Experiment 8: Gas Chromatography (GC)

In this experiment, mixtures of volatile organic compounds will be separated and

analyzed, and a sample containing an unknown percentage of ethanol will be quantitated by GC
analysis.

In GC, a liquid sample is injected into a separation column as sharp plug. The GC
column is usually coated with stationary phase of a given overall polarity. The column is
contained within an oven. The sample is vaporized to a degree and at a rate dependent on its
boiling point. It is carried by an inert gas (usually helium) through the column to a detector. The
detector signals a chart recorder, which records the response, ideally in the shape of a
Gaussian peak.

Two significant factors determine the rate at which an injected compound travels through
the column and reaches the detector. A compounds volatility at the column temperature
influences the distribution of the compound between the gaseous mobile phase and the
stationary phase. Other things being equal, a compound band will move through the column
more quickly if its distribution favors the mobile phase. Boiling points are often used as a
measure of the relative volatilities of compounds in a mixture. However, volatility alone does not
determine the distribution between the stationary and mobile phase. Specific interactions
between the injected compound and the stationary phase play an important role as well. Recall
that like dissolves like so a polar compound will tend to strongly distribute into a polar
stationary phase. Conversely, a polar compound will not have a strong affinity for a non-polar
stationary phase and, thus, will elute relatively quickly. In summary, components in a mixture
can be identified by analyzing the difference in their retention times, which is dependent upon
their volatility and polarity.

Quantitation is possible in GC methods by analyzing the peak area or peak height. For
the detector you will use in this experiment, peak area will provide better results. A larger peak
area indicates a larger amount of analyte present in the sample. Spiking samples with internal
standards helps to compensate for the imprecision inherent to GC methods and for variations in
sensitivity among different analytes. In this method, the analytical sample and standards are
spiked with an fixed amount of a solute whose retention time is similar to (but resolved from)
that of the sample. The ratio of the area of the analyte peak to that of the internal standard is
used to prepare a calibration curve and subsequently used to determine the analyte
concentration in an unknown mixture.

In this experiment, you will use a Gow-Mac GC equipped with two columns. Column A
is a polar column with a tradename of Carbowax 20M and column B is a non-polar column with
a tradename of DC 200. Only one column may be used at a time. The direction that the chart
recorder pen will deflect depends upon which column is in use; make certain that you change
the polarity of the pen direction whenever you change columns.

Several factors affect separation efficiency in GC, including column temperature and
carrier gas flow rate. The temperature for the GC oven has been preset for you; record the
temperature in your lab notebook. The carrier gas flow rate is adjustable for both columns and,
thus, the flow rate will not be the same for both columns. You will use a soap bubble flowmeter
to measure the flow rate of the carrier gas.

1
 Before starting, the detector and chart recorder must be zeroed. Set the Attenuation to
infinity and position the chart recorder pen to the desired baseline position using the recorder
Zero offset. Slowly turn the Attenuation control towards 1. If the chart recorder pens position
moves from the baseline setting, use the Zero control on the GC to reset the pen to its original
position. Once you are able to change the attenuation from infinity to 1 without significantly
changing the pens position, the instrument is zeroed. Begin your experiment by setting the
attenuation on 4 and the chart recorder sensitivity to 20 mV. Adjusting these parameters will
change the peak size on your chart paper. Setting the recorders sensitivity to a higher setting
will cause the peaks to become smaller; increasing the attenuation to a larger number will cause
the peaks to become dramatically smaller. Set the chart recorder speed to 5 cm/min. The TA
will demonstrate the proper use of the GC injection microsyringes. Treat these expensive
devices with care! Whenever a sample is injected into the GC, it is important to mark the chart
paper at the moment of injection. Also, you should inject as much air as possible with the
sample in order to obtain an air peak (this allows determination of adjusted retention times).

