Gas Chromatography

Method Development
Gas Chromatography
 Efficient separation is dependent on
 Compounds traveling thru the column at different
rates

 Rate at which a compound travels thru a

column is dependent on the factors below:

 1.Volatility of compound
 Low boiling point compounds travel faster than high
boiling point compounds
Gas Chromatography

 2.Polarity of Compounds
 Polar compounds will move slowly, esp. if
column is polar

 Polarity of organic functional group in

increasing order
 Aliphatic hydrocarbons < olefins < aromatic
hydrocarbons < halides < ethers < esters/
aldehydes / ketones < alcohols/amines < amides
< carboxylic acids < water
Gas Chromatography
 3.Column Temperature
 Raising the column temperature speeds up all the tR of the
compounds in the mixture

 4. Column Packing polarity

 Usually, all compounds will move slower on polar columns,
with polar compds showing a larger effect

 Polarity of stationary phase should match that of sample

components

 When the match is good, the order of elution is determined by

the boiling points of the eluents
Gas Chromatography
 5.Flow rate of gas
 Increasing flow rate, increases the speed of
compds moving thru the column

 6.Length of column
 Long columns causes compds to elute at a
longer time.
 Employed only to obtain better separation/
resolution
Gas Chromatography

 Generally the number one factor to

consider in obtaining good separation of
compds is it’s boiling point

 Differences in polarity of the compds is

only important if the mixture consists of
different polarity compds
Gas Chromatography
-- Method develop
 Areas to optimize in method development
are:
 Injector temperature
 Type of carrier gas
 Optimal range of velocities (van deemter’s
plot)
 Too low or high may result in loss of resolution
 Balance resolution vs analysis time

 Column / oven temperature

Gas Chromatography
--general elution problem

Solute signal

1 2

 3 4 5 6

(a)

Chromatogram shows 3 pairs of components with a widely different

distribution constant (K) and retention factors. (k’)
In the above , conditions are adjusted so that k’ for compds 1 and 2
are in the optimal range of 1 to 5, but other components are larger
than the optimal.
Hence the compds 5 and 6 appear after a significant period of time
and are very broad.
Gas Chromatography
--general elution problem
 3,4
1,2 5 6
Solute signal

(b)

As shown above changing conditions to optimize the separation of

components 5 and 6, bunches the peaks for the first four
components.
Resolution of 1 and 2 , and 3 and 4 are unsatisfactory
This condition however gives an ideal analysis time. (fast)
Gas Chromatography
--general elution problem
 1 2 3 4
Solute signal

(c)

5 6

a third set of condition, in which the value of k’ are optimal for 3 and
4.
However the k’ for the other peaks are not optimized.
This phenomenon is known as the general elution problem. (one gc
condition is used for all compds but resolution and efficiency is not
optimum for all the components)
Gas Chromatography
--general elution problem
 The solution for this problem
 Change the conditions that determines the
value of k’ as the separation proceeds.

 Can be performed in a stepwise manner or

continuously.
Gas Chromatography
--general elution problem

 To change the conditions that determines

the value of k’ as the separation proceeds.

 Can be performed in a stepwise manner or

continuously.
Gas Chromatography
--general elution problem
 Hence for chromatogram shown,
 In (a), immediately after the elution of
compds 1 and 2
 Conditions should be changed to that of
the optimal for separating compds 3 and
4, as in chromatogram (c).
 Once these are eluted,conditions should
be changed to those produced in (b).
Gas Chromatography
--general elution problem
 Often after this is done , the separation
of the compds will be done satisfactorily
in the minimum amount of time.
Gas Chromatography
--isothermal vs gradient elution
Isothermal at 45oC

Isothermal at 145oC

Programmed at 30oC to 180oC

Gas Chromatography
--isothermal vs tempt programming
 In general , optimum resolution is
associated with minimal temperature

 Low temperature, however, result in

longer elution times hence slower
analyses
Gas Chromatography
--isothermal vs tempt programming
 Isothermal Run
 Constant column tempt. Thru out
analysis.

 Refer to example in hand out pg 21

 Shows effects of isothermal run at 60°C.
Gas Chromatography
--isothermal vs tempt programming
 Isothermal Run
 The result is an increase in retention
time of all compounds.
 The heights of the later eluting peaks are
reduced and peak widths increased
because they are more affected by the
lower tempt. Program used.
Gas Chromatography
--isothermal vs tempt. programming
 Tempt. Programming
A chromatographic development
technique, used largely in GC.
 To accelerate the elution rate of late peaks.
(too long to elute)

 Achieved by increasing the column tempt.

