Activation of Rhizopus delemar C-lipase activity toward tripropionin by DDT and aldrin was studied by using a purified enzyme preparation. Titration experiments of lipase with the pesticide indicated that two molecules of the pesticides first bind to the lipase molecule without activation of the enzyme activity, but this binding facilitates the binding of further pesticide molecules to afford highly active pesticide-lipase complexes, a 9 : 1 DDT-lipase complex and a 7 : 1 aldrin-lipase complex, with 4.4 times and 16 times the activity of intact lipase, respectively. Polyacrylamide gel electrophoresis resolved the intact lipase and the pesticide-lipase complexes, giving each as a distinct protein band, suggesting the formation of irreversibly bound complexes. Gel filtration results also supported this hypothesis. The titration experiments also suggested that the binding of the first two molecules of the pesticides occurs at different specific sites of the lipase molecule for DDT and aldrin, influencing the nature of the enzyme ; the binding of the first two DDT molecules resulted in the formation of a DDT-activated complex, while treatment of the intact lipase with a mixture of DDT and aldrin gave an aldrin-activated complex.

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To understand the nature of the chemical substances responsible for the disease"Yusho, "a mixture of polychlorinated biphenyls (PCBs), polychlorinated dibenzofurans (PCQs) and polychlorinated dibenzofurans (PCDFs) having a composition similar to that found in the rice oil causing"Yusho"was prepared in our laboratory. Male Sprague-Dawley rats were daily given orally for 22 d a regimen consisting of PCBs, 1 mg/rat/d ; PCQs, 1 mg/rat/d ; PCDFs, 10 μg/rat/d ; or a mixture of PCBs, PCQs and PCDFs (Mixture, (1 mg+1mg+10 μg)/rat/d). Rats treated with PCBs, PCDFs and Mixture showed hepatic enlargement, reduction of serum corticoid level, increase of serum cholesterol level and increase in the hepatic microsomal drug-metabolizing enzyme activity, and the effect was larger in the order of Mixture > PCDFs > PCBs. In addition, when PCDFs or Mixture were administered to rats, atrophy of the thymus and suppression of body weight gain were observed. When Mixture was administered to rats, a significant decrease in serum glutamic pyruvic transaminase (GPT) level and a significant increase in serum triglyceride level were observed. In the PCQs-treated rats, increase in the hepatic microsomal drug-metabolizing enzyme activity and decrease in the adrenal corticoid levels were observed, but the degree of change was far less than that in the PCBs- or PCDFs-treated rats. PCDFs or Mixture showed induction of hepatic microsomal drug-metabolizing enzyme activity of the 3-methylcholanthrene (MC) type in the rat. On the other hand, PCBs and PCQs showed intermediary-type induction of the phenobarbital (PB) and MC types. Therefore, the predominant etiology of"Yusho"may involve PCDFs contained in the toxic rice oil.

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Exposure to cadmium induced the nematode Caenorhabditis elegans to synthesize metallothioneinlike proteins, which were isolated and characterized. The supernatant obtained after disruption of cadmium-exposed nematodes was applied to a Sephadex G-75 column, and cadmium was detected in a high-molecular-weight fraction and a lowmolecular-weight fraction, of which the latter corresponded in elution volume to the well-characterized metallothionein. From this low molecular weight cadmium-rich fraction, two major cadmium-binding proteins were separated by DEAE Sephadex A-25 ion exchange chromatography. These two isoproteins showed similar ultraviolet absorption spectra to mammalian metallothioneins, indicating that they can be categorized metallothioneins. The slower-eluted isoform, MT-II, was as homogeneous on polyacrylamide disc gel electrophoresis while the faster-eluted isoform, MT-I, contained some impurity. The amino acid composition of the purified MT-II was determined. The content of cysteic residues was 22.3 % and the next most abundant residues were aspartate, alanine, glutamate and lysine. The content of acidic amino acids was high and that of aromatic amino acids was very small. The properties were similar to those of metallothioneins found in lower phyla of the animal kingdom rather than in mammalian species.

