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CHM 510 GC - Gas Chromatography

Analytical Separation Methods (Universiti Teknologi MARA)

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CHM 510 LABORATORY REPORT

EXPERIMENT 1

TITLE: GAS CHROMATOGRAPHY

Name: MUHAMMAD SYAFIQ AKMAL BIN ZAMRI

Student No: 2018425444

Group: AS2452S1

Due Date: 30 Jun 2019

Lab Partner’s name:

Name Student No
AZIZAH SHAFIQAH BINTI ISMAIL 2018657276
WHITNEY SANDRA WALTER 2018224512
LORITA LAHAI AK WALTER 2018674564

Lecturer`s Name: DR WAN NAZIHAH BINTI WAN IBRAHIM.

Objectives:

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1. To explore the gas chromatogram instrument and learn the correct way on handling
the instrument.
2. To learn the concepts of retention time and resolution of the peak base on a mixture of
the standard been provided (methyl ester).
3. To study the effect of column temperature and flow rate on the separation compounds.
4. To identify the optimum temperature and optimum flow rate of the mixture standard.

Introduction:

Gas chromatography is an analytical separation technique that analyse compounds

eluted from a column, which carried through the column by a gas mobile phase. Efficiency
separation of compound by gas chromatography is dependent on the compounds travelling
through the column at different rate. There are our factors that influence the performance of
GC system. One of the factors is volatility of compound. The volatile the compound will
elute faster than compound that have high boiling points. Next is the column temperature of
GC. The higher the temperature of the column, the faster the separation of the compound in
the stationary phase. Carrier gas flow rate also give impact to the separation where increasing
the carrier gas flow will increase the speed of the compound mobilization through the
column. The longer the column, the longer the migration time of the analyte in it and take
longer time to be eluted. But the longer column will give better separation.

Resolution is the measure of how well the separation of compound in the column.
This definition can be proven by this formula:

2(tr 2−tr 1)
Rs = (Wb1+Wb 2)

The tr2-tr1 is the separation between two peaks example peak A and peak B; Wa and Wb are
the widths at the base of peak A and peak B respectively. The acceptable resolution is on

Rs = 1.0 and baseline resolution between two peak requires a Rs > 1.5.

Instrument:

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Gas chromatography (Agilent Technologies 6890N) equipped with flame ionization detector
(FID) and 30 m x 250 µm x HP5-MS capillary column.

Sample:

1. Individual methyl ester compound: methyl laurate, methyl myristate, methyl

palmitate, methyl stearate, methyl linoleate.
2. Standard mixture 1 (consist of first 3 individual standard) > for Isothermal method.

Procedure:

A. The instrument set up:

-Injection port – split (40:1)

-Injection port temperature – 250ºC

-Column temperature – 210ºC

-Carrier gas – 30 cm sec-1

-Detector temperature - 250ºC

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

1. 0.4 µL standard mixture was injected isothermally at 210ºC with carrier gas flow rate
of 30 cm sec-1.
2. The flow rate was then increased to 50 cm sec-1. The system was allowed to
equilibrate for a few minutes before injecting the standard again.
3. The procedure was repeated with flow rate at 70 cm sec -1.
4. The optimum flow rate was chosen to continue with the next steps.

C. Effect of column temperature on isothermal GC separation of methyl ester.

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1. 0.4 µL of standard mixture was injected isothermally at 170ºC, followed by 190ºC at

optimum carrier gas flow rate.
2. The effect of column temperature on separation, resolution and analysis time were
evaluated.

D. Identification of component in methyl ester mixture.

1. Each of the methyl ester were injected individually to identify the various compounds
in the standard mixture using optimized GC condition (optimum gas flow rate and
optimum column temperature)

Result and calculation:

Resolution calculation:

2(tr 2−tr 1)
Rs = (Wb1+Wb 2)

tr2 = Retention time of peak 3.

tr1 = Retention time of peak 2.

Wb1 = Peak Width of peak 2.

Wb2 = Peak Width of peak 3

Average resolution:

(Rs 1+ Rs 2)
Avg Rs =
2

Calculation of resolution is based on peak 2 and 3 as references.

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A. Effect of the variation of gas flow rate on the resolution.

Condition Injection Retention Peak Width Resolution, Average

time of peak of peak 2 Rs Resolution,
2 and 3, and 3 Rs
(min) , (min)
30 m/s 1 4.570, 6.491 0.0467, 34.77
210ºC 0.0638 35.07
2 4.568, 6.488 0.0450, 35.36
0.0636
50 m/s 1 2.740, 3.899 0.0351, 30.10
210ºC 0.0419 31.88
2 2.738, 3.896 0.0304, 33.66
0.0384
70 m/s 1 1.956, 2.789 0.0288, 25.09
210ºC 0.0376 25.31
2 1.959, 2.791 0.0314, 25.52
0.0338

From table A. Effect of the variation of gas flow rate on the resolution, the optimized
separation of methyl ester was at 70 m/s of gas flow rate.

