Showing papers in "Food Hygiene and Safety Science in 1976"

Separation and Detection of Polychlorodibenzofurans (PCDFs) on Japanese Commercial PCBs (Kanechlors) and Their Heated Preparation

Abstract: Modified analytical methods by Vos and Poter et al. were used for the separation of polychlorodibenzofurans (PCDFs) from Japanese commercial PCBs (Kanechlors) which were used in large quantities in Japan. Di, tri, tetra, penta, hexa and heptachlorodibenzofurans were detected on Kanechlors by GC-MS analysis. Total PCDF concentrations were 9, 33, 4 and 4ppm in Kanechlor 300 (KC-300), Kanechlor 400 (KC-400), Kanechlor 500 (KC-500) and Kanechlor 600 (KC-600), respectively. In addition, to investigate the possibility that PCDFs were formed during the usage of PCB as heat transfer medium, KC-400 was put into a pyrexglass ample and heated on a gas chromatograph oven. However, PCDFs did not generate by heating for 15 days at 300°C.

The Finiding of Polychlorodibenzofurans in Commercial PCBs (Aroclor, Phenoclor and Clophen)

Abstract: According to the modified methods of Vos and Poter et al., polychlorodibenzofurans (PCDFs) were separated and cleaned-up from commercial PCBs (Aroclor, Phenoclor and Clophen) which had been used in large quantities in the world. PCDFs were identified by gas chromatography-mass spectrometry and quantified by gas chromatography. PCDFs were found in all samples and the results were as follows.Aroclor: Tri to heptachlorodibenzofurans were found, and the total PCDF concentrations were 2-9ppm.Phenoclor: Tri to heptachlorodibenzofurans were found, and the total PCDF concentrations were 8-13ppm.Clophen: Di to hexaclorodibenzofurans were found, and the total PCDF concentrations were 5-17ppm.

Studies on the Media for Enterotoxin Production and Sporulation by Clostridium perfringens

Abstract: The present study Was aimed at obtaining a suitable medium for enterotoxin production and sporulation by Clostridium perfringens. Using both heat-stable (NCTC 8798) and heatlabile (TCW-224-1) strains, various nutritional ingredients were taken into considaration for enterotoxin production as well as sporulation.1. Various peptones, carbohydrates and meat infusions were examined for enterotoxin production, among which polypeptone S (Daigo), soluble starch and chicken meat infusion had the most enhancing effects on enterotoxin production. The appropriate pH range after cultivation was from 4.8 to 6.8, 6.2 being optimum.2. From the results described above, a new medium having following composition was established. Soluble starch, 5g; NaCl, 5g; Na2HPO4, 5g; FeCl3·6H2O, 10mg; sodium thioglycolate, 1g; in 1 liter chicken meat infusion. The pH of chicken meat infusion was adjusted to 7.5, autoclaved and filtered before the addition of other ingredients. This new medium was filtered after heating and autoclaved as usual.3. When various kinds of media were used, enterotoxin production by NCTC 8798 strain was detectable at spore numbers as small as 10/ml in culture, and the amount of enterotoxin seemed to increase as spore numbers increased. In a few instances, however, no enterotoxin was detected even at spore numbers of 105-7/ml in culture, meaning that, when enterotoxin is well produced, good sporulation usually occurs in the medium, and that the reverse is not always true. This implies that the quality of spores in addition to quantity or the degree of maturation in the sporulation stage, must also be taken into account for the enterotoxin production by this organism.

