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Biomedical subjects

Totaro Imasaka

Publications and source records attributed to Totaro Imasaka.

At least 19 recordsLinked to original sources
PubMed2007

Application of Hadamard transformation to MEKC.

The Hadamard transform (HT) technique, which permits the S/N in CE to be improved, was applied to MEKC. Multiple sample injection of fluorescent analytes according to a Hadamard code sequence was performed using an optically gated sample injection technique, in which a sample plug was produced based on photodegradation by irradiation with an intense laser beam. The capillary and reservoirs were filled with a sample solution containing buffer components and SDS as a pseudostationary phase. A preliminary study confirmed that fluorescein ion could be photobleached in the presence of SDS. The optically gated sample injection technique was then applied to multiple sample injection, based on a Hadamard matrix. The S/N in the electropherogram obtained by HT-MEKC was improved substantially compared to that obtained by a single injection method. When the technique was applied to the separation of several amino acids labeled with FITC, the S/N ratio for each amino acid was enhanced, without any evidence of degradation in separation resolution. Moreover, HT-MEKC was applied to the analysis of amino acids contained in a Japanese beverage, resulting in improved S/Ns for the amino acids.

Amino Acids↗
PubMed2006

On-column capture of a specific protein in capillary electrophoresis using magnetic beads.

A method for capturing specific molecules separated by CE has been explored. To demonstrate on-column capture of migrating analyte molecules, two detection windows were fabricated on a capillary. Magnetic beads containing immobilized molecules that react with the specific molecules under study were placed between the detection windows in the capillary using magnets. Molecules in a sample solution injected into the capillary were separated and detected at the first detection window. After passing through the first detection window, the separated molecules encountered the magnetic beads, where the specific analyte was captured. As a result, the peak area for those analyte molecules decreased or disappeared completely at the second detection window. Rabbit IgG and carbonic anhydrase were employed to demonstrate on-column capture of a specific molecule. For rabbit IgG, magnetic beads containing the immobilized antibody (anti-rabbit IgG) were used. Rabbit IgG molecules were captured on the magnetic beads during CE migration. Furthermore, the capture of carbonic anhydrase was demonstrated by the reaction between magnetic beads (containing immobilized anti-rabbit IgG) and anti-carbonic anhydrase (rabbit IgG), before the beads were packed in the capillary. After packing the magnetic beads in the capillary, a mixture of two proteins was injected into the capillary. Two proteins were detected at the first detection window, while the peak corresponding to carbonic anhydrase disappeared at the second detection window. The results show that using an appropriate antibody, the present technique would be applicable to any proteins.

Animals↗
PubMed2006

Fluorescence lifetime imaging microscope consisting of a compact picosecond dye laser and a gated charge-coupled device camera for applications to living cells.

An inverted microscope was combined with a compact dye laser with a pulse width of <190 ps and an intensified charge-coupled device (ICCD) camera with a minimum gate width of 200 ps. The resulting fluorescence lifetime imaging microscope, which has a temporal resolution of 340 ps, was used to measure the fluorescence lifetime of polymer microspherers. The results indicated a fluorescence lifetime of 0.9 ns. The present analytical instrument was also employed in an evaluation of biological cells after labeling them with SYTO 13, a fluorescent dye.

Cell Line, Tumor↗
PubMed2006

Gas chromatography/multiphoton ionization/mass spectrometry of polychlorinated dibenzofurans using nanosecond and femtosecond lasers.

Gas chromatography/multiphoton ionization/time-of-flight mass spectrometry (GC/MPI/TOF-MS) using a femtosecond laser source has been developed and applied to analysis of an authentic sample containing five polychlorinated dibenzofurans (PCDFs). The ionization efficiency was substantially improved for compounds such as tetrachlorinated dibenzofuran (tetraCDF) and pentachlorinated dibenzofuran (pentaCDF) using a femtosecond laser, instead of a nanosecond laser currently used for this purpose. The relatively poor efficiencies in nanosecond ionization can be attributed to the short lifetimes of the singlet excited states that are efficiently relaxed to triplet levels by intersystem crossing. The detection limits for pentaCDF isomers, i.e., 1,2,3,8,9-pentaCDF (non-toxic) and 1,2,3,7,8-pentaCDF (toxic), were determined to be ca. 100 pg, when the femtosecond laser was employed. The present method has the potential for use in the detection and determination of toxic PCDD/Fs in exhaust gas from an incinerator, because of the high inherent selectivity of GC and the high sensitivity of MPI/TOF-MS.

