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Home > Magazine > Liquid Chromatography
Expanding HorizonsThe throughput and speed of ultrahigh performance liquid chromatography systems are creating a proliferation of new products and suppliers.by Tim Studt, Editor in Chief
Waters Protein-Pak Hi Res IEX 4.6- x 100-mm columns were recently introduced for use on their ACQUITY UPLC System which is optimized for maximum sensitivity, component resolution, and speed. Each column features Waters ACQUITY UPLC eCord Technology for monitoring critical individual column-use parameters. | Ultrahigh performance liquid chromatography (UHPLC, or sometimes referred to as just UPLC) has, in a little more than six short years, become one of the leading tools in the chromatography marketplace. The use of sub-2-µm (micron) particle columns employed in systems running at higher pressures (up to 15,000 psi) than traditional HPLC systems has improved the resolution and sensitivity of these systems while increasing analytical speed.
A number of new systems, columns, and stationary phase materials have been introduced over the past several months, and even more are scheduled to be introduced at Pittcon 2010.
New systems
UHPLC systems have become so popular that intermediate systems and columns are now being introduced to entice traditional HPLC users into the UHPLC “camp.” Waters Corp., for example, recently introduced its ACQUITY UPLC H-Class system, which incorporates the performance of their ACQUITY UPLC with the operational familiarity of traditional HPLC systems. The basis of this system is a quaternary solvent manager (QSM) and sample manager (SM-FTN) with a flow-through needle design that mimics the traditional HPLC workflow.
According to Rohit Khanna, VP of Marketing at Waters, “the H-Class is meant to replace conventional HPLC systems because methods development can now be seamlessly transferred from an HPLC to a UPLC, or vice versa. Three particle substrates in 11 chemistries are available for the new systems.
At the same time, Waters introduced its Protein-Pak Hi-Res Ion Exchange (IEX) columns for the analysis of intact biomolecules, including monoclonal antibodies, recombinant proteins, DNA/RNA, and vaccine compounds. The IEX column family comprises one anion exchange—quaternary ammonium—column chemistry and two cation exchange column chemistries—a carboxy methyl and a sulfopropyl—each contained on monodisperse, non-porous particles. The unique multi-layered network of ion-exchange functional groups on the particles result in high sample loading capacities for proteins not found with use of typical non-porous particle IEX offerings.
The IEX 4.6- x 100-mm columns are ideally suited for use on Waters’ ACQUITY UPLC System, which is optimized for maximum sensitivity, component resolution, and speed. Each column features Waters UPLC eCord Technology for monitoring critical individual column-use parameters.
Varian also recently introduced new UHPLC-based columns. These are additions to their family of Pursuit LC columns for pharmaceutical applications. New materials address needs from drug discovery to bioanalysis, saving time on analysis and method development. The Pursuit range now covers a much broader array of bioapplications, from cutting edge, ultra high pressure and fast gradient UHPLC to more traditional analytical and preparative HPLC techniques.
The Pursuit UPS 1.9- and 2.4-micron columns excel under high pressures and fast gradients. Pursuit UPS (1.9) columns boast the highest efficiency of Varian’s phases for liquid chromatography, ensuring best resolution and sensitivity with UV and MS detection. For bioanalytical methods, where time is critical and thorough sample preparation is not always possible, the new Pursuit UPS (2.4) columns are also highly efficient and easy to use. They avoid sample particulate, clogging and backpressure issues to provide reduced instrument downtime and increased throughput.
“These new columns expand the Varian LC family to provide either generic UHPLC discovery or tailored (U)HPLC method development solutions, designed to fit the needs of distinct parts of the pharmaceutical industry,” says Martin O’Donoghue, SVP of Scientific Instruments at Varian. “We continue to provide life scientists with more high-quality choices for addressing their specific application challenges.”
Varian also offers a full range of UHPLC-compatible Pursuit XRs Ultra columns to transfer methods from UHPLC to traditional HPLC or preparative-scale HPLC. Used together with Pursuit XRs, Ultra enables smooth scale-up and easy transfer of UHPLC methods to sites or contract laboratories without UHPLC capability.
Small particles
Restek also recently announced its Ultra II LC columns with 1.9-micron particle sizes for UHPLC applications and 2.2-particles for ultrafast LC (UFLC) and rapid resolution LC (RRLC) applications. Ultra II columns are available in traditional phases (C18, aqueous C18, and silica) and unique chemistries, which provide selectivity (biphenyl, Aromax, and PFP Propyl). The unique selectivity of the Ultra II Aromax column separates all 17 EPA Method 8330B analytes with no coelutions under simple isocratic conditions.