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The elution patterns of silver ion from a household water purifier equipped with a silver-coated charcoal filter were investigated under domestic-type conditions and chromosomal aberration tests on human leukocyte cultures and cultured mammalian fibroblast cells were carried out in order to clarify the genetic toxicity of silver ion. New household water purifiers were used for the following elution studies. Two short-term elution tests showed that the concentration of silver ion in the eluate from a household water purifier was about 30 μg/l immediately after opening the tap, and fell to about 15 μg/l when 8 l of water had continuously passed through the purifier. The levels of silver ion in a long-term elution test for three months ranged from 13.0 to 28.5 μg/l. No silver ion was detected in tap water which had not been passed through the household water purifier (less than 1 μg/l). No significant chromosomal aberrations were induced by silver ion in human leukocyte cultures. However, the cytogenetic assay by using mammalian fibroblast cells suggested that silver ion might possess weak clastogenicity.

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A method for quantitative analysis of p-hydroxybenzoate esters (parabens) in cosmetic lotions and creams was developed by high-performance liquid chromatography (HPLC). Parabens were separated from the cosmetic base and other ingredients by bi-layer (upper ; celite, lower ; silica gel) column chromatography with Hexane-AcOEt (80 : 20). The eluate was extracted with acetonitrile. The acetonitrile layer was concentrated in vacuo and determined by HPLC on Develosil ODS by using a mobile phase of methanol-water (65 : 35, v/v). Recoveries of parabens obtained from model cosmetics were more than 95%. This method with clean-up procedure on bi-layer column chromatography and liquid extraction was much suitable for the determination of parabens in cosmetic lotions and creams.

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A simple and sensitive method for the determination of total phosphorus and orthophosphate in water was established by using atomic absorption spectrophotometry with a graphite furnace atomizer, and the method was applied to underground water samples. The proposed method is based on the atomic absorption spectrophotometric determination of molybdenum in a chelate complex produced by the reaction of orthophosphate with the molybdate reagent used in a spectrophotometric method. When the recoveries and coefficient values were estimated by applying this method to potassium dihydrogenphosphate as a standard compound in various kinds of water, they were within the range of 96.7-109.0% and below 6.1%, respectively, which are satisfactory. Since the proposed method is highly sensitive (detection limit : 0.0005 mg/l), a time-consuming concentration procedure of water samples could be omitted. When the proposed method and the spectrophotometric method described in the Official Gazette were applied to various kinds of water, a highly significant correlation (r=0.999) was observed between the two. The assayed values of total phosphorus in samples of underground water obtained from 77 stations situated in the southern area of Hyogo prefecture in Japan were distributed between 0.001 and 2.318 mg/l. The concentrations of total phosphorus in the underground water were confirmed to be influenced by the depth of the well, and there was a significant relationship between the concentration of total phosphorus in underground water and the degree of sewerage diffusion.

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Eighty nine kinds of environmental pollutants were subjected to LC₅₀ test by using small static system at low (10°C), medium (20°C) and high temperatures (30°C). Permissible volume of various dissolvents was determined at 30°C and 10% ether-dimethylsulfoxide solution was chosen as a suitable dissolvent for water-insoluble pollutants. Effects of temperature on the acute toxicity of various pollutants for fish were observed and categorized into four groups. First group : The acute toxicity for fish is not influenced by alterations in temperature. Second group : The acute toxicity for fish is the strongest at medium temperature (20°C). Third group : The acute toxicity for fish increased with a decrease in temperature. Fourth group : The acute toxicity for fish increased with an increase in temperature.

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The quantitative colorimetric determination of TiO₂ in food was investigated by comparing Pt crucible ashing described previously with porcelain crucible ashing proposed here. The estimation value between these two ashing methods agreed in recovery tests on test food and TiO₂ value in market foods. Therfore it was recognized that popular porcelain crucible can be used instead of Pt crucible in routine analysis.

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