Purification and characterization of an extracellular alpha-neoagarooligosaccharide hydrolase from Bacillus sp. MK03

An agarolytic bacterium was isolated from soil in Gifu prefecture, Japan, and identified as Bacillus sp. strain MK03. The strain secreted neoagarooligosaccharide hydroluse into the culture medium. The enzyme was purified 49.7-fold from the culture fluid by ammonium sulfate precipitation and anion-exchange and gel-filtration column chromatographic methods. The purified enzyme appeared as a single band on polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate (SDS)-PAGE. Estimations of the molecular mass by gel filtration and SDS-PAGE gave values of 320 kDa and 42 kDa, respectively, indicating that the enzyme is octametric. The enzyme cleaved the alpha-1,3 linkage in neoagarobiose to produce 3,6-anhydro-L-galactose and D-galactose. It also selectively cleaved the alpha-1,3 linkage at the nonreducing end in neoagarotetraose or neoagarohexaose to give 3,6-anhydro-L-galactose and agarotriose or agaropentaose. The optimum temperature and pH for the enzyme were 30 degrees C and 6.1, respectively. The N-terminal amino acid sequence showed no homology to sequences of other known neoagarooligosaccharide hydrolases and agarases.     

Highly dense protein layers confirmed by atomic force microscopy and quartz crystal microbalance

Protein adsorption on a gold surface is investigated by comparing the results of quartz crystal microbalance method and atomic force microscopy. The adsorption of streptavidin on functional gold surfaces is directly monitored by a quartz crystal microbalance, and confirmed by atomic force microscopy. For this investigation, a modified gold substrate is fabricated to obtain a topographic image of streptavidin molecules. Both methods show a correlation in terms of the highly dense protein single-layer formation, and the modified gold electrode shows a slightly denser protein layer formation because of the difference in substrate geometry as compared with that of a mica surface.     

Isolation and analysis of lipase-overproducing mutants of Serratia marcescens

We have isolated a lipase-overproducing mutant, GE14, from Serratia marcescens 8000 after three rounds of N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis. The mutant GE14 produced 95 kU/ml of extracellular lipase in the lipase medium, which was about threefold higher than that of produced by the original strain 8000. Enzymatic characteristics including specific activity of purified lipases from culture supernatants of GE14 and 8000 were almost same. The lipase gene (lipA) of GE14 contained two base substitutions; one in the promoter region and another in the N-terminal region of the lipA gene without an amino acid substitution. Promoter analysis using lipA-lacZ fusion plasmids revealed that these substitutions were responsible for the increase in the lipA expression level, independently. In contrast, no base substitution was found in the genes encoding the lipase secretion device, the Lip system. In addition, the genes coding for metalloprotease and the cell surface layer protein which are both secreted through the Lip system and associated with extracellular lipase production, also contained no base substitution. The strain GE14 carrying a high-copy-number lipA plasmid produced a larger amount of the extracellular lipase than the recombinant strains of 8000 and other mutants also did, indicating that GE14 was not only a lipase-overproducing strain, but also an advantageous host strain for overproducing the lipase by a recombinant DNA technique. These results suggest that the lipase-overproducing mutant GE14 and its recombinant strains are promising candidates for the industrial production of the S. marcescens lipase.     

Determination of alkylphenols,bisphenol A,benzophenone and phthalates in containers of baby food,and migration into food simulants

Infants, whose growth and development are highly dependent upon the endocrine system, are particularly vulnerable to endocrine disruptor exposure. In our study, we examined whether or not alkylphenols, bisphenol A, benzophenone, di(2-ethylhexyl)adipate and 10 phthalates are present in containers of baby food. Furthermore, the migration into food-simulating solvents was also examined. Sixteen types of containers of baby food, obtained on the Japanese market in 2001, were examined: 8 glass bottle types, 1 plastic type, 1 metallic package (laminated with plastic) and 6 multilayered laminated film packages. Alkylphenols, benzophenone, bisphenol A and di(2-ethylhexyl)adipate were not detected in any of the samples. On the other hand, dibutylphthalate (DBP) was detected in 7 samples (1.2-9.1 micrograms/sample), and di(2-ethylhexyl)phthalate (DEHP) was detected in 15 samples (1.3-18 micrograms/sample). Migration tests were carried out for 5 samples, and the levels of DBP and DEHP that migrated into n-heptane were 0.68-1.7 micrograms/sample and 0.33-3.5 micrograms/sample, respectively. From these results, a baby's intake levels of DBP and DEHP were estimated to be about 1/2,500 and 1/90-300 of the tolerable daily intake (TDI), respectively.     

