Enzyme-catalyzed signal amplification was introduced to the electrochemical detection of the OH radical. In the presence of phenol as a trapping agent, glucose as a substrate, and pyrroloquinoline quinone-containing glucose dehydrogenase (PQQ-GDH) as a catalyst, the current signal for the trapping adducts (catechol and hydroquinone) produced by the hydroxylation of phenol could be amplified and detected sensitively. The limit of detection (S/N = 3) for catechol was 8 nM. The trapping efficiency of phenol was also estimated.
Sequential injection immunoassay systems for environmental measurements based on the selective immunoreaction between antigen and antibody were described. A sequential injection analysis (SIA) technique is suitable to be applied for the procedure of enzyme-linked immunosorbent assay (ELISA), because the washing and the addition of reagent solutions can be automated by using a computer-controlled syringe pump and switching valve. We selected vitellogenin (Vg), which is a biomarker for evaluating environmental risk caused by endocrine-disrupting chemicals in the hydrosphere, and linear alkylbenzene sulfonates (LAS) and alkylphenol polyethoxylates (APEO), which are versatile surfactants, as target analytes in the flow immunoassay systems. For Vg monitoring, SIA systems based on spectrophotometric, chemiluminescence, and electrochemical determinations were constructed. On the other hand, chemiluminescence determination was applied to the detection of LAS and APEO. For APEO, an SIA system combined with surface plasmon resonance (SPR) sensor was also developed. These new sequential injection immunoassay systems are expected to be useful systems for environmental analysis.
UV Raman spectroscopy is a powerful tool for investigating the structures and interactions of the aromatic side chains of Phe, Tyr, Trp, and His in proteins. This is because Raman bands of aromatic ring vibrations are selectively enhanced with UV excitation, and intensities and wavenumbers of Raman bands sensitively reflect structures and interactions. Interpretation of protein Raman spectra is greatly assisted by using empirical correlations between spectra and structure. Many Raman bands of aromatic side chains have been proposed to be useful as markers of structures and interactions on the basis of empirical correlations. This article reviews the usefulness and limitations of the Raman markers for protonation/deprotonation, conformation, metal coordination, environmental polarity, hydrogen bonding, hydrophobic interaction, and cation-π interaction of the aromatic side chains. The utility of Raman markers is demonstrated through an application to the structural analysis of a membrane-bound proton channel protein.
A novel and simple interface for capillary gas chromatography–mass spectrometry (GC-MS) was developed using a piece of deactivated stainless-steel tubing. This interface eliminated the need to vent the MS ion source when changing columns. Various chromatographic performance indicators, such as inertness, and thermal and chemical stability, were confirmed to be unaffected by using this interface at an elevated temperature of around 300°C. The new interface should facilitate the characterization of polymeric materials using analytical pyrolysis techniques in which frequent switching is required in the measuring mode, such as evolved gas analysis-MS and flash pyrolysis-GC-MS.
An analytical method for determining bromate in drinking water was developed using liquid chromatography–tandem mass spectrometry (LC-MS/MS). The 18O-enriched bromate was used as an internal standard. The limit of quantification (LOQ) of bromate was 0.2 μg/L. The peak of bromate was separated from those of coexisting ions (i.e., chloride, nitrate and sulfate). The relative and absolute recoveries of bromate in two drinking water samples and in a synthesized ion solution (100 mg/L chloride, 10 mg N/L nitrate, and 100 mg/L sulfate) were 99 – 105 and 94 – 105%, respectively. Bromate concentrations in 11 drinking water samples determined by LC-MS/MS were <0.2 – 2.3 μg/L. The results of the present study indicated that the proposed method was suitable for determining bromate concentrations in drinking water without sample pretreatment.
Peak parking experiments were conducted to study the chromatographic behavior in a RPLC system consisting of a column packed with superficially porous C18-particles and a mixture of methanol and water (70/30, v/v). The values of the surface diffusion coefficient and the retention equilibrium constant of a column packed with superficially porous C18-particles were comparable to those of columns packed with a C18-silica monolith and full-porous C18-silica gel particles. The flow-rate dependence of HETP was hypothetically calculated by using moment equations to clarify the influence of the structural characteristics on the chromatographic behavior. The column efficiency of a column packed with the superficially porous particles is higher in the high flow-rate range than that with full-porous spherical particles. This is attributed to the smaller contribution of the intraparticulate mass transfer in the superficially porous particles to band broadening. The moment equations are effective for the quantitative analysis of chromatographic behavior of superficially porous particles.
