A new class of abasic site-binding fluorescence ligands, Naph-NBD in which 7-nitrobenzo-2-oxa-1,3-diazole (NBD) is connected to 2-amino-7-methyl-1,8-naphthyridine (Naph) by a propylene linker, is presented for the ratiometric assay for SNPs typing. In solutions buffered to pH 7.0 (I = 0.11 M, at 5°C), Naph-NBD is found to selectively recognize pyrimidine bases over purine bases opposite the abasic site in DNA duplexes (K₁₁/M^-1: T, 8.1 × 10⁶; C, 2.5 × 10⁶; G, 0.33 × 10⁶; A, 0.27 × 10⁶). The binding of Naph-NBD is accompanied by significant quenching of the fluorescence from the naphthyridine moiety (λ_max, 409 nm), while the emission from the NBD (λ_max, 544 nm) is relatively unaffected. Such a fluorescence response of Naph-NBD allows the emission ratio detection of pyrimidine/purine transversion.

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The tetraoctylammonium cation forms water-immiscible room temperature ionic liquids with dodecylsulfate and dodecylbenzenesulfonate anions. The ionic liquids are halogen-free and can be considered environmentally friendly solvents. At 25°C, the solubilities of water in tetraoctylammonium dodecylsulfate and tetraoctylammonium dodecylbenzenesulfonate were 2.8 and 4.0%, respectively, and the densities were 0.92 and 0.93 g cm^-3. The ionic liquids formed stable interfaces with water at 25°C. The possible use of tetraoctylammonium dodecylsulfate as chelate extraction solvent was evaluated by using 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione as extractant. Tetraoctylammonium dodecylsulfate showed high extraction performance for divalent transition metal cations.

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The recognition ability of pteridine derivatives for nucleobases opposite an abasic (AP) site in an oligodeoxynucleotide (ODN) duplex is enhanced by using a propylene residue (Spacer-C3) as an AP site. The recognition ability is further enhanced both by attaching methyl groups to a fluorescent ligand and by measuring the fluorescence response at 5°C; 6.2 × 10⁶ M^-1 of the binding constant is attained between 2-amino-6,7-dimethyl-4-hydroxypteridine and guanine opposite the AP site in water.

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Detection of hydrogen peroxide is of industry interest and of biological importance. Here we report a new approach to hydrogen peroxide measurement using multilayer modified microcantilevers. Through a layer-by-layer nanoassembly technique, horseradish peroxidase was intercalated into a nanoscale multilayer assembly on one surface of microcantilevers. These enzyme-functionalized microcantilevers deflected in response to hydrogen peroxide concentrations in the nanomolar level. The magnitudes of bending were proportional to the concentrations of hydrogen peroxide. Furthermore, our study also showed that microcantilever technique may be used as a novel, more sensitive tool for the study of the conformational or structural changes of enzymes or proteins on surfaces.

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Photoaffinity labeling enables the direct probing of a target protein through a covalent bond between a ligand and its binding protein, and even a complex formed by weak interactions can be isolated by the method. The photochemical fishing approach accelerates the throughput, isolating crosslinked complexes and analyzing the structure of the ligand binding site within the protein. We used carbene-generating phenyldiazirine for this approach because practical examinations had shown that the phenyldiazirine functioned as the powerful barb on the hook. Improving the synthetic pathways of the photoprobes and using chemoselective-integrated photoreactive units makes possible the easy and rapid preparation of carbene-generating photoreactive probes including the derivatives in peptides, proteins, DNAs, and carbohydrates. This review also shows several recent impacts of photoaffinity labeling, including the in vivo preparation of photoreactive proteins in living cells.

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The effects of nitrogenous synergists on the potentiometric responses to divalent transition metal ions were investigated concerning polymeric liquid membranes containing thenoyltrifluoroacetone (Htta) as an ionophore. The tested synergists were pyridine (py) and 4,4′-dioctyl-2,2′-bipyridyl (C8bpy). The potentiometric responses to metal ions, such as Cd²⁺, Co²⁺, Ni²⁺ and Zn²⁺, were induced by adding the synergists into the liquid membrane systems. The coexistence of Htta and a synergist was necessary for generating the membrane potential. The tta^- anion adsorbed at the liquid membrane/solution interface and the complex formation between the synergist and a given metal ion appeared to participate in preferential uptake of metal ions.

