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Harmful Algal Blooms (HABs) in freshwater systems and intensified aquaculture have increased the risk to human health through exposure to cyanotoxins such as microcystin-LR (MC-LR). To understand the uptake and processing of MC-LR in humans, the pig was chosen as an animal model. This was assessed by repeated exposure for 13 weeks of eight animals dosed daily with MC-LR at 0.04 µg/kg bw, repeated with six animals over five weeks at a dose 50 times higher at 2 µg/kg bw. An analytical method was developed for MC-LR in porcine serum and also to analyse levels of free MC-LR in harvested porcine tissues, with Lemieux Oxidation employed to determine bound MC-LR in these tissues. MC-LR was not detected in the serum of treated animals from either experiment but free MC-LR was observed in the large intestine and kidney from two animals from the higher dosed group at levels of 1.4 and 1.9 µg/kg dry weight (dw) respectively. The results indicated 50% of higher dosed animals accumulated bound MC-LR in liver tissue, averaging 26.4 µg, approximately 1.1% of the dose administered. These results point to the potential uptake and accumulation of MC-LR in human liver tissue exposed chronically to sub-acute doses.

Certain endogenous bile acids have been proposed as potential therapies for ameliorating Alzheimer's disease (AD) but their role, if any, in the pathophysiology of this disease is not currently known. Given recent evidence of bile acids having protective and anti-inflammatory effects on the brain, it is important to establish how AD affects levels of endogenous bile acids. Using LC-MS/MS, this study profiled 22 bile acids in brain extracts and blood plasma from AD patients (n = 10) and age-matched control subjects (n = 10). In addition, we also profiled brain/plasma samples from APP/PS1 and WT mice (aged 6 and 12 months). In human plasma, we detected significantly lower cholic acid (CA, p = 0.03) in AD patients than age-matched control subjects. In APP/PS1 mouse plasma we detected higher CA (p = 0.05, 6 months) and lower hyodeoxycholic acid (p = 0.04, 12 months) than WT. In human brain with AD pathology (Braak stages V-VI) taurocholic acid (TCA) were significantly lower (p = 0.01) than age-matched control subjects. In APP/PS1 mice we detected higher brain lithocholic acid (p = 0.05) and lower tauromuricholic acid (TMCA; p = 0.05, 6 months). TMCA was also decreased (p = 0.002) in 12-month-old APP/PS1 mice along with 5 other acids: CA (p = 0.02), β-muricholic acid (p = 0.02), Ω-muricholic acid (p = 0.05), TCA (p = 0.04), and tauroursodeoxycholic acid (p = 0.02). The levels of bile acids are clearly disturbed during the development of AD pathology and, since some bile acids are being proposed as potential AD therapeutics, we demonstrate a method that can be used to support work to advance bile acid therapeutics.

The recent detection of tetrodotoxins (TTXs) in European fish and shellfish has emphasized the urgent need to develop specific, selective, rapid and easy-to-use methods for their detection to assess the potential risk posed to human health. For this purpose, a dithiol self-assembled monolayer (SAM)-based immunoassay previously performed on maleimide plates (mELISA) has been adapted to gold electrode arrays for the development of an electrochemical immunosensor for TTX. The electrochemical SAM-based immunosensor designed herein, provided an oriented, stable and spaced sensing platform for the determination of TTX, attaining a limit of detection of 2.6 ng mL-1. The applicability of the biosensor array was demonstrated by the accurate quantifications obtained in the analysis of different tissues of several puffer fish species (Lagocephalus lagocephalus, L. sceleratus and Sphoeroides pachygaster) caught along the Mediterranean coast of Spain. The good agreements found between the TTX concentrations determined by the immunosensor array platforms and those determined by mELISA, surface Plasmon resonance (SPR) immunosensor and liquid chromatography-high resolution mass spectrometry (LC-HRMS) analysis, proved the feasibility of the approach. The electrochemical immunosensor enables the determination of TTXs at levels as low as 0.07 mg TTX equiv. kg-1 tissue, thus, well below the Japanese value of 2 mg TTX equiv. kg-1 tissue used as a criterion to consider puffer fish safe for consumption. Compared to the colorimetric SAM-based approach, the immunosensor array described herein shows promise towards the development of disposable, portable and compact analysis tools applicable in monitoring programs for the surveillance of fishery products.

The recent detection of tetrodotoxins (TTXs) in puffer fish and shellfish in Europe highlights the necessity to monitor the levels of TTXs in seafood by rapid, specific, sensitive and reliable methods in order to protect human consumers. A previous immunoassay for TTX detection in puffer fish, based on the use of self-assembled monolayers (SAMs) for the immobilization of TTX on maleimide plates (mELISA), has been modified and adapted to the analysis of oyster and mussel samples. Changing dithiol for cysteamine-based SAMs enabled reductions in the assay time and cost, while maintaining the sensitivity of the assay. The mELISA showed high selectivity for TTX since the antibody did not cross-react with co-occurring paralytic shellfish poisoning (PSP) toxins and no interferences were observed from arginine (Arg). Moreover, TTX-coated maleimide plates stored for 3 months at −20 °C and 4 °C were stable, thus when pre-prepared, the time to perform the assay is reduced. When analyzing shellfish samples, matrix effects and toxin recovery values strongly depended on the shellfish type and the sample treatment. Blank oyster extracts could be directly analyzed without solid-phase extraction (SPE) clean-up, whereas blank mussel extracts showed strong matrix effects and SPE and subsequent solvent evaporation were required for removal. However, the SPE clean-up and evaporation resulted in toxin loss. Toxin recovery values were taken as correction factors (CFs) and were applied to the quantification of TTX contents in the analysis of naturally-contaminated shellfish samples by mELISA. The lowest effective limits of detection (eLODs) were about 20 and 50 µg/kg for oyster extracts without and with SPE clean-up, respectively, and about 30 µg/kg for mussel extracts with both protocols, all of them substantially below the eLOD attained in the previous mELISA for puffer fish (230 µg/kg). Analysis of naturally-contaminated samples by mELISA and comparison with LC-MS/MS quantifications demonstrated the viability of the approach. This mELISA is a selective and sensitive tool for the rapid detection of TTX in oyster and mussel samples showing promise to be implemented in routine monitoring programs to protect human health. info:eu-repo/semantics/acceptedVersion

Aflatoxin, a human liver carcinogen, frequently contaminates groundnuts, maize, rice, and other grains, especially in Africa. The aim of this study was to evaluate the effectiveness of an educational intervention that involved training rural Gambian women on how to identify and remove moldy groundnuts to reduce aflatoxin B

The International Agency for Research on Cancer reported that eating processed meat can increase a person’s risk for colorectal cancer and classified processed meat as carcinogenic to humans. Nitrate and nitrite are used as additives to improve food quality and protect against microbial contamination and are sources of N-nitroso compounds (NOCs) which are known carcinogens. This review outlines the association between processed meat intake and colorectal cancer risk and discusses the use of nitrates and nitrites in processed meat as well as healthier alternatives. A wide range of factors affect the formation of NOCs including the amount of nitrite added, meat quality, fat content, processing, maturation and handling at home. Factors related to processing include additives, precursors (added via wood smoke, spices or other ingredients), heat applied during drying or smoking, storage/maturation conditions and packaging. NOC formation can be inhibited by the addition of ascorbic acid and α-tocopherol ingredients which are often added to processed meats. Studies have shown that plant polyphenols and alpha-tocopherol significantly decreased pH, lipid oxidation and residual nitrite content of processed meat. Plant polyphenols, especially green tea polyphenols can be used as alternatives to nitrates and nitrites to process meat improving the quality, shelf life and safety of processed meat products. These innovative meat products could potentially contribute to a reduction in cancer risk by means of nitrite reduction and phytochemical addition and should be explored further.