B. Effect of the variation of column temperature at optimized gas flow rate on the
resolution.

Condition Injection Retention Peak Width Resolution, Average

time of peak of peak 2 Rs Resolution,

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2 and 3, and 3, Rs
(min) (min)
70 m/s 1 3.644, 7.127 0.0390, 61.81
170ºC 0.0737 60.87
2 3.631, 7.116 0.0445, 59.93
0.0718
70 m/s 1 2.578, 4.279 0.0333, 38.18
190ºC 0.0558 36.75
2 2.579, 4.281 0.0356, 35.31
0.0608
70 m/s 1 1.956, 2.789 0.0288, 25.09
210ºC 0.0376 25.31
2 1.959, 2.791 0.0314, 25.52
0.0338

The optimized column temperature at 70 m/s of gas flow rate was at 210ºC column
temperature because it produced the resolution nearest to the ideal resolution value that is 1.5
and also shorter analysis time.

C. Retention time of standard compound of the methyl ester at optimum condition.

Standard Compound Retention time (min)

Methyl laurate 1.916
Methyl myristate 2.692
Methyl palmitate 4.289
Methyl linoleate 6.809
Methyl stearate 7.495

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Discussion:

Different flow rate of the mobile phase will affect the retention time of the compound.
The higher the flow rate of the carrier gas, it will elute the analyte faster (in shorter time) but
will cause band broadening due to mass transfer (C-term) of Van Deemter plot, where the
solute in the column are not completely interact and equilibrate. So, to reduce the band
broadening or the overlapping peak, the optimum gas flow rate must be used to achieved
better separation.

From the experiment results, the optimum gas flow rate achieved was at 70 m/s which
was at good resolution, 25.31 at average Resolution near to the ideal resolution. Other than
70m/s gas flow rate, the other flow rate has not achieved a good separation where there are
some peak overlapping and band broadening.

Besides, the column temperature also needs to be determined its optimum column
temperature for getting the good separation. Same with flow rate, the column temperature
with too high temperature will allow the analyte to elute fast but the volatile compound’s
peaks will overlapped and not well separated and vice versa with applying low column
temperature. The high column temperature will also lead to column bleeding.

From the experiment results, the optimum temperature achieved was at 210ºC. So, to
conclude, the Optimum GC condition to be well separated of the standard methyl ester was at
70 m/s with 210ºC.

The column used in the GC was HP-5 capillary column. This column consists of 5%
of phenyl group and can be assumed slightly less non-polar because in order to separate the
standard methyl ester (which is known as slightly less non -polar compound). This stationary
phase will retain the mixture longer in the column and separate at their retention time. The
most non-polar analyte will less retain in the column thus will make it elute first and vice
versa with the slightly less non-polar compounds.

From the standard mixture of methyl ester, there are three type of individual standard
which were methyl laurate, methyl myristate and methyl palmitate. These three standards
were eluted at 1.956min, 2.789min, and 4.486min respectively. These three standards’

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retention time were closed to the individual standard that have been analyse as reference to
the standard mixture methyl analyte. The first compound eluted was methyl laurate followed
by methyl myristate and methyl palmitate. These sequences of elution were related to the
polarity of the compounds. The methyl palmitate was the most retained compound in the
column and more less non-polar compared to methyl laurate and methyl myristate. Besides,
the GC also depend on the column temperature to separate. Methyl palmitate was less volatile
which have high boiling point than the other two compounds.

Conclusion:

From the results of the experiment, we can conclude that the optimized GC condition
for the separation of methyl ester were at 70m/s for the optimum gas flow rate and 210ºC for
the optimum column temperature. The first peak eluted in the column was identified as
methyl laurate followed by methyl myristate and methyl palmitate respectively. To conclude,
the experiment went well, and we had learned on the concept of the gas chromatography, the
correct way on setup and handling the GC instrument, learn the concepts of retention time
and resolution of the peak base on a mixture of the standard been provided, learned the effect
of column temperature and flow rate on the separation compounds and lastly been able to
identify the optimum temperature and optimum flow rate of the mixture standard.

References:

1. John.S,(2001). Principle of Instrumental Analysis, page 200-204: Gas

Chromatography.
2. Saim, N., Tajuddin, R., & Saaid, M. (2014). Experiment 1 GC – Gas
Chromatography: Optimum Flow Rate and Optimum Temperature. In
ANALYTICAL SEPARATION METHODS LABORATORY GUIDE 2nd edition
(pp. 1-2). Selangor: UiTM Press.
3. Gas Chromatography (GC) retrieved on 4 May 2019 from
https://en.wikipedia.org/wiki/Gas_chromatography

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Result Data:

Temp 210ºC, Flow rate 70 (1st injection)

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Temp 210ºC, Flow rate 70 (2nd injection)

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Temp 210ºC, Flow rate 50 (1st injection)

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Temp 210ºC, Flow rate 50 (2nd injection)

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Temp 210ºC, Flow rate 30 (1st injection)

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Temp 210ºC, Flow rate 30 (2nd injection)

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Temp 190ºC, Flow rate 70 (1st injection)

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Temp 190ºC, Flow rate 70 (2nd injection)

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Temp 170ºC, Flow rate 70 (1st injection)

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Temp 170ºC, Flow rate 70 (2nd injection)

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Individual Standard – Methyl Laurate

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Individual Standard – Methyl Myristate

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Individual Standard – Methyl Palmitate

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Individual Standard – Methyl Linoleate

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Individual Standard – Methyl Stearate

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