Further development and validation of innovative methods for traceability and authenticity of animal proteins in food and feed

Food and feed are strictly regulated and tightly controlled to ensure consumer safety, animal health, and to detect fraudulent activities. Enabling comprehensive feed and food control requires species- and tissue-specific, quantitative analytical methods. Within the project ANIMAL-ID2, previously established methods combining immunoaffinity enrichment and mass spectrometry able to determine animal proteins were to be validated according to FDAshort forFood and Drug Administration guidelines (being a prerequisite for market introduction): - three methods for qualitative and quantitative tissue and species discrimination of animal material in feed as well as - one additional method for qualitative and quantitative species discrimination in feed. Furthermore, - standard operating procedures are to be written for the established methods and - a test kit prototype is to be developend and tested in a ring trial.

Within the project ANIMAL-ID2, three tissue- and species-specific analytical methods quantifying animal proteins were validated according to United States Food and Drug Administration guidelines and proved to be fit for feed analysis purposes. The methods comply with general requirements regarding sensitivity, specificity, accuracy, and precisionClose dialogprecisionPositive Predictive ValueTo glossary. Remarkably, these methods are the first quantitative and thoroughly validated methods in the field of mass spectrometry-based feed analysis targeting animal protein. Their major advantage is their robustness to the presence of milk and milk products. This is highly relevant as milk proteins are known to often challenge standard mass spectrometric analysis workflows abstaining from selective analyte enrichment (here via immunoaffinity) or milk protein depletion. Furthermore, the methods enable to precisely distinguish non-authorised meat/bone meals from authorised ingredients such as milk powder, which current methods in official control are unable to.Results obtained in the project ANIMAL-ID2 show that the methodology is transferable to analysing food, e.g. minced meat products. However, the accuracy related to relative proportions of muscle and protein from the species included in the method are yet to be confirmed in binary and tertiary mixtures.Conducting a ring trial yielded valuable participant feed-back on test kit and standard operating procedure. Detailed instrument and performance parameters have been collected from the different mass spectrometry platforms available in the participating laboratories and will be used in further test kit development.A key point for further improving the quantitation limits is the sample homogeneity achievable with standard laboratory equipment in relation to the representativity of aliquots taken for analysis. Adressing this by modifying sample preparation protocols accordingly might enable to lower quantitation limits in real samples to the limits technically achieved with standard dilution.