Methods of analysis were presented for chemicals that promote or disrupt cellular signaling pathways. The developed analytical methods are based not only on receptor binding, but also on the following known molecular-level processes involved in signal transduction along signaling pathways, reconstituted in vitro or taken in part in living cells. The methods were discussed in relation to receptor binding assay and/or bioassay. Examples include: (1) Insulin signaling pathways; (1-i) Chemical selectivity of agonists for insulin signaling pathways based on agonist-induced phosphorylation of a target peptide; (1-ii) An SPR-based screening method for agonist selectivity for insulin signaling pathways based on the binding of phosphotyrosine to its specific binding protein; (1-iii) A fluorescent indicator for tyrosine phosphorylation-based insulin signaling pathways; (2) An optical method for evaluating ion selectivity for calcium signaling pathways in the cell; (3) Assay and screening of chemicals that disrupt cellular signaling pathways, potential endocrine disruptors in particular; (4) Protein conformational changes, and (5) A screening method for antigen-specific IgE using mast cells, based on intracellular calcium signaling.
Quinoline-8-ol, one of the most popular and versatile organic reagents, is known to react with several metals, because it has both a basic nitrogen and a phenolic group. Hence, quinoline-8-ol and its derivatives have been widely used as chelating and/or sorbent extraction agents in on-line flow injection analysis (FIA) along with a suitable detection technique. The state of the art for using quinoline-8-ol and its derivatives in FIA was studied in terms of the selectivity, sensitivity and precision. Emphasis was placed on validating these procedures by analyzing certified reference materials and applying these analytical methodologies to real samples for the determination of inorganics.
A high-performance liquid chromatography (HPLC) assay was developed for the determination of estazolam in human plasma. Estazolam and alprazolam as an internal standard were detected by ultraviolet absorbance at 240 nm. Estazolam in plasma was extracted by a rapid and simple procedure based on cyanopropyl bonded-phase extraction. Chromatographic separation was achieved with a reversed-phase C8-5 column using a mobile phase of 0.5% potassium dihydrogenphosphate(pH 4.5)-acetonitrile (70:30, v/v). The determination of estazolam was possible in the concentration range of 1.0 - 200.0 ng/mL. The mean recovery of estazolam added to plasma was 96.1 ± 1.5% with coefficients of variation of less than 5.5%. This method is applicable for accurately monitoring the plasma level of estazolam in healthy subjects participating in scientific research.
Poria cocos (Fuling), a popular Chinese medicinal (CM) herb of fungal origin, has been included in many combinations with other CM herbs for its traditionally claimed activities of inducing diuresis, excreting dampness, invigorating the spleen and tranquilizing the mind and its modern pharmacological use of modulating the immune system of the body. Dehydrotumulosic acid, one of the effective constituents of Fuling, was isolated from the chloroform-soluble material of ethanol extract of the fungus. After further purification by a high-performance liquid chromatographic method on a C18 column, the purified constituent was identified using modern analytical techniques, such as UV, 13C-NMR and EI-MS. A reversed-phase high-performance liquid chromatographic method has been developed for the determination of dehydrotumulosic acid in Poria cocos. The determination can be accomplished in less than 50 min using methanol-acetonitrile-2% glacial acetic acid as the mobile phase at a flow rate of 1.0 mL/min, with a UV detector setting at 242 nm and testosterone propionate used as an internal standard. This assay for dehydrotumulosic acid is simple, rapid and with good reproducibility.
A redox species was extracted from water (50 × 10-6 dm3) into a single micro-oil-droplet (30 × 10-12 dm3) in contact with a microelectrode using microcapillary injection and manipulation techniques. Further, an in situ microanalysis of the solute in a single oil droplet was demonstrated by differential pulse voltammetry. The redox species of 10-16 mol concentrated in the droplet could be quantitatively analyzed independently of the distribution coefficient of the solute between the oil and water phases. The potential of this technique was considered in terms of the preconcentration and separation as well as a microanalysis and an ultratrace analysis.
The preparation of a cholesterol amperometric biosensor using a platinized Pt electrode as a support for the electropolymerization of a polypyrrole film, in which cholesterol oxidase and ferrocene monocarboxylic acid (electron-transfer mediator) were co-entrapped, is described. All the biosensor preparation steps (platinization and electropolymerization) and the cholesterol determination take place in the same flow system. The presence of the mediator enhances the sensitivity and selectivity of the platinized biosensor without modifying the dynamic parameters of the response, and the platinized layer improves the operational lifetime of the mediated sensor. The sensitivity obtained was 88.51 nA mM-1 and the limit of detection was 12.4 µM of cholesterol. The analytical properties of the biosensor for the flow-injection determination of cholesterol were studied and compared with those of other more simple amperometric biosensor configurations.
