1. Basic Gas Chromatography
Prepared by: Mina S. Buenafe

2. Gas Chromatography Chromatography – A Very Brief History
Definitions / Terminologies in GC
Instrumentation Overview
System Modules
Mobile Phase (Carrier Gas)
Inlets
Stationary Phase(s)
Columns (Packed and Capillary)
Detector(s)
Troubleshooting

3. Chromatography – A (Very) Brief History
IN THE EARLY 1900’S
M. Tswett published his work on separation of plant pigments. He coined the term chromatography (literally translated as color writing) and scientifically described the process – earning him the title “Father of Chromatography”
W. Ramsey published his work on separation of mixture of gases and vapors on adsorbents like charcoal.
IN THE EARLY 1940’s
A. Martin and R. Synge first suggested the possibilities of gas chromatography in a paper published in Biochem. J., v.35, 1358, (1941). Martin won a Nobel Prize for his work in Partition chromatography.
IN THE EARLY 1950’s
A. Martin and A. James published the epic paper describing the first gas chromatograph

4. Definition of Terms Chromatography:
A physical method of separation in which the components to be separated are distributed between two phases, one of which is stationary while the other moves in a definite direction
“Official” IUPAC definition

5. Definition of Terms Chromatogram
It is the output signal from the detector of the instrument.

6. Definition of Terms Distribution Constant (KC)
It is the tendency of a given component to be attracted to the stationary phase. This can be expressed in chemical terms as an equilibrium constant. Also called the partition coefficient (KP) or the distribution coefficient (KD)
KC = [A]S/[A]M
Mathematically, it is defined as the concentration of solute A in the stationary phase divided by its concentration in the mobile phase.

7. Definition of Terms
The attraction to the stationary phase can also be classified according to the type of sorption by the solute.
Adsorption: sorption on the surface of the stationary phase
Absorption: sorption into the bulk of the stationary phase (usually called ‘partition’ by chromatographers)

8. Definition of Terms Retention Volume (VR)
It is usually defined as the distance between the point of injection to the peak maximum. It is the volume of the carrier gas necessary to elute the solute of interest.
Mathematically: VR = FC x tR
Where FC is the constant flow rate
tR is the retention time

9. Definition of Terms Phase Ratio (b) For packed columns:
b = Mobile Phase Volume
Stationary Phase Volume
For capillary columns:
b = rc/2df
Where rc is the radius of the column
df is the thickness of the film

10. Definition of Terms Retention Factor (k)
It is the ratio of the amount of the solute (NOT concentration) in the stationary phase to the amount in the mobile phase. It is also called capacity factor (k’), capacity ratio, or partition ratio
Mathematically:
k = (WA)S/(WA)M = KC/b
Also k = (tR - t0) = time in stationary phase
t time in carrier gas
k is temperature and flow dependent. Best separations occur when k is between 5 and 7

11. Definition of Terms Theoretical Plates (N)
This is the most common measure of column efficiency in chromatography
N = 16(tR/Wb)2 = 5.54(tR/ Wh)2
Where Wb is the peak width at the base
Wh is the peak width at half-height

12. Definition of Terms Height Equivalent to a Theoretical Plate (H)
This is a related parameter that also defines column efficiency. Also identified as HETP
Mathematically:
H = L/N
Where L is the Column Length
(An efficient column will have a large N and a small H)

13. Definition of Terms Separation Factor (a)
It is a measure of relative distribution constants. Also known as selectivity and/or solvent efficiency.
Mathematically: a = k2/k1 = (KC)2/(KC)1
It is dependent on:
Chemical composition of the phase
Partitioning between the two phases

14. Definition of Terms Resolution (Rs)
It is the degree to which adjacent peaks are separated.
Mathematically:
Rs = (tR)B – (tR)A
[(Wb)B + (Wb)A]/2
Also Rs =—L/H x k/(k+1) x a-1/a

