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Experiment 1: Ion Chromatography and Capillary Electrophoresis
Introduction:
The purpose of this lab is to become familiar with both the IC (Ion Chromatographer) and the CE
(Capillary Electrophoresis). The IC is an instrument used to separate anions from a liquid substance
through liquid chromatography. The CE separates ions based on their mobility in an electric field
through electroosmotic flow. This mobility is determined by charge-to-mass ratio and the additives
present in background electrolytes.
Procedure:
For Ion Chromatography:
A 50 ppm standard (based from 100ppm anion standard solution) was prepared by dispensing 7mL of
seven anion standard solution into a 100 mL beaker. 5mL of the sample was gathered and added to 5mL
distilled water. Along with this, three water samples from the Campbell Hall building were obtained.
These samples and the standard were tested by the IC using this SOP.
For Capillary Electrophoresis:
Five 1.5mL vials were obtained and filled with Buffer A (x2), Test Mixture B (mixture of p-hydroxybenzoic
acid and p-hydroxyphenylacetic acid), distilled water, and 1.0M HCl. Other various vials were also
collected and placed on a sample try according to the instruments SOP.
Data:
Data was collected using Ion Chromatography and Capillary Electrophoresis. Results are displayed here.
Results for Capillary Electrophoresis:
Time(minutes)
Area %
Height %
p-hydroxybenzoic acid
5.179
36.81
27.90
p-hydroxyphenylacetic
acid
5.529
62.22
71.43
Results for Ion Chromatography:
Sample
Standard
Lab Water
Bathroom Sink Water
Water Fountain
Fluoride
(ppm)
Chloride
(ppm)
20.000
47.298
64.617
88.8097
30.000
615.761
526.701
676.635
Nitrite
(ppm)
100.000
n.a.
42.418
n.a.
Bromide
(ppm)
100.000
n.a.
56.782
52.200
Nitrate
(ppm)
100.000
426.530
435.935
496.954
Phosphate
(ppm)
150.000
n.a.
203.948
71.517
Sulfate
(ppm)
150.000
433.134
552.581
441.088
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Experiment 1: Ion Chromatography and Capillary Electrophoresis
Discussion:
The results obtained from the IC were not even remotely close to that of the York Water
Company’s data. Most of our levels were around 23x higher than that of the York Water Company. This
error is mostly due in part by the replacement of the regenerate solution. Previously, we had analyzed
our sample about 7-8 times without any valid data. Because of this, we found that the regenerate
solution needed replaced in order to successfully run the experiment. However, because it was a new
solution, the machine needed a much longer time in order to flush the old regenerate out of its system.
Because we did not allocate enough time the proper calibration, our results were completely off, yet
precise. It has been included in our new SOP that when replacing the regenerate, to give the machine
around 2 hours of calibrating time in order to successfully run.
The results obtained from the CE indicate that the 4-hydroxybenzoic acid was the first to elute
at a time of 5.179 minutes, whereas the 4-hydroxyphenylacetic acid migrated at a time of 5.529
minutes. According to the graph, the 4-hydroxyphenylacetic acid had the higher of the two peaks. This
data is significant in showing that the 4-hydroxybenzoic acid has a lower charge-to-mass ratio, thus
giving the answer to why it eluted first. This also allows us to hypothesize that smaller compounds will
also migrate first from the electric field created.
Conclusion:
As a result of the IC data, the lab is considered to be a failure. Because the regenerate was
replaced, and peaks from each ion were off, we were unable to determine whether or not any other
anions were present in the samples. When looking at the data, there were unlabeled peaks that could
not be classified, due to the hardness (ppm) being off. Though the values for the anions were at an
extreme value compared to the York Water Companies data, the peaks from each water sample were
relatively precise. That being said, if the data from the seven-anion standard were accurate, there
would be a high chance that the sample data would have also been accurate and precise.
Looking at the data obtained from the CE, this portion of the lab was deemed to be a success.
Following the principles of particle migration, the particles with the smallest size and greatest charge are
the first to elute. This was the case in our experiment, as the 4-hydroxybenzoic acid (Molecular Mass =
138.12 g/mol) eluted before 4-hydroxyphenylacetic acid (Molecular Mass = 152.15 g/mol). This data is a
result of electroosmetic flow within the CE. Electroosmotic flow is where the walls of the capillary tube,
consisted of SiOH groups, lose a proton and become SiO- ions. Because of this, the wall obtains a
negative charge, attracting a double layer of cations. The inner layer of cations is stationary, but the
other layer is free to move along the capillary. Because of the electric field, these free-moving cations
begin to migrate toward the cathode, creating electroosmotic flow. In our experiment, the electric field
was produced at a charge of 25 kV. If we were to reverse this charge, the electroosmotic flow would
migrate in the opposite direction, towards the anode.