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A simple and rapid method using high-performance capillary electrophoresis (HPCE) for the simultaneous determination of the urinary metabolites of toluene, xylene and styrene, plus creatinine and uric acid in human urine specimens and standard solutions is described. The compounds were well separated from each other on a fused-silica capillary utilizing a 20 mM sodium tetraborate buffer (pH 9.65) with 15 mM β-cyclodextrin and UV detection at 200 and 225 nm. The total analysis time was less than 6 min per sample. The capillary zone electrophoresis (CZE) method shows a good correlation with the high-performance liquid chromatography (HPLC) method with respect to urinary hippuric acid concentrations in the urine specimens of subjects exposed to the vapors of a solvent mixture of toluene and xylene. In comparing these two techniques, HPCE was found to be superior to HPLC because the analysis time is shorter, and the separation of m-MHA and p-MHA takes a long time with HPLC.

事故時等の油汚染に対応する目的で, スクリーニング分析法の開発とナホトカ号から流出した重油中に含まれる化学成分の同定と漂着過程における成分の変化等について検討した。揮発性成分の分析では, ヘッドスペース法を用いた各種石油製品中のマスクロマトグラムパターンの解析と同定を行い, 重油中には揮発性のアルキルベンゼン類が多数存在することを見いだした。水質及び大気中の微量揮発性成分の分析法では, SPME法が効果的であることを見いだした。高沸点成分の分析では, 5%含水シリカゲルクロマトグラフィーを用いた油種分析のためのクリーンアップ法及びパラフィン類と多環芳香族炭化水素類の分別法を確立した。重油成分については, 多環芳香族炭化水素類の定量を行うとともに, 漂着の過程で, 重油は水溶性の高いアルキルベンゼン類及び低分子の多環芳香族炭化水素類を失うことを見いだした。

Oil mists at an automobile factory were measured size-selectively. Furthermore, subjective symptoms were surveyed through the distribution of a questionnaire. The measured levels of respirable-size, 2-10 micrometer oil mists at 3 selected workshops ranged 0.04-0.09 mg/m3 for the lower exposed group (17 men), 0.10-0.18 mg/m3 for the higher exposed group (16 men) to water-insoluble oil, and 0.08-0.29 mg/m3 for the higher exposed group (19 men) to water-soluble oil. The higher the level of exposure to respirable-size particulates, the more workers complained of dissatisfaction with air quality. As to the subjective symptoms, the "yes" rate for "irritated nose" and "sneezing" showed significant differences among the unexposed, lower exposed, and higher exposed groups. Workers using water-soluble oil complained of "uncomfortable smell" more than those who used water-insoluble oil. The results of this study suggested that it is necessary to measure the levels of oil mists size-selectively.

To evaluate the intensity of worker exposures to mixed solvents, the concentrations of hippuric acid (HA), methylhippuric acids (MHAs), and methyl isobutyl ketone in end-of-shift urine were examine...

In Japan, industrial health control is accomplished by control of the working environment, work practice management and health care. Periodical biomonitoring of workers exposed to lead and 8 popular organic solvents, including mixed solvents, became mandatory as of October 1, 1989, with an ordinance issued by the Ministry of Labor. This ordinance states that the results of measurements of lead and the 8 solvents in each work place must be classified into 3 categories, named distributions 1, 2 and 3 based on the biological level of the determinant, and then must be reported to the Labor Standards Inspection Office. Distribution 3 encompasses workers having concentration levels above the BEIs of ACGIH except for the concentration of lead in the blood. The National Federation of Industrial Health Organizations surveyed the state of biological monitoring in Japan by a questionnaire to the laboratories. The total number of cases examined in the fiscal year 1990 was about 110,000 for the biomonitoring of lead, and about 520,000 for the monitoring of urinary metabolites of the 8 organic solvents. The ratio of the number of workers in distribution 3 to that of the total workers examined was 0.4 % for urinary delta aminolevulinic acid and 0.2∼3.5 % for the urinary metabolites of each organic solvent. Similar results were obtained from 7 major laboratories. To achieve large-scale biological monitoring in future, a method is necessary for evaluating the exposure or health risks of workers exposed to mixed organic solvents by estimating their urinary metabolites.