The academic discussion of power in supply chains has changed from a discussion of the use of coercive power to one which emphasizes the role of trust in embedding co-operation and disincentivizing opportunism. Whilst a number of empirical studies have suggested the former is alive and well, this paper argues that power relations may also be constituted by the self-perceptions of weaker actors as much as by the explicit actions of more powerful ones. This study explores the role of power through the perceptions of subjugated actors, which set the ‘rules of the game’. Our case centres on perceptions of Northern Irish beef farmers and their reflections on their ‘powerlessness’ in relation to the larger, more consolidated processors that they sell to. We find that the way farmers make sense of the power relations they encounter is influenced by the individuating character of the power relations exercised by the processors, which debilitates their ability to collaborate and resist collectively. What emerges is a story about the process of accommodation whereby farmers pragmatically resign themselves to play by ‘the rules of the game’ to remain ‘part of the game’.

Nitrofurans are a broad group of drugs once widely used for the treatment of microbial and proto-zoal infections in many livestock species. However, as concerns grew globally with regard to the potentially carcinogenic and mutagenic effects of their residues in foods, they have been banned from use in many parts of the world. In order to monitor compliance to these bans, it is essential to have fit-for-purpose testing meth-ods. Immunoassays are the screening tool of choice for many testing laboratories due to their relative low cost, ease of use and high sensitivity. As is the case with all immunoassays, the most important reagents required are high quality, high affinity antibodies that exhibit the required sensitivity and specificity. Generating such antibodies for the nitrofuran family of compounds has required a great deal of effort in the design of immuno-gens, as the compounds, due to size, are not capable of eliciting an immune response in hosts and are not eas-ily conjugated to carrier proteins. This article reviews the range of strategies used to successfully generate suitable antibodies to a wide range of these drugs and their metabolites. In addition, the platform technologies for nitrofuran detection have moved from simple enzyme-linked immunosorbent assay (ELISA)-based proce-dures to more sophisticated multiplexing systems which can undertake faster and broader spectrum testing for the parent drugs and their metabolites. Reviews of the technologies used for immunochemical detection of the nitrofurans and of commercially available test kits are also presented.

Practical solutions for multiple antibiotic determination in food are required by the food industry and regulators for cost-effective screening purposes. This study describes the feasibility in development and preliminary performance of a novel multispot nanoarray for antibiotic screening in honey. Using a multiplex approach, the metabolites of the four main nitrofuran antibiotics, including morpholinomethyl-2-oxazolidone (AMOZ), 3-amino-2-oxazolidinone (AOZ), semicarbazide (SEM), 1-aminohydantoin (AHD) and chloramphenicol (CAP), were simultaneously detected. Antibodies specific to the five antibiotics were nano-spotted onto microtitre plate wells and a direct competitive assay format was employed. The assay characteristics and performance were evaluated for feasibility as a screening tool for antibiotic determination in honey to replace traditional ELISAs. Optimisation of the spotting and assay parameters was undertaken with both individual and multiplex calibration curves generated in PBS and a honey matrix. The limits of detection as determined by the 20% inhibitory concentrations (IC20) were determined as 0.19, 0.83, 0.09, 15.2 and 35.9 ng ml–1 in PBS, 0.34, 0.87, 0.17, 42.1 and 90.7 ng ml–1 in honey (fortified at the start of the extraction), and 0.23, 0.98, 0.24, 24.8 and 58.9 ng ml–1 in honey (fortified at the end of the extraction) for AMOZ, AOZ, CAP, SEM and AHD respectively. This work has demonstrated the potential of multiplex analysis for antibiotics with results available for 40 samples within a 90-min period for antibiotics sharing a common sample preparation. Although both the SEM and AHD assay do not show the required sensitivity with the antibodies available for use to meet regulatory limits, with further improvements in these particular antibodies this multiplex format has the potential to show a reduction in cost with reduced labour time in combination with the high-throughput screening of samples. This is the first 96-well spotted microtitre plate nanoarray for the semi-quantitative and simultaneous analysis of antibiotics.

Nanotechnology has emerged as a technological advancement that could develop and transform the entire agri-food sector, with the potential to increase agricultural productivity, food security and economic growth for industries. Though as still a relatively new concept there are concerns over its safety, regulation and acceptance by the industry and consumers. This review set out to address the implications of nanotechnology for the agri-food industry by examining the potential benefits, risks and opportunities. Existing scientific gaps in knowledge are believed to be a prohibitive factor in addition to uncertainties in the level of awareness and attitudes towards the use of nanotechnology by the industry.

Aflatoxin B1 (AFB1), ochratoxin A (OTA) and fumonisin B1 (FB1) are contaminants which have been shown to regularly co-occur in a range of foods. However, only a small number of studies have evaluated the interactive effect of binary and tertiary mycotoxins. The present study evaluated the effects of low levels of each mycotoxin in combination at their EU regulatory limits. Toxic effect with respect to cell viability was measured by MTT and neutral red assays, assessing mitochondria and lysosome integrities respectively. Individual toxicity showed that OTA (10 μg/ml) was the most cytotoxic mycotoxin in all three cell lines studied (caco-2, MDBK and raw 264.7). Binary combinations were cytotoxic to the MDBK cell line in the order [OTA/FB1] > [AFB1/FB1] > [AFB1/OTA], whilst all effects observed were classified as being additive. Tertiary combinations of AFB1, FB1 and OTA at the EU regulatory limits were tested and not found to exhibit measurable cytotoxicity in MDBK, caco-2 or raw 264.7 cells. However by increasing these concentrations above the legal limits to OTA (3 μg/ml), FB1 (8 μg/ml) and AFB1 (1.28 μg/ml), cytotoxicity was observed with up to 26% reduction in cell viability and synergistic effects were evident with regard to mitochondrial integrity.

A novel multiplex microarray has been developed for the detection of five groups of harmful algal and cyanobacterial toxins found in marine, brackish, and freshwater environments including domoic acid (DA), okadaic acid (OA, and analogues), saxitoxin (STX, and analogues), cylindrospermopsin (CYN) and microcystins (MC, and analogues). The sensitivity and specificity were determined and feasibility to be used as a screening tool investigated. Results for algal/cyanobacterial cultures (n = 12) and seawater samples (n = 33) were compared to conventional analytical methods, such as high performance liquid chromatography (HPLC) and liquid chromatography tandem mass spectrometry (LC-MS/MS). Detection limits for the 15 min assay were 0.37, 0.44, 0.05, 0.08, and 0.40 ng/mL for DA, OA, STX, CYN, and MC, respectively. The correlation of data obtained from the microarray compared to conventional analysis for the 12 cultures was r(2) = 0.83. Analysis of seawater samples showed that 82, 82, 70, 82, and 12% of samples were positive (IC20) compared to 67, 55, 36, 0, and 0% for DA, OA, STX, CYN, and MC, respectively, for conventional analytical methods. The discrepancies in results can be attributed to the enhanced sensitivity and cross-reactivity profiles of the antibodies in the MBio microarray. The feasibility of the microarray as a rapid, easy to use, and highly sensitive screening tool has been illustrated for the five-plex detection of biotoxins. The research demonstrates an early warning screening assay to support national monitoring agencies by providing a faster and more accurate means of identifying and quantifying harmful toxins in water samples.

