Aflatoxins and ochratoxin A in dried eggplant and green bell pepper

In this study, 50 dried eggplant and 50 dried green bell pepper samples were analyzed in terms of their aflatoxin and ochratoxin A (OTA) content. Aflatoxins G₂, G₁, B₂, and B₁, and OTA contents were analyzed using high-performance liquid chromatography with a flame ionization detector (HPLC–FID). Total aflatoxin and, as well as aflatoxin G₂, G₁, B₂, and B₁ content in dried eggplant samples were ranged between 0.82 and 2.58, 0.10–0.23, 0.32–1.35, 0.12–0.67, and 0.17–0.71 μg kg⁻¹, respectively. Total aflatoxin and, as well as aflatoxin G₂, G₁, B₂ and B₁ content in dried green bell pepper samples were 0.81–2.42, 0.11–0.22, 0.32–1.38, 0.13–0.66, and 0.18–0.91 μg kg⁻¹, respectively. OTA content was varied from 8.88 to 21.35 μg kg⁻¹ in eggplant samples, and from 15.38 to 24.70 μg kg⁻¹ in dried green bell pepper samples. Of the dried eggplant samples and dried green bell pepper samples, 36% and 24% of them, respectively, had aflatoxin B₁ values which were below the minimum limit of detection (LOD) of 0.05 μg kg⁻¹. None of the analyzed samples exceeded the legal limit values of 10 μg kg⁻¹ for total aflatoxin content, and 5 μg kg⁻¹ for aflatoxin B₁ content. However, 80% of the dried eggplant samples and 100% of the dried green bell pepper samples exceeded the legal limit value for OTA content (15 μg kg⁻¹). According to the results, it was concluded that dried vegetables should be examined in terms of their aflatoxins. It is essential to analyze OTA content more thoroughly, as it has the potential to pose a risk for public health, as well as for the economy.     

Determination of aflatoxins in walnut sujuk and Turkish delight by HPLC-FLD method

This study aims to assess the risk of aflatoxins (AFs) in traditional confectionery products (walnut sujuk and Turkish delight) of Turkey. A high performance liquid chromatography coupled with fluorescence detection (HPLC-FLD) method was used for the determination of AFs. Evaluation of the method showed good selectivity, linearity, recovery and precision. The limit of quantification (LOQ) ranged from 0.106 to 0.374 μg kg⁻¹. The expanded measurement uncertainty was less than 40% for all target analytes. The validated method was successfully applied to the determination of AFs in 112 traditional confectionery products containing nuts (hazelnuts and walnuts). AFs were detected in 43.8% of walnuts and 60.9% of hazelnuts used as ingredients in walnut sujuk and Turkish delight and at levels ranging from 0.58 to 15.2 μg kg⁻¹ and 0.43–63.4 μg kg⁻¹, respectively. This means that AFs levels in walnut sujuk and Turkish delight were up to levels of 6.1 and 9.5 μg kg⁻¹, respectively. Six walnut samples and twenty-one hazelnut samples were above the EU maximum limits (MLs) of 2 and 5 μg kg⁻¹ for aflatoxin B₁ (AFB₁), respectively.     

A survey on the occurrence of aflatoxin M1 in raw milk produced in Adana province of Turkey

During 2012, a total of 176 samples of raw milk obtained from dairy plants of Adana province of Turkey were analysed for the presence of aflatoxin M₁ (AFM₁). Aflatoxin M₁ analysis was carried out by centrifugation, liquid–liquid extraction, immunoaffinity column clean-up and high performance liquid chromatography with fluorescence detection (HPLC-FD). The limits of detection (LOD) and quantification (LOQ) of the analytical method were 0.021 μg kg⁻¹ and 0.025 μg kg⁻¹. Accuracy of the method obtained from bias ranged from 2.94 to 8.70. Aflatoxin M₁ was detected in 53 out of 176 samples analysed (30.1%). The ranges for positive samples were 0.042–0.552, 0.033–1.01, 0.047–0.150 and 0.025–0.102 μg kg⁻¹ in autumn, winter, spring and summer seasons, respectively. Thirty samples of raw milk (17%) were above the legal limits of Turkey and EU regulations.     