The molecular defects in the catalase gene, levels of m-RNA and properties of the residual catalase studied by scientists are reviewed in human (Japanese, Swiss and Hungarian) and non-human (mouse and beagle dog) acatalasemia with reference to the bioinformatics. Japanese acatalasemia-I, the G to A transition at the fifth position of intron 4 of the catalase gene, limited the correct splicing of the mRNA and synthesized trace catalase with normal properties. Hungarian acatalasemia type C showed a splicing mutation. In the Japanese acatalasemia II and the type A and B of Hungarian acatalasemia, the deletion or insertion of nucleotides was observed in the coding regions, and the frame shift altered downstream amino acid sequences and formed truncated proteins. In the Hungarian acatalasemia D, the substitution of a nucleotide in the exon was found. In mouse and beagle dog acatalasemia, the substitution of nucleotides in the coding regions was also observed. Studies of residual catalase in Swiss, mouse and beagle dog acatalasemia showed that aberrant catalase protein degrades more quickly than normal catalase in cells. The experimental research in genetic toxicology concerning the effect of oxidative stressors (nitrogen monoxide, nitrogen dioxide and so on) on Japanese acatalasemic blood and acatalasemic mice is described. The clinical features of Japanese and Hungarian acatalasemic subjects are also described.

A simple method for the determination of 4-chlorocatechol (ClCh, 4-chloro-1,2-benzenediol) and chlorophenols (ClPh), metabolites of monochlorobenzene (ClBz), in urine by high performance liquid chromatography (HPLC) is described. Enzymatic hydrolysates of urine were applied to a stainless-steel column packed with octadecyl-silanized silica gel, and a mixed solution of 20 mM potassium phosphate monobasic: acetonitryl (75:25, v/v) was used as a mobile phase. The procedures for ether extraction and evaporation of extract could be omitted. The accuracy and precision of the present HPLC method were satisfactory. The excretion kinetics of ClCh and p-ClPh were investigated over 35 h after cessation of ClBz inhalation. Proportional relationships between concentrations of ClBz in air and of its metabolites in urine were observed. The slopes of regression lines predicting the levels of ClCh, p-ClPh and total ClPh in urine taken during the last 2 h of exposure to ClBz in air were 6.56, 1.13 and 2.83 mg/g creatinine for 1 ppm ClBz, respectively. ClBz in the blood and the end exhaled air of subjects at the end of exposure were identified by gas chromatography (GC) and mass spectrometry. A proportional relationship was observed between the concentration of ClBz in air and that in blood. The validity of the threshold limit value (TLV) for ClBz as evaluated from the subjective and objective symptoms is discussed.

Maximal expiratory flow-volume (MEFV) curves have been utilized in the diagnosis of chronic obstructive lung diseases, in the survey of air pollution, and in studies of respiratory functions in school children. Indices for the lower lung volumes of MEFV curves have been used in statistical analysis or individual evaluation because of their effort independence, while indices for higher lung volumes of MEFV curves, which were considered to be effort-dependent, have not been used. A few reports using indices for higher and lower lung volumes have been described because the indices for higher lung volumes are as reproducible to the maximal expiration as those for lower lung volumes. Recently, MEFV-type analysis has been studied and this analysis has been useful in assessing the effects of smoking on MEFV curves, or in assessing subjects with nasal allergies or those with prior nasal allergies. From the aspects of hygiene, this review mainly describes MEFV-type analysis and the quantitative analysis of discriminant analysis.

Heptachlor is a major component of the insecticide, chlordane. It is a health hazard but is still in use in some countries of Southeast Asia. To elucidate the toxicity of heptachlor its effects on mice after oral and intraperitoneal administration were studied. A 3-day group, 92-day group and 180-day group were given heptachlor intraperitoneally, orally and ad libitum, respectively. Results showed increased levels of serum alanine aminotransferase and decreased levels of serum cholinesterase activity. Serum creatine phosphokinase levels increased significantly. These may be due to the disruption of muscle membrane by chlordane. Results also showed significant variations of serum lipid levels from control as heptachlor has a known effect on lipid metabolism. Also the lipid peroxide levels expressed as TBA values were increased significantly, showing heptachlor's role in causing liver injury. These results suggest that the deterioration of membranes due to lipid peroxidation leads to liver and muscle injuries caused by heptachlor.