Studies on Metabolism of Food Additives by Microorganisms Inhabiting Gastrointestinal Tract (IV)

Abstract: Cyclohexylamine (CHA) is a metabolite of the artificial sweetening agent Cyclamate which was rejected from the food additives. The notice about cyclamate is subsiding, but studies with this substance have been continued. To determine sodium cyclamate (CHS-Na) assimilating bacteria inhabiting gastrointestinal tract of animals, a quantitative and qualitative bacteria analysis for faecal materials have been conducted in a monkey administered orally with 20% CHS-Na solution (1.25ml/kg daily), and in untreated monkey. The faecal flora of the normal monkeys predominently constituted Catenabacteria and Peptostreptococci, and Clostridia, Bacteroidaceae, Bifidobacteria, and Veillonella, all of which belong to the obigate anaerobe, were commonly found in order. Facultative anaerobes like Lactobacilli and Streptococci were abundantly found, and Enteroba cteria and Staphylococci could be detected in the faecal homogenates. On the other hand, the constitution of faecal flora in the monkey excreting CHA in the urine was not quite different from that of the normal monkeys. In addition, any marked alteration of the flora in the treated monkey was not observed throughout this analysis. In parallel with this, in vitro, culture study was carried out by using the modified BC broth containing 0.25% of CHS-Na. Initial mass cultures obtained from the faecal homogenates were inoculated into the BC broth and incubated at 37°C for 14 days under anaerobic condition, and production of CHA periodically examined by the chemical procedure. The production of CHA by the initial mass culture was gradually increasing in progressively. After 14 days culture, bacteria survived were Clostridia, Bacteroidaceae, Lactobacilli, and Streptococci, those which may have CHA productivity. These bacteria and the initial mass culture were subcultured again on the BC broth but no production of CHA was observed in these subcultures. After the cultivation on differential media for identification, these organisms exposed to the air (O2) may have lost their CHA productivity. From this study, it is indicated that although CHA converting bacteria was not definitely detected in vitro, some anaerobic bacteria such as Clostridia, Bacteroidaceae, Lactobacilli and Streptococci seemed to have sodium cyclamate metabolizing activity in gastrointestinal tract of the monkeys.

Determination of bromate in foods by gas chromatography

Abstract: An improved method for quantitative determination of bromate in bread and fish paste products was established.A sample was homogenized with distilled water and made up to 200ml after centrifugation. The supernatant was filtered and n-butanol-ethanol (2:1) mixture was added to 10ml of the filtrate. The mixture was allowed to stand for 10min then centrifuged to remove the precipitate. DEAE-Sephadex A-25 suspension and acetic acid were added to the supernatant, and the mixture was stirred for 5min. After centrifugation, the supernatant was subjected to the same procedure with DEAE-Sephadex A-25 suspension. The DEAE-Sephadex A-25 was mixed with ethanol, and rinsed twice with ethanol, 5.0% acetic acid and distilled water. The solid was packed in a chromatocolumn, and bromate was eluted with 30ml of 30% potassium chloride solution. Two ml of 4×10-3M styrene monomer solution (washed with 1% sodium hydroxide solution before use), 1ml of 0.01M potassium bromide solution and 1ml of sulfuric acid were added to the effluent, and the whole was shaken vigorously. Styrene bromo derivative was extracted with 2ml of n-hexane, and bromate was determined by ECD-GC.In this method, it was found that bromate in foods could be determined with a recovery of 80-92.3% and a variation coefficient of 6.2-2.6%. The detection limit was 0.01μg/g.

Accumulation of Mercury in Sea bream (Chrysophrys major) and Horse mackerel (Trachurus japonicus)

Abstract: Rate of accumulation of mercury in the different tissues of marine fish species was followed by setting two series of rearing experiments using sea bream (Chrysophrys major) and horse mackerel (Trachurus japonicus). In the first series, fish were fed with ground flesh of horse mackerel in the sea water added by mercuric chloride at the levels of 0.1ppm, 0.01ppm, and 0.001ppm. In the second one, fish were fed with pellets prepared from a variety of deep sea serranids (Malakichthys griseus) which accidentally gave a high concentration of methyl mercury; 1.4ppm. Both groups consisting 75 fish were held in aquaria with capacity of 3 tons of sea water for 91 days. In another series of experiment, the same number of fish were also fed by methyl mercury-containing pellets in the sea water added HgCl2 at level of 0.1ppm.In the first group of fish, mercury mostly accumulated in spleen, followed by brain, liver and muscle in the order of concentration. At the end of rearing period, mercury level of muscle was 0.3ppm and 3.0ppm, when they were held in sea water containing mercuric chloride of 0.01ppm and 0.1ppm, respectively. Almost negligible level of mercury in organic form was noticed in these tissues of fish after 13 weeks and this may suggest methylation of mercury hardly occur in these organs.By oral intake of the pellets, mercury levels were much lower than those of branchial invasion. A level of 0.8ppm in the muscle and brain and 1.2ppm in the liver and spleen was detected respectively after 13 weeks of rearing. Mercury concentration in spleen was not so remarkable after oral feeding of methyl mercury.

