Chapter 31

Dong-Sun Lee/ CAT-Lab / SWU 2012-Fall version

Gas Chromatography (GC)

Introduction

Gas chromatography is a chromatographic technique that can be used to separate volatile organic compounds.

Two types of GC are encountered: gas-solid chromatography(GSC) and gas-liquid chromatography(GLC). GLC is finds widespread use in all fields of science, where its name is usually shortened to GC.

A gas chromatograph consists of a flowing mobile phase, an injection port, a separation column containing the stationary phase, and a detector.

GSC is based on a solid stationary phase on which retention of analytes is the consequence of physical adsorption. GLC is based on partitioning behavior of the analyte between the mobile gas phase and the liquid stationary phase in the column.

Characteristics of GLC

1. Sensitivity : mg ~ pg (10–3 ~ 10–9 g)

2. Versatility : from rare gases to liquids and solids in solution with 800~1000 MW

3. Speed of analysis : typically 5 ~ 30 min , complex 3 ~ 30 mixture separation

4. Reproducibility : qualitative Accuracy : quantitative

GC-14A gas chromatograph (Shimadzu)with an integrator.

HP 5890 gas chromatograph (Hewlett Packard) with an integrator

GC-MS : GC-Q plus ion-trap GC-MSn ( Thermoquest - Finnigan ) Xcalibur software

Fuel gas

Carrier gas

Regulator

Trap Make-up gas

Split vent

Injector Detector

Column oven

Column

Fuel gas

Electrometer

Recorder Integrator Computer

Valve/ Gage

Schematic diagram of a gas chromatograph system.

Basic components of Gas Chromatograph

Carrier gas supply Sample introduction inlet Column and controlled-temperature oven Detector & oven Recorder

Carrier gas system in gas chromatograph

The purpose of the carrier is to transport the sample through the column to the detector.

The selection of the proper carrier gas is very important because it affects both column and detector performance. The detector that is employed usually dictates the carrier to be used.

From a column performance point of view a gas having a small diffusion coefficient is desirable (high molecular weight, e.g., N2, CO2, Ar) for low carrier velocities while large diffusion coefficients (low molecular weight, e.g., H2, He) are best at high carrier velocities.

The viscosity dictates the driving pressure. For high-speed analysis, the ratio of viscosity to diffusion coefficient should be as small as possible. H2 would be the best choice, followed by helium.

The purity of the carrier should be at least 99.995% for best results. Impurities such as air or water can cause sample decomposition and column and detector deterioration. In temperature programmed runs, impurities in the carrier gas such as water can be retained at low temperatures but are then eluted at higher temperatures impairing the baseline. Many instrument problems have been traced to contaminated carrier gases.

The carrier must also be inert to the components of the sample and the column.

Properties of common carrier gases

Gas molecular weight Thermal conductivity Viscosity

× 105 at 100oC × 10–6 at 100oC

(g-cal/sec-cm- oC ) (P)

Ar 39.95 5.087 270.2 *

CO2 44.01 5.06 197.2

He 4.00 39.85 234.1

H2 2.016 49.94 104.6**

N2 28.01 7.18 212.0

O2 32.00 7.427 248.5***

* at 99.6oC ** at 100.5oC **** at 99.74oC

Using the correct carrier and detector gases are an important factor in installing a new GC. The five gases commonly used as carrier gas and detector fuels in capillary gas chromatography are helium, hydrogen, nitrogen, argon-methane, and air. The types of gases necessary are partly determined by the detection system used. Factors to consider for each individual gas are discussed below.

Carrier Gas Choice

Carrier gases that exhibit a broad minimum on a van Deemter profile are essential in obtaining optimum performance. Volumetric flow through a capillary column is affected by temperature. When temperature programming from ambient to 300oC, the flow rate can decrease by 40 percent. A carrier gas that retains high efficiency over a wide range of flow rates and temperatures is essential in obtaining good resolution throughout a temperature programmed run. Figure 1 shows the van Deemter profile for hydrogen, helium, and nitrogen carrier gases.

