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PubMedDate not supplied

Use of the Mycosep multifunctional cleanup column for liquid chromatographic determination of aflatoxins in agricultural products.

A liquid chromatographic (LC) technique has been developed that uses the Mycosep multifunctional cleanup (MFC) column. MFC columns provide a rapid 1-step extract purification. They are designed to retain particular groups of compounds that may create interferences in analytical methods. At the same time, MFC columns allow compounds of interest to pass through. In the method presented, test samples are extracted in a blender with acetonitrile-water (9 + 1). A portion of the extract is forced through an MFC column designed especially for analysis of numerous mycotoxins. Analytical interferences are retained, while aflatoxins pass through the column. Aflatoxins B1 and G1 are converted to their hemiacetals by heating a mixture of purified extract and water-trifluoroacetic acid-acetic acid (7 + 2 + 1) at 65 degrees C for 8.5 min. An aliquot of this mixture is analyzed by isocratic LC with acetonitrile-water mobile phase and fluorescence detection. A detection limit of less than 0.5 ng/g for aflatoxin B1 was obtained. Average recoveries greater than 95% total aflatoxins (B1, B2, G1, and G2) and coefficients of variation of less than 3% were obtained. The method was successfully applied to the following commodities: corn, almonds, pista-chios, walnuts, peanuts, Brazil nuts, milo, rice, cottonseed, corn meal, corn gluten meal, fig paste, and mixed feeds.

Aflatoxins↗

PubMed1991

[A screening method for simultaneous immunochemical detection of aflatoxins and ochratoxin A in cereals and feed].

An immunochemical method was proposed for simultaneous determination of aflatoxin and ochratoxin A with the use of mixed solutions of the following reagents: standards of both mycotoxins, antiserums against the mycotoxins, and radioligands of 125I-aflatoxin B1 and 125I-ochratoxin A. The result of the analytical procedure is the value of concentration of the aflatoxin + ochratoxin A sum in the sample. The procedure needs half the amount of reagents as separate determination of each of the two mycotoxin, and is far less laborious. The proposed simultaneous immunoanalysis is suitable for large-are inspection of grain and feed safety from the viewpoint of aflatoxin and ochratoxin A levels.

Aflatoxins↗

PubMedDate not supplied

Liquid chromatographic determination of aflatoxin levels in peanut butters using an immunoaffinity column cleanup method: international collaborative trial.

Thirteen laboratories in 7 different countries participated in a collaborative trial to evaluate the immunoaffinity column cleanup procedure with quantitation by fluorescence liquid chromatography (post-column derivatization) for the determination of aflatoxins in peanut butters. Participants were sent 10 randomly numbered samples of roasted peanut butter for analysis (5 pairs of undisclosed duplicates). Two of the pairs were "blank" peanut butters to which aflatoxin standards had been added; these "spiked" samples were used for recovery purposes. The other 3 pairs of samples were a nominal "blank" and 2 naturally contaminated peanut butters. A full statistical presentation of the results is given. Coefficients of variation (CVs) for the total aflatoxin determinations for mean levels of 4, 15, and 38 microns/kg were between 32 and 44% for the blank and 2 trial samples. Recovery levels for the 2 spiked samples were 51-67%, with aflatoxin B1 recovery of 60%. Relative standard deviations for method repeatability (RSDr) and reproductibility (RSDR) for the 3 trial samples were 15-26% and 33-45%, respectively.

Aflatoxins↗

PubMed1990

Nonassociation of aflatoxin with primary liver cancer in a cross-sectional ecological survey in the People's Republic of China.