A reliable and sensitive time-resolved fluorescent immunochromatographic assay (TRFICA) for ochratoxin A in agro-products

Immunochromatographic assays (ICAs) are considered as a suitable diagnostic tool for the detection of mycotoxins. Mycotoxins and especially, ochratoxin A are analytes with more demanding sensitivity requirements. To enhance the sensitivity of current immunochromatographic assays for ochratoxin A (OTA), a novel sensitive ICA was developed in this study. In the assay, microspheres enclosing fluorescent europium (III) [Eu(III)] nanoparticles (EuNPs) were used as a label for OTA monoclonal antibody (OTA-mAb) conjugation. Accordingly, assay was called time-resolved fluorescent immunochromatographic assay (TRFICA). The test strip was composed of three parts: a sample pad, nitrocellulose membrane and an absorbent pad. As for detection, a proper concentration of conjugated microspheres was pipetted into the microtube and sample extract was added to it. Then the strip was inserted into the tube and the fluid flow along the strip. The TRFICA results were obtained in 8 min and read by a portable TRFICA strip reader. The established method allows quantitative determination of OTA with limit of detection as low as 1.0 μg kg⁻¹ in the samples. For validation, spiked samples including wheat, maize, soybean and rice were respectively assayed by TRFICA and a standard high performance liquid chromatography equipped with a fluorescence detector (HPLC-FLD), and good agreement of results was obtained between two methods.     

EU legislation on cereal safety: An update with a focus on mycotoxins

Cereals are still by far the world's most important source of food, both for direct human consumption and indirectly, as inputs to livestock production. FAO's latest forecast for world cereal production in 2011 stands at nearly 2313 million tonnes. Total EU-27 grain production forecast was 283 and 272 million tonnes for 2011 and 2012, respectively. Cereal contamination has an important impact on human and animal health. The European Union has established the most comprehensive regulations for food and cereal safety to facilitate world trade and protect consumer health. This paper reviews the existing legislation associated with cereal safety, with a focus on mycotoxin contamination. Regulations and Directives were classified into the following topics: general food legislative framework, official controls (sampling and analysis), maximum levels for contaminants, prevention and reduction. To give the reader a rapid first approach to the topic of his interest, a synoptical presentation of all laws related to the above-mentioned topics is given, and the main points of each law, cited in conjunction with its effect on previous laws (repeal, modification, amendments, replacement, related acts), are reported in tables. Moreover, data regarding the worldwide occurrence of mycotoxins in cereals were reported.     

Presence of mycotoxins in animal milk: A review

Mycotoxins can cause toxicity when ingested by humans and animals. Although the rumen is supposed to be a barrier against mycotoxins, some studies demonstrate that carry-over of mycotoxins to milk is possible. Different studies have found mycotoxin levels in animal milk, mainly related to contaminated feed for ruminants. Aflatoxin M1 is the most studied mycotoxin in milk and levels exceeding the EU maximum level for this mycotoxin in this matrix (0.050 μg/kg) have been found. Maximum levels in milk for other mycotoxins have not been established; however ochratoxin A, aflatoxins G1, G2, B1, B2 and M2, fumonisin B1, cyclopiazonic acid, zearalenone and its metabolites and deepoxy-deoxynivalenol have also been found in milk samples. Taking into account that multi-exposure to mycotoxins is the most likely scenario and co-occurrence of mycotoxins could affect their toxicological effects in humans and animals, there is a need to determine the co-occurrence of mycotoxins in milk.     