Van Deemter curves for GC of n-C17H36 at 175oC, using N2, He, or H2 in a 0.25 mm diameter × 25 m long wall coated column with OV-101 stationary phase

Hydrogen is the fastest carrier gas (uopt), with an optimum linear velocity of

40cm/sec, and exhibits the flattest van Deemter profile. Helium is the next best choice, with an optimum linear velocity of uopt = 20cm/sec. Nitrogen's performance is inferior with capillary columns because of its slow linear velocity, uopt = 12cm/sec. Argon-methane has a slower optimum linear velocity than nitrogen and is not recommended for use as a carrier gas with capillary columns. Air is not recommended as a carrier gas because it can cause stationary phase oxidation.

      With hydrogen and helium as carrier gases, the minimum H.E.T.P. values can be maintained over a broader range of linear velocities than with nitrogen, and high linear velocities can be used without sacrificing efficiency. Nitrogen is beneficial only when analyzing highly volatile gases under narrow temperature ranges where increasing stationary phase interaction is desirable. Otherwise, the use of N2 results in longer analysis times and a loss

of resolution for compounds analyzed on a wide temperature range.

http://www.restekcorp.com/gcsetup/gcsetup3.htm

Exert Caution when Using Hydrogen as a Carrier Gas

Hydrogen is explosive when concentrations exceed 4% in air. Proper safety precautions should be utilized to prevent an explosion within the column oven. Most gas chromatographs are designed with spring loaded doors, perforated or corrugated metal column ovens, and back pressure/flow controlled pneumatics to minimize the hazards when using hydrogen carrier gas. Additional precautions include:

• Frequently checking for leaks using an electronic leak detector.

• Using electronic sensors that shut down the carrier gas flow in the event of pressure loss.

• Minimizing the amount of carrier gas that could be expelled in the column oven if a leak were to occur by installing a flow controller (needle valve) prior to the carrier inlet bulkhead fitting to throttle the flow of gas (for head pressure controlled systems only) as shown Fig. 2.

• Fully open the flow controller (needle valve) and obtain the proper column head pressure, split vent flow, and septum purge flow rates. Decrease the needle valve flow rate until the head pressure gauge begins to drop (throttle point). Next, increase the flow controller (needle valve) setting so that the right amount of flow is available to the system. Should a leak occur, the flow controller will throttle the flow, preventing a large amount of hydrogen from entering the oven.

Make-up and Detector Fuel Gases

Gas added to the stream after the column is called makeup gas. Choosing the correct make-up and detector gases will depend on both the detector and application. Most GC detectors operate best with a total gas flow of approximately 30ml/min. to ensure high sensitivity and excellent peak symmetry. Refer to your GC manual for optimum flow rates on different instruments. Carrier gas flows for capillary columns range from 0.5 to 10ml/min. which are well below the range where most detectors exhibit optimal performance. To minimize detector dead volume, make-up gas is often added at the exit end of the column to increase the total flow entering the detector. Make-up gas helps to efficiently sweep detector dead volume thereby enhancing detector sensitivity.

Make-up gas can be added directly to the hydrogen flame gas for flame ionization detectors (FID), nitrogen phosphorous detectors (NPD), and flame photometric detectors (FPD) or added to the column effluent by an adaptor fitting. However, GCs such as Perkin-Elmer and Fisons do not require make-up gas.      

Combustion type detectors (FID, NPD, FPD) use three gases: make-up, hydrogen (fuel gas), and air (combustion/oxidizing gas). For non-combustion detectors, such as the thermal conductivity detector (TCD), electron capture (ECD), and photo ionization detector (PID), only carrier and make-up gases are required. In the case of the electrolytic conductivity detector (ELCD), the make-up gas is hydrogen, as a reaction gas in the halogen and nitrogen mode or air in the sulfur mode. Table I shows recommended gases for various detectors.

Carrier gases and detector fuel gases for use with various GC detectors

TCD ECD FID NPD FPD ELCD PID

Carrier Gases He O O O O O O O

H2 O - O - O O O

N2 O O O O O - O

Combustion/Reaction Gases    

H2 - - O O O O -

Air - - O O O - -

Make-up Gases   N2 O O O O O - O

He O - O O O - O

ArCH2 - O - - - - -

http://www.restekcorp.com/gcsetup/gcsetup3.htm

Effect of impurities

- Impurities such as hydrocarbon, oxygen, water contribute to unwanted noise levels, excessive baseline drift.

