Chapter 27 Gas Chromatography
Problems: 1, 2, 3, 4, 6, 7, 8, 9, 10,11,13, 14, 20, 21, 22
Sample is vaporized and injected onto head of column
Elution comes from a gas flowing through the column
this gas does not interact with the sample, only acts to make it flow along
In principle 2 major types
Gas-solid chromatography (GSC)
gas-liquids chromatography (GLC)
GSC based on physical adsorption of solute molecules onto a solid
usually lots of tailing due to non-linear process
so not used very much outside of a few small molecules
only discussed lightly in section27-E
GLC, GC
based on adsorption of solute onto a liquid coating on a solid
GLC is found in all fields of science and name is usually shortened to just GC
theory first proposed in 1941
first lab demonstration in 1952
first commercial instrument 1955
in 1985 estimated that 200,000 in the world
Widely used, well understood, bulk of the chapter is devoted to this
27A Principles of Gas-Liquid Chromatography
Only minor changes from theory of last chapter due to compressibility of gaseous
mobile phase
27A-1 Retention Volumes
Because pressure and T affect gas flow through column, usually talk about
retention volume
volume of a substance retained on a column
=
tRF
OR
tR retention time
F flow rate in column
VM=tmF
Vm = volume of an unretained species
OR
Can’t measure flow rate of gas inside the column directly
but can easily measure flow rate as gas leaved the column with a bubble meter
(figure 27-2) In this case we have to adjust for the fact that the bubble meter is at
one temperature and the column is probably heated, and that the gas gets
saturated with water in the bubble, and this changes its pressure slightly so we
have:
2
Fm is Flow measured
TC is column temp (in K)
T is ambient T (in K)
P is ambient P
PH2O is partial pressure of water at this T
Both OR and VM depend on pressure inside the column
But there is a pressure difference between the head and the foot of the column
(Inlet and outlet)
To correct for this pressure factor you have
V0R = jtRF and V0M= jtM F
where j s calculated from
Pi is inlet pressure
P is outlet pressure (usually ambient P)
Specific retention volume Vg is then
Where W is the mass of the stationary phase
determined when you make (or buy) the column
Tc is temp of column in K
27A-2 Relationship between Vg and K
Rather than deriving as the book does, let’s just cut to the chase
Vg the specific retention volume of a substance
Vg = K/DS x 273/TC
Where K is the distribution constant
3
And DS is the density of liquid on the stationary phase
Note that the Vg depends only on the distribution constant and the density of the
stationary phase. Thus it might be a useful parameter for identifying
compounds. The literature does have these numbers recorded, but they aren’t
condensed into one table, and the data isn’t that good
27A-3 Effect of Mobile-Phase Flow rate
The equation we derived in the last chapter apply here
One of the more critical is the longitudinal diffusion term because diffusion in a
gas is very fast (10,000 more than in liquid)
This makes minima in H vs flow rate broad
27B Instruments
30 manufactures
130 different GC’s to choose from
1,500-40,000 (ours is in the 15-20,000 range)
Basic components figure 27-1
note flow splitter our instrument does not use in this way
we do split flow to stat of column, just so don’t overload column
but detector does not need a reference so isn’t piped to detector
27B-1 Carrier Gas Supply
Carrier gas must be chemically inert
typical are He, N2, H2
(We use He on the mass spec, N2 -cheaper, on the non-mass spec)
associated with gas are pressure regulators, gauges, flowmeters
Often molecular sieves to remove trace O2 or H2O
Flow rates controlled by 2 stage regulator at the cylinder
inlet pressures usually 10-50 psi
flow rates 25-150 ml/min - packed columns
1-25 ml/min open tubular capillary column
Flow rates established by bubble gauge or rotameter
(actually on our machine is monitored and controlled electronically so don’t have
these devices, just have to assume they are right!)