The effects of oxygenation aeration treatment on dissipation behaviors of tricaine mesylate in carp (Cyprinus carpio) muscle and water

Tricaine mesylate (MS-222) is one of the most used anesthetics in fish. It can be absorbed by the human body via food consumption, with related detriments to human health. In this study, a modified quick, easy, cheap, effective, rugged, and safe (QuEChERS) method was developed for the determination of MS-222 in carp muscle and water samples using liquid chromatography-tandem mass spectrometry (LC-MS/MS). For cleanup procedure, multiplug filtration cleanup (m-PFC) method with n-Hexane delipidation was adopted. The extraction solvent, cleanup methods and sorbents were optimized. All method validation parameters were in the acceptable range. The dissipation behavior study was followed by the method development. Firstly, the anesthesia dose and time were optimized in application study. Secondly, carps were revived for different period of time with (experimental group) and without (control group) the oxygenation aeration treatment to compare the dissipation rate of MS-222. After being anesthetized for 6 h at 50 mg/L and 12 h of elimination, the concentrations of MS-222 in crap muscle and water of experimental group was lower than those of control group. After 36 h of elimination under oxygenation aeration, over 90% of MS-222 was dissipated in carp muscle. The results showed that the half-life of MS-222 in carp muscle was 6.2 h. The findings suggested that the commonly-used oxygenation aeration treatment in aquaculture production had potential effects in accelerating the dissipation of MS-222 in carp and water. In this study, three days of withdrawal period was recommended in carps after MS-222 administration under oxygenation aeration.     

Determination of water-soluble vitamins in energy and sport drinks by micellar electrokinetic capillary chromatography

A method for the determination of water-soluble vitamins in several energy and sport drinks by micellar electrokinetic chromatography (MEKC) has been developed in this work. The separation of vitamins was studied in terms of background electrolyte composition (borate content, pH, surfactant type and content) and in other MEKC parameters. A study of the possible compounds found in the vitamin-enriched drinks that could interfere in vitamin determination was also performed, and a modified procedure with enhanced resolution was developed. The proposed method was successfully applied to the analysis of water-soluble vitamins in a variety of energy and sport drinks and also in fruit nectars. The method implies minimal sample preparation and reagent consumption, being environmentally sustainable. Thus, the proposed methodology could be useful for quality control purposes in the soft drink industry.     

Real-time pixel based early apple bruise detection using short wave infrared hyperspectral imaging in combination with calibration and glare correction techniques

High speed data processing for online food quality inspection using hyperspectral imaging (HSI) is challenging as over hundred spectral images have to be analyzed simultaneously. In this study, a real-time pixel based early apple bruise detection system based on HSI in the shortwave infrared (SWIR) range has been developed. This systems consists of a novel, homogeneous SWIR illumination unit and a line scan camera. The system performance was tested on Jonagold apples bruised less than two hours before scanning. Partial least squares-discriminant analysis was used to discriminate bruised pixel spectra from sound pixel spectra. As the glossiness of many fruit and vegetables limits the accuracy in the detection of defects, several reflectance calibrations and pre-processing techniques were compared for glare correction and maximizing the signal to noise ratio. With the best combination of first derivative and mean centering, followed by image post-processing, this system was able to detect fresh bruises in thirty apples with 98% accuracy at the pixel level with a processing time per apple below 200 ms.     

Aflatoxin B1 production by Aspergillus parasiticus and strains of Aspergillus section Nigri in currants of Greek origin

Aflatoxin B₁ (AFB₁) mostly produced by Aspergillus flavus and Aspergillus parasiticus, is an extremely toxic and carcinogenic metabolite. Currants are used in the Mediterranean diet as a food with antioxidant properties. Four strains of Aspergillus section Nigri have been isolated from currants originated from Crete and Corinth. In this study AFB₁ production by A. parasiticus and the four strains of Aspergillus section Nigri in Cretan and Corinthian currants (Vitis vinifera L.) is investigated. AFB₁ determination was performed by HPLC–FID. Results revealed that the four strains Aspergillus section Nigri, as well as the aflatoxigenic strain A. parasiticus produced AFB₁ (0.0052–1.31 μg AFB₁ 15 g⁻¹, corresponding to 0.0003–0.087 μg AFB₁ g⁻¹) in both type of currants (Cretan and Corinthian) on the 12th day of observation. Moreover, AFB₁ production, by A. parasiticus in the synthetic Yeast Extract Sucrose (YES) medium was also studied. The ability of AFB₁ production has been affected by the special characteristics of each isolate and the currants substrate.     