- Molecular sieve --- Moisture trap, Oxygen trap, Chemical filter

Effect of water on column efficiency

- Carrier gas dryness is very important !! (use anhydrous sodium sulfate)

- Water can and usually does react with some portion of the column. This results in loss of resolution and tends to produce asymmetric or tailing peaks.

Unwanted components or ghost peaks may also appear.

Another effect is a net loss of sensitivity.

Gas purifiers

The trap will remove any water vapor or oils that may have been introduced in the filling process since a number of gases are water pumped. The contaminants removed by the trap could otherwise interact with the column packing material to produce spurious peaks. In addition the contaminants can cause increased detector noise and drift.

The traps should be reconditioned (about twice a year ) by heating to 300 oC for 4~8 hr with a stream of gas passing through it or in a vacuum oven.

Carrier gas purifiers

GC1 2 3 4

Gas cylinder

1. Hydrocarbon trap

2. Moisture trap

3. Oxygen trap

4. Indicating oxygen trap

Gas purifier recommendation for GC applications

Capillary column GC Carrier Hydrocarbon, Moisture, Oxygen

with any detector Make-up None - all detector but

ECD moisture & oxygen

Air for FID Hydrocarbon

H2 for FID None

ELCD reaction gas Hydrocarbon

Packed column GC Carrier Hydrocarbon, Moisture, Oxygen

with FID or TCD

Packed column GC Carrier Hydrocarbon, Moisture, Oxygen

with ECD, FPD, NPD,

MSD

Flow requirements

1. Stable 2. Reproducible 3. Convenient

The more constant the flow rates, the more precise and accurate the results.

Flow controller ( Pressure controller ) --- to maintain precise and accurate flow rates

Effect of decreased flow rate or lower temperature

- All peaks have shifted to longer retention times- Apparent loss of peak height- The base of each peak is wider, however, individual peak area remain constant.

Effect of increased flow rate

- Sample components are squeezed toward the injection point- Cause two components to elute together, appearing as single peak

Regulations of carrier gas

Carrier cylinder bottled at about 2500 psi(150-160 atm)

Two stage pressure regulator :

- first stage : high inlet pressure

- second stage : low outlet pressure

( set at 40~100psi)

Gas generators

Gas flow rate control

A 1 % change in carrier gas flow rate will cause a 1% change in retention time. For all these reasons it is important to keep the flow of the carrier gas constant.

1. Control of carrier gas inlet pressure

2. Control of carrier gas flow rate

In isothermal operation the means of regulation is immaterial because both means provide constant inlet pressure as well as constant flow rate. In temperature programmed runs, however, the situation is quite different. If one maintains the inlet pressure constant the flow rate will change. Therefore, with temperature programming of the column, the flow rate must be controlled.

Pressure controllers

1. The second stage regulator on the cylinder

2. A pressure regulator mounted in the GC

3. A needle valve(variable restrictor) mounted in the GC

4. A fixed restrictor mounted in the GC

Flow measurement

1) Rotometer

The column flow rate is typically indicated by a rotometer . ( Calibrate equilibrium position indicating the flow )

Rotometer is operated by the volume of gas passing a ball in a tapered cell.

2) Bubble meter

3) Electronic flow sensor

Relationships between inside diameter, column length, mesh size, and carrier gas flow for packed column

Inside diameter Mesh size for Mesh size for Carrier flow

mm length up to 3m length over 3m N2, ml/min He or H2, ml/min

2 100~120 80~100 8~15 15~30

3 100~120 80~100 15~30 30~60

4 80~100 60~80 30~60 60~100

John A. Dean, Analytical Chemistry Handbook, McGraw-Hill, 1995, p. 4.31 .

Comparison of 1/8-in(0.316 cm) packed, wide bore, and WCOT columns

1/8 in packed Wide bore WCOT

Inside diameter, mm 2.2 0.53 0.25

Film thickness, m 5 1~5 0.25

Phase volume ratio() 15~30 130~250 250

Column length, m 1~2 15~30 15~60

Flow rate, ml/min 20 5 1

Effective plates(Neff) per meter 2000 1200 3000

Effective plate height (Heff),mm 0.5 0.6 0.3

Typical sample size 15 g 50 ng

John A. Dean, Analytical Chemistry Handbook, McGraw-Hill, 1995, p. 4.31 .

