Which Key UHPLC Characteristics Are Required for a High Throughput LC/MS Assay?

Significance of the Topic

LC/MS/MS is a cornerstone technique in analytical chemistry offering exceptional selectivity and sensitivity for quantitative assays. In high-throughput environments such as drug metabolism, pharmacokinetic studies and clinical testing, reducing total cycle time per sample is essential to maximize laboratory productivity and data output.

Objectives and Study Overview

This white paper examines the key front-end UHPLC characteristics required to achieve ultra-fast, high-throughput LC/MS assays. It outlines critical design features, carryover control strategies and multiplex configurations that together minimize non-data-acquisition time.

Methodology

Three major approaches were evaluated:

• Injection Speed and Sample Capacity – Utilizing sub-2 µm or core-shell columns coupled with the SIL-40 autosampler to achieve 6.7 s injection cycles and support up to 17 000 samples per 384-well plate.
• Low-Carryover and Rinsing Function – Incorporating a needle-in-flow-path design, zero-dead-volume connections and multi-solvent internal rinsing steps (acidic, organic, ionic) to suppress carryover below 0.0015 % without rinse and to 0.00007 % with optimized internal wash.
• Multiplex Configuration – Employing Nexera MX dual-stream technology to overlap data acquisition and column wash/equilibration phases, effectively halving the non-acquisition time and doubling sample throughput.

Used Instrumentation

• Shimadzu Nexera LC-40 UHPLC system
• SIL-40 series autosampler with Plate Changer units
• Shimadzu LCMS-8050 triple quadrupole mass spectrometer
• Nexera MX system with MX Dual Stream Technology

Main Results and Discussion

The SIL-40 autosampler achieved 6.7 s injection cycles, reducing non-data-acquisition time for 100 one-minute assays from over 50 min to approximately 11 min. In plasma assays of verapamil and its deuterated analog, 300 consecutive injections at an 18 s cycle time yielded 2.4 % RSD for the internal standard. Carryover tests with glucose-6-phosphate demonstrated 0.0015 % without rinse and 0.00007 % with internal multi-solvent washing. The Nexera MX system completed a cytochrome P450 biomarker panel in 38 s versus 1 min 22 s with conventional UHPLC, effectively doubling throughput.

Benefits and Practical Applications

These front-end UHPLC advances deliver higher sample throughput, reduced downtime, excellent quantitative accuracy and stable performance. Laboratories engaged in DMPK, clinical assays, quality control and high-volume screening can leverage these features to increase efficiency, conserve resources and accelerate project timelines.

Future Trends and Opportunities

Emerging directions include expanded multiplexing capabilities, deeper integration with laboratory automation and informatics, AI-driven method optimization, novel column chemistries for greener solvents, and further miniaturization of flow paths to boost speed and sustainability.

Conclusion

Optimizing UHPLC front-end characteristics—rapid injection, minimal carryover and multiplexed operation—enables ultra-fast, high-throughput LC/MS assays. The Shimadzu Nexera solutions exemplify how tailored system design can dramatically improve laboratory productivity and data quality.

Reference

1. T. Uchikata, D. Vecchietti, Ultra-Fast Analysis of Drugs in Biological Fluids with the SIL-40 Autosampler, Shimadzu Technical Report (C190-E228)
2. Nexera Series Brochure (C196-E096)
3. Nexera MX Brochure (C190-E190)
4. K. Watanabe, C. Campbell et al., Prominent Features of Shimadzu UHPLC for an LC/MS Assay, Shimadzu Technical Report (C190-E282)