Novel sol-gel titania film coated needles for solid-phase dynamic extraction (SPDE)-GC/MS analysis of desomorphine and desocodeine are described. The high thermal stability of titania film permits efficient extraction and analysis of poorly volatile opiate drugs. The influences of sol-gel reaction time, coating layer, extraction and desorption time and temperature on the SPDE needle performance were investigated. The deuterium labeled internal standard was introduced either during the extraction of analyte or directly injected to GC after the extraction process. The latter method was shown to be more sensitive for the analysis of water and urine samples containing opiate drugs. The proposed conditions provided a wide linear range (from 5 – 5000 ppb), and satisfactory linearity, with R2 values from 0.9958 to 0.9999, and prominent sensitivity, LOQs (1.0 – 5.0 ng/g). The sol-gel titania film coated needle with SPDE-GC/MS will be a promising technique for desomorphine and desocodeine analysis in urine.
The aim of the present work was to develop a new method using synchronous fluorescence spectrometry (SFS) to determine the concentration of isoliquiritigenin (ISL) in mouse blood and tissues, and to investigate ISL’s distribution among organs after an intraperitoneal (IP) dose of ISL. The synchronous fluorescence method was optimized with the sample pH, stability, metal ions, concentration of Al3+, and surfactants. The proposed method was used to determine the ISL concentration in mouse blood, brain, heart, kidney, liver, spleen and lung after an IP injection of ISL. The optimal conditions for the determination of ISL using SFS were found to be: excitation and emission wavelengths of 469 and 557 nm, respectively; the use of 3% AlCl3 as a fluorescence intensity enhancer; measuring samples within 1 h of collection, sample pH 7 – 8, isolation of samples from surfactants; and wavelength interval (Δλ) = 70 nm. After IP injection, the distribution of ISL in mouse organs was: liver > kidney > spleen > blood > lung > brain > heart. The blood concentration of ISL peaked at 60 min; concentrations of ISL in liver, kidney and spleen achieved maxima at 120 min. SFS provides a simple, but effective analytical method that will benefit the study of in vivo biological effects of ISL, including absorption, distribution, metabolism, and excretion.
A water-soluble fluorescence dye, bis(2,2′-bipyridine)-5-amino-1,10-phenanthroline ruthenium complex (Ru(bpy)2(5-NH2-1,10-phen)), was synthesized and used as a fluorescence probe for detecting dissolved oxygen in water. The fluorescence intensity of the probe in different dissolved-oxygen concentrations was investigated. The sensitivity of the probe was evaluated in terms of the ratio IN2/IO2, where IN2 and IO2 correspond to the detected fluorescence intensity of nitrogen and oxygen-saturated solutions, respectively. The experimental results showed that the probe yielded a linear Stern–Volmer plot, and had a IN2/IO2 ratio of about 5.2. The detection limit, defined as three-times the standard deviation, was 8.6 × 10−7 mol L−1 after eleven determinations of nitrogen-saturated blank solutions. Additionally, the probe was pH-insensitive and ionic strength-independent with good characteristics of practicality and selectivity.
We report on a novel method for the quantitation of proteins specifically bound on a ligand-presenting biochip by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). The bound protein was digested by trypsin, and the resulting peptide fragments were analyzed by MALDI-TOF MS in the presence of an isotope-labeled internal standard (IS). The IS has the same sequence as a reference peptide (RP) of the target protein digest, but a different molecular weight. The absolute amount of the specifically bound protein on a biochip is then quantitated by comparison of mass intensities between the RP and the IS. Because they have the same molecular milieu, the mass intensities of these two analytes represent the real amounts of analytes on the chip. As a model system, we tested glutathione s-transferase (GST) and a GST-fusion protein, which were captured on glutathione-presenting biochips. We observed that the glutathione densities on biochips showed a good correlation with the absolute quantity of the proteins. We believe that our method will provide an alternative to currently existing tools for the absolute quantitation of surface-bound proteins.
In the present paper, the sorption properties of caffeine (CAF) onto polyether type polyurethane foam (PUF) as solid phase sorbent were investigated with UV determination at 274.3 nm. Batch and column methods were used to optimize chemical, flow, kinetic and isothermal conditions for preconcentration of CAF. Results indicated quantitative sorption of CAF at pH 8 and 30 min shaking time. The maximum sorption capacity was found to be 4.1 mg g−1. Column preconcentration was recommended at a flow rate of 1.5 mL min−1 and desorption with 4 mL from 0.15 mol L−1 hydrochloric acid. The procedure provided a linear analytical range of 0.05 – 30 mg L−1. The detection and quantification limits are 0.016 and 0.047 mg L−1, respectively. The procedure was applied to determination of CAF in spiked human plasma. The obtained recoveries were 98 – 101% and RSD values were from 0.05 to 9.5%.