Sports supplements are becoming a regular dietary addition for consumers who view such products as a means of improving their health and performance. Previously estrogenic endocrine disruptors (EDs) were detected in 80% of 116 sports supplements investigated by biological in vitro reporter gene assays (RGAs). The aim of this study was to quantify the hormonal activity in 50 of these sports supplement samples using a validated estrogen RGA and perform an exposure and risk assessment for human health. Results showed that 17β-estradiol equivalent levels were higher than those reported as being present in the typical human omnivore diet in 33 of the sports supplements and higher than the acceptable daily intake (ADI) in 13 of these products. The highest activity samples presented a potential to influence the human daily exposure to 17β-estradiol like activity in various risk groups with a predicted hormonal impact of greatest concern in young boys and postmenopausal women. In conclusion, consumers of sports supplements may be exposed to high levels of estrogenic EDs.

Patulin (PAT) is a mycotoxin produced by various species of fungi, with Penicillium expansum being the most commonly occurring. Apples and apple products are the main sources of PAT contamination. This mycotoxin has been shown to induce toxic effects in animals, a few of which include reproductive toxicity and interference with the endocrine system. Here the endocrine disrupting potential of PAT has been investigated in vitro to identify disruption at the level of oestrogen, androgen, progestagen and glucocorticoid nuclear receptor transcriptional activity, and to assess interferences in estradiol, testosterone and progesterone steroid hormone production. At the receptor level, 0.5–5000 ng/ml (0.0032–32 μM) PAT did not appear to induce any specific (ant) agonistic responses in reporter gene assays (RGAs); however, nuclear transcriptional activity was affected. A >6 fold increase in the glucocorticoid receptor transcriptional activity was observed following treatment with 5000 ng/ml PAT in the presence of cortisol. At the hormone production level, despite cytotoxicity being observed after treatment with 5000 ng/ml PAT, estradiol levels had increased >2 fold. At 500 ng/ml PAT treatment, an increase in progesterone and a decrease in testosterone production were observed. The findings of this study could be considered in assessing the health risks following exposure to PAT.

Refined vegetable oils are widely used in the food industry as ingredients or components in many processed food products in the form of oil blends. To date, the generic term ‘vegetable oil’ has been used in the labelling of food containing oil blends. With the introduction of new EU Regulation for Food Information (1169/2011) due to take effect in 2014, the oil species used must be clearly identified on the package and there is a need for development of fit for purpose methodology for industry and regulators alike to verify the oil species present in a product. The available methodologies that may be employed to authenticate the botanical origin of a vegetable oil admixture were reviewed and evaluated. The majority of the sources however, described techniques applied to crude vegetable oils such as olive oil due to the lack of refined vegetable oil focused studies. Nevertheless, DNA based typing methods and stable isotopes procedures were found not suitable for this particular purpose due to several issues. Only a small number of specific chromatographic and spectroscopic fingerprinting methods in either targeted or untargeted mode were found to be applicable in potentially providing a solution to this complex authenticity problem. Applied as a single method in isolation, these techniques would be able to give limited information on the oils identity as signals obtained for various oil types may well be overlapping. Therefore, more complex and combined approaches are likely to be needed to identify the oil species present in oil blends employing a stepwise approach in combination with advanced chemometrics. Options to provide such a methodology are outlined in the current study.

In recent years distillers dried grains and solubles (DDGS), co-products of the bio-ethanol and beverages industries, have become globally traded commodity for the animal feed sector. As such it is becoming increasingly important to be able to trace the geographical origin of commodities in case of a contamination incident or authenticity issue arise. In this study, 137 DDGS samples from a range of different geographical origins (China, USA, Canada and European Union) were collected and analyzed. Isotope ratio mass spectrometry (IRMS) was used to analyze the DDGS for 2H/1H, 13C/12C, 15N/14N, 18O/16O and 34S/32S isotope ratios which can vary depending on geographical origin and processing. Univariate and multivariate statistical techniques were employed to investigate the feasibility of using the IRMS data to determine botanical and geographical origin of the DDGS. The results indicated that this commodity could be differentiated according to their place of origin by the analysis of stable isotopes of hydrogen, carbon, nitrogen and oxygen but not with sulfur. By adding data to the models produced in this study, potentially an isotope databank could be set up for traceability procedures for DDGS, similar to the one established already for wine which will help in feed and food security issues arising worldwide.

A B S T R A C T Blooms of Alexandrium occur annually during the summer months in the North Channel of Cork Harbour on the south coast of Ireland. This study monitored an extensive bloom of the toxin producing Alexandrium minutum during the summer of 2011 with the use of the MIDTAL (Microarrays for the Detection of Toxic Algae) microarray and a prototype multiplex surface plasmon resonance (multi SPR) biosensor. Microarray signal intensities and toxin results from three testing platforms of the prototype multi SPR biosensor, commercial (CER) enzyme-linked immunosorbent assay (ELISA) and high performance liquid chromatography (HPLC) were compared against light microscopy counts. The main aim was to demonstrate the use of these methodologies to support national monitoring agencies by providing a faster and more accurate means of identifying and quantifying the harmful phytoplankton community and their toxins in natural water samples. Both the microarray signals and multi SPR biosensor results followed a significant trend with light microscopy results and both techniques indicated detection limits of

Ethnopharmacological relevance The ethnobotanical use of Aframomum melegueta in the treatment of urinary tract and soft tissue infection suggested that the plant has antimicrobial activity. Materials and methods To substantiate the folkloric claims, an acetone, 50:50 acetone:methanol and 2:1 chloroform:methanol extracts were tested against Escherichia coli K12 ; acetone extract and the fractions of acetone extracts were tested against Listeria monocytogenes . Bioassay-guided fractionation was performed on the extract using L. monocytogenes as the test organism to isolate the bioactive compounds which were then tested against all the other organisms. Results Four known labdane diterpenes (G3 and G5) were isolated for the first time from the rhizomes of A. melegueta and purified. These were tested against E. coli , L. monocytogenes , methicillin resistant Staphylococus aureus (MRSA) and S. aureus to determine antibacterial activity. The result showed that two compounds G3 and G5 exhibited more potent antibacterial activity compared to the current clinically used antibiotics ampicillin, gentamicin and vancomycin and can be potential antibacterial lead compounds. The structure of the labdane diterpenes were elucidated using nuclear magnetic resonance (NMR) spectroscopy and Mass spectrometry. A possible mode of action of the isolated compound G3 and its potential cytotoxicity towards mammalian cells were also discussed. Conclusion The results confirmed the presence of antibacterial compounds in the rhizomes of A. melegueta with a favourable toxicity profile which could be further optimized as antibacterial lead compounds.

Despite considerable advances in reducing the production of dioxin-like toxicants in recent years, contamination of the food chain still occasionally occurs resulting in huge losses to the agri-food sector and risk to human health through exposure. Dioxin-like toxicity is exhibited by a range of stable and bioaccumulative compounds including polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs), produced by certain types of combustion, and man-made coplanar polychlorinated biphenyls (PCBs), as found in electrical transformer oils. While dioxinergic compounds act by a common mode of action making exposure detection biomarker based techniques a potentially useful tool, the influence of co-contaminating toxicants on such approaches needs to be considered. To assess the impact of possible interactions, the biological responses of H4IIE cells to challenge by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in combination with PCB-52 and benzo-a-pyrene (BaP) were evaluated by a number of methods in this study. Ethoxyresorufin-O-deethylase (EROD) induction in TCDD exposed cells was suppressed by increasing concentrations of PCB-52, PCB-153, or BaP up to 10 μM. BaP levels below 1 μM suppressed TCDD stimulated EROD induction, but at higher concentrations, EROD induction was greater than the maximum observed when cells were treated with TCDD alone. A similar biphasic interaction of BaP with TCDD co-exposure was noted in the AlamarBlue assay and to a lesser extent with PCB-52. Surface enhanced laser desorption/ionization-time of flight mass spectrometry (SELDI-TOF) profiling of peptidomic responses of cells exposed to compound combinations was compared. Cells co-exposed to TCDD in the presence of BaP or PCB-52 produced the most differentiated spectra with a substantial number of non-additive interactions observed. These findings suggest that interactions between dioxin and other toxicants create novel, additive, and non-additive effects, which may be more indicative of the types of responses seen in exposed animals than those of single exposures to the individual compounds.

