For analysis of methylguanidine (MG) which can be nitrosated to highly mutagenic and carcinogenic methylnitrosocyanamide, preliminary experiments were made to determine the most appropriate analytical procedures. Extraction of sample materials with a mixture of concentrated hydrochloric acid and 5% solution of ethanol in acetone (2:1000) and column chromatographic fractionation of the extract, first using microcrystalline cellulose as adsorbent and a mixture of concentrated hydrochloric acid and ethanol (2: 1000) as eluant, and then using Amberlite CG-50 resin as adsorbent and 1.6N sodium hydroxide as eluant were thought to give an adequate isolation of MG from other related compounds. Amount of MG was then quantitatively determined by Sakaguchi reaction. The isolated MG was confirmed by paper chromatography of the above fractions using a mixture of butanol, acetic acid, and water (4:1:2) as a developer. For gas chromatographic examination, conversion of methylguanidine to its pyrimidine derivative by heating with anhydrous sodium carbonate and acetyl acetone in n-amyl alcohol was tried but the conversion was not well quantitative.
The determination of migrated vinyl chloride monomer (VCM) in foods by internal standard method was studied. As internal standard substance, methyl vinyl ether (MVE) was used, VCM and MVE could be concurrently collected into a test tube cooled with dry ice-ethanol by nitrogen bubbling. The concentration of VCM in the cooling test tube was determined by gas chromatography. VCM added in the samples such as soy sauce, wine, edible oil and jam were determined by the internal standard method and by the absolute calibration curve method. The values of VCM obtained by the proposed method were reasonably greater than those obtained by the absolute calibration curve method.
A method for the determination of pentachlorophenol (PCP) in wooden chips was investigated. PCP was distilled with steam and extracted with n-hexane from acidified distillate. PCP in n-hexane was extracted with 0.1M sodium carbonate solution and then acetylated with acetic anhydride. PCP acetate was extracted with n-hexane and purified by silica gel column chromatography. The eluate was analyzed by gas chromatography using an electron capture detector. Acetyl derivative of PCP gave heigher recorder response than free PCP. The recovery of PCP from PCP added wooden chips was 98.4%.
The amount of 3, 4-benzopyrene was determined in vegetable oils, butter, and margarine according to the method for the foods that had been already published. 3, 4-Benzopyrene in vegetable oils was extracted with dimethylsulfoxide after dissolving in n-hexane using a separatory funnel, and one in butter and margarine was extracted using the wax extraction apparatus which had been contrived by Shiraishi, et al. The clean up was proceeded through a silica gel column (1×30cm, petroleum ether: ether (9:1)) and a 1% water containing alumina column (1×30cm, n-hexane: ether (2:1)) in succession and then 3, 4-benzopyrene was determined by fluorophotometry. The recovery of 3, 4-benzopyrene by the proposed method was 94.9% for vegetable oils and 95.2% for butter, when 3ppb of 3, 4-benzopyrene was added. In commercial samples, 3, 4-benzopyrene was detected at levels of 0.08 to 1.24ppb in 9 out of 12 samples of vegetable oils, 0.35 to 0.60ppb in all the 3 samples of butter, and 0.46 to 0.83ppb in all the 3 samples of margarine on the market.
Fluorometric assay of malonaldehyde in oxidized oil was investigated. The mixture of 20g of whale oil or rape seed oil and 20ml of n-hexane was prepared. To the mixture was added to 5ml of pH 7.8 phosphate buffered solution (ionic strength, 0.05) and was shaken for 10min at room temperature. The shaken mixture was centrifuged at 4, 000rpm for 20min and was separated the buffered solution extracts from the mixture. A part of the extract was reacted with DL-valine in 50% acetic acid (A), and the other part was added on a polyamide dry column and eluted with water. The eluate was reacted with DL-valine in 50% acetic acid (B). Fluorescence of (A) and (B) were measured at 460nm when samples were excited at 390nm. Malonaldehyde in oil was determined from (fluorescence (A)-fluorescence (B)). Malonaldehyde in rape seed oils was more than 1.9μg/kg.
The purpose of this study was to clarify whether dietary calcium levels would be related to the depositions of PCB and calcium in dam and fetus of rats fed with various diets containing 100ppm PCB (Kanechlor 500). The results obtained were as follows: 1) The increase of liver weight in pregnant rats treated with PCB was significantly promoted in the deficiency of calcium in diets. 2) It was recognized that the amounts of ash and calcium decreased markedly in the bone and fetus of pregnant rats fed with calcium deficient diet containing PCB. 3) In the liver and bone of pregnant rats, the PCB deposition of calcium deficient group was higher about two times than that of calcium normal group. 4) Under the administration of calcium deficient diet to pregnant rats, PCB transferred from the dam to the fetus during gestation increased remarkably. 5) No significant change was found in external and skeletal abnormalities of the fetuses from all experimental groups.
In order to dissolved the substances to be estimated by the chicken embryo test, various solvents have been used. This study was performed to ascertain whether these solvents affect on the toxic strength of the samples for the chicken embryos. In this experiment, aflatoxin B1 was used and water, DMSO, propylene glycol and olive oil were selected as its solvents. Aflatoxin B1 dissolved in each solvent was inoculated into the air cells or yolk sacs of White Leghorn fertile eggs aged four to seven days. The toxicity was estimated by the mortality of the embryos three days after inoculation. In the air cell inoculation, though all the solvents showed by themselves none of the toxic effect within their doses, the toxicity of aflatoxin B1 varied by the kind of the solvent. That is, the toxic strength came in the order of the sample dissolved in water, DMSO, propylene glycol and olive oil, corresponding with the amount of aflatoxin B1 incorporated into the egg through its membrane. Thus it was found that the changes in the toxicity were dependent on the membrane permeability of aflatoxin B1 affected by the solvents. In the yolk sac inoculation, aflatoxin B1 dissolved in water and olive oil showed almost the same toxicity, but that dissolved in DMSO and propylene glycol did not prove any toxicity at all. It is assumed that those decrease of the toxicity in latter solvents were due to disturbed diffusion of the toxin from its injected site, since they were noticed to denature the yolk proteins.
