Development of Tetrahydrofuran/Water Optical Waveguide and Its Application to the Observation of Extraction Behavior of 1-Anilino-8-naphtalene Sulfonate at the Tetrahydrofuran/Water Interface.

A stable two-phase sheath flow using tetrahydrofuran (THF) for an inner flow and water for an outer flow was formed in a glass capillary, and worked as a stable liquid-core/liquid-cladding optical waveguide (THF/water LLW). Although THF and water were miscible with any ratio, the length of the stable THF/water LLW at 0.9 - 2.1 cm s-1 reached at least 150 mm. The THF/water LLW was applied to the observation of extraction behavior of solvatochromic fluorescence dye, 1-anilino-8-naphtalene sulfonate (ANS), through the THF/water interface. ANS was added to the water phase (clad solution) and its fluorescence, which was excited with the guided light (355 nm) through the LLW, was observed by changing the position of the detector. While the ANS stayed in the region of 70% THF to the end of the LLW without the addition of cationic surfactant, hexadecyltrimethylammonium ion (CTA+) at pH 3 and 11, the ion-pair of ANS and CTA+ was extracted into the higher concentration region of THF with the addition of CTA+ at pH 11.

Flow injection spectrophotometric determination of sub-mg dm(-3) silver(I) in a strongly acidic solution containing concentrated copper(II) using a pyridylazo reagent.

A novel spectrophotometric flow injection method for determination of silver(I) in a strongly acidic solution containing concentrated copper(II) was developed using a coloring ligand, 4-(3,5-dibromo-2-pyridylazo)-N-ethyl-N-(3-sulfopropyl)aniline (3,5-diBr-PAESA). The method was first investigated by batch method. The interference from copper(II) chelate could be eliminated by the masking effect of EDTA. By utilizing the large formation constant (K = 12.3) of AgBr, one could determine silver(I) as a decrease of absorption by silver(I) chelate due to formation of AgBr by addition of KBr. Based on the results of batch experiments, two types of flow injection analysis (FIA) systems were constructed. Sub-mg dm(-3) determination of silver(I) was attained without interference from excess copper(II). The proposed method was successfully applied to determination of silver in a copper plating solution used in a plant to manufacture copper printed circuit boards, where the concentration of silver was critically important in the process control.

Slab optical waveguide high-acidity sensor based on an absorbance change of protoporphyrin IX.

A sensitive and fast-responsive evanescent wave absorption sensor has been constructed for pH measurements in highly acidic ranges. This sensor is based on a pH-dependent color change of protoporphyrin IX (PPIX). For the sensitive detection, a visible attenuated total reflection spectrometer with a slab optical waveguide (SOWG) was laboratory-made, and the guiding layer surface was modified with a PPIX-immobilized acrylamide-based thin membrane. The sensing membrane with a thickness of approximately 1 mum was directly fabricated on the SOWG glass surface by copolymerization of acrylamide, N,N'-methylene bisacrylamide, and PPIX in the narrow space confined by a cover plate. PPIX possesses two double bonds in its structure, and so it can be covalently incorporated into the membrane. The response characteristics of the PPIX-immobilized optode membrane were explored using aqueous solutions with different concentrations of HNO(3) or HCl. The optode membrane provided characteristic Soret band absorption spectra depending on the hydrogen ion concentration; the absorbance at 410 nm increased with increasing the concentrations in the range of 0.15-2 M, corresponding to the range of pH -0.3 to 0.8. The absorption signal reached 90% of its final value within 10 s, while the absorption signal was quite readily returned to background level simply by passing 2 mL of distilled water through a flow cell with a volume of 16.5 muL placed on the SOWG. Due to the rapid response and reversibility, this sensor could be operated in a flow-through mode as well as in a conventional static mode, where deionized water was conveniently used as a carrier and conditioning solution. In terms of the stability and precision, this sensor showed no significant change in response even after 100 assays and after being stored in a dry condition for over 6 months. Relative standard deviations for 10 replicate measurements were less than 1.8% in the linear range, and the detection limit calculated from 3 times of the standard deviation was 0.02 pH unit.

