Miniaturized multiresidue method for determination of 267 pesticides, their metabolites and polychlorinated biphenyls in low mass beebread samples by liquid and gas chromatography coupled with tandem mass spectrometry.

Current work presents developed and validated miniaturized method for residue analysis of 261 pesticides and their metabolites as well as 6 congeners of non-dioxin like polychlorinated biphenyls (ndl-PCB) in a very low mass beebread sample. Sample preparation is based on modified QuEChERS protocol with all steps miniaturized to enable multiresidue analysis of sample with extremely low weight. Sample of beebread (0.3 g) was extracted with 1 mL of acetonitrile containing 5% formic acid and ammonium formate salt were added, then extract was subjected to clean-up by freezing and two-step dispersive solid phase extraction (dSPE) with a Supel QuE Verde sorbents (Supelclean ENVI-Carb Y; Supelclean PSA; Z-Sep+; magnesium sulfate). After 1st step dSPE a portion of extract was analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS) for 200 pesticide residues. Remaining extract was subjected to 2nd step dSPE clean-up by another Supel QuE Verde and then after concentration and solvent exchange it was analyzed by gas chromatography tandem mass spectrometry (GC-MS/MS) for another 61 pesticide and 6 ndl-PCB residues. Method enables determination of residues of 101 insecticides, 72 herbicides, 67 fungicides, 10 acaricides, 6 growth regulators, 5 veterinary drugs and 6 ndl-PCB's. Particular attention was paid to the pesticides being active substances of plant protection products recommended for the protection of winter oilseed rape and apple orchards which during their blooming periods are one of the most attractive sources of food for pollinators and could serve as representatives of other economically important crops. Method was validated according to the Guidance document SANTE/12682/2019 at six concentration levels from 0.001 to 0.5 mg kg-1. The analysis of beebread samples spiked at the level of 0.01 mg kg-1showed mean recovery (trueness) value of about 98% and RSDr (precision) below 20%. The small weight of the sample did not adversely affect the limits of quantification and 75% of analytes could be quantified at least at concentration of 0.005 mg kg-1. Developed mini-method was tested in the analysis of beebread samples, each extracted from individual cell of honeycomb. It is the first time when analyses at single comb cell level were possible.

Determination of neonicotinoids and 199 other pesticide residues in honey by liquid and gas chromatography coupled with tandem mass spectrometry.

Current work presents a modified QuEChERS method for the determination of 207 pesticide residues in honey by LC-MS/MS and GC-MS/MS. Acetate buffered acetonitrile extraction with Z-Sep+ and PSA dispersive-SPE clean-up were used for sample preparation. Optimised conditions allows determination of neonicotinoids as well as other insecticides, fungicides, herbicides, acaricides, growth regulators and veterinary drugs in honey samples. Validated method enable sensitive analysis at least at concentrations from 0.001, 0.005 or 0.01 mg/kg for 45%, 41% and 14% of pesticides, respectively. Method was utilised for the analysis of 155 honey samples from Poland during 2015-2017. Residues of 21 pesticides were determined in honey. Cyano-substituted neonicotinoids (acetamiprid, thiacloprid) were quantified in 77% of samples and were the most frequently detected pesticides. Concentrations of acetamiprid was from 0.001 to 0.13 mg/kg whilst thiacloprid from 0.001 to 0.2 mg/kg. Fungicides were determined in 50% and amitraz metabolites in 35% of honey samples.

Multi-residue method for the determination of pesticides and pesticide metabolites in honeybees by liquid and gas chromatography coupled with tandem mass spectrometry--Honeybee poisoning incidents.

A method for the determination of 200 pesticides and pesticide metabolites in honeybee samples has been developed and validated. Almost 98% of compounds included in this method are approved to use within European Union, as active substances of plant protection products or veterinary medicinal products used by beekeepers to control mites Varroa destructor in hives. Many significant metabolites, like metabolites of imidacloprid, thiacloprid, fipronil, methiocarb and amitraz, are also possible to detect. The sample preparation was based on the buffered QuEChERS method. Samples of bees were extracted with acetonitrile containing 1% acetic acid and then subjected to clean-up by dispersive solid phase extraction (dSPE) using a new Z-Sep+ sorbent and PSA. The majority of pesticides, including neonicotionoids and their metabolites, were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS) but some of pesticides, especially pyrethroid insecticides, were analyzed by gas chromatography tandem mass spectrometry (GC-MS/MS). The procedure was validated according to the Guidance document SANCO/12571/2013 at four concentration levels: 1, 5, 10 and 100 ng/g bees and verified in the international proficiency test. The analysis of bee samples spiked at the limit of quantification (LOQ) showed about 98% mean recovery value (trueness) and 97% of analytes showed recovery in the required range of 70-120% and RSDr (precision) below 20%. Linearity and matrix effects were also established. The LOQs of pesticides were in the range of 1-100 ng/g. The developed method allows determination of insecticides at concentrations of 10 ng/g or less, except abamectin and tebufenozide. LOQ values are lower than the median lethal doses LD50 for bees. The method was used to investigate more than 70 honeybee poisoning incidents. Data about detected pesticides and their metabolites are included.