15. Instrumentation Overview
Schematic of a Typical Gas Chromatograph

16. Very dry Nitrogen or Argon, 5% Methane
System Modules
Carrier Gas
Main purpose: carries the sample through the column
Secondary purpose: provides a suitable matrix for the detector to measure the sample component.
Detector
Carrier Gas
Thermal Conductivity
Helium
Flame Ionization
Helium or Nitrogen
Electron Capture
Very dry Nitrogen or Argon, 5% Methane
Carrier gases should be of high purity (minimum of %).
Oxygen & water impurities can chemically attack the liquid phase of the column and destroy it.
Trace water content can desorb other column contaminants and produce high detector background or ‘ghost peaks’.
Trace hydrocarbon contents can cause high detector background with FID’s and limit detectability.

17. System Modules Carrier Gas Flow Measurements and Control:
Essential for column efficiency and qualitative analysis (e.g. reproducibility of retention times)
Average linear flow velocity (ū) in OT columns:
ū = L/tm
where L is column length in cm
tm is the retention time of an unretained peak (e.g. methane) in sec
To convert linear flow velocity to flow rate (Fc) in mL/min:
Fc = ū x Pr2 x 60sec/min

18. System Modules Carrier Gas
Effect of mobile phase (carrier gas) density on column efficiency. Van Deemter plots for the 3 common carrier gases for a column of capacity factor k’ = The low density gases (H2 & He) have optimum efficiency at slightly higher flow rates than N2. The much lower slopes of H2 and He curves allow them to be used at higher flow rates (compared to N2) with very little loss of separation efficiency.

19. System Modules Inlets Inlets are the points of sample introduction
Ideal Sample Inlets for Column Type:
Packed Columns
Capillary Columns
Flash Vaporizer
Split
On-Column
Splitless

20. Cross section of a typical split injector
System Modules
Inlets
Split Injector
The oldest, simplest, and easiest injection technique.
Cross section of a typical split injector
Advantages to Split Injection:
High resolution separations
Neat samples can be introduced.
Dirty samples can be introduced by putting a deactivated glass wool plug in the liner to trap non-volatile components
Disadvantages:
Trace analysis is limited
Process sometimes discriminates between high molecular weight solutes so that the sample entering the column is not representative of the sample injected.

21. Cross section of a typical splitless injector
System Modules
Inlets
Splitless Injector
Samples have to be diluted in a volatile solvent and 1-5mL is injected in the heated injection port. Septum purge is essential in splitless injections.
Cross section of a typical splitless injector
Advantages to Splitless Injection:
Improved sensitivity over a split injector
Disadvantages:
Time consuming
Initial temperature and time of opening the split valve needs to be optimized.
Not well suited for volatile compounds (boiling points of peaks of interest have to be about 30oC higher than solvent.

22. System Modules Inlets Other Types of Inlets:
Direct Injection: involves injecting a small sample into a glass liner where vapors are carried directly into the column.
On-Column Injection: inserting the precisely aligned needle into the capillary column and making injections inside the column.
Flash Vaporization: involves heating the injection port to a temperature well above the boiling point to ensure rapid volatilization
Static Headspace: concentrates the vapors over a solid or liquid sample (best for residual solvent analysis)

23. System Modules Stationary Phase Sub-classification of GC Techniques
GSC: gas solid chromatography - stationary phase is solid
GLC: gas liquid chromatography -stationary phase is liquid

24. System Modules Stationary Phase Gas Solid Chromatography (GSC)
Solids used are traditionally run in packed columns
These solids should have small and uniform particle sizes (e.g. 80/100 mesh range)
Some Common GC Adsorbents
Commercial/Trade Names
Silica Gel
Chromasil®, Porasil®
Activated Alumina
Alumina F-1, Unibeads-A®
Zeolite Molecular Sieves
MS 5A, MS 13X
Carbon Molecular Sieves
Carbopack®, Carbotrap®, Carbograph®, Graphpac®
Porous Polymers
Porapak®, HayeSep®, Chromosorb®
Tenax Polymers
Texan TA®, Tenax GR®
Some of these solids have been coated on the inside walls of capillary columns and are called “Support Coated Open Tubular” or SCOT columns.

