Supercriticalfluid chromatography


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Supercriticalfluid chromatography

  1. 1. Supercritical fluid chromatography
  2. 2. Supercritical Fluid Chromatography Contents • History • What is a supercritical fluid? • Theory of SFC • Instrumentation • Applications
  3. 3. History  The use of a supercritical fluid mobile phase in chromatography was first proposed in 1958 by J. Lovelock.  The first actual report use of this in a chromatographic system was in 1962 by Klesper et al, who used it to separate thermally-labile porphyrins.
  4. 4. S.F.C • S.F.C is a column chromatographic technique in which supercritical fluid is used as a mobile phase. • Cost efficient • User friendly • Better resolution • Faster analysis
  5. 5. What is a supercritical fluid? Supercritical Fluid exists at temperatures and pressures above its critical temperature and pressure and have densities, viscosities and other properties that are intermediates between those of the substance in its gaseous and liquid state.
  6. 6. Properties of Supercritical Fluid • High densities so they have a remarkable ability to dissolve large, non-volatile molecules . • Lower viscosities relative to liquid solvents. • Inexpensive • Ecofriendly • Non-toxic. • Higher diffusion coefficients and lower viscosities relative to liquids.
  7. 7.    SFC Advantages vs HPLC Supercritical fluids have low viscosities - faster analysis (5 to 10 X faster). - the use of open tubular columns is feasible. Column lengths from 10 to 20 m are used. Resolving power is ~5X that of HPLC.  SFC Advantages vs GC Can analyze non-volatile, polar, or adsorptive solutes without derivatization.  Can analyze thermally labile compounds.  Can analyze solutes of much higher molecular weight.
  8. 8. Theory :     based on the density of the supercritical fluid which corresponds to solvating power. As the pressure in the system is increased, the supercritical fluid density increases and correspondingly its solvating power increases. Therefore, as the density of the supercritical fluid mobile phase is increased, components retained in the column can be made to elute. This is similar to temperature programming in GC or using a solvent gradient in HPLC.
  9. 9. SFC Instrumentation  Gas Supply or Mobile Phase  Stationary Phase  Pumps  Injectors  SFC Columns  Detector  Restrictor
  10. 10. Gas supply or Mobile Phase: Liquid CO2 Pump Cost  interference with chromatographic detectors  physical properties like nontoxic nature nonflammable  are considered while selecting a mobile phase.
  11. 11. Fluid Critical Temperature (K) Critical Pressure (bar) Carbon dioxide 304.1 73.8 Ethane 305.4 48.8 Ethylene 282.4 50.4 Propane 369.8 42.5 Propylene 364.9 46.0 Trifluoromethane (Fluoroform) 299.3 48.6 Chlorotrifluoromethane 302.0 38.7 Trichlorofluoromethane 471.2 44.1 Ammonia 405.5 113.5 Water 647.3 221.2 Cyclohexane 553.5 40.7 n-Pentane 469.7 33.7 Toluene 591.8 41.0
  12. 12. Carbon dioxide is the ideal to satisfy all the above properties.  Safe to use nontoxic nonflammable noncorrosive inert.  Detector compatible • The main disadvantage of it is very polar or ionic compounds are not able to be eluted. • This can be overcome by adding a small portion of a second fluid called a Modifier fluid. (alcohols, cyclic ethers, acetonitrile and chloroform). 
  13. 13. Modifiers    CO 2 is not a very good solvent for high molecular weight, ionic and polar analytes This can be overcome by adding a small portion of a second fluid called modifier fluid This is generally an organic solvent, which is completely miscible with carbon dioxide  methanol, acetonitrile, ethanol and 1-propanol.