To search for the molecular defect of Japanese-type acatalasemia, we cloned the mutant catalase gene from a person with this deficiency. The nucleotide sequence of the mutant gene was determined for all exons, exon/intron junctions, and 5' and 3' flanking regions, and the findings were compared with the sequence from the normal gene. Seven base differences were found between the two genes. Among them, a G to A substitution at the fifth position of intron 4 (a splicing mutation) seemed most likely to be responsible for the defective catalase synthesis in the subject. To obtain suggestive evidence, we constructed chimeric genes that contained a segment of either the normal or mutant catalase gene, encompassing a 3' part of exon 4, the entire intron 4 and a 5' portion of exon 5, within the third exon of the human alpha-globin gene. When this chimeric gene construct was introduced into simian virus 40-transformed simian cells (COS-7), the transcript of the normal catalase/alpha-globin chimeric gene was spliced correctly, as revealed by Northern blotting and RNase mapping techniques. In contrast, the splicing of the mutant chimeric pre-mRNA occurred between the 5' donor site of the preceding intron and the 3' acceptor site of the intron containing the substitution, thereby skipping one entire exon sequence. Thus, the G to A transition at the fifth position of intron 4 of the catalase gene indeed severely limits the correct splicing of the RNA product. The same splice site mutation was found in the genomic DNA of another acatalasemic individual from an unrelated family. We suggest that this base substitution is the causal mutation of these cases of Japanese-type acatalasemia.

Machine oils are widely used in metal processing and many workers are exposed to oil mists in the work environment. Some investigators have pointed out such health hazards due to prolonged exposure to oil mist as respiratory disorders, dermatitis, and possible carcinogenesis. In Japan, a permissible limit of oil mists of 3 mg/m3 was recommended in 1977 by the Japan Association of Industrial Health (JAIH) mainly based on the hazardous effects on the respiratory system. After this recommendation was made, only a few studies have been made on the measurement of oil mists and on the health effects in machine workers to determine whether the permissible limit is justifiable or not. In the present study, the levels of oil mists in the air of machine workshops were measured together with personal exposure levels by personal samplers. Oil mists were collected by a sampler head with 2 stages which enabled differentiation of distribution of particulates between larger than 10 microns and 2-10 microns in size. Oil components collected on the stainless steel stages were washed out by sonication in CCl4 solution and measured by an oil meter with infra-red spectrometry against a standard solution of heavy oil class B according to procedures reported elsewhere by the authors. Questionnaire surveys were also conducted on 308 male machine workers composed of 221 workers exposed to oil mists and 87 nonexposed controls. The questions were composed of five items about air quality in the work environment and 18 subjective symptoms during work and daily lives. The symptoms included nasopharyngeal, muco-dermal, gastrointestinal and neuro-muscular symptoms. Statistical analysis was made by the Manthel-Haenszel method for a comparison of "yes" rates of complaints between the exposed and the non-exposed by adjusting the underlying confounding factor of age distribution. 1. The levels of oil mists measured here ranged from 94 to 813 micrograms/m3 in the ambient air and from 107 to 483 micrograms/m3 of personal exposure. There was no obvious difference between the level in the ambient air and that of personal exposure. All these measured levels were under the permissible limit (3 mg/m3) recommended by JAIH in 1977. 2. The observed distribution of particulate size of oil mists from 2 to 10 microns referred to as respirable size was 32.8 +/- 16.1% generated from the grinding machines using water-soluble type of machine oils and 50.0 +/- 12.4% from those using insoluble type.(ABSTRACT TRUNCATED AT 400 WORDS)

Personal air samplers were attached to workers wearing protective masks to determine the levels of toluene vapor in the breathing zone. Concentrations of toluene in exhaled air, blood and urine; and hippuric acid and o-cresol concentrations in the urine of the workers were determined. Subsequently, toluene concentrations in the air inhaled by workers with and without gas masks were estimated by single and multiple regression equations. Analysis of single regression equations revealed that, compared with toluene concentrations in air, masks decreased the concentrations of the four biological exposure indicators: toluene in exhaled air, urinary toluene, urinary hippuric acid and urinary o-cresol by about 29% in average. Analysis by multiple regression equations showed a decrease of 38% in four biological indicators. Since average exposure to toluene in the shop was relatively low, the workers wore the masks only during high concentrations of toluene; they were, however, exposed to direct inhalation when the masks were removed in lower concentrations.

