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Final Research Paper Ciaramitaro Doerr Weidemann
Final Research Paper Ciaramitaro Doerr Weidemann
Introduction………………………………………………………………………...1
Review of Literature……………………………………………………………….3
Problem Statement………………………………………………………………..9
Experimental Design……………………………………………………………...10
Conclusion……………………………………………………………………….…29
Works Cited…………………………………………………………………………34
Introduction:
Ciaramitaro-Doerr-A.Weidemann
Before the early 20th century, there were limited ways to separate and
and identified. This remained true until 1903, when Mikhail Tswett separated the
the retention factors of each chromatogram. This was done by allocating highs
and lows to the factors, polarity and porosity. Once all the factors were
established, the trials were conducted and observed to calculate the Rf value of
each chromatogram. These Rf values were then averaged with its corresponding
experiment was used to help the collection of data in an organized fashion and
The research was conducted to test the use of different solvent polarities
and porosity on the effect of the developed chromatogram to help indicate the
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because the less of the new medium is needed. Other applications may be the
improved processes can be used, and materials will not be wasted trying to find
Review Of Literature:
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Heterogeneous mixtures. This image was found on the article Chemistry for Kids
the early 20th century. Founded by Russian botanist Mikhail Tswett in 1903,
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Today, it is one of the most widely used techniques for many analytical and
mixture. It is relatively easy to use because it only needs a small amount of the
involve a liquid solvent carrying a stationary solvent, which is the mixture that is
being analyzed, with it. Although this method slightly changes with each type of
Academy”). The ratio of the distance travelled between the two is what is known
look at the retention factors (Rf values for short) of certain substances. Every
compound has its own retention factor for the solvent used. This value is
solute and the distance travelled by the solvent front, the endpoint of the
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Ciaramitaro-Doerr-A.Weidemann
𝑅𝑒𝑡𝑒𝑛𝑡𝑖𝑜𝑛 𝐹𝑎𝑐𝑡𝑜𝑟
Since these values are ratios, they will always be between 1 and 0 because the
solvent front will always travel farther. These calculations are very important
when identifying mixtures and the compounds that make them up.
front travelled 2.8 cm from the origin while the compound front only travelled 2.1.
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substances. Notice that the dyes are separated because they traveled different
distances up the paper. The solvent front carried each component up the paper
before the components stopped moving and the solvent front carried on. This
Compound B in the figure above will have a larger retention factor because it
polar, its charges are not properly distributed. Polarity will have an impact on how
far the solution moves up the paper. Non-polar molecules in the mixture that are
trying to separate will have little attraction towards water molecules attached to
the cellulose on the paper sheet, and will dissolve in the solvent. These
molecules will therefore travel a greater distance upwards on the paper carried
by the solvent and, they will have relatively high retention factor. Polar molecules
are quite different and will have a high attraction towards the water
therefore tend to dissolve in the thin layer of water around the cellulose fibers
much more than in the agitating solvent. They spend more time dissolved in the
stationary phase and less time in the mobile phase ,so they aren't going to travel
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This paper comes in many shapes and grades. This experiment will use strips of
instead of sheets or circles. Each grade of paper is slightly different from the
next. Grade 1 qualitative paper has a paper like feel and is smooth. Grade 3 is
rough and thicker. As the grade of paper increases, the paper gets more porous.
High porosity paper will have more small pores which allows the solvent and the
compound to grip better. The two fronts will travel farther if the paper has more
pores.
System, they are able to analyze more samples and identify more fatty acids at a
Acids”). The experiment that will be conducted is similar to this. It will use paper
the type of chromatography used, and the substance that will be identified.
chromatography, very similar to the method used in the current experiment, but
than chemical compounds which would allow the interpolation of data. These
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The experiment that will be explained in this paper will use similar
methods to the two labs described above. This experiment will use paper
different dyes in a solution, and it will talk about the concepts behind the
individual retention factors and any trends or effects that show up. The
experiment will also focus more upon why the solvents and qualitative paper
real world applications. This lab is designed to show how useful it can be in
identifying parts in a mixtures that could not have been separated even over one
Problem Statement:
What effect will different combinations of polarity of the solvent and type of
Hypothesis:
The highest polarity solvent, water, and the most porous filter paper, grade
3, will produce the highest accuracy retention factor value results because the
high polarity and the great porosity will allow the dye solution to move faster and
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more efficiently along the filter paper and create more observable/accurate
results.
Data Measured:
The independent variable used was the specific FD&C food dyes which
will be Red 40, while the dependent variable measured was the Rf values of the
FD&C Red 40 dye. The Rf values can be defined as the distance ratio in
millimeters of the ionized solvent, the FD&C food dyes, compared to the distance
of the solutions across the selected chromatogram. The Rf value will be analyzed
using a two factor D.O.E. method. The two factors include the type of filter paper,
and the polarity of the solvent. The D.O.E. will determine which level of each
Experimental Design:
Materials:
(0.5 g) FD&C Red 40 (7) Beaker, 600-mL
Water, Distilled or Deionized, 3.8-L (2) Graduated Cylinder, 25-mL
(36) Grade 1 Qualitative Paper (1) Graduated Cylinder, 100-mL
(54) Grade 2 Qualitative Paper (2) Ruler
(36) Grade 3 Qualitative Paper (3) Capillary Tube
(7 mL) Ethyl Alcohol (95%), C2H6O (10 mL) Isopropyl Alcohol (70%), C3H7OH
(2) Scoopula
Procedure:
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Pre-Lab:
Lab:
4. Using a ruler and a pencil, draw a faint line 15 mm from the bottom of the
paper across the width of the strip. Measure 9.5 mm from the edge and
place a dot on the line. This is the starting point for the sample. See figure
1.
