Determination of 2-MCPD and 3-MCPD Fatty Acid Esters in Infant Formula Using an Agilent 8890 GC System with an Agilent 5977B GC/MSD

Importance of the topic

Monochloropropanediol (MCPD) fatty acid esters are heat-induced contaminants found in processed foods, including infant formula. These compounds can release free MCPDs during digestion, raising potential toxicological concerns. Accurate, reliable quantification of 2- and 3-MCPD esters is critical for quality control, regulatory compliance and safeguarding infant health.

Objectives and Study Overview

This study aimed to develop and validate a robust gas chromatography–mass spectrometry (GC/MS) method using an Agilent 8890 GC system coupled with a 5977B MSD for simultaneous determination of 2- and 3-MCPD fatty acid esters in infant formula. Two derivatization approaches, heptafluorobutyrylimidazole (HFBI) and phenylboronic acid (PBA), were compared to optimize sensitivity, selectivity and ease of use.

Methodology

The workflow comprises four main steps:

• Sample extraction: QuEChERS salt packet and hexane extraction with Geno/Grinder agitation.
• Acid transesterification: Sulfuric acid/methanol cleavage at 40 °C for 16 hours.
• Sample cleanup: Supported liquid extraction (Agilent Chem Elut S) followed by solvent evaporation.
• Derivatization: HFBI (75 °C, 30 min) or PBA (room temperature, 5 min) prior to GC/MS analysis.

Instrumentation

The analytical setup included:

• Agilent 8890 GC with split/splitless inlet (280 °C) and DB-5ms Ultra Inert column (30 m × 0.25 mm, 1 µm film).
• Agilent 5977B GC/MSD operated in electron ionization and selected-ion monitoring (SIM) mode.
• Auxiliary equipment: SPEX Geno/Grinder, Eppendorf centrifuge, ultrasonic bath, and vacuum manifold.
• Chemicals: HFBI, PBA, acid/methanol solution, QuEChERS salts, Chem Elut S cartridges and standard MCPD ester compounds.

Results and Discussion

Both HFBI and PBA workflows achieved baseline separation of target compounds and internal standards. HFBI delivered higher signal intensity but required complete water removal, while PBA offered faster, water-tolerant derivatization. Method performance with HFBI showed linearity over 10–1 000 µg/kg (R² > 0.997), detection limits of 1.30–1.35 µg/kg and recoveries of 86.9–106.7% (RSD < 15%). PBA yielded recoveries of 100–103% (RSD < 10%). Supported liquid extraction improved throughput and minimized emulsions compared to liquid–liquid extraction.

Benefits and Practical Applications

• High sensitivity and specificity for MCPD esters in complex dairy matrices.
• Two derivatization options to suit laboratory constraints and throughput requirements.
• Comprehensive method validation supports routine quality control and regulatory monitoring.

Future Trends and Potential Applications

Emerging trends include direct LC-MS/MS assays for intact esters, automation of sample preparation, integration with high-throughput screening platforms, and extension to other processing contaminants (e.g., glycidol esters). Real-time monitoring and miniaturized workflows may further enhance food safety analytics.

Conclusion

The developed GC/MS method using Agilent 8890/5977B provides reliable, validated workflows for simultaneous quantification of 2- and 3-MCPD fatty acid esters in infant formula. HFBI and PBA derivatization each offer distinct advantages, enabling flexible implementation for routine food safety testing.

Reference

1. AOCS. 2- and 3-MCPD Fatty Acid Esters and Glycidol Fatty Acid Esters in Edible Oils and Fats by Acid Transesterification. 2013.
2. AOCS. Determination of Bound Monochloropropanediol- and Bound 2,3-Epoxy-1-Propanol by GC/MS. 2013.
3. AOCS. Fatty-Acid-Bound 3-MCPD and 2,3-Epoxypropane-1-ol Determination in Oils and Fats by GC/MS. 2013.
4. ISO 18363-1. Animal and Vegetable Fats and Oils—Part 1: Fast Alkaline Transesterification. 2015.
5. ISO 18363-2. Methods Using Slow Alkaline Transesterification. 2018.
6. ISO 18363-3. Acid Transesterification and Measurement. 2017.
7. GB 5009.191-2016. National Food Safety Standard for Chloropropanediols. 2016.
8. SN/T 5220-2019. Determination of 3-MCPDE and Glycidol Esters in Food. 2019.
9. Arisseto A., Marcolino P. Determination of 3-MCPD Diesters and Monoesters in Vegetable Oils. Agilent application note 5991-3406EN. 2013.