Agilent Technologies - Utilizing Fast HPLC Screening Methods for the Detection of Steroids

As the importance of steroids in the regulation of bioprocesses has increased so has the demand for fast screening methods for their detection. HPLC is a highly regarded analytical technique for pharmaceutical applications due to its unrivalled analytical validation characteristics; including accuracy, precision, limit of detection, specificity, linearity and range, and ruggedness.

The high throughput demands of quantitative steroid analysis require faster run times and as a result there has been a focus on the development of fast HPLC screening methods to detect steroids such as hydrocortisone, testosterone and progesterone. Recent developments in HPLC column technology, such as sub-2 μm totally porous and sub-3 μm superficially porous columns, have offered improvements in separation speed and performance through reduced analysis times and increased separation efficiency. It is these improvements in column technology which mean that fast HPLC screening for the detection of steroids can be achieved.

Selectivity is the most powerful tool to optimize separations in HPLC and can be manipulated to improve separations. As a result, the analysis of steroids can be dramatically improved by choosing the optimal HPLC column for the separation. Selectivity can be increased by changing the mobile phase composition, column temperature and the composition of stationary phase; for example, the short column length and high efficiency of Poroshell 120 HPLC columns  provide fast HPLC analysis times and rapid equilibration leading to fast investigations of selectivity.

A key feature of Poroshell 120 columns is their superficially porous  microparticulate column packing. Poroshell 120 particles have a 1.7 μm solid silica core with a 0.5 μm porous outer layer. This unique configuration delivers all the performance advantages of sub-2 μm particles with the backpressure of a sub-3 μm particle. With Poroshell 120 columns, it is possible to achieve 80–90% or more of the efficiency expected from a sub-2 μm Fast LC/UHPLC column — but at HPLC pressures (below 400 bar). This ability to perform fast separations at low pressures can dramatically enhance productivity by allowing more samples to be run in less time — even using existing HPLC systems. This allows laboratories to generate better chromatographic results quicker so they can increase their throughput of samples. 

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