Recent Developments in HPLC/UHPLC

When choosing the articles to include in this supplement, my selection was based on a very simple criterion. My invitations went to the researchers that typically have something to say that I am interested in learning. The selections here are broad based; hopefully, to give you an overall perspective of current directions in the key separation areas.

Mary Ellen McNally, PhD, DuPont Crop Protection

Joe DeStefano, Bill Johnson, Stephanie Schuster, and Jack Kirkland have outlined a new perspective on an age-old torment: how to improve resolution. Those of us in the laboratory have often looked at beautifully resolved peaks only to examine one critical pair that simply co-elute. This frequently happens when there are a large number of sample components or if the components are chemically similar, perhaps positional isomers. The authors offer several viable options to solve this problem. Particle size, column length, flow rate, pressure, column temperature, and solvent strength relationships are described as possible options to improve resolution from this competent group. They are presenting solutions, I believe, you will all need and use.

Researchers from the Molnár Institute for Applied Chromatography led by Imre Molnár outline the use of their predicted tool, a design of experiment model that uses gradient time and temperature and one other factor, either pH or ternary composition, successfully. Twelve experiments are necessary to complete this model — the advantage is fast evaluation using gradients and within a few hours more than 100 virtual experiments can be evaluated. Absolutely awesome!

The question of whether or not chromatographers have gone too far with the use of high pressures, which create frictional heating, a source of interference to accurate temperature control, is addressed by Dick Henry and his coauthors, Hillel Brandes, Dan Nowlan, and John Best. They postulate that popular column dimensions, labeled since the early 2000s as ultrahigh pressure, ultrafast, and ultra-performance, along with current oven designs can lead to this problem. Investigating instrument bandspreading and instrument dispersion, the report also describes, via theoretical equations, the instrument optimizations required for these higher performance "ultra" columns. They conclude that future trends for both columns and instruments are hard to predict, and if higher operating pressures are used, they question how this will affect accuracy and precision in ultrahigh-pressure liquid chromatography (UHPLC). The authors request two very basic improvements: better control of column temperature and independent monitoring of both column and instrument pressure, ultimately, achieved without adding to instrument dispersion. Instrument manufacturers, please take heed!

The analysis of anions and cations in the same separation is an objective desired in many instances. Often, industry either switches instrumentation back and forth between two modes or purchases two pieces of equipment to accommodate these measurements. Foley and Blackney have reported on the requirements to eliminate biases in the resolution and analysis time during cation and anion determination by dual opposite injection capillary zone electrophoresis (CZE). When a voltage is applied, electro-osmotic flow is cathodic and cations travel faster through the capillary than anions. As a result, the cations are less resolved. Their paper describes corrections to address this and is a unique development in CZE, which should be widely employed for its practicality.

Those of us involved in supercritical fluid chromatography (SFC) in the late 1980s and early 1990s are skeptical of the resurgence over the last decade. Larry Taylor reports what is different now and predicts the future of SFC. His scheme of four protocols for the elution of four groups: hydrocarbons, ethers, and esters; alcohols, amides, and anilines; ionizable acids and bases; and amphoterics, peptides, carbohydrates, water soluble vitamins, and inorganic ions, illustrate the breath of materials that SFC is capable of analyzing. The technology has moved beyond a niche technique for chiral molecules and preparative separations — move over HPLC!

Finally, in my laboratories, we have long been interested in improving the precision associated with the assay measurement of the technical active ingredient. In our experiments, we found several areas that needed control to review with our laboratories world-wide. Details about optimum back pressure, wavelength selection, internal versus external standard use, and how to make sure carryover is under control are presented in this back to the basics article.

In reading this supplement on the advancements and predictions in separations, my hope is you will take time to think about the future directions in your laboratory. My special thanks to the authors, for a job well done.