This paper describes the validation of an LC-MS/MS-based method for the quantification of > 500 secondary microbial metabolites. Analytical performance parameters have been determined for seven food matrices using seven individual samples per matrix for spiking. Apparent recoveries ranged from 70 to 120% for 53–83% of all investigated analytes (depending on the matrix). This number increased to 84–94% if the recovery of extraction was considered. The comparison of the fraction of analytes for which the precision criterion of RSD ≤ 20% under repeatability conditions (for 7 replicates derived from different individual samples) and intermediate precision conditions (for 7 technical replicates from one sample), respectively, was met (85–97% vs. 93–94%) highlights the contribution of relative matrix effects to the method uncertainty. Statistical testing of apparent recoveries between pairs of matrices exhibited a significant difference for more than half of the analytes, while recoveries of the extraction showed a much better agreement. Apparent recoveries and matrix effects were found to be constant over 2–3 orders of magnitude of analyte concentrations in figs and maize, whereas the LOQs differed less than by a factor of 2 for 90% of the investigated compounds. Based on these findings, this paper discusses the applicability and practicability of current guidelines for multi-analyte method validation. Investigation of (apparent) recoveries near the LOQ seems to be insufficiently relevant to justify the enormous time-effort for manual inspection of the peaks of hundreds of analytes. Instead, more emphasis should be put on the investigation of relative matrix effects in the validation procedure.

This work provides a proposal for proper determination of matrix effects and extraction efficiencies as integral part of full validation of LC-MS/MS based multi-class methods for complex feedstuff. Analytical performance data have been determined for 100 selected analytes in three compound feed matrices and twelve single feed ingredients using seven individual samples per matrix type. Apparent recoveries ranged from 60–140% for 52–89% of all compounds in single feed materials and 51–72% in complex compound feed. Regarding extraction efficiencies, 84–97% of all analytes ranged within 70–120% in all tested feed materials, implying that signal suppression due to matrix effects is the main source for deviation from 100% of the expected target deriving from external calibration. However, the comparison between compound feed and single feed materials shows great variances regarding the apparent recoveries and matrix effects. Therefore, model compound feed formulas for cattle, pig and chicken were prepared in house in order to circumvent the issue of the lack of true blank sample material and to simulate compositional uncertainties. Results of this work highlight that compound feed modelling enables a more realistic estimation of the method performance and therefore should be implemented in future validation guidelines.

Monitoring of food contaminants and residues has undergone a significant improvement in recent years and is now performed in an intensive manner. Achievements in the area of chromatography-mass spectrometry coupling techniques enabled the development of quantitative multi-target approaches covering several hundred analytes. Although the majority of methods are focusing on the analysis of one specific group of substances, such as pesticides, mycotoxins, or veterinary drugs, current trends are going towards the simultaneous determination of multiclass compounds from several families of contaminants and residues. This work provides an overview of relevant multiclass concepts based on LC-MS/MS and LC-HRMS instruments. Merits and shortcomings will be critically discussed based on current performance characteristics of the EU legislation system. In addition, the discussion of a recently developed multiclass approach covering >1000 substances is presented as a case study to illustrate the current developments in this area.

An informed opinion to a hugely important question, whether the food on the Europeans’ plate is safe to eat, is provided. Today, the Europeans face food-borne health risks from non-communicable diseases induced by excess body weight, outbreaks caused by pathogens, antimicrobial resistance and exposures to chemical contaminants. In this review, these risks are first put in an order of importance. Then, not only potentially injurious dietary chemicals are discussed but also beneficial factors of the food. This review can be regarded as an attempt towards a dietary-exposome evaluation of the chemicals, the average European adult consumers could chronically expose to during their life-times. Risk ranking reveals that currently the European adults are chronically exposed to a mixture of potentially genotoxic-carcinogenic contaminants, particularly food process contaminants, at the potential risk levels. Furthermore, several of the contaminants whose dietary exposures pose risks appear to be carcinogens operating with a genotoxic mode of action targeting the liver. This suggests that combined health risks from the exposure to a mixture of the chemical contaminants poses a greater potential risk than the risks assessed for single compounds. Over 100 European-level risk assessments are examined. Finally, the importance of a diversified and balanced diet is emphasized.

The first quantitative multiclass approach enabling the accurate quantification of >1200 biotoxins, pesticides and veterinary drugs in complex feed using liquid chromatography tandem mass spectrometry (LC-MS/MS) has been developed. Optimization of HPLC/UHPLC (chromatographic column, flow rate and injection volume) and MS/MS conditions (dwell time and cycle time) were carried out in order to allow the combination of five major substance classes and the high number of target analytes with different physico-chemical properties. Cycle times and retention windows were carefully optimized and ensured appropriate dwell times reducing the overall measurement error. Validation was carried out in two compound feed matrices according to the EU SANTE validation guideline. Apparent recoveries matching the acceptable range of 60-140% accounted 60% and 79% for all analytes in cattle and chicken feed, respectively. High extraction efficiencies were obtained for all analyte/matrix combinations and revealed matrix effects as the main source for deviation of the targeted performance criteria. Concerning the methods repeatability 99% of all analytes in chicken and 96% in cattle feed complied with the acceptable RSD ≤ 20% criterion. Limits of quantification were between 1-10 µg/kg for the vast majority of compounds. Finally, the methods applicability was tested in >130 real compound feed samples and provides first insights into co-exposure of agro-contaminants in animal feed.