A rapid and simplified detection method for enterotoxin-producing Staphylococus aureus in food was devised. The procedure is as follows: a homogenized food sample is added to Tellulite-Lithium-Starch-Pyruvate Brain Heart Infusion broth in a Petri dish, and incubated for 18 hrs at 37°C. Enterotoxins produced in the broth are detected by using a “SET-RPLA” kit (Reversed Passive Latex Agglutination method).
Numerous bacteria coexisting with S. aureus in raw milk, minced meat and potato salad did not interfere with the growth and enterotoxin production of S. aureus. Egg-yolk reaction-negative S. aureus was also detected. An excellent correlation in bacterial count of S. aureus was found between MPN value obtained by the above procedure and the usual plate count. When this method was applied to various commercial foods, S. aureus was detected more effectively than by the conventional plate method.
This method may be useful for routine laboratory tests for the detection of S. aureus in foods.

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The fates of cadmium, copper, and zinc in rats were examined after administrations of cadmium chloride (80mg/kg/day) alone and cadmium chloride (80mg/kg/day) with L-cysteine (1, 500mg/kg/day) for 20 consecutive weeks.
At this high dose of cadmium, cadmium levels in the kidney reached a plateau by the 6th week of administration in both groups, while cadmium levels in the liver reached a plateau of about 400-500μg/g at about 16 weeks in the cadmium chloride plus L-cysteine administration group and about 20 weeks in the cadmium chloride administration group. Urinary excretion of cadmium increased markedly in proportion to cadmium accumulation in the liver in both groups.
The fates of copper and zinc in rats during repeated oral administration of cadmium were examined by evaluating (1) intestinal absorption, (2) interactions with metallothionein and (3) urinary excretion. Copper levels in the organs (liver and kidney) and blood were found to decrease below the control values, while simultaneous administration of L-cysteine with cadmium suppressed the decrease in copper levels. The decrease of copper levels was considered to be attributable partly to cadmium-induced promotion of urinary copper excretion and mainly to cadmium-induced inhibition of intestinal copper absorption. Zinc levels in organs rapidly increased in the early period of cadmium administration and showed a gradual decrease at and after the middle period of the administration. Zinc levels in organs did not decrease below the control values.

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The fates of essential elements, including iron, manganese, calcium and magnesium, were examined after oral administration of cadmium chloride (80mg/kg/day) alone and cadmium chloride (80mg/kg/day) with L-cysteine (1, 500mg/kg/day) in rats for 20 consecutive weeks.
Iron levels in the liver and kidney were greatly decreased below the control values in the 4th week of repeated oral administration of cadmium, but the decrease was suppressed by the simultaneous administration of L-cysteine with cadmium. Urinary excretion of iron remained almost unchanged during the repeated oral administration of cadmium. Iron levels in organs did not decrease much after subcutaneous administration of cadmium. Therefore, the decrease of iron levels was considered to have been induced by the inhibition of intestinal iron absorption by cadmium. Moreover, development of iron deficiency anemia was observed as the iron levels in organs and blood decreased.
Calcium levels in organs did not change greatly, indicating that the inhibition of intestinal calcium absorption by cadmium was far less than in the case of iron absorption. The levels of manganese and magnesium in organs and urine remained close to the control values suggesting that oral administration of cadmium had no effect on them.

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The mutagenicities of 122 kinds of processed foods were tested in the Salmonella/microsome system of Ames. The mutagens were purified by using blue cotton, which is adsorbent cotton bearing covalently bound trisulfo-copper-phthalocyanine. Coffee, red wine, smoked dried bonito (katsuobushi), canned broiled fishes and flavor seasoning (dashinomoto) were mutagenic in Salmonella typhimurium TA98 with S9 mix. The characteristics of the mutagenicity of coffee differed from those reported earlier. The mutagenicity of katsuobushi may be due to compounds other than benzo [a] pyrene. The mutagens found in these processed foods may be polycyclic aromatic mutagens, because blue cotton has specific affinity for them.

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Basic investigations for the determination of Diarrhetic Shellfish Poison (DSP) were carried out with Tetrahymena pyriformis in place of the mouse lethal assay, and the following findings were obtained.
1. The growth of T. pyriformis was inhibited in proportion to the toxicity.
2. The growth inhibition of T. pyriformis increased with the progress of purification of the DSP fractions.
3. Heavy metals, pesticides, and PCBs in the hepatopancreas of scallops were effectively removed during DSP extraction. Therefore, DSP is responsible for the growth inhibition of T. pyriformis.
4. The 50% growth-inhibition value of T. pyriformis by DSP fractions was found to be equivalent to 0.5 Mouse Unit (0.05MU/ml medium) with satisfactory reproducibility.

