. Also the present method is less interfered by high concentrations of potassium and ammonium cations and oxidative nitrite anion as well as silicate anion.
A screen-printed dissolved oxygen sensor was fabricated using cerium oxide-supported silver catalyst and polydimethylsiloxane (PDMS) film. A PDMS film of 3 μm thickness showed good permeability for oxygen and impermeability for hydrogen peroxide. The calibration curve has shown a linear relationship with a correlation coefficient of 0.996 for the dissolved oxygen concentration. The sensitivity and detection limit of the present sensor were calculated at –158 μA mM−1 and 8.4 μM, respectively.
Thermochemolysis–gas chromatography in the presence of tetramethylammonium acetate was applied to the direct determination of terephthalic acid (TPA) contained in solid decomposition products obtained from the hydrothermal recycling process of poly(ethylene terephthalate) (PET). On the chromatograms of the hydrothermal decomposition products of PET, a sharp peak of the TPA component was clearly observed as its corresponding dimethyl ester formed through the thermochemolysis reaction. Based on the peak intensities, the contents of TPA in the decomposition products were determined precisely and rapidly without using any cumbersome sample pretreatments.
An automated fluid-transport device for a chip-based capillary electrophoresis system has been developed. The device mainly consists of six peristaltic micropumps, two vacuum micropumps, microvalves, multi-way joints, titanium tubes, and a macro-to-micro connector. Various solutions used for the cleaning and activation of chip channels, and electrophoresis separation, are allowed to automatically transport to chip reservoirs by the electric control module. The performance of the whole system was characterized by the analysis of fluorescein sodium using chip electrophoresis with LED-induced fluorescence detection. The peak-height variation (RSD) was 3.8% in six cycles of analyses. Additionally, compared with conventional manual operation, the developed device can spare 60% time for chip pretreatment. This microdevice offers high-efficiency pretreatment for microchips, thereby resulting in a remarkable improvement of analytical capacity for batch samples.