Vertically integrated human P450 and oxygen sensing film for the assays of P450 metabolic activities

An assaying method of cytochrome P450 (P450 or CYP) monooxygenase activities for toxicological evaluation of drugs and environmental pollutants was developed by immobilizing P450 on an oxygen sensoring layer. Membrane fractions from E. coli expressing human P450 were entrapped in agarose or silica-based gels and immobilized on 96-well microarrays having an oxygen sensing film at the bottom. The oxygen sensing film was made of an organically modified silica film (ORMOSIL) doped with Tris(4,7-diphenyl-1,10-phenanthroline) ruthenium dichloride (Ru(dpp)(3)Cl(2)). P450 activity toward the substrates was monitored through the fluorescence intensity enhancement due to the oxygen consumption by the metabolic reactions. For the metabolism of chlortoluron, a selective herbicide used to control grass weeds, CYP1A1 immobilized in agarose gel showed a higher activity and stability compared with those in silica gels and free suspensions. The luminescence changing rate evaluated by the dynamic transient method (DTM) could be correlated with the substrate concentration. We also compared the metabolic responses of human P450s (CYP1A1,CYP2C8, CYP2E1, CYP3A4) toward various substances. The use of immobilized P450 on an oxygen sensing layer provides a versatile assaying platform owing to the following features. First, the oxygen sensor can detect metabolic reactions of any P450 species, in contrast with assays using fluorogenic substrates. Second, vertical integration of the oxygen sensor and immobilized P450 enhanced the sensitivity because of the effective depletion of oxygen in the vicinity of the oxygen sensing layer. Third, immobilization enables repeated use of P450 by replacing the substrate solutions using a flow cell. Furthermore, the activity of immobilized P450 was retained at least for 3 weeks at 4 °C, suggesting its long-term stability, which is an additional attractive feature.     

Multiscale seamless‐domain method for solving nonlinear problems using statistical estimation methodology

This article presents a nonlinear solver combining regression analysis and a multiscale simulation scheme. First, the proposed method repeats microscopic analysis of a local simulation domain, which is extracted from the entire global domain, to statistically estimate the relation(s) between the value of a dependent variable at a point and values at surrounding points. The relation is called regression function. Subsequent global analysis reveals the behavior of the global domain with only coarse‐grained points using the regression function quickly at low computational cost, which can be accomplished using a multiscale numerical solver, called the seamless‐domain method. The objective of the study is to solve a nonlinear problem accurately and at low cost by combining the 2 techniques. We present an example problem of a nonlinear steady‐state heat conduction analysis of a heterogeneous material. The proposed model using fewer than 1000 points generates a solution with precision similar to that of a standard finite‐element solution using hundreds of thousands of nodes. To investigate the relationship between the accuracy and computational time, we apply the seamless‐domain method under varying conditions such as the number of iterations of the prior analysis for statistical data learning.     

Fabrication of particle free epitaxal AlN thin films by reactive PLD combined with an electron beam and a rotating crucible

An AlN epitaxial film without projections or spiral growth features was successfully fabricated on a (0 0 0 1) sapphire substrate by reactive laser ablation of a liquid Al target in NH₃ using a 248 nm laser. The liquid Al target was prepared with an electron beam in a rotating crucible. The surface of the rotating liquid Al target was always smooth. Spiral growth features were greatly suppressed. AlN films have a surface roughness less than 0.3 nm. The X-ray rocking curve's narrowest FWHM was 180 arcsec, which is nearly identical to that recently reported in films grown by MOCVD or reactive MBE.     

Analytical solutions to eddy current in carbon fiber-reinforced composites induced by line current

This paper presents analytical solutions to eddy current distribution in carbon fiber-reinforced plastics induced by line current. Derived solutions show that eddy current distribution in unidirectional carbon fiber composite is dependent only on electrical conductivity in the drive current direction when the drive current is directed in the fiber direction or the transverse direction. Moreover, according to the derived analytical solution, skin depth of eddy current depends not only on frequency of drive current but on lift-off and width of the drive current unlike general expression of skin depth. Finally, we discuss improvement of sensitivity of eddy current conductivity sensor using analytical solutions. It is found that analytical solutions can clearly divide effects of magnetic field from drive current and that from eddy current. Sensitivity of eddy current sensor can be improved by 10 times placing pickup coil at region where the effect of magnetic field from eddy current is large.     