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A rapid sample-digestion method for the determination of toxic metals, cadmium, chromium, and lead, in polyethylene and polyvinyl chloride has been developed by using a microwave oven for household use. An appropriate amount of the sample taken in a PTFE decomposition vessel was mixed with nitric acid or nitric and sulfuric acids. The vessel was heated in a microwave oven by a predetermined operating program. The digested sample was diluted to a definite volume with water after evaporating most of the nitric acid. The precipitate, if formed, was filtered off by a membrane filter. The metals were determined by ICP-AES. The sample digestion required 5 min (for 20-mg sample) to 25 min (for 60-mg sample). The analytical results obtained for cadmium, chromium, and lead in a polyethylene certified reference material, BCR-680, digested with nitric acid, were in good agreement with the certified values.

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A novel method for separating fulvic acid (FA) from soil extracts is proposed. The FA, defined as the acid-soluble fraction of an alkaline extract of soil, was separated based on the precipitation of an ion-pair with a cationic surfactant, such as domiphen bromide. The precipitate was dissolved in aqueous HCl to produce H⁺-type FA and a surfactant chloride (SUR-Cl). SUR-Cl, in the aqueous solution, was removed by extraction with CHCl₃. After the aqueous phase was passed through a cation-exchanger (H⁺-type), H⁺-type FA (FA-SUR) was obtained as a powder by lyophylization. The chemical characteristics of FA-SUR were compared with an FA sample separated according to the method of the International Humic Substances Society using a DAX-8 resin (FA-DAX). The oxygen content, O/C atomic ratio and total acidity of the FA-SUR were significantly larger than the corresponding values for FA-DAX. The solid-state CPMS ¹³C NMR spectra indicated that the higher oxygen content of the FA-SUR could be due to alcoholic hydroxyl groups and polysaccharides as well as carboxylic groups. These results show that FA-SUR is more polar and hydrophilic than FA-DAX.

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The application of an ultraviolet (UV) light-emitting diode (LED) to on-line sample concentration/fluorescence detection in capillary electrophoresis (CE) is described. The utility of a UV-LED (peak emission wavelength at 380 nm, ∼2 mW) for fluorescence detection was demonstrated by examining both a naturally fluorescent (riboflavin) compound and a non-fluorescent compound (tryptophan), respectively. The detection limit for riboflavin was determined to be 0.2 ppm by the normal MEKC mode, which was improved to 3 - 7 ppb when dynamic pH-junction technique was applied. On the other hand, the detection limit of the tryptophan derivative was determined to be 1.5 ppm using the MEKC mode, which was improved to 3 ppb when the sweeping-MEKC mode was applied. In an analysis of an actual sample, the concentrations of riboflavin in beer, and tryptophan in urine and milk samples were determined, respectively.

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A rapid and simple sampling approach using solid phase microextraction (SPME) for pyrethroid residues analysis in flue-cured tobacco was studied. The fibers coated with poly-dimethylsiloxane (PDMS) at 100 µm thickness were chosen. Extraction time of 180 s, desorption time of 120 s and desorption temperature of 280°C were selected. The whole sampling process, only including an ultrasonic assisted extraction step and a solid phase microextraction step, can be completed within 15 min. The associated SPME and ultrasonic assisted extraction using water as extracting solvent shows good results for tobacco pyrethroid residues determination. Results indicated that four pyrethroids can be determined simultaneously, and the limits of detection are below 35 ng g^-1 using GC/MS in selected ion monitoring (SIM) mode. The reproducibility of the technique is found to be better than 11.8% RSD.

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By using an ICP optical emission spectrometer having two different observation modes, the authors compared the spectrochemical characteristics of various emission lines as viewed from the axial direction and the direction radial to the long axis of the plasma. The excitation temperature, the emission intensity, and the degree of ionization were investigated when iron and chromium were employed as the test sample and further potassium was added as an interfering element. These observations could lead to a similar conclusion that the emission intensities from the axial direction were more easily affected by the potassium addition. The reason for this effect is probably because the portion of the plasma observed from the axial direction includes the tail zone which is apart from the induction zone and thus has lower temperatures. On the other hand, in the radial observation, one can observe the emission intensities from a narrow portion of the plasma just above the load coil. The axial observation mode gave better analytical performance, including a lower detection limit as well as a better signal-to-background ratio, compared to the radial observation mode. However, interferences from co-existing elements should be noted if the axial observation is employed in practical applications.

