Determination of Bisphenol A in Infant Formula by Automated Sample Preparation and Liquid Chromatography-Mass Spectrometry

Significance of the topic

A sensitive and rapid method for detecting Bisphenol A (BPA) in infant formula is crucial given the compound’s prevalence in plastic food containers and its known health risks, especially for infants. Low-level exposure has been linked to endocrine disruption, metabolic disorders, and developmental issues, driving the need for reliable analytical techniques in food safety laboratories.

Objectives and Study Overview

The study aimed to develop a fully automated sample preparation and liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for trace-level quantitation of BPA in canned infant formula powder. Key goals included reducing sample handling, improving throughput, and achieving low parts-per-billion detection limits using a deuterated internal standard (BPA-d16).

Methodology and Instrumentation

Sample Preparation:

• Infant formula powder was spiked with BPA standards and BPA-d16 internal standard.
• Each gram of powder was extracted with 10 mL of 3 % ammonium acetate in 70:30 acetonitrile–water and centrifuged at 10 000 rpm for 30 minutes.
• Serial dilutions yielded calibration standards ranging from 0.78 to 1000 ng/mL BPA.

Automated Online Cleanup and Analysis:

• Thermo Scientific Aria TLX-1 system equipped with TurboFlow Cyclone P extraction column for removal of large matrix molecules.
• Analyte captured on the extraction column, then back-flushed onto a Hypersil GOLD aQ C18 analytical column (4 × 50 mm, 3 µm).
• Gradient elution delivered to a Thermo Scientific TSQ Vantage triple quadrupole MS with APCI in negative ion mode.
• Monitored transitions: BPA m/z 227→133 for quantitation and 227→212 for confirmation; BPA-d16 m/z 241→223 for internal standard.
• Total run time: 5.6 minutes per injection, 25 µL injection volume.

Instrument Parameters:

• APCI discharge current: 4.0 µA, vaporizer 60 °C, capillary 275 °C.
• Sheath gas 30 units, auxiliary gas 5 units, sweep gas 2 units (N₂).
• Collision gas: 1.2 mTorr; collision energies: 27 V (227→133) and 19 V (227→212).

Main Results and Discussion

The method demonstrated a lower limit of quantitation (LLOQ) of 0.78 ng/mL and a dynamic range up to 1000 ng/mL with r² > 0.992. Repeatability was high, with %CV < 20 % at the LLOQ and < 10 % at higher concentrations. Background subtraction using double-blank correction effectively removed pervasive BPA contamination. Matrix interference at 12.5 and 25 ng/mL was minimal. Comparison with an alternative online SPE approach showed similar preparation times but a fourfold shorter LC-MS/MS run time with TurboFlow technology.

Benefits and Practical Applications

The fully automated protocol eliminates laborious offline extraction steps, reducing operator variability and contamination risk. High throughput (5.6 min per sample) and low detection limits (7.8 µg/kg dry powder LOD; 31.3 µg/kg LLOQ) make this method well suited for routine QA/QC screening of BPA in infant formula and other complex food matrices.

Future Trends and Applications

• Implementation of multiplexed Aria TLX-2 or TLX-4 systems to further increase sample throughput.
• Extension to simultaneous monitoring of other bisphenols and phenolic contaminants.
• Integration with high-resolution MS for broader non-targeted screening.
• Miniaturization of extraction columns for reduced solvent consumption and waste.

Conclusion

A rapid, robust, and sensitive automated LC-MS/MS method using TurboFlow online cleanup has been established for BPA determination in infant formula. The protocol offers high accuracy, low detection limits, minimal matrix effects, and significantly reduced sample preparation time compared to traditional approaches.

References

• US Food and Drug Administration. www.fda.gov
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• Ye X, Kuklenyik Z, Needham LL, Calafat AM. Measuring environmental phenols and chlorinated organic chemicals in breast milk using automated on-line column-switching HPLC-isotope dilution tandem MS. J Chromatogr B. 831 (2006) 110–115.