RAPID DETERMINATION OF LACTULOSE IN DAIRY PRODUCTS USING ULTRAPERFORMANCE CONVERGENCE CHROMATOGRAPHY COUPLED WITH MASS SPECTROMETRY

Significance of the Topic

Milk heat treatment can induce conversion of lactose into lactulose, a key marker for assessing thermal processing intensity and product quality. Accurate, rapid quantification of lactulose in dairy matrices such as milk, yogurt, and beverages supports food safety control, authentication of processing methods, and regulatory compliance.

Objectives and Overview of the Study

This work aimed to develop and validate a sensitive ultra-performance convergence chromatography–mass spectrometry (UPC2–MS) assay for rapid determination of lactulose in various dairy products. Specific goals included baseline separation of isomeric sugars (lactulose, lactose, sucrose), low detection limits, and a streamlined sample workflow suitable for quality control laboratories.

Methodology and Instrumental Setup

• Sample Preparation: 0.2 g of homogenized dairy sample extracted with water–methanol–acetonitrile mixture, vortexed, centrifuged, and directly injected.
• Chromatographic Conditions: Waters ACQUITY UPC2 Torus DEA column (3 mm × 100 mm, 1.7 µm), run at 40 °C for 10 min; mobile phase A: supercritical CO₂; B: methanol/water (97/3, v/v) with formic acid; gradient elution and backpressure at 2,000 psi.
• Mass Spectrometry: ACQUITY QDa detector in positive ESI mode, selected ion recording of sodium adduct (m/z 365), capillary voltage 0.8 kV, cone voltage 20 V, probe temperature 600 °C, 5 Hz acquisition.

Main Results and Discussion

• Separation: Lactulose (RT 6.15 min) was resolved from sucrose (RT 6.96 min) and lactose (RT 7.34 min) with resolution > 1.5 and minimal tailing.
• Calibration and Performance: Quadratic calibration over 0.1–10 mg L⁻¹ (r² ≥ 0.99); retention time RSD 0.2 %, peak area RSD 0.5 %.
• Sensitivity: LOD 0.03 mg L⁻¹ in matrix, LOQ 0.1 mg L⁻¹ (equivalent to 0.75 and 2.5 mg L⁻¹ in undiluted samples).
• Recovery and Precision: Average recovery 93 % at 100 mg kg⁻¹ fortification, RSD < 2.1 % for pasteurized milk.
• Application to Commercial Samples: Analysis of milk beverages, yogurts, pasteurized and reconstituted milks showed lactulose presence in most products but concentrations remained below the 600 mg L⁻¹ upper limit for UHT classification.

Benefits and Practical Applications

• Speed: Total runtime of 10 minutes per sample accelerates throughput.
• Efficiency: Reduced solvent use and simplified sample prep compared with HPLC methods.
• Sensitivity and Selectivity: Low detection limits and clear isomer separation support reliable quality control.
• Regulatory Use: Suitable for monitoring heat treatment markers in dairy production and verifying processing claims.

Future Trends and Opportunities

• Integration with high-resolution MS for broader profiling of heat-induced and process-related markers.
• Automation of sample prep and data analysis for real-time in-line monitoring in dairies.
• Application of convergence chromatography to other complex food matrices and isomeric analytes.

Conclusion

The UPC2–MS approach delivers rapid, robust, and sensitive quantification of lactulose in diverse dairy matrices. Its high throughput, minimal solvent usage, and clear separation of isomeric sugars make it well suited for industrial quality control and regulatory compliance in heat-treated milk products.

References

1. E. Marconi et al. Enzymatic and GC/HPLC methods for lactulose in milk. Food Chemistry, 84(3), 447–450 (2004).
2. J.L. Chávez-Servín et al. Chromatographic analysis of dairy sugars. Journal of Chromatography A, 1043(2), 211–215 (2004).
3. European Commission Dairy Chemists’ Group Doc. VI/5726/92. Brussels (1992).
4. IDF/ISO B-Doc. 222. Brussels (1992).
5. IDF/ISO B-Doc. 235. Brussels (1993).