Method For Detection Of Aflatoxins Research Articles

Determination of aflatoxin biomarkers in excreta and ileal content of chickens

Aflatoxins are carcinogenic secondary metabolites frequently detected in food and feed stuff based on maize and other crops susceptible to infection with the fungal pathogen Aspergillus flavus. We investigated the metabolization of aflatoxins in chickens by analyzing excreta and ileal content and developed and validated a biomarker method for detection of aflatoxins and their metabolites in these matrices. Analysis of ileal content served to distinguish between urinary and fecal excretion combined in the excreta samples. During a 3-wk animal trial, one hundred sixty-eight 1-day-old chicks were randomly allocated to 24 pens with 7 chicks per pen and subjected to different feed regimens with: A) toxin-free feed, B) feed supplemented with 18ng of total aflatoxins/g, and C) feed supplemented with 515ng of total aflatoxins/g. Chicken excreta and ileal content were sampled after 7, 14, and 21 D. An analytical method based on liquid chromatography coupled to tandem mass spectrometry was validated for the determination of aflatoxin B1, B2, G1, G2, M1, P1, Q1, and aflatoxin B1-N7-guanine (AFB1-N7-Gua) in chicken's samples. Comparing chicken excreta, which contain urine and feces, to ileal content, which contains no urine, we explored the secretion pathway of aflatoxin metabolites. The AFB1-N7-Gua was only detected in excreta, whereas aflatoxin M1 (AFM1) was detected both in ileal content and excreta. Aflatoxin M1 was detected in excreta in concentrations 5 times higher than in ileal content, suggesting primary excretion via urine. Although chickens are relatively resistant to aflatoxins, contamination of feed can lead to adverse effects and thus economic losses in farming. Therefore, a biomarker method to estimate the exposure of chickens to aflatoxins can play an important role to monitor the animals' health.

Potential of near infrared spectroscopy as a rapid method to detect aflatoxins in brown rice

Brown rice is a main popular health food with high nutritional value and health benefits. As a result of poor post-harvest drying and inappropriate storage conditions, rice grains are often damaged through fungal spoilage as well as mycotoxin production. The objective of this research was to evaluate the possibility of using the near infrared spectroscopy, with a wavenumber range between 12500 and 4000 cm−1 (800–2500 nm), as a rapid method for detection of aflatoxins in brown rice. Firstly, storage trials were carried out to generate representative of samples contaminated and non-contaminated with aflatoxins. These data were used to create a partial least squares regression model using 120 brown rice samples with the required near infrared spectral data and aflatoxin concentration levels that were determined using a standard enzyme-linked immunosorbent assays method. The accuracy of developed models was externally validated using the test set. The statistical model developed from the treated spectra (vector normalization; SNV) provided the best accuracy in prediction with a coefficient of determination of prediction ( r2) of 0.95, a root mean square error of prediction of 415.00 µg kg−1 and a bias −54.00 µg kg−1. The model developed showed good predictive performance which suggests that it could have practical applications as a rapid method to detect aflatoxins in brown rice.

Intra-laboratory Development and Evaluation of a Quantitative Method for Measurement of Aflatoxins B1, M1 and Q1 in Animal Urine by High Performance Liquid Chromatography with Fluorescence Detection.

Mycotoxins negatively impact animal health. Aflatoxins (AFs) are the most common mycotoxins affecting both large and small animals and are a common cause of toxin-related pet food recalls. Definitive diagnosis of aflatoxicosis is constrained by a lack of validated ante-mortem analytical methods for detection and quantitation of AFs and their metabolites in biological specimens. Herein, we developed and evaluated a urine-based quantitative method for measurement of aflatoxin B1 (AFB1) and its metabolites aflatoxin M1 (AFM1) and aflatoxin Q1 (AFQ1) in animal urine. (Some of the results have been presented at 59th AAVLD conference, Greensboro, North Carolina, October 13-19th, 2016.) This method uses an immuno-affinity column for clean-up and pre-column derivatization followed by high performance liquid chromatography analysis with fluorescence detection. The method has high selectivity, recovery (>81%) and sensitivity with an instrument limit of detection of 0.20-1.02 pg; instrument limit of quantitation of 0.77-4.46 pg; and a method lower limit of quantitation of 0.30-2.5 ng/mL. The method has high accuracy, repeatability, and is rugged against minor changes. However, because of poor sensitivity of AFQ1 at low concentrations we recommend this method for quantitative determination of AFB1 and AFM1, and for qualitative measurement of AFQ1 in animal urine for diagnosis of aflatoxicosis.

Antibody-free colorimetric determination of total aflatoxins by mercury(II)-mediated aggregation of lysine-functionalized gold nanoparticles

Currently known immunoassays for aflatoxins (AFs) usually are not capable of simultaneous determination of total AFs (i.e., aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2) due to the lack of group-specific antibodies. We are presenting here a colorimetric method for determination of total AFs by exploiting the Hg(II) ion-mediated aggregation of lysine-functionalized gold nanoparticles (Lys-AuNPs). AFs bind Hg(II) ion to form stable complexes, and this prevents the Hg(II) induced aggregation of Lys-AuNPs. Hence, the color change of solutions of AuNPs that occurs after aggregation of the AuNPs is suppressed. This effect was exploited to design a method for quantitation of AFs that can be detected by UV-Vis spectrophotometry (by measuring the ratio of absorbances at 725 nm and 525 nm), and even with bare eyes. The method has a very low detection limit (1.1 ppb) and is highly selective for AFs over other mycotoxins. Colorimetric analysis of rice samples by this method gave results that were in good agreement with those obtained by HPLC. To the best of our knowledge, this is the first method for visual detection of total AFs based on the distance-dependent optical properties of AuNPs.

