We previously described our systematic progress that eventually resulted in a commercially available immunoassay based biosensor (PCB biosensor) for detecting PCBs in oil. However, IC50 of the commercialized PCB biosensor was approximately 2 ppb for PCBs, and did not achieve the theoretical detection limit (TDL) which would represent an IC50 of approximately 0.5 ppb. In this study, we characterize the effects of the antibody concentration, flow volume and flow rate on the PCB biosensor’s response. Using the optimum operating conditions, the PCB biosensor achieved the TDL and its performance as a screening test was improved. Working at the stringent maximum residue limit specified by Japanese law (0.5 ppm total PCBs), the optimized biosensor exhibited excellent performance (0% false negatives and 7% false positives) in the screening of 110 samples of used Japanese transformer oil. The general approach for optimization described here is expected to benefit immunoassay researchers attempting to achieve optimum performance.
The synthesis of functionalized naphthalimide (compound 1) and its application for the preparation of pH chemosensor was described. The fluorescence enhancement of compound 1 with the increase of hydrogen ion concentration is based on the hindering of photo-induced electron transfer (PET) from aliphatic amine group to the naphthalimide. The comparison of this method with some other fluorescence methods based on naphthalimide for the measurement of pH indicated that the method can be applied in aqueous solutions rather than organic solutions. The analytical performance characteristics of the proposed pH chemosensor were also investigated. The chemosensor can be applied to the determination of pH between 3.20 and 7.80. Common inorganic ions do not show obvious interference for pH measurements. Moreover, the response of the chemosensor for pH is fast (response time less than 3 min). In addition, the chemosensor has been used for determination of pH in urine samples with satisfactory results.
A new fluorescent chemosensor for Cu(II) ions was designed and synthesized on the basis of the sequence-specific cleavage of the peptide bond by the peptidase (metal or metal complexes). In the chemosensor system, the substrate was labeled with a FAM fluorophore (6-carboxyfluorescein) at the N-terminal and with a Dabcyl quencher 4-(4′-dimethylaminophenylazo)benzoic acid at the ε-N of C-terminal Lys. In the presence of Cu(II), the substrate strand is cleaved, and the release of the cleaved fragment results in a significant fluorescence increase. The design was aided by the FRET study that showed a “turn-on” response for Cu(II) in an aqueous medium. Under optimum conditions, the novel chemosensor described here had a linear response range for Cu(II) from 1.0 × 10−8 to 1.0 × 10−6 mol dm−3 with a detection limit of 1.0 × 10−8 mol dm−3.
To overcome the sensitivity limit for small molecules (haptens) in immunoassays based on antigen-horseradish peroxidase (HRP) conjugates as labels, a novel approach was established that afforded very low detection limits. Biotinylated anti-HRP antibody was utilized in order to attach, via a streptavidin bridge, liposomaly entrapped HRP. Fentanyl, used as a model antigen, could be determined via the generation of a high-intensity and relatively stable chemiluminescence (CL) signal of a HRP-catalyzed luminol/H2O2/enhancer system, immediately after the addition of a substrate solution. 4-(1-Imidazolyl)phenol (4-IMP) was used as an enhancer, and the outcome of this combination was a very low detection limit (0.895 pg mL−1) in plasma samples. The respective detection limit with the use of just the classical HRP-antigen conjugate was > 5-times higher. Intra- and inter-assay RSDs of the novel assay were 6.8 – 9.9 and 11 – 17%, respectively. The proposed method could be utilized for a wide range of molecules without replacing existing antigen-HRP based kits.
A DC current, which was driven by the self-bias voltage, could be conducted in a radio-frequency-powered glow discharge plasma by connecting a low-pass filter circuit and a load resistor with the discharge tube. This current enhanced the intensity of emission spectra from the plasma largely. The intensities of iron atomic lines increased 35 – 50 times, whereas the sputtering rate was not changed by the current introduction. Boltzmann plots for iron atomic (Fe I) and ionic lines (Fe II) were investigated when the bias current was conducted, so that the excitation process relating to the intensity increase could be clarified. While the excitation temperature of the Fe I lines was slightly changed (3000 – 3600 K), that of the Fe II lines was drastically reduced from 7600 to 4300 K, which was close to the temperature of the Fe I lines at higher bias currents. Therefore, the plasma was changed towards an LTE condition so that both the Fe I and the Fe II lines could be excited through a common major process. The bias-current enhanced the density of electrons enabling low-lying excited energy levels (3 – 5 eV) of iron atom/ion to be much more populated, and they became the major colliding partners for the excitation of these iron species.
