5.5 Pesticide and Metabolites Residues in Honeybees: A 2014-2017 Greek Compendium

In the period between 2014 mid-2017, more than 200 samples of honeybees were sent by authorities and individuals in Benaki Phytopathological Institute after incidents of unexpected deaths of bees in various parts of Greece. The samples were analyzed for pesticides and breakdown products, by two multi-residue methods based on an expanded HPLC-ESI-MS/MS and a newly developed GC-MS/MS method. Sample preparation was optimized and based on modified QuEChERS using for clean-up C18 and PSA. Until mid-2017, 293 detections were registered in a total of 205 honeybee samples, resulting in a 76% percent of positive samples, to at least one active substance. Concentrations’ range varied from 1 to 160000 ng/g bee body weight . In some cases, these levels surpassed LD50 values indicating intoxication events. Predominant substances were clothianidin, coumaphos, imidacloprid, acetamiprid and dimethoate. In less extent, other acaricides such as amitraz (mostly its breakdown products DMF, DMPF), tau-fluvalinate and certain pyrethroids exemplified by cyhalothrin, cypermethrin and deltamethrin were also recorded. In several samples, more than one active substance was detected.Overall, this work aspires to provide valuable insight to pesticides and metabolites occurrence in honeybees in Greece between 2014-2017 and in parallel assist research community and apiculturists in this pivotal Mediterranean region that bee health and pollination services have prolific importance.In the period between 2014 mid-2017, more than 200 samples of honeybees were sent by authorities and individuals in Benaki Phytopathological Institute after incidents of unexpected deaths of bees in various parts of Greece. The samples were analyzed for pesticides and breakdown products, by two multi-residue methods based on an expanded HPLC-ESI-MS/MS and a newly developed GC-MS/MS method. Sample preparation was optimized and based on modified QuEChERS using for clean-up C18 and PSA. Until mid-2017, 293 detections were registered in a total of 205 honeybee samples, resulting in a 76% percent of positive samples, to at least one active substance. Concentrations’ range varied from 1 to 160000 ng/g bee body weight . In some cases, these levels surpassed LD50 values indicating intoxication events. Predominant substances were clothianidin, coumaphos, imidacloprid, acetamiprid and dimethoate. In less extent, other acaricides such as amitraz (mostly its breakdown products DMF, DMPF), tau-fluvalinate and certain pyrethroids exemplified by cyhalothrin, cypermethrin and deltamethrin were also recorded. In several samples, more than one active substance was detected.Overall, this work aspires to provide valuable insight to pesticides and metabolites occurrence in honeybees in Greece between 2014-2017 and in parallel assist research community and apiculturists in this pivotal Mediterranean region that bee health and pollination services have prolific importance

Headspace Solid Phase Micro Extraction Gas Chromatographic Determination of Fenthion in Human Serum

A simple and effective analytical procedure was developed for the determination of fenthion residues in human serum samples. The sample treatment was performed using the headspace solid-phase micro extraction with polyacrylate fiber, which has the advantage to require low amount of serum (1 mL) without tedious pre-treatment. The quantification of fenthion was carried out by gas chromatography-mass spectrometry and the recoveries ranged from 79 to 104% at two spiking levels for 6 replicates. Detection and quantification limits were calculated as 1.51 and 4.54 ng/mL of serum respectively. Two fenthion metabolites fenoxon and fenthion–sulfoxide were also identified

Standard addition method based on four-way PARAFAC decomposition to solve the matrix interferences in the determination of carbamate pesticides in lettuce using excitation–emission fluorescence data

