Direct Determination of Trace Hormones in Drinking Water by Large Volume Injection using the LCMS-8050 Triple Quadrupole Mass Spectrometer

Significance of the Topic

The presence of trace steroid hormones in drinking water raises significant environmental and public health concerns due to their endocrine-disrupting potential. Even at sub-nanogram per litre levels, these compounds can interfere with reproductive, developmental, neurological, and immune functions in humans and wildlife. Ensuring accurate, sensitive detection is therefore critical for water quality monitoring and regulatory compliance.

Study Goals and Overview

This study aimed to develop a rapid, robust method for direct determination of seven target hormones in drinking water without relying on solid-phase extraction (SPE). Key objectives included:

• Quantifying estrone, estriol, 17-β-estradiol, equilin, androstenedione, testosterone, and 17-α-ethynylestradiol.
• Meeting US EPA UCMR3 and EPA Method 539 reporting levels (ng/L to pg/L).
• Minimizing sample preparation time through large-volume injection.

Methodology and Used Instrumentation

• Liquid Chromatography: Shimadzu Nexera UPLC with Shim-pack XR-ODS III column (150×2 mm, 2.2 µm) and Kinetex XB-C18 guard at 45 °C.
• Mobile Phase: A – 0.15 mM ammonium fluoride in water; B – methanol; gradient from 10% to 100% B over 15 min at 0.3 mL/min.
• Injection Mode: Three successive 400 µL injections (total 1200 µL) via a 500 µL loop.
• Mass Spectrometry: Shimadzu LCMS-8050 triple quadrupole; heated electrospray ionisation; rapid polarity switching (5 ms); MRM transitions tailored to each hormone.

Main Results and Discussion

• Ammonium fluoride (0.15 mM) significantly enhanced signal-to-noise ratios for all analytes compared to ammonium hydroxide (signal gains of 2.8–4.8×).
• Calibration was linear over two and a half orders of magnitude (R² > 0.999 with 1/C weighting).
• Detection limits ranged from 0.005 ng/L for testosterone to 0.330 ng/L for 17-α-ethynylestradiol, meeting or exceeding regulatory requirements.
• Peak area repeatability at low and high levels was typically <5% RSD; long-term robustness over 62 h yielded <5% drift for key analytes.

Benefits and Practical Applications

• Elimination of SPE reduces analysis time, consumable costs, and potential sample loss.
• High sensitivity and selectivity support routine monitoring of endocrine disruptors in drinking water.
• Method is adaptable to high-throughput laboratory environments requiring consistent performance.

Future Trends and Potential Applications

• Expansion of MRM panels to include emerging micropollutants and transformation products.
• Integration with automated sample preparation techniques (e.g., online SPE, microfluidics).
• Application to other environmental matrices such as wastewater effluent and surface water.

Conclusion

The developed direct large-volume injection LC–MS/MS method on the Shimadzu LCMS-8050 provides a fast, sensitive, and reproducible solution for quantifying trace steroid hormones in drinking water. It complies with EPA Method 539 and UCMR3 criteria while streamlining workflow by eliminating traditional SPE steps.

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