Journal Article↗
PubMed2005

Suppression of electroosmotic flow and its application to determination of electrophoretic mobilities in a poly(vinylpyrrolidone)-coated capillary.

A hydrophilic polymer, poly(vinylpyrrolidone) (PVP), was employed for suppressing the electroosmotic flow (EOF). A capillary was filled with aqueous PVP solution for coating the capillary wall with PVP; the PVP solution was then replaced by a migration buffer solution containing no PVP. Three types of PVP with different molecular weights were examined. The EOF was suppressed more effectively as the molecular weight of PVP increased. The EOF in the coated capillary was approximately 10-fold smaller than that of a bare capillary and was constant in the pH range of 6-8. The suppressed EOF was stable even when no PVP was added to the migration buffer. However, the EOF increased significantly when sodium dodecyl sulfate was added into the migration buffer. The method was applied for determining the electrophoretic mobilities of inorganic anions that have negative electrophoretic mobilities larger than the electroosmotic mobility of the bare capillary. A novel method for determining the electrophoretic mobilities was proposed based on the linear relationship between electric current and electrophoretic mobility. The electrophoretic mobility was proportional to the electric current. Therefore, the intercept of the regression equation represents the electrophoretic mobility at room temperature. The electrophoretic mobilities were in good agreement with the absolute electrophoretic mobilities.

Electrophoresis, Capillary↗
PubMed2005

Capillary electrophoresis immunoassay based on an on-column immunological reaction.

An on-column immunological reaction was employed to achieve simple and rapid analysis in an immunoassay based on capillary electrophoresis using semiconductor laser-induced fluorescence detection. Human serum albumin (HSA) labeled with sulfoindocyanine succinimidyl ester (Cy5), a fluorescent compound with an absorption maximum at 649 nm, was used as a fluorescent probe for the immunoassay. In a binding assay, with anti-HSA as the analyte molecule, Cy5-HSA was injected in a capillary column followed by the injection of anti-HSA so as to form individual zones. By applying a potential, the anti-HSA reacted with Cy5-HSA at the boundary between Cy5-HSA and anti-HSA zones, since anti-HSA has a higher electrophoretic mobility than Cy5-HSA. Furthermore, the on-column method enhances the sensitivity by injecting a large volume of the sample. Free Cy5-HSA and its immunocomplex with anti-HSA were separated with less degradation in resolution than that predicted from the injection time of anti-HSA, even when the injection time for anti-HSA was increased. The ratio of the peak area of the complex to that of the total Cy5-HSA (free Cy5-HSA and the complex) increased in proportion to the injection time of anti-HSA. As a result, the detection limit was improved up to eight-fold (the concentration detection limit, 0.007 mg mL(-1), for an injection time of 240 s, compared to that obtained using an off-column sample preparation. Furthermore, the on-column reaction method was applicable to an immunoassay to determine native HSA, in which native HSA and Cy5-HSA react with anti-HSA stepwise. The detection limit in the stepwise reaction immunoassay was 0.005 mg mL(-1), which is 14 times lower than that in an off-column method, with the analysis time less than 10 min as the result of increasing the injection time of native HSA. In addition, the present on-column immunoassay was applied to the sample containing a high concentration of salts for investigating the effect of salts in the sample solution.

Carbocyanines↗
PubMed2005

Chlorine substitution pattern, molecular electronic properties, and the nature of the ligand-receptor interaction: quantitative property-activity relationships of polychlorinated dibenzofurans.