The simultaneous determination of two carbamate pesticides (carbaryl and carbendazim) and of the degradation product of carbaryl (1-naphthol) in iceberg lettuce was achieved by means of PARAFAC decomposition and excitation–emission fluorescence matrices. A standard addition method for a calibration based on four-way data was applied using different dilutions of the extract from iceberg lettuce as a fourth way that provided the enough variation of the matrix to carry out the four-way analysis. A high fluorescent overlapping existed between the three analytes and the fluorophores of the matrix. The identification of two fluorescent matrix constituents through the four-way model enabled to know the matrix contribution in each dilution of the extract. This contribution was subtracted from the previous signals and a subsequent three-way analysis was carried out with the tensors corresponding to each dilution. The PARAFAC decomposition of these resulting tensors showed a CORCONDIA index equal to 99%. For the identification of the analytes, the correlation between the PARAFAC spectral loadings and the reference spectra has been used. The trueness of the method, in the concentration range studied, was guaranteed because there was neither constant nor proportional bias according to the appropriate hypothesis tests. The best recovery percentages were obtained with the data from the most diluted extract, being the results: 127.6% for carbaryl, 125.55% for carbendazim and 87.6% for 1-naphthol. When the solvent calibration was performed, the decision limit (CCα) and the capability of detection (CCβ) values, in x0=0, were 2.21 and 4.38 μg L−1 for carbaryl, 4.87 and 9.64 μg L−1 for carbendazim; and 3.22 and 6.38 μg L−1 for 1-naphthol, respectively, for probabilities of false positive and false negative fixed at 0.05. However, these values were 5.30 and 10.49 μg L−1 for carbaryl, 18.05 and 35.73 μg L−1 for carbendazim; and 1.92 and 3.79 μg L−1 for 1-naphthol, respectively, when the matrix-matched calibration using the most diluted extract was carried out in the recovery study.Ministerio de Economía y Competitividad(CTQ2011-26022) and JuntadeCastillayLeón(BU108A11-2)

Introducing the INSIGNIA project: environmental monitoring of pesticide use through honey bees

INSIGNIA aims to design and test an innovative, non-invasive, scientifically proven citizen science environmental monitoring protocol for the detection of pesticides by honey bees. It is a 30-month pilot project initiated and financed by the EC (PP-1-1-2018; EC SANTE). The study is being carried out by a consortium of specialists in honey bees, apiculture, statistics, analytics, modelling, extension, social science and citizen science from twelve countries. Honey bee colonies are excellent bio-samplers of biological material such as nectar, pollen and plant pathogens, as well as non-biological material such as pesticides or airborne contamination. Honey bee colonies forage over a circle of 1 km radius, increasing to several km if required, depending on the availability and attractiveness of food. All material collected is accumulated in the hive.The honey bee colony can provide four main matrices for environmental monitoring: bees, honey, pollen and wax. Because of the non-destructive remit of the project, for pesticides, pollen is the focal matrix and used as trapped pollen and beebread in this study. Although beeswax can be used as a passive sampler for pesticides, this matrix is not being used in INSIGNIA because of its polarity dependent absorbance, which limits the required wide range of pesticides to be monitored. Alternatively, two innovative non-biological matrices are being tested: i) the “Beehold tube”, a tube lined with the generic absorbent polyethylene-glycol PEG, through which hive-entering bees are forced to pass, and ii) the “APIStrip” (Absorbing Pesticides In-hive Strips) with a specific pesticide absorbent which is hung between the bee combs.Beebread and pollen collected in pollen traps are being sampled every two weeks to be analysed for pesticide residues and to record foraging conditions. Trapped pollen provides snapshots of the foraging conditions and contaminants on a single day. During the active season, the majority of beebread is consumed within days, so beebread provides recent, random sampling results. The Beehold tube and the APIStrips are present throughout the 2-weeks sampling periods in the beehive, absorbing and accumulating the incoming contaminants. The four matrices i.e. trapped pollen, beebread, Beehold tubes and APIStrips will be analysed for the presence of pesticides. The botanical origin of trapped pollen, beebread and pollen in the Beehold tubes will also be determined with an innovative molecular technique. Data on pollen and pesticide presence will then be combined to obtain information on foraging conditions and pesticide use, together with evaluation of the CORINE database for land use and pesticide legislation to model the exposure risks to honey bees and wild bees. All monitoring steps from sampling through to analysis will be studied and rigorously tested in four countries in Year 1, and the best practices will then be ring-tested in nine countries in Year 2. Information about the course of the project, its results and publications will be available on the INSIGNIA website www.insignia-bee.eu and via social media: on Facebook (https://www.facebook.com/insigniabee.eu/); Instagram insignia_bee); and Twitter (insignia_bee). Although the analyses of pesticide residues and pollen identification will not be completed until December 2019, in my talk I will present preliminary results of the Year 1 sampling.info:eu-repo/semantics/publishedVersio

Introducing the INSIGNIA project: Environmental monitoring of pesticides use through honey bees