Recommended ranges of gas flow rates

Detector Gases Range Capillary Packed

FID Carrier 2 ml/min 40 ml/min

Hydrogen 30~50 ml/min 35 ml/min 40 ml/min

Air 300~600 ml/min 350 ml/min 500 ml/min

Make-up(N2) 10~60 ml/min 30 ml/min not used

NPD Hydrogen 2~4 ml/min

Air 40~80 ml/min

Make-up(N2, He) 10~20 ml/min not necessary

PID Make-up 5~10 ml/min

Sheath 30~40 ml/min

FPD Carrier 1~3 ml/min 30~50 ml/min

Hydrogen 85~100 ml/min 100~120 ml/min

Air 100~120 ml/min 110~135 ml/min

Inlet requirements

1. Temperature controlled 2. Low volume (total swept by carrier ) 3. Inert construction

Column Overload

If too large an sample were allowed to enter small bore(capillary) columns column overload and a loss of resolving power would like occur.

Liner

All liners help protect the vaporized sample form contacting the metal wall of the inlet as sample flows onto the column. Deactivated glass wool may be used as an aid for sample vaporization, to minimize discrimination based on boiling point, and to provide a surface on which non0volatiles can be trapped. The simplest liner is a straight tube, which gives all-around good performance at low cost. Single-taper liners improve on a straight tube by minimizing sample vapor contact with metal at the bottom of the injection port, although they are somewhat more expensive. Liners are deactivated borosilicate glass, except quartz where noted. Liners are guaranteed inert for phenols, organic acids and bases.

Why is Glass Wool Added to an Injector Liner? - General GC

The Glass wool serves three major purposes. The Glass wool will prevent the small pieces of septa from reaching the column. The presence of Glass wool will help the injected sample stay in the liner a little longer which will help the sample to vaporize and mix more thoroughly with the carrier gas. If positioned properly it will wipe the outer surface of the syringe needle and improve the precision of the liquid injection.

Inlet configuration

1. Direct column inlet --- 1/8 " OD or larger column

sampling syringe is actually inserted into the end of the column needle guide / cap / spring / septum mounting holes / carrier gas in / inlet body column Swagelock ferrules / Swagelock nut

2. Splitter inlet --- open tubular column or less than 1/8" OD column

Because of the limited capacity for sample of these small bore columns and the difficulty of injecting extremely small volume samples, a large portion of the injected sample is vented to atmosphere by the inlet.

septum / preheated carrier gas / mixing tube / restrictor buffer volume / tapered needle / gold gasket / column fitting

Injection port for split injection into an open tubular column. The glass liner is slowly contaminated by nonvolatile and decomposed samples and must be replaced periodically. For splitless injection, the glass liner is a atraight tube with no mixing chamber. For dirty samples, split injection is used and a packing material can be replaced inside the liner to adsorb undesirable components of the sample.

Common injection techniques

1) Hot flash vaporization

Direct

Cold-trap

Split

Splitless

2) Direct cold : on column

Split or on-column

Split 1) Simple 2) High column efficiency 3) Column may be protected

On-column 1) Best accuracy 2) Thermolabile compounds 3) Trace analysis

Representative injection conditions for split, splitless, and on-column injection into an open tubular column.

Direct injector 1) Good sensitivity 2) Low column efficiency 3) Best for thick films, widebore column ( 0.53 mm )

Hot on-column injectors 1) Reduced column efficiency 2) Best with thick films, widebore columns 3) Nonvolatiles may damage column 4) Cold on-column injector may be used with 0.1 to 0.53 mm i.d. columns

Advantage of on-column 1) Best reproducibility : Quantitative results 2) No split, no loss of high boilers 3) "Cold" on-column injection available

Advantage of splitless

1) High sensitivity ( 95 % of sample on column ) 2) Solvent effect produces narrow sample bands 3) Same hardware as split injection

Disadvantage of splitless 1) Slow sample transfer to column 2) Must dilute sample with volatile solvent 3) Time consuming : must cool column 4) Poor for thermolabile compounds

Split and splitless injections of a solution containing 1 vol % methyl isobutyl ketone (bp 118 oC) and 1 vol % p-xylene (bp 138 oC) in dichloromethane (bp 40 oC) on a BP-10 moderately polar cyanopropyl phenyl ,ethyl silicone open tubular column(0.22 mm I.d., 10 m long, 0.25 m, column temperature =75 oC).

