Putting Packed Column Supercritical Fluid Chromatography into Perspective

Introduction

What Is Packed Column SFC?

Packed column supercritical fluid chromatography (SFC or pSFC) is an analysis technique similar to liquid chromatography (LC) that uses supercritical fluids (SFs), instead of liquids, as the mobile phase (MP) (supercritical fluids are defined in the next section). The MP solvates the solutes. The stationary phase (SP) consists of a bed of very small particles packed in a tube capable of withstanding high pressures. Some SPs are the surfaces of uncoated particles. Some are organic films bonded to the surface of the particles. Solutes are separated by differential attraction to the SP.

Compared with LC, packed column SFC is faster, more efficient, has a wider range of selectivity, and detection options, and produces less toxic waste. Not surprisingly, the fields most likely to be affected by packed column SFC in the future are traditional LC application areas. In particular, pharmaceutical and agricultural chemical development will derive significant benefits. Chiral separations will likely be a major application area for packed column SFC. This list is likely to surprise many readers since the application areas most often associated with capillary SFC have involved less polar but perhaps more complex solute mixtures such as homologous series, of surfactants, polymers, and the like.

In reality, the characteristics of interest in 'SFC' have more to do with intermolecular interactions in the MP than the name of the fluid. Many of the characteristics that make SFs interesting to chromatographers (e.g., high diffusivity, low viscosity) are also available from some fluids defined as gases or liquids. Unfortunately, the name SFC is somewhat misleading. SFC differs from LC in that the MP is a dense compressed fluid which will dramatically expand if external pressure is removed.

The most widely used supercritical fluids (like carbon dioxide, nitrous oxide, or CHF3) are inorganic and do not produce a response in some GC detectors, like the FID. This combination of characteristics allows some LC-like separations with more GC-like figures of merit, such as high speed, high resolution, and multiple detection options.

Several modern packed column SFC chromatograms may help convey the features that make the technique desirable. In Figure 1.1, the 11 carbamates of EPA Method 531.1 are separated in 9 minutes and directly detected using both a UV and an NPD (Nitrogen–Phosphorus Detector).

The methanol/carbon dioxide (MeOH/CO2) MP flow rate is 2.5 ml min-1, producing near optimum chromatographic efficiency. This is approximately 3.5 times the optimum flow rate in LC (on this column) and illustrates the superior diffusion rate in supercritical fluids.

The standard method uses gradient elution LC2 followed by two postcolumn reactions to yield fluorescent products. Although the separation takes ca. 40 minutes, the column must then be re-equilibrated. The whole process requires ca. 1 hour between injections. A representative chromatogram is shown in Figure 1.2.

Cumulatively, the SFC separation and detection options produce a throughput approximately six times that of the LC standard method, and avoid the complexity of the postcolumn reactions.

An alternative example of the unusual characteristics of SFC is shown in Figure 1.3. The separation in Figure 1.3 was developed to suggest the feasibility of using SFC for screening pesticides not amenable to GC analysis. A 10 ml water sample containing 6–22.5 p.p.b. of 31 carbamate, sulfonylurea, phenylurea, and triazine pesticides was injected into a precolumn mounted in place of an external loop on a six port valve. The water was blown off with helium, and then the precolumn was switched into the flowing stream. The solutes were eluted by a gradient of 1–16% MeOH in CO2, 90–140 bar, from a 1.6 m long LC column packed with 5 µm particles. At 2 ml min-1 of 20% MeOH, the pressure drop was 150 bar. After the column, the flow was split, diverting a fraction to an ECD and an NPD while most passed through the UV diode array detector. The detection limits for some solutes were a few tens of parts per trillion (1/10).

One trend in LC is toward the use of smaller diameter packed columns, requiring less MP. Major reasons are a desire to reduce solvent cost and minimize toxic waste generation. In some locations, it is already more expensive to dispose of solvents than purchase them. Unfortunately, smaller columns require more stringent instrumental design. In general, it is more difficult to achieve the same high efficiencies, and high sensitivity on a small column as on a large column. Packed column SFC offers an attractive alternative. Inert CO2 replaces most of the liquid solvent. Modifiers typically represent 2–20% of the mobile phase. An SFC method on a 4.6 mm column creates the same or less liquid waste as a 2 or even 1 mm LC column. By retaining the larger column format, SFC allows relaxed constraints on extra column effects, while often providing higher capacity, better detection, and reproducibility.

What is a Supercritical Fluid?

It is important to understand that 'Supercritical' is only a defined state. Supercritical fluids are not a separate state of matter (there are only gases, liquids, and solids). To be 'supercritical', a fluid must be above BOTH its critical temperature, Tc, and critical pressure, Pc. The combination of Tc and Pc is known as the 'critical point'. Above its critical point, a fluid cannot be liquified, no matter how high the pressure is raised. Note that the definition only deals with T >Tc and P >Pc. The definition ignores what happens at conditions BELOW the critical point.

There has been a great deal of confusion about transitions from subcritical to supercritical conditions. Such transitions are NOT phase transitions. They are only transitions from one DEFINED state to another. This ambiguity is dealt with in depth in Chapter 3.

Supercritical fluids lack adequate intermolecular interactions which would otherwise condense them to liquids. This low intermolecular energy gives the fluids certain advantageous characteristics compared with normal liquids familiar as mobile phases in LC.

With SFs (and some similar fluids), the pressure can be increased until the molecules are as close to each other as the molecules in a condensed liquid. This molecular...