Kazuo Nakamura

Elucidation of optimum conditions for immobilization of viable cells by using calcium alginate

Abstract Different factors which affect the stability of calcium alginate gel beads entrapping viable cells during fermentation were investigated. It was found that among others, the initial population of cells per ml of gel beads, the length of period of incubation in CaCl 2 solution, and the concentration of sodium alginate used for the immobilization were the most important factors affecting the stability of the gel beads during fermentation. By using an initial cell population of about 10 5 cells per ml of 2.0% sodium alginate, and incubating the beads for at least 22 h in a CaCl 2 solution after immobilization, the percentage of beads which developed cracks during fermentation was highly reduced. Also, without the addition of CaCl 2 into the fermenting broth, the gel beads were stable for nine consecutive batch fermentations.

Simultaneous removal of hydrogen sulfide and trimethylamine by a bacterial deodorant

Abstract Simultaneous removal of hydrogen sulfide (H2S) and trimethylamine (TMA) using a bacterial deodorant was investigated. The bacterial deodorant consisted of Thiobacillus thiooxidans JCM7814 cells and sodium citrate buffer. H2S was biologically removed by T. thiooxidans cells, which oxidized the H2S at a maximum oxidation rate of 0.84 mmol-H2S/g-cell·min in a reaction mixture based on the sodium citrate buffer. TMA was chemically removed by a neutralizing reaction with the citric acid contained in the buffer. The removability of both malodorous gases depended on the pH and the citrate concentration in the buffer. Appropriate buffer conditions for simultaneous removal of the gases were: pH range, 2.0–4.0; citrate concentration, 0.2 M. The bacterial deodorant simultaneously removed 14.0 μl/l H2S and 25.7 μl/l TMA over a period of 30 min in a sealed glass vessel (gas-phase volume, 3 l), in which the initial concentrations of the H2S and TMA generated were 20 and 30 μl/l, respectively. No decrease removability in removal capability was observed over 50 test repetitions.

Microbial sensor for selective determination of sulphide

A microbial sensor consisting of immobilized Thiobacillus thiooxidans, a gas-permeable membrane, and an O2 electrode was prepared for the determination of sulphide. When a sample solution containing sulphide was passed into the flow cell, the output of the microbial sensor decreased markedly with time until a steady state was reached. The total time required for an assay was 20–30 min by the steady-state method. In the pulse method, the total time required for an assay was about 5 min. A linear relationship was obtained between the sensor output and the concentration of sodium sulphide below 0.40 mm. The minimum detectable concentration of sodium sulphide was 0.02 mm. Selectivity of the sensor was satisfactory. The microbial sensor was applied to the determination of sulphide in spring water. A good agreement was obtained between the microbial sensor and the methylene blue method. The regression coefficient was 0.97 for five experiments. The activity of the microbial membrane was stable for more than 25 days. The response was reproducible with 2.5% of the relative standard deviation when a sample solution containing 0.2 mm sodium sulphide was employed. *** DIRECT SUPPORT *** AG903053 00005

Determination of free sulphite in wine using a microbial sensor

SummaryA microbial sensor of immobilized Thiobacillus thiooxidans S3 cells was assembled to determine free sulphite in wine. Sulphite oxidation activity of the immobilized cells was sufficiently high for use even after 3 months storage at 4° C. The sensitivity of this sensor was 116 nA·1·mg-1 for sulphur dioxide. The relationship between the current decrease and the sulphur dioxide concentration was linear up to 17 mg·1-1. The sampling rate achieved was 10 min per sample including washing time. This sensor method needed no pretreatment of wine samples, and wines diluted with 5 mM sulphuric acid solution could be directly introduced in the computer-aided analysis system. The pigments in red wine did not disturbed the analysis.

