GAS LIQUID CHROMATOGRAPHY

1. GAS LIQUID CHROMATOGRAPHY Principles Partition of molecules between gas (mobile phase) and liquid (stationary phase).

2. Most Common Stationary Phases • 1. Separation of mixture of polar compounds • Carbowax 20M (polyethylene glycol) • 2. Separation of mixtures of non-polar compounds • OV101 or SE-30 (polymer of methylsilicone) • Methylester of fatty acids • DEGS (diethylene glycol succinate)

3. H RESET Air Hydrogen Gas Carrier Gas Chromatography Filters/Traps Data system • gas system • inlet • column • detector • data system Regulators Syringe/Sampler Inlets Detectors Column

4. Schematic Diagram of Gas Chromatography

5. Schematic Diagram of Gas Chromatography

6. DETECTORS Flame Ionization Detector (Nanogram - ng) High temperature of hydrogen flame (H2 +O2 + N2) ionizes compounds eluted from column into flame. The ions collected on collector or electrode and were recorded on recorder due to electric current.

7. Exhaust Chimney Collector Electrode Igniter Polarizing Electrode Column Hydrogen Effluent Inlet Schematic Diagram of Flame Ionization Detector

8. Schematic Diagram of Flame Ionization Detector Collector Detector electronics  - 220 volts Flame Chassis ground Jet Signal output Column

9. Thermal Conductivity Detector Measures the changes of thermal conductivity due to the sample (mg). Sample can be recovered.

10. Thermal Conductivity Detector Principal: The thermal balance of a heated filament Electrical power is converted to heat in a resistant filament and the temperature will climb until heat power loss form the filament equals the electrical power input. The filament may loose heat by radiation to a cooler surface and by conduction to the molecules coming into contact with it.

11. The TCD is a nondestructive, concentration sensing detector. A heated filament is cooled by the flow of carrier gas . Thermal Conductivity Basics When the carrier gas is contaminated by sample , the cooling effect of the gas changes. The difference in cooling is used to generate the detector signal. Flow Flow

12. Thermal Conductivity Detector When a compound elutes, the thermal conductivity of the gaseous mixture of carrier gas and compound gas is lowered, and the filament in the sample column becomes hotter than the other control column. Its resistance increased, and this imbalance between control and sample filament resistances is measured by a simple gadget and a signal is recorded

13. Thermal Conductivity Detector

14. Relative Thermal Conductivity

15. Thermal Conductivity Detector • Responds to all compounds • Adequate sensitivity for many compounds • Good linear range of signal • Simple construction • Signal quite stable provided carrier gas glow rate, block temperature, and filament power are controlled • Nondestructive detection

16. Electron Capture Detector For pesticide analysis (picogram). Accept electrons of carrier gas.

17. Electron Capture Detector ECD detects ions in the exiting from the gas chromatographic column by the anode electrode. 3H or 63Ni which emits  particles. Ionization : N2 (Nitrogen carrier gas) +  (e) = N2+ + 2e These N2+ establish a “base line” X (F, Cl and Br) containing sample +  (e)  X- Ion recombination : X- + N2+ = X + N2 The “base line” will decrease and this decrease constitutes the signal. Insecticides, pesticides, vinyl chloride, and fluorocarbons

18. Electron Capture Detector

19. Electron Capture Detector

20. Gas Chromatography Application

21. SEMI- QUANTITATIVE ANALYSIS OF FATTY ACIDS

22. TENTATIVE IDENTIFICATION OF UNKNOWN COMPOUNDS

23. Retention Times

24. GLC ADVANTAGES 1. Very good separation 2. Time (analysis is short) 3. Small sample is needed - ml 4. Good detection system 5. Quantitatively analyzed

25. DISADVANTAGES OF GAS CHROMATOGRAPHY Material has to be volatilized at 250C without decomposition.

26. Gas Chromatogram of Methyl Esters of Fatty Acids

27. 6 6 CH CH OH OH 2 2 O O 5 5 4 4 1 1 OH OH HO HO OH OH 2 2 3 3 OH OH The Effects of OH groups of Carbohydrates

28. CH 3 Si 5Cl CH 3 CH 3 6 CH OH 2 O 5 4 1 OH HO OH 2 3 OH Derivation of Glucose with Trimethylchlorosilane + Glucose Trimethylchlorosilane 6 CH O-Si(CH3)3 2 O 5 + 5HCl 4 1 O-Si(CH3)3 (CH3)3-Si-O O-Si(CH3)3 2 3 O-Si(CH3)3

29. Effects of Derivation 1. Time consumption 2. Side reaction 3. Loss of sample

30. THIN LAYER CHROMATOGRAPHY Stationary Phase ---------> Silica Gel Mobile Phase -------------> Solvent (developing)

31. THIN LAYER CHROMATOGRAPHY

33. Thermal Conductivity Detector The detector contains two filaments: one exposed only to carrier gas, while the other is exposed to the carrier gas for sample analysis. When the gas for the sample analysis is only carrier gas , the two filaments can be balanced. Instead of a direct measurement of filament temperature, the filament resistant, which is a function of temperature, is measured.

34. Thermal Conductivity Detector The ability of a colliding molecule to carry off heat depending on its thermal conductivity. Hydrogen and helium have high thermal conductivity and therefore will be more efficient at “cooling” a heated filament than other gases will

35. Thermal Conductivity Detector The TCD will respond to any substance different from the carrier gas as long as its concentration is sufficiently high enough.

36. Thermal Conductivity Detector

37. Thermal Conductivity Detector

38. Electron Capture Detector Electron capture compound, X (highly electonegative element), tends to capture free electrons and increase the amount to ion recombination X (F, Cl and Br) + e  X- Ion recombination : X- + N2+ = X + N2 The current will decrease and this decrease constitutes the signal. Halogens, lead, phosphorous, nitro groups, silicone and polynuclear aromatics. Insecticides, pesticides, vinyl chloride, and fluorocarbons

39. Electron Capture Detector