Natsuki Ikuta

Operational modes for transient isotachophoretic preconcentration-capillary zone electrophoresis and detectable concentration of rare earth ions.

Operational modes for transient isotachophoretic preconcentration capillary zone electrophoresis (tr‐ITP‐CZE) were studied by using 5 νM and 0.5 νM rare earth mixtures as analytes in comparison with field‐enhanced sample stacking. After examination of several operational modes for tr‐ITP, it was found that tr‐ITP effectively occured even if both the leading electrolyte and the terminating electrolyte were injected after the sample plug. This was explained as the result of a field‐enhanced stacking for both sample components and the leading and terminating ions. The observed theoretical plate numbers were 4–20 times higher than those obtained by normal stacking; and the estimated low limit of detectable concentration of rare‐earth elements (REE) was ca. 0.1 νM which was 2.5 times lower compared to normal stacking. For the 0.5 νM sample, a concentration factor of 20 000 could be achieved after only tr‐ITP.

Numerical simulation for capillary electrophoresis: I. Development of a simulation program with high numerical stability

A computer program was developed for the simulations of capillary electrophoresis on the basis of unsteady partial differential equations. Since a new algorithm (shifted upwinding approximation) was used to suppress numerical divergence and oscillation, the program succeeded in generating only physically meaningful peaks at the cost of slight overestimation of ionic diffusion. The mathematical model and equations used in the program were described in detail, and simulation performance was compared among the previously reported programs, confirming the numerical stability of our program. As an application example, a system peak in indirect UV detection was simulated.

Generation of an X-ray microbeam for spectromicroscopy at SPring-8 BL39XU

A pair of elliptical mirrors (KB mirror) was designed and fabricated to realize an energy tunable x-ray microbeam for spectromicroscopy at SPring-8 BL39XU. As is commonly recognized, the obtainable beam size with the aspherical total reflection mirrors is strongly affected with the slope error of the mirror. Considering that the extremely high brilliance of the undulator radiation from the SPring-8, the small mirror size and the small mirror-to-focus distance were employed to minimize effects of the slope error. Preliminary evaluation of the KB mirror was carried out using 10 keV monochromatized undulator radiation. Alignment of the mirror was assisted by the beam monitor system composed of a scintillator and a CCD, and the beam size less than 5 microm can be easily achieved even when the source was fully used. The beam size obtained with this experiment was 2 x 4 microm2 with the photon flux of 1 x 10(10) photons/s. Smaller beam size may be expected with the use of intermediate slits. Characterization of trace elements with the spatial resolution will be carried out by using x-ray fluorescence (XRF) analysis and x-ray absorption fine structure (XAFS) measurements with XRF yield method.

Change of migration time and separation window accompanied by field-enhanced sample stacking in capillary zone electrophoresis.

When field‐enhanced sample stacking was used in capillary zone electrophoresis (CZE) analysis of cations, the decrease of migration time and the reduction of separation window was observed with increase of sample plug length. A simple equation expressing the migration velocity in the stacking process was derived to explain the above phenomenon. From experiments and theoretical consideration, we confirmed that this effect was caused by the higher potential gradient and larger eletroosmotic flow (EOF) mobility at the sample plug than those at the supporting electrolyte. A mathematical model appropriate for the computer simulation of such a system was studied considering the experimental results, and it was concluded that electroosmotic velocity (veof) should be introduced to the equation of continuity as a constant.

Standardization of capillary zone electropherograms obtained by using field enhanced sample stacking.

Migration times in a capillary zone electropherogram obtained by using the field enhanced sample stacking technique are strongly affected by the injected sample volume. That is, the migration times significantly decrease with the increase of the sample volume. To avoid inaccurate qualitative analysis due to the above phenomena, the time axis of the electropherograms was converted into an effective mobility axis using our conversion method taking account of the temperature increase in the separation tube and relaxation of the potential gradient of the separation field. After the conversion, accurate qualitative analysis was possible in spite of drastic change of the migration time, suggesting our conversion method could be successfully used for the standardization of electropherograms obtained even by using the stacking effect. The cause of the decrease of the migration time in the stacking process was briefly discussed.

New method for standardization of electropherograms obtained in capillary zone electrophoresis.

