EXPERIMENT 1: GAS CHROMATOGRAPHY (GC) OPTIMIZATION OF FLOW

RATE AND COLUMN TEMPERATURE.

NAME

MUHAMMAD RAHIMI BIN ZAHURI

(2015829866)

PARTNERS NAME

AKMAL ARSYAD BIN MOHD RAFFI

(2015875404)

MOHD IQBAL BIN NORAZMAN

(2015840192)

MUHAMMAD RAHIMI BIN ZAHURI

(2015829866)

SYARAFUDDIN BIN MOHD SHAMSIBY

(2015872898)

DATE OF EXPERIMENT:
DATE OF SUBMISSION OF REPORT:
Introduction:

In this experiment, there are several objectives which are firstly, to optimize the flow rate and
column temperature of the gas chromatography, to determine the retention time and resolution of
the mixture of methyl esters, and to determine the effects of column temperature and flow rate on
the separation of the methyl esters compound. Gas chromatography is a versatile and unique
technique at which a liquid or gas is introduced into the injection port through rubber septum into
a stream of inert gas which is called mobile phase or carrier gas. The gas is then passes through a
chromatographic column kept in an oven that controls the column temperature. The column is
lined or packed with stationary phase and as the analytes pass through the column, they partition
between the mobile phase and stationary phase. The analytes which has low affinity for the
stationary phase will spend more time in the column which then pass through more slowly
compared to the analytes which is not retained onto the stationary phase. At the end of the
separation, the analytes are then detected by the detector quantitatively which is in the form of a
chromatogram. The solutes retention time is the characteristic interval required for it to pass
through the entire column under particular conditions such as temperature, gas flow rate, and
stationary phase composition. Under a given conditions, a single analyte will always have the
same retention time. The height of the peak is proportional to the concentration of the solute. In
this experiment, the main objective is to explore gas chromatography, including the concepts of
retention time and resolution using a mixture of methyl esters which are methyl laurate, methyl
myristate, methyl palmitate, methyl stearate, and methyl linoleate. The effects of column
temperature and flow rate on the separation of these compounds will also be investigated. The
factors that contribute to the efficient separation of mixture included the affect of carrier gas flow
rate on the isothermal and temperature programming GC separation of methyl esters. The elution
rate of the compound depends on the volatility of the compounds, column temperature, carrier
gas flow rate, and the length of the column of the particular GC system.

Experimental:
Instrument used: Gas Chromatograph (Agilent Technologies 6890N) equipped with flame
ionization detector (FID) and 30m x 250m x 0.25m HP5-MS capillary column.
a. Instrument set up
Injection port
Injector port temperature
Column temperature
Carrier gas flow rate
Detector temperature

: Split (40:1)
: 250C
: 210C
: 30 cm sec-1
: 250C

b. Effect of carrier gas flow rate on isothermal GC separation of methyl esters.

0.4L of standard mixtures were injected isothermally at 170C at carrier gas flow rate of
20ml/s. Then, the flow rate was increased to 50cm 3/sec. The system was allowed to
equilibrate before the standard mixture was injected again.

c.

Effect of column temperature on isothermal GC separation of methyl esters.

0.4L of standard mixtures were injected isothermally at 170C, 190C, and 210C at the
optimal carrier gas flow rate. The effect of column temperature on the separation,
resolution, and analysis time was investigated.

d.

Separation of methyl esters using column temperature programming.

Standard mixture at the optimal carrier gas flow rate was injected using linear
temperature ramp from 100C to 290C at 40C/min.

e.

Identification of components in methyl esters mixture.

Each methyl esters was injected individually to identify the various compounds in the
standard mixture using the optimized GC conditions.

Results and Discussion:

For standard mixture 1
Table 1: Flow Rate 30cm3/sec and column temperature 210C

Repeatability

Peak

Retention
time, tR

Rs1, peak
Width

2.721

0.0775

3.801

0.1391

6.058

0.2459

2.721

0.0865

3.798

0.1446

6.048

0.2491

1 & peak

Rs2, peak
Average

2 & peak

9.97

11.72
9.65

9.32

Table 2: Flow rate 50cm3/sec and column temperature 210C

Average

11.58
11.43

Repeatability

Peak

Retention
time, tR

Rs1, peak
Width

1.636

0.0534

2.291

0.0947

3.654

0.1822

1.636

0.0563

2.290

0.0965

3.651

0.1823

1 & peak

Rs2, peak
Average

2 & peak

8.85

9.84
8.71

8.56

Average

9.80
9.76

Table 3: Flow rate 70cm3/sec and column temperature 170C

Repeatability

Peak

Retention
time, tR

Rs1, peak
Width

2.073

0.1094

4.091

0.2438

9.055

0.4818

2.074

0.1035

4.097

0.2410

9.049

0.4810

1 & peak

Rs2, peak
Average

2 & peak

11.43

13.68
11.59

11.74

Table 4: Flow rate 70cm3/sec and column temperature 190C

Average

13.70
13.72

Repeatability

Peak

Retention
time, tR

Rs1, peak
Width

1.464

0.0654

2.396

0.1296

4.496

0.2731

1.463

0.0609

2.394

0.1292

4.488

0.2755

1 & peak

Rs2, peak
Average

2 & peak

9.56

10.43
9.68

9.80

Average

10.39
10.34

Table 5: Flow rate 70cm3/sec and column temperature 210C

Repeatability

Peak

Retention
time, tR

Rs1, peak
Width

1.171

0.0459

1.642

0.0779

2.618

0.1498

1.172

0.0474

1.642

0.0787

2.620

0.1526

1 & peak

Rs2, peak
Average

2 & peak

7.61

8.57
7.53

7.45

Table 6: Comparison of Resolution in Different Flow Rate and Temperature

Average

8.57
8.56

Flow Rate and Temperature

Temp 210C
Flow rate 30cm3/sec
Temp 210C
Flow rate 50cm3/sec
Temp 170C
Flow rate 70cm3/sec
Temp190C
Flow rate 70cm3/sec
Temp 210C
Flow rate 70cm3/sec

Resolution, Rs1

Resolution, Rs2

9.65

11.68

8.71

9.80

11.59

13.70

9.68

10.39

7.53

8.57

For individual methyl ester

Table 7: Retention time for individual methyl ester at temperature of 210C, flow rate of
70cm3/sec
Methyl Ester

Retention Time, tR

Width

Stearate

4.585

0.2784

Laurate

1.175

0.0486

Palmitate

2.613

0.1510

Myristate

1.643

0.0807

Linoleate

4.234

0.2720

Discussion:
From the experiment conducted, the results show that the chromatogram of any compounds or
samples is dependent of the condition of the gas chromatography (GC). Different condition of
the gas chromatography such as the column temperature and the mobile phase flow rate will

affect the efficiency of the separation between two components in the mixture. By analyzing and
comparing the results of the experiment conducted, an optimum condition to separate the
components of the mixture can be obtained. In this experiment, GC was used to identify various
components in the standard mixture of methyl esters using the optimized GC conditions. The
standard methyl ester contains several individual components which are methyl laurate, methyl
myristate, methyl palmitate, methyl stearate, and methyl linoleate. In this experiment, the
instrument is set to be in split injection mode because only small amount of sample is introduced
into the column. This type of injection also produced sharper and narrower peak compared to the
splitless injection. In terms of optimizing the condition of the GC, the optimum condition to
separate the methyl ester is determined by the injection of sample at different temperature and
flow rates. The standard mixture is injected at different flow rates which are at 30, 50, and
70cm3/sec while the temperature is 170, 190, and 210C. Different types of condition produced
different resolution and retention time of the components in the chromatogram at which this two
determine the optimum condition of the separation. Based on the chromatogram results obtained
from this experiment, by comparing between all of the results, it can be concluded that the
optimum condition for the separation of the methyl ester is at the flow rate of 70cm 3/sec and at
the temperature of 210C. This is because at the flow rate of 70cm 3/sec and at the temperature of
210C, the lowest resolution value is obtained compared to the others. The best resolution value
is between the values of 1 to 20 at which both components are well separated with the best
analysis time. If the value is above 20, the separation is good but in terms of analysis time, the
separation takes longer time to complete which is not desirable in the chromatography analysis.
For the individual methyl ester, each component is identified by comparing with the
chromatogram of the standard mixture. For the separation of the individual sample, the optimum
condition is used with the flow rate of 70cm 3/sec and temperature of 210C. The average
retention times of the individual peaks of methyl ester are shown in table 7. Based on the results
obtained, it shows that methyl laurate elutes first followed by methyl myristate, methyl palmitate,
methyl linoleate, and lastly methyl stearate. In terms of precaution, in handling of the instrument,
the injection process must be done immediately and the analysis button on the instrument must
be pressed immediately after the sample is injected. The tip of the injector must not be placed too
long in the injector port. This is due to the high temperature of the injector port which may cause
the sample to be vaporized before injects into the column. This may affect the result of the

chromatogram which causes the band to broaden. The injector must also be washed and cleaned
before being used in order to avoid any contaminants which may cause unneeded peaks to exist
in the resulting chromatogram. Besides, certain result of the individual methyl ester
chromatogram produces peak which is not smoothly resolved. This are due to the problem of the
software of the PC used which connected to the GC instrument.

Conclusion:
From this experiment, it can be concluded that the efficiency of the separation between two
components in a mixture using GC-FID is affected by changing the column temperature and the
flow rate of the carrier gas or mobile phase. All of the five individual compounds are separated
well at high temperature which is at 210C with the flow rate of 70cm 3/sec which is the optimum
condition to separate the components from the given mixture.

References:

Gas Chromatography. (n.d.). Retrieved from

http://teaching.shu.ac.uk/hwb/chemistry/tutorials/chrom/gaschrm.htm
Saim, N., Tajuddin, R., & Saaid, M. (2014). Experiment 1 Gas
Chromatography(GC): Optimization of Flow Rate and Column
Temperature. In ANALYTICAL SEPERATION METHODS LABORATORY
GUIDE 2nd edition (pp. 1-2). Selangor: UiTM Press.
Skoog, W. H. (2000). Analytical Chemistry: An Introduction, 7th edition.