25. System Modules Stationary Phase
One major application of Packed Column GSC is in Gas Analysis.
Reasons:
Adsorbents provide high surface areas for maximum interaction with gases that may be difficult to retain on liquid stationary phase.
Large samples can be accommodated, providing lower absolute detection limits.
Some packed column GC’s can be configured to run below ambient temperature which will also increase the retention of the gaseous solutes.
Unique combinations of multiple columns and/or valving make it possible to optimize analysis of a particular sample.
Packed Columns also provide the flexibility of allowing mixed packings for special applications (e.g. 5% Fluorcol on Carbopack B® for analysis of Freons)

26. System Modules Stationary Phase Gas Liquid Chromatography (GLC)
To use liquid as stationary phase,techniques were applied to hold the liquid in a column.
For packed columns: liquid is coated onto a solid support, chosen for its high surface area and inertness. The coated support is then dry-packed into a column as tightly as possible.
For capillary or open tubular (OT) columns: liquid is coated on the inside of the capillary. To make it adhere better, the liquid phase is often extensively cross-linked and sometimes chemically bonded to the fused silica surface.
Schematic representation of (a) packed column and (b) capillary column

27. System Modules Stationary Phase Gas Liquid Chromatography (GLC)
Requirements for the stationary liquid phase:
Low vapor pressure
Thermal stability
(if possible) Low viscosity (for fast mass transfer)
Should interact with the components of the sample to be analyzed (“Like dissolves like”)

28. System Modules Stationary Phase Gas Liquid Chromatography (GLC)
Types of Capillary Columns (OT)
WCOT: Wall-coated open tubular column (provides the highest resolution of all OT’s – i.d.’s range from 0.1mm to 0.53mm and film thickness from 0.1 – 5.0m)
PLOT: Porous layer open tubular column (less than 5% of all GC use these days)
SCOT: Surface-coated open tubular column (no longer available in fused silica)

29. System Modules Detectors
The part of the system that ‘senses’ the effluents from the column and provides a record of the analysis in the form of a chromatogram. The signals are proportional to the quantity of each analyte.

30. System Modules Detectors FID: Flame Ionization Detector
The most common GC detector used.
The column effluent is burned in a small oxy-hydrogen flame producing some ions in the process. These ions are collected and form a small current that becomes the signals. When no sample is being burned, there should be little ionization, the small current is produced from impurities from the from the hydrogen and air supplies.
Hydrogen flow rate is commonly set to 40 – 45mL/min, Air, mL/min, and for OT columns (with flows of about 1 mL/min), Make-Up gases is added to carrier gas (to make up the flow to 30mL/min)

31. System Modules Detectors TCD: Thermal Conductivity Detector
This is a differential detector that measures the thermal conductivity of the analyte in the carrier gas compared to the thermal conductivity of the pure gas. At least two cavities are required. These cavities are drilled into a metal block and each contain a hot wire or filament. The filaments are incorporated into a Wheatstone Bridge Circuit (for resistance measurements).
The choice of carrier gas will depend on the thermal conductivity of the analyte (H2 and He have highest TC’s, N2 gives rise to unusual peak shapes)

32. System Modules Detectors NPD: Nitrogen Phosphorus Detector
A bead of Rb or Cs is electrically heated when flame ionization occurs. The detector shows enhanced detectability for nitrogen-, phosphorus-, or halogen- containing samples.

33. System Modules Detectors MSD: Mass Spectrometric Detector
Analyte molecules are first ionized in order to be attracted or repelled by the proper magnetic or electrical fields.