INSIGNIA aims to design and test an innovative, non-invasive, scientifically proven citizen science environmental monitoring protocol for the detection of pesticides via honey bees. It is a pilot project initiated and financed by the European Commission (PP-1-1-2018; EC SANTE). The study is being carried out by a consortium of specialists in honey bees, apiculture, chemistry, molecular biology, statistics, analytics, modelling, extension, social science and citizen science from twelve countries. Honey bee colonies are excellent bio-samplers of biological material such as nectar, pollen and plant pathogens, as well as non-biological material such as pesticides or airborne contamination. Honey bee colonies forage over a circle of about 1 km radius, increasing to several km if required depending on the availability and attractiveness of food. All material collected is concentrated in the hive, and the honey bee colony can provide four main matrices for environmental monitoring: bees, honey, pollen and wax. For pesticides, pollen and wax are the focal matrices. Pollen collected in pollen traps will be sampled every two weeks to record foraging conditions. During the season, most of pollen is consumed within days, so beebread can provide recent, random sampling results. On the other hand wax acts as a passive sampler, building up an archive of pesticides that have entered the hive. Alternative in-hive passive samplers will be tested to replicate wax as a “pesticide-sponge”. Samples will be analysed for the presence of pesticides and the botanical origin of the pollen using an ITS2 DNA metabarcoding approach. Data on pollen and pesticides will be then be combined to obtain information on foraging conditions and pesticide use, together with evaluation of the CORINE database for land use and pesticide legislation to model the exposure risks to honey bees and wild bees. All monitoring steps from sampling through to analysis will be studied and tested in four countries in year 1, and the best practices will then be ring-tested in nine countries in year 2. Information about the course of the project and its results and publications will be available in the INSIGNIA website www.insignia-bee.eu.info:eu-repo/semantics/publishedVersio

Mixture-risk-assessment of pesticide residues in retail polyfloral honey

[EN] The presence of even tiny quantities of pesticide residues in honey, a traditional healthy product, is a matter of concern for producers, packers and consumers. This paper aims to quantify pesticides in retail brands of polyfloral honey, and to calculate the mixture risk assessment of honey for consumers according to the results obtained from the analyzed samples. A LC-MS/MS multi-residue method was developed and validated for 13 compounds: 11 pesticides (chlorfenvinphos, coumaphos, tau-fluvalinate, amitraz, which are very common in veterinary treatments, and imidacloprid, acetamiprid, simazine, cyproconazole, tebuconazole, chlorpiryphos-methyl, chlorpiryphos, widely used in agricultural practices), and 2 metabolites of amitraz (2,4-DMA and 2,4-DMF). Results showed that the samples contained pesticide residues at different concentrations; however, the MRL in honey for each of the 11 pesticides was never exceeded. The most common were amitraz (from 1 to 50 ¿Ýg/kg) present in 100% of the samples, and coumaphos (up to 14 ¿Ýg/kg) in 63%. The hazard index (HI) for adults was less than 0.002 in all cases, a long way from 1, the value established as the limit of acceptability. Therefore, commercial honey does not represent any significant risk to health. However, considering that residue levels should be present ¡§as low as reasonably achievable¡¿ it is deemed necessary to make an effort to reduce their presence by appropriate agricultural and, above all, beekeeping practices due to acaridae treatments.This study forms part of a project funded by the Ministerio de Economia y Competitividad of Spain (Programa Estatal de Investigacion Desarrollo e Innovacion Orientada a los retos de la sociedad; Project number AGL2013-48646-R), for which the authors are grateful.Juan Borrás, MDS.; Doménech Antich, EM.; Escriche Roberto, MI. (2016). Mixture-risk-assessment of pesticide residues in retail polyfloral honey. Food Control. 67:127-134. https://doi.org/10.1016/j.foodcont.2016.02.051S1271346

Determination of antioxidant compounds in foodstuff

© 2017 Scrivener Publishing LLC. Phenolic compounds, vitamins and carotenoids are naturally found in different foodstuff. These antioxidant compounds play an important role in human health and are of interest for the food, pharmaceutical and cosmeceutical industries. Modern chromatographic and spectrometric techniques have made analysis easier than ever before, but their success depends on the extraction method used. In fact, the different antioxidants are identifified using chromatographic techniques coupled to diff erent specififi c detectors according to the characteristics of each molecule. Beyond their well-known health-promoting effects, antioxidant molecules can also be used to functionalize or preserve the freshness, nutritive value, flflavor and color of foodstuff s, which justify their incorporation into several matrices. In this chapter, the most common antioxidant compounds in foodstuff will be described, as well as the methodologies involved in their extraction, separation, identifification and quantifification. The bioactive properties and industrial applications of these compounds through innovative techniques will also be taken into account.info:eu-repo/semantics/publishedVersio