A comprehensive cross-sectional survey was undertaken in The People's Republic of China of possible risk factors for primary liver cancer (PLC) to include 48 survey sites, an approximately 600-fold aflatoxin exposure range, a 39-fold range of PLC mortality rates, a 28-fold range of hepatitis B virus surface antigen (HBsAg+) carrier prevalence, and estimation of exposures for a large number of other nutritional, dietary, and life-style features. PLC mortality was unrelated to aflatoxin intake (r = -0.17) but was positively correlated with HBsAg+ prevalence (P less than 0.001), plasma cholesterol (P less than 0.01), frequency of liquor consumption (P less than 0.01), and mean daily intake of cadmium from foods of plant origin (P less than 0.01). Multiple regression analyses for various combinations of risk factors showed that aflatoxin exposure consistently remained unassociated with PLC mortality regardless of variable adjustment. In contrast, associations of PLC mortality with HBsAg+, plasma cholesterol, and cadmium intake remained, regardless of model specification, while the association with liquor consumption was markedly attenuated (was nonsignificant) with adjustment for plasma cholesterol. The sharp contrast between the findings of no aflatoxin effect upon PLC prevalence in this survey and the positive correlation reported for previous but more restricted surveys is discussed. Based on the results of this survey and the data of laboratory animal and in vitro studies, an explanatory model for the etiology of PLC is proposed, taking into consideration the role of nutrition in the etiology of this disease.

Aflatoxins↗

PubMedDate not supplied

Comparative evaluation of commercially available aflatoxin test methods.

Five qualitative methods and 1 quantitative aflatoxin analytical method were compared with the Holaday-Velasco (HV) minicolumn and thin-layer chromatography (TLC) methods for corn in an evaluation involving 4 U.S. Department of Agriculture Federal Grain Inspection Service (USDA-FGIS) laboratories, 1 laboratory at the University of Georgia, and 1 laboratory at the University of Arizona. Samples analyzed included 1 set of artificially contaminated corn containing both aflatoxin B1 and B2 (ratio of B1:B2 of 92:8), 1 set of artificially contaminated corn containing only aflatoxin B1, and 1 set of naturally contaminated corn. Levels of total aflatoxin tested were 0, 10, 15, 20, 25, 30, and 40 ppb. Results of analysis of these samples with each method evaluated are reported. Chi-square analyses indicated that performance of the Afla-20-Cup, Aflatest, EZ-Screen, OXOID, and SAM-A methods was not statistically different from that of the HV minicolumn. Agri-Screen results were not statistically different from those obtained with TLC.

Aflatoxins↗

PubMedDate not supplied

Survey of aflatoxin content in Swiss concentrate feeds for dairy cattle.

Within the context of restrictive Federal measures, the Swiss concentrate feeds for dairy cattle have been periodically surveyed and analyzed for aflatoxin content. In these commercial feeds, the average aflatoxin B1 content decreased sharply from 47 micrograms/kg in the winter of 1976-1977 (period prior to restrictive measures) to 24 micrograms/kg in the winter of 1977-1978 and to 3 micrograms/kg in the winter of 1978-1979. Work made public by the Federal Office of Public Health, reveals a corresponding downward trend for aflatoxin M1 concentration levels in milk from 109 ng/kg in the winter of 1976-1977 to 44 ng/kg in the winter of 1977-1978 and to 10 ng/kg in the winter of 1978-1979. This record indicates that the feed control measures taken were effective in reducing milk contamination by aflatoxins.

Aflatoxins↗

PubMed1978

Effect of aflatoxin on the humoral and cell-mediated immune systems of the chicken.

Chickens fed 2.5 microgram of aflatoxin/g of diet from 2 to 4 weeks of age or from hatching to 4 weeks were deficient in cell-mediated immunity, as measured at 4 weeks of age by the graft-versus-host reaction. Delayed-type hypersensitive skin reactions to tuberculin were also reduced in chickens given dietary aflatoxin from hatching to 7 weeks of age. Humoral immunity, as measured by the ability of 4-week-old chicks to produce natural agglutinins to rabbit red blood cells, was not significantly altered by dietary aflatoxin. A significant decrease in concentrations of serum immunoglobulins (Ig) IgG and IgA, but no IgM, however, did occur in chicks given dietary aflatoxin from hatching to 4 weeks or between 2 and 4 weeks of age. Aflatoxin consumption from 0 to 2 weeks of age produced no marked effect on either cell-mediated or humoral immunity in 4-week-old chicks.

Aflatoxins↗

PubMedDate not supplied

[Presence of aflatoxin in products intended for human consumption: the choice of methods and the results obtained].