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A new ECD gas chromatographic determination method for kanamycin (KM) is proposed.
KM in beef was extracted with 10% trichloroacetic acid, and the extract was washed with ether twice before being absorbed on an Amberlite CG-50 (H form) resin column and eluted with 0.1N hydrochloric acid. The eluate was concentrated to dryness and dissolved in methyl alcohol-water (1:1) mixture to prepare a test solution.
A portion of the test solution was dried, and N-acylation of KM in the residue was carried out with heptafluorobutylyl-imidazole, followed by trimethylsilylation with trimethylsilylimidazole. The resulting derivative was determined by ECD-gas chromatography on a glass column (1.5m×3mm id) packed with 4% OV-101 coated onto Gaschrom Q.
A linear calibration curve was obtained in the range from 0.05 to 1.0μg of KM. Recoveries of KM added to beef at levels of 0.2, 0.5 and 1.0ppm were 74%, 86% and 83%, respectively. The limit of detection was 0.1μg/g.

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The teratogenicity of sodium D-erythro-3-oxohexonate lactone (sodium erythorbate), which is used as a food additive, was studied in Wistar rats. The pregnant rats were fed a diet containing 5, 0.5 or 0.05% sodium erythorbate from day 7 through day 14 of pregnancy. Neither any adverse effect on body weight gain nor any clinical sign of toxicity was observed in the dams of any group during pregnancy. No significant difference between the sodium erythorbate-treated groups and the control group was found in the incidence of intrauterine fetal death, the number of live fetuses per dam, the sex ratio of fetuses, the fetal body weight and the placental weight. External, skeletal and internal examinations of the fetuses revealed no evidence of teratogenesis in any group. In the postnatal development of the offspring from the dams given sodium erythorbate, a high survival rate and good growth of the offspring were observed.
From these results, it could be concluded that sodium erythorbate has no teratogenic effect in rats under the present experimental conditions.

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The teratogenicity of potassium metabisulfite (K₂S₂O₅), which is used widely as a food additive, was studied in Wistar rats. The pregnant rats were fed a diet containing 10, 1 or 0.1% potassium metabisulfite from day 7 to day 14 of pregnancy. The maternal body weight gain during pregnancy and food intake during the administration period were markedly suppressed but no clinical signs of toxicity were observed in the 10% group. In this group, the fetal body weight was significantly lower than that of the control group and the postnatal survival rate of the offspring was slightly decreased. Maternal malnutrition during pregnancy might be one of the reasons for these adverse effects in the 10% group. External, skeletal and internal examinations of the fetuses revealed no evidence of teratogenesis in any group. No adverse effects on the pre- and postnatal development of the offspring were found in the 1 and 0.1% groups. It could be concluded that potassium metabisulfite has no teratogenic effect in rats under the present experimental conditions.

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The teratogenicity of disodium glycyrrhizinate (NaG) was examined in Wistar rats. Pregnant rats were fed diet containing 2%, 0.4% or 0.08% NaG ad libitum from day 0 to 20 of pregnancy. A comparison of the control and all groups of rats treated with NaG revealed no significant differences in food intake and in body weight gain during pregnancy. However, body weight gains after delivery were significantly lower in the two highest level groups. No significant differences between the control and NaG-treated groups were found in the numbers of corpora lutea and implants of dams, in the number of live fetuses and intrauterine dead fetuses per litter, in the sex ratios, in fetal body weight of both sexes, in the placental weight, in the degrees of ossification in the sternebrae and caudal vertebrae of the fetuses, or in the live birth index, survival rate or body weight gain of the offspring within 8 weeks after birth. Several kinds of skeletal variation of the fetus were observed in all the groups treated with NaG, but the incidences showed no significant differences as compared with the control. Except for one fetus with dilatation of the renal pelvis in the 0.08% group, no fetus with external, skeletal or internal malformations was found at any level examined. From these results, it can be concluded that NaG has no teratogenic effects on the fetus in rats.

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An analytical method was developed for quantitative determination of residual chlorine in foods using head space gas chromatography.
The principles of the proposed method are as follows. Phosphate buffer solution of pH 7.0 and potassium cyanide solution were placed in a vessel for head space analysis, a sample was added, and the resultant mixture was left standing in the vessel (covered with a lid) for 30 minutes. Cyanogen chloride formed therein was determined by ECD-GC. The values of residual chlorine in tap water measured by the present method were compared with those obtained by the o-tolidine colorimetric method. The respective values were quite consistent. The determination limit was 0.1μg/g.
Analysis of 67 specimens of vegetables and processed fish and shellfish products available in the market revealed 14.0μg/g and 11.6μg/g of residual chlorine in two specimens of soybean sprouts.

