The discovery of supercritical fluids led to novel analytical applications in the fields of chromatography and extraction known as supercritical fluid chromatography (SFC) and supercritical fluid extraction (SFE). Supercritical fluid chromatography is accepted as a column chromatography methods along with gas chromatography (GC) and high-performance liquid chromatography (HPLC). Due to to the properties of supercritical fluids, SFC combines each of the advantages of both GC and HPLC in one method. In addition, supercritical fluid extraction is an advanced analytical technique.
Definition and formation of supercritical fluids
A supercritical fluid is the phase of a material at critical temperature and critical pressure of the material. Critical temperature is the temperature at which a gas cannot become liquid as long as there is no extra pressure; and, critical pressure is the minimum amount of pressure to liquefy a gas at its critical temperature. Supercritical fluids combine useful properties of gas and liquid phases, as it can behave like both a gas and a liquid in terms of different aspects. A supercritical fluid provides a gas-like characteristic when it fills a container and it takes the shape of the container. The motion of the molecules are quite similar to gas molecules. On the other hand, a supercritical fluid behaves like a liquid because its density property is near liquid and, thus, a supercritical fluid shows a similarity to the dissolving effect of a liquid.
The characteristic properties of a supercritical fluid are density, diffusivity and viscosity. Supercritical values for these features take place between liquids and gases. Table demonstrates numerical values of properties for gas, supercritical fluid and liquid.
| Gas | Supercritical fluid | Liquid | |
|---|---|---|---|
| Density (g/cm3) | 0.6 x 10-3 – 2.0 x 10-3 | 0.2 – 0.5 | 0.6 – 2.0 |
| Diffusivity (cm2/s) | 0.1 – 0.4 | 10-3 – 10-4 | 0.2 x 10-5 – 2.0 x 10-5 |
| Viscosity (cm/s) | 1 x 10-4 – 3 x 10-4 | 1 x 10-4 – 3 x 10-4 | 0.2 x 10-2 – 3.0 x 10-2 |
The formation of a supercritical fluid is the result of a dynamic equilibrium. When a material is heated to its specific critical temperature in a closed system, at constant pressure, a dynamic equilibrium is generated. This equilibrium includes the same number of molecules coming out of liquid phase to gas phase by gaining energy and going in to liquid phase from gas phase by losing energy. At this particular point, the phase curve between liquid and gas phases disappears and supercritical material appears.
In order to understand the definition of SF better, a simple phase diagram can be used. Figure displays an ideal phase diagram. For a pure material, a phase diagram shows the fields where the material is in the form of solid, liquid, and gas in terms of different temperature and pressure values. Curves, where two phases (solid-gas, solid-liquid and liquid-gas) exist together, defines the boundaries of the phase regions. These curves, for example, include sublimation for solid-gas boundary, melting for solid-liquid boundary, and vaporization for liquid-gas boundary. Other than these binary existence curves, there is a point where all three phases are present together in equilibrium; the triple point (TP).
There is another characteristic point in the phase diagram, the critical point (CP). This point is obtained at critical temperature (Tc) and critical pressure (Pc). After the CP, no matter how much pressure or temperature is increased, the material cannot transform from gas to liquid or from liquid to gas phase. This form is the supercritical fluid form. Increasing temperature cannot result in turning to gas, and increasing pressure cannot result in turning to liquid at this point. In the phase diagram, the field above Tc and Pc values is defined as the supercritical region.
In theory, the supercritical region can be reached in two ways:
- Increasing the pressure above the Pc value of the material while keeping the temperature stable and then increasing the temperature above Tc value at a stable pressure value.
- Increasing the temperature first above Tc value and then increasing the pressure above Pc value.
The critical point is characteristic for each material, resulting from the characteristic Tc and Pc values for each substance.
Supercritical fluid chromatography (SFC)
Just like supercritical fluids combine the benefits of liquids and gases, SFC bring the advantages and strong aspects of HPLC and GC together. SFC can be more advantageous than HPLC and GC when compounds which decompose at high temperatures with GC and do not have functional groups to be detected by HPLC detection systems are analyzed.
There are three major qualities for column chromatographies:
- Selectivity.
- Efficiency.
- Sensitivity.
Generally, HPLC has better selectivity that SFC owing to changeable mobile phases (especially during a particular experimental run) and a wide range of stationary phases. Although SFC does not have the selectivity of HPLC, it has good quality in terms of sensitivity and efficiency. SFC enables change of some properties during the chromatographic process. This tuning ability allows the optimization of the analysis. Also, SFC has a broader range of detectors than HPLC. SFC surpasses GC for the analysis of easily decomposable substances; these materials can be used with SFC due to its ability to work with lower temperatures than GC.