The contamination of food resources by aflatoxin is a major problem for those countries where the meteorological and sanitary conditions allow the growth of Aspergillus Flavus and the contaminated food is not discarded because of the critical economical and social situation. However this phenomenon is also recorded in those Nations where the environmental and meteorological conditions are not favourable to Aspergillus Flavus growth. In Europe it has been possible to measure low aflatoxin (M1) concentrations in milk destined to humans. This kind of contamination may represent a source of chronic exposure to an important risk factor for hepatocancerogenesis. In Italy an appropriate law regulating the maximum aflatoxin concentration in the food destined to human nutrition is lacking, therefore the purpose of this paper is to contribute to the knowledge of a possible human exposure from the milk consumed in the Turin area (north western Italy). The results obtained display a lower (but more diffused) contamination in milk sampled after processing treatments (1.9-15 ppt), than in raw milk (5.3-94 ppt). Moreover it has been possible to show a positive relationship between dry animal feed and aflatoxin concentrations in milk.

Aflatoxins↗

PubMedDate not supplied

Comparison of two ELISA screening tests with liquid chromatography for determination of aflatoxins in raw peanuts.

A study was conducted to evaluate the performance of 2 enzyme-linked immunosorbent assays (ELISA) for rapidly screening samples of peanuts for the presence of aflatoxin. The EZ-Screen Quick Card Test and the Afla-10 Cup Test were compared with liquid chromatography in duplicate analyses of common extracts of peanuts contaminated in the range of 0-70 ppb (ng/g). Each assay properly identified 95% of samples containing no detectable aflatoxin as negative and greater than 97% of samples containing greater than 10 ppb aflatoxin as positive. The card test, which had a 20 ppb detection threshold, identified as positive 32 of 34 samples in the 11-20 ppb range. This indicates that the card test might actually have a detection threshold closer to 10 ppb. Most of the errors associated with the assays occurred on samples containing less than 10 ppb aflatoxin. The cup and card tests identified 76 and 67% of the samples, respectively, as negative, in the range of 4-10 ppb. For samples either negative or contaminated above their detection thresholds for the assays, the methods are well suited for use as rapid screening tests.

Aflatoxins↗

PubMed1989

Complement, bacteriostatic, and enzymatic activities in sera from guinea pigs given aflatoxin and/or rubratoxin.

The relationship of serum complement activity and bacteriostatic activity was investigated in male guinea pigs given aflatoxin and/or rubratoxin. In experiment 1, guinea pigs were given 0.6 mg of aflatoxin/kg of body weight, PO, once. In experiment 2, guinea pigs were given 0.02 mg of aflatoxin/kg, PO, and/or 8 mg of rubratoxin, PO, 11 times. Aflatoxin (0.02 mg/kg) had no effect given alone, but potentiated the effect of rubratoxin. In both experiments, changes in complement activity were accompanied by similar but not always significant (P less than 0.05) changes in bacteriostatic activity of serum. Guinea pigs given 0.06 mg of aflatoxin/kg had significant (P less than 0.05) changes in complement titers and in serum alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase activities. Guinea pigs given repeated oral doses of aflatoxin and/or rubratoxin had changes in complement titers, bacteriostasis, and alkaline phosphatase and aspartate aminotransferase activities, but not in alanine aminotransferase activities. Significant differences were detected only when average values for all guinea pigs given rubratoxin or rubratoxin with aflatoxin were compared with average values for guinea pigs not given rubratoxin.

Aflatoxins↗

PubMed1978

Reduced nicotinamide adenine dinucleotide phosphate-dependent formation of 2,3-dihydro-2,3-dihydroxyaflatoxin B1 from aflatoxin B1 by hepatic microsomes.