Common injection methods

Syringe injection

Valve injection

Sampling Syringe

0 ~ 1 μL --- the sample is totally confined to the needle 0 ~ 5 / 0 ~ 10 μL needle / barrel / plunger

Gas sampling valve

Sample in Sample loop --- 1/4 or 10 mL loop size compatible with needs Sample vent Carrier gas to column

Solvent effect

t-1 t-2

time t-1 --- just after injection, solvent and sample are condensed in a long plug at the front of the column. The column temperature must be cold enough to condense the solvent. time t-2 --- after some time, the column temperature has been raised, most of the solvent has evaporated, and the solvent effect has left the sample molecules concentrated in a narrow band. As the column is further heated, the remaining solvent and sample molecules are rapidly vaporized --- resulting in high column efficiency and narrow peak.

Syringe for solid phase microextraction.

Sampling by SPME and desorption of analyte from the coated fiber into a gas chromatograph.

Purge and trap apparatus for extracting volatile substances from a liquid or solid by flowing gas.

GC column

Parts of Column

1) Tubing material

Stainless steel--- reactive ( steroids, amines, free acids ) Glass ------------ can be made inert, difficult handling Fused silica ---- flexible most inert most popular high resolution2) Stationary phase

Solid support --- carefully sized granular

Liquid phase --- active portion of the column

Porous polymers

Adsorbents

Important column parameters 1) Inside diameter

2) Length

3) Film thickness

4) Stationary phase composition

5) Flow rate

Column diameter

i.d. Resolution Speed Capacity Ease

100 micrometer +++ +++ + + (narrow bore ) 250, 320 ++ ++ ++ ++ (mid bore) 530 + ++ +++ +++ (wide bore)

Column length

N œ L R œ L1/2 tR œ L

24-foot 1/8" packed column  wound on 6" coil 6-foot 1/4"

packed column  wound on 5" coil

60-meter 0.53mm metal

wide bore  capillary column wound on

3.5" coil 

15-meter 0.53mm fused silica wide bore capillary column

wound on 7" cage

30-meter .25mm metal narrow bore capillary column wound

on 3.5" coil http://www.srigc.com/catalog/columns.htm

ColumnGlass wool --- both ends of the column 1- ½” (inlet side) 1/4” (detector side)

Fused silica surfacemade by the reaction of SiCl4 and water vapor in a flame - SiO2 contains 0.1 % –OH groups - Very inert - Uniform chemical surface

Fused silica - High tensile strength - Flexible - Sheath of polyimide - Very inert

Fused Silica Capillary ColumnsA fused silica capillary column is comprised of three major parts (Figure 1). Polyimide is used to coat the exterior of fused silica tubing. The polyimide protects the fused silica tubing from breakage and imparts the amber-brown color of columns. The stationary phase is a polymer that is evenly coated onto the inner wall of the tubing. The predominant stationary phases are silicon based polymers (polysiloxanes), polyethylene glycols (PEG, Carbowax) and solid adsorbents.

Figure 1.

Capillary columns have to be properly installed to maximize their performance and lifetime. You can obtain enhanced column performance and lifetime by following these recommended installation guidelines. More detailed installation, operational and troubleshooting information can be found in the following references

WCOT = Wall Coated Open Tubular

invented and patented by Dr Marcel Golay

Tubing - Fused silica - Glass - Stainless steel Liquid phase coating

WCOT - - - High resolution

Film thickness : 0.5 to 5.0 micrometer i.d. : 0.10, 0.25, 0.32, 0.53 mm Length : 10 to 60 m

Open tubular GC column

Operational guideline for open tubular GC columns

WCOT

narrow intermediate wide bore

Column inner diameter, mm 0.25 0.32 0.53

Maximum sample volume, l 0.5 1 1

Maximum amount for

one component, ng 2~50 3~75 5~100

Effective plates(Neff) per meter 3000~5000 2500~4000 1500~2500

Trennzahl(separation) number

per 25 m 40 35 25

Optimum flow for N2, ml/min * 0.5~1 0.8~1.5 2~4

Optimum flow for He, ml/min ** 1~2 1~2.5 5~10

Optimum flow for H2, ml/min *** 2~4 3~7 8~15

* Optimum velocity is 10 to 15 cm/s for each column

** Optimum velocity is 25 cm/s for each column

*** Optimum velocity is 35 cm/s for each column

Other types of capillary columns SCOT = Support Coated Open Tubular Solid support : Celite Liquid phase Not available fused silica tubing

PLOT = Porous Layer Open Tubular Porous adsorbent : alumina or molecular sieve

* Molecular sieve --- efficient for H2, Ne, Ar, O2, N2, CO, CH4.