Surface plasmon resonance (SPR)-based biosensor is a popular platform for real-time monitoring and sensitive detection for a myriad of targets. However, only a few studies have reported the use of bacteriophages as specific binders for SPR-based detection. This study aimed to demonstrate how filamentous M13 bacteriophages expressing 12-mer peptides can be employed in an SPR-based assay, using a Salmonella -specific bacteriophage as a model binder to detect the foodborne bacterium Salmonella. Several important factors (immobilization buffers and methods, and interaction buffers) for a successful bacteriophage-based SPR assay were optimized. As a result, a Salmonella -specific bacteriophage-based SPR assay was achieved, with very low cross reactivity with other non-target foodborne pathogens and detection limits of 8.0 × 10 7 and 1.3 × 10 7 CFU/mL for one-time and five-time immobilized sensors, respectively. This proof-of-concept study demonstrates the feasibility of using M13 bacteriophages expressing target-specific peptides as a binder in a rapid and label-free SPR assay for pathogen detection.

Protein G-coated magnetic particles (MPs) were used as immobilisation supports for an antibody against okadaic acid (MAb OA ) and carriers into a surface plasmon resonance (SPR) device for the development of a direct competitive immunosensor for okadaic acid (OA). SPR analysis of MAb OA –MP conjugates demonstrated that conjugations were successful with complete immobilisation of all the antibody biomolecules onto the MPs. Moreover, MAb OA –MP conjugates provided up to 11-fold higher SPR signals, compared to free MAb OA . The use of conjugates in the direct competition assay provided a 3-fold lower LOD μg/L (2.6 μg of OA/L, equivalent to 12 μg of OA/kg mussel meat). The presence of mussel matrix did not interfere in the OA quantification as seen in the calibration curves. Mussel samples, obtained from Ebro Delta's bays (NW Mediterranean) during a diarrheic shellfish poisoning (DSP) event and in the presence of Dinophysis sacculus , an OA producer, in the shellfish production area, were analysed with the MP-based SPR immunosensor. The OA contents correlated with those obtained by liquid chromatography–tandem mass spectrometry (LC–MS/MS) ( y = 0.984 x − 5.273, R 2 = 0.789, p

The need for safe and quality food, free from the presence of hazardous contaminants such as mycotoxins is an on-going and complex challenge. Cold atmospheric pressure plasma (CAPP) has the potential to contribute to achieving this goal. Decontamination efficacy of CAPP against six of the most common mycotoxins found in foods and feedstuffs was assessed herein. Concentration reduction of up to 66% was achieved in maize for both aflatoxin B1 and fumonisin B1. Degradation products were detected only in the case of aflatoxin B1 and zearalenone and were tested on human hepatocarcinoma cells with no increase in cytotoxicity observed. Analysis of treated maize revealed substantial changes to small molecular mass components of the matrix. While CAPP shows promise in terms of mycotoxin detoxification important questions concerning potential changes to the nutritional and safety status of the food matrix require further investigations.

Plant secondary metabolites are protective dietary constituents and rol genes evidently increase the synthesis of these versatile phytochemicals. This study subjected a globally important vegetable, lettuce (Lactuca sativa) to a combination of untargeted metabolomics (LC-QTof-MS) and in vitro bioactivity assays. Specifically, we examined the differences between untransformed cultured lettuce (UnT), lettuce transformed with either rolABC (RA) or rolC (RC) and commercially grown (COM) lettuce. Of the 5333 metabolite features aligned, deconvoluted and quantified 3637, 1792 and 3737 significantly differed in RA, RC and COM, respectively, compared with UnT. In all cases the number of downregulated metabolites exceeded the number increased. In vitro bioactivity assays showed that RA and RC (but not COM) significantly improved the ability of L. sativa to inhibit α-glucosidase, inhibit dipeptidyl peptidase-4 (DPP-4) and stimulate GLP-1 secretion. We putatively identified 76 lettuce metabolites (sesquiterpene lactones, non-phenolic and phenolic compounds) some of which were altered by several thousand percent in RA and RC. Ferulic acid levels increased 3033–9777%, aminooxononanoic acid increased 1141–1803% and 2,3,5,4′tetrahydroxystilbene-2-O-β-d-glucoside increased 40,272–48,008%. Compound activities were confirmed using commercially obtained standards. In conclusion, rol gene transformation significantly alters the metabolome of L.sativa and enhances its antidiabetic properties. There is considerable potential to exploit rol genes to modulate secondary metabolite production for the development of novel functional foods. This investigation serves as a new paradigm whereby genetic manipulation, metabolomic analysis and bioactivity techniques can be combined to enable the discovery of novel natural bioactives and determine the functional significance of plant metabolites.

Context: Freshwater cyanobacterial toxins, microcystins, may be a contributing factor to the development of hepatocellular cancer and colorectal cancer.Objectives: This review summarizes the toxicity data, exposure routes and the methodologies available to determine exposure to elucidate the relationship to liver and colorectal cancer.Methods: Literature searches were conducted using Medline, PubMed and Web of Science.Results: There is evidence of human poisonings resulting from exposure to microcystins, however current methods rely on targeted approaches only suitable for acute exposure. No methods exist for the determination of chronic exposure to microcystins.Conclusions: With the growing evidence of exposure to microcystins and the possible links to cancer, methods to measure medium to long-term human exposure are needed. The identification and validation of candidate biomarkers are key to undertaking urgently required epidemiological studies.

Coccidiostats are authorized in the European Union (EU) to be used as poultry feed additives. Maximum (residue) levels (M(R)Ls) have been set within the EU for consumer and animal protection against unintended carry-over, and monitoring is compulsory. This paper describes the single-laboratory validation of a previously developed multiplex flow cytometric immunoassay (FCIA) as screening method for coccidiostats in eggs and feed and provides and compares different approaches for the calculation of the cut-off levels which are not described in detail within Commission Decision 2002/657/EC. Comparable results were obtained between the statistical (reference) approach and the rapid approaches. With the most rapid approach, the cut-off levels for narasin/salinomycin, lasalocid, diclazuril, nicarbazin (DNC) and monensin in egg, calculated as percentages of inhibition (%B/B0), were 60, 32, 76, 80 and 84, respectively. In feed, the cut-off levels for narasin/salinomycin, lasalocid, nicarbazin (DNC) and monensin were 70, 64, 72 and 78, respectively, and could not be determined for diclazuril. For all analytes, except for diclazuril in feed, the rate of false positives (false non-compliant) in blank samples was lower than 1 %, and the rate of false negatives (false compliant) at the M(R)Ls was below 5 %. Additionally, very good correlations (r ranging from 0.994 to 0.9994) were observed between two different analysers, a sophisticated flow cytometer (FlexMAP 3D(®)) and a more cost-efficient and transportable planar imaging detector (MAGPIX(®)), hence demonstrating adequate transferability.