Thermal treatments and their effects on the fumonisin B1 level in rice

The objective of this study was to evaluate the effects of dry and hydrothermal treatment on FB₁ level in polished, parboiled and whole grain rice collected in Brazilian market. The effect of thermal treatment on FB₁ level was carry out by applying conventional cooking, autoclaving and dry heat treatment. Conventional hydrothermal treatment (cooking) reduced the initial natural contamination by 80%. However, no significant reduction was obtained by autoclaving. Dry heat treatment produced the reduction (70%), at temperatures range from 150 to 200 °C.     

Mass spectrometry quantification of beef and pork meat in highly processed food: Application on Bolognese sauce

Food frauds have become a very important issue in the field of food quality and safety. The risk of food adulteration is higher in highly processed food and mainly affects high added value foodstuff. The methods currently available to face this issue, PCR and ELISA, are very sensitive and specific, but they have some limitations. In the present work, tandem mass spectrometry is presented as an emerging approach to detect beef and pork meat in very complex and highly processed food matrices, such as Bolognese sauce, both in qualitative than in quantitative way. The detection is achieved using two different marker peptides, specific for beef and pork meat, both deriving from α2-collagen chain. Then, a calibration curve is set up using real sauces made by different percentages of pork and beef meat in a working range from 0 to 100%. The method here developed allows to quantify beef and pork meat in a complex product such as Bolognese sauce.     

Bioaccessibility and changes on cylindrospermopsin concentration in edible mussels with storage and processing time

The alkaloid cylindrospermopsin has been recognized of increased concern due to the global expansion of its main producer, Cylindrospermopsis raciborskii. Previous studies have shown that bivalves can accumulate high levels of cylindrospermopsin. Based on the potential for human health risks, a provisional tolerable daily intake of 0.03 μg/kg-body weight has been recommended. However, the human exposure assessment has been based on the cylindrospermopsin concentration in raw food items. Thus, this study aimed to assess the changes on cylindrospermopsin concentration in edible mussels with storage and processing time as well as cylindrospermopsin bioaccessibility. Mussels, (Mytilus galloprovincialis) fed cylindrospermopsin-producing C. raciborskii, were subjected to the treatments and then analyzed by LC-MS/MS. Mussels stored frozen allowed a significantly higher recovery of cylindrospermopsin (52.5% in 48 h and 57.7% in one week). The cooking treatments did not produce significant differences in cylindrospermopsin concentration in the mussel matrices (flesh), however, cylindrospermopsin was found in the cooking water, suggesting that heat processing can be used to reduce the availability of cylindrospermopsin. The in vitro digestion considerably decreased the cylindrospermopsin availability in uncooked and steamed mussels, highlighting the importance in integrating the bioaccessibility of cylindrospermopsinin in the human health risk assessment.     

Determination of methenamine residues in edible animal tissues by HPLC-MS/MS using a modified QuEChERS method: Validation and pilot survey in actual samples

A modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) method was developed for the determination of methenamine in edible animal tissues by high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). Samples were extracted with acetonitrile, and cleaned with anhydrous sodium sulfate and primary secondary amine (PSA) sorbent. An isotope dilution mass spectrometry technique was applied to compensate for matrix effect. The separation was performed on a HILIC column, and the mobile phase composed of acetonitrile-0.1% of formic acid and 5.0 mmol/L of ammonium acetate in water. The method showed a linear relationship in the range of 1.0–20.0 μg/L for methenamine, and the determination coefficients (R²) ranged from 0.9939 to 0.9995. The limit of detection (LOD) and quantification (LOQ) for methenamine in animal tissues sample was 1.5 μg/kg and 5.0 μg/kg, respectively. The average recoveries were 86.7–109.5% at spiked levels of 5.0, 25.0, 100.0 μg/kg. The intra-day precision ranged from 2.6 to 7.0%, and the inter-day precision ranged from 4.9 to 11.3%. The validated method was successfully applied to determination of methenamine in swinish muscle, kidney and liver.     