Continuously (linear function of time) or in
step wise manner.
Gas Chromatography
--isothermal vs tempt programming
 Oven
 Samples with wide boiling range, necessary
for temperature programming

 a technique in which the temperature of a gas

chromatographic column is increase
continuously or in steps during elution
Gas Chromatography
--isothermal vs tempt programming
 Tempt. programming
 Involves increasing temperature of a gas-
chromatographic column as a function of time.

 Low boiling point constituents are separated

initially at temperatures that provide best
resolution

 As separation proceeds, column temperature is

increase so that the higher boiling constituents
come off the column with good resolution and at
reasonable lengths of time.
Gas Chromatography

 Refer to hand out pg 22

 for effects of split ratio, on pg 23

 Overloaded Chromatography
 Pg 24 & 25
Gas chromatography

 Applications of GC
 Gc is an effective tool for separations of
 Complex organics
 Metal- organics

 Biochemical species

 Made of volatile species or species that

can form volatile derivatives
Gas chromatography
 GC can also give info after separation
has been completed, such as:
 Retention time
 Retention volumes

 Used for qualitative analysis

 Peak heights
 Peak areas

 Used for quantitative analysis

Gas chromatography

 Trend now is to combine GC’s superior

separation qualities with
 Superior identification properties of

 Mass spectrometers
 IR spectrometers

 NMR spectrometers
Gas chromatography

 Qualitative analysis
 GC often used to establish organic
compounds purity
 Appearance of additional peaks
 Reveal contaminations are present

 Area under the peaks

 Represent estimate of contaminations
Gas chromatography
 In theory, retention times
 should be useful to identify components in mixtures

 However applicability is limited by

 a number of variables that must be controlled to
obtain reproducible results

 Still GC provides an excellent means of

 confirming presence or absence
 of a suspected compound in a mixture, provided an
authentic sample of the compound is available
Gas chromatography
 When authentic sample is added to the
compound mixture
 No new peaks will appear
 Instead compound peak will increase in size

 Evidence is more convincing if the effect

is duplicated
 On different column
 and at different temperature.
Gas chromatography
Retention Index
 Retention Index (I)
 Was proposed by E. Kovats in 1958 for
 Identifying solutes from chromatograms

 The retention index for any solutes can be

calculated from a chromatogram of
 Mixture of solute with at least two normal alkanes
 Whose retention index bracket that of the solute
Gas chromatography
--retention index
 Retention Index Scale is based on
normal alkanes

 By definition retention index of normal

alkanes is
 Equal to 100 times the number of carbons in
the compound
 Regardless of column packing, temperature,
or other chromatographic conditions
Gas chromatography
--retention index
 Retention indexes of all compounds
other than normal alkanes
 vary (by several 100 retention index units)

 A plot of the homologous series of the

 log of adjusted retention time (tR’=tM – tR)
versus the number of carbon atom is linear
 Provided the lowest member of the series is
excluded
Gas chromatography
--retention index

Toluene I=7.49x100
n-nonane
Benzene I=6.44x100 I=900
Log (adjusted tR, s)

2,2-dimethyl butane n-hexane

I=5.37x100 I=600

6.44 7.49
5.37

4 5 6 7 8 9
Number paraffinic carbon atoms
Gas chromatography
--retention index
 A plot of C4 to C9 is shown in fig 27-18
 Also shown are the adjusted tR of 3
compounds on the same column and at the
same temperature
 Their retention index is obtained by
multiplying the abscissa value by 100.
 Toluene I = 749
 Benzene I= 644
 2,2-dimethylbutane I= 537
Gas Chromatography
--Retention index
 Normally, a graphical procedure is not
required to determine retention index

 Instead adjusted retention time data are

calculated by interpolation from a
chromatogram of a mixture of the solute
of interest and two or more alkane
standard
Gas Chromatography
--Retention index
 EX.
 Use the retention index data given to
calculate the retention index of 1-
hexene.

Sample Retention time , min

Air 0.571

n-pentane 2.16

n-hexane 4.23

1-hexene 3.15
Gas Chromatography
--Retention index
 Note*- retention Index for normal alkane
 Is independent of temperature and column
packing
 Therefore: I for heptane is always 700.

 However I for other solutes , vary widely

from one column to another.
Gas Chromatography
--Retention index
 Eg: I for acenathene on a cross linked
polydimethyl siloxane stat. phase at 140°C
is 1460.

 with 5% phenyl-polydimethyl siloxane as

stat. phase, it’s 1500 at same tempt.

 With polyethylene glycol as stat. phase, it’s

2084
Gas Chromatography
--Retention index
 Retention Index System
 Has advantage of being based on readily available
standard reference material
 That cover a wide range of boiling point

 Furthermore it is temperature independent.

 In 1984, Sadtler Research Laboratories

introduced a library of I for 4 types of fused
silica open tubular column