The objective of this study was to examine the oestrogen and androgen hormone removal efficiency of reactive (Connelly zero-valent iron (ZVI), Gotthart Maier ZVI) and sorptive (AquaSorb 101 granular activated carbon (GAC) and OrganoLoc PM-100 organoclay (OC)) materials from HPLC grade water and constructed wetland system (CWS) treated dairy farm wastewater. Batch test studies were performed and hormone concentration analysis carried out using highly sensitive reporter gene assays (RGAs). The results showed that hormonal interaction with these materials is selective for individual classes of hormones. Connelly ZVI and AquaSorb 101 GAC were more efficient in removing testosterone (Te) than 17β-estradiol (E2) and showed faster removal rates of oestrogen and androgen than the other materials. Gotthart Maier ZVI was more efficient in removing E2 than Te. OrganoLoc PM-100 OC achieved the lowest final concentration of E2 equivalent (EEQ) and provided maximum removal of both oestrogens and androgens.

Low emission slurry spreading techniques such as the trailing shoe are known to increase nitrogen use efficiency and to increase the available window for slurry application, whilst minimising sward contamination. However, little is known about the ability of the trailing shoe to mitigate against incidental phosphorus (P) losses, especially following slurry application to mature swards. This 3 × 2 randomised block design experiment investigates the impact of sward cover on P losses in runoff following application of manure via simulated trailing shoe apparatus. Treatments comprised of three stages of grass growth (0, 10 and 20 day re-growths; equivalent to 1550, 2050 and 2900 kg DM/ha, respectively), and two slurry application rates: 25 m 3 /ha (Slurry) and 0 m 3 /ha (No slurry). Runoff was generated at day 2, 9 and 16 post slurry application using a drip type rainfall simulator. Throughout the experiment P concentrations in runoff were significantly greater from the Slurry treatments than from the No slurry treatments ( P P P > 0.05) on particulate P concentrations in runoff. No significant differences ( P > 0.05) between the three re-growth treatments for DRP, PP, or TP concentrations in runoff on day 9 and 16 post slurry application with the exception of the Slurry plots at day 16 when the 0 day re-growth treatment exhibited significant higher levels of DRP in runoff than the 10d ( P = 0.006) and 20d ( P = 0.003) RG treatments. This experiment demonstrated that allowing a grass sward to recover for at least 10 days post-harvest provides a potential mitigation strategy to reduce P losses in runoff following slurry application.

Multiplexed immunochemical detection platforms offer the potential to decrease labour demands, increase sample throughput and decrease overall time to result. A prototype four channel multiplexed high throughput surface plasmon resonance biosensor was previously developed, for the detection of food related contaminants. A study focused on determining the instruments performance characteristics was undertaken. This was followed by the development of a multiplexed assay for four high molecular weight proteins. The protein levels were simultaneously evaluated in serum samples of 10-week-old veal calves (n = 24) using multiple sample preparation methods. Each of the biosensor's four channels were shown to be independent of one another and produced multiplexed within run repeatability (n = 6) ranging from 2.0 to 6.7%CV, for the four tested proteins, whilst between run reproducibility (n = 4) ranged from 1.5 to 8.9%CV. Four calibration curves were successfully constructed before serum sample preparation was optimised for each protein. Multiplexed concentration analysis was successfully performed on four channels revealing that each proteins concentration was consistent across the twenty-four tested animals. Signal reproducibility (n > 19) on a further long term study revealed coefficient of variation ranging from 1.1% to 7.3% and showed that the multiplexed assay was stable for at least 480 cycles. These findings indicate that the performance characteristics fall within the range of previously published data for singleplex optical biosensors and that the multiplexing biosensor is fit-for-purpose for simultaneous concentration analysis in many different types of applications such as the multiplexed detection of markers of growth-promoter abuse and multiplexed detection of residues of concern in food safety.

Alternariol (AOH) is a mycotoxin commonly produced by Alternaria alternata on a wide range of foods. Few studies to date have been performed to evaluate the effects of AOH on endocrine activity. The present study makes use of in vitro mammalian cellular based assays and gene expression to investigate the ability of AOH to act as an endocrine disruptor by various modes of action. Reporter gene assays (RGAs), incorporating natural steroid hormone receptors for oestrogens, androgens, progestagens and glucocorticoids were used to identify endocrine disruption at the level of nuclear receptor transcriptional activity, and the H295R steroidogenesis assay was used to assess endocrine disruption at the level of gene expression and steroid hormone production. AOH exhibited a weak oestrogenic response when tested in the oestrogen responsive RGA and binding of progesterone to the progestagen receptor was shown to be synergistically increased in the presence of AOH. H295R cells when exposed to 0.1–1000 ng/ml AOH, did not cause a significant change in testosterone and cortisol hormones but exposure to 1000 ng/ml (3.87 μM) AOH resulted in a significant increase in estradiol and progesterone production. In the gene expression study following exposure to 1000 ng/ml (3.87 μM) AOH, only one gene NR0B1 was down-regulated, whereas expression of mRNA for CYP1A1, MC2R, HSD3B2, CYP17, CYP21, CYP11B2 and CYP19 was up-regulated. Expression of the other genes investigated did not change significantly. In conclusion AOH is a weak oestrogenic mycotoxin that also has the ability to interfere with the steroidogenesis pathway.

The presence of paralytic shellfish poisoning (PSP), diarrheic shellfish poisoning (DSP), and amnesic shellfish poisoning (ASP) toxins in seafood is a severe and growing threat to human health. In order to minimize the risks of human exposure, the maximum content of these toxins in seafood has been limited by legal regulations worldwide. The regulated limits are established in equivalents of the main representatives of the groups: saxitoxin (STX), okadaic acid (OA), and domoic acid (DA), for PSP, DSP, and ASP, respectively. In this study a multidetection method to screen shellfish samples for the presence of these toxins simultaneously was developed. Multiplexing was achieved using a solid-phase microsphere assay coupled to flow-fluorimetry detection, based on the Luminex xMap technology. The multidetection method consists of three simultaneous competition immunoassays. Free toxins in solution compete with STX, OA, or DA immobilized on the surface of three different classes of microspheres for binding to specific monoclonal antibodies. The IC50 obtained in the buffer was similar in single- and multidetection: 5.6 ± 1.1 ng/mL for STX, 1.1 ± 0.03 ng/mL for OA, and 1.9 ± 0.1 ng/mL for DA. The sample preparation protocol was optimized for the simultaneous extraction of STX, OA, and DA with a mixture of methanol and acetate buffer. The three immunoassays performed well with mussel and scallop matrixes displaying adequate dynamic ranges and recovery rates (around 90% for STX, 80% for OA, and 100% for DA). This microsphere-based multidetection immunoassay provides an easy and rapid screening method capable of detecting simultaneously in the same sample three regulated groups of marine toxins.

Thiamphenicol and florfenicol are antibacterial agents permitted for use as veterinary drugs in animals used for food production. However, as the EU has established maximum residue limits for both and the metabolite florfenicol amine, there is a requirement to monitor animal food products for their residues. In this study antisera were generated which can simultaneously detect thiamphenicol, florfenicol and florfenicol amine in an immunoassay. Details of the various coupling techniques employed to prepare immunogens and enzyme labels are provided and the antibodies produced have been assessed, in homologous and heterologous ELISA formats, with respect to sensitivity and specificity. It was found that while the antisera raised to thiamphenicol and florfenicol generally performed better in a heterologous set up, those raised to florfenicol amine were not only less affected by the assay format but also produced the most sensitive antibodies to all three target analytes. Antisera matched previous sensitivity (IC 50 − 1 ) but had improved cross-reactivity (> 100%) to thiamphenicol and florfenicol.