The following chart lists some important properties of the classes of compounds used in
this experiment:

Compound Polarity Boiling Point

alcohols polar boiling point increases with

molecular weight

ketones polar lower boiling point than alcohols of

comparable molecular weight

alkanes non-polar boiling point is lower than ketones of

comparable molecular weight

alkyl benzenes non-polar high degree of double bonds increases

stability, thus, relatively high boiling points

Procedure: Sample Prep

1. Each group needs one of each of the following solutions:
Mix A: equal amount of straight chain alcohols from C2-C6
Mix B: equal amounts of n-heptane, 2-butanone, 1-butanol and o-xylene
pure n-heptane
pure 2-butanone
pure 1-butanol
pure o-xylene

Each student must obtain an individual unknown.

2
2. Prepare the following standards in a 10-mL volumetric flask. Fill the flasks to the mark with
water after adding the ethanol and n-butanol. (volumes in mL):

% ethanol (v/v) vol. ethanol vol. n-butanol

20% 2.00 1.00
40% 4.00 1.00
60% 6.00 1.00
80% 8.00 1.00

3. Pipette 5 mL of your unknown into a 10-mL volumetric flask. Add 1 mL n-butanol and fill to
the mark with water.

Procedure: Chromatography
For each set of chromatograms, record the following:
solution injected
recorder sensitivity (mV)
flow rate (mL/min)
chart speed
injection volume (L)
column temperature (oC)
type of column
GC attenuation

4. Inject 1 L of the alcohol mixture onto column A. Inject as much air as possible with the
sample. Adjust parameters as necessary until retention times for each peak can be analyzed.

5. Individually analyze each compound in mix B on each column by injecting 2 L of each pure
substance. REMEMBER: you cannot operate both columns simultaneously.

6. Inject 1 L of mix B onto each column.

7. Inject 1 L of the ethanol standards onto column A. Adjust parameters so that the peak
areas can be easily measured.

8. Inject 1 L of the unknown onto column A; repeat 3-4 times.

Report:
You will write formal report for this laboratory. For your unknown report, provide your name,
your unknown number, the percent ethanol in your unknown sample, the standard deviation.
Although this experiment was performed in groups, each student must write his/her own
report.

1. Title page: Include name, title, date, unknown number, and lab partners.

2. Introduction: Discuss principles of GC and the purpose of this lab.

3
3. Experimental: Briefly describe solution preparations and experimental procedures.

4. Data: Neatly organize relevant data into tables. Include GC parameters, elution order and
retention times. Do not include your chromatograms in this report.

5. Results and Discussion. Include the following points within your discussion:

5a) Explain the elution order observed with mix A.

5b) Plot log adjusted retention time vs. carbon number.
5c) Plot log adjusted retention time vs. boiling point.
5d) Comment on these plots.
5e) Estimate the adjusted retention time for CH3(CH2)9OH. Explain how you did this.
5f) Explain the elution order for mix B on the two columns.
5g) Calculate the resolution of 2-butanone and 1-butanol on the two columns. Which
column yields better resolution? Why?
5h) Calculate the efficiency of 1 butanol on the two columns. Which column has better
efficiency? Why?
NOTE: equations are given on next page.
5i) Prepare a calibration plot (peak area ethanol)/(peak area butanol) vs. % ethanol.
Remember, do a best-fit line for a calibration plot; do not force line through the origin.
5j) Use your calibration plot to determine the % ethanol in your original unknown sample
(NOT your diluted aliquot).

6. Conclusions. Discuss sources of errors and how they could affect your results, along with
the conclusions drawn from this experiment. Within this section, answer the following
questions:
6a) How could you have improved the resolution achieved?
6b) What changes would you predict in retention time and peak shape if the following
occurs:
The column temperature is increased between injections?
The carrier gas flow rate is decreased between injections?
6c) Explain the effect of each of the following three processes on band width.
Eddy diffusion
Longitudinal diffusion
Resistance to mass transfer

7. References

Useful equations:

Resolution (Rs) = 2(t2 - t1)/(W1 + W2)

where t1 = retention time of one component
t2 = retention time of second component
W1 = width of peak at baseline of first component
W2 = width of peak at baseline of second component

Efficiency (n) = 5.54 (tr/W1/2)2

where tr = retention time
W1/2 = width of peak at half-height