Conceived as a separations tool for biomolecules, high-performance liquid chromatography (HPLC) has expanded its scope significantly into chemistry, pharmaceuticals, forensics, and organic chemical analysis. The need for speed and quality has led to faster, more efficient HPLC separations. But as the time between injection and elution shrinks to less than one minute for high-pressure UHPLC (ultra high-performance LC), cycle time becomes the principal bottleneck.

Terry Adams, life science business manager at Shimadzu Scientific Instruments (Columbia, MD), cites several areas of improvement that have made a difference in throughput and quality:

  • Easy-to-clean autosamplers reduce the time between samples
  • Fused-core or “core shell” columns and media perform like sub- 2-micron systems but at much lower pressures
  • Multiplexing two or more HPLC systems to a single mass detector utilizes detector downtime
  • Walk-up access allows any lab worker to queue samples onto a communal instrument and walk away
  • Networked instruments e-mail chromatograms or instrument service reports to scientists and technical personnel
  • Multidimensional HPLC provides peak capacity that is the product of the capacities of both columns
  • Increased use of derivitization for fluorescence and mass detection produces extremely clean fluorescence and mass signals, even from “dirty” mixtures that do not resolve well on the column

Although traditional HPLC still dominates methods and new system sales, customers increasingly opt for high-pressure UHPLC systems, says Phil DeLand, pharmaceutical market manager at Dionex (Sunnyvale, CA). “Many are still using 3- or 5-micron columns, but they’re hedging a bet that they may want or need the speed or resolving capabilities of UHPLC down the road.”

Pharmaceutical industry analysts, for example, will stick with existing HPLC methods for products or pipeline molecules because those have likely already been validated on low-pressure instruments. “They probably would not switch to UHPLC until a new compound comes along that has already been validated with newer methods,” DeLand adds.

DeLand believes that UHPLC has reached a point of diminishing returns in terms of speed since cycle time often takes substantially longer than a chromatography run. Increasing pressure from 1,000 to 1,200 bar, which is substantial, may reduce elution time from two minutes to one minute forty seconds, a gain of twenty seconds. “How much time does that really save you within the context of sample prep and data analysis?” DeLand asks. “Not much.” Still, other experts feel that for very high-throughput situations, every second counts.

Yet many users are eager to upgrade, which brings cost issues to the forefront. At one time, conventional wisdom dictated that purchasers budget 10 percent of system cost per year for maintenance. Users are now challenging that idea, says DeLand. Twenty years ago laboratory personnel took a hands-on approach to instrumentation. This was followed by a period when soup-to-nuts service contracts dominated. “During the last five years, as sample loads have increased and staff has been cut, interest in training for in-house troubleshooting has risen, and with that, demand for more reliable HPLC systems. We have come full circle.”

1200 Infinity Series

• Available in three models: 1220, 1260 and 1290
• 1220 and 1260 LCs are standardized on 600 bar pressure and 80 Hz data acquisition speed
• 1260 Quaternary Bioinert LC offers highest performance in bio-analysis and bio-purification
• 1290 LC enables users to deploy any particle type, any column dimensions and any mobile/stationary phases

Agilent Technologies
www.agilent.com


ExpressLC-ultra®

• Designed to run 0.5 mm ID micro-columns at pressures up to 10,000 psi
• Allows for the use of separation columns packed with <2 μm particles
• CCD-based detector offers higher UV sensitivity with full spectral capability
• Features a binary gradient pump (flow rate range 1-50 μl/min) with solvent selection

Eksigent
www.eksigent.com

ACQUITY UPLC® H-Class

• Combines flexibility of ternary or quaternary solvent blending with simplicity of flow-through needle injections
•Quaternary solvent manager (QSM) and sample manager (SM-FTN) mimic traditional HPLC system workflows, allowing a user to make the switch to UHPLC when he/she is ready

Waters
www.waters.com

Nexera

•Optimized to enable analysis at pressures up to 130 MPa
• Features a 10-second injection time and 4,600-plus sample capacity
• Autosampler offers fixed-loop injection for ultra-fast separation and minimized peak width
• Intelligent Heat Balancer (IHB) minimizes band broadening during high-temperature analysis

Shimadzu Scientific Instruments
www.ssi.shimadzu.com