Spiramycin, tylosin, bacitracin and virginiamycin are among a group of antibiotic growth promoters that have been banned in the European Union since the 1999 Council. This was due to concerns over the development of resistant bacteria emerging between humans and animals with the threat of antibiotics no longer being able to be used effectively to treat human infections. A sensitive and fast immunochemical method is presented for the determination of these four antibiotic growth promoters simultaneously in poultry tissue. The method employs methanol extraction followed by sample clean-up by solid-phase extraction (SPE) with determination by enzyme-linked immunoabsorbant assay (ELISA). The limit of detection (LOD) was less than 1 ng g(-1) and the detection capability (CCβ) was 3 ng g(-1) or less for all four antibiotic growth promoters. Validation was completed with both raw and cooked chicken, therefore either matrix could be used for the monitoring of these banned drugs. In a feeding trial no residues of either bacitracin or virginiamycin were found in medicated birds even without a withdrawal period. In the case of tylosin and spiramycin much higher residues level were detected immunochemically than was the case by mass spectrometry.

A novel multiplexed immunoassay for the analysis of phycotoxins in shellfish samples has been developed. Therefore, a regenerable chemiluminescence (CL) microarray was established which is able to analyze automatically three different phycotoxins (domoic acid (DA), okadaic acid (OA) and saxitoxin (STX)) in parallel on the analysis platform MCR3. As a test format an indirect competitive immunoassay format was applied. These phycotoxins were directly immobilized on an epoxy-activated PEG chip surface. The parallel analysis was enabled by the simultaneous addition of all analytes and specific antibodies on one microarray chip. After the competitive reaction, the CL signal was recorded by a CCD camera. Due to the ability to regenerate the toxin microarray, internal calibrations of phycotoxins in parallel were performed using the same microarray chip, which was suitable for 25 consecutive measurements. For the three target phycotoxins multi-analyte calibration curves were generated. In extracted shellfish matrix, the determined LODs for DA, OA and STX with values of 0.5±0.3 μg L(-1), 1.0±0.6 μg L(-1), and 0.4±0.2 μg L(-1) were slightly lower than in PBS buffer. For determination of toxin recoveries, the observed signal loss in the regeneration was corrected. After applying mathematical corrections spiked shellfish samples were quantified with recoveries for DA, OA, and STX of 86.2%, 102.5%, and 61.6%, respectively, in 20 min. This is the first demonstration of an antibody based phycotoxin microarray.

Tetrodotoxin (TTX) is a potent neurotoxin emerging in European waters due to increasing ocean temperatures. Its detection in seafood is currently performed as a consequence of using the Association of Analytical Communities (AOAC) mouse bioassay (MBA) for paralytic shellfish poisoning (PSP) toxins, but TTX is not monitored routinely in Europe. Due to ethical and performance-related issues associated with this bioassay, the European Commission has recently published directives extending procedures that may be used for official PSP control. An AOAC-accredited high-performance liquid chromatography (HPLC) method has now been accepted by the European Union as a first action screening method for PSP toxins to replace the MBA. However, this AOAC HPLC method is not capable of detecting TTX, so this potent toxin would be undetected; thereby, a separate method of analysis is required. Surface plasmon resonance (SPR) optical biosensor technology has been proven as a potential alternative screening method to detect PSP toxins in seafood. The addition of a similar SPR inhibition assay for TTX would complement the PSP assay in removing the MBA. The present report describes the development and single laboratory validation in accordance with AOAC and IUPAC guidelines of an SPR method to be used as a rapid screening tool to detect TTX in the sea snail Charonia lampas lampas, a species which has been implicated in 2008 in the first case of human TTX poisoning in Europe. As no current regulatory limits are set for TTX in Europe, single laboratory validation was undertaken using those for PSP toxins at 800 μg/kg. The decision limit (CCα) was 100 μg/kg, with the detection capability (CCβ) found to be ≤200 μg/kg. Repeatability and reproducibility were assessed at 200, 400, and 800 μg/kg and showed relative standard deviations of 8.3, 3.8, and 5.4% and 7.8, 8.3, and 3.7% for both parameters at each level, respectively. At these three respective levels, the recovery of the assay was 112, 98, and 99%.

Aims The objective of this study was to develop a novel screening method for detection of viable Mycobacterium avium subsp. paratuberculosis (Map) in milk and faeces, as a rapid alternative to Map culture. Methods and Results The new method couples Map-specific peptide-mediated magnetic separation technique with an optimized phage amplification assay followed by detection of released progeny phage by ELISA in a competition assay format using polyclonal antibody produced against the D29 mycobacteriophage involved in the phage assay. Sample matrices were found not to interfere with the developed method, and the dynamic range of the assay was 3 × 102–6 × 108 phage ml−1. When low numbers of Map were present (102 CFU ml−1), the burst size of a single host Map cell was maximal (103 phage per cell) resulting in a highly sensitive screening assay. Conclusion A rapid, sensitive immuno-based screening method suitable for the detection of viable Map in milk and faeces was developed. Significance and Impact of the Study The novel PMS-phage-ELISA permits sensitive, qualitative detection of viable Map in milk or faeces samples within 48 h, representing a substantial decrease in time to detection compared with current culture methods for Map.

Aims The objectives of this study were to produce Salmonella-specific peptide ligands by phage display biopanning and evaluate their use for magnetic separation (MS). Methods and Results Four-phage display biopanning rounds were performed, and the peptides expressed by the two most Salmonella-specific (on the basis of phage-binding ELISA results) phage clones, MSal020401 and MSal020417, were chemically synthesized and coupled to MyOne™ tosylactivated Dynabeads®. Peptide capture capability for whole Salmonella cells from nonenriched broth cultures was quantified by MS + plate counts and MS + Greenlight™ detection and compared to capture capability of anti-Salmonella (antibody-coated) Dynabeads®. MS + Greenlight™ gave a more comprehensive picture of capture capability than MS + plate counts and showed that Peptide MSal020417-coated beads exhibited at least similar, if not better, capture capability to anti-Salmonella Dynabeads® (mean capture values of 36·0 ± 18·2 and 31·2 ± 20·1%, respectively, over Salmonella spp. concentration range 3 × 101–3 × 106 CFU ml−1) with cross-reactivity of ≤1·9% to three other foodborne pathogens: Escherichia coli, Listeria monocytogenes and Campylobacter jejuni. Conclusions One of the phage display-derived peptide ligands was demonstrated by MS + Greenlight™ to be a viable antibody alternative for MS of Salmonella spp. Significance and Impact of the Study This study demonstrates an antibody-free approach to Salmonella detection and opens substantial possibilities for more rapid tests for this bacterium.

We have made a comparison of (a) different surface chemistries of SPR sensor chips (such as carboxymethylated dextran and carboxymethylated C1) and (b) of different assay formats (direct, sandwich and subtractive immunoassay) in order to improve the sensitivity of the determination of the model bacteria Acidovorax avenae subsp. citrulli (Aac). The use of the carboxymethylated sensor chip C1 resulted in a better sensitivity than that of carboxymethylated dextran CM5 in all the assay formats. The direct assay format, in turn, exhibits the best sensitivity. Thus, the combination of a carboxymethylated sensor chip C1 with the direct assay format resulted in the highest sensitivity for Aac, with a limit of detection of 1.6 × 106 CFU mL-1. This SPR immunosensor was applied to the detection of Aac in watermelon leaf extracts spiked with the bacteria, and the lower LOD is 2.2 × 107 CFU mL−1.

