The target organ of inorganic tin and its toxic action are described in this review. Tin has some toxic effects on gastrointestinal function, blood cells, hepatic enzymes, endocrine secretion from pancreas, renal calcium metabolism and its related enzymes, bone metabolism, central nervous system, and reproduction function. In bone, however, a marked deposition is caused by the ingestion of inorganic tin compounds, while a slight deposition is observed in other soft tissues. The acute toxic studies of inorganic tin on various biochemical indices suggest that the critical organ of tin is bone with a decrease in the femoral calcium content. Furthermore, from the subacute toxic experiment by oral administration of stannous chloride for 90 days in weanling rats, the decrease of calcium content in the femoral epiphysis is regarded as the most specific biochemical manifestation of the toxic action of inorganic tin. On the basis of the sign of this manifestation, the nonobserved-effect level of inorganic tin is estimated to be lower than 0.6 mg Sn/kg/d. Also, the maximal allowable concentration of inorganic tin contained in the diet would be lower than 50 ppm Sn. On the other hand, as a possible mechanism of the toxic action of inorganic tin on bone metabolism, it seems that tin induces the obstruction of proliferation of the bone cells accompanied with the prevention of DNA synthesis succeeded to the inhibition of protein synthesis, i.e. alkaline phosphatase and collagen, and as the result the bone formation is suppressed. This review emphasizes that a primary target organ of inorganic tin is bone, and the metal may cause fragile bone.

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In recent years, the introduction of improved anaerobic culture technique and media, and the utilization of gnotobiotic animal technique have greatly promoted the studies of microecological research of intestine. Intestinal flora participates in normal physiological processes and also contributes to complex pathological states. The flora is capable of metabolizing a variety of substances such as foods, drugs or endogenous compounds, thus influencing drug efficacy and toxicity, carcinogenesis, and aging. However, little is known about which bacterial species implicate in these phenomena, and about the mechanism of these microbe-host interactions. A solution of these problems will only be achieved by the cooperation of workers of different research areas such as microbiology, physiology, biochemistry, pathology, pharmacology, toxicology and clinical medicine.

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In order to elucidate the mechanisms involved in the formation of the Hg-Se complex in plasma in vivo, the interaction of mercury and selenium in rat plasma in vitro was studied by a gel filtration method. When mercuric chloride was added to rat blood within 20 min after the addition of selenite, two distinct overlapping peaks of mercury and selenium were found in the gel chromatographic fractionation of plasma. One of the overlapping peaks was found in the void volume fraction, while the other overlapping peak of both elements appeared in the elution region of the Hg-Se complex formed in plasma in vivo. When mercury was added 40 min after the addition of selenite, only the former overlapping peak appeared. When the plasma obtained from an incubation mixture of blood and selenite (incubation time : 5 or 40 min) was treated with mercury, the selenium level in the void volume fraction increased, whereas that in the albumin fraction decreased. A mercury peak appeared only in the void volume fraction. These results suggested that the latter overlapping peak of both elements was produced by the interaction of mercury with labile metabolites of selenite, other than albumin-bound selenium, existing in the blood within 20 min after the addition of selenite.

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Fenitrothion (FT) at low concentration of around 10^-6M reacted with mouse liver homogenate and disappeared immediately similarly to fenitrooxon (FO). This occurred mostly in the soluble fraction of mouse liver and was dependent on glutathione (GSH). The main metabolite detected was desmethyl-FT in the case of the reaction with a small amount of mouse liver homogenate, while desmethyl-FO was predominant metabolite in the reaction with relatively large amount of homogenate. In both cases all desmethylated metabolites were obtained more than 50% of added FT. These facts reveal that at such low concentration, FT is detoxified mainly through desmethylation reaction catalyzed by GSH S-transferase (GTase). On the other hand the amount of 4-nitro-m-cresol was less than 10%, so the oxidative dearylation by mixed function oxidase (MFO) contributed little to the detoxication of FT at low concentration. The Km and Vmax values of both GTase and MFO are well consistent with this reaction mechanism.

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Formation of dimethyl disulfide (DMDS) from methionine added in the mixed liquor of activated sludge was investigated. Though the formation of DMDS was influenced by culture time, the content of methionine added, concentration of mixed liquor suspended solids, pH and temperature of the liquor, the condition of activated sludge influenced more significantly the formation of DMDS. As the condition of activated sludge became worse, the formation of DMDS increased, which was found on addition of not only L-methionine but also D-methionine.