34. Troubleshooting Common GC Problems: Retention Time Problems
Resolution Problems
Baseline Problems
Peak Problems

35. Retention Time Problems
Retention Time Shift
Possible Cause
Solution
Comments
Change in carrier velocity
Check the carrier gas velocity
All peaks will shift in the same direction by approximately the same amount
Change in column temperature
Check the column temperature
Not all peaks will shift by the same amount
Change in column dimensions
Verify the column identity
Large change in compound concentration
Try a different sample concentration
May also affect adjacent peaks. Sample overloading is corrected with an increased split ratio, sample dilution, or decreased injection volume.
Leak in the injector or column connection
Leak-check the injector and column installation
Usually accompanied by peak size change.

36. Retention Time Problems
Retention Time Shift
Possible Cause
Solution
Comments
Blockage in a gas line
Clean or replace the plugged line
More common for the split line; also check flow-controllers and solenoids.
Septum leak
Replace the septum
Check for needle barb.
Sample solvent incompatibility
Change solvent. Use a retention gap.
For splitless injector.
Contamination
Trim the column.
Solvent-rinse the column.
Remove ½ - 1 meter from the front of the column.
Only for bonded and cross-linked phase.

37. Resolution Problems Loss of Resolution Decrease in separation
Possible Cause
Solution
Comments
Different column temperature
Check column temperature
Differences in other peaks will be visible
Different column dimensions or phase
Verify column identity
Co-elution with another peak
Change the column temperature
Decrease column temperature and check the appearance of a peak shoulder or tail.
Column contamination – resulting in a change in column selectivity
Trim the column
Solvent-rise the column
Remove ½ - 1 meter from the front of the column.
Only for bonded and cross-linked phase.

38. Resolution Problems Loss of Resolution Increase in peak width
Possible Cause
Solution
Comments
Change in carrier gas velocity
Check carrier gas velocity
A change in retention time also occurs
Column contamination
Trim the column
Solvent-rise the column
Remove ½ - 1 meter from the front of the column.
Only for bonded and cross-linked phase.
Inlet liner contamination
Clean or replace liner
Change in the injector
Check the injector settings
Typical areas: split ratio, liner, temperature, injection volume
Change in the sample concentration
Try a different sample concentration
Peak width increases at higher concentration
Improper solvent effect lack of focusing
Lower oven temperature.
Choose different solvent for better solvent/sample/phase polarity match.
Use a retention gap
For splitless injection.

39. Baseline Problems Excessive Column Bleed Possible Cause Solution
Comments
Thermal damage to the column
Remove column from detector and bake-out overnight, reinstall and condition as usual
Use GC maximum temperature function
Oxygen damage to column
Columns damaged by oxygen will usually need to be replaced although an overnight bake-out may be attempted
Perform periodic leak checks. Change septa regularly. Use high quality carrier gases. Install and maintain oxygen traps
Chemical phase damage to column
Remove ½ to 5 meters from the front of the column
Perform sample prep to remove inorganic acids and bases from the sample. Install guard column and trim frequently. If acids or bases must be used, choose HCl or NH4OH, or an organic alternative.

40. Baseline Problems Erratic Baseline (drift, wander) Possible Cause
Solution
Comments
Inlet contamination
Clean the injector
Try a condensation test; gas lines may also need cleaning. Take steps to prevent sample backflash (reduce injection volume, lower inlet temperature, use larger volume liner
Column contamination
Bake-out column.
Solvent-rinse the column
Limit bake-out to 1 – 2 hours
Only for bonded and cross-linked phases.
Check for inlet contamination
Incompletely-conditioned column
Fully condition the column
More critical for trace analysis
Un-equilibrated detector
Allow the detector to stabilize
Some detectors may require up to 24 hours to fully stabilize
Change in carrier gas flow-rate during the temperature program
Normal in many cases
MS, TCD, and ECD respond to carrier gas flow rate changes

41. Baseline Problems Erratic Baseline (drift, wander) Possible Cause
Solution
Comments
Contaminated gases
Use appropriate purifier to remove contaminants
More of a problem for detector gases.
Column and inlet liner misaligned
Check installation of column end and inlet liner, adjust if necessary
Causes a baseline change after a large peak
Large leak at the septum during injection and for a short time thereafter
Replace septum
Use smaller diameter needle
Causes a baseline change after a large peak.
Common with large diameter needles.
Sample decomposing
Remove inlet liner and check cleanliness.
Use new, deactivated liner or replace glass wool and packing.
Causes a baseline rise before and after a peak.