27B-2 Sample Injection System
Want the sample to be injected in a small, discrete volume of gas or a ‘plug’
if the plug is too big, or if fades on and off, then get band spreading and poor
resolution
4
Figure 27-3 a typical injection port
sample injection of a few ul (.1 to 20, may be gas or liquid)
needle pierces as self sealing septum
goes into a heated injector
(Usually 50 C above B.P of highest BP component
Capillary columns require even less sample
Sample splitter is used to take a small fraction of flow to column and
dump the rest
For more reproducibility can use a rotary valve or autoinjector
27B-3 Column Configurations + Ovens
2 major types of columns
1. Packed
2. Open tubular or capillary
Of the two capillary is faster and most efficient, so is beginning to replace
the packed for almost all purposes
Length <2 m to > 50 m
stainless steel, fused silica, glass or Teflon
usually formed into coils so they can fit in the small oven
Talk more about their composition in next section (27C)
Column is important parameter, need to be controlled to a 1/10th of a degree
Optimum column temp equal or slightly > BP of solute is required for elution in
the 2-20 min range
Sample with a broad range of BP, need to use temperature programming so
change T of oven during the run so compounds come out more quickly
In general better resolution with lower T, however longer run time so have to
make a trade off here
27B-4 Detection Systems
Dozens of different detectors the bok describes the most common ones, I will
look at just one or two
Characteristics of the Ideal Detector
1. Must be sensitive enough for your sample
Sensitivities of detectors can vary by 107 so it ll depends on what
you need for your experiment
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2. Good stability and reproducibility
3. linear response over several orders of magnitude
4. Operate anywhere between Rt and 400C
5. Short response time independent of flow rate
6. High reliability and ease of use - foolproof
7. Similar response for all solutes or at least predicable response
8. Nondestructive ?? (not always necessary)
Not such single detector in existence
Flame Ionization Detectors (FID’s)
This is what we have
Probably the most generally used and generally applicable
Figure 27-6
Simply a burner
Column effluent mixed with H2 and air and burned
When burned most organic compounds produce both ions and
electrons
A potential of a few hundred V applied between flame jet and tip of
burner
These ions and electrons allow a small current to flow, so set up
electronics to measure this current
Ionization of C compounds in a flame is not well understood
But observe that # of ions is proportional to reduced C in
compound
IE CH3 most signal, COOH oxidized least signal
Flame will detect number of C entering detector, so is mass
sensitive (not concentration sensitive)
Response of functional groups is variable
Insensitive to H2O, CO2 SO2 and Nox
That actually makes it a good detector for samples with these
contaminants
High sensitivity 10-13 g/s
Large linear response range 107
Low noise
Rugged and easy to use
6
Only down size is that it does destroy the sample
Thermal Conductivity Detectors (TCD)
One of the earlier detectors, so found on many older machines
we don’t have
Figure 27-7
Boils down two 2 wires, one sits in column effluent, one sits in flow of gas
that did not go through column
When the two gases are the same, the wires have the same electrical
resistance, and the circuit has zero output
When column effluent contains an added compound, this half of the cell
has a higher thermal conductivity, so its resistance changes, and the
circuit ‘sees’ a difference
Simple and rugged
large dynamic range 105 (not quite as good as FID
Responds to both organics and inorganics
Nondestructive
Low sensitivity (10-8 g/ml carrier gas
So FID thousands to million times better
Essentially can’t be used in capillary columns because not sensitive
enough
Skip all others,
27C Gas Chromatographic Columns and Stationary phases
First GC, 1950's done in packed columns
Stainless steel or glass tube
packed with an inert powder
thin film of a liquid adhered to surface of powder
Theory showed, however, that unpacked columns with diameters of <1mm
should be better in both efficiency and speed
This is the capillary column, with the stationary phase as uniform coating
on the inside of the tube
First demonstrated that this would work in late 1950's
but manufacturing problems weren’t solved until late 1970's
27C-1 Open Tubular Columns
Open Tubular or Capillary Columns 2 major types
7
Wall-coated open tubular (WCOT)
Support-coated open tubular (SCOT)
Wall -Coated
Capillary tube with thin layer of stationary phase on inside of tube
Support Coated
Inner surface of tube is coated with a thin (30 um) film of support
material. Stationary phase coated on that
Lots more stationary phase
So greater sample capacity
Not quite as good efficiency as WCOT, but still very good
Early WCOT made of stainless steel, Al, Cu, or plastic
Then made of glass with inside etched with acid to give it a rough surface
Now Fused-silica
Specially purified silica containing metal oxides
Much thinner walls, so coated with polyimide coating
Result is fairly flexible, can be bent
This is what is generally used now
Most widely used silica have diameters of .32 and .26 mm
High resolution diameters .2 to ,15 mm
A but more difficult to use
Need to split sample so not all goes on column
.53 mm capillaries - ‘megabore’ columns for large samples
Table 27-1 compares properties of columns
27C-2 Packed Columns
glass metal or teflon tubes
2-3m length, but made in a coil
2 to 4 mm diameter
densely packed with a fine packing material
packing material coated with .05 to 1 um of stationary phase
Solid Support Material
Ideal, small, uniform, inert spheres with good mechanical strength
And surface area of at least 1m2/g
Must be uniformly wetted by liquid phase
No such stuff
Diatomaceous earth is widely used
Skeletons of diatoms that inhabited ancient lakes and seas
8
Actually pretty close to ideal
Particle Size of Supports
Efficiency of column increases with decreasing particle size
However decreased particle size means better packing so flow rate
decreases
Since having pressures > 50 psi causes problems this puts a
practical lower limit on particle size
60-80 mesh (260 to 170 um)
80 to 100 mesh (170 to 149 um)
27C-3 Adsorption to packings or walls (not the stationary phase)
Silica used in column and found in diatomaceous earth has a problem
Surface is coated with silanol groups
OH OH OH
O-Si-O-Si-O-Si
|
|
|
These groups tend to bind polar or polarizable groups
Makes peaks tail
Need to remove or cover up surface Si-OH groups
react with dimethylchlorosilane (DMCS)
|
CH3
|
CH3
-Si-OH + Cl-Si-Cl 6
-Si-O-Si-Cl + HCl
|
CH3
|
CH3
Then remove Cl with MeOH
|
CH3
|
CH3
Si-O-Si-Cl + CH3OH 6 Si-O-Si-OCH3 + HCl
|
CH3
|
CH3
Diatomaceous earth still slightly reactive due to metal oxide impurities
These can be washed off with an acid was BEFORE the sialination
Purified Si used for columns doesn’t have this impurity.