Liquid core waveguide spectrophotometry for the sensitive determination of nitrite in river water samples.

A flow injection analysis system was built with a liquid core waveguide spectrophotometric detector using an 80 cm Teflon AF-1600 capillary tube (2,2-bistrifluoromethyl-4,5-difluoro-1,3-dioxole/tetrafluoroethylene). The system was applied to determine nitrite ion in river water samples. The lower limit of detection for nitrite was 2.1 nmol dm(-3) (0.1 ng dm(-3) as NO(2)(-)) and the relative standard deviation of measurements was typically 0.56% (n = 5) at 0.21 micromol dm(-3).

Monitoring of intercellular messengers released from neuron networks cultured in a microchip.

A cellular biochemistry analysis system was integrated on a quartz glass microchip with a microchamber for cell culture followed by a microchannel for detecting with a thermal lens microscope (TLM). Nerve cells from rat hippocampus were successfully cultured to form neural networks in the microchip. An aqueous solution of glutamate, which is known as a neurotransmitter, was introduced to stimulate the cultured neuron to release a retrograde messenger, arachidonate which is considered to be critical for neuronal plasticity, especially for long-term potentiation (LTP). After the introduction, the solution that flowed through the culture chamber was analyzed using the UV-TLM (excitation wavelength, 244 nm). The measured signal intensity was dependent on glutamate solution concentration, and the neurons were considered to release the retrograde messenger according to the glutamate concentration. This system is suitable for time-course monitoring of ultra trace amounts of chemicals released from very small amount of cultured cells.

Development and optimization of a lab-on-a-chip device for the measurement of trace nitrogen dioxide gas in the atmosphere.

We propose the use of lab-on-a-chip technology for measuring gaseous chemical pollutants, and describe the development of a microchip for the detection of nitrogen dioxide (NO2) in air. A microchip fabricated from quartz glass has been developed for handling the following three functions, gas absorption, chemical reaction and fluorescence detection. Channels constructed in the microchip were covered with porous glass plates, allowing nitrogen dioxide to penetrate into the triethanolamine (TEA) flowing within the microchannel beneath. The nitrogen dioxide was then mixed with TEA and reacted with a suitable fluorescence reagent in the chemical reaction chamber in the microchip. The reacted solution was then allowed to flow into the fluorescence detection area to be excited by an ultraviolet light-emitting diode (UV-LED), and the fluorescence was detected using a photomultiplier tube (PMT). The reaction time, reagent concentration, pH, flow rate and other measurement conditions were optimised for analysis of nitrogen dioxide in air. Preliminary studies with standardized test solutions revealed quantitative measurements of nitrite ion (NO2-), which corresponded to atmospheric nitrogen dioxide in the range of 10-80 ppbv.

Preparation of low flow-resistant methacrylate-based monolithic stationary phases of different hydrophobicity and the application to rapid reversed-phase liquid chromatographic separation of alkylbenzenes at high flow rate and elevated temperature.

Low flow-resistant alkyl methacrylate-based monolithic stationary phases of different hydrophobicity were constructed for reversed-phase capillary liquid chromatography by thermally initiated radical polymerization of respective methacrylate ester monomer with different alkyl chain (C2, C4, C6, C12, C18) and ethylene glycol dimethacrylate (EDMA) in a 250 microm i.d. fused silica capillary. The hydrophobicity was basically controlled by changing the length and/or the density of the alkyl-chain, while the composition and the ratio of porogenic solvent were adjusted to obtain highly permeable rigid monoliths with adequate column efficiency. Among the prepared monolithic stationary phases, C18-methacrylate monoliths polymerized from a binary porogenic solvent of isoamyl alcohol and 1,4-buthandiol exhibited the most promising performance in terms of hydraulic resistance and column efficiency. The pressure drops of 20-cm long monolithic columns were below approximately 0.4 MPa at a normal linear velocity of 1mm/s (a flow rate of 3 microL/min), and the numbers of theoretical plates for alkylbenzenes mostly exceeded 3000 plates/20 cm. The produced monolithic columns had good mechanical strength for high pressure and temperature, and could be properly operated even at a temperature of 80 degrees C and at a pressure of at least 33 MPa. At 80 degrees C, the theoretical plate numbers reached 6000 plates/20 cm because of the enhanced mass transfer. Due to the novel hydraulic resistance and mechanical strength, the separation time could be reduced 120-fold simply by raising the flow rate and column temperature.