Data from the large-scale biological monitoring program in Japan were assembled and analyzed and the following results were obtained. All workers handling lead and eight kinds of major organic solvents received physical examinations and biological monitoring at the same time. Therefore, the number of workers handling industrial chemicals and that received physical examinations and the number of workers been examined by biological monitorings were similar to each other. The total number of cases examined from 1989 to 1994 was about 661,000 for lead in the blood and about 4,173,000 for the urinary metabolites of eight organic solvents. The results were classified into three categories and category 3 consists of workers having exposure concentrations above the 1988-1989 biological exposure indices of the ACGIH with the exception of lead concentration in the blood where the limit in Japan was set at 40 micrograms/100 ml. The percentage of exposed workers in category 3 was 1.4% for blood lead and 0.2-2.4% for the urinary metabolites of the eight organic solvents. The percentage of exposed workers in category 3 for blood lead, delta-aminolevulinic acid, urinary mandelic acid, N-methylformamide and 2,5-hexanedione in the urine has decreased with time. In ambient monitoring, the percentage of workplaces in classification 3 for lead and styrene also has decreased with time.

An attempt was made to compare the toxic effects of the organochlorine insecticide ‘chlordane’ in man and rats. Analysis of blood for chlordane metabolites showed their presence in the descending order of trans-nonachlor, oxychlordane, hepatachlorepoxide and cis-nonachlor. The total range of chlordane and its metabolites in the sera of workers was 9.84 ± 4.47 ng/g. Serum levels of triglycerides (TG), creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) activities were also found to be higher in pestcontrol operators. In a simultaneous study, rats were administered 100 mg/kg body wt. of chlordane by stomach tube once a day for 4 days, whereas 50 mg/kg body wt. of chlordane was injected intraperitoneally once a day for 4 days. The data show that total cholesterol and serum TG as well as CPK and LDH activities are increased after chlordane treatment. The isoenzyme patterns suggest that an increase in CPK and LDH is related to skeletal muscle. Furthermore, the hepatotoxicity of chlordane was also studied in rats only. A significant increase in liver weight, its water content, total lipids, triglycerides and phospholipids was recorded. Chlordane induced lipid peroxidation in the liver, exhibiting a dose-response relationship. Although no appreciable effect on mitochondrial function and latent ATPase activity was observed, 2,4-dinitrophenol-stimulated ATPase activity was inhibited. Histological examination of the liver confirmed fatty infiltration induced by chlordane in rats.

High-performance liquid chromatography (HPLC), gas chromatography (GC) and spectrophotometry were used to examine 2,5-dimethylpyrrole, derived from 2,5-hexanedione in the acid-hydrolyzed urine of subjects exposed to n-hexane. The urine of a subject exposed to n-hexane was hydrolyzed with hydrochloric acid and then neutralized. Thereafter, 2,5-hexanedione in the hydrolysate was heated with ammonium carbonate to yield 2,5-dimethylpyrrole. A methanol extract of the 2,5-dimethylpyrrole formed was subjected to HPLC and an ethylacetate extract of it was subjected to GC, with a flame thermoionic detector (FTD). For spectrophotometry, the ethylacetate extract was allowed to react with Ehrlich's reagent and then the absorbance of the colored compound formed was read at 525 nm. Pyrrole condensation with ammonia is specific to 1,4-diketones and these three assay methods can be used for determination of 2,5-hexanedione in the urine of workers exposed to n-hexane. The hydrolytic conditions for urine of subjects exposed to n-hexane for analysis of 2,5-hexanedione were discussed.

Analyse des hydrocarbures aromatiques polycycliques de benzothiophenes et de dibenzothiophenes

In a study of the hepatotoxicity of heptachlor (1,4,5,6,7,8,8-heptachlor 3a, 4,7,7 a-tetrahydro-4,7-methanoindene), a major compound of chlordane, the effect of heptachlor on the respiratory activity (oxidative phosphorylation and electron transport) of rat liver mitochondria was investigated. Heptachlor at a final concentration of 50 microM with succinate as substrate decreased the respiratory control index (RCI) due to a marked inhibition of state 3 respiration and a slight inhibition of state 4 respiration. One hundred microM heptachlor with succinate as substrate suppressed the states 3 and 4 respiration almost completely. On the other hand, heptachlor at a final concentration ranging from 50 to 100 microM with beta-hydroxybutylate (beta-HB) slightly decreased the RCI and decreased the RCI hardly at all with ascorbate plus N,N,N',N', -tetramethylphenylene diamine (TMPD) as substrate. Heptachlor at a concentration of 50 microM in the presence of succinate also decreased the ADP/O ratio of mitochondria. The mode of inhibition of succinate oxidation by heptachlor apparently is a noncompetitive inhibition, as shown by Lineweaver-Burk plot.