Development and in-house validation of a competitive ELISA for the quantitative detection of gluten in food

Wheat gluten contains peptide sequences, which activate specific T cells causing a chronic inflammation of the small intestine in celiac disease patients. It is well established that next to wheat gluten, the gluten-like proteins in barley and rye are similarly harmful to celiac disease patients whereas oat is generally considered safe. This study focuses on the development of an ELISA method for the detection of native and processed gluten proteins. The developed test utilizes a monoclonal antibody specific for the DQ2.5-glia-α3 T cell epitope present in the α-gliadins, which are part of wheat gluten that triggers celiac disease. The developed competitive ELISA uses a synthetic DQ2.5-glia-α3 peptide standard for calibration. The conversion from the measured DQ2.5-glia-α3 peptide concentration to gliadin content is achieved by using the experimentally determined multiplication factor of 250. The gluten content can be then calculated by multiplying the gliadin concentration by a factor of 2. A simple sample preparation method with 60% ethanol is used to extract the disease-causing proteins from cereals and processed foods. The assay was found to be specific for the detection of gluten from wheat, barley and rye with no cross-reaction with 8 tested oat varieties. The LOD and LOQ for gliadin were calculated based on the results obtained for 60 blank oats samples and they were 2.9 and 3.6 ppm, respectively. The assay could detect as little as 0.01% wheat gluten and gluten-like proteins from rye and barley in oats. The ELISA was also found to be applicable to the analysis of a range of processed food such as sauces, beers, soups and bread. In conclusion, the developed assay is a sensitive, specific and cost-effective tool for screening cereals and processed foods for the presence of harmful wheat gluten and gluten-like proteins from barley and rye.     

Development of an aptasensor for the fast detection of Versicolorin A

Aflatoxins (AFs) are secondary metabolites produced by Aspergillus flavus and Aspergillus parasiticus. The molds may contribute to pre-harvest aflatoxin contamination of susceptible crops. For the customer and food producer, a predictive model for aflatoxin detection is very desirable. Versicolorin A (VerA), which is the first precursor in the pathway of aflatoxin B₁ (AFB₁) biosynthesis, shares similar toxic group with the furofuran structure in aflatoxin B₁. VerA exhibits a much lower teratogenic toxicity than AFB₁ and may be used as a predictive indicator for aflatoxin B₁ contamination of storage crops. Therefore, the development of a fast detection method for VerA is important. One of the randomly computer-generated aptamers of VerA was confirmed by isothermal titration calorimetry with K_d = 9.26 × 10⁻⁶ mol l⁻¹. In addition, a simple and sensitive label-free aptasensor was developed for the electrochemical detection of VerA. According to the results from differential pulse voltammetry (DPV), a linear relationship existed between the log conc. of VerA (ranged from 0.01 to 100 ng ml⁻¹) and the current (△I_p) with a limit of detection (LOD) of 10 pg ml⁻¹. The resulting aptasensor exhibited good reproducibility for detecting VerA and stability after storage for 15 days at 4 °C with acceptable anti-interference against ZEN, OTA, DON, and FB₁. When used in corn samples, the recoveries of VerA were determined to be in the range of 81.3%–104.4 %. Although with some intercross, result suggests that the obtained aptamer for VerA is potentially used as a sorbent for the preparation of solid-phase-extraction procedure to clean up food samples in conjunction with high-performance liquid chromatography analysis.     

Species determination of pine nuts in commercial samples causing pine nut syndrome

Consumption of pine nuts from the species of Pinus armandii has been reported to cause dysgeusia, commonly known as pine mouth, or pine nut syndrome (PNS). However, the number of reports on pine nut consumptions of the different species and PNS is limited. This leaves open the possibility that other pine species than P. armandii could be involved in PNS as well. This study investigated 18 samples involved in PNS and received at the Danish Veterinary and Food Administration in 2011 through 2012. Samples were subjected to gas chromatographic analysis of fatty acids. The content of 11 individual fatty acids was used together with the diagnostic index and the sum of Δ5-fatty acids as diagnostic parameters. Diagnostic parameters from samples were then compared to reference material and literature data to determine the species. In a limited number of samples, the diagnostic parameters matched neither our reference materials nor literature data. However, the morphology, the fatty acid analysis, and externally obtained DNA sequencing data suggest a P. armandii subspecies or a variety. With these possible P. armandii subspecies, P. armandii was identified in all analyzed samples. The application of principal component analysis (PCA) to the data set showed a satisfactory separation of the majority of the 13 pine species included in the study.     