In this study data generated by 1H NMR were combined with chemometrics to analyse brain and plasma samples from APP/PS1 and wild type mice with the aim of developing a statistical model capable of predicting the features of Alzheimer’s disease (AD) displayed by this animal model. APP/PS1 is a well characterised double transgenic mouse model of AD and the results here demonstrate the potential of NMR technology as a platform for the detecting this disease. Using partial least squares discriminant analysis a model was built using both brain extracts (R2 = 0.99; Q2 = 0.66) and a high throughput method of plasma analysis (R2 = 0.98; Q2 = 0.75) capable of predicting AD in APP/PS1 mice. Analysis of brain extracts led to the elucidation of 20 metabolites and 16 of these were quantifiable. Relative brain levels of ascorbate, creatine, γ-aminobutyric acid and N-acetyl aspartic acid were significantly altered in APP/PS1 mice (p < 0.05). Analysis of plasma identified 14 metabolites and the levels of acetate, citrate, glutamate, glutamine, methionine, and an unknown signal were significantly altered in APP/PS1 mice (p < 0.05). Combining 1H NMR spectral data with chemometrics has been previously used to study biochemical disturbances in various disease states. This study further indicates the translational potential of this technology for identifying AD in people attending the memory clinic.

Okadaic acid (OA) is a marine toxin ingested by shellfish. The consumption of contaminated seafood causes diarrheic shellfish poisoning, responsible for gastrointestinal troubles in humans. The development of fast, reliable and sensitive methods for OA analysis is an evident necessity in order to guarantee the seafood safety and to protect human health. In this work, competitive indirect immunoassays were developed for the toxin detection. Indirect and direct labeling were used and compared. With indirect labeling, the unlabeled anti-OA antibody binds to the toxin and a labeled secondary antibody is used for the signal generation. Alternatively, using the direct labeling, the label is attached via a covalent bond to the anti-OA antibody. In this work, peroxidase was conjugated to the anti-OA antibody through a periodate activation. The peroxidase–antibody conjugate is used for both toxin recognition and signal generation. In the last case, ELISA test is simplified and the colorimetric competitive indirect immunoassay based on direct labeling is more sensitive to OA than the test based on indirect labeling. In optimized conditions, the IC 50 value and detection limit were respectively 0.066 μg L −1 and 0.0061 μg L −1 . Then, the colorimetric immunoassay based on direct labeling was validated with certified reference mussel samples, demonstrating the efficiency of the approach.

Low emission slurry spreading techniques are known to improve nitrogen use efficiency, but their impact on phosphorus (P) losses in surface runoff has received little attention. The current study was designed to examine the effect of slurry spreading technique on P losses in runoff. Twelve treatments were examined on 0.5- m by 1.0-m plots in a nominal 2 × 6 factorial design experiment. Treatments comprised grass swards at two different stages of growth, a stubble and a 4-wk regrowth, and six different slurry application treatments: control (no slurry), and slurry applied to simulate splash-plate, injection (across and down slope), and trailing shoe (across and down slope) spreading. Slurry was applied by hand (40 m ha). Rainfall simulations (40 mm h) were conducted at 2, 9, and 28 d post-slurry application. When slurry was applied to the stubble, dissolved reactive P (DRP) concentrations in runoff at Day 2 were 47 and 37% lower ( < 0.05) from the injection and trailing shoe treatments compared with the splash-plate treatment. Similarly, at Day 2, TP concentrations in runoff from the injection treatments were 27% lower ( < 0.05) than the splash-plate treatment. In contrast, application technique had no effect ( 0.05) on P concentrations in runoff following slurry application to the regrowth treatment. Phosphorus concentrations in runoff were unaffected by direction of slurry spreading (across or down) at both applications. Our results indicate that trailing shoe and injection techniques offer the potential to reduce DRP concentrations in runoff during the period immediately after slurry application.

Marine dinoflagellates of the genera Alexandrium are well known producers of the potent neurotoxic paralytic shellfish toxins that can enter the food web and ultimately present a serious risk to public health in addition to causing huge economic losses. Direct coastal monitoring of Alexandrium spp. can provide early warning of potential shellfish contamination and risks to consumers and so a rapid, sensitive, portable and easy-to-use assay has been developed for this purpose using an innovative planar waveguide device. The disposable planar waveguide is comprised of a transparent substrate onto which an array of toxin-protein conjugates is deposited, assembled in a cartridge allowing the introduction of sample, and detection reagents. The competitive assay format uses a high affinity antibody to paralytic shellfish toxins with a detection signal generated via a fluorescently labelled secondary antibody. The waveguide cartridge is analysed by a simple reader device and results are displayed on a laptop computer. Assay speed has been optimised to enable measurement within 15 min. A rapid, portable sample preparation technique was developed for Alexandrium spp. in seawater to ensure analysis was completed within a short period of time. The assay was validated and the LOD and CCß were determined as 12 pg/mL and 20 pg/mL respectively with an intra-assay CV of 11.3% at the CCß and an average recovery of 106%. The highly innovative assay was proven to accurately detect toxin presence in algae sampled from the US and European waters at an unprecedented cell density of 10 cells/L. © 2012 Elsevier B.V. All rights reserved.

A prototype fluorescent based biosensor has been developed for the antibody based detection of food related contaminants. Its performance was characterised and showed a typical antibody binding signal of 200-2000 mV, a short term noise of 9.1 mV, and baseline slope of -0.016 mV/s over 4 h. Bulk signal detection repeatability (n=23) and reproducibility (n=3) were less than 2.4%CV. The biosensor detection unit was evaluated using two food related model systems proving its ability to monitor both binding using commercial products and inhibition through the development of an assay. This assay development potential was evaluated by observing the biosensor's performance whilst appraising several labelled antibody and glass slide configurations. The molecular interaction between biotin and an anti-biotin antibody was shown to be inhibited by 41% due to the presence of biotin in a sample. A food toxin (domoic acid) calibration curve was produced, with %CVs ranging from 2.7 to 7.8%, and a midpoint of approximately 17 ng/ml with further optimisation possible. The ultimate aim of this study was to demonstrate the working principles of this innovative biosensor as a potential portable tool with the opportunity of interchangeable assays. The biosensor design is applicable for the requirements of routine food contaminant analysis, with respect to performance, functionality and cost. (C) 2012 Elsevier B.V. All rights reserved.

Tetrodotoxin (TTX) is one of the most potent marine neurotoxins reported. The global distribution of this toxin is spreading with the European Atlantic coastline now being affected. Climate change and increasing pollution have been suggested as underlying causes for this. In the present study, two different sample preparation techniques were used to extract TTX from Trumpet shells and pufferfish samples. Both extraction procedures (accelerated solvent extraction (ASE) and a simple solvent extraction) were shown to provide good recoveries (80–92%). A UPLC–MS/MS method was developed for the analysis of TTX and validated following the guidelines contained in the Commission Decision 2002/657/EC for chemical contaminant analysis. The performance of this procedure was demonstrated to be fit for purpose. This study is the first report on the use of ASE as a mean for TTX extraction, the use of UPLC–MS/MS for TTX analysis, and the validation of this method for TTX in gastropods.

