Experiment 5: Gas chromatography (GC)

Introduction

Chromatographic analysis is used to separate complex moistures of compounds. First used in

the early 1900s chromatography got its name because it was used to separate different
mixtures of colored compounds. By the 1930s the popularity of chromatography had
increased as chemists realized that this experimental technique could also be used to separate
mixtures of colorless compounds. All chromatographic systems have two phases, a mobile
phase and a stationary phase. The mobile phase is a liquid or a gas that carries a sample
through a solid stationary phase. As the sample in the mobile phase passes through the
stationary phase the compound in the mixture will separate because of differences in their
affinities for the stationary phase, and differences in their solubilities in the mobile phase

In chromatographic analysis, the eluted compounds are characterized by retention times, t R.

Qualitative analysis involves determination of tR of analytes and comparing then with tR of
standards. Quantitative analysis is accomplished by comparing the areas of the analyte peaks
with those of standards

Instrumentation

Mobile phases are generally inert gases such as helium, argon, or nitrogen. The injection port
consists of a rubber septum through which a syringe needle is inserted to inject the sample.
The injection port is maintained at a higher temperature than the boiling point of the least
volatile component in the sample mixture. Since the partitioning behavior is dependent on
temperature, the separation column is usually contained in a thermostat-controlled oven.
Separating components with a wide range of boiling points is accomplished by starting at a
low oven temperature and increasing the temperature over time to elute the high boiling point
components. Most columns contain a liquid stationary phase on a solid support. Separation of
low-molecular weight gases is accomplished with solid adsorbents. The sample is swept
through an open tubular column by a carrier gas, and the separated eluents (the compounds
exiting the column) flow through a detector, whose response is displayed on a computer
screen. The column must be hot enough to produce sufficient vapor pressure for each solute
to be eluted in a reasonable time. The detector is maintained at a higher temperature than the
column so that all the solutes are gaseous at the point of detection. Figure 5 below shows the
main components of a gas chromatograph.
Some GC detectors are as follows:

Flame-ionization detector (FID)

FID stands for flame ionization detector. What that means is that as the effluent (carrier gas
and any organic compounds) comes out of the column they are ignited in the flame made of
hydrogen and air. The compounds produce ions as they burn. These ions conduct electricity.
Changes in current within the flame are measured and sent to the computer to be seen as
peaks on the chromatogram.

FID is a good general detector for organic compounds and is able to detect at the nanogram
level. The FID is extremely sensitive with a large dynamic range and its only disadvantage is
that it destroys the sample.

Electron-capture detector (ECD)

ECD stands for Electron Capture Detector. This detector is very sensitive to halogenated
compounds, as well as compounds with very electronegative functional groups such as nitro
groups and peroxides. The detection limit of this detector for halogenated compounds can be
as low as the picogram level. This detector cannot detect compounds such as hydrocarbons,
amine, and alcohols, making it very useful in quantifying herbicides and insecticides.

The ECD is as sensitive as the FID but has a limited dynamic range and finds its greatest
application in analysis of organic molecules that contain electronegative functional groups,
such as halogens, phosphorus, and nitro groups

Thermal conductivity detector (TCD)

The TCD is not as sensitive as other detectors but it is non-specific due to its response to both
organic and inorganic species and is non-destructive.
Objective:

1. To determine the retention times tR of n-butanol and 2-propanol.

2. To identify the components present in a standard mixture based on the tR.
3. To identify the component(s) present in an unknwon sample.
4. To determine the effect of temperature on tR and Rs.

Apparatus

Syringe

Beaker

Dropper

Chemicals

2-propanol

n-butanol

Standard mixture of 2-propanol and n-butanol (1:1) ratio)

Unknown sample

Procedure

A. Sample Handling
1. The syringe was rinse before filling it with the sample. The volume of the sample may
be more than the required volume. There should be no air bubbles in the syringe. To
remove any air bubbles, it should be tap gently
2. The syringe was hold vertically, needle up and push the plunger to the required
volume at eye level. The excess sample was removed using a tissue.
B. Experimental
1. The instrument was turned on.
2. The GC is then set using the following conditions:
a. Initial oven temperature : 70oC
b. Final oven temperature: 70oC
c. Injection temperature: 180oC
d. Detector temperature: 180oC
3. The component sample was then injected and the retention time of each component
individually was determined.
4. The standard mixture was injected and each component was identify by comparing
the retention time of each component with the retention time of each single
component determined previously.
5. The unknown sample was injected and the component(s) present in the unknown was
determine
6. The column temperature was changed as follows:
a. Initial column temperature: 100OC
b. Final column temperature: 100oC

