Performance characteristics of the Agilent 1100 Series capillary LC system using diode-array UV and MS for detection

Importance of the topic

Capillary liquid chromatography (capillary LC) has become essential when sample volume is limited or when enhanced sensitivity is required, such as in bioanalytical and pharmaceutical research. By reducing column diameter, peak volumes shrink while mass remains constant, leading to higher peak concentration and better signal-to-noise ratios. Coupling capillary LC with UV diode-array detection and mass spectrometry (MS) addresses the need for low-level quantification and identification of peptides, proteins, small molecules and metabolites in complex matrices.

Objectives and study overview

This study evaluates the performance of the Agilent 1100 Series capillary LC system equipped with a diode-array detector (DAD) and an MSD Ion Trap (MSn) for sensitivity, precision and robustness. Key goals include measuring retention time reproducibility, system delay volume, gradient mixing quality, injection volume precision, carry-over and limits of detection (LOD) across UV and MS detection modes. The system was also assessed after modification to handle higher flow rates for standard column formats.

Methodology and instrumentation

A range of capillary columns (0.3–1.0 mm id) and standard columns (2.1–4.6 mm id) were tested using water and organic solvents with formic acid or TFA modifiers. Gradient and isocratic methods were applied, and injection volumes spanned from tens of nanolitres up to tens of microlitres. Performance metrics were collected using repeated tracer and peptide standard injections. Key instrumentation included:

• Agilent 1100 Series capillary pump with electronic flow control down to 1 µl/min
• Diode‐array detector with 500 nl flow cell
• MSD Ion Trap with optimized capillary sprayer for ESI and MSn
• Micro well‐plate sampler with dual capillary loops (30 nl–8 µl and 200 nl–40 µl ranges) and vial bottom sensing
• Thermostatted column compartment (ambient to 80 °C)
• Micro vacuum degasser and Upchurch mixer options

Main results and discussion

The Agilent 1100 Series capillary LC system demonstrated:

• Retention time precision <0.5 % RSD for peptide standards across extended gradients
• System delay volume of ~80 nl (column-to-detector) and ~600 µl for the total flow path; bypassing autosampler reduced delay time from 16 min to ~5 min
• Gradient step accuracy within 0.04–0.07 % and precision (RSD) of 0.02–0.04 % for 1 % step increments
• Mixing noise below 0.03 % when using dedicated capillary mixer
• Injection volume precision of 2–3 % RSD for 30–500 nl and <1 % for larger volumes up to 8 µl
• Injection linearity with correlation coefficients >0.9999 and response factor RSDs below 4 %
• Accessible sample volumes down to ~2 µl in low-volume vials with appropriate needle positioning
• Carry-over as low as 0.01 % for challenging compounds (e.g., azithromycin) using optimized needle and valve washing sequences
• Limits of detection by DAD around 50 fmol for peptides, improved to <50 fmol using MS Ion Trap with extracted ion monitoring
• Post-modification performance for higher flow rates up to 2.5 ml/min with standard columns, showing stable retention, mixing and precision comparable to microbore operation

Benefits and practical applications

This capillary LC-MS platform combines high sensitivity, low carry-over, precise retention and small delay volumes, making it ideal for proteomics, pharmacokinetics, metabolite profiling and trace analysis in QC laboratories. Its modular design allows easy transition between nano, capillary and microbore modes and compatibility with both UV and MS detectors. The robust sampler with bottom sensing expands its usability for limited-volume and low-abundance samples.

Future trends and opportunities

Emerging applications include deeper integration with high-resolution mass spectrometry, multidimensional separations, and real-time process monitoring. Further miniaturization and automation will support single-cell analyses, point-of-care testing and enhanced throughput in drug discovery. Advanced data-handling and predictive maintenance algorithms will improve operational efficiency and method development speed.

Conclusion

The Agilent 1100 Series capillary LC system delivers excellent robustness, sensitivity and precision for capillary LC coupled with diode-array UV and MS detection. It addresses the challenges of low-volume, low-concentration analyses and can be tailored for higher flow rate applications, offering a versatile solution for modern analytical workflows.

Reference

1. Agilent Technologies. Technical Note: Performance characteristics of the Agilent 1100 Series capillary LC system using diode-array UV and MS for detection. Publication Number 5988-7511EN, May 1, 2007.