The Preservative Effects of Sorbic Acid for Fish Sausage

Abstract: Sorbic acid has been used as antiseptic for fish sausage, since the use of AF-2 [2-(furyl)-3-(5-nitro-2-furyl) acrylamide], which had been one of the effective preservatives for fish sausage, was prohibited in July of 1974. Therefore, the preservative effects of sorbic acid for fish sausage was reinvestigated in connection with pH value and storage temperature of the sausage. The results obtained are as follows.1) At storage temperature of 30°C, the shelf life of fish sausage with the pH values of lower than 5.9 was about one week. Additions of sorbic acid to the sausages at the level of 0.1 and 0.2% prolonged their shelf lives for one and two weeks respectively.2) At storage temperature of 10°C, the sausages with about 6.2 of pH did not show any sign of spoilage for the test period of 7 weeks without regard to the addition of sorbic acid. The same results were obtained at the elevated storage temperature of 15°C.3) From these results, it was presumed that fish sausages added at the level of 0.2% of sorbic acid could be preserved for at least two months at about 10°C, if the pH is adjusted to the suitable pH range in which sorbic acid shows effective bacteriostatic action.

Studies on Analysis of L-Aspartyl-L-Phenylalanine Methyl Ester in Foods

Abstract: An analytical method was investigated for detection and determination of L-aspartyl-L-phenylalanine methyl ester (APM) in foods by thin layer and high speed amino acid chromatography.The presented method was composed of the following steps: Sample was dialized using a cellulose tube against water (adjusted to pH 4) or extracted with methanol. APM in the outer solution obtained by dialysis or the methanol extract was separated by thin layer chromatography using silica gel or cellulose plate. APM spot was detected on TLC plate under an ultraviolet light (365nm) after spraying with fluorescamine reagent. The sensitivity limit of detection was 12ng of APM on the TLC plate.High speed liquid chromatography of APM carried out using the column LCR-2 (Buffer pH 4.25).The analytical conditions were as follows:Column temperature; 52°CReaction bath; 95°CChromogenic reagent; ninhydrineThe recovery of APM added in orange juice and other soft drinks were 94-99%.The presented method was found to be sufficientry satisfactory for determination of APM in beverages, coffee, tea, pudding, topping, and cereals.

Proteolytic and Lipolytic Activities of Psychrotrophic Bacteria Originated from Raw Milk

Abstract: The proteolytic (in skim milk agar) and lipolytic (in Tributyrin agar and Clossry's agar) activities of the bacteria isolated at 5°C for 10 days and 20°C for 4 days incubations from raw milk were tested at 5°C for 10 days, 20°C for 4 days, and 35°C for 2 days incubations respectively.Of the 792 strains isolated at 5°C incubation, 23.1% showed the proteolytic activity, and 40.9% showed the lipolytic activity in the Tributyrin agar, and 48.0% in Clossry's agar. Of the 588 strains isolated at 20°C incubation, 37.9, 17.9, and 23.6% showed the proteolytic or lipolytic activity respectively.In the 792 strains isolated at 5°C incubation, the number of strains showed both proteolysis and lipolysis at 20°C incubation was the most, and the one at 5°C incubation was next. On the other hand, in the 588 strains isolated at 20°C incubation, the greatest number of strains showed both proteolysis and lipolysis was at 20°C incubation, and the next was at 35°C incubation.The percentage of strains showed proteolytic activity was high in Bacillus, coliform bacteria, Pseudomonas isolated at 5°C incubation, and was high in Pseudomonas, Alcaligenes-Achromobacter, Flavobacterium, Streptococcus, coliform bacteria isolated at 20°C incubation. Most of gram-negative bacilli showed proteolytic activity at 5 and 20°C incubation, and, most of gram-positive bacilli and cocci showed at 20°C and 35°C.The percentage of strains showed lipolytic activities to both tributyrin and butter-fat was more than 70% in Bacillus, Acinetobacter, Alcaligenes-Achromobacter, unclassified gram-negative bacilli and coliform bacteria isolated at 5°C incubation, also Pseudomonas showed lipolysis to butter-fat in more than 70% of strains. In the strains isolated at 20°C incubation, Acinetobacter, Pseudomonas, Staphylococcus, Aeromonas, and coliform bacteria had respectively high percentage of strains showed lipolysis in both tributyrin and butter-fat.Coliform bacteria and Pseudomonas were main proteolytic and lipolytic activators, and most strains of them showed strongly their activities at 20°C incubation.