2,3-Dihydro-2,3-dihydroxyaflatoxin B1 (dihydrodiol) was formed as a major metabolite in the incubation of aflatoxin B1 with rat and hamster liver microsomes. The yield of the dihydrodiol was maximal at pH 6.5, was reduced nicotinmide adenine dinucleotide phosphate- and cytochrome P-450-dependent, and was increased 2- to 4-fold by pretreatment of the animals with phenobarbital; pretreatment with 3-methylcholanthrene did not alter the activity of rat hepatic microsomes. Inhibitors of epoxide hydrase did not lower the yield of the dihydrodiol in these systems. Negligible yields of the dihydrodiol were formed from aflatoxin B1 and rat liver microsomes in the presence of DNA. Little or no formation of the dihydrodiol was noted with microsomes from rat intestinal mucosa, kidney, or lung. These results further support the formation of aflatoxin B1 2,3-oxide as a major electrophilic metabolite of aflatoxin B1 in rat and hamster liver microsomal systems, since this highly reactive epoxide would be expected to hydrolyze readily to form the dihydrodiol.

Aflatoxins↗

PubMed1985

Measurement of individual aflatoxin exposure among people having different risk to primary hepatocellular carcinoma.

The establishment of the carcinogenic role of aflatoxins has been impeded by the lack of suitable tests to measure individual exposure for long-term studies. It was shown that the use of immuno-concentration followed by high pressure liquid chromatography (HPLC) or immunoassay can regularly detect aflatoxins down to pg/ml in fluids including urine. Aflatoxin M1 (AFM1) in free form was identified as the major metabolite in urine suitable for use as an approximate dosimetric indicator of recent exposure to aflatoxin B1 (AFB1). A panel of monoclonal antibodies of IgG class were developed in the murine system against AFB1 and/or AFM1. Their affinity constants are at the level of 10(8)-10(9) L/mol, and they are suitable for use in either effective immuno-concentration or in high sensitivity immunoassays. It was shown in a high risk area of liver cancer (Qidong of China) that 10% or more of the local inhabitants had a urinary output of AFM1 one or two orders of magnitude higher than that of Beijing people, especially during the wet seasons. The ingestion of AFB1 among these local people was estimated to exceed 1 mg/year. The major source came from contaminated corn and rice, but local alcoholic beverages also contributed. The increased excretion of AFM1 was more pronounced in patients with chronic active hepatitis. This observation of very low AFM1 content in urine of a significant percentage of local people implies their food storage practices may offer a feasible method for such prevention. The value and possible problems in conducting long-term studies on primary prevention are discussed.

Aflatoxins↗

PubMedDate not supplied

Liquid chromatographic determination of aflatoxins in feedstuffs containing citrus pulp.

A liquid chromatographic (LC) method was developed for the determination of aflatoxins in feedstuffs containing citrus pulp. The feed-stuff sample is extracted with chloroform, followed by Sep-Pak Florisil cartridge cleanup and Sep-Pak C18 cartridge cleanup. The final eluate (water-acetone, 85 + 15, v/v) is submitted to reverse-phase liquid chromatography with water-methanol-acetonitrile (130 + 70 + 40, v/v/v) as mobile phase and postcolumn derivatization with iodine. Citrus components are removed from the extract efficiently. The limit of detection for aflatoxin B1 is less than 1 microgram/kg. Other aflatoxins can also be detected and measured. Recoveries of aflatoxins B1, B2, G1, and G2 for dairy rations spiked at 13, 5, 10, and 4 micrograms/kg were 87, 86, 81, and 82%, respectively. Corresponding coefficients of variation were 3.1, 3.6, 5.2, and 3.8%, respectively.

Aflatoxins↗

PubMed1977

In vitro stimulation of bovine peripheral blood lymphocytes: suppression of phytomitogen and specific antigen lymphocyte responses by aflatoxin.

The effects of aflatoxin B1 on responses of the peripheral blood lymphocytes (PBL) of 2 normal animals to phytohemagglutinin, concanavalin-A, and pokeweed mitogen and of the PBL of 2 Mycobacterium bovis-infected animals to phytohemagglutinin and purified protein derivative of M bovis (PPD) were studied. Aflatoxin concentrations of greater than or equal to 10 microgram/ml significantly suppressed the lymphocyte response of normal animals to the phytomitogens. Lymphocyte response of M bovis-infected animals to specific antigen PPD was significantly suppressed at aflatoxin concentration of 0.5 microgram/ml. Fifty- to 100-fold higher concentrations of aflatoxin were required to produce 50% suppression of lymphocyte response to phytomitogens, as compared with that produced to PPD.

