Agilent 1290 Infinity II LC with ISET – Emulation of the Agilent 1100 Series LC Through Waters Empower Software

Importance of the Topic

The ability to transfer chromatographic methods seamlessly between conventional HPLC and modern UHPLC platforms is essential for pharmaceutical analysis, quality control, and method development. Intelligent System Emulation Technology (ISET) enables laboratories to preserve retention times and resolution without modifying existing methods or hardware.

Study Objectives and Overview

This application note evaluates the use of ISET to emulate an Agilent 1100 Series Binary LC method on an Agilent 1290 Infinity II LC under Waters Empower 3 control. A mixture of five analgesic compounds was analyzed to compare retention times, resolution, and run-to-run precision between both systems.

Methodology and Instrumentation

The original analysis employed an Agilent 1100 Series Binary LC with binary pump, micro vacuum degasser, autosampler, thermostatted column compartment, and diode array detector. Emulation was performed on the Agilent 1290 Infinity II LC using a high-speed pump, multisampler, multicolumn thermostat, and diode array detector. Both setups were controlled through Waters Empower 3 with Agilent Instrument Control Framework and ISET modules.

Solvents and Sample Preparation

• Solvents: LC-grade acetonitrile, methanol, ultra-pure water.
• Analytes: acetaminophen, antipyrine, acetanilide, hydroxyantipyrine, phenacetin.
• Stock solutions: 1 mg/mL in 20% acetonitrile/0.1% formic acid, diluted to final concentrations.

Chromatographic Conditions

Column: Agilent ZORBAX RRHD SB C18 (4.6 × 150 mm, 3.5 µm) at 30 °C. Mobile phase A (water) and B (acetonitrile) gradient from 15% to 80% B over 9 min at 1.5 mL/min. Detection at 254 nm with 360 nm reference.

Main Results and Discussion

Without ISET, UHPLC retention times were up to –13% lower than the HPLC reference, exceeding the ±5% specification. Activating ISET restored retention times within ±1% and maintained resolution within acceptable limits. Run-to-run precision was excellent (RSD < 0.13% for all analytes). Chromatogram overlays confirmed near-identical elution profiles.

Benefits and Practical Applications

• No method or hardware changes are required when migrating HPLC methods to UHPLC.
• ISET ensures reproducible retention and resolution for routine analysis and validation.
• Enhanced method development throughput by screening columns and solvents on UHPLC and emulating to legacy systems.

Future Trends and Opportunities

Broader adoption of instrument emulation across multiple vendor platforms will improve method portability. Advancements in third-party control frameworks and dynamic method adaptation will further enhance laboratory productivity in pharmaceutical and industrial settings.

Conclusion

Agilent ISET combined with the 1290 Infinity II LC enables direct transfer of chromatographic methods from HPLC to UHPLC, preserving analytical performance under third-party software control. This approach delivers consistent retention, resolution, and precision without modifying established workflows.

References

• Agilent Instrument Control Framework (ICF), Customer Letter, Pub. 5990-5756EN, 2010.
• Agilent 1290 Infinity with ISET, User Manual, Pub. G4220-90313, 2014.
• Huesgen AG, Seamless method transfer using ISET, Technical Overview, Pub. 5990-9113EN, 2011.