Simultaneous Analysis of Food Allergens, Including Nuts and Fruits, Using a Triple Quadrupole Mass Spectrometer

Significance of the Topic

Food allergies trigger severe immune responses to specific proteins and require accurate identification of allergenic ingredients for consumer safety and regulatory compliance. Traditional methods such as ELISA, PCR and Western blotting can be limited by cross reactivity, inability to quantify multiple targets simultaneously, and difficulties in protein level determination. The development of an LC MS MS based approach offers high specificity, multiplex capacity and quantitative performance for food allergen analysis.

Objectives and Study Overview

This study aimed to establish and validate a simultaneous LC MS MS method for the quantification of seven regulated allergens and ten equivalent allergens in processed foods. Key goals included selection of allergen specific peptides, optimization of multiple reaction monitoring transitions, calibration curve preparation from 1 ppm, and evaluation of method specificity and linearity in spiked food matrices.

Methodology and Instrumentation

The workflow involved extraction of total proteins from standard allergen powders, enzymatic digestion with trypsin, peptide identification using data dependent acquisition on a Q TOF system, transition development in Skyline, and optimization on a triple quadrupole mass spectrometer. Sample preparation for processed foods consisted of spiking standards, protein extraction, reduction and alkylation, tryptic digestion, cleanup by solid phase extraction, concentration, filtration, followed by LC MS MS analysis.

Used Instrumentation

• Nexera X3 ultrafast liquid chromatograph
• LCMS 8060RX triple quadrupole mass spectrometer
• LCMS 9030 Q TOF mass spectrometer for peptide discovery
• S Traps for protein digestion and Shim pack GIST HP C18 analytical column

Main Results and Discussion

New MRM transitions were successfully developed for walnut and eight additional allergens. Specificity tests showed minimal false positives for all peptides except those from orange, which require further optimization. Calibration curves prepared in allergen free chicken rice matrix from 1 to 10 mg/kg demonstrated excellent linearity (R2 > 0.995) and sensitivity for all targets. Spiked processed food samples yielded clear quantifier and qualifier ion peaks without interference in unspiked controls.

Benefits and Practical Applications

• Single sample preparation and analysis of 17 allergens reduces time and cost
• High specificity through peptide level discrimination minimizes false positives
• Quantitative performance from 1 ppm supports compliance with labeling regulations
• Applicable to quality control in food manufacturing and regulatory testing

Future Trends and Potential Applications

Further refinement of transitions for challenging allergens such as orange will expand coverage. Integration with high throughput sample preparation platforms and automated data processing will enhance routine monitoring. The approach may be extended to emerging allergens and applied in global regulatory environments, research on cross contamination, and allergen risk management in complex food matrices.

Conclusion

The LC MS MS method developed here enables simultaneous quantitative analysis of 17 food allergens with high specificity and sensitivity using a single sample protocol. This tool offers significant advantages for food safety laboratories and supports stringent allergen labeling requirements.

Reference

• Nozomi Maeshima, Manami Kobayashi. Simultaneous Analysis of Food Allergens, Including Nuts and Fruits, Using a Triple Quadrupole Mass Spectrometer. Shimadzu Application News 01-00955-EN, Jan. 2026
• Shimadzu Corporation. Development of a Simultaneous Analysis Method for Allergens in Food Using a Triple Quadrupole Mass Spectrometer. Application News 01-00665-EN