Simultaneous Determination of 14 Paralytic Shellfish Toxins using LC-MS/MS on the SCIEX QTRAP® 6500+ System

Significance of the Topic

Paralytic shellfish toxins are potent neurotoxins produced by marine microalgae that accumulate in shellfish and threaten consumer health. Regulatory limits are set at 800 μg STX eq/kg, necessitating rapid and reliable detection methods.

Objectives and Study Overview

This study introduces a streamlined LC-MS/MS approach using the SCIEX ExionLC AD and QTRAP 6500+ systems to quantify 14 paralytic shellfish toxins simultaneously. The goal was to improve sensitivity, throughput, and reproducibility while simplifying sample preparation.

Methodology and Instrumentation

• Sample preparation: Five grams of homogenized shellfish were extracted with aqueous acetic acid, sonicated, centrifuged, and cleaned using a Cleanert PEP-2 SPE cartridge.
• Chromatography: A 10 μL injection was run on a TSK-GEL Amide-80 HILIC column (150 × 2.0 mm, 3 μm) with mobile phases of water and acetonitrile containing 2 mM ammonium formate and 0.05% formic acid. The gradient program delivered a full 18-minute cycle at 0.4 mL/min.
• Mass spectrometry: The SCIEX QTRAP 6500+ with fast polarity switching and IonDrive Turbo V source (450 °C) monitored two MRM transitions per analyte for confident identification.
• Instrument platform: SCIEX ExionLC AD coupled to QTRAP 6500+.

Main Results and Discussion

The method achieved baseline separation of isomeric toxins within 18 minutes. Matrix-matched calibration curves exhibited linearity (R2 > 0.99). Limits of quantitation ranged from 1 to 50 ng/g, aligning with regulatory requirements. Recovery at low and high spike levels was 70–120% with RSDs below 30%, and system precision tests returned %RSD values under 15%.

Benefits and Practical Applications of the Method

This LC-MS/MS workflow eliminates the need for derivatization, reducing labor and analysis time compared to LC-FLD. Its high throughput and sensitivity make it well suited for routine monitoring of PSTs in diverse shellfish matrices in food safety and regulatory laboratories.

Future Trends and Applications

Advancements may include high-resolution mass spectrometry, automated sample preparation, and integration of online SPE or microfluidic platforms. Broadening toxin panels and applying machine learning for data analysis could further strengthen marine biotoxin surveillance.

Conclusion

A rapid and robust LC-MS/MS method was validated for the simultaneous detection of 14 paralytic shellfish toxins in shellfish. It demonstrates high sensitivity, reproducibility, and compliance with regulatory standards, offering a practical alternative to bioassays and derivatization-based techniques.

References

1. European Commission (EC) No. 853/2004
2. GB 5009.213-2016
3. Food safety supervise department 0305, No2 (Japan)
4. Michael J. Boundy, Andrew I. Selwood, D. Tim Harwood et al. Development of a sensitive and selective liquid chromatography-mass spectrometry method for high throughput analysis of paralytic shellfish toxins using graphitized carbon solid phase extraction. Journal of Chromatography A, 1387 (2015) 1-12
5. Choonshik Shin, Haerim Jang, Hyejin Jo et al. Development and validation of an accurate and sensitive LC-ESI-MS/MS method for the simultaneous determination of paralytic shellfish poisoning toxins in shellfish and tunicate. Food Control, 77 (2017) 171-178