Analysis of Natural and Synthetic Estrogens at Sub-PPT Levels in Surface Water and Crude Influent Water Utilizing the ACQUITY UPLC System with 2D LC Technology and Xevo TQ-S

Importance of the Topic

Estrogenic compounds such as 17α-ethinylestradiol, 17β-estradiol and estrone are introduced into aquatic environments through municipal wastewater discharge and can disrupt endocrine systems in aquatic organisms. Regulatory bodies have placed these substances on watchlists to understand their occurrence at sub-parts-per-trillion levels. Sensitive, reliable analytical approaches are vital for monitoring and managing environmental risks.

Objectives and Study Overview

This study aimed to develop and fully validate an analytical protocol for the confirmation and quantification of natural and synthetic estrogens in surface water, crude influent, and final effluent at sub-ppt concentrations. The method integrates off-line solid phase extraction (SPE) with two-dimensional liquid chromatography (2D LC) and tandem mass spectrometry to meet stringent performance criteria for environmental monitoring.

Methodology and Instrumentation

The analytical workflow comprises two main sample preparation steps and online concentration and separation:

• Off-line SPE: Surface and effluent water samples were first concentrated using Oasis HLB cartridges, followed by cleanup on Sep-Pak silica cartridges to reduce matrix interferences and enhance detection limits.
• Online Concentration and 2D LC: Extracts were loaded onto a Direct Connect Oasis HLB trapping column for further preconcentration, then eluted onto an ACQUITY UPLC BEH C18 analytical column.
• Detection: A Waters ACQUITY UPLC System with 2D LC Technology coupled to a Xevo TQ-S mass spectrometer operating in electrospray negative ion mode. Two multiple reaction monitoring (MRM) transitions per compound were recorded, and the RADAR™ function collected simultaneous full-scan and MRM data.

Main Results and Discussion

The method achieved excellent linearity (R2 > 0.999) over the calibration range of 0.120–1.20 ng/L, with precision (%RSD) below ±3%. Limits of detection and quantification were below 1.2 ng/L for 17α-ethinylestradiol, 12 ng/L for 17β-estradiol, and 40 ng/L for estrone in surface waters. Spiked recovery studies in river and wastewater matrices demonstrated robust accuracy, while chromatographic examples confirmed clear separation and sensitive detection at low levels.

Benefits and Practical Applications of the Method

The validated approach offers:

• Ultra-sensitive detection of estrogens in complex water matrices.
• High throughput through online concentration and 2D LC separation.
• Reliable quantification for environmental monitoring, regulatory compliance and water treatment performance evaluation.

Future Trends and Potential Applications

Advanced 2D LC platforms and high-resolution mass spectrometry are expected to expand the scope of trace organic contaminant analysis. Real-time monitoring, automated sample preparation, and broader compound panels will support comprehensive assessment of emerging pollutants.

Conclusion

The combination of off-line SPE extraction, on-line trapping, 2D UPLC separation, and Xevo TQ-S detection delivers a fully validated, ultra-sensitive method for quantifying natural and synthetic estrogens at sub-ppt levels in surface, influent, and effluent waters, meeting rigorous performance criteria for environmental analysis.

Reference

Ross Euan; Boag Angela; Todd Hamish; Gatward Neil. Analysis of Natural and Synthetic Estrogens at Sub-PPT Levels in Surface Water and Crude Influent Water Utilizing the ACQUITY UPLC System with 2D LC Technology and Xevo TQ-S. Waters Corporation, 2016.