The standard mixture was injected and the effect of reducing the temperature on the
retention time and Rs of the component was recorded in the report

7. The oven temperature was changed as follows:

a. Initial oven temperature: 140oC
b. Final oven temperature: 140oC
The standard mixture was injected and the effect of increasing the temperature on
the retention time and Rs of the components was commented.
C. Operation of the GC
Instrument: Agilent 6890
Operation instructions
1. For manual injection, the sample was injected into the septa(liner).
2. The oven column was checked. Make sure the instrument is in the off mode
3. The instrument was switched on. Warning tone will ring If the gas is not enough
4. ‘Instrument 1 online’ was clicked in the window screen. The instrument is
allowed to setup itself
5. After the setup is completed, click ‘method’  ‘edit entire method’ (make sure all
the items in this column are correct)  ‘ok’
6. The title was entered.
7. To select the injection, click ‘manual’. To select injection location, click ‘back’.
At instrument, edit inlet, choose ‘back’ and keep in information
8. If sample is in high concentration, choose ‘split’ mode. IF sample is in low
concentration or in small quantities, choose ‘splitless’ in inlet.
9. Go to the column section, choose ‘column no. 2’. At inlet column, choose ‘back’.
At mode column, choose ‘constant pressure’. At outlet column, choose ‘ambient’
10. Go to the detector column. Use FID (‘front’)
11. Inject parameter (time:5min, 1 µL)
12. After doing all of the above, click ‘ok’
13. Click ‘apply’ ’ok’ signal detail ’ok’ ’edit integration events’
’ok’’specify report’’ok’’run time checklist’’ok’
14. Go to ‘file’, click ‘save’, ’method’. Keep in the command.
15. During injection, make sure no bubbles appear in the syringe
16. Go to ‘run method’ and the instrument will wait for the injection
17. Set signal 2,fill in the sample name
18. Click ‘run method’ and the instrument will wait for injection
19. Inject the sample quickly(use both hands!) and then press start button quickly on
the instrument. Then quickly pull out the syringe from the septa
20. Go to data analysis, click ‘file’ then ‘load signal’
21. Enter ‘calibration’, click ‘new calibration table’. Enter your name and print the
document
22. For the next sample, click ‘data analysis’ and repeat steps 17-21
Reports

1. Include in your report the chromatograms you obtained and explained how you
interpreted the peaks

Diagram 1

Diagram 1(enlarge)
 Diagram 2

Diagram 2(enlarge)
 Diagram 3

Diagram 3(enlarge)
Diagram 1
This is the reading of standard mixture.
70oC(tR of 1.46, 1.60, 1.69, 1.80, 1.85)
At this temperature,the peaks that are fully resolves are at the retention time of 1.46,1.60, and
1.69. The two peaks, 1.80 and 1.85 are closely together,and it is not fully resolves.

100oC(tR of 1.39, 1.45, 1.52, 1.54)

At this temperature, the retention time of 1.39, and 1.45 and fully resolves while 1.52 and
1.54 are not fully resolved.

140oC(tR of 1.34, 1.39, 1.56, 1.77, 3.05)

At this temperature,all of the peaks,which are 1.34, 1.39, 1.56, 1.77 are fully resolves thus
this is a good temperature for compound to be fully distinguished.

Diagram 2
This is the reading of n- butanol.
70oC(tR of 1.81, 1.90, 1.95, 3.71)
At this temperature,it seems that the peaks are closely together,broad and not fully resolves.
The retention time are 1.81,1.9 and 1.95. At peak of 3.71,there seems to be a small broad
peak,and take a longer time to resolve.

100oC(tR of 1.45, 1.52, 1.55, 2.12, 3.00)

At this temperature,at the retention time of 1.45,2.12 and 3.00,it seems that the component
are fully resolves but at peaks of 1.52 and 1.55,the component are not fully resolves

140oC(tR of 1.39, 1.56, 1.77)

At this temperature,all of the peaks are full resolves,where the peaks are sharp,not
broad and take a shorter time to resolves. The peaks are 1.39,1.56 and 1.77
Diagram 3
This is the reading of 2-propanol.
70oC(tR of 1.48, 1.59, 1.68, 1.79)
At this temperature, the component at the peaks of 1.48 and 1.59 are not fully resolves while
tR for 1.68 and 1.79 are a good peaks,where it is fully distinguished and resolve at this
temperature and takes a short time too.