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The analytical performances of coupled hydride generation, integrated atom trap (HG-IAT) atomizer flame atomic absorption spectrometry (FAAS) systems were evaluated for determination of As and Se in biological and environmental reference materials. Arsenic and Se hydrides are atomized in air-acetylene flame-heated IAT. A new design of HG-IAT-FAAS hyphenated technique that would exceed the operational capabilities of existing arrangements (a water-cooled single silica tube, double-slotted quartz tube or an “integrated trap”) was investigated. A dramatic improvement in detection limit was achieved compared with that obtained using anyone of the above atom trapping techniques separately. The concentration detection limits were 4 and 3 ng ml^-1 for As and Se, respectively. For a 2 min in situ preconcentration time, sensitivity enhancement, compared to FAAS, were 875 and 833 folds for As and Se, respectively, using hydride generation, atom trapping technique. The sensitivity can be further improved by increasing the collection time. The relative standard deviations (RSDs) are of the order of 5 - 9% for this hyphenated technique. The designs studied include slotted tube, water-cooled single silica tube and integrated atom trap. The accuracy was assessed by analyses of NRCC DOLT-2 (Dogfish Liver) and NIST SRM 1648 (Urban Particulate Matter) reference materials. The measured As and Se contents in two reference materials were in satisfactory agreement with the certified values.

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A luminol chemiluminescence method was used to evaluate the porphyrin-induced photogeneration of hydrogen peroxide (H₂O₂). This method enabled us to detect H₂O₂ in the presence of a high concentration of porphyrin, which was not possible using conventional colorimetry. The limit of detection was about 1 µM. We compared the ability to generate H₂O₂, using uroporphyrin (UP), hexacarboxylporphyrin (HCP), coproporphyrin (CP), hematoporphyrin (HP), mesoporphyrin (MP), and protoporphyrin (PP). The amount of H₂O₂ photoproduced was strongly related to the state of the porphyrin in the aqueous solution. UP and HCP, which existed predominantly in a monomeric form, had a good ability to produce H₂O₂. HP and MP, existing as dimers, showed weak activity. CP, forming a mixture of monomer and dimer, had a moderate ability to produce H₂O₂. PP, which was highly aggregated, had a good ability. These results demonstrated that the efficiency of porphyrins to produce H₂O₂ was strongly dependent on their aggregated form, and the dimer suppressed the production of H₂O₂.

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The results of a study and application of leucocrystal violet for the determination of antimony in parts per million levels is described here. The proposed method is based on the reaction of antimony(III) with acidified potassium iodate to liberate iodine. The liberated iodine selectively oxidizes leucocrystal violet to crystal violet dye. The formed dye shows maximum absorbance at 590 nm. The color system obeys Beer's law in the concentration range from 0.4 - 3.6 µg antimony per 25 ml of final solution. The molar absorptivity and Sandell's sensitivity were found to be 7.32 × 10⁵ l mol^-1 cm^-1 and 0.0016 µg cm^-2, respectively. All variables were studied in order to optimize the reaction. The proposed method is satisfactorily applicable for the analysis of antimony in various environmental and biological samples. The method is simple, highly sensitive, accurate and reliable.

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The application of generalized rank annihilation method (GRAM) to the analysis of fluorescence excitation-emission matrices of mixtures of piroxicam and pyridoxine is described. The input of GRAM consists of two bilinear data matrices, i.e. one for unknown and one for the calibration sample. The excitation wavelength range was from 290 to 340 nm and the emission was recorded from 370 to 560 nm. Piroxicam and pyridoxine were determined in the concentration ranges 0.33 - 4.00 µg ml^-1 and 0.66 - 8.00 µg ml^-1, respectively. To check the accuracy of the proposed method, several binary synthetic mixtures and one real sample were analyzed successfully. No matrix effect was observed in mixture analysis, so a single external calibration sample was used for each analyte. The ability of GRAM to quantify the studied compounds and the comparability of GRAM results were evaluated by comparing them with those of PLS regression as a standard first-order calibration.

