Jens Hinge Andersen

Optimization of solid phase extraction clean up and validation of quantitative determination of corticosteroids in urine by liquid chromatography-tandem mass spectrometry

A solid phase extraction (SPE) method for extraction and clean up of 9 synthetic corticosteroids was optimized for quantification by reversed-phase high-performance liquid chromatography/negative electrospray ionisation mass spectrometry. Clean up was accomplished using a mixed mode polymeric strong anion exchange SPE column. The final method was validated according to EU regulations for determination of residues of veterinarian drugs in products of animal origin. Initial results showed a large difference in ion suppression between samples of porcine and bovine urine. The aim of optimisation was to design a procedure that minimised this difference while using a single SPE procedure and a fast HPLC method that enabled sufficient separation of the epimers beta- and dexamethasone. To include conjugated corticosteroids in the analysis, the sample was hydrolysed with Helix Pomatia beta-glucuronidase/aryl sulfatase. For the final method, which included fluocinolone acetonid, triamcinolone acetonid, beclomethasone, flumethasone, dexamethasone, betamethasone, 6alpha-methylprednisolone, prednisone and prednisolone, a quantification based on spiked samples carried through the entire analytical procedure was used. For quantification of triamcinolone acetonid an internal standard (triamcinolone acetonid-D6) was used. Relative average recoveries from 96 to 103% were found, except for beclomethasone (113%). Absolute average recoveries were 81-99%. Quantification limits (decision limits, CCalpha) were demonstrated to be not higher than 1 microg L(-1) (3 microg L(-1) for prednisone and prednisolone). The internal reproducibility, determined by triplicates from spiking at three different levels in six analytical series was 7-19% (at 2-4 microg L(-1)) except for prednisone and prednisolone (26-27% at 3-6 microg L(-1)).

Results from the monitoring of pesticide residues in fruit and vegetables on the Danish market, 1998?99

To check compliance with the maximum residue levels in foods and to monitor the residue levels to enable an evaluation of the exposure of the Danish population to pesticides, a monitoring programme for pesticides residues in fruit and vegetables was performed. Sampling plans were designed based on previous findings and on consumption data. Samples (n = 4404) of mainly fresh conventionally and organically grown fruit and vegetables were collected at wholesalers and importers and at food processing companies. Of the samples, 3% were frozen products and 5% organically grown. Of the samples, 34% were of Danish origin, with 66% from other countries. Two accredited laboratories performed analyses of the samples using capillary gas chromatography, liquid chromatography and a spectrophotometric method. Reporting levels were set at the lowest calibration level or at the limit of determination. Residues were found in 60% of the samples of fruit but in only 18% of the vegetable samples. Residues above the maximum residue levels were found in 6% of all samples of fruit and in 2% of the vegetable samples.

Survey of pesticide residues in table grapes: Determination of processing factors, intake and risk assessment

The differences in residue pattern between Italy and South Africa, the main exporters of table grapes to the Danish market, were investigated. The results showed no major differences with respect to the number of samples with residues, with residues being found in 54–78% of the samples. Exceedances of the European Union maximum residue limit (MRL) were found in five samples from Italy. A number of samples were rinsed to study the possible reduction of residues. For copper, iprodione, procymidone and dithiocarbamates a significant effect of rinsing was found (20–49% reduction of residues). However, no significant effect was found for organophosphorus pesticides and pyrethroids, whereas the number of samples with residues of benzilates, phenylamids and triazoles was insufficient to demonstrate any significant effects. An intake calculation showed that the average intake from Italian grapes was 3.9 µg day−1 for pesticides and 21 µg day−1 for copper. Correspondingly, the intakes from South African grapes were 2.6 and 5.7 µg day−1, respectively. When the total exposure of pesticides from grapes were related to acceptable daily intake, expressed as the sum of Hazard Quotients, the exposure were approximately 0.5% for Italian samples and 1% for South African samples.

Dietary exposure assessment of Danish consumers to dithiocarbamate residues in food. A comparison of the deterministic and probabilistic approach

Probabilistic and deterministic estimates of the acute and chronic exposure of the Danish populations to dithiocarbamate residues were performed. The Monte Carlo Risk Assessment programme (MCRA 4.0) was used for the probabilistic risk assessment. Food consumption data were obtained from the nationwide dietary survey conducted in 2000–02. Residue data for 5721 samples from the monitoring programme conducted in the period 1998–2003 were used for dithiocarbamates, which had been determined as carbon disulphide. Contributions from 26 commodities were included in the calculations. Using the probabilistic approach, the daily acute intakes at the 99.9% percentile for adults and children were 11.2 and 28.2 μg kg−1 body weight day−1, representing 5.6% and 14.1% of the ARfD for maneb, respectively. When comparing the point estimate approach with the probabilistic approach, the outcome of the point estimate calculations was generally higher or comparable with the outcome of the probabilistic approach at the 99.9 percentile (consumers only). The chronic exposures for adults and children were 0.35 and 0.76 μg kg−1 body weight day−1 at the 99.9 percentile, representing 0.7% and 1.5%, respectively, of the acceptable daily intake for mancozeb and maneb at 50 μg kg−1 body weight.

Cumulative dietary exposure of the population of Denmark to pesticides.