100oC(tR of 1.39, 1.41, 1.45, 1.53)

At this teperature,The paks are closely together,not fully resolves except at the retention time
of 1.53. The retention time of 1.39 and 1.41 are not fully resolves

140oC(tR of 1.33, 1.35)

At this temperature, there are only two peaks shows in this graph which are at the retention
time of 1.33 and 1.35. The peaks are not fully resolves and is not a good peak although it is
sharp. Thus it only takes a short time for compound to resolves.
 2. Explain the reasoning used to determine which peak in the ‘unknown’
chromatogram correspond to each component in the standard mixture

This is to determine what compounds are exist in the unknown chromatogram. As we

compared with the known chromatogram, we definitely can distinguish and take the
similarity peaks of known and unknown chromatograms, which is correspond to each
component in standard mixture. In an unknown chromatogram, there exist a compound and
this is what was used to determine it. By comparing the known chromatogram, we can detect
and select the similarity of peaks of known and unknown chromatograms. There will surely
be a corresponding to each component in the standard mixture.

3. Discuss the effects of reducing and increasing the column temperatures on t R and
Rs

(In discussion).
Questions

1. State the types of compounds which are suitable for analysis using GC

-The characteristics of the compound must be sufficient volatility and thermal stability.

2. Why is FID a suitable detector for this analysis?

-This is because when it comes to sensitivity and also the range of the linearity is wide,
FID would be suitable for it.

3. List two factors which can increase the efficiency of a GC column.

-Lengthening the column.

-Increase analysis time.

 DATASHEET EXPERIMENT 5

GAS CHROMATOGRAPHY (GC)

Name:Marvellis anak Machillies Date: 24/10/18

Student ID:2016248128 Group: AS1205B

Name of the instrument:

Table 5.1: Retention times of standards and unknown sample

Temperature(oC) Compounds Retention time,tR(min)
70 Standard mixture 1.46, 1.60, 1.69, 1.80, 1.85
70 2-propanol 1.48, 1.59, 1.68, 1.79
70 n-butanol 1.81, 1.90, 1.95, 3.71
70 Unknown sample
100 Standard mixture 1.39, 1.45, 1.52, 1.54
140 Standard mixture 1.34, 1.39, 1.56, 1.77, 3.05

Unknown number:

Lecturer’s signature,

______________________________
Discussion:
The purpose of the experiment is to:

1. To determine the retention times tR of n-butanol and 2-propanol.

2. To identify the components present in a standard mixture based on the tR.
3. To identify the component(s) present in an unknwon sample.
4. To determine the effect of temperature on tR and Rs.

Along the experiment,GC-FID was used instead of GC-ECD because of the extremely
sensitive of the instrument and it also has a wide range of linearity. GC-ECD is sensitive as
GC-FID but the dynamic range is limited but the advantages part of GC-ECD is that it does
not destroy the sample like GC-FID instead it finds its greatest application in analysis of
organic molecules. The mobile phase are gas and the stationary phase is the liquid. As the
reading of the standard mixture,2-propanol and n-butanol were recorded, it seems that the
standard mixture has the best peaks separation and fully resolves at temperature of
140oC,while for n-butanol is at temperature of 140oC and propanol is at temperature of
70oC.There are some factors that can increase the efficiency of the GC, which are lengthening
the column and increase the analysis time, so that the result that were collected is more
accurate.

There is some effect happen when reducing and increasing the column temperature or t R and
Rs.

If the column temperature reduced,

The retention time and column resolution would both increase.

If the column temperature increased,

The retention time and column resolution would decrease.

Conclusion:

In conclusion,the retention time of the n-butanol and 2-propanol are recorded in the table of
datasheet and results.The compound that prsents in the standard mixture based on the
retention time is 2-propanol.Lastly the effect of the temperature to Rs and tR are:

If the column temperature reduced,retention time will increase and column resolution will
increase too.

If the column temperature increased,Retention time will decrease,and column resolution will
decrease too.
References:

Brechbühler, B., Gay, L., & Jaeger, H. (1977). A micro electron capture detector for
temperature programmed analysis with capillary columns for a wide range of
applications. Chromatographia, 10(8), 478–486. https://doi.org/10.1007/BF02257363

GOLAY, M. J. E. (1958). Gas Chromatographic Terms and Definitions. Nature, 182(4643),

1146–1147. https://doi.org/10.1038/1821146a0

HARLEY, J., NEL, W., & PRETORIUS, V. (1958). Flame Ionization Detector for Gas
Chromatography. Nature, 181(4603), 177–178. https://doi.org/10.1038/181177a0

Zuo, H.-L., Yang, F.-Q., Huang, W.-H., & Xia, Z.-N. (2013). Preparative Gas
Chromatography and Its Applications. Journal of Chromatographic Science, 51(7), 704–
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