New Features of the Agilent Method Scouting Wizard for Automated Method Development of Complex Samples

Significance of the topic

Automated chromatographic method development is critical in pharmaceutical, environmental, food, and chemical analysis, where complex mixtures and large numbers of analytes demand efficient screening of columns, solvents, gradients, and temperatures. Reducing manual effort accelerates time-to-result, ensures robust separations, and supports quality control and research workflows.

Objectives and Study Overview

This technical overview evaluates the Agilent Method Scouting Wizard within the 1290 Infinity II Method Development Solution. The goal is to identify optimal conditions for separating 16 pesticides in a complex sample by screening multiple column chemistries, dimensions, solvent combinations, and temperatures. Automated adjustment of flow rates and gradient times for varying column sizes is demonstrated.

Methodology and Instrumentation

The experimental setup included:

• Agilent 1290 Infinity II Solution modules: Multisampler, Flexible Pump, Diode Array Detector, Valve Drive, and two Multicolumn Thermostats with selector valves.
• Software: Agilent OpenLAB CDS ChemStation with Method Scouting Wizard.
• Columns: ZORBAX SB-C18, SB-C8, Bonus-RP, Eclipse Plus C8 RRHD (2.1×150 mm, 1.8 µm) and shorter 4.6×50 mm/100 mm variants (3.5–5 µm).
• Mobile phases: binary gradients of water + 0.1 % formic acid with either acetonitrile or methanol + 0.1 % formic acid.
• Gradient times and flow rates automatically scaled to column length and inner diameter (initial method: 2 mL/min for 4.6 mm ID, 30 min on 150 mm columns; 0.417 mL/min for 2.1 mm ID with proportional gradient times).
• Sample: mixture of 16 pesticides at 1 mg/mL each in acetonitrile/acetone (4:1).

Main Results and Discussion

A screening matrix of 42 scouting runs (88 methods including washes and equilibrations) was generated. For small datasets, bubble-chart reports quickly highlighted that acetonitrile-based gradients on the 2.1×150 mm Eclipse Plus C8 column at 40 °C yielded the highest resolution sum and complete separation in 30 min. Temperature variations (40–60 °C) revealed selectivity shifts: some coelutions at 50 °C resolved at 40 °C and 60 °C with acetonitrile, while methanol produced incomplete separations. For larger datasets, the Summary Performance Report enabled filtering by maximum resolution sum, minimum resolution threshold, and number of detected peaks, efficiently identifying top methods without manual chart inspection.

Benefits and Practical Applications

The Method Scouting Wizard streamlines LC method development by:

• Automating method creation for diverse column–solvent–gradient–temperature combinations.
• Scaling flow rates and gradient durations to column dimensions without manual calculations.
• Providing integrated reports to filter and rank hundreds of runs based on resolution metrics.
• Reducing development time from days to hours, enhancing productivity in QA/QC and R&D laboratories.

Future Trends and Potential Applications

Upcoming developments may include:

• Integration of mass spectrometric detection within scouting workflows for compound identification.
• Machine-learning-driven optimization to predict optimal conditions from prior datasets.
• Expanded solvent and column chemistries to support green analytical methods.
• High-throughput coupling with 96-well plate autosamplers for large-scale screening.

Conclusion

The Agilent Method Scouting Wizard, combined with the 1290 Infinity II platform and ChemStation reports, provides a powerful automated solution for complex LC method development. Automated scaling of flow and gradient parameters, coupled with built-in performance reports, enables rapid identification of robust separation conditions, significantly reducing manual workload.

Reference

• Automated Scouting of Stationary and Mobile Phases Using the Agilent 1290 Infinity II Method Development Solution, Agilent Technologies Application Note 5991-5679EN, 2015.
• The Agilent 1290 Infinity II Multi-Method Solution, Agilent Technologies Application Note 5991-5934EN, 2015.