Stabilization of freeze-dried Thiobacillus thiooxidans cells as a bacterial deodorant for removal of hydrogen sulfide

Abstract Freeze-dried cells of Thiobacillus thiooxidans exhibited sulfide-oxidizing activity similarly to intact cells, but the activity level was reduced during storage. The decrease in the activity was more marked when the freeze-dried cells were stored at higher temperature than −20°C and under more aerobic conditions than 0% of oxygen partial pressure (pO2). Nonreducing disaccharides such as trehalose and sucrose were effective in maintaining the sulfide-oxidizing activity of the freeze-dried cells. The freeze-dried cells prepared in the presence of 10% (w/v) nonreducing disaccharides showed more than 80% of the initial activity for 60 d even though the cells were stored at 25°C under atmospheric conditions. The freeze-dried cells were used as a bacterial deodorant for removal of hydrogen sulfide (H2S) from gas phase.

Purification and properties of isocitrate lyase from Candida brassicae E-17

Abstract Isocitrate lyase (EC 4.1.3.1) was purified from acetate-grown cells of Candida brassicae E-17, by ammonium sulfate fractionation and DEAE-cellulose and Sephadex G-200 gel filtration column chromatographies. The purified enzyme was electrophoretically homogeneous. The molecular weight of this enzyme was 290,000 by gel filtration, and it was composed of four identical subunits whose molecular weights were 71,000 each. The pH and temperature optima were 6.8 and 37°C, respectively. The enzyme was stable from pH 6.0 to 7.0. The enzyme was activated by Mg 2+ and the maximum activity was obtained with a concentration of 8 mM Mg 2+ . The enzyme was also activated by Mn 2+ and Ba 2+ . The activity of this enzyme was stimulated by reducing agents. The K m values for dl -isocitrate were 1.5 mM in sodium phosphate buffer and 0.62 mM in imidazole-HCl buffer.

A microbial method using whole cells of Thiobacillus thiooxidans for measuring sulphate in waters

Abstract A microbial method to determine sulphate concentration in water was developed on the basis of sulphate-dependent acid phosphatase (APase) in whole cells of Thiobacillus thiooxidans. The activity of the APase was determined colorimetrically by using p-nitrophenylphosphate as substrate. The APase was activated by sulphate. A linear relationship was obtained between the activity of the APase and the concentration of sulphate in the range 0–0.6 mM. Therefore, the sulphate concentration was estimated from the APase activity, represented by the absorbance (A400). The microbial method was applied to the determination sulphate in water. The lower limit of detection was 0.02 mM, the relative standard deviation being 2% for 10 measurements on a standard sample. As for practical samples, which were taken from rain, river and tap water, good agreement was obtained between the values measured by the microbial method and those given by a conventional barium chloranilate method. The relative standard deviation was 2.1% for 12 measurements of tap water. The activity of the APase was stable over a period of more than 100 days when the cells were stored in 0.1 M sodium acetate/acetic acid buffer (pH 5.0) at 4 °C.

Reduction of horse heart ferricytochrome c at platinum and gold electrodes modified by various ad-atoms

Abstract The electrochemical behaviour of cytochrome c was examined on platinum and gold electrodes modified by various ad-atom species (Cu, Ag, Hg, Sn, As, Sb, Bi, S, Se and Te). It was found that direct electron transfer occurs between cytochrome c molecules and platinum electrodes modified by Ag, Sn and S ad-atoms or gold electrodes modified by Ag, Hg, Sn, Sb, S and Se. The enhancement effects caused by these ad-atoms exhibit maxima at a coverage between 0.35 and 0.45. The primary effect caused by ad-atoms results from the suppression of the strong bond strength of cytochrome c molecules to the surface, and the secondary effect results from the electrostatic attraction.

Microwave Sterilization by TiO2 Filter Coated with Ag Thin Film

Microwave sterilization for E.coli and/or B. subtilis was investigated. We prepared the filters made of A₂O₃ wool coated with Ag and/or TiO₂ and the urethane foams filter. Ag and TiO₂ played an important role in the sterilization system, because OH radical (*OH) may be generated on the surface of the filter. We discussed about the MW sterilization mechanism by detecting the *OH from XPS analysis of V₂O₈ thin films. We proposed the mechanisms of microwave sterilization, which was mentioned about *OH effects.