Aflatoxins↗

PubMed1977

Effects of aflatoxin on the cellular immune system.

Observations of aflatoxin induced thymic involution were followed by studies of other parameters of cellular immunity. Guinea pigs treated with 0.03 to 0.35 mg aflatoxin B1 for 3 to 5 weeks were tested for the development of cutaneous hypersensitivity, migration inhibition factor activity, and stimulation of lymphocyte transformation. A significant suppression of cutaneous hypersensitivity to a Nocardia asteroides sensitin in sensitized aflatoxin recipients was demonstrated. In vitro reductions in migration inhibition factor (MIF) and depressed stimulation indices (Phytohemagglutinin) of peripheral blood lymphocytes were also demonstrated in aflatoxin recipients but did not reach significant levels.

Aflatoxins↗

PubMedDate not supplied

Optimum methanol concentration and solvent/peanut ratio for extraction of aflatoxin from raw peanuts by modified AOAC method II.

The amount of aflatoxin extracted from raw peanuts by using the water-slurry modification of AOAC Method II was determined for 49 different combinations of methanol concentrations and solvent/peanut ratio. Results indicate that the amount of aflatoxins B1 and B2 extracted from raw peanuts is a function of both methanol concentration and solvent/peanut ratio, and a cubic equation was developed, using regression techniques, to describe the combined effects. From the functional relationship, the predicted methanol concentration and solvent/peanut ratio that extracts the most aflatoxin B1 was computed to be 60.0% and 10.8 mL solvent/g peanuts, respectively. This combination extracted 12.1% more aflatoxin than did AOAC Method II.

Aflatoxins↗

PubMedDate not supplied

Ability of ethoxyquin and butylated hydroxytoluene to counteract deleterious effects of dietary aflatoxin in chicks.

The antioxidants ethoxyquin and butylated hydroxytoluene (BHT) were added to diets of chicks in concentrations three and eight times above that usually found in poultry feed beginning 15 days after hatch; the chicks had been placed on feed containing 1000 or 3000 ppb aflatoxin on the day of hatch. These diets were continued until chicks were 6 weeks of age. At that time, deleterious effects of aflatoxin on weight gain, feed efficiency, and organ weights (spleen, bursa) were evident. BHT alleviated these effects, but ethoxyquin did not. Pretreatment with ethoxyquin did not protect chicks either. Ethoxyquin was not able to induce the activities of chick liver enzymes that detoxify aflatoxin and other foreign compounds. Lack of effect of ethoxyquin on these enzymes may hinder ability of this antioxidant to protect chicks from aflatoxin.

Aflatoxins↗

PubMed1977

Species comparison of in vitro metabolism of aflatoxin B1.

The metabolism of [14C]aflatoxin B1 by 9000 X g supernatant fraction of livers of duck, rat, mouse, monkey, and humans was compared by incubating the compound and liver fractions in the presence of cofactors and air for 30 min. The incubation medium was extracted with chloroform, and the soluble metabolites were separated on thinlayer plates and quantified; radioactivity remaining in the aqueous phase was determined in order to quantify metabolism to water-soluble derivatives. Duck, monkey, and human livers were most active in total conversion, each metabolizing approximately 80% of available substrate in 30 min. Rat and mouse livers had lower activites, metabolizing 15 to 20%. Duck liver produced mainly (60%) chloroform-insoluble derivatives, but all other species produced larger quantities of chloroform-soluble than insoluble metabolites. Aflatoxin Q1 was the principle chloroform-soluble metabolite produced by monkey, human, and rat liver, whereas duck liver produced mainly aflatoxicol in that fraction. Aflatoxin P1 was produced by monkey, human, and mouse liver, but not by duck and rat. The chromatographic region containing M1 and B2alpha contained low levels of radioactivity in all species except human. No consistent pattern of metabolism emerged which could be correlated with species differences in response to aflatoxin B1 toxicity or carcinogenicity.

Adult↗