It was confirmed that both quadrupole moments and polarizabilities of polychlorinated dibenzofurans (PCDFs), which were calculated using Hartree-Fock theory and/or density functional theory, change systematically with the chlorination pattern governing molecular charge distribution. A mathematical model based on ligand-receptor binding and solute-solvent interaction is reported to explain the difference in toxicity between PCDFs. Multiple regression analysis demonstrated that the difference in the potency of aryl hydrocarbon hydroxylase (AHH) and 7-ethoxyresorufin O-deethylase (EROD) inductions is mostly determined by the polarizabilities of PCDFs. This suggests that the interaction of a PCDF with the aryl hydrocarbon receptor (AhR) and its interaction with surrounding molecules in the cytosol are dispersion interactions rather than electrostatic interactions. Quadrupole moment, electron affinity, and absolute hardness do not appear to be significantly correlated with the differences in AHH and EROD activities among PCDFs. The entropy change of dissolution is important in predicting the AHH and EROD activities with good accuracy. A mathematical model is also used to study the differences in AhR binding between PCDFs.

Benzofurans↗
PubMed2005

On-line concentration of a protein using denaturation by sodium dodecyl sulfate.

A novel method for the on-column sample stacking of proteins is described. The strategy takes advantage of interactions between protein molecules and sodium dodecyl sulfate (SDS) monomers. A long plug of a protein sample (either acidic or basic) is injected into a capillary filled with a background electrolyte (BGE) containing SDS. When a potential is applied, the proteins interact with SDS monomers in the BGE to form protein-SDS complexes that migrate more slowly than the corresponding uncomplexed protein, resulting in protein stacking. Both acidic and basic proteins migrate at an almost identical electrophoretic velocity after stacking, which indicates that the protein-SDS complexes formed in the BGE zone have a similar charge/mass ratio. The mechanism of stacking was investigated using a sample consisting of a basic protein, lysozyme, and a small molecule, methylene blue. The findings clearly show that two interactions with SDS occur, a stepwise binding interaction between protein molecules and SDS monomers and an interaction in which the small molecules enter into micelles formed by SDS molecules. The method was also applied to the detection of a protein labeled with a fluorescent labeling reagent at trace levels. The labeled protein was detected even under labeling conditions where the labeling efficiency was too low to detect by short-plug injection.

Electrophoresis↗
PubMed2005

Development of a narrow-band tunable picosecond dye laser and its application to excited-state lifetime measurement of a chlorinated aromatic hydrocarbon.

The development of a distributed-feedback dye laser, with a pulse width and a line width of 25 ps and 8.78 pm, respectively, is described. Using this nearly Fourier-transform-limited pulse, we measured the first singlet-excited-state lifetime of 1,2,4-trichlorobenzene. The tunable picosecond dye laser developed herein has a potential for the lifetime measurement and the efficient multiphoton ionization of aromatic hydrocarbons with a larger number of chlorine atoms and shorter excited-state lifetimes.

Journal Article↗
PubMed2004

Generation of a sub-20-fs single optical pulse by four-wave Raman mixing using two Raman cells filled with molecular hydrogen.

To attempt to expand the spectral domain for pulse shortening, we generated several emission lines by stimulated Raman scattering and subsequent four-wave Raman mixing. The efficiency of generation of the Raman emission was improved by passing the beam through two Raman cells that were connected in series. The group-velocity dispersion induced by a Raman cell window and hydrogen was compensated for by means of a pair of chirped mirrors for pulse compression. By a phase lock of the emission lines in the process of four-wave Raman mixing, the pulse width was reduced from 114 to 17 fs in the second Raman cell.

Journal Article↗
PubMed2004

Selective multiphoton ionization of coplanar polychlorobiphenyls using 266-nm laser emission by gas chromatography/mass spectrometry.

Polychlorinated biphenyls (PCBs), used as a heat exchange oil (Kanechlor, KC-300), can be measured by gas chromatography combined with multiphoton ionization mass spectrometry (GC/MPI-MS). Several compounds, e.g., 4,4'-dichlorobiphenyl and 3,4,4'-trichlorobiphenyl, have nearly the same retention time but can be selectively determined by MPI-MS. Coplanar PCBs are more efficiently ionized using the fourth harmonic emission of a Nd:YAG laser (266 nm), compared to noncoplanar PCBs. Thus, the approach reported herein may be useful in the selective as well as the sensitive analysis of toxic PCBs, contained in old transformers and capacitors that have been mandated by the government to be properly disposed of within 10 years in Japan.