Advanced energy saving in the reaction section of the hydro-desulfurization process with self-heat recuperation technology

The reaction section of the naphtha hydro-desulfurization (HDS) process is a heating and cooling thermal process consisting of a feed/effluent heat exchanger and a fired heater. Energy savings are fundamentally made as a result of the maximized heat recovery in the heat exchanger and the reduced heat duty of the fired heater. To achieve further energy saving in the process, “self-heat recuperation technology” (SHRT) was adopted. In this technology, a compressor was introduced. The suction side of the compressor needed a lower pressure and the feed stream evaporated much easily. The discharged side of the compressor satisfied the operating conditions of both pressure and temperature at the inlet of the reactor. And the reactor effluent stream was able to be used completely to preheat and vaporize the feed stream. All the heat in the process stream was re-circulated without using a fired heater. SHRT was applied to the naphtha HDS process of 18,000 barrel per stream day (BPSD) in the refinery and the mass and energy balance of the process was calculated using commercially available simulation software, Invensys PROII version 8.1. This process-simulation case study confirmed that despite there being no more energy saving potential in the conventional process that makes use of a fired heater, the advanced process with SHRT can reduce the energy consumption significantly by using the recuperated heat of the feed stream.     

A 557-mW, 2.5-Gbit/s SONET/SDH regenerator-section terminating LSIchip using low-power bipolar-LSI design

A regenerator-section terminating digital large-scale-integration chip for an STM-16 (2.5-Gbit/s synchronous optical network/synchronous digital hierarchy) regenerator has been developed using low-power bipolar technologies. The high-speed performance of bipolar devices enabled four or more chips, including a demultiplexer and a multiplexer, to be integrated into a single chip. The low-power dissipation of 557 mW, only about one-tenth that of previously reported chips, was achieved through the use of four design steps: one-chip integration architecture, power management, 2.5-V emitter-coupled logic, and power optimization     

An innovative modularity of heat circulation for fractional distillation

A novel modularity of heat circulation for distillation process, which reduces the energy consumption, is proposed. By incorporating compressors and heat exchangers, the heat of the distillate is recuperated and exchanged with the heat of the feed streams. The proposed technology achieves the reduction in the required energy more than 75% as compared with a benchmark process which uses external heat source for heating. This shows the proposed modularity of heat circulation for distillation process is very promising technology to drastically reduce energy demand for distillation.     

Giant electric double-layer capacitance of heavily boron-doped diamond electrode

The physical and electrochemical properties of boron-doped polycrystalline diamond electrodes, prepared with various B/C ratio, i.e., 0.1%, 0.5% (BDD-A), 1% (BDD-B), and 5% (BDD-C), were investigated. Electrochemical measurements of the heavily boron-doped films (BDD-C) showed giant electric double-layer capacitance and activity which is significantly larger than BDD-A and BDD-B as well as glassy carbon electrodes. However, interestingly, actual boron concentration of BDD-C was observed to be almost the same as that of BDD-B by secondary ion mass spectroscopy (SIMS) and glow discharge optical emission spectroscopy (GDOES) analysis. It is suggested that the large capacitance is due to a few sp²-bonded carbon impurities, which was observed only in BDD-C, although the amount of the sp²-bonded nondiamond species are very small. In the present work, the reason for the interesting electrochemical properties of heavily boron-doped diamond electrodes is discussed. Furthermore, dimensional stability of the electrodes was also confirmed by conducting harsh anodic treatment.     

First results on U levels in global fallout

The global fallout ²³⁶U level in soil was deduced from measurements of ²³⁶U, ^239+240Pu and ¹³⁷Cs in surface soils which are solely influenced by global fallout. A total of 12 soil cores from the depths of 0-10, 0-20 and 0-30 cm were collected at a flat forest area in Japan. Concentrations of ^239+240Pu and ²³⁸U were determined by α-particle spectrometry, while the ²³⁶U/²³⁸U ratio was measured with accelerator mass spectrometry (AMS). Consistent ²³⁶U/²³⁹Pu ratios between 0.212 and 0.253 were found. Using this ratio, the total global fallout of ²³⁶U on the earth is estimated to be as much as ca. 900 kg. This knowledge will contribute to the promotion of research on U isotopes, including ²³⁶U, for the fields of geo-resources, waste management and geochemistry.