Methods for Detection of Aflatoxins in Agricultural Food Crops

Aflatoxins are toxic carcinogenic secondary metabolites produced predominantly by two fungal species: Aspergillus flavus and Aspergillus parasiticus. These fungal species are contaminants of foodstuff as well as feeds and are responsible for aflatoxin contamination of these agro products. The toxicity and potency of aflatoxins make them the primary health hazard as well as responsible for losses associated with contaminations of processed foods and feeds. Determination of aflatoxins concentration in food stuff and feeds is thus very important. However, due to their low concentration in foods and feedstuff, analytical methods for detection and quantification of aflatoxins have to be specific, sensitive, and simple to carry out. Several methods including thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), mass spectroscopy, enzyme-linked immune-sorbent assay (ELISA), and electrochemical immunosensor, among others, have been described for detecting and quantifying aflatoxins in foods. Each of these methods has advantages and limitations in aflatoxins analysis. This review critically examines each of the methods used for detection of aflatoxins in foodstuff, highlighting the advantages and limitations of each method. Finally, a way forward for overcoming such obstacles is suggested.

An empirical evaluation of three vibrational spectroscopic methods for detection of aflatoxins in maize

Three commercially available vibrational spectroscopic techniques, including Raman, Fourier transform near infrared reflectance (FT-NIR), and Fourier transform infrared (FTIR) were evaluated to help users determine the spectroscopic method best suitable for aflatoxin analysis in maize (Zea mays L.) grain based on their relative efficiency and predictive ability. Spectral differences of Raman and FTIR spectra were more marked and pronounced among aflatoxin contamination groups than those of FT-NIR spectra. From the observations and findings in our current and previous studies, Raman and FTIR spectroscopic methods are superior to FT-NIR method in terms of predictive power and model performance for aflatoxin analysis and they are equally effective and accurate in predicting aflatoxin concentration in maize. The present study is considered as the first attempt to assess how spectroscopic techniques with different physical processes can influence and improve accuracy and reliability for rapid screening of aflatoxin contaminated maize samples.

Determination of aflatoxins in animal feeds by liquid chromatography/tandem mass spectrometry with isotope dilution

The objective of the present study is to develop a simple, fast method for detection of aflatoxins in animal feeds. Simultaneous quantitation of four aflatoxins (AFB(1), AFB(2), AFG(1) and AFG(2)) in animal feeds was achieved in a single liquid chromatography/tandem mass spectrometry (LC/MS/MS) run. The solid-phase extraction cleanup step is eliminated with the stable isotope dilution method. Matrix effects were observed and overcome by isotope dilution. The method was tested in a variety of animal feed matrices and proved to be accurate and reliable. Method ruggedness tests resulted in recoveries of 78% to 122% with an intra-day assay precision of 2% to 15% and an inter-day assay precision of 3% to 17%. These results indicate that this method is suitable for quantitation of aflatoxins in animal feeds.

THE EFFECT OF DIFFERENT STABILIZERS ON STABILITY OF HORSERADISH PEROXIDASE- BOVINE SERUM ALBUMIN-AFLATOXIN B1, A CONJUGATED TRACER FOR DETECTION OF AFLATOXIN B1 IN IMMUNOASSAY-BASED METHODS

Aflatoxins are a group of fungal toxic metabolites, which are contaminated certain food commodities. ELISA is one of the sensitive methods for detection of aflatoxins. Preparation and stabilizing of a proper conjugated tracer for detection of aflatoxins is probably the main step for designing an ELISA method. In current study, different stabilizers were applied to stabilize a newly prepared conjugated molecule, Horseradish peroxidase-Bovine serum albumin- Aflatoxin B1 (HRP-BSA-AFB1). Stabilizing effects of six different stabilizers were compared during 10 months verification at room temperature and 6 weeks verification at 37°C. Based on the results, it was concluded that trehalose-containing stabilizers especially those contain casein in their compositions show the best stabilizing effects on HRP-BSA-AFB1 conjugated tracer.

Rapid Detection Of Aflatoxins By Pressure Minicolumn Technique

Abstract A simple, rapid, reliable method for detection of aflatoxins in food and feeds by Pressure Minicolumn has been developed. The technique employs commonly available glass syringes which are tightly packed with certain adsorbants. The presence of aflatoxins is observed as a compact bluish fluorescent hand under long range UV light. Further, chemical confirmatory tests have also been incorporated in the same and the combination of trifluoroacetic acid-sulphuric acid was found to be the best. The technique has been successfully put to use for detecting contaminated samples of food and feed.

Development of a sensitive high-performance liquid chromatographic method for detection of aflatoxins in pistachio nuts

A very sensitive method is described for the detection of aflatoxins in pistachio nuts. The method employs the extraction procedure used in the BF method (described in the Association of Official Analytical Chemist' Book of Methods) followed by an acid alumina column clean-up to remove sample components which would otherwise interfere in the final determinative step. The quantitation of the afiatoxins is carried out by high-performance liquid chromatography on a C 18 reversed-phase column after conversion of aflatoxins B 1 and G 1 to their more fluorescent derivatives, B 2a and G 2a. The limit of detection was found to be 0.5 μ/kg. The chromatograms were found to be free of interfering peaks and multiple injections showed excellent reproducibility.

A rapid method for detection of aflatoxins from xylitol

A rapid method for detection of aflatoxins from xylitol