Fortis Technologies recently introduced 2.1-micron particles for UHPLC systems in a range of phase chemistries, which allow the analyst to maximize selectivity as well as increase resolution and speed due to efficiency. Chemistries available include C18, C8, Cyano, and Fortis’ unique di-phenyl functionality that’s specifically targeted at positional isomers and metabolite profiling separations. These particles are designed to optimize the operation of commercial UHPLC instruments that include the Jasco X-LC, Agilent 1290, and Waters ACQUITY UPLC systems.
In 2009, Phenomenex introduced its Kinetex core-shell columns in 2.6- and 1.7-micron sizes, along with three chemistries (C18, PFP, and HILIC). The unique design of the Kinetex particles enables chromatographers to get performance comparable to sub2-micron columns without investing in UHPLC systems. With Kinetex, researchers can obtain UHPLC results on any LC instrument platform, and any chromatographic method can now be completed in a fraction of the time previously required, without sacrificing performance.
Phenomenex also recently announced a new Web-based tool that optimizes customer HPLC methods for use with their Kinetex core-shell columns. The specific benefits to be gained by switching from fully porous particle columns are also calculated.
With the Kinetex calculator, the researchers input their method running conditions and details regarding their current chromatographic results. The calculator will then deliver an optimized method with the recommended Kinetex column and optimal run conditions for the user’s system, whether it’s HPLC or UHPLC. The device provides expected backpressures, efficiency gains, solvent reduction, timesavings, and resolution improvements, and can also process details of both isocratic and gradient methods.
Thermo Fisher Scientific also continues to support the UHPLC trend with its Accela UHPLC system and Hypersil Gold 1.9-micron columns. Hypersil Gold 1.9-micron particles give higher efficiency than 3- or 5-micron particles, and this efficiency is delivered over a greater range of optimum linear velocities. This makes it possible to operate at higher flow rates without losing performance. Because shorter columns packed with 1.9-micron particles give equivalent efficiency to longer columns packed with 5-micron particles, faster analyses and solvent savings become a reality for the chromatographer.
Agilent Technologies launched its 1290 UHPLC in 2009, along with its companion Zorbax Rapid Resolution High Definition SB-C18 threaded columns. While Agilent has been offering UHPLC systems since 2006, the company considers the 1290 and the Zorbax RRHD columns a new option for researchers rather than a replacement for another Agilent LC platform.
ZORBAX RRHD columns are an expansion of the ZORBAX 1.8-µm particle column line. The RRHD columns used improved packing processes to achieve stability up to 1200 bar for use with the Agilent 1290 Infinity LC. The RRHD 1.8-µm columns are available in 50, 100 and 150 mm lengths for fast or high-resolution—truly high definition—separations of researchers’ most complex samples.
The RRHD columns facilitate easy and secure method transfer from HPLC to UHPLC through identical column chemistries. These high-pressure (1200 bar) columns are designed for optimum results with the 1290 Infinity LC or other UHPLC instruments. And the 1.8-µm particles deliver maximum resolution for the most defined separations.
They are available in ZORBAX Eclipse Plus C18 for superior peak shape and ZORBAX StableBond C18 for alternate selectivity and low pH stability. They also achieve the same selectivity on the 3.5- and 5-µm ZORBAX columns with the same bonded phase for compatibility with any LC.
Higher productivity
The Flexar UHPLC Systems from PerkinElmer feature the new FX-15 UHPLC system, which can provide as much as a 10-fold gain in productivity, while reducing mobile phase solvent consumption by as much as 15-fold, depending on the application. Smaller particle size (<2 µm) columns offer new possibilities for resolution and separations and allow for faster analysis time.
PerkinElmer’s Chromera chromatography data system, which controls the Flexar platform, provides an extensive suite of tools designed to allow users to more easily control instruments, rapidly visualize data and efficiently communicate results through an accessible worktop. The platform uses a patented technology for instrument control that allows flexibility of configurations and multiple detection capabilities to help address the increasing need for laboratory productivity. Chromera also extends Flexar’s flexibility by supporting multi-detector system configurations often used in environmental and food safety applications. The Flexar line offers complete LC systems ranging from 6k- to 10k- to 15k-psi operation, allowing for highly customized LC solutions for specific applications.
Shimadzu Scientific Instrument’s Prominence UFLCXR high-separation, high-resolution detection of fine peaks enables the highly reliable analysis in which the slightest presence of impurities in a sample can be detected. The UFLCXR is optimized to enable high-speed analysis using Shimadzu’s Shim-pack XR-ODS II series of columns (2 and 3 micron with 8-nm pores). The UFLCXR is capable of operating at system pressures up to 9,500 psi for special situations that demand extra resolution (XR). Unlike many other high-pressure systems, the UFLCXR delivers data with high integrity and reproducibility, a result of fully re-engineered components in the injection valve.
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