High-Throughput Determination of Multiple Toxic Alkaloids in Food by UHPLC/MS/MS

Importance of the Topic

Alkaloid residues in food represent a significant safety concern due to their high bioactivity and potential toxicity. Regulatory limits for certain alkaloids are set very low in many regions, reflecting their risk. Rapid, reliable screening methods are essential for ensuring food safety across diverse matrices such as milk, wine, bread, and rice powder.

Objectives and Study Overview

This study aimed to develop and validate a high-throughput UHPLC/MS/MS workflow for simultaneous quantitation of 18 toxic alkaloids in four food matrices. The goal was to achieve sensitive, accurate, and precise results that meet stringent regulatory criteria, with a simple sample preparation adapted for routine analysis.

Methodology and Instrumentation

Sample Preparation:

• Solid samples (2 g) hydrated with water, extracted with acetonitrile and Na2EDTA–McIlvaine buffer at pH 3.
• Liquid samples (milk, wine) extracted similarly without hydration step.

UHPLC/MS/MS Conditions:

• UHPLC: Agilent ZORBAX Eclipse Plus C18 column (3.0 × 50 mm, 1.8 µm), 0.1% formic acid in water/methanol gradient, 0.5 mL/min, 40 °C, 2 µL injection.
• MS/MS: Agilent 6470A triple quadrupole with Jet Stream ESI in positive mode, MRM acquisition for 18 alkaloids.

Used Instrumentation

• Agilent 1290 Infinity II binary pump with degasser
• Agilent 1290 Infinity II autosampler with needle seat backflush
• Agilent 1290 Infinity II column compartment
• Agilent 6470A triple quadrupole LC/MS with Jet Stream ESI

Main Results and Discussion

Method Sensitivity and Linearity:

• Calibration range: 0.5–50 µg/L (0.5 mL extract) corresponding to 5–500 µg/kg in matrix.
• Correlation coefficients > 0.995 for all analytes.
• Limit of quantitation set at 5 µg/kg (S/N > 10).

Recovery and Precision:

• Recoveries across wine, milk, rice powder, and bread ranged 82–119% at spiking levels of 5, 50, 250 µg/kg.
• Relative standard deviations between 2.3 and 7.9% (n = 6).

Matrix Effects:

• Matrix-matched calibration compensated for signal suppression/enhancement, especially in bread extracts.

Benefits and Practical Applications

This method combines a straightforward extraction protocol with fast UHPLC/MS/MS detection, enabling high-throughput analysis. It achieves regulatory compliance levels for multiple alkaloid classes in diverse foods, making it suitable for routine monitoring in quality control, research laboratories, and regulatory agencies.

Future Trends and Potential

Advances in UHPLC/MS/MS sensitivity and speed will further reduce analysis time and detection limits. Integration with automated sample preparation and data processing will enhance throughput. Expansion of target panels to include emerging alkaloids and other contaminants will broaden applicability in comprehensive food safety screening.

Conclusion

The developed UHPLC/MS/MS method delivers robust, sensitive, and accurate multi-alkaloid quantitation in four food matrices. Its high recovery, precision, and throughput support effective routine surveillance of toxic alkaloid residues.

Reference

1. Commission Regulation (EU) 2016/239 on tropane alkaloids in food for infants.
2. Commission Regulation (EU) No 37/2010 on permitted substances in animal-derived food.
3. Meng P.J. The Properties of Toxic Alkaloids and Analysis. Chin. J. Chem. Bull. 2010, 6, 484–489.
4. Jain V. et al. Analysis of Alkaloids in Areca Nut Products by LC-MS/MS. J. Agric. Food Chem. 2017, 65(9), 1977–1983.
5. Lv C. et al. Determination of Five Pyrrolidine Alkaloids in Honey by HPLC-MS/MS. Chin. J. Chromatogr. 2013, 31(11), 1046–1050.