27C-4 The stationary phase
Ideal
low volatility (BP >100 higher than max T
Heat stable
chemically inert
solvent characteristics such that k’ and " allow all solutes to be resolved
Hundred have been used
9
presently about 10 will do for most applications
guidelines to help you choose what should work
but always have to do the experiment
retention time depends on K (distribution constant)
Related to chemical nature of solute and stationary phase
need different k’s for each solute in the mix
K’s can’t be too big or too small
Use like dissolves like rule
polar solutes (alcohols, acids, amines)need polar stationary phases -CN, CO, and -OH
Nonpolar solutes (saturated hydrocarbons) need hydrocarbon like
stationary phase or dialkyl silanes
Intermediate solutes ( ethers ketones aldehydes) something in between
Widely used stationary phases
Table 27-2
in order of increasing polarity
these 6 can do about 90% of the sample you come up with
Five of these are polydimethylsiloxanes
general structure
Just change the R groups to modify properties
polydimethyl -R are both methyls makes nonpolar
phenyl -C6H5
cyanopropyl C3H6CN
trifluoropropyl C3H6CF3
Vary percentages to get intermediate qualities
#5 polyethylene glycol
HO-CH2-CH2-(O-CH2-CH2)nOH
is a widespread polar type column
Bonded and Cross-linked Stationary phases
Bonging and cross linking is used to form a chemical bond between the
stationary phase and the Si support. This makes the stationary phase
more stable and longer lasting. (Prevents Bleeding)
10
Sometimes if stationary phase is contaminated can even rise it off with a
solvent dn not lose it
Bonding process in commercial columns is proprietary
Can do it yourself with peroxide free radical reactions
Film Thickness
commercial columns 0.1 ro 0.5 um
affects retention
thicker films for more volatile analytes so retained longer
Most applications .26 or .32 mm columns use 0.26 mm films
Megabore columns 1 to 1.5 um film
Chiral Stationary phases
how do you resolve enantiomers?
Use a chiral column!
27D Applications of G(L)C
2 major roles
1. To do separations for any volatile species including Organic, metal-organic,
biochem
2. Last step in an analysis for identification of unknowns
ID based on retention time alone is very limited
but hook into mass spec or IR have a great hypenated method
27D-1 Qualitative Analysis
GC used for purity check or organics
Any stray peaks indicate an impurity
Area of peaks roughly tell amount of contamination
Retention times can be used to identify compounds
But lots of variables, so time alone is not a great ID parameter
OK if combine with experiments with knowns compounds to
confirm or deny the identity of a peak
Selectivity Factors
From earlier
Selectivity factor "
11
If you choose some standard factor for A
then relative retention of B or the above " factor can be used to identify B
and this number is relatively independent of all column variables except
temperature
No single great standard, so not standardized use of this factor
Retention Factor
Retention Index I
Proposed by Kovats in 1958
based on normal alkanes
(CH4, C2H6, CH3CH2CH3...)
In a particular system find the t alkanes that bracket you compound
Retention # for alkane is 100x n
(IE for methane =100, ethane=200, etc)
When yours is in between it gets an in between number
This retention index is independent of column packing, temp, or just about
anything else, so is a nice useful number
Easily available reference compounds, so fairly well used
27D-2 Quantitative analysis
signals from most GC detectors can be used for quantitative analysis
good to about 1% when done right
27D-3 Interfacing with spectroscopic methods
With GC alone, only way to identify a peak is to collect it (run it through a
cold trap to change if from a gas back to a liquid or solid) then run the sample into an
MS or IR or NMR. Trapping was difficult, and getting enough material for an analysis
was difficult
Now design machines so column effluent flows directly into a detector
designed for gases
Best is GC/MS
Gas Chromatography/Mass Spectrometry (GC/MS)
Since output of a GC is a gas, it is the ideal sample to run directly into an
MS which needs a molecular gas as a starting material
12
With smaller capillary columns can put column effluent directly into mass
spec
With larger megabore columns need to reduce volume a bit (preferably by
getting rid of carrier gas instead of sample. In this case use a jet
separator (figure 27-14)
Quad, iontrap and FTMS are all fast enough that you can scan the
complete molecular spectrum in less than a second, so you can get MS of
every component coming off the GC
First GC/MS appeared in 1970s
(Ours showed up in 2002)
Generally have 2 different display modes
Total mass (just shows total of all ions- use to keep track of where
the peaks are)
Spectral mode (show each mass spectrum as it occurs - use to
identify a given peak)
Examples shown in figure 27-16
GC/FTIR -skip
27E Gas-Solid Chromatography - skip