Biomedical subjects

Totaro Imasaka

Publications and source records attributed to Totaro Imasaka.

12 recordsLinked to original sources

PubMed2003

Hadamard transform microchip electrophoresis combined with diode laser fluorometry.

The application of a Hadamard transform technique to microchip electrophoresis is described. The sample is electrokinetically injected into a separation channel and is then detected by diode laser-induced fluorometry. The sample and buffer solutions are introduced into the channel by controlling the high voltages applied to the solutions, according to a code determined by a Hadamard matrix. The S/N ratio of the signal in the electropherogram can be improved by a factor of 5 in comparison with that obtained by a conventional single-injection method, although an 8-fold improvement is theoretically predicted when a 255-order matrix is used.

Journal Article↗

PubMed2003

A sample introduction technique for obtaining a multiphoton ionization spectrum using picogram quantities of analyte in supersonic jet spectrometry.

A new sample introduction techniquefor ensuring the safety of a researcher during the measurement of hazardous compounds, such as dioxins, by supersonic jet spectrometry is described. The solution containing the analyte is introduced beneath the nozzle through a narrow capillary with a glass frit at the end, which is formed to vaporize only the solvent and to deposit the analyte. The first laser, that is, the second harmonic emission of a Nd: YAG laser, vaporizes the analyte and entrains it into a supersonic jet. The analyte molecule is ionized by the second laser, that is, an optical parametric oscillator. Ion measurement is achieved by a time-of-flight mass spectrometer. The analyte molecule is localized in the jet, thus permitting the amount of sample to be minimized. The minimum amount of phenol necessary for measurement of a multiphoton ionization spectrum by scanning the wavelength of the ionization laser in the 3-nm range was 180 pg, which is similar to or slightly less than the tolerable daily intake (TDI) of dioxin.

Journal Article↗

PubMed2003

Development of a compact supersonic jet/multiphoton ionization/time-of-flight mass spectrometer for the on-site analysis of dioxin, part I: Evaluation of basic performance.

A new type of compact supersonic jet/resonance-enhanced multiphoton ionization/time-of-flight mass spectrometer is described. The analytical instrument, consisting of a single turbo molecular pump equipped with a rotary pump, was maintained at < 2 x 10(-3) Pa when a 0.3-atm sample was injected into a vacuum at 10-Hz using a 200-micros pulse valve. The diameters of the extraction and ground skimmer electrodes were expanded to 30 mm in order to avoid strong focusing and defocusing of the ion, and the optimum conditions for the system were investigated. The mass spectrometer functioned as expected: (1) no defocusing of the ion beam was observed even when the potential of the einzel lens was adjusted to zero; (2) the direction of the ion beam to an assembly of microchannel plates deviated in the expected manner when the potential of the defection electrode was changed from 0 to 30 V.

Journal Article↗

PubMed2003

Development of a compact supersonic jet/multiphoton ionization/time-of-flight mass spectrometer for the on-site analysis of dioxin, part II: Application to chlorobenzene and dibenzofuran.

A compact supersonic jet/multiphoton ionization/mass spectrometer (SSJ/MPI/MS) equipped with a single pump system was developed for use in the analysis of dioxin. A mass spectrum and multiphoton ionization spectrum were obtained for chlorobenzene and dibenzofuran using an optical parametric oscillator (OPO) for excitation and subsequent ionization. The detection limit for chlorobenzene (S/N = 3) was 0.3 ppb, which corresponds to an absolute amount of 0.1 pg. The rotational temperature was 35 K. This slightly high temperature can be attributed to the long nozzle throat used in the instrument.

Journal Article↗

PubMed2003

Resonance-enhanced multiphoton ionization mass spectrometry using a two-color laser beam generated by stimulated Raman scattering.

Two-color (1+1') resonance-enhanced multiphoton ionization/time-of-flight mass spectrometry (REMPI/TOF-MS) combined with supersonic jet (SSJ) spectrometry has been demonstrated. The methodology uses a two-color laser beam consisting of a Stokes beam generated by stimulated Raman scattering (SRS) and a fundamental beam for the excitation and subsequent ionization of p-dichlorobenzene. The MPI signal was found to be substantially increased when a two-color laser beam was employed. This approach greatly simplifies the analytical system and reduces the cost of the instrumentation required for two-color ionization. The potential advantage of applying this method for the analysis of polychlorinated dibenzo-p-dioxin (PCDD) and polychlorinated dibenzofuran (PCDF) emitted from an incinerator is discussed.

Journal Article↗

PubMed2003

Absorption spectrometry using a diffractive optical element constructed by gel electrophoresis.

A novel diffractive optical element was constructed by means of gel electrophoresis. Five capillaries were filled with a methylene blue solution and inserted into a slab gel. An electrical potential was then applied to the slab gel. Methylene Blue in the capillaries underwent electrophoretic migration, forming clear blue bands in the gel. The bands functioned as a transmission grating, and diffracted a laser beam. The intensity of the diffracted light increased with increasing the concentration of methylene blue.