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This study was performed in order to investigate the effects of gonyautoxins (GTXs) and tetrodotoxin (TTX) on the movement of the excised small intestine of a guinea pig.
The results may be summarized as follows:
1) The peristaltic movement appeared when pressure was applied to the internal wall of the excised intestinal canal by the method of Trendelenburg. However, after pretreatment of the intestine with GTXs (1.0MU/ml) for 2min, this movement was not observed.
2) The tonic contractions of the intestinal preparation induced by adding acetylcholine (ACh, 1×10^-7g/ml) to the organ bath were not suppressed by pretreatment with GTXs (1.0MU/ml) or TTX (1.0MU/ml) for 3min.
3) The contractions induced by nicotine (1×10^-5g/ml) were markedly suppressed by pretreatment with GTXs (1.0MU/ml) or TTX (1.0MU/ml) for 3min.
4) The contractions of the intestinal preparation elicited by transmural electrical stimulation (0.5msec duration, 40Hz supramaximal voltage) were completely suppressed by pretreatment with GTXs (1.0MU/ml) or TTX (1.0MU/ml) for 3min.
From these results, it may be concluded that GTXs have inhibitory effects on the liberation of ACh from the nerve endings, but not on the ACh receptor of the smooth muscle.

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An automatic colony counter (ACC), Artek 880, was compared with the manual technique for accurate and rapid evaluation of the viable bacterial counts in foods.
For 30 to 300 colonies on a plate, from either pure culture strains or raw milk samples, the accuracy of repetition of ACC and personal errors in the manual technique almost satisfied the requirements of Standard Methods for the Examination of Dairy Products (14th ed.) recommended by the American Public Health Association.
The correlation coefficient between ACC counts and manual counts of pure culture strains and raw milk samples were 0.985 and 0.972, respectively, and the slopes of regression lines were 0.903 and 0.942, respectively. Furthermore, acceptable arithmetic mean counts by ACC (within ±10% of the corresponding mean manual counts) were 74% and 46%, respectively.
These results indicated that ACC is useful for routine work on large numbers of samples in food quality control.

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A sensitive and simple separation method for methylguanidine (MG) and agmatine (AGM) in various foods by high performance liquid chromatography with a fluorescence detector is described. Food samples were homogenized in 4% trichloroacetic acid solution and filtered. The guanidino compounds were extracted with n-butanol from the filtrate under alkaline conditions and re-extracted with 1N hydrochloric acid. They were converted into fluorescent derivatives by reaction with benzoin under alkaline conditions. Guanidino compounds were separated and determined on a Nucleosil 5C₁₈ column with methanol -0.5M Tris-Hydrochloric acid Buffer (pH 8.6) (8:2) as the mobile phase. The recoveries of MG and AGM added to smoked dried fish and wiener sausage were 85.0% (MG), 82.5% (AGM) and 88.2% (MG), 82.3% (AGM), respectively. The limits of determination were 0.1ppm for MG and 0.5ppm for AGM. Smoked-dried bonito (Katsuo-bushi) contained 12 to 238ppm of MG. The amounts of MG and AGM determined in almost all samples were small or below the detection limits.

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Eighty-six specimens (58 males, 28 females) of pufferfish, Fugu vermicularis vermicularis, were collected off Futtsu, Tokyo Bay, and the toxicity of various organs was assessed.
More than half of the specimens were found to possess a toxic liver, the highest and average toxicity scores being 1, 500 and 132MU/g, respectively. The testis of this pufferfish, which has been reported to be non-toxic, was found to be weakly toxic, depending upon the specimen. Most ovaries were toxic, with the highest score of 5, 900MU/g and the average score 874MU/g. The muscles were non-toxic, except for a single, weakly toxic example.
It was noted that the Tokyo Bay specimens maintained a fairly high toxicity level even after the spawning season, in contrast to the observations on other pufferfishes, in which the toxicity levels sharply decreased down after spawing.

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The accumulation and excretion of the herbicides molinate, benthiocarb (thiobencarb), chlornitrophen (CNP) and chlomethoxynil in bivalve mussels (Mytilus edulis) were studied in the laboratory. An inverse correlation (r=-0.96) was observed between the bioconcentration ratio and the water solubility of these chemicals and it was found that they are concentrated at higher levels in adipose tissues.
Concerning excretion from the mussel, molinate was excreted quickly; the biological half-lives of the other herbicides in the whole body were 0.7 day for benthiocarb, 4-8 days for CNP and 1.6 days for chlomethoxynil. The amounts of individual chemicals in various tissues did not differ markedly. It was suggested that over 90% of each chemical taken into mussels was excreted into the surrounding sea water without being metabolized.