  14. 14. Stationary Phase Same as those for GC and LC, with some modification. • Silica/Alumina Useful for non-polar compounds Lead to irreversible adsorption of some polar solutes • Widely used polar Stationary Phase are Polysiloxanes:- stable, flexible Si--O bond lead to good diffusion. Substituted with chemical groups for selective interaction with analyte Polymethylsiloxanes:- increase efficiency in separating closely related polar analytes Cyanopropyl polysiloxanes:-useful for compounds with -COOH
  15. 15. Pumps:  Here mainly flow control is necessary.  So, syringe pumps are used for capillary SFC for consistent pressure.  For Packed columns for easier blending of the mobile phase or introduction of modifier fluids reciprocating pumps are used. Syringe Pump
  16. 16. Injector •   For packed column SFC, a conventional HPLC injection system is adequate, but for the capillary column SFC, the sample volume depends on column diameters and small sample volumes must be quickly injected into the column, therefore pneumatically driven valves are used. Injector Volumes Open tubular columns ◦ Injection volumes >96nL ◦ Greater volume affects resolution Packed columns ◦ Injection volumes >1uL
  17. 17. Oven : Conventional GC or LC ovens are used
  18. 18. SFC Columns • Open tubular (derived from GC) -Large theoretical plates ~X500 -Easier to control pressure (low P drop) • Packed (derived from HPLC) - Faster analysis - Higher flow rates - Higher sample capacity
  19. 19. Detectors The choice of detectors will depend upon • • • • mobile phase composition column type flow rate ability to withstand the high pressures of SFC.
  20. 20. Contd.. • • • It is compatible with both HPLC and GC detectors. Flame photometric detectors Flame ionization detectors • liquid-phase detectors like RID, ultraviolet-visible spectrophotometric detectors and • light scattering detectors have been employed for SFC.
  21. 21. Back-Pressure Device or Restrictor • This is a device, which is used to maintain desired pressure in the column by - a pressure-adjustable diaphragm or - controlled nozzle so that the same column-outlet pressure is maintained irrespective of the mobile phase pump flow rate. • It keeps the mobile phase supercritical throughout the separation and often must be heated to prevent clogging. • The pressure restrictor is placed either after the detector or at the end of the column.
  22. 22. Applications of SFC By now SFC has been applied to wide variety of materials.         natural products drugs foods pesticides herbicides surfactants polymers and polymer additives chiral compound
  23. 23. Natural products •   separation of underivatized triterpene acids estimation of caffeine from tea and conjugated bile acids analysis of panaxadiol / panaxatriol in ginseng . Pesticides • analysis of pesticide residues in canned foods, fruits and vegetables wherein pyrethroids, herbicides, fungicides and carbamates have been tested .
  24. 24. Surfactants Separation of the oligomers in a sample of the nonionic surfactant Triton X100 . Lipids for the analysis of high molecular weight lipids like triacylglycerols. Separation of fatty acid methyl esters, biosynthetic polyunsaturated fatty acids (PUFA) 37 , nonsaponifiable lipids , cholesterol and its esters in human serum and food samples
  25. 25. Drugs        Separation of drugs like phenothiazine antipscychotics, beta blockers felodipine clevidipine methylated betacyclodextrins vasodialators like isosorbide mononitrate, isosorbide dinitrate, cyclandelate, nimodipine, amlodipine , oestrogens combinations of various nonsteroidal antiinflammatory drugs like flufenamic acid, mefenamic acid, fenbufen, indomethacin mixtures, acetyl salicylic acid, ketoprofen and fenbufen
  26. 26. Chiral compounds SFC has been applied to separation of a large number of enantiomers, diasterioisomers and geometrical isomers like achiral and chiral analysis of camazepam and its metabolites, chiral separation of 1,3 dioxolane derivatives Organometallics Separation of metal chelates and organometals of thermally labile category, chelates of transition metals, heavy metals, organometallic compounds of lead, mercury and tin has been carried out by SFC.
  27. 27. Conclusion In the overall ranking of chromatographic techniques, it has been judged that SFC falls somewhere between HPLC and GC as the chromatographic method of choice. Lately SFC has found a niche in the field of pharmaceutical chemistry and has gained much support in the field of bioanalytical applications
  28. 28. References  http://www.chemcenter/org     Instrumental analysis by Skoog Heller  Instrumental methods of analysis by Kaur   