LC-HRMS/MS for the simultaneous determination of four allergens in fish and swine food products.

The inclusion on the label of packed foods of any ingredient or technological adjuvant causing allergies is required by EU food legislation. In this study a targeted proteomics method for detecting four allergens in animal-derived food matrices was developed. Liquid chromatography coupled with high-resolution tandem mass spectrometry (LC-HRMS/MS) was used to select marker peptides from four allergens and develop a quantitative method able to simultaneously detect the presence of milk, egg, crustaceans and soy. The method was validated on fish or swine processed food products contaminated at 5 µg g-1 for milk and egg and 10 µg g-1 for soy and crustaceans. The method was tested by analyzing commercial food products with high protein content and was compared to the ELISA technique. Our results indicated the presence of soy not reported on the food label of some products, pointing out the need for efficient controls to protect allergic consumers.

Development and validation of a QuEChERS method coupled to liquid chromatography and high resolution mass spectrometry to determine pyrrolizidine and tropane alkaloids in honey.

Awareness about pyrrolizidine alkaloids (PAs) and tropane alkaloids (TAs) in food was recently raised by the European Food Safety Authority stressing the lack of data and gaps of knowledge required to improve the risk assessment strategy. The present study aimed at the elaboration and validation of a method to determine PAs and TAs in honey. QuEChERS sample treatment and liquid chromatography coupled to hybrid high resolution mass spectrometry, were used. The method resulted in good linearity (R2>0.99) and low limits of detection and quantification, ranging from 0.04 to 0.2µgkg-1 and from 0.1 to 0.7µgkg-1 respectively. Recoveries ranged from 92.3 to 114.8% with repeatability lying between 0.9 and 15.1% and reproducibility between 1.1 and 15.6%. These performances demonstrate the selectivity and sensitivity of the method for simultaneous trace detection and quantification of PAs and TAs in honey, verified through the analysis of forty commercial samples.

The Status of Honey Bee Health in Italy: Results from the Nationwide Bee Monitoring Network.

In Italy a nation-wide monitoring network was established in 2009 in response to significant honey bee colony mortality reported during 2008. The network comprised of approximately 100 apiaries located across Italy. Colonies were sampled four times per year, in order to assess the health status and to collect samples for pathogen, chemical and pollen analyses. The prevalence of Nosema ceranae ranged, on average, from 47-69% in 2009 and from 30-60% in 2010, with strong seasonal variation. Virus prevalence was higher in 2010 than in 2009. The most widespread viruses were BQCV, DWV and SBV. The most frequent pesticides in all hive contents were organophosphates and pyrethroids such as coumaphos and tau-fluvalinate. Beeswax was the most frequently contaminated hive product, with 40% of samples positive and 13% having multiple residues, while 27% of bee-bread and 12% of honey bee samples were contaminated. Colony losses in 2009/10 were on average 19%, with no major differences between regions of Italy. In 2009, the presence of DWV in autumn was positively correlated with colony losses. Similarly, hive mortality was higher in BQCV infected colonies in the first and second visits of the year. In 2010, colony losses were significantly related to the presence of pesticides in honey bees during the second sampling period. Honey bee exposure to poisons in spring could have a negative impact at the colony level, contributing to increase colony mortality during the beekeeping season. In both 2009 and 2010, colony mortality rates were positively related to the percentage of agricultural land surrounding apiaries, supporting the importance of land use for honey bee health.

Retrospective evaluation of lead contamination in honey from 2005 to present in northeastern Italy and future perspectives in the light of updated legislation.

Lead is a naturally occurring element but largely originating from human activities. Food is the major source of exposure to lead for humans and the publication of a scientific opinion from the European Food Safety Authority on the risks to human health related to the presence of lead in foodstuffs, led European Union to establish more restrictive limits for this contaminant in food from 1 January 2016. In particular, a maximum level of 0.10 mg kg(-1) was established for honey. The retrospective evaluation of 995 honey samples analysed since 2005, revealed a progressive reduction in the concentration of lead, with a mean value of 0.045 mg kg(-1) in 2015. Total 1.5% of honeys analysed in 2015 exceeded the maximum lead level and therefore will no longer be marketable. Interested beekeepers should clarify the causes of honey contamination and adopt corrective actions to keep their honey production within the legal levels of lead.

Food safety concerns deriving from the use of silver based food packaging materials.

The formulation of innovative packaging solutions, exerting a functional antimicrobial role in slowing down food spoilage, is expected to have a significant impact on the food industry, allowing both the maintenance of food safety criteria for longer periods and the reduction of food waste. Different materials are considered able to exert the required antimicrobial activity, among which are materials containing silver. However, challenges exist in the application of silver to food contact materials due to knowledge gaps in the production of ingredients, stability of delivery systems in food matrices and health risks caused by the same properties which also offer the benefits. Aims of the present study were to test the effectiveness and suitability of two packaging systems, one of which contained silver, for packaging and storing Stracchino cheese, a typical Italian fresh cheese, and to investigate if there was any potential for consumers to be exposed to silver, via migration from the packaging to the cheese. Results did not show any significant difference in the effectiveness of the packaging systems on packaged Stracchino cheese, excluding that the active packaging systems exerted an inhibitory effect on the growth of spoilage microorganisms. Moreover, silver migrated into the cheese matrix throughout the storage time (24 days). Silver levels in cheese finally exceeded the maximum established level for the migration of a non-authorised substance through a functional barrier (Commission of the European Communities, 2009). This result poses safety concerns and strongly suggests the need for more research aimed at better characterizing the new packaging materials in terms of their potential impacts on human health and the environment.

Expression of CYP4 and GSTr genes in Venerupis philippinarum exposed to benzo(a)pyrene.