A new method for standardization of electropherograms obtained by capillary zone electrophoresis was proposed, where the migration time axis was replaced by the effective mobility axis. The mobility increase due to temperature increase by Joule heating and the relaxation effect of the potential gradient were eliminated successfully by introducing a temperature coefficient for mobility expression and a delay time, respectively. The precision of the mobility evaluated by the proposed conversion methods was evaluated for a model sample. By using the conversion method, almost the same electropherograms could be obtained even from the electropherograms originally obtained by using different hardware conditions.

Separation of rare-earth ions by isotachophoresis and capillary zone electrophoresis

Abstract The simultaneous separations of 15 rare-earth (RE) ions including yttrium and lanthanide both by isotachophoresis (ITP) and by capillary zone electrophoresis (CZE) were achieved by using the developed electrolyte systems containing α-hydroxyisobutyric acid (HIBA) as main complexing agent and malonic acid as the assistant one. The successful separation of Y3+ from Dy3+ without loss of the separation of Eu3+ and Gd3+ was briefly summarized. Based on the assumed equilibria existed in steady state zone while using such an electrolyte system with binary complexing agents, the effective mobilities of the RE complex ions and the qualitative indicies (RE, the ratio of the specific conductivity of the sample zone to that of the leading zone) for the RE complex zone were theoretically simulated. The reason for the successful separation was clarified from the view of complex formation, and the simulated isotachopherogram for an optimum electrolyte system was then obtained. The validity of the simulation was checked by comparison of the simulated RE with the observed ones. Furthermore, the historic sample didym prepared from a RE ore by Brauner was analyzed by ITP–particle-induced X-ray emission as well as by CZE, where the supporting electrolyte for CZE analysis is 30 mM creatinine containing 4 mM HIBA and 0.4 mM malonic acid buffered by 2-ethylbutryric acid to pH 4.8.

High-sensitive analysis of DNA fragments by capillary gel electrophoresis using transient isotachophoresis preconcentration and fluorescence detection

In this report aimed on further development of a high-sensitivity capillary gel electrophoresis (CGE) method for analysis of DNA fragments, we firstly explored online transient isotachophoresis (tITP) preconcentration combined with fluorescence detection (FD). The fluorescence signal (excitation: 488 nm; emission: 590 nm) was generated using the intercalating dye of ethidium bromide (EB). It was found when the leading electrolyte (LE) was injected behind the sample zone, such a special tITP mode has significant advantages to solve the bubble formation issue and to improve the analytical performance stability. Two standard DNA samples, a 50 bp DNA step ladder and the phiX174/HaeIII digest, were used to evaluate the qualitative and quantitative abilities of the tITP-FD approach. A highly diluted sample (10,000-fold in the water, e.g. the phiX174/HaeIII digest diluted from 500 microg/ml to the 50 ng/ml level) was enriched and detected; the LOD was down to 0.09 ng/ml for the 72 bp fragment, apparently improved more than 1000-fold in comparison with UV detection. Although the RSD of peak areas (n=3) was around 15.5% for the sample was electrokinetically injected, good linearity of peak area response showed that the proposed method is suitable for quantitative analysis.

NUMERICAL SIMULATION FOR CAPILLARY ELECTROPHORESIS. II. RELAXATION EFFECT OF POTENTIAL GRADIENT IN CAPILLARY ZONE ELECTROPHORESIS

Abstract Using our simulation program for CE based on partial differential equations of unsteady state, we found the phenomena at the initial stage of the capillary zone electrophoretic process that a transient zone with very high potential gradient was produced at the front-end of the sample plug and consequently the potential gradient of the rest part without samples decreased. The potential gradient of the zone and the rest part gradually reached the averaged potential gradient (relaxation of potential gradient). This phenomena causes delay of migration time to some degree affecting its reproducibility. This effect is due to discontinuities of the CE system and low mobility of a counter ion in a background electrolyte (BGE). Therefore, in order to improve the delay, discontinuities of solution should be minimized or a counter ion with high mobility in BGE should be selected.

Electropherogram of capillary zone electrophoresis with effective mobility axis as a transverse axis and its analytical utility. I. Transformation applying the hypothetical electroosmotic flow.

For capillary zone electrophoresis, a new method of transformation from migration time to effective mobility was proposed, in which the mobility increase due to Joule heating and the relaxation effect of the potential gradient were eliminated successfully. The precision of the mobility evaluated by the proposed transformation was discussed in relation to the analysis of rare earth ions. By using the transformation, almost the same pherograms could be obtained even from the pherograms obtained originally at different applied voltages.