Effect of Coal Tar Dyes on Pancreatic Lipase

Abstract: The inhibitory effects of different coal tar dyes, most of which are used for food additives at present time, on pancreatic lipase were examined in vitro by the titration method using a pH stat.Xanthene dyes except acid red showed the significant inhibition of lipase activity in order of eosine, erythrosine, phloxine and rose bengale. The inhibition was gradually enhanced during preincubation of dye with enzyme or substrate, almost independent of pH in preincubation. On the other hand, other dyes tested did not inhibit the enzyme activity at neutral pH, but most of them showed remarkable inhibition at pH 2.The inhibitory effect of rose bengale was considerably enhanced by exposure of preincubation mixture under a fluorescent lamp, suggesting a photo-inactivation of the enzyme by halogen atom in the excited molecule of dye.The inhibition was also influenced by the co-exsistence of other substances such as protein, sodium chloride and sucrose. From the kinetic study, the type of inhibition of pancreatic lipase by rose bengale was non-competitive.

Studies on Removal of Residual Pesticides from Fruits and Vegetables by Washing

Abstract: For the purpose of finding a method for optimization of removal effect on the residual Bordeaux from tomato, an investigation has been made by use of tomato sprayed with Bordeaux in a laboratory.It was found that the combined use of sugar fatty acid ester and potassium pyrophosphate exhibited good detergency to remove the residual Bordeaux on tomato.Using this combined detergent, it was also recognized that almost complete removal of the residual Bordeaux was achieved through the following conditions: more than 0.2% of detergent concentration, over 2 minutes washing, above 25°C as washing temperature, and stirring of washing solutions with above 50r.p.m.

Studies on the Formation of Nitrosamines (IV)

Abstract: The rates of carboxymethylnitrosourea (CMNU) formation from glycocyamine (GC) and sodium nitrite were studied at pH 2.5 and 37°C, considering the condition in the human stomach after ingestion of foods. The initial rates of CMNU formation were calculated from the absorbance at 394nm after decomposition of nitrite with sulfamic acid. They were proportional to the concentration of glycocyamine and nitrite under experimental condition.The effects of thiocyanate, citrate, tartrate, sodium chloride and ascorbate on the rates of CMNU formation from GC and nitrite were also studied. Thiocyanate, citrate and tartrate accelerated CMNU formation, and ascorbate inhibited it. Sodium chloride did not affect CMNU formation under experimental condition.

Studies on the Formation of Nitrosamines (III)

Abstract: The reaction products of glycocyamine, one of the naturally occurring guanidines, and nitrite under acidic conditions were explored. The main product was identified as carboxymethylnitrosourea, a derivative of hydantoic acid whose secondary amino group was nitrosated, by elemental analysis and NMR spectrum. The structure was further confirmed by mixed analysis and by comparing IR and absorption spectra with the reaction product of hydantoic acid and nitrite.A nitrosocyanamide was expected to be one of the reaction products, but it could not be detected in various conditions examined. Stability and the decomposition products of carboxymethylnitrosourea were also discussed.