An electrochemical sensor for amperometric detection of hydroxylamine was developed by electrodeposition of hybrid copper-cobalt hexacyanoferrate films on a glassy carbon electrode. The modified electrode could reduce the overpotential of the hydroxylamine oxidation by about 400 mV. This sensor exhibited a linear relationship between the response current and the hydroxylamine concentration in the range of 1.8 × 10-3 M to 4.6 × 10-6 M with a detection limit of 2.1 × 10-7 M. Good agreement was obtained between results from the present method and conventional UV-visible spectrophotometry results. The sensor had good recovery, stability and anti-interference ability.
In this study, a wall-jet flow cell with a GRC (graphite reinforced by carbon) electrode was designed for the amperometric detection of phenol and chlorophenols in liquid chromatography. The voltammetric responses of these analytes at the GRC electrodes are very similar to those at conventional glassy carbon electrodes. As the GRC electrodes were made of the same materials as commercially available mechanical pencil leads, they exhibit the advantages of low cost, simple surface renewability, lower residual current, and good electrode-to-electrode reproducibility, and thus can be used as disposable-type electrodes. Chromatographic separations of phenol, o-chlorophenol (o-CP), 2,4-dichlorophenol (2,4-DCP), 2,4,6-trichlorophenol (2,4,6-TCP), and pentachlorophenol (PCP) were achieved with an ODS column using a mobile phase containing a mixture of CH3CN and H2O (40:60) containing 25 mM L-(+)tartaric acid (pH = 4.5). Amperometric detections were based on the electrochemical oxidation of these compounds around +0.9 V vs. Ag/AgCl. Under the optimized conditions, linear calibrations were obtained in a range up to 100 µM for phenol, o-CP, 2,4-DCP, 2,4,6-TCP, and 200 µM for PCP, with the correlation coefficients r2 of 0.9992, 0.9997, 0.9986, 0.9992, and 0.9968, respectively. The chromatographic detection limits for the tested analytes were obtained at pmol levels.
In this study, an infrared (IR) hollow waveguide sampler was developed to detect organic compounds in aqueous samples with sample volumes less than 50 µL. This sampler was prepared by coating a thin hydrophobic film inside the IR hollow waveguide. After injecting a certain amount of aqueous solution, organic compounds could be absorbed into the hydrophobic film by partitions. By removing the residual water in the hollow waveguide sampler with a nitrogen purging gas, the absorbed organic compounds could be sensed using IR radiation. To investigate the applicability of this hollow waveguide sampler in the detection of small amounts of aqueous samples, an analytical working function was developed following an examination of the parameters which influence the analytical signals. Such factors as the volume of the aqueous solution, the sample concentration, the length of the hollow waveguide, and the sensitivity of this method were investigated. Excellent agreement between the analytical and theoretical predicted values was observed. Upon examining the linear relationship between the analyte signals and the concentration, the regression coefficients were generally higher than 0.998 in the examined concentration range of 1 to 100 ppm. Under the condition that the sample volume was 300 µL and based on three-times the spectra noise level, the calculated detection limits for this method were found at around 1 ppm for the examined analytes.
The presented work provides baseline data on the existing airborne conditions in the Miyagi Museum of Art in Sendai, Japan, during the summer of 2000. The chemical composition, size and indoor and outdoor origin of the suspended particulate matter were identified using a number of advanced X-ray techniques, such as Electron Probe X-Ray Microanalysis (EPXMA) and Energy Dispersive X-Ray Fluorescence Analysis (EDXRF). Our results, to the best of our knowledge, represent the first detailed study of the chemical nature of the indoor particulate matter in a Japanese museum and, as such, may contribute to future improvements of the air quality inside museums and to the lasting conservation of works of art.
The design and construction of a new type of time-of-flight mass spectrometer is described. The instrument was designed to improve the ionization efficiency and to efficiently detect ions. The system performance was evaluated using the SIMION software, and was compared with the experimental results. The shapes of the repeller, extraction, and ground electrodes had a strong effect on the trajectory of ions in the processes of ion acceleration and focusing. The major difference between the theory and the experiment can be attributed to space-charge effects.