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Recently, the use of human head hair has become of interest as one of the biological indicators of enviromental pollution. Though hair is easily collected and multi-element analysis of hair is easily carried out by applying instrumental neutron activation analysis, it is not always clear how the trace element contents in hair represent those in organs, and there are a number of unclarified points regarding the correlation between them. As one of the first steps in approaching the problems, the authors made a survey on the mercury movement in body hair and in tissues of mice and obtained the correlation coefficients between them, by using ²⁰³Hg tracer in HgCl₂. The maximum concentration of mercury was obtained in kidney and the minimum value in blood. The mercury concentration in body hair tended to be higher than that in every tissue except kidney. The correlation coefficient between the concentrations in body hair and tissues were 0.864 in kidney, 0.869 in liver, 0.843 in lung, 0.933 in brain, 0.883 in spleen, 0.892 in intestine, 0.890 in thighbone and 0.839 in blood, and a significant correlation was observed between the mercury concentration in hair and in every tissue. By applying the compartmental analysis to the data obtained, the difference of turnover characteristics was examined between body hair and every tissue. Further, the regression equation was introduced to calculate the mercury concentration in every tissue from that in body hair.

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The uptake of cadmium (Cd) and the production of cadmium binding protein (Cd-BP) in Chlorella ellipsoidea were examined. To the medium in which chlorella was growing to the stationary phase CdCl₂ (2 mg Cd/l) was added and the cultivation in the presence of CdCl₂ was run for 0-120 h under light intensity of 4 kLx at 25°C with bubbling 5% CO₂ gas. An aliquot of chlorella was harvested every 24 h from the culture medium and its Cd concentration was estimated by atomic absorption spectrophotometry. Cadmium taken up by the cells increased with increasing the cultivation period and the concentration of CdCl₂ added to the medium. Cadmium in the cells was detected in the cytosol fraction and estimated to bind to the protein. The Cd-BP produced in the cytosol was eluted at the position of Ve/Vo 2.3 on a Sephadex G-75 column. The production of Cd-BP was found to be dependent on the concentration of Cd in chlorella cells. Moreover, the production of Cd-BP was not stimulated by ZnSO₄ (5.36×10^-5 mol in the medium). Cadmium bound to the protein was partly substituted by Cu when 1.07×10^-5 mol CuSO₄ was added to the medium.

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Two metabolites formed by rat caecal contents or human E. coli (W3110 strain) incubated with 4, 6-dinitro-2-sec-butylphenol (DNBP), one of the agricultural chemicals of dinitrophenol homologous series, were detected by thin-layer chromatography. These metabolites, 6-amino-4-nitro-2-sec-butyl-phenol (6A4NBP) and 6-acetylamino-4-nitro-2-sec-butylphenol (6AA4NBP), were identified by comparing their infrared and mass spectral data with those of authentic samples. The amounts of 6A4NBP and 6AA4NBP which were formed after 48 h incubation were determined by densitometric analysis to be about 63 and 7% for rat caecal contents ; 1.7 and 0.9% for human E. coli, respectively. The results of this investigation indicated that a primary metabolic reaction of DNBP in the rat caecal microflora was the reduction of a nitro group to an amino group, and that N-acetylation of the amino group occurred as the secondary reaction. In addition, densitometric analysis adopted in the present study was confirmed to be very effective for the determination of DNBP and its metabolites.

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At 18 h after the single injection of lead acetate (100 mg/kg) in male Wistar rats, zinc content in the low molecular-weight protein fraction (zinc binding protein, Zn-BP) in the hepatic 105000g soluble fraction markedly increased, whereas lead was detected only in the high molecular-weight protein fraction by using Sephadex G-75 gel filtration. The properties of this Zn-BP were investigated and the following results were obtained. 1) Molecular weight was estimated about 10000 daltons. 2) Zinc in Zn-BP was substituted with cadmium. 3) UV spectrum of Zn-BP showed similar spectrum to metallothionein. 4) The formation of Zn-BP induced by lead was almost inhibited by the pretreatment with actinomycin D. 5) Zn-BP was heat-stable, when treated at 80°C for 1 min. These findings suggest that Zn-BP is thought to be zinc metallothionein (Zn-MT) and consequently lead has an inductive potency to Zn-MT in the liver of rats.