We used the Hazard Index (HI) method to carry out a cumulative risk assessment after chronic dietary exposure to all monitored pesticides in fruit, vegetables and cereals for various consumer groups in Denmark. Residue data for all the pesticides were obtained from the Danish monitoring programme during the period 2004-2011. Food consumption data were obtained from DANSDA (the DAnish National Survey of Diet and physical Activity) for the period 2005-2008. The calculations were made using three different models to cope with residues below the limit of reporting (LOR). We concluded that a model that included processing factors and set non-detects to ½ LOR, but limited the correction (Model 3), gave the most realistic exposure estimate. With Model 3 the HI was calculated to be 0.44 for children and 0.18 for adults, indicating that there is no risk of adverse health effects following chronic cumulative exposure to the pesticides found in fruit, vegetables and cereals on the Danish market. The HI was below 1 even for consumers who eat more than 550 g of fruit and vegetables per day, corresponding to 1/3 of the population. Choosing Danish-produced commodities whenever possible could reduce the HI by a factor of 2.

Confirmatory analysis of steroids in muscle using liquid chromatography-tandem mass spectrometry

A method is described for screening and confirmation of synthetic and endogenous steroids in muscle tissue. The method is sensitive, selective, and rapid and the consumption of organic solvents is low, compared to previously published methods. The procedure involves hydrolysis, defattening with heptane and final clean up with SPE using C18 cartridge. After filtration, the analytes are analysed by LC/MS/MS and quantification is performed using deuterated internal standards. Decision limits (CCα) varied from 0.02 to 0.33 µg kg−1 and the detection capabilities (CCβ) were <0.50 µg kg−1. The mean within-laboratory reproducibility ranged 5–22% (%RSDIR). Endogenous steroids (e.g. testosterone, epitestosterone and androstenedione) have been included in the method, to provide an insight into their levels, as the presence of these steroids was detected several times during analysis of imported meat.

An intercomparison study of the determination of glyphosate, chlormequat and mepiquat residues in wheat

An intercomparison study of the determinations of glyphosate, chlormequat and mepiquat residues in cereals was performed. Four samples comprising one blank, two incurred and one spiked sample were sent to six participating laboratories. For glyphosate, two laboratories reported considerably lower results than the other four. One of the two laboratories with low results also reported low recoveries. The results of a sample spiked with 0.80 mg kg−1 glyphosate and an incurred sample, ranged from 0.23–0.87 mg kg−1 and 0.11–0.25 mg kg−1 respectively. The strong correlation between the two samples (r 2 = 0.95) indicates a systematic between-laboratory variation. Several different principles were used for the analysis of glyphosate using different clean-up techniques and GC/MS, HPLC-fluorescence or LC/MS for detection. The results of the chlormequat residues showed more consistency. All but one laboratory obtained comparable results. However the correlation between the results for the sample spiked with 0.38 mg kg−1 (range: 0.26–0.65 mg kg−1) and the incurred samples (range: 0.19–0.45 and 0.15–0.23 mg kg−1, respectively) again showed a strong correlation (r 2 = 0.99 and 0.88) indicating a systematic component. For mepiquat, results above the limit of quantification were only reported for the spiked sample. The results ranged from 0.29–0.92 mg kg−1 (spiked concentration = 0.38 mg kg−1). Three laboratories had results that deviated less than 25% from the fortified concentration. Two laboratories reported results 38% and 141% above the fortified concentration, respectively.

Optimised deconjugation of androgenic steroid conjugates in bovine urine

ABSTRACT After administration of steroids to animals the steroids are partially metabolised in the liver and kidney to phase 2 metabolites, i.e., glucuronic acid or sulphate conjugates. During analysis these conjugated metabolites are normally deconjugated enzymatically with aryl sulphatase and glucuronidase resulting in free steroids in the extract. It is well known that some sulphates are not deconjugated using aryl sulphatase; instead, for example, solvolysis can be used for deconjugation of these aliphatic sulphates. The effectiveness of solvolysis on androgenic steroid sulphates was tested with selected aliphatic steroid sulphates (boldenone sulphate, nortestosteron sulphate and testosterone sulphate), and the method was validated for analysis of androgenic steroids in bovine urine using free steroids, steroid sulphates and steroid glucuronides as standards. Glucuronidase and sulphuric acid in ethyl acetate were used for deconjugation and the extract was purified by solid-phase extraction. The final extract was evaporated to dryness, re-dissolved and analysed by LC-MS/MS.

Pesticide Residues in Commercial Lettuce, Onion, and Potato Samples From Bolivia—A Threat to Public Health?

Bolivia does not have a surveillance program for pesticide residues in food. The few published studies have suggested that pesticide contamination in food may present a public health problem. Data are lacking for all foods except tomatoes and breast milk. In this study 10 potato, 10 onion, and 10 lettuce samples from La Paz were sampled on August 15, 2015 at a local market and screened for 283 pesticides. Residues of cypermethrin, chlorpyrifos, difenoconazol, or/and λ-cyhalothrin were detected in 50% of the lettuce samples, whereas no pesticides were found in potatoes and onions. In 20% of the lettuce samples, the measurements were above the maximum residue limits, and 2 or 3 pesticides were identified simultaneously. Washing almost halved the pesticide levels, but still 20% of the samples showed measurements above the limits. No samples contained concentrations of pesticides which alone or together would lead to exposures that exceeded the acceptable daily intake or the acute reference dose. To protect consumers from pesticide poisonings and chronic effects, the development of measures for prevention, control, and monitoring of food contamination by pesticides in Bolivia is suggested.