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There is a possibility that the color development of the dye-binding method based on a protein error of a pH indicator is affected by the coexisting inorganic salt. Thus, the author theoretically and experimentally investigated the effect of the inorganic salt on the protein error. In a theoretical analysis, the anion of an inorganic salt, like the dissociated dye and buffer anions, was assumed to react with the protein, forming a colorless anion-protein complex. The calculated results were compared with those obtained by experiments using three pH indicators and various kinds of inorganic salts. The calculated results obtained are as follows: (1) The color development decreases with increasing the concentration of the inorganic salt and the equilibrium constant of the reaction between the inorganic salt and protein; (2) The rate of the absorbance decrease is larger for a lower concentration of the inorganic salt than for a higher one; (3) The larger is the equilibrium constant, the larger is the absorbance decrease. The absorbance decrease was caused by the anion, and was increased by increasing the anion concentration. The magnitude of the effect of the anion was iodide > bromide > chloride, which was associated with their ionic radius. The difference in the effect of the anion was thought to indicate that the equilibrium constant, in other words, the bonding strength of the anion to protein is iodide > bromide > chloride.

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A simple and sensitive method was conducted for the determination of trace amounts of proteins with benzeneazo-8-acetylamino-1-naphthol-3,6-disulfonic acid sodium salt (azophloxine, AP) using a Rayleigh light-scattering (RLS) technique. At pH 2.60 and in the presence of an emulsifier OP microemulsion, the RLS of AP can be greatly enhanced by proteins, owing to the interaction between AP and protein. The enhanced intensity is proportional to the concentration of proteins. Four proteins, including bovine serum albumin (BSA), human serum albumin (HSA), lysozyme (Lys) and γ-globulin (γ-G) have been tested. For example, the linear range of BSA was 0 - 0.06 µg mL^-1 with detection limits of 2.38 ng mL^-1. The method was applied to the analysis of protein in human urine and penicillin samples with satisfactory results. The relative standard deviation was in all instances less than 4.0%, and the recovery was in the range of 97.5 - 104%.

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In this paper we describe a highly sensitive and selective liquid chromatographic method for the determination of 5-methoxyindoles (5-methoxyindole-3-acetic acid, 5-methoxytryptamine, 5-methoxytryptophol, and melatonin) using a post-column technique involving electrolytic demethylation followed by fluorescence derivatization with benzylamine. We separated these compounds within 30 min by reversed-phase liquid chromatography using acetate buffer (pH 6.5)-acetonitrile-methanol [8:1:1 (v/v); isocratic elution] and then demethylated them, using a commercial coulometric system, to give the corresponding 5-hydroxyindoles. Next, we converted the 5-hydroxyindole products into fluorescent derivatives by their reactions with benzylamine in the presence of potassium hexacyanoferrate(III). We detected the derivatives spectrofluorometrically at 480 nm upon excitation at 345 nm. The detection limits (signal-to-noise ratio = 3) of the 5-methoxyindoles were in the range from 12 to 93 fmol per 20-µL injection.

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A simple and rapid gradient reversed-phase high-performance liquid chromatographic method for simultaneous separation and determination of paracetamol and its related compounds in bulk drugs and pharmaceutical formulations has been developed. As many as nine process impurities and one degradation product of paracetamol have been separated on a Symmetry C₁₈ column (4.6 × 250 mm i.d., particle size 5 µm) with gradient elution using 0.01 M potassium dihydrogen phosphate buffer (pH 3.0) and acetonitrile as mobile phase and photo diode array detection at 215 nm. The chromatographic behavior of all the compounds was examined under variable compositions of different solvents, temperatures, buffer concentrations and pH values. The correlation coefficients for calibration curves for paracetamol as well as impurities were in the range of 0.9951 - 0.9994. The proposed RP-LC method was successfully applied to the analysis of commercial formulations; the recoveries of paracetamol were in the range of 99 - 101%. The method could be of use not only for rapid and routine evaluation of the quality of paracetamol in bulk drug manufacturing units but also for detection of its impurities in pharmaceutical formulations.