Soya bean products are used widely in the animal feed industry as a protein based feed ingredient and have been found to be adulterated with melamine. This was highlighted in the Chinese scandal of 2008. Dehulled soya (GM and non-GM), soya hulls and toasted soya were contaminated with melamine and spectra were generated using Near Infrared Reflectance Spectroscopy (NIRS). By applying chemometrics to the spectral data, excellent calibration models and prediction statistics were obtained. The coefficients of determination (R(2)) were found to be 0.89-0.99 depending on the mathematical algorithm used, the data pre-processing applied and the sample type used. The corresponding values for the root mean square error of calibration and prediction were found to be 0.081-0.276% and 0.134-0.368%, respectively, again depending on the chemometric treatment applied to the data and sample type. In addition, adopting a qualitative approach with the spectral data and applying PCA, it was possible to discriminate between the four samples types and also, by generation of Cooman's plots, possible to distinguish between adulterated and non-adulterated samples.

A panel of reporter gene assays (RGAs) coupled with a single solid phase extraction (SPE) step was developed and used to screen bottled mineral water for the presence of four classes of endocrine disruptors (EDs), oestrogens, androgens, progestagens and glucocorticoids. Fourteen brands of bottled mineral water in triplicate (42 samples) were analysed. Overall, hormonal activity was found in 78% of the samples. Oestrogenic, androgenic, progestagenic and glucocorticoid activity was found in 38%, 38%, 36% and 55% of the samples, respectively at an average concentration of 10 ng/l 17β-estradiol equivalent (EEQ), 26 ng/l testosterone equivalent (TEQ), 123 ng/l progesterone equivalent (PEQ) and 13.5 ng/l hydrocortisone equivalent (HEQ). The level of oestrogenic, androgenic and progestagenic activity observed is not considered a matter of concern for the consumers' health. It is unknown whether the glucocorticoid levels observed are safe. The ED source, long term exposure and mixture effects remain to be investigated.

Pyrrolizidine alkaloids (PAs) are known secondary plant metabolites which can cause hepatotoxicity in both humans and livestock. PAs can be consumed through the use of plants for food, medicinal purposes and as contaminants of agricultural crops and food. PA contaminated grain has posed the largest health risk, although any PA contamination in our food chain should be recognised as a potential health threat. For this purpose, retail honeys were tested by LC-MS/MS. The method allows for specific identification of toxic retronecine and otonecine-type PAs by comparison to reference compounds via a spectral library. In total, 50 honey samples were matched to the reference spectra within a set of tolerance parameters. Accurate data analysis and quick detection of positive samples was possible. Positive samples contained an average PA concentration of 1260 μg kg(-1) of honey. Good linear calibrations were obtained (R(2)>0.991). LOD and LOQ ranged from 0.0134 to 0.0305 and 0.0446 to 0.1018 μg mL(-1), respectively.

A study combining high resolution mass spectrometry (liquid chromatography-quadrupole time-of-flight-mass spectrometry, UPLC-QTof-MS) and chemometrics for the analysis of post-mortem brain tissue from subjects with Alzheimer's disease (AD) (n = 15) and healthy age-matched controls (n = 15) was undertaken. The huge potential of this metabolomics approach for distinguishing AD cases is underlined by the correct prediction of disease status in 94-97% of cases. Predictive power was confirmed in a blind test set of 60 samples, reaching 100% diagnostic accuracy. The approach also indicated compounds significantly altered in concentration following the onset of human AD. Using orthogonal partial least-squares discriminant analysis (OPLS-DA), a multivariate model was created for both modes of acquisition explaining the maximum amount of variation between sample groups (Positive Mode-R2 = 97%; Q2 = 93%; root mean squared error of validation (RMSEV) = 13%; Negative Mode-R2 = 99%; Q2 = 92%; RMSEV = 15%). In brain extracts, 1264 and 1457 ions of interest were detected for the different modes of acquisition (positive and negative, respectively). Incorporation of gender into the model increased predictive accuracy and decreased RMSEV values. High resolution UPLC-QTof-MS has not previously been employed to biochemically profile post-mortem brain tissue, and the novel methods described and validated herein prove its potential for making new discoveries related to the etiology, pathophysiology, and treatment of degenerative brain disorders.

The performance of spring calving Holstein-Friesian (HF) and Jersey×Holstein-Friesian (J×HF) dairy cows was examined during three successive years (mean of 35 HF cows and 31 J×HF cows per year). Throughout the experiment cows were managed on one of three grassland-based systems of milk production, namely low concentrate (LC), medium concentrate (MC) or high concentrate (HC). Post-calving, cows were housed and offered grass silage, supplemented with 6.0, 8.0 and 10.0 kg concentrate/cow/d in systems LC, MC and HC, respectively (mean period from calving to start of full time grazing, 69 days). During the grazing period target concentrate feed levels were 0, 2.5 and 5.0 kg/cow/d for systems LC, MC and HC, respectively (mean period from start of full time grazing to start full time re-housing, 206 days). Full lactation concentrate inputs were 530, 1092 and 1667 kg/cow, in systems LC, MC and HC, respectively. There were no significant genotype×system interactions for any of the milk production parameters examined. Food intake during the confinement and grazing periods was unaffected by genotype. Milk yield was highest with HF cows while milk fat and milk protein content were highest with the J×HF cows (P

Saxitoxin (STX) is a low molecular weight neurotoxin mainly produced by certain marine dinoflagellates that, along with its family of similarly related paralytic shellfish toxins, may cause the potentially fatal intoxication known as paralytic shellfish poisoning. Illness and fatality rates are low due to the effective monitoring programs that determine when toxins exceed the established regulatory action level and effectuate shellfish harvesting closures accordingly. Such monitoring programs rely on the ability to rapidly screen large volumes of samples. Many of the screening assays currently available employ antibodies or live animals. This research focused on developing an analytical recognition element that would eliminate the challenges associated with the limited availability of antibodies and the use of animals. Here we report the discovery of a DNA aptamer that targets STX. Concentration-dependent and selective binding of the aptamer to STX was determined using a surface plasmon resonance sensor. Not only does this work represent the first reported aptamer to STX, but also the first aptamer to any marine biotoxin. A novel strategy of using a toxin-protein conjugate for DNA aptamer selection was successfully implemented to overcome the challenges associated with aptamer selection to small molecules. Taking advantage of such an approach could lead to increased diversity and accessibility of aptamers to low molecular weight toxins, which could then be incorporated as analytical recognition elements in diagnostic assays for foodborne toxin detection. The selected STX aptamer sequence is provided here, making it available to any investigator for use in assay development for the detection of STX.

© 2016 Elsevier Ltd Blooms of the toxic dinoflagellate Alexandrium tamarense (Group 1) seriously impacted the Tasmanian shellfish industry during 2012 and 2015, necessitating product recalls and intensive paralytic shellfish toxin (PST) product testing. The performance of four commercial PST test kits, Abraxis™, Europroxima™, Scotia™ and Neogen™, was compared with the official AOAC LC-FLD method for contaminated mussels and oysters. Abraxis and Europroxima kits underestimated PST in 35–100% of samples when using standard protocols but quantification improved when concentrated extracts were further diluted (underestimation ≤18%). The Scotia kit (cut off 0.2–0.7 mg STX-diHCl eq/kg) delivered 0% false negatives, but 27% false positives. Neogen produced 5% false negatives and 13% false positives when the cut off was altered to 0.5–0.6 mg STX-diHCl eq/kg, the introduction of a conversion step eliminated false negatives. Based on their sensitivity, ease of use and performance, the Neogen kit proved the most suitable kit for use with Tasmanian mussels and oysters. Once formally validated for regulatory purposes, the Neogen kit could provide shellfish growers with a rapid tool for harvesting decisions at the farm gate. Effective rapid screening preventing compliant samples undergoing testing using the more expensive and time consuming LC-FLD method will result in significant savings in analytical costs.