vMethod™ Application for Food Testing

Significance of the topic

Accurate and sensitive detection of undeclared meat species in food products is critical for consumer protection, religious compliance, and ethical transparency. Regulatory agencies set a 1% (w/w) threshold for cross‐contamination, driving the need for reliable analytical methods capable of identifying multiple meat types in both raw and processed matrices.

Objectives and Study Overview

This study presents a single‐injection LC-MS/MS workflow on the QTRAP® 4500 system for simultaneous screening and speciation of six complementary meat species (pork, beef, lamb, chicken, duck, horse). The method aims to achieve a detection limit of 1% (w/w) in complex food matrices, supporting both qualitative screening and relative quantitation.

Methodology and Instrumentation

Sample Preparation:

• Homogenization of 10 g meat or product, optional defatting with hexane.
• Protein extraction with ammonium bicarbonate buffer, reduction at 60 °C and alkylation of cysteines.
• Tryptic digestion (4–12 h), quenching with formic acid, desalting on Cleanert PEP SPE cartridges.

LC Separation:

• ExionLC™ AC system with Phenomenex Kinetex C18 column (100 × 4.6 mm, 2.6 μm).
• 15-minute linear gradient (0.1% formic acid in water/acetonitrile) at 500 μL/min, 10 μL injection.

MS Acquisition and Data Analysis:

• TripleTOF® 6600 employed for protein identification and selection of candidate peptides via IDA and ProteinPilot™ database search.
• Skyline and NCBI BLAST to confirm species-specific signature peptides.
• Scheduled MRM™ on QTRAP® 4500 with Turbo V source (positive ESI, 650 °C) to monitor 48 transitions (two peptides per protein, two transitions per peptide).

Main Results and Discussion

Signature Peptide Panel:

• Selection of two unique proteins and two peptides per protein for each species (total 24 peptides).
• Scheduled MRM reduces concurrent transitions, maximizes dwell time and sensitivity.

Stability and Precision:

• All peptide markers detected with consistent sensitivity in raw vs. pan-fried meat mixtures.
• Calibration curves from 0–100% (w/w) demonstrated linearity (r2 > 0.99) for all species.
• Reliable detection at 1% (w/w) in combined matrix, with no interference in 0% controls.

Screening of Commercial Samples:

• Halal‐certified products confirmed absence of pork peptides.
• Positive identification aligned with declared ingredients in non-halal sausages.

Benefits and Practical Applications

This LC-MS/MS approach surpasses PCR and ELISA in processed meat analysis, offering high specificity, robust multiplexing, and applicability across raw and heat-treated samples. The single-injection format and vMethod™ package streamline sample preparation, data acquisition, and reporting for routine quality control and food authenticity laboratories.

Future Trends and Potential Applications

Expanding the signature peptide library to additional species, integrating ultra-high-sensitivity platforms (e.g., QTRAP® 6500+), and automating data workflows will support higher throughput and lower detection limits. Coupling with advanced software for real-time screening could further enhance surveillance against food fraud.

Conclusion

A robust LC-MS/MS method on the QTRAP® 4500 system enables reliable detection of six meat species at regulatory thresholds in a single injection. The workflow balances sensitivity, specificity, and practicality for broad application in food authenticity testing.

References

1. Department for Environment Food & Rural Affairs. Processed beef products and horse meat. 2013.
2. Wearne S. Setting a threshold for contamination of processed meat products with undeclared meat species. Food Standards Agency, 2014.