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Activated carbon fiber (ACF) as extraction fiber for solid-phase microextraction (SPME) and its application for the analysis of banned organochlorine pesticides (OCPs) were investigated. Firstly, ACF was activated by different concentration of zinc chloride, which indicated that ACF activated by 60% zinc chloride had a reasonable specific surface area, pore volume and pore distribution. Secondly, the parameters for the ACF-SPME procedure, the adsorption and desorption conditions, were also optimized when coupled with gas chromatography-mass spectrometry (GC-MS). Thirdly, the ACF-SPME was used to analyze 17 kinds of OCPs in water. The linearity of most pesticides was found to be between 0.2 and 50 µg/l with GC-MS under the selected ion monitoring (SIM) acquisition mode. The limits of detection (LOD) at the sub µg/l were obtained. The work demonstrated here shows that ACF is a promising alternative for the SPME procedure.

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A novel and highly sensitive stopped-flow kinetic spectrophotometric method for the determination of nitrite, based on monitoring the variation in the absorbance of the intermediate within a very short period, has been developed. The optimum conditions for various parameters on which the reaction of nitrite with perphenazine depends, were investigated. It was found that the initial reaction rate increased linearly with increasing nitrite concentration in the range from 1.0 × 10^-8 to 6.0 × 10^-6 M. The detection limit was calculated to be 4.8 × 10^-9 M. This method was used for the determination of nitrite in natural and drinking-water with satisfactory results. The influence of cationic, non-ionic and anion surfactants was also studied in this work.

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We have established a simple method for assaying the quantity of soluble bromine in the discharge tubes of an extra-high-pressure mercury discharge lamp. Each discharge tube is destroyed in 5 ml of 10 mM sodium hydroxide, and the recovered sodium hydroxide solution is analyzed by suppressed-ion chromatography using gradient elution. We have clarified that this method can assay less than 1 µg of soluble bromine in a discharge tube.

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A rapid method for the determination of Pu isotopes in a soil sample within 60 min of starting sample pretreatment was developed. The large reduction in the analysis time was attained by the rapid and perfect digestion of the sample using an alkaline fusion method with an induction heating machine. Pu concentrations were then determined by flow injection/ICP-MS using a solid extraction resin after preconcentration by batch extraction with a chelate resin. The limits of detection for ²³⁹Pu and ²⁴⁰Pu were 9.2 fg and 4.3 fg, corresponding to 0.03 and 0.05 Bq kg^-1, respectively, under our analytical conditions, which satisfy the lower detection limits (0.5 Bq kg^-1 of ²³⁹Pu, and 2 Bq kg^-1 of ²⁴⁰Pu) required for rapid analysis techniques by the Ministry of Education, Culture, Sports, Science and Technology, Japan. This method provides a powerful and practical technique for emergency monitoring in and around nuclear facilities that handle large amounts of plutonium.

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A simple and highly sensitive spectrophotometric method for the determination of anthracycline anticancer agents, such as Daunorubicin hydrochloride (DAU), was established by using aluminum(III) and Chromazurol S (CAS) in a nonionic surfactant micellar medium. In the case of determination of DAU, the apparent molar absorptivity was 1.3 × 10⁵ dm³ mol^-1 cm^-1 at 615 nm. Beer's law was obeyed in the concentration range of 0.028 - 2.82 µg ml^-1 for DAU. Owing to no need for solvent extraction, this method could be applied to assays of DAU and related drugs in pharmaceutical preparations.

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A zeolite NaA (A-type zeolite of ca. 0.4 nm pore size; Linde Type A, LTA) membrane for the dehydration of alcohol was characterized by X-ray diffraction analysis (XRD). Also, the relationship between the X-ray absorption and the EtOH/H₂O pervaporation (PV) dehydration performance (water selectivity and permeation flux) of the LTA membrane was first investigated. The LTA membranes used here were gel-synthesized hydrothermally on an alumina porous support tube. Since diffraction lines from the alumina generate from a deeper layer than those of the LTA crystal, and are absorbed by both the surface LTA crystal and materials embedded in the alumina porous support, the alumina (113) diffraction line was intensively monitored to estimate the overall X-ray absorption by the LTA membrane. The intensity of the alumina (113) diffraction line showed a good correlation with the PV dehydration performance of the LTA membrane, that is, lower values with the water selectivity and higher values with the permeation flux. The lower diffraction intensity means stronger X-ray absorption by the LTA membrane. The major factor causing the difference in the X-ray absorption is the thickness or quantity of materials embedded in an alumina porous support, rather than those of the surface LTA crystal. These phenomena can be used conveniently (without real PV experiments) to determine the EtOH/H₂O PV dehydration performance of the LTA membrane.

