Analysis of Microcystins RR, LY, and YR in Bottled, Tap, and Surface WaterUsing ACQUITY UPLC Systems with 2D-LC Technology

Importance of the Topic

Algal blooms driven by cyanobacteria produce microcystins, potent cyclic peptide toxins that threaten human and animal health through contaminated drinking and recreational waters. Trace-level detection of microcystins RR, LR, and YR is essential for meeting WHO guidelines and protecting ecosystems and public health.

Objectives and Study Overview

This study evaluates a rapid microextraction protocol combined with ACQUITY UPLC 2D-LC and Xevo TQD MS detection to quantify microcystins RR, LR, and YR in bottled, tap, and surface water. Key goals include achieving sub-ppb detection, streamlined sample preparation, and robust performance across varying water matrices.

Methodology

The protocol uses a 15 mL water sample loaded onto a 3 cc Oasis HLB SPE cartridge at 10 mL/min. Sequential elution with 10–50% acetonitrile + 1% formic acid isolates the three toxins in a 15 min workflow. An at-column dilution trap-and-elute 2D-LC setup permits high-volume injections (250 µL) and on-line enrichment. Chromatographic separation employs an ACQUITY BEH C18 column (2.1×50 mm, 1.7 µm) at 60 °C with a 5 min 5–95% B gradient (water/ACN + 0.5% formic acid) at 0.5 mL/min. MS detection is performed on a Xevo TQD in ESI+ mode using optimized MRM transitions (e.g., 520>135.2 for RR, 995.5>135.2 for LR, 1045.5>135.2 for YR).

Instrumentation Used

• ACQUITY UPLC 2D-LC system with at-column dilution
• Xevo TQD triple quadrupole mass spectrometer (ESI+)
• Oasis HLB SPE cartridges (3 cc, 20 µm)
• ACQUITY UPLC BEH C18 column, 2.1×50 mm, 1.7 µm

Main Results and Discussion

The method achieved a 50 ppt detection limit with 10:1 microextraction enrichment and up to 2000:1 overall enrichment via on-line trapping. Calibration over 50–5000 ppt was linear (r2>0.995) using nodularin as an internal standard. Sequential elution experiments revealed that microcystin RR elutes at low organic content (20% ACN) under acidic conditions, while LR and YR require higher organic percentages (~40%). Recovery studies demonstrated 90–104% for bottled water and 75–85% for tap and surface water with 10:1 enrichment, indicating effective cleanup across diverse matrices.

Benefits and Practical Applications

This workflow offers a complete sample preparation and analysis in under 15 minutes, trace-level sensitivity in the low ppt range, and robust performance across bottled, tap, and surface waters. Its high throughput, minimal solvent consumption, and ease of automation make it well suited for routine water quality monitoring and regulatory compliance.

Future Trends and Applications

Further developments may include coupling with high-resolution MS for expanded toxin panels, fully automated microextraction platforms, green solvent strategies, and advanced data analytics powered by machine learning to enhance sensitivity, throughput, and multi-analyte screening in environmental water testing.

Conclusion

The described microextraction and 2D-LC-MS/MS method delivers rapid, sensitive, and reproducible quantification of microcystins RR, LR, and YR in various water matrices. It meets or exceeds WHO guideline requirements and provides a streamlined solution for laboratories engaged in water safety and environmental monitoring.

References

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3. World Health Organization. Guidelines for Drinking-Water Quality, 2nd ed., 1998.
4. Mallet CR et al. Time De-Coupled Chromatography, Waters White Paper No. 720005125EN, 2014.
5. Mallet CR. Analysis of Pharmaceuticals and Pesticides in Water using ACQUITY UPLC 2D Tech, Waters App Note No. 720005167EN, 2014.