An improved UHPLC-MS/MS assay for the determination of acrylamide from foodstuffs

Importance of the Topic

Acrylamide is a probable human carcinogen and reproductive toxicant formed in carbohydrate rich foods during high temperature processing. Regulatory bodies have established benchmark levels across a variety of food products, creating a critical need for rapid, robust analytical methods capable of quantifying trace levels of acrylamide in complex food matrices.

Objectives and Study Overview

This study presents an improved UHPLC-MS/MS assay aimed at quantifying acrylamide in diverse foodstuffs including potato chips, ground coffee, and baby food. The goals were to enhance sample clean-up, minimize matrix effects, increase throughput, and demonstrate applicability to additional heat-processed foods.

Methodology

The protocol employs a two-stage extraction. Stage one removes lipids and other nonpolar interferents by water and dichloromethane partitioning. Stage two uses supported liquid extraction (pH 9) in a 96-well plate format with ethyl acetate/tetrahydrofuran elution. Extracts are dried under nitrogen, reconstituted in water, and centrifuged before analysis. UHPLC separation is performed on a Hypercarb column (100×2.1 mm, 5 μm) at 60 °C using a 4.5-minute gradient (0–10% B to 100% B, then re-equilibration) at 0.5 mL/min. MS/MS detection uses a HESI II probe in positive mode monitoring transitions m/z 72→55 for acrylamide and m/z 75→58 for the d3 internal standard.

Instrumentation

• Thermo Scientific Vanquish Horizon UHPLC system
• Thermo Scientific TSQ Endura triple-stage quadrupole mass spectrometer
• Thermo Scientific Hypercarb HPLC column (100×2.1 mm, 5 μm)
• Thermo Scientific HyperSep SLE 96-well plates (200 mg, pH 9)

Main Results and Discussion

The method yielded a linear calibration range of 100–5000 ng/g with 1/x² weighting and an LLOQ of 100 ng/g. Accuracy bias remained within ±13% and precision CV below 5% across matrices. Absolute recovery ranged from 82.5% to 106%, and no ion suppression or carryover was observed. Specificity was confirmed by absence of interfering peaks in blanks. Application to burnt toast samples demonstrated concentrations of approximately 150 ng/g in white toast and 318 ng/g in brown toast, illustrating the method’s suitability for diverse heat-processed foods.

Benefits and Practical Applications

• Two-stage extraction ensures removal of lipids and reduces matrix effects
• High throughput with a 4.5-minute run time
• Automation potential using 96-well SLE format
• Wide applicability across varied food matrices including snacks, beverages, and baby foods

Future Trends and Possibilities

Further developments may include integration of online extraction workflows, coupling with high resolution mass spectrometry for broader contaminant screening, miniaturization of sample preparation, and extension to a wider spectrum of heat-processed products to support evolving regulatory requirements.

Conclusion

The improved UHPLC-MS/MS assay provides a fast, robust, and sensitive solution for quantifying acrylamide in complex food matrices, meeting the demands of regulatory monitoring and quality control laboratories.

References

• Woodmansey K Application Note 22061 Thermo Fisher Scientific 2020