Preparation of ultrafine particles of silicon base intermetallic compound by arc plasma method

The purpose of this paper was to prepare ultrafine particles of Si-based intermetallic compounds, TiSi₂, MoSi₂ and VSi₂, by an arc plasma method with hydrogen addition. The properties of the prepared particles were affected by the vapor pressure ratio (Ti/Si, V/Si: 10⁻¹; Mo/Si: 10⁻³) of the constituent metals. The vaporization and condensation rates of the constituent metals should be controlled to prepare intermetallic compound particles in the case of large different vapor pressures. The vaporization rate can be controlled by H₂ concentration in the arc; for example, an increase in H₂ concentration leads to an increase in Ti fraction in the prepared particles in a Si---Ti system. The preparation of ultrafine particles of TiSi₂ was most successful from the 60-wt.% Ti raw material with the 50%-H₂ arc. MoSi₂ particles were prepared from the 85-wt.% Mo raw material with the 50%-H₂ arc. A single phase of VSi₂ particles was prepared successfully from the 60-wt.% V raw material with the 50%-H₂ arc. Another purpose was to investigate the mechanism of vaporization enhancement of particular metals from a metal mixture by hydrogen in arc plasmas. The vaporization enhancement was mainly attributed to the formation of intermediate products such as hydride and/or activity modification by hydrogen in molten metals.     

Biomimetic coating of an apatite layer on poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactic acid); improvement of adhesive strength of the coating

A biodegradable polymer coated with a bonelike apatite layer on its surface would be useful as a scaffold for bone tissue regeneration. In this study, poly(l-lactic acid) (PLLA) was treated with oxygen plasma to produce oxygen-containing functional groups on its surface. The plasma-treated specimen was then alternately dipped in aqueous CaCl₂ and K₂HPO₄·3H₂O solutions three times, to deposit apatite precursors onto the surface. The surface-modified specimen then successfully formed a dense and uniform bonelike surface apatite layer after immersion for 24 h in a simulated body fluid with ion concentrations approximately equal to those of human blood plasma. The adhesive strength between the apatite layer and the specimen surface increased as the power density of the oxygen plasma used increased. The maximum adhesive strength of the apatite layer to the specimen was significantly higher than that to the commercially available artificial bone, HAPEX^TM. The resultant bonelike apatite–PLLA composite would be useful as a scaffold for bone tissue regeneration.     

Piezoresistive effect of plain-weave CFRP fabric subjected to cyclic loading

The present study deals with electrical resistance changes in woven-fabric CFRP during loading. Four kinds of plain weave woven-fabric CFRP laminated specimens are prepared and subjected to cyclic tensile loading that does not cause any damages, and the electrical resistance changes of the specimens are measured experimentally by the four-probe method. As a result, the present study shows that the electrical resistance of a specimen comprised of six ±45° plies decreases remarkably with increasing number of loading cycles. The decrease is caused by shear plastic deformation of ±45° plies. The thickness shrinkage caused by shear plastic deformation increases the number of fiber contacts, and this decreases the interlaminar contact resistance between the plies. For a single ±45° ply, the same electrical resistance decrease caused by the shear plastic deformation is observed, and the magnitude of the decrease is smaller than that of the six-ply laminate tested. This is because the effect of interlaminar contact resistance decrease does not exist for a single ±45° ply. For the six 0°/90° plies, the present study shows that electrical resistance in the through-thickness direction is decreased by out-of-plane plastic deformation of carbon fiber and misalignment of the plies.     

Novel wet electrospinning system for fabrication of spongiform nanofiber 3-dimensional fabric

Electrospinning method is a simple and efficient method to fabricate the nanofiber non-woven mat from a rich variety of functional materials. However, it is difficult to control the 3-dimensional structure of nanofiber fabric via electrospinning method. On that point of view, novel wet electrospinning system was established for the 3-dimensional structure control of nanofiber fabric by combining electrospinning system and wet spinning system. Spongiform Poly (glycolic acid) (PGA) nanofiber 3-dimensional fabric controlling the fiber density was successfully formed with simplicity and cheapness using novel wet electrospinning system. Comparing with usual PGA nanofiber non-woven mat prepared by conventional electrospinning method, spongiform PGA nanofiber fabric had low apparent density and high porosity. The novel nanofiber fabrication system would be very useful for the structure control of the nanofiber fabric.     

Self-Sharpening of Thin Tungsten Electrode in Single, High-Current Discharge: Its Dynamics and Mechanism

Self-sharpening of a thin tungsten electrode occurs in single, high-current discharge. This phenomenon has been observed with a custom-made, high-speed imaging system. The system allowed us to take one hundred images of the electrode shape during and after the discharge with 16 μs intervals. The following selfs-harpening mechanism has been proposed based on observations: (1) During the discharge, when the melt front descends towards the axis of the electrode, while the melt forms a sphere and moves along it due to the surface tension. (2) The melt continues moving even after the discharge, revealing a needle-shaped unmelted part and then solidifies. Detailed studies of discharge conditions to the diameter of the needle-shape formed by the process have been carried out. Results supporting the proposed mechanism are obtained.