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A simple and sensitive method for the determination of anionic surfactant in water was established by using atomic absorption spectrophotometry with graphite furnace atomizer. The proposed method was based on the atomic absorption spectrometric determination of cobalt in a chelate complex produced by the reaction of anionic surfactant with bis [2-(5-chloro-2-pyridylazo)-5-diethylaminophenolato] cobalt (III) chloride (Co-PADAP) proposed by Taguchi et al. A water sample (surfactant content <30μg) was taken in a beaker, adjusted to pH 1 with sulfuric acid and transferred into a separating funnel. One ml of 7.9×10^-4M Co-PADAP solution was added to the sample solution and mixed for a few seconds. After allowed to stand for 1 min, 5.0 ml of benzene was added to the solution and shaked for 1 min. The benzene layer was subjected to atomic absorption spectrophotometry. The recoveries of sodium di-(2-ethylhexyl) sulphosuccinate as a standard compound added to various kinds of water were in the range of 92.0-110.0% and the coefficient variations were within 4.7%. The interference by ClO^-could be eliminated by the addition of Na₂SO₃. There were high significant correlations between the proposed method and the methylene blue method or the Taguchi's colorimetric method. The assayed values of anionic surfactants in the water samples obtained from the rivers in Kobe city were distributed 0.026 mg/l and 2.562 mg/l.

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Bonding between iron ions and soybean protein (produced by the hydrolysis of soybean meal) in an iron-protein compound, which was prepared by mixing the soybean protein and iron (II) sulfate solution, was studied by infrared spectroscopy and an X-ray powder diffraction method. The oxidation state of the predominant iron ions within the compound was +3, but some Fe (II) ions were also present. In the infrared spectrum of the iron-protein compound, the band ascribable to stretching of the COO^-group was shifted to the higher frequency side by 40 cm^-1 in comparison with that of the free protein. No evidence was found for the formation of any crystalline iron compounds such as Fe₂O₃, Fe₃O₄ and FeO in the X-ray powder patterns. The possibility of the formation of gel-like compounds of iron (II) and iron (III) such as Fe(OH)₂ and Fe(OH)₃, respectively, was excluded, because the hydrolyzed product of iron (II) is easily oxidized to iron (III) and Fe(OH)₃ transforms, at least partly to Fe₂O₃ or Fe₃O₄ after aging or storing the product in air. Thus, we concluded that the iron ion is bound to the soybean protein molecule through one or more carboxyl groups. Although other coordination sites within the protein molecule could also combine with the iron ions, we could not find evidence for other bondings.

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Caffeine and sodium benzoate were determined by high performance liquid chromatography with a UV detector by use of stainless column (4.0 mm×0.15 m) packed with Nucleosil ₅C₁₈. Mobile phase was M/30 KH₂PO₄+M/30 Na₂HPO₄·2H₂O-methanol (70 : 30). The sample was dissolved in 20% aqueous methanol. Determination was carried out successfully by using antipyrine as an internal standard and by maintaining the temperature at 40°C. The recovery of sodium benzoate and caffeine from synthetic mixtures were 97.2±1.0% and 99.8±1.1%. This method was applied to three kinds of pharmaceutical preparations. The contents of sodium benzoate and caffeine were 48.4-50.1% and 49.0-49.6%.

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Some properties of Cd-binding protein (Cd-BP) in chlorella cells (Chlorella ellipsoidea) were examined. The Cd-BP was isolated and purified from the 105000×g supernatant of chlorella cells grown in a medium containing CdCl₂ (1 mg Cd/l) by gel filtration on Sephadex G-75 and Sephadex G-25 and by ion exchange chromatography on DEAE-Sephadex A-25. In the DEAE-Sephadex A-25 chromatography a high concentration (0.6 M) of NaCl was required for eluting the Cd-BP. The molecular weight of the Cd-BP was estimated as approximately 8500 by gel filtration on Sephadex G-75. The 50% dissociation of Cd from the Cd-BP was given at pH 3.6. Absorption intensity of Cd-BP at 250 nm decreased with Cd dissociation from Cd-BP by lowering pH. These results suggest that the property of Cd-BP in chlorella cells is similar to that of mammalian metallothioneins except that the Cd-BP showed highly anionic behavior on the ion exchange chromatography.

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