Determination of Polychlorinated Dibenzo-p-dioxins and Polychlorinated Dibenzofurans by Exhaustive Chlorination Method

Abstract: A method of quantitative determination of minute amount of polychlorinated dibenzo-p-dioxins (PCDD) and polychlorinated dibenzofurans (PCDF), complex mixtures of isomers, was described. PCDD and PCDF (1-10μg) were exhaustively chlorinated to octachlorodibenzo-p-dioxin (OCDD) and octachlorodibenzofuran (OCDF) by heating with perchlorinating agent at 68-69°C for 2 hours. The perchlorinated compounds were quantitated by gas chromatography equipped with an electron capture detector using XE-60 as a liquid phase. Yields of perchlorinated compounds from dibenzo-p-dioxin and dibenzofuran both having two or more chlorine atoms were more than 75%, while from those having no or one chlorine were 4-16%. This method was also applicable to quantitation of polychlorinated biphenyls.By this perchlorination method, the contents of PCDD and PCDF in pesticide PCP, Kanechlor (KC) and Yusho oil were determined. From two brands of PCP, 430 and 0.6μg/g of OCDD and 290 and 1.3μg/g of OCDF were quantitated, respectively. Only PCDF were detected in KC-300, KC-400, KC-500, KC-600 and Yusho oil (3 samples), and their estimated contents calculated from the amounts of OCDF produced were 1.5, 16.6, 2.5, 2.7 and 4.9μg/g, respectively.

Effects of Dibutyl Phthalate on Lipid Metabolism and Drug Metabolising Enzyme System in Rats

Abstract: Effects of dibutyl phthalate (DBP) on the liver constituents and the drug metabolizing enzyme system were investigated in rats.1. In the experiments at a single oral dose of DBP (630 or 1260mg/kg), the glycogen content was decreased only at the high dose, but no effects were observed on the contents of glycogen, triglyceride, microsomal protein and cytochromes, and on the activities of drug metabolizing enzymes.2. In the repeated oral dose of DBP (630 or 1260mg/kg/day) for 5 days, the ratio of liver weight to body weight was increased in both female and male rats, whereas the increases of cytochrome P-450 content and aniline hydroxylase activity were noted only in male rats. However, the contents of liver triglyceride, phospholipids, and cholesterol were unchanged. On the other hand, serum cholesterol content which showed the tendency to be decreased at the low dose was significantly decreased at the high dose.3. In the incorporation of 1-14C-acetate into liver and serum lipids after repeated oral dose of DBP (630mg/kg/day) for 5 days in male rats, the incorporation into triglyceride showed tendency to be increased, whereas the incorporation into cholesterol and cholesterol ester remained unchanged in vivo and in vitro.

Studies on Analysis of Mycotoxins (V)

Abstract: Systematic analysis for the simultaneous determination of fluorescent mycotoxins, aflatoxin B1, B2, G1, G2, sterigmatocystin, ochratoxin-A and citrinin from polluted grains was studied.The mycotoxins were extracted with the solvent consisted of 2ml of 20% sulfuric acid, 20ml of 4% potassium chloride solution and 178ml of acetonitrile. They were transfered to chloroform and freed from solvent. By means of silica gel column chromatography the former five toxins were cleaned-up and eluted by chloroform-methanol mixture (97:3) and the remaining ochratoxin-A and citrinin were recovered in the fraction eluted by benzene-acetone-acetic acid (75:20:5) after the intermediate washing of the column with chooroform-methanol (93:7).Each fraction was concentrated and applied to thin layer chromatography for final determination, in which the mutual separation of ochratoxin-A and citrinin was performed by employing DC-Fertigplatten Kieselgel for solid layer and chloroform-methanol-water-90% formic acid (90:10:1:1) as developing solvent.The fluorescent intensity of thin layer spots of sterigmatocystin, ochratoxin-A and citrinin were amplified by spraying 20% alminium trichloride solution, exposing in ammonia vaper, and spraying 20% alminium trichloride after exposure in ammonia vaper, respectively.