A novel cellulose-based resin functionalized with polyallylamine was synthesized. It was applied to the collection of phosphate in environmental water samples, followed by concentration determination using an inductively coupled plasma–atomic emission spectrometer (ICP/AES). The synthesized resin, cellulose-glycidylmethacrylate-polyallylamine (CGP), showed good adsorption behavior toward trace amounts of phosphate over a wide pH range. The adsorbed-analyte can be easily eluted using 0.5 M hydrochloric acid; its recoveries was found to be 80 – 100%. The CGP resin synthesized was packed in a mini-column, which was then installed in a computer-controlled auto-pretreatment system for on-line collection/concentration and determination of trace phosphate by ICP/AES. The limit of detection for phosphate was found to be 0.6 μg P l−1. The sample volumes were only 5 ml and the total analysis time was about 4 min. The developed method with CGP resin was successfully applied to the determination of phosphate in river water and tap water samples with satisfactory results. The recovery test showed that common matrices that may exist in environmental waters did not interfere with the determination of phosphate.
An aqueous extraction of inorganic As species, such as arsenite (As(III)) and arsenate (As(V)), was developed for monitoring inorganic As in the edible brown alga Hizikia fusiforme (hijiki). The ultrasonic extraction with water, even without heating, was found to be an acceptable monitoring method for an evaluation of water-soluble inorganic As, since it could extract about 80% of total As. Such an extraction efficiency was almost the same as those of enzyme assisted extraction methods. The developed extraction procedure was applied to 15 hijiki samples that had been collected at different coasts in Japan. All samples contained a substantial proportion of As as arsenosugars; the relative amounts of the different As species extracted were dependent on the sample. The percentages of extractable As species in the hijiki samples were in the range from 70 to 90%, and the sums of the concentrations of As(III) and As(V), which was defined as i-As, were in the range from 36 to 79% of the total As concentration in each sample. The proposed method is appropriate for environmental monitoring for inorganic As speciation in algae.
An automatic system for simultaneous determination of K+, Na+, Cl− and Ca2+ in serums was established on the basis of a negative-pressure flow-injection using micro-electrodes, and parameters of the system were optimized. The total ionic strength adjustment buffer consisted of 0.25 mmol L−1 K+, 48.6 mmol L−1 Na+, 2.56 mmol L−1 Cl−, 0.25 mmol L−1 Ca2+ and 23 mmol L−1 Na2B4O7–H3BO3; the flow rate was 1.58 mL min−1, the sampling volume was 45 μL, and the mixing coil length was 30 cm. The system could conduct 480 detections h−1, and its RSD was less than 1.6%. Recoveries were 97.3 – 103.6%, and linear responses were 2.0 – 22.0 mmol L−1 for K+, 89.6 – 253.0 mmol L−1 for Na+, 20.1 – 248.2 mmol L−1 for Cl−, and 0.35 – 10.0 mmol L−1 for Ca2+, respectively. The ion concentration ranges were in ranges of human serum electrolytes. The system features minimized sorption of fibrin in sensors and prolonged sensor life.
Lipopeptide is one of the most important biosurfactants. The content of each isoform of a lipopeptide is crucial to the study of the properties of metabolites as well as the biological and biochemical characters of microbes. However, this information has not been accurately provided by the current analysis method. A new method for the quantitative determination of each isoform in the surfactin family has been established. The surfactin was firstly hydrolyzed in an acid solution at 90°C for 20 h, dried and then treated with bis(trimethylsilyl)trifluoroacetamide at 60°C for 20 min. The derived hydrolysates were then analyzed by GC-MS for a quantitative determination via comparison with working curves made with amino acids. GC-MS analyses show that the nine isoforms with an amount of 4.80 × 10−7 mol (493 μg) in a 500 μg surfactin sample were detected, in which the mole fractions of surfactin isoforms with different fatty acid chains were 0.32% (n C12), 4.89% (iso C13), 6.27% (anteiso C13), 23.05% (iso C14), 8.95% (n C14), 17.69% (iso C15), 38.69% (anteiso C15), 0.07% (iso C16), and 0.07% (n C16), respectively. This approach can be applied to quantitative analyses for other families of lipopeptides as long as the sequence of amino acid residues in the peptide is determined.
