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Why Have High Performance When You Can Have Ultra-High Performance?

Team TFS
Team TFS
HPLCLiquid chromatography plays an extremely important part in the development and production of biotherapeutic drugs. From separation of large quantities of antibodies in cell culture media to minute quantities of product from biological fluids during clinical studies, there is a growing need to be able to separate and measure these substances.

The Russian botanist Mikhail Tsvet is considered to have invented the chromatographic technique (link to Lab Manager article) when he reported separations of different plant pigments into a series of colored bands on a packed column (see image below). He called this technique chromatography. HPLC became popular in the 1960s and 1970s, with UHPLC becoming popular in the 1990s. A clue as to the longevity of the technique is that the HPLC meeting (link to 2015 conference) is now in its 42nd year!




Regulators take HPLC and UHPLC seriously as well. In fact, the U.S. Food and Drug Administration (link to FDA website) lists 1,970 documents that include the search term “UHPLC”! The European Medicines Agency (link to EMA website) is similarly well versed in the usefulness of HPLC and its variants. Here is a link to a great resource detailing the guidelines for bioanalytical method development (link to article), which has some really useful reading about performance expectations for HPLC instruments used in conjunction with mass spectrometry for clinical studies.



Despite modern complexities, the principles behind HPLC and indeed UHPLC remain the same, neatly described by Wikipedia (link to article) as “passing a pressurized liquid solvent containing the sample mixture through a column filled with a solid adsorbent material. Each component in the sample interacts slightly differently with the adsorbent material, causing different flow rates for the different components and leading to the separation of the components.” The obvious difference is the operational pressure, being typically 50-350 Bar for HPLC and up to or even beyond 1,000 Bar for UHPLC. But what drives the requirement for pressure?


Particle Size

Basically, it’s all about the particle size. By reducing the particle size, it’s possible to increase the resolution and speed of analysis. Today’s columns with 1.7 micron particles are incredibly high-resolution, but require vast pressures to force the mobile phase through the tightly-packed stationary phase. The future may bring even smaller particle sizes and even greater pressures, all driven by the demands of industries such as the biopharmaceutical sector.

As a consequence, this sector of the industry also helps to drive the technology behind the UHPLC instruments. We’ve recently launched a new UHPLC instrument that was purpose-designed for biotherapeutic applications (link to biopharma page), so I thought it was worth a closer look. Vanquish Flex™ is the second in the Vanquish range of UHPLC products, designed and built at our Germering facility in Germany.

I spoke to Mauro de Pra (link to LinkedIn profile), one of the solutions team responsible for the design of Vanquish Flex. Mauro told me that “the Vanquish range of products was designed from the ground up for the most challenging workflows that involve UHPLC. In particular, Vanquish Flex was created to be the separations solution for all of the major biotherapeutic workflows, including glycans, intact proteins, charge variants, aggregates and peptide mapping”. Impressive stuff, but what in particular marks Vanquish out for being suitable for these challenges?

Without going in to too many technical details, it’s the overall design that makes the difference. Here’s a link to a video about the Vanquish platform (link to Youtube video) to see how this system is built for biopharma.


Additional Resources

For more information about UHPLC and its role in biotherapeutics, visit the resources below.


Do you have any insight into the analysis of biotherapeutic drugs by HPLC/UHPLC? Or, do you have an interesting story about the regulated environment that these drugs are produced in? If so, please let us know.