STD‐NMR‐Based Protein Engineering of the Unique Arylpropionate‐Racemase AMDase G74C

Structure‐guided protein engineering achieved a variant of the unique racemase AMDase G74C, with 40‐fold increased activity in the racemisation of several arylaliphatic carboxylic acids. Substrate binding during catalysis was investigated by saturation‐transfer‐difference NMR (STD‐NMR) spectroscopy. All atoms of the substrate showed interactions with the enzyme. STD‐NMR measurements revealed distinct nuclear Overhauser effects in experiments with and without molecular conversion. The spectroscopic analysis led to the identification of several amino acid residues whose substitutions increased the activity of G74C. Single amino acid exchanges increased the activity moderately; structure‐guided saturation mutagenesis yielded a quadruple mutant with a 40 times higher reaction rate. This study presents STD‐NMR as versatile tool for the analysis of enzyme–substrate interactions in catalytically competent systems and for the guidance of protein engineering.     

Negative Pore Air Pressure Generation in Backfill of Retaining Walls During Earthquakes and its Effect on Seismic Earth Pressure

In order to investigate the seismic behavior of conventional type and geosynthetic-reinforced soil retaining walls, 1-g model shaking tests were conducted. Model walls having a height of about 50 cm were placed on a subsoil layer and backfilled with a layer of dense dry Toyoura sand. They were subjected to several steps of horizontal irregular excitations. As a result, generation of negative pore air pressure in the backfill was observed. The maximum amplitude of the negative pore air pressure during each shaking step increased with the base acceleration. Based on analyses of the measured data, it was inferred that such negative pore air pressure was caused by outward wall displacement relative to the backfill and not by dilative behavior of the backfill. It would cause a reduction in the seismic earth pressures exerted from the backfill. This feature suggests an advantage of a rigid full-height facing for reinforced soil walls over the segmental types of facing. A simplified numerical procedure to evaluate earth pressure was applied while considering the effects of the negative pore air pressure, and it could qualitatively simulate the measured behavior in terms of the seismic earth pressure and the angle of failure plane in the backfill.     

Ca-α SiAlON hollow spheres prepared by carbothermal reduction–nitridation from different SiO2 powders

The formation process of hollow spheres composed of nanosized Ca-α SiAlON particles was investigated using SiO₂ starting powders with different characteristics in particle size, shape and crystalline state. TEM observations showed Ca-α SiAlON hollow spheres composed of a large number of nanosized particles in the products prepared at 1450 °C for 120 min in nitrogen. In all systems, the Ca-α SiAlON hollow spheres were always produced through an intermediate Si–Al–Ca–O liquid phase in the same mechanism, regardless of the characteristics of SiO₂ starting powders used. Spherical solid particles consisted of amorphous phase containing Si, Al, Ca, O and a small amount of N were generated at the initial stage of carbothermal reduction–nitridation. These spherical solid particles changed into hollow particles with the progression of the reaction from the liquid phase to the crystalline Ca-α SiAlON with increasing temperature.     

砂土地基地震液化沉降的两种简易计算方法的对比分析

利用两种简易方法对日本某核电站的砂性土地基在地震液化后的大变形沉降进行了计算和对比分析。这两种简易的实用方法只需要利用标准贯入试验数据就可以操作。计算结果表明,该核电站的砂性土地基在L2级地震作用下会发生较大的地面沉降,并可能对地下管线造成损坏。由于两种方法的计算原理相同,因此计算所得到的地基分层沉降趋势基本一致,但是在数值上存在着较大的差异,这主要是由于两种方法建立的基础试验数据不同以及具体计算方法上存在着差异所造成的。     

Study on the determination of pesticide residues in crops by ion-trap GC/MS/MS

Ion-trap GC/MS/MS was evaluated for the multi-residue determination of pesticides in agricultural products. Matrices were extracted from samples (spinach, carrot, onion and brown rice) with acetone and submitted to gel permeation chromatography, followed by a clean-up step through a graphite carbon cartridge. Thirty-five pesticides were added to either matrix, and analyzed by GC/MS/MS. Detection limits of pesticides by GC/MS/MS was almost the same as those by GC/MS (SIM). Coefficients of variation of peak area in 5 measurements of each pesticide at 0.1 microgram/mL or 0.05 microgram/mL with or without matrices were mostly acceptable, though those of 20 pesticides out of 35 were higher than 10% at a concentration of 0.02 microgram/mL. It was indicated that matrix artifacts, which interfere with GC/MS-Scan analysis, could be eliminated in some cases by using GC/MS/MS.     

Use of multivariate analysis for the improvement in prediction accuracy of bacterial aerobic plate count by flow cytometry

Flow cytometry (FCM) and aerobic plate count (APC) by the culture method were performed on green tea samples spiked with Escherichia coli type strain NCTC9001 (ATCC11775) solutions of different concentrations. In FCM, fluorescence signals from multiple stained bacteria and other fluorophores are detected using detector channels, and recorded as events with a voltage at each channel. FCM data were analyzed in two ways: conventional and multivariate analysis. In the former, the number of events with voltages larger than the defined threshold values was regarded as the predicted APC. In the latter, voltage histograms of all channels were obtained and merged horizontally to serve as explanatory variables. Then a partial least squares regression (PLSR) model was built to predict APC from the histogram data. The coefficient of determination (R²) and the root mean square error (RMSE) between APC by the culture method and that predicted by conventional FCM were 0.916 and 1.08 cfu/ml². The APC values predicted by the PLSR model and those measured were in good agreement with R² of 0.982 and RMSE of 0.417 cfu/ml, which verified the potential of the proposed method for improving APC prediction accuracy by FCM.     

RADIOLYTIC DEGRADATION OF A CROWN ETHER FOR EXTRACT ABILITY OF STRONTIUM

ABSTRACT

Dicyclohexano- 18-crown-6 (DC18C6), dissolved in chloroform, 1-octanol, toluene or cyclohexane, was irradiated by ⁶⁰Co γ-rays. Radiolytic destruction of DC18C6, measured by gas chromatography, decreased in the order: chloroform > cyclohexane > 1-octanol > toluene. Distribution coefficients in the solvent extraction of strontium from nitric acid solution were measured. These distribution coefficients were minimally affected by γ-irradiation in toluene solution, but decreased with the absorbed dose in chloroform solution. It was suggested that the distribution coefficient for the liquid-liquid extraction system was influenced by radiolytic products. The effect of radiolytic products on the distribution coefficients was evaluated by considering DC18C6 remaining in the organic phase after irradiation. The distribution coefficient measured for the organic phase with radiolytic products was larger than that without radiolytic products. The radiolytic products were thus shown to contribute to the increase in the distribution coefficient.     

Improvement in D-E Hysteresis Curve Squareness of PZT Thin Films—Effect of Oxygen Partial Pressure in High-Pressure Post-Annealing

A high-pressure annealing was applied to a post-annealing process for sol-gel derived PZT thin films. The squareness of D-E hysteresis curves changes depending on both total pressure and oxygen concentration. Moreover, the change follows the product of the total pressure and the oxygen concentration, which correspond to oxygen partial pressure P_O2. Where the P_O2 is higher than 0.03 MPa, few of the squareness of the hysteresis curve are excellent. The squareness of the hysteresis curves dramatically improve as the P_O2 decreases. Where the P_O2 is lower than 0.01 MPa, the squareness deteriorates slightly. These changes in the D-E hysteresis curves are thought to be explained by the generation of lead and oxygen vacancies as a function of the P_O2.