Fast and Ultrafast HPLC on sub-2 μm Porous Particles — Where Do We Go From Here?

Jun 01, 2006

Figure 1
Since the beginning of modern high performance liquid chromatography (HPLC) in the late 1960s, users have required continually new and improved columns to tackle more difficult separation problems or to improve their overall productivity and sample throughput. Column researchers and manufacturers have responded to these needs with the development of more efficient and more reliable packing materials. One of the areas in which improvements have been made is in particle size reduction. Figure 1 shows a series of H versus ν curves that I developed in the early 1970s that showed the influence of the particle size of silica gel on column efficiency.1 Known by theoreticians for years, this data systematically showed that the use of smaller size particles resulted in more efficient columns. At that time, all we had to use were irregularly shaped particles; spherical microparticulate silicas had not yet been developed. To make an easier comparison, Figure 2 provides a plot of this data for H at 1.0 cm/s linear velocity versus average particle size of the silica packing. This log–log plot suggested that even smaller irregular particles would result in further performance improvements but at the time, smaller particles were not available in narrow particle size distributions.

Figure 2
Figure 3 gives a rough chronological order of the introduction of commercial column packings since the beginning of HPLC. The first high-performance packings were the pellicular (porous-layer bead) ion-exchange packings developed by Horvath and coworkers.2 These particles were used for the separation of nucleotides and resulted in the first commercial high-performance packing Pellosil (Northgate Laboratories, Hartford, Connecticut, USA). Pellicular packings were rather large compared with today's particles — 40–50 μm. They had a thin porous coating that allowed rapid solute mass transfer into and out of the packing, resulting in improved chromatographic efficiency relative to the large porous particles that were generally used for liquid chromatography (LC) separations at the time. However, these pellicular packings had a big disadvantage — low surface area and, thus, very low sample capacity. For more details on this historical perspective, consult reference 3.

Figure 3
At the time, column researchers knew that small porous particles (less than 20 μm) would provide even better efficiency and maintain the high capacity of the earlier porous packings. Some earlier work by Piel4 in 1966 and Bidlingmeyer and Rogers5 in 1969 showed promise, but the particles that they used were commercially available Cabosil (Cabot, Billerica, Massachusetts, USA) fumed-silica packings that were sub-1.0 μm sizes, were fairly inert, were difficult to handle and required extremely high pressures to operate. In short, these particles were unsuitable for the current needs at the time. Small porous particles in the range of 10 μm were not available in narrow particle-size distributions and in commercial quantities and packing procedures for micrometer-size particles into usable columns were not available. This all ended when Merck (Darmstadt, Germany) produced 5–10 μm narrow cuts of their thin-layer chromatography (TLC) grade silica gel and made them commercially available, and high-pressure slurry techniques were developed to reproducibly pack them.6 The first microparticulate column, MicroPak Si-10 silica gel, was introduced in 1972 by Varian Associates (Walnut Creek, California, USA).