42. Baseline Problems ·Noisy Baseline Possible Cause Solution Comments
Inlet contamination
Clean the injector, replace liner, gold seal
Try a condensation test; gas lines may also need cleaning.
Column contamination
Bake-out the column.
Solvent-rinse the column.
Limit bake-out to 1 – 2 hours
Only for bonded and cross-linked phases.
Check for inlet contamination
Detector contamination
Clean the detector.
Usually the noise increases over time and not suddenly.
Contaminated or low quality gases
Replace spent gas purifier.
Use purifiers to remove contaminants.
Use better grade gases.
More of a problem for detector gases.
Column inserted too far into the detector
Reinstall the column.
Consult the GC manual for the proper installation distance.

43. Baseline Problems ·Noisy Baseline Possible cause Solution Comments
Incorrect detector gas flow rates
Adjust the flow rates to the recommended values.
Consult the GC manual for appropriate flow rates
Leak when using an MS, ECD, or TCD
Find and eliminate the leak.
Usually at the column fittings or injector.
Old detector filament, lamp, or electron multiplier, NPD head
Replace appropriate part.
Septum degradation
Replace septum.
For high temperature applications, use appropriate septum

44. Baseline Problems ·Ghost Peaks Possible Cause Solution Comments
Contaminants introduced with sample
Sample or solvent clean-up
Contaminants in sample process or solvent.
Inlet contamination
Clean the injector, replace liner, gold seal, and septum
Try a condensation test; gas lines may also need cleaning. Take steps to prevent sample backflash (reduce injection volume, lower inlet temperature, use larger volume liner)
Septum bleed
Replace septum.
Use a high quality septum appropriate for the inlet temperature.
Contamination of sample prior to introduction to the GC
Check sample handling steps for potential contamination sources: sample clean-up, handling, transfer, and storage
Semi-volatile contamination (peak widths will be broader than sample peaks with similar retention
Bake out column,
Solvent-rinse the column.
Check for contamination in the inlet, carrier gas, or carrier gas lines.
Limit bake-out to 1 – 2 hours.
Only for bonded and cross-linked phases.

45. Peak Problems ·Fronting Peaks Possible cause Solution Comments
Column overload
Reduce mass amount of the analyze to the column. Decrease injection volume, dilute sample, increase split ratio
Most common cause for fronting peaks.
Improper column installation.
Reinstall the column in the injector.
Consult the GC manual for the proper installation distance
Injection technique.
Change technique.
Usually related to erratic plunger depression or having sample in the syringe needle.
Use an autosampler.
Compound very soluble in injection solvent
Change solvent. Using a retention gap may help.
Mixed sample solvent
Change sample solvent.
Worse for solvents with large differences in polarity or boiling points.

46. Peak Problems ·Tailing Peaks Possible cause Solution Comments
Severe column contamination
Trim the column.
Solvent rinse the column
Remove ½ - 1 meter from the front end of the column.
Only for bonded or cross-linked phase.
Check for inlet contamination. Tailing will sometimes increase with compound retention.
Active column
Cut off 1-meter from the front end of the column.
Replace column.
Only affects active compounds. Usually produces tailing that increases with retention.
Improper column installation, leak, or column end poorly cut
Re-cut and reinstall the column into the inlet.
Replace ferrule.
Confirm installation is leak-free
Make a clean square cut with a reliable cutting tool.
Consult GC manual for the proper installation distance.
More tailing for early eluting peaks.
Contaminated or active liner or gold seal
Use new, deactivated liner.
Clean or replace gold seal.
Only affects active compounds.