Instrumentation for SFC
As it can be seen in Figure, SFC has a similar setup to an HPLC instrument. They use similar stationary phases with similar column types. However, there are some differences. Temperature is critical for supercritical fluids, so there should be a heat control tool in the system similar to that of GC. Also, there should be a pressure control mechanism, a restrictor, because pressure is another essential parameter in order for supercritical fluid materials to be kept at the required level. A microprocessor mechanism is placed in the instrument for SFC. This unit collects data for pressure, oven temperature, and detector performance to control the related pieces of the instrument.
Stationary phases
SFC columns are similar to HPLC columns in terms of coating materials. Open-tubular columns and packed columns are the two most common types used in SFC. Open-tubular ones are preferred and they have similarities to HPLC fused-silica columns. This type of column contains an internal coating of a cross-linked siloxane material as a stationary phase. The thickness of the coating can be 0.05-1.0 μm. The length of the column can range from of 10 to 20 m.
Mobile phases
There is a wide variety of materials used as mobile phase in SFC. The mobile phase can be selected from the solvent groups of inorganic solvents, hydrocarbons, alcohols, ethers, halides; or can be acetone, acetonitrile, pyridine, etc. The most common supercritical fluid which is used in SFC is carbon dioxide because its critical temperature and pressure are easy to reach. Additionally, carbon dioxide is low-cost, easy to obtain, inert towards UV, non-poisonous and a good solvent for non-polar molecules. Other than carbon dioxide, ethane, n-butane, N2O, dichlorodifluoromethane, diethyl ether, ammonia, tetrahydrofuran can be used. Table shows select solvents and their Tc and Pc values.
| Solvent | Critical Temperature (°C) | Critical Pressure (bar) |
|---|---|---|
| Carbon dioxide (CO2) | 31.1 | 72 |
| Nitrous oxide (N2O) | 36.5 | 70.6 |
| Ammonia (NH3) | 132.5 | 109.8 |
| Ethane (C2H6) | 32.3 | 47.6 |
| n-Butane (C4H10) | 152 | 70.6 |
| Diethyl ether (Et2O) | 193.6 | 63.8 |
| Tetrahydrofuran (THF, C4H8O) | 267 | 50.5 |
| Dichlorodifluoromethane (CCl2F2) | 111.7 | 109.8 |
Detectors
One of the biggest advantage of SFC over HPLC is the range of detectors. Flame ionization detector (FID), which is normally present in GC setup, can also be applied to SFC. Such a detector can contribute to the quality of analyses of SFC since FID is a highly sensitive detector. SFC can also be coupled with a mass spectrometer, an UV-visible spectrometer, or an IR spectrometer more easily than can be done with an HPLC. Some other detectors which are used with HPLC can be attached to SFC such as fluorescence emission spectrometer or thermionic detectors.
Advantages of working with SFC
The physical properties of supercritical fluids between liquids and gases enables the SFC technique to combine with the best aspects of HPLC and GC, as lower viscosity of supercritical fluids makes SFC a faster method than HPLC. Lower viscosity leads to high flow speed for the mobile phase.
Thanks to the critical pressure of supercritical fluids, some fragile materials that are sensitive to high temperature can be analyzed through SFC. These materials can be compounds which decompose at high temperatures or materials which have low vapor pressure/volatility such as polymers and large biological molecules. High pressure conditions provide a chance to work with lower temperature than normally needed. Hence, the temperature-sensitive components can be analyzed via SFC. In addition, the diffusion of the components flowing through a supercritical fluid is higher than observed in HPLC due to the higher diffusivity of supercritical fluids over traditional liquids mobile phases. This results in better distribution into the mobile phase and better separation.
Applications of SFC
The applications of SFC range from food to environmental to pharmaceutical industries. In this manner, pesticides, herbicides, polymers, explosives and fossil fuels are all classes of compounds that can be analyzed. SFC can be used to analyze a wide variety of drug compounds such as antibiotics, prostaglandins, steroids, taxol, vitamins, barbiturates, non-steroidal anti-inflammatory agents, etc. Chiral separations can be performed for many pharmaceutical compounds. SFC is dominantly used for non-polar compounds because of the low efficiency of carbon dioxide, which is the most common supercritical fluid mobile phase, for dissolving polar solutes. SFC is used in the petroleum industry for the determination of total aromatic content analysis as well as other hydrocarbon separations.
Resource:
- D. A. Skoog and J. J. Leary, Principles of Instrumental Analysis, Saunders College Publishing, Philadelphia (1992).
- M. Caude and D. Thiebaut, Practical Supercritical Fluid Chromatography and Extraction, Harwood Academic Publishers, Switzerland (1999).
