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Advanced LC techniques focus on ultrafast analysis

8th June 2015

Posted By Paul Boughton


Waters’ new in vitro diagnostic system is designed to tackle the most demanding of quantitative analyses

Ongoing developments in ultra performance liquid chromatography and ultra high performance liquid chromatography technology is making seamless integration into existing laboratory systems much simpler, says Eugene McCarthy

Clinical laboratories today typically use LC-MS technology to perform qualitative and quantitative analyses of patients' samples. Such tests cover a variety of uses, for example: to confirm a clinical suspicion (including making a diagnosis); assist in the selection, optimisation and monitoring of treatment; provide a prognosis; screen for disease in the absence of clinical signs or symptoms; and establish and monitor the severity of physiological disturbance.

Liquid chromatography separates analytes and interferents within a given sample, while mass spectrometry technology is used for detection and confirmation of those analytes.

The new Waters ACQUITY UPLC I-Class IVD/Xevo TQ-S in vitro diagnostic (IVD) system features the company’s UltraPerformance LC (UPLC) technology coupled with a tandem quadrupole mass spectrometer designed for the most demanding UPLC-MS/MS quantitative analysis. This system uses StepWave, which Waters describes as a breakthrough off-axis ion source technology, and RADAR, an information-rich acquisition method, enabling the system with sensitivity and robustness.

One early adopter of the new system is the Chinese Food and Drug Administration (CFDA) which has approved it for IVD use in the country. So it can now be used in China for the clinical analysis of a variety of compounds including diagnostic indicators and compounds for treatment monitoring.

According to Jeff Mazzeo, senior director, health sciences for Waters Division, improved diagnostic and predictive tools such as this are essential to help medicine move from a reactive approach to treating disease to one that is more proactive and preventive.

In related news, Waters ACQUITY UltraPerformance UPLC received an award at the recent Pittcon conference on analytical chemistry and applied spectroscopy as a result of good customer reviews and ratings. The technology allows chromatographers to work at higher efficiencies with a much wider range of linear velocities, flow rates, and backpressures. What differentiates from other UPLC systems is Waters’ patented 1.7 micron hybrid particle chemistry.

Agilent technologies is also expanding its UHPLC portfolio with the 1290 Infinity II LC system – the latest addition to its 1200 Infinity series LC portfolio.

The company has particularly focused on analytical quality, ease of use, and seamless integration with its new system as it strives to help laboratory scientists reach new levels of efficiency. In development, Agilent worked with a variety of customers in order to identify their most challenging liquid chromatography workflow issues and bottlenecks.

This involved three main priority areas for development. First is analytical efficiency. Here it has combined chromatographic resolution/precision and broad dynamic range detection with low carry-over. The second priority is instrument efficiency.

Maximum flexibility to accommodate changing separation conditions reduces turnaround times, while fast throughput is enabled by high sample capacity per bench space and fastest injection cycles.

Lastly is laboratory efficiency, and this is enabled by allowing the smooth transfer from any legacy equipment – always an important factor when laboratories are seeking to lower costs and increase their return of investment.

The analysis of complex samples is the target of the new Altus UPLC from PerkinElmer. The company says its new technology represents a breakthrough in LC particle technology and system design, combining as it does advanced fluidics with sub-2-μm particle columns for what PerkinElmer describes as superb performance at high pressures, with minimal volumes and optimised flow paths.

Combining the flexibility of quaternary solvent blending with the flow-through needle injector, this elite LC system provides high resolution, sensitivity, and improved throughput with exceptional, reliable results, adds the company.

Preparation time

One of the key issues tackled with Altus is preparation time. This is reduced significantly with AutoBlend technology, which automates the formulation of mobile phases from reservoirs of pure solvents or concentrated stock solutions. AutoBlend Plus also manages pH and ionic strength requirements for the selected mobile phase automatically, so users can take full advantage of the system’s quaternary blending capabilities. Plus, users can transfer methods to other laboratories quickly and easily, reducing the possibility of errors.

The company cites a number of additional benefits, including multi-solvent blending, direct inject sampling, next-generation column compartments, managed dwell volume, and a full complement of detection modes.

Also, it is easy to transfer methods from current instruments to the Altus UPLC system. Users simply select a column comparable to their HPLC column, enter existing HPLC conditions into the system’s UPLC Columns Calculator, and run.

For Shimadzu, high performance across a wider application range and improving flexibility and reliability of HPLC/UHPLC analyses have been the driving forces behind development of its Nexera X2 series. 

Like its predecessor, the Nexera X2 is optimised to enable analysis at pressures up to 19,000 psi without compromising performance and data quality. Nexera X2 autosamplers provide near-zero carryover, support ultrafast analysis with what the company says is the world’s fastest cycle time (14 secs), and deliver excellent injection reproducibility down to 0.1µL injection volumes. In addition, the Nexera X2 systems can be configured with a variety of solvent delivery units and column ovens to meet any laboratory workflow.

An extra benefit is its modular construction. This allows a solvent blending function that can mix up to four solvents per pump at a desired ratio, letting users change the proportion of the mobile phase components from run to run so there is no solvent wasted.

What limits your HPLC system?

When was the last time that you couldn’t sufficiently separate some peaks of interest? How did this affect your analysis?

If you own a standard HPLC system and normally perform separations on 5 µm or 3 µm columns, you have a great chance for improving this lack of separating power or chromatographic resolution. However, according to Knauer, there some issues you should take into account before investing time and money in new methods or equipment.

First is the fitness of the LC system. For example, says the company, if you don’t own a system with low dead volume, capable of delivering solvent at over 600 bar, there will only be a few applications on fully porous sub-2 µm columns which are suitable for your HPLC. Core-shell or superficially porous columns might be worth a try, because of their lower back pressure, but you need to be sure that system dead volume and your detector flow cell don’t spoil the improved separation results of the column. If the gain in resolution with the new column is high enough you could even speed up your analysis by using shorter columns.

Then there is scalabililty. For example, if you use your system for screening and the methods you develop are intended for being easily transferred to a preparative scale HPLC, fully porous particle columns are your best choice, adds the company.

For a solution that is capable of mastering both of these situations and offering laboratories a much greater freedom of choice, Knauer says that its PLATINblue UHPLC/HPLC Plus system is ideal.





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