An enzymatic assay method for the determination of urinary formic acid is described. Formic acid in urine was cleaved to carbon dioxide and water by formic acid dehydrogenase, whereby NAD+ was converted to NADH, which reacted with INT (p-iodonitrotetrazolium violet) in the presence of NAD-diaphorase. The color thus produced was determined at 500 nm. In addition, a simple gas chromatographic method of urinary formic acid is described, in which head space gas of formic acid methylester was applied into the wide bore column. The urinary formic acid concentrations by the enzymatic method agreed well with that by the gas chromatographic method. A simple gas chromatographic method for urinary methanol assay is also described. Acetonitrile was added to an equal volume of urine containing methanol. After centrifugation, the supernatant was injected into gas chromatography (GC). The peaks of urinary methanol and ethanol were separated by GC. Formic acid and methanol in urine of unexposed healthy subjects and workers exposed to methanol were analyzed by the colorimetric and gas chromatographic methods. Geometric mean concentrations of urinary formic acid and methanol in the healthy subjects were 7.82 mg/g creatinine and 1.34 mg/l, respectively. The concentration ratio of formic acid to methanol in the urine of the workers exposed to methanol was calculated to be 3.67 +/- 2.10, which agreed with the ratio under a controlled exposure experiment. A slower excretion of formic acid than that of methanol in the urine of a volunteer was also observed.

Six female students were orally given 175 and 350ml of a soft drink containing 190mg/L of sodium benzoate (BA-Na) and three male volunteers were also orally given BA-Na solution at a dosage of 20mg/kg body weight. Urinary hippuric acid (HA) and creatinine excreted by the subjects given either the soft drink or the BA-Na solution were determined simultaneously by high performance liquid chromatography (HPLC) from the end of the administration until 4 or 5hr later, at appropriate intervals.In the case of soft drink administration, the concentration of HA attained a maximum in urine taken from 0 to 30min and recovered to the predose level after 3hr. The molar ratios of HA excreted in the 0-3hr urine samples to the administered dose were estimated to be 66-86%, by subtracting the amount of HA excreted in the predose period on the day of the administration. In the case of BA-Na solution administration, the concentration of HA attained a maximum in the 0-1hr urine samples and did not recover to the predose level within 5hr. The molar ratio of HA excreted in the 0-5hr urine samples to the administered dose was estimated to be about 89%, by subtracting the amount of HA excreted in the predose period on the day of the administration. Based upon these findings, it is suggested that the amount of absorbed BA-Na in human subjects can be estimated by determining the amount of HA excreted in the urine.

A high performance liquid chromatographic method for the determination of phenylsulfate and phenylglucuronide, metabolites of benzene, or phenol in urine is described. For the determination of urinary phenylsulfate, a stainless-steel column packed with octadecyl-silanized silica gel was used, and the mobile phase was mixed with a solution of methanol/water acetic acid (20/80/0.2) containing 0.05 M tetra-n-butylammonium bromide. For the exact determination of urinary phenylglucuronide, a stainless-steel column packed with the same resin was used and the mobile phase was a solution of methanol/water/acetic acid (50/50/0.2) containing 0.05 M tetra-n-butylammonium bromide. The method is simple and specific. Urine can be analyzed directly, without solvent extraction or pretreatment. The method has a lowest detection limit of 0.2 μg on the column. The whole analysis and quantitative determination can be performed within about 15 min for samples containing phenylsulfate and phenylglucuronide in the urine of rats.

Urinary metabolites of styrene, i.e., mandelic acid, phenylglyoxylic acid and hippuric acid, were determined by high performance liquid chromato-graphy. Female Wistar rats were administered various amounts of styrene intraperitoneally, and the metabolites in the urine were determined at suitable intervals after the injection. The amounts of urinary mandelic and phenyl-glyoxylic acids were in proportion to the amount of administered styrene. The amount of urinary metabolites per body weight per hour reached a maximum level in the urine from the 10-24 hr period. In another experiment, tetrachlo-romethane was injected intramuscularly into rats prior to the administration of styrene. The amounts of urinary mandelic, phenylglyoxylic and hippuric acids in the tetrachloromethane-administered group were much less than those of the control group.