Journal Article↗
PubMed2004

Hadamard transform capillary electrophoresis combined with absorption spectrometry.

A novel injection device for applying absorption spectrometry to Hadamard transform (HT) capillary electrophoresis is described. A small hole, at the center part of the capillary, functions as an inlet port for the sample. The hole is immersed in a sample solution and the end of the capillary that is usually employed for sample introduction is immersed in a buffer solution. An ultraviolet absorption detector is placed between the sample injection port and the other end of the capillary filled with a buffer solution. A high potential is continuously applied between the injection port and the end of the capillary, which allows the sample solution to be introduced into the separation capillary. By application of a higher potential modulated according to a Hadamard code between both ends of the capillary, the buffer solution is injected into the separation capillary. In some preliminary experiments, this injection device was utilized to introduce a single sample segment into a capillary. As expected, a single peak was observed in the electropherogram for a sample containing a single component. This device was then employed for multiple sample injection in HT capillary electrophoresis. An 8-fold improvement in the S/N ratio was observed when the HT technique was used, in which a 255-order of a Hadamard matrix was used, as expected from theory. The present approach was also utilized for the sensitive detection of a sample comprised of multiple components.

Journal Article↗
PubMed2004

A two-color three-photon ionization scheme for the efficient and selective ionization of a chlorinated aromatic hydrocarbon.

A two-color three-photon ionization scheme, for the efficient and selective ionization of a chlorinated aromatic hydrocarbon that has an ionization potential higher than the two-photon energy of the laser used for excitation, is described. In this technique, an ultraviolet (UV) laser, i.e., the second harmonic emission of a fundamental (VIS) laser, is used for excitation and a UV and VIS laser for the subsequent two-photon ionization from the electronic excited state. A sample of o-chlorophenol was used as a model compound to demonstrate the advantage of this technique. The signal in supersonic jet/resonance-enhanced multiphoton ionization/mass spectrometry was increased approximately 4 times by the introduction of the VIS beam, when the polarization was adjusted to be parallel to the UV beam. Thus, the two-color three-photon (2UV+VIS) ionization scheme is more sensitive than one-color three-photon (3UV) ionization. The merits of this method over other ionization schemes such as two-color two-photon (UV(1)+UV(2)) ionization are discussed in terms of sensitivity and selectivity in spectrometric analysis.

Letter↗
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

Multiphoton ionization mass spectrometry of chlorophenols as indicators for dioxins.

Mono-, di- and trichlorophenols were measured using resonance-enhanced multiphoton ionization mass spectrometry (MPI-MS) combined with supersonic jet (SSJ) or effusive molecular beam (EMB) spectrometry. All mono- and dichlorophenols, except 2,6-dichlorophenol, provided sharp and structured MPI spectra for the S1 <-- S0 transition. Selectivity and sensitivity were both enhanced when SSJ spectrometry was used, compared with EMB spectrometry, because of a narrower linewidth in the MPI spectrum, given by molecular cooling by supersonic jet expansion. The ionization efficiency decreased with increasing number of chlorine substituents for the chlorophenols, since they have shorter excited-state lifetimes and require three photons for ionization. Some of the chlorophenols, which are toxic themselves, have the potential for use as indicators for analysis of polychlorinated dibenzo-p-dioxin/dibenzofurans in flue gases emitted from an incinerator.

Air Pollutants↗
PubMed2003

Detection of 27 molecules in a single injection volume by Hadamard transform capillary electrophoresis.

A concentration detection limit of 100 fM was achieved for the fluorescein ion by improving the experimental setup used for Hadamard transform capillary electrophoresis. Two argon-ion lasers, a gating laser for sample injection and a probe laser for the excitation of analyte molecules, were employed for the efficient photodegradation of analyte molecules in laser-induced fluorescence detection using an optically gated sample-injection method. In addition, a dichroic mirror, located in the pathway of the probe laser was used to exclude the other lines of the argon-ion laser. Using a Hadamard matrix on the order of 2046, the concentration limit of detection for fluorescein ion was determined to be 100 fM at S/N = 3, in which the average number of molecules in a single injection volume was calculated to be 27. The influences of the output power in both the gating and probe lasers on the sensitivity are also discussed.

Journal Article↗