Porous carbon stationary phase ( 2 m thick) on inside wall of fused silica open tubular column.

Capillary column vs Packed columnCapillary Packed

Length 60 m 2 m

Theoretical plates(N/m) 3000-5000 2000

Total plates

length×(N/m)

180000-300000 4000

Advantages of capillary column

1) Not packed : long lengths : high resolution2) Thin film ; efficient, fast3) Used silica --- inert surface, better results

Disadvantages of capillary column

1) More expensive2) Limited liquid phase Requires very small samples3) Dedicated instrumentation --- capillary inlets, septum purge

(Left) Gas chromatogram of alcohol mixture at 40 oC using packed column ( 2mm I.D., 76 cm long containing 20 % Carbowax 20 M on a Gas-Chrom R support and FID.

(Right) Chromatogram of vapors from headspace of beer can, obtained with 0.25 mm diameter, 30 m long porous carbon column oerated at 30 oC for 2 min and then ramped up to 160 oC at 20 oC/min.

Component Separation with the Column < The process of separation >

A series of partitions : Dynamic In-and Out (or Stop-and-Go) All differential migration process. The most volatile components usually pass through the column first, the least volatile or highest boiling emerges last.

Mobile phase( Driving force)

Stationary phase ( Resistive force)

Analytes

Capillary Column Installation Steps

1. Check traps, carrier gas, septum, liner2. Place the nut and ferrule on the column and carefully cut the column end3. Install the column into the injector4. Turn on the carrier gas5. Install the column into the detector6. Inspect for leaks7. Confirm carrier gas flow and proper column installation8. Condition the column9. Accurately set the carrier gas velocity10. Bleed test 11. Run test mix

Click here for a complete listing of tools available from J&W, including magnifiers and cutting tools.http://www.jandw.com/gccolumn.htm#Fused Silica Capillary Columns

Recommended Installation Tools and Supplies

1. Cutting tool such as a diamond or carbide tipped pencil, sapphire tipped pencil or ceramic cleaving wedge2. Magnifier (10-20X)3. Ruler4. Wrench5. Ferrules6. Vial of solvent7. Clean syringe8. Supply of an appropriate non-retained compound9. Column test mixture10. Flow meter11. Other supplies: septa, clean injector liners, liner ferrules/O-rings, etc.

Conditioning of the ColumnOnce the column has been checked for proper installation and the absence of leaks, it is ready for conditioning. Heat the column to its isothermal, upper temperature limit (temperature limits listed below) or a temperature 10-20 oC above the highest operating temperature of your particular method. Do not exceed the upper limit or column damage will result. Heat the column rapidly - slow temperature programming is not necessary. After the column has reached the conditioning temperature, plot the baseline. Keep the baseline on scale so that it can be observed. The baseline should be elevated at first then start to drop after 5-10 minutes at the conditioning temperature. The baseline will continue to drop for 30-90 minutes then stabilize at a constant value. If the baseline does not stabilize after 2-3 hours or does not start to significantly decrease after 15-20 minutes, either a leak is present or a contamination problem exists. In either case, immediately cool the oven down below 40 oC and resolve the problem. Continued conditioning will result in column damage or the inability to obtain a stable baseline. Excessive conditioning of the column may result in a shortened lifetime.

In general, polar stationary phases and thick film columns usually require longer times to stabilize than less polar and thinner film columns. GS PLOT columns require a different conditioning procedure than liquid stationary phase columns.

Conditioning Procedure for GS-Q PLOT Columns

GS-Molesieve, GS-Alumina  and GS-Q  columns require special conditioning procedures. The following conditions are recommended.

 Column: Temperature:

 GS-Q:  250 oC for a minimum of 8 hours

 GS-Molesieve: 300 oC for 3 hours or 250 oC for 12 hours

 GS-Alumina:  200 oC for a minimum of 8 hours

GS-Alumina  and GS-Molesieve are susceptible to retention shifts from reversible absorption of water. If retention shifts are observed after analyzing high water content samples, re-condition the column to remove any water trapped by the stationary phase.