Journal Article↗

PubMed2002

Band broadening caused by the multiple labeling of proteins in micellar electrokinetic chromatography with diode laser-induced fluorescence detection.

When a labeling reagent is used, in the determination of proteins by capillary electrophoresis with laser-induced fluorescence detection, the multiple labeling of proteins frequently occurs, which can degrade the separation efficiency. In order to understand the influence of the multiple labeling of proteins on separation efficiency, the band broadening caused by a labeling reaction between bovine serum albumin (BSA) and a cyanine fluorescent dye (Cy5) was investigated using micellar electrokinetic chromatography in conjunction with diode laser-induced fluorometry. With the aid of an internal standard, methylene blue, the height equivalent to the theoretical plate (HETP) ratio of BSA to methylene blue was used as an indicator for band broadening under optimum separation conditions. Labeling conditions, including reaction buffer pH, reaction time, and initial concentration of Cy5 to bovine serum albumin, were found to influence the HETP ratio. The separation efficiency for the labeled protein was degraded by experimental conditions employed in the labeling, which indicates an increase in the heterogeneity of the final products.

Chromatography, Micellar Electrokinetic Capillary↗

PubMed2002

Ultratrace analysis based on Hadamard transform capillary electrophoresis.

Hadamard transform capillary electrophoresis, which is based on a multiple sample injection technique, was combined with laser-induced fluorometry and utilized in the determination of analytes at subpicomolar levels. The sensitivity was substantially improved by increasing the order, i.e., the number of elements, of the Hadamard matrix. In fact, the signal-to-noise ratio was enhanced 18-fold by the use of a matrix of order 2047. A feasibility study was carried out by computer simulation to study the detection of an average of less than a single molecule in a single injection volume. The signal peak was clearly observable even under conditions at which only 0.3 molecule is present in the volume. Thus, this approach is potentially useful for ultratrace analysis, in which conventional single-injection capillary electrophoresis cannot be applied.

Journal Article↗

PubMed2002

Micellar electrokinetic chromatography with diode laser-induced fluorescence detection as a tool for investigating the fluorescence labeling of proteins.

We describe the use of micellar electrokinetic chromatography with diode laser-induced fluorometry (MEKC-DLIF) as a tool to characterize a labeling reaction between sulfoindocyanine succinimidyl ester (Cy5), a cyanine fluorescent dye, and a model protein, bovine serum albumin (BSA). To decrease the influence of imprecise injection, methylene blue was added as an internal standard. Labeled BSA was completely separated from the unconjugated Cy5, and methylene blue under optimized conditions. A kinetic study of the reaction was performed by changing some parameters, such as reaction buffer pH, reaction buffer concentration and the initial concentration of BSA. A comparison between the current method and traditional methods was made.

Animals↗

PubMed2002

Determination of dye/protein ratios in a labeling reaction between a cyanine dye and bovine serum albumin by micellar electrokinetic chromatography using a diode laser-induced fluorescence detection.

The degree of labeling, i.e., dye/protein ratio (D/P) is important for characterizing properties of dye labeling with proteins. A method for the determination of this ratio between a fluorescent cyanine dye and bovine serum albumin (BSA), based on the separation of the labeling mixture using micellar electrokinetic chromatography with diode laser-induced fluorescence detection, is described. Two methods for the determination of D/P were examined in this study. In these methods, a hydrolysis product and impurities, which are usually unfavorable compounds that are best excluded for protein analysis, were utilized to determine the amounts of dye bound to BSA. One is a direct method in which a ratio of the peak area of BSA to the total peak area of all the products produced in the labeling reaction was used for determining the average number of dye molecules bound to a single BSA molecule. The other is an indirect determination, which is based on diminution of all peak areas related to the products except for the labeled BSA. These methods were directly compared by means of a spectrophotometric method. The experimental results show that the indirect method is both reliable and sensitive. Therefore, D/P values can be determined at trace levels using the indirect method.

Animals↗

PubMed2002

A tunable picosecond dye laser for use in dioxin analysis.

A distributed-feedback dye laser with a quenching cavity was designed and constructed for generating a tunable picosecond pulse with a narrow spectral linewidth. This nearly transform-limited pulse was succeedingly amplified by a triple-pass off-axis amplifier. The pulse duration and the spectral linewidth were 60 ps and 9.4 pm, respectively. The amplified pulse was frequency-doubled by second-harmonic generation, producing a 0.5-mJ pulse with no background emission. The potential advantage of this laser in the analysis of dioxin based on supersonic jet/resonance-enhanced multiphoton ionization/mass spectrometry is discussed.

Journal Article↗

PubMed2002

Use of ring-repeller, double-skimmer electrodes for efficient ion focusing in mass spectrometry.

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.

Journal Article↗