The suitability of 5,10,15,20-tetrakis(4-carboxyphenyl)porphine as a complexing agent for the on-line preconcentration of cadmium using an anion exchanger (Amberlite IRA-904) in a microcolumn has been tested. Various parameters which affect complex formation and its sorption, as well as elution into the nebulizer of flame atomic absorption spectrometry (FAAS), were optimized. A 5 × 10-4 mol l-1 reagent was on-line mixed with an aqueous sample solution and flowed through the microcolumn for 2 min. The sorbed complexes were then eluted with 2 mol l-1 of nitric acid into the nebulizer of FAAS. A good precision (2.1% RSD for 40 µg l-1 cadmium) and a high enrichment factor (36) with a detection limit (3σ) of 1.4 µg l-1 were obtained. The achieved analytical results for a standard reference material (SRM 1643d) were in good agreement with the certified values.
The adsorption curves of divalent metal ions (M2+) to an iminodiacetate(IDA)-type chelating resin (-L2-) under the condition of metal ions in excess against IDA groups clearly indicated the difference in contribution between two species. Copper and nickel are adsorbed only as [(-L)MII], while cadmium and calcium are adsorbed as [(-L)MII] and [(-LH)2MII]. Addition of salts may enhance the deprotonation of IDA groups, interfere with the adsorption as [(-LH)2MII], and yield remarkable changes in adsorption curves, depending on the metal ions.
A thermospray interface was modified for the on-line coupling of normal- and reversed-phase high-performance liquid chromatography (HPLC) and Fourier transform infrared (FTIR) spectrometry. An LC/FTIR assembly was used to evaporate the column effluents and the solutes were deposited as a series of individual spots on a stainless-steel moving belt, which continuously transferred the solutes into a diffuse reflectance accessory of FTIR, enabling the identification of deposited solutes by measuring the IR spectrum. A lowered desolvation temperature of reversed-phase HPLC eluents, a higher deposition efficiency, such as 69%, and a reduction of the thermospray capillary voltage were achieved by using a heated gas flow and a heating plate. The thermospray temperature and the distance between the tip of the thermospray tubing and the surface of the belt were shown to influence the area of deposition of spots. A variation of ±1°C could be used for a sensitive and reproducible deposition of Irganox 565 with a relative standard deviation (RSD) of 1.8 to 2.5%. The UV and FTIR chromatograms gave similar features for the HPLC-separated constituents. The interface-derived IR spectra of the constituents showed excellent agreement of the spectral features with those of the standard FTIR spectra, and no thermal degradation was found to occur.
The catalytic behavior of iron tetrasulfonatophthalocyanine (FeTSPc) for the oxidation reaction of L-tyrosine with H2O2 in a di-2-ethylhexyl sodium sulfosuccinate (AOT) reversed-micellar system (AOT/cyclohexane) was studied. It was indicated that the reversed micelles could not only enhance the catalytic activity of FeTSPc, but could also increase the fluorescence intensity of the product. Factors that may influence the catalytic reaction, including the concentration of AOT, the cosolubilized water, temperature and pH, were further examined. The possibility of its analytical application was also tested. Experimental results show that the calibration graphs for the determinations of FeTSPc and H2O2 under optimum conditions are linear over the range of 1.0 × 10-8 - 1.0 × 10-6 mol L-1 and 0.0 - 3.0 × 10-6 mol L-1, respectively, with detection limits of 1.1 × 10-9 mol L-1 and 3.1 × 10-9 mol L-1 for FeTSPc and H2O2, respectively.
The absorption spectra of the 4f electron transitions of neodymium, praseodymium, holmium and erbium complexes with fleroxacin in the presence of cetylpyridinium chloride were studied by normal and derivative spectrophotometry. Their molar absorptivity at the maximum absorption bands are about 5.3 (at 571 nm) times greater for neodymium, 2.8 (at 483 nm) times greater for praseodymium, 12.6 (at 448.5 nm) times greater for holmium and 9.7 (at 519 nm) times greater for erbium than those in the absence of complexing agents. The second-derivative spectrum is used both to eliminate the interference from other rare earths and to improve the sensitivity. Beer’s law is obeyed from 3.0 - 70 µg ml-1 for neodymium and holmium, from 6.0 - 70 µg ml-1 for erbium, and from 12.0 - 70 µg ml-1 for praseodymium. The relative standard deviations are 1.9% and 1.5% for 7.5 µg ml-1 of neodymium and holmium, and 2.1% and 1.6% for 15.0 µg ml-1 of praseodymium and erbium, respectively. Their detection limits (signal-to-noise ratio = 2) are 3.2 µg ml-1, 1.3 µg ml-1, 1.1 µg ml-1 and 2.5 µg ml-1 for praseodymium, neodymium, holmium and erbium, respectively. A new system for the simultaneous determinations of the praseodymium, neodymium, holmium and erbium in rare earth mixtures with good accuracy and selectivity is proposed.