In the leadoff article, columnist Ron Majors provides an overview of column developments. He looks at various alternatives
to high-throughput separations including small porous particles, monoliths and superficially-porous particles. Microfluidics
and parallel column systems provide further alternatives. An alternative approach to isocratic method development uses optimized
stationary phase combinations. Brief coverage of new phases for hydrophilic interaction chromatography, high temperature operation,
chiral and mixed mode columns and finally supercritical fluid chromatography columns round out the overview. At the conclusion,
Majors speculates on future directions in column technology.
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Since my article on the developments in high performance liquid chromatography (HPLC) column packing design (1) in the last
supplement published just before HPLC 2006 (2), there has been a continuing interest in column developments with improvements
in small, porous particle technology, monoliths (especially silica-based), superficially porous particles, and increased use
of high-temperature LC with new phases capable of operating up to 200 °C. For handling complex samples, there has been a renewed
interest in multidimensional and comprehensive LC×LC. In this update on column technology, I will cover some of the packing
and column developments and highlight observations made in the past two years and speculate in future directions in column
technology. Elsewhere in this special supplement other column topics will expand on changes that have occurred in stationary phases and
in multidimensional separations.
Further Developments in Small Porous Packings
Table I: Commercial two and sub-2-mm totally porous HPLC columns*
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One of the driving forces for continued improvements in HPLC column technology is the requirement for higher throughput but
without sacrificing analytical performance. One of the trends noted in the last two years is the strong interest in smaller
particles, especially those with average particle diameters less than 2 μm. In 2006, only a few companies were promoting the
virtues of sub-2-μm columns (1). Since further introductions at Pittcon 2008 (3), there are 14 companies that have commercialized
these small particles (Table I). When sub-2-μm particles are packed into short columns, separations can be performed faster,
sometimes in just a minute or two, than longer columns packed with larger particles without sacrificing chromatographic resolution.
The flat van Deemter curves noted for the sub-2-μm columns allow relatively high flow rates to be used, in the range of 2–3
mL/min if necessary. Even though the column pressure increases with the inverse square of the average particle diameter, these
shorter columns (usually 50 mm and under) can be used with most conventional HPLC pumping systems, even at these increased
flow rates.
For more demanding separations, longer columns of 100- and 150-mm lengths may be required. With such lengths, column back
pressure may increase beyond the capabilities of conventional pumping systems (~400 bar upper limit). Thus, in recent years,
pumping systems capable of operation up to 20,000 psi (1330 bar) have come onto the market. However, it is rare to see a separation
run at such high pressures, even at 1000 bar. User concerns about stress on instrument hardware and on the columns themselves
have limited widespread applications at these high pressures. The demands of sample cleanliness and easier "pluggability"
of the small porosity frits terminating the sub-2-μm columns also have made some users cautious into jumping into their routine
application. Nevertheless, these columns have proven to be rugged if used properly by ensuring particle-free samples are injected.
With the coming of low-dead-volume, high-pressure guard columns to help to protect the analytical column, liquid chromatographers
should become more comfortable using these columns.
Table II: Commercial two to three mm totally porous HPLC columns*
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Even so, some column and instrument suppliers have "backed-off" in joining the sub-2-μm bandwagon and have made more moderate
reduction in the particle size from the popular 3–3.5 μm columns (see Table II). Since for packed columns in the range of
2–3 μm, the particle diameter is larger than the sub-2-μm particles, the pressure drop is lower but efficiency is better than
the more popular 3–3.5 μm particles. The arguments for using packings in the 2–3 μm range evolve around considering the entire
separation cycle time where higher temperatures are used to improve efficiency and lower back pressure, improved liquid chromatograph
hydraulics to lower band dispersion and gradient reequilibration time, and faster autosamplers, detectors and data systems.
The use of lower pressures also places less stress on the instrumentation (for example, pump seals, pistons, check valves,
and injector valve cores) and on column materials.
So, it should be interesting to see if even smaller porous particles will be in vogue in the next two years. Currently, the
smallest particle size that is commercially available is 1.5 μm. The pressure "race" seems to be on in HPLC instrumentation
so there is no reason to believe that the instruments could not keep up with further reductions in particle size. However,
there are other competing technologies that do not expose the column and instrumentation to such dramatic pressure conditions:
monoliths and superficially porous packed columns.
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