Prevalence and antimicrobial resistance of Salmonella isolated from an integrated broiler chicken supply chain in Qingdao,China

The present study analyzed the prevalence and antimicrobial resistance of Salmonella along an integrated broiler chicken supply chain. A total of 172 Salmonella isolates were recovered from 1148 samples collected from four sample sources (breeder farms, broiler farms, abattoir, and retail markets), representing nine production stages. These Salmonella isolates were examined for antimicrobial susceptibility to 12 different antimicrobial agents using a disk diffusion assay. Among them, 168 were identified as six different serotypes of Salmonella enterica. The predominant serotype was S. Enteritidis (n = 116), followed by S. Infantis (n = 18), S. Gueuletapee (n = 16), S. Derby (n = 12), S. Meleagridis (n = 4), and S. London (n = 2). The remaining four isolates were serogroup-untypeable. A majority of the 172 isolates (96.51%) was resistant to one or more antibiotics and 61.05% of the Salmonella isolates showed a multidrug resistance phenotype. Statistical analysis indicated the one risk product stage for Salmonella contamination occurred in the sample source at the abattoir, specifically the stage of Carcasses after chilling. The majority of S. Enteritidis isolates shared the same pulsed-field gel electrophoresis (PFGE) cluster, suggesting that the S. Enteritidis strain might spread along the broiler chicken supply chain. The prevalence and antimicrobial resistance of Salmonella in different production stages suggest the importance of controlling Salmonella in the broiler chicken supply chain for public health, underlying the need for improved measures of reducing carcass contamination in abattoirs and the appropriate use of antimicrobials in broiler flocks.     

Authentication of Iberian dry-cured ham: New approaches by polymorphic fingerprint and ultrahigh resolution mass spectrometry

Foods with high added value, such as Iberian dry-cured products, are susceptible to fraud. Many attempts have been made to differentiate the commercial/quality categories of Iberian dry-cured hams by analytical determinations. However, as discrimination by such means is not fully reliable, legislation to prevent fraudulent practice is based on administrative controls and certification. Here, new analytical approaches based on ultrahigh resolution mass spectrometry (UHRMS) and crystallographic techniques applied to the lipid fraction, in combination with chemometrics, are studied. The results of the triacylglycerol profile determined by UHRMS and the fingerprint provided by the thermograms obtained by differential scanning calorimetry offer the promise of analytic discrimination of Iberian dry-cured ham categories. In addition, these determinations, in combination with chemometrics, may prove extremely useful to authenticate many foods containing high to moderate amounts of lipids.     

Antibacterial mechanism of bifidocin A,a novel broad-spectrum bacteriocin produced by Bifidobacterium animalis BB04

Bifidocin A, a novel broad-spectrum bacteriocin produced by Bifidobacterium animalis BB04, was isolated from the feces of a healthy centenarian. To understand the mechanism of the antibacterial action of bifidocin A against gram-negative bacteria, its effects at a minimum inhibitory concentration on cell morphology, intracellular organization, membrane permeability, membrane integrity, and membrane proton motive force (PMF) of Escherichia coli 1.90 were investigated. Scanning and transmission electron microscopy analyses showed that bifidocin A induced alterations in the morphology and intracellular organization of E. coli cells. The intracellular organization was more susceptible to changes induced by bifidocin A than the morphology. Bifidocin A treatment caused the leakage of K⁺ and inorganic phosphate, the release of ATP and UV-absorbing materials, and a collapse of the transmembrane electrical potential and pH gradient in E. coli cells. Confocal laser scanning microscopy images showed that E. coli cells treated with bifidocin A took up propidium iodide. These results suggested that the mechanism of action bifidocin A against E. coli involved dissipation of the PMF of the cytoplasmic membrane, an increase in membrane permeability, pore formation in the cell membrane, a change in membrane integrity, and complete cell disintegration.