In order to establish a systematic method for the determination of DP, OPP and TB in citrus oil, this investigation was carried out. Alumina column clean up followed by either FID or FPD gas chromatographic technique was adopted. 0.5g of sample was taken and was eluted through activated alumina (Merck, Art. 1097 Aluminiumoxid standardisiert heated for more than 15hr at 110°C was used) with each 30ml of n-hexane, ethyl acetate and 28% ammonia water-methanol mixture (2:8), in turn. Since it had been confirmed that β-carotene was eluted together with DP while that OPP and TB moved together with Sudan G, these two dyes served as tracers of the three preservatives. DP recovered from ethyl acetate fraction was determined by use of FID-GC (5% DEGS+1%H3PO4), while OPP and TB were recovered from the last eluate. OPP was determined by use of FID-GC, while TB was measured with FPD-GC fitted with columns packed with 0.5% OV-17 on Neosorb NG 40/60. The detection level of each preservative was around 5ppm. More than 98% of recoveries were obtained on addition of from 20 to 100ppm of the preservatives on lemon oil. Residue levels of these three preservatives in thirty four imported citrus oils were analyzed.
Furylfuramide (2-(2-furyl)-3-(5-nitro-2-furyl) acrylamide) is a chemical which had been used widely in such foods as soybean caud, fish paste product and fish sausage as a food preservative in Japan from 1965 to 1974. The effects of furylfuramide on pregnant mice, fetuses and neonates were studies in ICR: JCL mice for three generations. Furylfuramide at dietary concentations of 0.1 or 0.5% was fed to both male and female weanling mice (F0). Thereafter the same dietary concentrations were administered continuously to each generation until F2 offspring were 60 days of age. Hepatic enlargement and degenerative hepatic changes were consistently found in the treated dams and their offspring. High mortality in F1 neonates and poor conception in F1 dams occurred in the group fed 0.5% furylfuramide. Suppression of body weight gain was also observed in pregnant mice and offspring at the 0.5% level. However, continuous administration of furylfuramide for three generations did not increase fetal mortality nor did it cause major malformations.
An analytical method was investigated for the quantitative determination of water-soluble acid dyes permitted in Japan in foods by thin layer chromatography. The presented method was composed for the following: The water-soluble acid dyes were extracted from foods with a liquid anion exchange resin (Amberlite LA-2), and washed with water. The dyes could be re-extracted from the resin solution by extracting with ammonia water. The dyes in the extracting solution was separated by thin layer chromatography using cellulose or silica gel plate. Each spot was scanned by the dual wave length and zig-zag scanning type densitometer. The sensitivity limit of detection were 0.05μg of the dyes on the TLC plate. The analytical conditions were as follows: Slit; 1.25×1.25mm Mode; Zig Zag scan Wave length; Reference. λR=700nm Sample. λS=a maximum wave length of each dyes The recoveries of dyes added in juice, kamaboko (made from fish paste) and suhama (made from rice powder) were more than 93% except indigo carmine. As the results of the survey of commercial foods by the use of above mentioned procedure, concentration of dyes was used under about 100ppm in most of samples. The presented method was found to be sufficiently satisfactory for quantitative determination of dyes in foods.
Three lots of Mysost-type whey cheese were subjected to the determination of nitrate and nitrite. Nitrite content was measured by use of the well-known diazotization-coupling reaction, while cadmium column reduction method by use of metallic cadmium was applied for the determination of nitrate. The results revealed that nitrite contents were within the range of 1.8-2.7mg NO2/kg cheese, while nitrate contents were 48.6, 100.0 and 390.0mg NO3/kg cheese, respectively. No difficulty was encountered during the application of the cadmium column reduction method formerly settled for the analysis of natural and processed cheese on Mysost-type whey cheese.
Corn oil was autoxidized at room temperature by UV irradiation and four samples were obtained in the course of autoxidation. Peroxide values of the four samples were 202 (SO200), 399 (SO400), 805 (SO800), and 1117 (SO1100) respectively. Each sample was administered to male Wistar rats and its effects on lipid metabolism and drug metabolizing enzyme system were investigated. The results obtained were as follows: (1) In the experiments at a single oral dose (4ml/kg) of autoxidized oils, aniline hydroxylase activity was significantly decreased at SO400 administration, but was increased at SO1100. The content of liver triglyceride was decreased at SO1100. The activity of hepatic δ-aminolevulinic acid synthetase was increased both at SO800 and at SO1100 administration. (2) In the repeated oral dose of SO800 or SO1100 (4ml/kg/day) for 10days or 7days respectively, aniline hydroxylase activity and the ratio of liver weight to body weight were increased at SO1100 administration. However, the activity of drug metabolizing enzymes, the contents of glycogen, microsomal protein, triglyceride, and cytochromes were not altered at SO800 administration. (3) In the experiments of a single intraperitoneal injection (4ml/kg) of SO800, aminopyrine demethylase activity, the contents of microsomal protein and cytochrome P-450 were increased in 2hr after the injection, whereas the content of liver triglyceride was decreased approximately 50% in 6 or 12hr after the injection. On the other hand, the contents of liver triglyceride and serum cholesterol were increased in 24hr after the injection.