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The boron in plant cell walls, which is water-insoluble and in the solid state, is solubilized by pectinase digestion to give a dimeric rhamnogalacturonan II-borate (dRG-II-B) complex. To clarify the nondestructive structure of boron present in plant cell walls (as represented by sugar beet fiber), we performed 192- and 96-MHz ¹¹B solid state NMR measurements. The use of a high field magnet frequency of 192-MHz enabled us to observe ¹¹B isotropic chemical shifts at -9.7 and -9.6 ppm for dRG-II-B and sugar beet fiber in the solid state, respectively, demonstrating that the boron in isolated dRG-II-B and in plant cell walls is present as a borate-diol ester (1:2). The observation of the magnetic field dependence of the chemical shift and lineshape for the borate-diol ester (1:2) by quadrupolar interaction suggested that the borate complex had a distorted tetrahedral boron structure.

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For the dehydration process of biomass alcohol using a zeolite NaA (LTA) membrane supported by an alumina porous support tube, it is very important to understand damage of the surface crystal, which may be caused by acetic acid. Therefore, the surface structure and the chemical composition of the LTA membrane that reacted with water or acetic acid (pH 3 - 4) were investigated by the Fourier Transform Infrared Attenuated Total Reflectance method (FTIR-ATR), with a diamond prism as the waveguide, scanning electron microscope and an energy-dispersive X-ray analysis. For leaching experiments using water and acetic acid, ultrasonication was applied to promote the leaching rate and to surely prepare damaged membranes as a model experiment. The Si-O asymmetric stretching vibration spectra of the original LTA membranes showed a bimodal peak. LTA membranes after water leaching also showed the same peak. However, for the LTA membrane reacted with acetic acid of pH 3, damage of the surface LTA crystal and the loss of sodium by dissolution were clearly observed. Also, its Si-O spectral shape was broad, which suggests amorphous-like substances secondarily formed on the membrane surface. Sodium acetate was not detected for all LTA membranes reacted with acetic acid. To evaluate surface-damaged and sodium-loss membranes, FTIR-ATR can sensitively detect a Si-O spectral change corresponding to the surface structure and the chemical composition of the LTA membrane.

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A simple, precise and accurate method is proposed for rapid determination of trace amounts of hydroxylamine based on the reaction of hydroxylamine with iodate in acidic media. The reaction of neutral red by the produced nitrite ion was used to monitor the reaction spectrophotometrically at 525 nm by a fixed time method. Hydroxylamine in the range of 0.0400 - 1.200 µg mL^-1 could be determined. The relative standard deviation for 10 determinations of 0.500 µg mL^-1 hydroxylamine was 1.81% and the limit of detection was 0.010 µg mL^-1. The proposed method was applied to the determination of hydroxylamine in water samples with satisfactory results.

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A new, simple and inexpensive kinetic catalytic spectrophotometric method for the determination of oxalate is described. The method is based on an activation effect of oxalate on a catalytic effect of iron(II) on the oxidation of iodide by bromate. The reaction is monitored by measuring the absorbance of triiodide ion at λ_max = 352 nm. A calibration graph was obtained from 0.10 - 7.0 µg cm^-3 of oxalate with a detection limit of 0.080 µg cm^-3. The standard deviations for ten replicate determinations of 0.50, 1.0 and 5.0 µg cm^-3 of oxalate were 4.0, 2.6 and 1.8%, respectively. The applicability of the method was demonstrated by the determination of oxalate ion in real samples.

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With the help of nanocrystalline TiO₂, the direct electrochemistry of xanthine oxidase (XOD) was achieved and two pairs of redox waves were observed. The interaction between XOD and nitric oxide (NO) was also investigated. The experimental results reveal that NO can be reduced at a XOD-nano TiO₂ film modified electrode. When the NO concentration was low, the reduced product, HNO, would inactivate the protein. However, when the NO concentration was high, HNO would continue to react with NO to form N₂O₂- and N₃O₃-, which would not inhibit XOD, and thus the amount of active protein did not decrease any further.

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