Characteristics of a liquid/liquid optical waveguide using sheath flow and its application to detect molecules at a liquid/liquid interface.

The formation conditions and characteristics of a liquid/liquid optical waveguide (LLW) were studied using a two-phase sheath flow, where the inner organic phase flow acted as the core and the outer aqueous flow acted as the clad. In immiscible solvent systems, i.e., toluene/water and diethyl ether/water systems, the LLWs were formed in the range of higher than ca. 600 of the Reynolds number (Re), where the linear velocity of the organic solvent was much higher than that of the aqueous solution. On the other hand, in a miscible solvent system, i.e., a tetrahydrofuran/water system, a stable LLW was formed in the range of a much lower Re than in immiscible systems. Moreover, the molecules at the toluene/water interface of the LLW were observed with both fluorescence and absorbance measurement systems. In particular, the change in the fluorescence spectrum of 1-anilino-8-naphthalenesulfonate (ANS) at the interface within 1 ms was observed by this method, indicating the usefulness of the LLW for a fast kinetic study of a liquid/liquid interface.

Laser-induced fluorescence microscopic system using an optical parametric oscillator for tunable detection in microchip analysis.

A laser-induced fluorescence microscopic system based on optical parametric oscillation has been constructed as a tunable detector for microchip analysis. The detection limit of sulforhodamine B (Ex. 520 nm, Em. 570 nm) was 0.2 mumol, which was approximately eight orders of magnitude better than with a conventional fluorophotometer. The system was applied to the determination of fluorescence-labeled DNA (Ex. 494 nm, Em. 519 nm) in a microchannel and the detection limit reached a single molecule. These results showed the feasibility of this system as a highly sensitive and tunable fluorescence detector for microchip analysis.

A sub-second, time-resolved, linear dichroism measurement system for visible attenuated total reflection spectroscopy with a slab optical waveguide.

A sub-second, time-resolved, linear dichroism (LD) measurement system was constructed, fitted with a slab optical waveguide (SOWG), and utilized to observe the adsorption process of methylene blue (MB) onto silica surfaces. In the system, a semiconductor laser (670nm) was used as the light source and the out-coupled beam from the SOWG was split by a polarizing beam splitter into two polarized beams (TE and TM modes) to allow sequential linear dichroic ratio data to be obtained; the acquisition rate was 11 data sets per second. For an MB solution that contained no sodium dodecylbenzenesulfonate (DBS), in contact with a bare silica surface, a clear decrease occurred in the average orientation angle of adsorbed MB in the initial stages of the adsorption process. This result may correspond to a change in the chemical form of MB from monomer to dimer.

Preparation and application of methacrylate-based cation-exchange monolithic columns for capillary ion chromatography.