The effects of elevated hydrostatic pressure on four representative proteins, lysozyme, human serum albumin, ubiquitin and RNase A, were investigated by using Fourier transform infrared (FTIR) spectroscopy, by principal component analysis (PCA) and by moving-window two-dimensional (MW2D) correlation analysis. In addition, we revealed the pressure-induced changes of secondary structure elements using curve fitting. With pressure increase, the amide I band shifted to lower wavenumbers, with a transition at 200 MPa, which was indicative of hydration enhancement. Moreover, the pressure-induced behavior of pure water was studied, similar transition pressure was observed with protein in aqueous solution, suggesting that structure change of water around 200 MPa caused a hydration enhancement of protein. Under pressure higher than 200 MPa, the structural changes of the four proteins were obviously different except for the common features shifting to lower wavenumbers with pressure, basically due to the distinct structural differences among them.
The effects of neutron scattering by matrix hydrogen in geological samples were examined in order to accurately determine their chemical compositions by neutron-induced prompt gamma-ray analysis (PGA). Three different matrix materials including basaltic reference rock sample (JB-2) mixed with chemical reagents including H-containing ones were analyzed by using thermal and/or cold neutron-guided beams of JRR-3 at Japan Atomic Energy Agency. The sensitivity change of elements with the variation of the H concentrations was evaluated for disk and spherical target geometries. The results show that the analytical sensitivities of B, Cl and Cd in disk samples seem to increase with increasing the matrix H concentrations by the irradiation of both thermal and cold-neutron beams. The sensitivity enhancement of B for disk-shaped JB-2 mixed with up to 2% mass H is within the analytical uncertainty associated with PGA with a thermal-neutron beam.
A certified reference material (CRM) for trace elements in tea leaves has been developed in National Metrology Institute of Japan (NMIJ). The CRM was provided as a dry powder (<90 μm) after frozen pulverization of washed and dried fresh tea leaves from a tea plant farm in Shizuoka Prefecture, Japan. Characterization of the property value for each element was carried out exclusively by NMIJ with at least two independent analytical methods, including inductively coupled plasma mass spectrometry (ICP-MS), high-resolution (HR-) ICP-MS, isotope-dilution (ID-) ICP-MS, inductively coupled plasma optical emission spectrometry (ICP-OES), graphite-furnace atomic-absorption spectrometry (GF-AAS) and flame atomic-absorption spectrometry (FAAS). Property values were provided for 19 elements (Ca, K, Mg, P, Al, B, Ba, Cd, Cu, Fe, Li, Mn, Na, Ni, Pb, Rb, Sr, Zn and Co) and informative values for 18 elements (Ti, V, Cr, Y, and all of the lanthanides, except for Pm whose isotopes are exclusively radioactive). The concentration ranges of property values and informative values were from 1.59% (mass) of K to 0.0139 mg kg−1 of Cd and from 0.6 mg kg−1 of Ti to 0.0014 mg kg−1 of Lu, respectively. Combined relatively standard uncertainties of the property values were estimated by considering the uncertainties of the homogeneity, analytical methods, characterization, calibration standard, and dry-mass correction factor. The range of the relative combined standard uncertainties was from 1.5% of Mg and K to 4.1% of Cd.
Helix-coil transitions of poly α-aspartic acid (PASP) were studied by dc polarography in the presence of Zn2+ as a marker attached to the polymer. The diffusion current (id) of Zn2+ declined markedly in the pH range of 3.5 – 7.4 due to a formation of metal ion-PASP macromolecular complexes. The complex formation also reflects on an increase of the magnitude at ca. 222 nm of CD spectrum, suggesting that PASP forms the helix structure by coordination of Zn2+ in the corresponding pH region. Helix content, determined by the decrease in id of Zn2+, corresponds favorably to that by CD measurements. In the lower acidic pH region, the coordination mode of Zn2+ to PASP is different from that at neutral pH region. The decrease in id of Zn2+ is independent of the further formation of helix structure. Zn2+ coordinates with sparsely dissociated carboxylate groups of the helical part of PASP, which bring about an aggregation of polypeptide strands. The diffusion current of the ion attached to the polymer, therefore, is a parameter sensitive to conformational changes of PASP from acidic through neutral pH region.