Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Chapter 10 Supercritical fluid chromatography and Extraction (SFC & SFE) References: Roger M. Smith, Supercritical Fluid Chromatography, Skoog book, p768 S.A. Westwood, Supercritical Fluid Extraction and its use in chromatographic sample preparation,

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE 1.Principles 1.1Brief history To be able to use supercritical fluid (SF) as a chromatographic mobile phase, the SF has to have ability to dissolve substances. The dissolving ability of SE was discovered probably first by Hannay and Hogarth in 1879, when they studied the solubility of cobalt and iron chlorides in supercritical ethanol. Lovelock in 1958 suggested that a SF might be used as a mobile phase in chromatography.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE E. Klesper was first demonstrated in 1962 SFC by separation of nickel porphyrins using supercritical chlorofluoromethanes as mobile phases. In the following years (1960s), further developments were carried out both practically and theoretically by number of groups, among many, Sie, Rijnders, and Giddings.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Another important development is the demonstration of SFC with capillary columns by Novotny and M. Lee in One of the most interesting patents in the history of SFC was filed for in early 1982 by Novotny, Lee, Peaden et al. for the fundamental use of open-tubular capillary columns for SFC. This patent had aroused some controversy in the SFC industry, and consequently it was subjected to a review in late 1985 and On December 31, 1986 the claims of the patent were preliminary rejected. However, after amendments were made, the fundamental patent for capillary SFC patent was validated in March It is held by Brighham Young University and licensed exclusively to Lee Scientific and these groups have done much to promote the subsequent interest in SFC.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE

1.2Supercritical fluid

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE The critical temperature of a substance is the temperature above which a distinct liquid phase cannot exist, regardless of pressure. The vapor pressure of a substance at its critical temperature is its critical pressure. At temperature and pressures above its critical temperature and pressure (its critical point), a substance is called a supercritical fluid.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Several important properties of SF High density: remarkable ability to dissolve large, nonvolatile molecules (e.g extraction caffeine from coffee bean to give decaffeinated coffee and extracting nicotine from cigarette tobacco). Viscosity: reduced pressure drop across the column. Liquid viscosity is greater by a factor of ca. 100, the pressure drop in HPLC is between 10 to 100 times greater than in SFC and GC.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Solute diffusion coefficient: the mass transfer properties resulting from the solute diffusion coefficients in SF lead to the analysis speeds which increase in the sequence HPLC, SFC, and GC. Analyte recovery: in most cases analyte dissolved in the SF can be recovered easily by simply allowing the solution to equilibrate with the atmosphere at relatively low temperature.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE

1.3Supercritical Fluid Chromatography 1.3.1Mobile Phase in SFC The most common used mobile phase for supercritical fluid chromatography is carbon dioxide. Major advantages: Nontoxic, odorless Inexpensive Suitable critical temperature and pressure (31  C and 72.9 atm) Readily available

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Modify the solubility or the partition coefficient of the analyte: Change pressure, hence the density of the SF. Add polar organic modifiers such as methanol It is important to be aware of the modifier- fluid phase diagram to ensure that the solvent is in one phase.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE

2.Instrumental considerations Capillary SFC was developed on principles based on capillary GC Nearly all previous SFC instruments employed components normally used in conventional HPLC system, including high pressure pumps, stainless steel tubing, injection valves, and columns, with few modifications, or not at all.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE 2.1Stationary phase Both open-tubular and packed columns are used for SFC, although currently the former is favored. Both columns are similar to those used in GC and HPLC. 2.2Detectors –A major advantage of SFC over HPLC that the universal detector: flame ionization detector of gas chromatography can be employed. –Conventional detectors from both GLC and HPLC have been successfully adapted to SFC, such as UV, FID, FPD et al. –SFC/MS

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE

Challenges in SFC SFC faces a strong competition from the high- temperature Gas-liquid chromatography (GLC) and HPLC. By using high-temperature Gas-liquid chromatography (GLC), many high molecular weigh compounds can be separated. It is also facing competition from HPLC and CE. The future needs for SFC would be the development of instrumentation, particularly in sample introduction system, column and restrictor techniques, and the use of polar mobile phases.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE 3.Supercritical Fluid Extraction (SFE) An offline separation technique, or a sample preparation technique. 3.1Principles of SFE Three interrelated factors influence recovery of the target compounds as shown in the SFE triangle:

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Solubility Diffusion Matrix

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Solubility The solute must, firstly, be sufficiently soluble in the supercritical fluid. Solubility: –Change pressure –Add modifier

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Diffusion The solute must be transported sufficiently rapidly by “diffusion” from the interior of the matrix in which it is contained.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Matrix The third factor is that of the matrix (other than its effects on diffusion). Matrix effects mean that, although in many cases SFE will extract all of a particular compound in a sample, in some cases not all of a compound is “extractable”; the rest being locked into the structure of matrix, or too strongly bound to its surface.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Of these three factors, that of the matrix is the least well understood at present and a detailed scientific discussion cannot be given. Of the two remaining factors, solubility and diffusion, the latter is of more concerned in analytical extractions. This firstly because in most applications, the analyte is present in small quantities in the matrix and secondly a fluid and conditions will have to chosen in which the analyte is soluble at least to some extent.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE % Extracted Time Solubility is only important at the beginning of an extraction in a typical flow system.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE 3.2Instrumentation Offline method Dynamic The sample is continuously supplied with fresh supercritical fluid and the extracted analytes are constantly swept into the collection device. Static The out let of the extraction cell is shut off and the cell is pressed under static (non-flowing) conditions. Following an appropriate extraction time, the analytes are recovered from the static extraction, generally by opening a value at the outlet of the cell and performing a short dynamic extraction.

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE Dynamic mode is preferred for solubility controlled extraction Static mode is preferred for diffusion controlled extraction

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE

on-line SFE extraction

Advanced Analytical Chemistry – CHM 6157® Y. CAIFlorida International University Updated on 11/6/2006Chapter 10SFC-SFE