GAS LIQUID CHROMATOGRAPHY
• Principles
Partition of molecules between gas (mobile phase) and liquid (stationary phase).
Gas Liquid Chromatography
Most Common Stationary Phases
1. Separation of mixture of polar compoundsCarbowax 20M (polyethylene glycol)
2. Separation of mixtures of non-polar compoundsOV101 or SE-30 (polymer of methylsilicone)
3. Methylester of fatty acidsDEGS (diethylene glycol succinate)
Filters/Traps
Air
Hyd
rog
en
Gas C
arrier
Column
Gas Chromatography
• gas system• inlet• column• detector• data system
Data system
Syringe/Sampler
Inlets
Detectors
Regulators
H
RESET
Schematic Diagram of Gas Chromatography
Detector
• 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.
Exhaust
Chimney
Igniter
Hydrogen Inlet
Column Effluent
Collector Electrode
Schematic Diagram of Flame Ionization Detector
Measures the changes of thermal conductivity due to the sample (g). Sample can be recovered.
Thermal Conductivity Detector
Thermal Conductivity Detector
Principal: The thermal balance of a heated filament
Electrical power is converted to heat in a 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 by conduction to the molecules which contact with the filament.
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.
The TCD is a nondestructive, concentration sensing detector. A heated filament is cooled by the flow of carrier gas.
Flo
w
Flo
w
When a separated compound elutes from the column , the thermal conductivity of the mixture of carrier gas and compound gas is lowered. The filament in the sample column becomes hotter than the control column.
The imbalance between control and sample filament temeprature is measured by a simple gadget and a signal is recorded
Thermal Conductivity Detector
Relative Thermal Conductivity
Compound Relative Thermal Conductivity
Carbon Tetrachloride 0.05
Benzene 0.11
Hexane 0.12
Argon 0.12
Methanol 0.13
Nitrogen 0.17
Helium 1.00
Hydrogen 1.28
Thermal Conductivity Detector
• Responds to all compounds
• Adequate sensitivity for many compounds
• Good linear range of signal
• Simple construction
• Signal quite stable if carrier gas glow rate, block temperature, and filament power are effectively controlled
• Nondestructive detection
Thermal Conductivity Detector
Electron Capture Detector
Analyses for pesticide, Insecticides, vinyl chloride, and fluorocarbons in foods.
Most sensitive detector (10-12 gram)
Electron Capture Detector
ECD detects positive ions of carrier gas by the anode electrode.
63Ni emits particles.
Ionization : N2 (Carrier gas) + (e) = N2+ + 2e. The N2
+ establish a “base line”
X (F, Cl and Br) containing sample + (e) X-
Ion recombination: X- + N2+ = X + N2, The “base line” due to the
N2+ will decrease and this decrease constitutes the signal.
The more the halogen containing X compounds in the sample, the less the N2
+ in the detector
Electron Capture Detector
Electron Capture Detector
Chromatogram of Compounds from Fermented Cabbage
Chromatogram of Orange Juice Compounds
Gas Chromatography Application
Semi-Quantitative Analysis of Fatty Acids
C
C
C
Dete
ctor
Resp
onse
Retention Time
14
16
18
Pea
k A
rea
Sample Concentration (mg/ml)
2
4
6
8
10
0.5 1.0 1.5 2.0 2.5 3.0
The content % of C fatty acids =C
C + C + C
= the content % of C fatty acids14
14
Tentative Identification of Unknown Compounds
Res
pons
e
GC Retention Time on Carbowax-20 (min)
Mixture of known compounds
Hexane
Octane Decane1.6 min = RT
Res
pons
e
Unknown compound may be Hexane
1.6 min = RT
Retention Time on Carbowax-20 (min)
Res
p on s
e
GC Retention Time on SE-30
Unknown compound
RT= 4 min on SE-30
Res
pons
e
GC Retention Time on SE-30
HexaneRT= 4.0 min on SE-30
Retention Times
Advantages of Gas Chromatography
• Very good separation
• Time (analysis is short)
• Small sample is needed - l
• Good detection system
• Quantitatively analyzed
Disadvantages of Gas Chromatography
Material has to be volatilized at 250C without decomposition.
R C OH CH3OH H2SO4
O
R C O CH3
O
CH2 O C R
CH O C R
CH2 O C R
O
O
O
CH3OH
O
R C O CH3
CH3ONa
Fatty Acids Methylester
Reflux
+ 3
Volatile in Gas Chromatography
Volatile in Gas Chromatography
+ +
Gas Chromatogram of Methyl Esters of Fatty Acids
Effects of OH groups of Carbohydrates
OH
O
OH
OHHO
CH2
OH
1
23
4
5
6
OH
O
OH
OHHO
CH2OH
1
23
45
6
+ Si
CH3
CH3
CH35Cl
O-Si(CH3)3
O
O-Si(CH3)3
O-Si(CH3)3(CH3)3-Si-O
CH2O-Si(CH3)3
1
23
45
6
5HCl+
Derivation of Glucose with Trimethylchlorosilane
Glucose Trimethylchlorosilane
Effects of Derivation
• Time consumption
• Side reaction
• Loss of sample
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