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Development, optimisation and validation of a non-targeted proteomics method for meat species identification - FA0166

This project aimed to develop and validate a multi-species proteomics screening tool for meat species verification in processed meat products. Incidents of food and drink mislabelling and fraud can damage public trust in the integrity of the food chain and impact on the competitiveness and resilience of UK businesses. The development of robust analytical tools to help verify product labelling and support food law enforcement is fundamental to improving confidence in food labelling for consumers, businesses and the food industry. The development of methods and technologies to detect food fraud supports Defra¡¯s policy and vision for a world leading food and drink industry with the highest standards of food safety and quality.
Defra¡¯s food authenticity research programme is at the forefront of using cutting edge technology to develop fit for purpose methods to detect food fraud and help enforcers verify compliance with food labelling law. A recent project (FA0157) successfully assessed the applicability of modern PCR-based technologies for detection and quantitation of meat species. Traditional DNA-based methods tend to be applied in a species-targeted way and so intelligence on the adulterants is needed. Multiplex PCR methods are less advanced due to analytical challenges and higher costs. An alternative approach suitable for food testing is the analysis of proteins and their constituent peptides by liquid chromatography/mass spectrometry (LC-MS). Peptide mass spectrometry analysis is emerging as a powerful tool that can complement and support molecular biology methods. A proteomics method using high resolution mass spectrometry and a bespoke database of meat species-specific peptides was developed under Defra projects FA0138 and FA0163 (Claydon et al., 2015). This project builds on that previous work, optimising the list of species-unique peptides that can be used as markers of individual meat species, and developing and validating a multispecies method for simultaneous detection of nine meat species using mass spectrometry instrumentation available to Public Analysts (liquid chromatography/triple quadrupole mass spectrometry).
The main objectives of the project were to develop a fit for purpose method for meat speciation in processed meat products representative of those available from retailers. For this purpose, a cooking method (rag¨´ sauce, slow cooked for three hours) and ingredients were sourced from industry and utilised to prepare meat products for method validation. In line with the current European Commission recommended action limit for meat species adulteration, the desired limit of detection for all species was 1% (w/w) level. The method covers nine meat species: beef, pork, horse, goat, lamb, donkey, rabbit, chicken and turkey. Beef, pork, horse and chicken have been used for the complete intra-laboratory method validation.
Key outputs of the projects:
- Improvement of the original database of marker peptides for the nine animal species studied. Further work would be required for complete optimisation. Additional information on the peptides identified in this project can be provided on request.
EVID4 Evidence Project Final Report (Rev. 06/11) Page 3 of 28
- Expansion of the high-resolution mass spectrometry work from previous projects to achieve detection of pork, horse, donkey, lamb, rabbit and chicken at 1% (weight in weight) adulteration levels.
- Development and validation of a triple quadrupole mass spectrometry method suitable for simultaneous identification of nine meat species in processed meat products.
- The method has been validated across four separate instruments.
- The limit of detection for all the species is ¡Ü 1% (weight in weight).
- A scoping plan for a future inter-laboratory validation is presented.
- Standard Operating Procedures (SOPs) have been produced for all methods involved.
Options for future work:
- Ring trial of the triple quadrupole method with public analysts to further validate the method.
- Knowledge transfer, e.g. training course, to set up LC-MS methods for meat speciation.
- Transfer of the method to quantitation. The best marker peptides for each species could be used as standards to develop a quantitative method.
- Additional species could be included in the method. The work carried out in this project has demonstrated successful identification of nine species of meats, even when mixed and cooked together, indicating that the triple quadrupole method possesses sufficient sensitivity and specificity. Markers for other species of interest could be investigated and, if proven to provide the required specificity, added to the method.
- The method can be used to confirm speciation results obtained using DNA analysis.
The proposed project builds on a previously commissioned Defra study which scoped out a number of options and the associated laboratory work required for further refining, developing and validating a non-targeted proteomic method for detecting meat species in highly processed foodstuffs. The new project will carry out validation work to ensure that non-targeted proteomic methods are fit for purpose and will include the development of a Standard Operating Procedure (SOP). Validation of this method will allow enforcement authorities and industry to verify the labelling of meat ingredients in food products to detect food fraud and protect consumers.
Project Documents
• EVID4 - Final project report : Evid 4 FA0166 Proteomic Screening Tool Final Report   (1338k)
Time-Scale and Cost
From: 2017

To: 2019

Cost: £149,825
Contractor / Funded Organisations