47. Peak Problems ·Tailing Peaks Possible cause Solution Comments
Solid particles in liner
Clean or replace liner.
Needle hitting and breaking packing in inlet liner
Partially remove packing from liner or use without packing.
Solvent/column not compatible
Use a different solvent.
Use a retention gap.
More tailing for the early eluting peaks or those closest to the solvent front.
3 – 5 meter retention gap is sufficient.
Split ratio too low
Increase split ratio.
Flow from split vent should be  20mL/min
Solvent effect violations for splitless or on-column injections
Decrease the initial column temperature to °C below solvent boiling point
Peak tailing decreases with retention.
Poor injection technique
Change technique
Usually related to erratic plunger depression or having sample in the syringe needle.
Use an autosampler.

48. Peak Problems ·Tailing Peaks Possible cause Solution Comments
Inlet temperature too high
Decrease inlet temperature by 50°C
Tailing generally worse for early eluters.
Inlet temperature too low
Increase inlet temperature by 50°C
Tailing usually increases with retention
Dead volume in system
Reduce dead volume. Transfer line connections, fused silica unions, etc.
Peak tailing decreases with retention.
Cold spots (condensation)
Eliminate cold spots. Commonly at transfer lines.
Overloading of PLOT columns
Reduce the amount injected onto column.

49. Peak Problems ·Split Peaks Possible cause Solution Comments
Column installation
Reinstall the column in the injector
Consult the GC manual for the proper installation distance
Injection technique
Change technique.
Usually related to erratic plunger depression or having sample in the syringe needle.
Use an autosampler.
Mixed sample solvent
Change sample solvent.
Worse for solvents with large differences in polarity or boiling points
Poor sample focusing
Use a retention gap
For splitless, on-column, and PTV injectors
Solvent/column not compatible
Use a different solvent.
Use a retention gap.

50. Peak Problems ·Split Peaks Possible cause Solution Comments
Sample degradation in injector (only some peaks show splitting)
Reduce inlet temperature.
Derivatize sample to make compounds thermally stable.
Change to an on-column injector.
Peak broadening or tailing may occur if the temperature is too low.
Requires an on-column injector
Severe detector overload
Reduce the amount of sample on-column.
May only affect some peaks.

51. Peak Problems ·Changes in Peak Size Possible cause Solution Comments
Change in detector response
Check gas flow, temperature and settings.
Check background level or noise
All peaks may not be equally affected
May be caused by the system contamination, not the detector.
Change in the split ratio
Check the split ratio
All peaks may not be equally affected.
Change in the purge activation time
Check the purge activation time
For splitless injectors
Change in injection volume
Check injection technique
Injection volumes are not linear.
Change in injector discrimination
Maintain the same injector parameters: flows, temperatures, liners, etc.
Most severe for spit injections. All peaks may not be equally affected

52. Peak Problems ·Changes in Peak Size Possible cause Solution Comments
Change in sample concentration
Check and verify sample concentration
May be caused by degradation, evaporation, or variances in sample temperature or pH
Leak in the syringe
Use a different syringe
Sample leas past the plunger or around the needle; leaks are often not readily visible
Column contamination
Trim the column
Solvent-rinse the column
Remove ½ - 1 meter from the front of the column.
Only for bonded and cross-linked phases
Column activity
Trim or replace the column
Only affects active compounds

53. Peak Problems ·Changes in Peak Size Possible Cause Solution Comments
Co-elution
Change column temperature or stationary phase
Decrease column temperature, and check for the appearance of peak shoulder or tail
Sample backflash
Inject less, use larger liner, or reduce the inlet temperature
Less solvent and higher flow rates are most helpful
Decomposition from inlet contamination
Clean the inlet, replace the liner, replace the gold seal
Only use deactivated liners and glass wool in the inlet.
Loss of sample prior to introduction into the GC
Check sample handling, sample preparation, transfer and storage

54. T H E E N D