Bivalve molluscs, such as Venerupis philippinarum, are often used as bioindicators of environmental pollution since they can bioaccumulate a large variety of pollutants because of their filter feeding. The Polycyclic Aromatic Hydrocarbon (PAH) benzo(a)pyrene (B(a)P) is an important contaminant, commonly present in the marine environment. Pollutants are generally metabolized by enzymes of phase I, mainly CYPs enzymes, and by conjugation enzymes of phase II like GST. In this study, we investigated by Real Time PCR the expression of CYP4 and GSTr (GST class rho) in the digestive gland of V. philippinarum exposed to different concentrations of B(a)P for 24 h and after a 24 h depuration period. Accumulation of B(a)P by clams has been confirmed by the HPLC-FLD analyses. Moreover, HPLC-FLD analyses evidenced that after depuration, B(a)P concentrations decreased in animals subjected to 0.03 mg/l and 0.5mg/l exposures but did not decrease in animals subjected to 1mg/l exposure. B(a)P exposure and depuration did not cause histopathological lesions in the different organs. The analysis of GSTr expression in the digestive gland showed a significant increase in mRNA in animals subjected to 1 mg/l exposure, whereas the analysis of CYP4 expression did not evidence differences among treatments. Moreover, the expression of both genes did not exhibit any differences after the purification treatment. The results demonstrate that B(a)P significantly affects the expression of GSTr mRNA in the digestive gland of V. philippinarum and suggest that GSTr gene could play an important role in the biotransformation of B(a)P.

Distribution of sulfathiazole in honey, beeswax, and honeybees and the persistence of residues in treated hives.

This study was performed to evaluate the distribution and depletion of sulfathiazole in different beehive matrices: honey, honeybees, "pre-existing" honeycomb, "new" honeycomb, and capping wax. Sulfathiazole was dissolved in sugar syrup or directly powdered on the combs, the matrices were sampled at different time points, and sulfathiazole residues were quantified by high-performance liquid chromatography with fluorescence detection. In honey, the higher concentration of sulfathiazole (180 mg kg(-1)) occurred 2 weeks after the last treatment in syrup. In beeswax, drug concentration was higher than in honey, particularly with powder administration, with a maximum level (340 mg kg(-1)) 3 days following the last treatment. The strongest contamination in honeybees (28 mg kg(-1)) was achieved with sulfathiazole administered in powder 3 days after the second treatment. The high persistence of sulfathiazole in the different beehive matrices suggests that it could be a reliable marker of previous treatments performed by beekeepers.

Canine nephrotoxicosis induced by melamine-contaminated pet food in Italy.

Two young dogs belonging to the same kennel placed nearby Treviso (north-eastern Italy) died at the end of 2008 with clinical signs of renal failure. They were subjected to necropsy and were evaluated for histopathological and toxicological changes. Both the animals had same clinical signs and laboratory evidence of uremia. Post mortem investigations revealed severe nephrotoxicosis, associated with uroliths deposition within renal tubules and pelvis. The predominant crystal type was identical to those observed in the kidneys of animals involved in the 2004 and 2007 melamine-associated renal failure epidemic in Asia and US, providing evidence that they share the same causative agent. High doses of melamine were detected in the pet food administered to the dogs, likewise melamine was identified in renal tissue from one dead dog and in urine samples from both the animals. Therefore, a diagnosis of melamine-related nephrotoxicosis was made. To the author's knowledge this is the first report about melamine contamination of pet food from EU.

Identification and semi-quantitative determination of gum Arabic in wines by GC-MS and size exclusion chromatography.

A method for the identification and the semi-quantitative determination of the food additive gum Arabic in wines is described. Tests carried out on solutions spiked with known amounts of wine and gum Arabic polysaccharides allowed to define the suitable conditions for their quantitative precipitation and size exclusion analysis. CG-MS analyses of the different recovered fractions allowed to discriminate between gum Arabic and wine polysaccharides through the identification of glucose and mannose present only in wine polysaccharides. The proposed method was based on the wine polysaccharides free peak area obtained by size exclusion chromatography. The same cut-off time was always used both in the preparation of the calibration plot and in the analysis of the real samples. Gum Arabic was determined in a ratio of 1/10 w/w with wine polysaccharides with a detection limit of 0.074 mg ml(-1) which is lower than the lowest gum Arabic amount usually added into wines. Owing to the moderately low natural variability of the gum Arabic standards the described procedure is suitable for a semi-quantitative analysis even if its accuracy allowed a quite reliable determination of the gum Arabic amount usually added to wine.

The use of cyclic voltammetric measurements for the determination of kinetic constants in enzyme-based electrocatalysis. The case of the redox mediated dihydronicotinamide adenine dinucleotide oxidation catalyzed by flavoproteins.

On the basis of simulated voltammetric curves used as testing data, practical methods to extract the bimolecular rate constant for a mediated enzymatic catalysis with an excess of the substrate are examined. The evaluation criteria are the simplicity of treatment and the agreement between the expected and the obtained values of the rate constants. The results obtained indicate that the different approaches examined are equivalent.

Kinetic considerations on the electrogenerated luminescence of luminol at platinum electrode in the presence of hydrogen peroxide and oxygen.

The ECL behavior of the luminol/H2O2 and luminol/O2 systems was evaluated at Pt electrode by using different electroanalytical techniques such as chronoamperometry, cyclic and rotating disk electrode (RDE) voltammetry. Diffusive and kinetic parameters such as the diffusion coefficient of luminol, D, the number of exchanged electrons, n, and the apparent heterogeneous rate constant, kap, were determined in the maximum light emission conditions achieved at pH 11, at an electrode potential of 750 mV vs. SCE. The experimental order of reaction were determined from the relation between the reactant concentrations and the emitted light intensity.