If the column is conditioned with the detector end disconnected, a small portion of the column’s end may be damaged. Remove 10-20 cm of the exposed column end before installing the column into the detector.

Temperature Limits

The temperature limits define the range over which the column can be safely used. If the oven is operated below the lower temperature limit, poor separation and peak shape problems will be evident, but no column damage will occur.

Upper temperature limits are usually given as two numbers. The first or lower temperature of the two is the isothermal limit. The column can be maintained at this temperature for indefinite periods of time. The second or higher temperature is the program limit. The column can be heated to this temperature for short periods of time (<10 minutes). Exceeding the upper temperature limits will significantly reduce column lifetime.

Liquid Phase(=Stationary phase) Classes

1. Non-polar phase gives boiling point order separation

2. Selective phase separates components that have close b.p. and small structural differences

3. Polar phase depends on internal functional groups to separate compounds that have reactive –OH, –NH2 or other polar radicals

4. Each stationary phase retains solutes in its own class best

Raising percentage of stationary phase leads to

1) Greater capacity for solute2) Longer retention time3) Increased HETP

Column Temperature

Dimethylpolysiloxane

DB-1; 0.1, 0.25 and 1.0 m -60 to 325/350 oC

DB-1; 3.0 and 5.0 m -60 to 280/300 oC

DB-1; Megabore  0.1 m -60 to 360 oC

DB-1; Megabore  1.5 m -60 to 300/320 oC

DB-1; Megabore  3.0 and 5.0 m -60 to 260/280 oC

DB-1ht -60 to 400 oC

(5%-Phenyl) Methylpolysiloxane

DB-5 -60 to 325/350 oC

DB-5; Megabore  -60 to 300/320 oC

DB-5; Megabore  5.0 m -60 to 260/280 oC

DB-5ht -60 to 400 oC

DB-5ms -60 to 325/350 oC

DB-5ms; Megabore  -60 to 300/320 oC

DB-5.625 -60 to 325/350 oC

Polyethylene Glycol

DB-WAX 20 to 250/260 oC

DB-WAX; Megabore 20 to 230/240 oC

Base Deactivated Polyethylene Glycol

CAM 60 to 220/240 oC

Carbowax 20M

Carbowax 60 to 220/240 oC

-Cyclodextrin for chiral column.

Column oven and temperature control

Oven size : column sits in an oven Inner volume : laboratory GC ; about 22-25 L process GC ; about 40 L

Isothermal The oven temperature is kept constant during the entire analysis Practical temperature < 250oC Maximum temperature < 400oC

Temperature programming The oven temperature is varied during the analysis Linear Non linear

High temperature GC (HTGC) Working limit : conventional GC --- 330oC HTGC --- 450oC Masses of analysed substrates: 600-1000

Temperature programming

With homologues, the retention time increases exponentially with the number of carbon.

As retention time increases, width increase and the height decreases, making detection impossible after a few peaks have eluted.

Since solubility of gas in a liquid decreases as temperatures goes up, we can reduce the retention of a compound by increasing column temperature.

General steps to create a program assuming that the separation is possible

1) Determine initial temperature and time based on best possible separation offirst few peaks

2) Report for the last few peaks to find the best final temperature and time

3) Experiment with various ramps to account for the rest of the components

Temperature programming

Factors to consider :

Variations in solubility of solutes

Changes in volatility of solutes

Stability of solutes

Flow rate changes

Stability of stationary phase

Must stay within Tmin/Tmax of column

Other factors are found experimentally

A temperature program

Ex.

40 oC(5 min) – 10oC/min – 250oC(10 min)

A: initial temperature and holding time

B: ramp (oC/min)

C: final temperature and holding time

Some GCs will allow for a more complex program.

A

B

C

Raising column temperature

1) Decrease retention time

2) Decrease resolution

3) Sharpens peaks

Comparison of isothermal and programmed temperature chromatography. Each sample contains linear alkanes run on a 1.6 mm × 6 m column containing 3% Apiezon L (liquid phase) on a 100/200 mesh VarAport 30 solid support with He flow rate of 10 ml/min.