This study reports on a method for the speciation of iron in aqueous samples by the simultaneous analysis of divalent and trivalent iron ions with ion chromatography equipped with chemiluminescence detection (IC-CLD). Ferrous and ferric ions are first chelated by pyridine-2,6-dicarboxylic acid (PDCA) to form complexed anions, and separated by a mixed-bed ion-exchange column. The separated complexed ions are then detected with a CLD system containing luminol and hydrogen peroxide in a basic solution. This luminescence system has a linear dynamic range of ca. 3 orders of magnitude, with method detection limits as low as 7 μg L−1 for Fe(II) and 3 μg L−1 for Fe(III), measured in the simultaneous detection mode. This system resists interferences from common cations such as Cd, Ca, Cr, Cu, Mg, Ni, Pb, and Zn. Evaluation by analyzing real samples shows that this method is rapid, accurate, sensitive, and selective.
In metabolomic research, it is important to reduce systematic error in experimental conditions. To ensure that metabolomic data from different studies are comparable, it is necessary to remove unwanted systematic factors by data normalization. Several normalization methods are used for metabolomic data, but the best method has not yet been identified. In this study, to reduce variation from non-biological systematic errors, we applied 1-norm, 2-norm, and quantile normalization methods to liquid chromatography–mass spectrometry (LC-MS)-based metabolomic data from human urine samples after oral administration of cyclosporine (high- and low-dose) in healthy volunteers and compared the effectiveness of the three methods. The principal component analysis (PCA) score plot showed more obvious groupings according to the cyclosporine dose after quantile normalization than after the other two methods and prior to normalization. Quantile normalization is a simple and effective method to reduce non-biological systematic variation from human LC-MS-based metabolomic data, revealing the biological variance.
By exposure to appropriate UV intensities, rapid and quantitative oxidation/reduction of inorganic selenite, selenate and several organoselenium compounds representative of those of biochemical/metabolic interest, including selenomethionine, selenobetaine, L-selenocystine, selenomethylselenocysteine, γ-glutamyl-seleno-methylselenocysteine and selenocystamine, is achieved. In the presence of acetic acid, quantitative conversion to volatile SeH2 and SeCO occurs using a flow-through system comprising a highly efficient 40 W UV lamp for oxidation in tandem with a lower power 8 W UV photocatalytic reactor utilizing a thin-film coating of titania. The volatile reduced species are detected by atomic absorption spectrometry using a heated quartz tube atomizer. Direct photochemical conversion of selenite, selenomethionine, L-selenocystine, γ-glutamyl-Se-methylselenocysteine and selenocystamine occurs in the presence of 5% acetic acid, following exposure to an 8 W UV field, to yield volatile detectable species, whereas selenobetaine and selenate are unresponsive unless the latter is first subjected to oxidation by exposure to a highly efficient 40 W UV lamp and the selenate reduced in the presence of titania.
Plasmodium berghei strain NK65 is a rodent malaria parasite species widely used as a model of the human-infectious malaria parasite. Because a rodent malaria parasite model allows issues to be addressed which would not be possible with human-infectious species, e.g., mode of action and in vivo screening, a convenient method to analyze the malaria parasite proteome is required. The proteins of P. berghei separated using two-dimensional polyacrylamide gel electrophoresis were analyzed using matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight tandem mass spectrometry.
A steel-slag/compost fertilizer can be useful in supplying complex Fe(II) species to barren coastal regions. Seawater extractable organic matter (SWEOM) was examined for use as a novel chelator of Fe(II) in the compost. The dissociation kinetics for Fe(II)-SWEOM were evaluated, based on the rate of ligand-exchange with ortho-phenanthroline. The ΔH‡ for the Fe(II)-SWEOM (19 kJ mol−1) was significantly smaller than the corresponding values for Fe(II) complexes with humic substances (27 kJ mol−1), suggesting that the Fe(II)-SWEOM is kinetically less stable.
The chromatographic behavior of penicillins, cephalosporins and carbapenems has been studied on the thin layers of transition-metal ion (viz. Ni2+/Zn2+/Cu2+/Co2+) silicate modified silica. Transition-metal silicate (3.92%) and silica (96.08%) were found to be optimum and resulted in spherical-compact spots and improved resolution of the analytes. The effect of various mobile phases was also investigated. The chromatograms were visualized as yellow spots by placing in an I2-chamber. The method has been found to be reproducible and convenient for routine analysis.