Chromatographers are always interested in higher efficiency. This is motivated by the resolution equation, which shows that
increases in column efficiency always result in improved resolution. Chromatographic efficiency is affected by a large number
of experimental variables and its optimization can be achieved in many different ways, depending upon how many variables one
is willing to adjust. These include pressure, temperature, particle size, column length, and eluent velocity. In the early
days of high performance liquid chromatography (HPLC), the selection of column formats (particle size, type, and column diameters)
was rather limited and thus, optimization often was done by adjusting operational variables such as eluent velocity, column
temperature, and operating pressure. Nowadays the selection of column formats is substantially wider and one can find a number
of particle sizes between 1.7 μm and 5 μm, and numerous column lengths are achievable by coupling columns in series. This
makes optimization of these nominally "discrete" variables possible (that is, particle size and column length).
Optimization of performance (meaning efficiency versus time) in HPLC has been studied for decades. Van Deemter was clearly
among the first who understood how plate count on a given column could be optimized by varying the eluent velocity (1). When
column length is also allowed to vary, one can use the Poppe plot (2) or kinetic plot (3,4) techniques that consider pressure
and time constraints as part of the optimization process. Ultimately, one also can vary particle size, and this optimization
has been studied by several LC pioneers including Halasz (5), Knox (6), Horvath (7), and Guiochon (8). We recently summarized
these distinct optimization schemes and emphasized the different chromatographic results that these approaches entail (9).
The purpose of this column is to propose a simple, stepwise optimization procedure that is solidly founded and to demonstrate
its practical utility by applying it to the development of an ultrafast isocratic separation for pharmaceutical applications.
As the march toward more-efficient separations continues, so does the demand for faster separations without sacrificing efficiency.
This has been stimulated by the present and persistent need to deal with more and more samples in the same or less time. Clearly
"fast" means different things in different applications. For example, "fast" can be a 10-min separation for organic impurities
analysis in pharmaceuticals, or it can be a 1-min separation for cleaning analysis in drug manufacturing. In this column,
we focus on the implementation of fast LC in pharmaceutical drug product dissolution testing. In such a test, one is primarily
interested in quantifying the active pharmaceutical ingredient (API) released from dosage forms, thus, only moderate plate
counts are required; here, analysis speed is more important than higher efficiency. Dissolution samples often are analyzed
by two techniques, direct UV or HPLC–UV, in almost equal proportions as shown by an online survey of dissolution studies (10).
Table I compares these two techniques. A thorough review of the use of HPLC in dissolution studies can be found in a chapter
written by Wang and Gray (11). While UV–vis spectroscopy is cheaper and simpler, HPLC offers many advantages, such as better
specificity and a greater linear dynamic range. HPLC is also more versatile especially for early drug development when different
formulations and strengths are screened. The main disadvantage of traditional HPLC is its relatively slower speed (about 3–5
times slower than UV analysis). Clearly, increasing the speed of HPLC makes it more attractive in this context.
This work focuses on using theoretical guidance to develop better ultrafast isocratic separations (approximately 30 s) for
dissolution testing, with the goal of making the speed of HPLC competitive with UV analysis. In addition, ultrafast HPLC is
critical to the implementation of automated dissolution testing systems with online HPLC analysis (11). There are many approaches
to speed up HPLC analysis. One popular way is to use monolithic columns at very high eluent velocities; sub-1-min separations
have been achieved (12,13). However, our recent review of different technologies for enhancing speed suggests that the most
effective way of achieving very high speed is the combined use of higher temperatures, higher pressures, and smaller particles (14). In addition, substantially lower solvent consumption
can be achieved with narrow-bore columns packed with small particles compared to monolithic columns. Therefore, in this column,
we focus on optimizing an ultrafast separation under high temperature and high pressure conditions.
Table I: Criteria of UV and traditional HPLC analysis for dissolution test*