The authors determined that the organic sulfur compounds usually contained in crude oil can be used as a marker of oil pollution in shellfish and fish. Short-necked clams (Tapes Angdala phillipinarum A. et Reeve) and eels were maintained in a controlled laboratory environment in water with suspension of crude oil. Mass spectra and mass chromatograms of short-necked clam extract showed the presence of organic sulfur compounds of alkyl- (from C1 to C10) benzothiophenes, dibenzothiophene, alkyl- (C1 to C8) dibenzothiophene, naphthalene, and alkyl- (C1 to C6) naphthalene. Alkyl benzothiophenes and alkyl dibenzothiophenes of low carbon numbers were more easily transferred to short-necked clams than those of higher carbon numbers. Capillary column gas chromatography-mass chromatograms of crude oil and extract from the soft body of a short-necked clam showed the presence of organic sulfur compounds of alkyl benzothiophene (from C2-C6), dibenzothiophene, and alkyl dibenzothiophene (C1-C4). Besides sulfur components, various other compounds were contained in crude oil and short-necked clam, namely alkyl benzene (C3-C9), naphthalene, alkyl naphthalene (C1-C5), phenanthrene, alkyl phenanthrene (C1-C6), n-paraffin (C12-C21), and probably pyrene and alkyl pyrene (C1-C2). Also, mass chromatograms of crude oil and the extract from eel flesh showed the presence of alkyl benzothiophene (C1-C5), dibenzothiophene, naphthalene, and alkyl naphthalene (C1-C5). Data indicated that the organic sulfur compounds and polyaromatic compounds could serve as markers of oil pollution in shellfish and fish. for the identification of sulfur compounds by capillary gas chromatography/mass spectrometry, a new intensity matching method was devised.

The effects of the chlordane-related compounds, cis -chlordane, trans -chlordane, heptachlor, and heptachlor epoxide, on stimulation responses of guinea pig polymorphonuclear leukocytes (PMNs) were examined. Treatment of PMN with these compounds stimulated superoxide (O 2 − ) generation, altered membrane potential, and increased intracellular Ca 2+ concentration ([Ca 2+ ] i ). However, there was no definite tendency among the stimulating effects of chlordane-related compounds, therefore the relationship between the effect and molecular structure of these substances remains unknown. Of these response reactions of PMN stimulated by chlordane-related compounds, stimulation of O 2 − generation lagged behind others. Increase in [Ca 2+ ] i was due both to acceleration of extracellular Ca 2+ release from the intracellular pool. These results indicate that chlordane-related compounds stimulate PMN and suggest a causal relationship between the stimulation of O 2 − generation by these substances and their toxicity.

This paper describes the sampling of air and its analysis for mixtures of organic solvent vapours by personal and room air samplers. The application of a potassium carbonate to remove water vapour, and silica gel adsorption tube cooled by dry ice for collection of organic solvents were investigated. The development of a method for the extraction of organic solvents from silica gel by cumene, ethyl ether, pyridine and dimethyl sulfoxide and subsequent estimation of their amounts by gas chromatography or colorimetry is reported. This method was useful for the personal air sampler in the workshop using thinner. Ethyl cellosolve acetate, and N,N-dimethyl formamide being cooled by dry ice and mixture of dimethyl sulfoxide and N,N-dimethyl formamide being cooled by ice were useful as absorbing agents of organic solvents particularly in the room air sampler.

A protein of low molecular weight (MW 12, 000) was detected in the urine of a rabbit given cadmium for 17 months, by polyacrylamide gel electro-phoresis in the presence of sodium dodecyl sulfate. This protein was identified as β2-microglobulin by electrosyneresis using standard β2-microglobulin, a urine sample and anti rabbit β2-microglobulin mouse serum. The urinary concentration of β2-microglobulin in the rabbit fed a diet containing cadmium for 17 months was estimated to be 8-40 mg/l by staining the gel with silver and to be 10-20 mg/l (3.6-7.2 mg/day) by electrosyneresis.