Ecological Survey on Bacterial Contamination of Bottled Milk

Abstract: Recently, remarkable progress has been made on the sanitary control of the milk plant, especially in the countermeasure against air microbes, by introducing such equipment as an air-conditioner. Then a survey was conducted to analyze the effect of such progress upon the aspect of bacterial contamination of bottled milk and the environment of the plant. Since a similar survey was carried out in 1964 before such progress was made, the present survey was performed almost under the same conditions as it in 1974. After that, results were compared between the two surveys with the following findings.1) When bottled milk was examined for bacterial contamination, psychrotrophic, mesophilic, or coliform bacteria were not detected from it immediately after bottling. When bottled milk was stored at 37°C for 24 hours or at 6°C for 7 days, the number of its samples showing a bacterial count exceeding 50, 000, or the lowest allowable limit, was much lower in the present survey than in the 1964 survey. The frequency of appearance of milk samples which failed to meet the minimum requirements after storage was higher in summer than in winter.2) When bottled milk samples were stored at 37°C in the winter of 1964, their bacterial flora contained various types of thermolabile bacteria simultaneously with Bacillus organisms. When the same samples were stored at 37°C in the winter of 1974, their bacterial flora was composed exclusively of Bacillus organisms, which were presumed to have withstood pasteurization and disinfection. When the same samples were stored at 37°C in the summer of 1974, their bacterial flora contained thermolabile bacteria in addition to Bacillus organisms.3) The count of falling bacteria and the bacterial count of finished products (after storage) were compared between the line with an air-conditioner introduced and that without such equipment introduced in the same plant conducting UHT pasteurization. As a result, much more satisfactory results were obtained from the former line.

Gaschromatographic Determination of Propionic Acid in Foods by Phenacyl Esterification

Abstract: In this paper a method of phenacyl esterification determination of propionic acid after steam distillation for foods was described.Para-chlorophenacyl bromide (PCPB) was used as a phenacylation agent. Outline of the procedure was as follows: The mixture of the distillate and dimethylformamide solution of PCPB was stood for 30 minutes in a boiling water bath. After cooling of phenacyl ester of propionic acid was extracted with n-hexane. The phenacyl ester of propionic acid was determined by a gas chromatograph equipped with electron capture detector. The glass column for analysis was 2m long and was packed with 5% OV-17 on gaschrom Q 80-100mesh. The column operated at 200°C.Because of including chlorine, the compound could be measured with high sensitivity by electron capture detector. The lower limit of identification of the concentration of propionic acid was 0.1ppm solution.

Gas Chromatography of p-Hydroxybenzoate Esters in Soy Sauce

Abstract: Trifluoroacetyl (TFA) derivatives of ethyl (PHBA-Et), n-propyl (PHBA-Pr), iso-propyl (PHBA-isoPr), n-butyl (PHBA-Bu) and iso-butyl p-hydroxybenzoate (PHBA-isoBu) in soy sauce were determined by gas-liquid chromatography equipped with hydrogen-flame ionization detector on 2% OV-1 column at 135°C.Determination was made successfully using benzophenone as the internal standard.These esters were separated by steam distillation from samples and extracted with ether.The recovery rates of PHBA-Et, PHBA-Pr, PHBA-isoPr, PHBA-Bu and PHBA-isoBu added to soy sauce were 61.3%, 79.0%, 91.2%, 92.3% and 94.9%, respectively.This method was applied to ten kinds of commercially available soy sauce, and PHBA-isoPr, PHBA-isoBu and PHBA-Bu were found in seven samples and only PHBA-Bu in three samples.

Studies on the Formation of Volatile Amines from Trimethylamine Oxide by γ-Irradiation (I)

Abstract: The simplified method for determination of ammonia and volatile amines, especially methylamines formed from trimethylamine oxide (TMAO) by γ irradiation, was established.The method involved the extraction procedure by means of Conway's micro diffusion apparatus and the determination by TLC-fluorodensitometry of DNS-derivatives as well as by gas-liquid chromatography.The satisfactory quantitative results were obtained using ammonia, monomethylamine and dimethylamine in the concentration of 1×10-7-3×10-6mol/ml and trimethylamine in the concentration of 8×10-8-1×10-5mol/ml.