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A simple method for the determination of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in “health food” contained EPA by gas liquid chromatography (GC) was developed.
A sample of 100mg was methyl-esterified by the boron trifluoride-methanol (BF₃-MeOH) method and the product was extracted with hexane containing triphenyl ethylene as an internal standard. The sample solution was determined by GC on a 10% Silar 10C column. Recoveries of EPA and DHA added to oils were in the range of 98.8-107% and 98.8-122%, respectively. The present method was applied to “health food” contained EPA. The results showed thet the quantities of EPA and DHA (mg) per gram of oils and fats of the supplements were 20.3 and 11.4 on average, respectively.
Moreover, the fatty acid composition, acid value (AV), peroxide value (POV) and tocopherols (Toc) in the “health food” were investigated. Though the “health food” is easily taken, from the viewpoint of safety, nutritional balance and price, EPA and DHA should be taken in the diet in the form of sardines, mackerels, and so on.

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The effects of coexisting substances (inorganic salts, glucose, sucrose, sodium salts of carboxylic acids and amino acids) on the value of tetrodotoxin (TTX) toxicity as determined by the mouse bioassay method were investigated. The relative toxicity decreased linearly with increase of the concentration of inorganic salts such as sodium chloride. In the presence of glucose or sucrose, the relative toxicity was independent of the concentration. The effects of TTX concentration were also examined. The relative toxicity decreased with increasing TTX concentration in the presence of sodium chloride and with decreasing TTX concentration in the presence of glucose.
Sodium salts of mono- and dicarboxylic acids progressively-decreased the relative toxicity as the size of the molecule became larger. The more hydrophobic and larger amino acids also decreased the relative toxicity, but the decrease in the presence of amino acids was small compared with that caused by the sodium salts of carboxylic acids.
These results suggest that the mouse bioassay method may be affected by coexisting substances (inorganic salts, organic acids, etc.).

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The cadmium-binding capacity of the soybean protein fraction was determined by equilibrium dialysis. The aqueous supernatant was extracted from the soybeans with 0.01M Tris-hydrochloric acid buffer (pH 7.4) by ultracentrifugation. The protein fraction (F-I, >10, 000 in molecular weight) was separated from the extract by Sephadex G-50 chromatography. Binding measurement was carried out in 0.01M Tris-hydrochloric acid buffer (pH 7.4). The binding data were analyzed according to the Scatchard equation. A Scatchard plot of the binding data showed several phases, indicating heterogeneity of the binding sites. The maximum amount of cadmium bound by F-I was calculated to be 43, 000μg/g. When cadmium was added to the aqueous supernatant, five cadmium-binding fractions (at molecular weights of>100, 000, 75, 000, 38, 000, and two low-molecular species) were detected by chromatography on Sephadex G-100. The major cadmium-binding fraction in F-I was the macromolecular species of>100, 000 in molecular weight.

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A sensitive method for the determination of aspartame (APM) in foods by high performance liquid chromatography with a fluorescence detector is described.
APM was extracted from a food sample with water. The extract was passed through a C₈ column (LiChroprep RP-8, 40-63μm, 1g). The column was washed with 0.05M acetate buffer solution (pH 6.0), then APM was eluted with a mixture of acetonitrile-0.05M acetate buffer solution (3:7). APM in the eluate was derivatized with fluorescamine and injected onto a LiChrosorb RP-8 column; a mixture of acetonitrile-0.05M acetate buffer solution (pH 6.0) (22:78) was used as a mobile phase.
The recoveries of APM added to foods were more than 92%. The detection limits of APM in liquid food and solid food were 2ppm and 4ppm, respectively.

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By using a rapid and sensitive method for simultaneous detection of nivalenol (NIV), deoxynivalenol (DON), and zearalenone (ZEN), their natural occurrence in 18 samples of scabby wheat grains harvested in 1984 from eighteen farms in the Tokachi district of Hokkaido, a northern island of Japan, was detected. DON and ZEN were detected in all 18 samples, among which 7 (39%) samples were co-contaminated with NIV. The contents averaged 205ng/g of NIV, 3812ng/g of DON, and 189ng/g of ZEN. Thus, NIV, DON, and ZEN coexisted in the scabby wheat grains harvested in Hokkaido, where severe scabby wheat intoxication breaks out sporadically. Interestingly, the average content of DON in wheat was about 20-fold higher than that of NIV.

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