An attempt was made to determine the organic sulfur compounds, usually contained in crude oil, as a marker of oil pollution in shellfish. Short-necked clam ( Tapea Angdala phillipinarum A. et Reeve) were maintained in a controlled laboratory environment and in water with a suspension of crude oil. Capillary column gas chromatography-mass chromatography of short-necked clam extract showed the presence of organic sulfur compounds of alkyl-benzothiophene (from C 1 –C 6 ) and dibenzothiophene and alkyl-dibenzothiophene (C 1 –C 4 ). These organic sulfur compounds are a marker of oil pollution of shellfish. Other organic aromatic compounds, naphthalene, alkyl-naphthalene (C 1 –C 3 ), phenanthrene, alkyl-phenanthrene (C 1 –C 3 ), pyrene and alkyl-pyrene (C 1 ) in soft body of short-necked clam were transferred from oil suspension to shellfish. The concentration factors of organic sulfur compounds according with lapse of time were determined by gas chromatography with an attached flame ionization detector.

In order to study the factors which have influenced to β2-microglobulin levels in urine, the effect of hard loading on the concentration of urinary β2-microglobulin was studied.Increase in both actual and corrected β2-microglobulin concentration and slight increase in albumin was observed with volunteers in the urine after marathon running.Maximum excretion of urinary β2-microglobulin was observed 30min after cession of exercise.And increase in actual and corrected β2-microglobulin concentration, and the ratio of β2-microglobulin to creatinine in urine of volunteers loaded 210 steps/min for 1 hour were recognized 30 min after cession of exercise by training machine. Similar result of urinary excretion to β2-microglobulin was obtained with those of creatinine.Concentration of β2-microglobulin in serum had a tendency to increase, but concentration of creatinine in serum was not increased.Results of Sephadex G-100 gel filtrations indicated that urinary β2-microglobulin after exercise, which measured by radioimmunoassay, was contained in the filtrated fraction having molecular weight of approximately 11, 800, suggesting that similar substance to HL-A did not excreted in urine after hard loading.

To investigate the toxicity of chlordane, an organochlorine insecticide, effects of cis-Chlordane, trans-Chlordane, Heptachlor and Heptachlor epoxide were examined on stimulus responses of guinea pig polymorphonuclear leukocytes (PMNs). Results obtained were as follows. These chlordane-related compounds stimulated superoxide (O2-) generation, altered membrane potential and increased intracellular Ca2+ concentration ((Ca2+]i). As a significant tendency was not found in the stimulating effects of these compounds, the relationship between the effect and molecular structure of these substances remains unknown. Of these response reactions of PMN stimulated by chlordanes, stimulation of O2- generation lagged behind the others. Increase in [Ca2+]i was due to both acceleration of extracellular Ca2+ influx and Ca2+ release from intracellular pool. These results indicate that these chlordane-related compounds stimulate PMN and suggest a causal relationship between the stimulation of O2- generation by these substances and their toxicity.

Human acatalasemic erythrocytes had only 0.01 to 0.06X the uptake found in normal erythrocytes with hydrogen peroxide, and 0.06 to 0.24X the uptake without hydrogen peroxide in regard to their ability of in vitro mercury uptake from air saturated with mercury vapor. Normal erythrocytes with hydrogen peroxide took up 9.3 to 37.4X more mercury than cells without hydrogen peroxide, whereas acatalesemic cells with hydrogen peroxide took up 2.4 to 5.8X more mercury than cells without hydrogen peroxide. Similar results were obtained from normal and acatalasemic hemolysates. In addition, the concentration of hydrogen peroxide affected the uptake of mercury by both normal and acatalasemic erythrocytes. Data indicate that the catalase-hydrogen peroxide system plays a role in mercury uptake, presumably by mercury oxidation, since oxidation states of the mercury were never determined in these studies.

Antiserum specific for rabbit β2-microglobulin was obtained from mouse. The antiserum was used for measuring concentration of this protein in rabbit urine by electrosyneresis. Its lower limit of the detection was 50 mg/l. Excretion of urinary β2-microglobulin in the rabbit administered sodium chromate increased temporarily and reached to the maximum (above 800 mg/l) 2 days after the administration.

The organic sulfur compounds usually contained in crude oil were used as a marker of oil pollution in shellfish. The oyster (Crassostrea gigas) and mussel (Mytilus edulis) were caught in the Seto Inland Sea of Japan. Capillary-column gas chromatography/mass spectrometry (GC/MS) of extracts showed the presence of organic sulfur compounds of dibenzothiophene and alkyl dibenzothiophenes (C1-C3). The concentration factor of organic sulfur compounds in the oyster and mussel was determined by gas chromatography using a flame photometric detector.