Polymer-based strong cation-exchange monolithic capillary columns with different capacities were constructed for ion chromatography by radical polymerization of glycidyl methacrylate (GMA) and ethylene dimethacrylate in a 250-microm-i.d. fused-silica capillary and its subsequent sulfonation based on ring opening of epoxides with 1 M Na(2)SO(3). The cation-exchange capacities can easily and reproducibly be controlled in the range of up to 300 microequiv/mL by changing the immersion time of the epoxy-containing polymer in the Na(2)SO(3) solution. The chromatographic performance of the produced monolithic capillary columns was evaluated through the separation of a model mixture of common cations such as Na(+), NH(4)(+), K(+), Mg(2+), and Ca(2+). As an example, these cations could be well separated from one another on a 15-cm-long cation-exchange monolithic column (column volume, 7.4 microL) with a capacity of 150 microequiv/mL by elution with 10 mM CuSO(4). The pressure drop of this 15-cm column was approximately 1 MPa at a normal linear velocity of 1 mm/s (a flow rate of 3 microL/min), and the numbers of theoretical plates for the cations were above 3000 plates/15 cm. This GMA-based cation-exchange monolithic column could withstand high linear velocities of at least 10 mm/s. Over a period of at least two weeks of continuous use, no significant changes in the selectivity and resolution were observed. The applicability of a flow rate gradient elution and the feasibility of direct injection determination of major cations in human saliva sample were also presented.

A new isoelectric focusing system for fast two-dimensional gel electrophoresis using a low-concentration polyacrylamide gel supported by a loose multifilament string.

A new isoelectric focusing (IEF) system for two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) has been proposed. In this system, a super-soft and tough IEF gel was achieved by casting polyacrylamide gel down to 2.0% T using a loose multifilament string (LMS) of nylon as a gel support. The IEF apparatus for the LMS-gel, fabricated from acrylic boards, had a cooling water chamber, and eliminated the need of electrode solutions by directly connecting the two ends of individual gels to platinum electrodes. The carrier ampholyte-generated pH gradients using the new IEF system was stable over a long duration of time and a wide range of voltages, and the IEF time became shorter using a 2.0% T gel than using a 4.0% T gel. Also, the LMS-gels prepared in different runs exhibited excellent reproducibility. The new IEF system was applied to 2-D PAGE of a chicken skeletal muscle extract, and it was found that the protein loading capacity, protein entry into the LMS-gels, and protein transfer efficiency from the first-dimensional to the second-dimensional gels were significantly improved by using a low-concentration (2.5% T) gel. Also, proteins of high molecular weight of more than 200 kDa were observed in the 2-D maps, and therefore the new IEF system has a very good potential to be applied for fast 2-D PAGE of high molecular-weight proteins.

Liquid/liquid optical waveguides using sheath flow as a new tool for liquid/liquid interfacial measurements.

A liquid/liquid optical waveguide was constructed using a sheath flow. Since the refractive index of an organic solvent is generally higher than that of water (nD = 1.33), light introduced into the inner organic flow should proceed with total multi-reflection within the inner flow, so that the inner part of the sheath flow acts as the core of an optical waveguide. This sheath flow liquid/liquid optical waveguide was stable and showed no substantial background scattering. Moreover, it is applicable to both miscible and immiscible liquid/ liquid interfaces. Thus, it may become a new tool for studying liquid/liquid interfaces as well as for sensitive optical measurements.

Sensitive measurement of methylene blue active substances by attenuated total reflection spectrometry with a trimethylsilane-modified glass slab optical waveguide.

Attenuated total reflection spectrometry with a slab optical waveguide (SOWG) was explored for the simple, rapid and sensitive measurement of total anionic surfactants by the methylene blue active substance (MBAS) method. A fused-silica sheet used as a guiding layer was modified with trimethylsilane (TMS) to extract and concentrate the MBASs on the SOWG surface. Based on preliminary studies of the adsorption behavior and visible ATR spectrum of MB on the modified silica surface, a detection wavelength of 600 nm was chosen for the sensitive measurement of anionic surfactants. When the concentration of MB was set at 10 microM in the final measurement solution, the calibration curve for a typical anionic surfactant (sodium dodecylbenzenesulfonate) was linear up to 0.6 microM and the detection limit was 0.07 microM. The proposed method was applied to the determination of total anionic surfactants in river water.