5. Using the same ruler, measure 21 mm from the top of the strip and fold
across the width of the strip. This will allow the strip to hang on the lip of
the flask. See figure 1.
6. Repeat steps 3 and 4 for each strip of each grade of filter paper
7. Using the capillary tube, spot the chromatography strip by placing it into
the dye mixture solution then touching the tip of the tube gently onto the
designated pencil dot. Allow the sample to dry. Note: This step is
necessary to increase the concentration of the sample but do not allow the
size of the spot to increase.
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9. After the dye has completely dried (1 min), carefully hang 3 of the
corresponding chromatography strips into the beaker with the dye dot end
down. Do not get any solvent on the upper portion of the strip. The sample
spots must remain above the level of the solvent. If the solvent level is too
high, the samples will dilute into the solvent. (Try not to touch the strip to
the edge of the beaker when it is hanging). See figure 2.
10. Allow the the chromatogram to develop for 25 minutes. Record the
observations of the dye sample as the solvent travels up the paper and
the chromatogram develops. See figure 3.
11. Repeat steps 8 and 9 using the other chromatographic solvents for each
trial.
12. After the 25 minutes is up, carefully take out the paper strips.
13. Immediately, with a pencil, lightly draw a line to mark the distance
travelled by the solvent. This is the solvent front. Draw a line marking the
furthest the compound traveled. This is the compound front. (This will be
measured in millimeters).
15. Record results into a data table that calculates the Rf Value for each
chromatogram. (Compound Front / Solvent Front).
Diagrams:
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Figure 4. Materials
The materials used in the experiment are shown above. The deionized
water was used as one of the solvents and also to mix the other solvents. The
other solvents consisted of the isopropyl alcohol and the ethyl alcohol. The
deionized water was also used to mix the Red 40 FD&C Food Dye. The test
tubes were used to measure out the solvents that were then placed into the
600mL beakers. Rulers were used to measure the three different grades of
qualitative paper. The capillary tubes were used to dot the paper with the food
dye. The 100 mL graduated was used primarily for mixing solutions.
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The above figures shows the line that was drawn 15mm from the bottom.
This line was then used as a reference to place the two dots of Red 40 FD&C
Dye. This figure also depicts the folds that were done 21 mm from the top of the
strip. These folds were tasked at 21 mm because it was a perfect balance for the
strips not to touch the side of the beaker and to have the perfect height into the
solvent.
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soaking in the solvent. This picture was taken a 25 minutes after being placed
into the beaker. Notice each dye is relatively at the same distance. A trend in
Grade 3 strips, depicted in this figure, is that the retention factor is higher than
the others. This figure supports this claim as the compound front is close to the
solvent front.
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chromatogram right after it is removed from the beaker. The marks were quickly
placed to prevent incorrect data due to development after taking out the paper.
The dashes were then measured and taken as a ratio. This was labeled as the Rf
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Table 1
Compound Front Measurements
+,+ +,- -,+ -,- Stan 1 Stan 2 Stan 3
April DOE
28th 1 101 101 85 73 81 88 88
DOE
2 110 100 86 71 87 89 89
DOE
3 114 103 87 79 84 87 95
May DOE
1st 4 101 97 75 88 83 87 80
DOE
5 109 92 78 76 88 82 80
DOE
6 108 103 74 82 86 116 92
May DOE
3rd 7 100 99 94 81 79 75 81
DOE
8 100 94 91 82 80 82 79
DOE
9 99 100 86 76 86 76 84
Table 1 shows the gathered compound fronts from the trials performed.
Table 2
Solvent Front Measurements
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April DOE 1
103 102 88 77 99 90 88
28th
DOE 2
112 102 88 74 89 91 118
DOE 3
117 105 90 83 85 89 97
May DOE 4
103 99 84 97 85 90 85
1st
DOE 5
111 95 83 97 90 86 85
DOE 6
110 105 81 89 87 125 95
May DOE 7
103 101 96 84 80 84 81
3rd
DOE 8
102 97 93 84 82 84 81
DOE 9
102 102 97 79 87 77 85
Table 2 shows the gathered solvent fronts from the trials performed. The
Table 3
Rf Value Calculations
+,+ +,- -,+ -,- Stan 1 Stan 2 Stan 3
April DOE 1
28th 0.9806 0.9902 0.9659 0.9481 0.8605 0.9778 1.0000
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DOE 2
0.9821 0.9804 0.9773 0.9595 0.9775 0.9780 0.7542
DOE 3
0.9744 0.9810 0.9667 0.9518 0.9882 0.9775 0.9794
May DOE 4
1st 0.9806 0.9798 0.8929 0.9072 0.7953 0.9667 0.9412
DOE 5
0.9820 0.9684 0.9398 0.7835 0.9778 0.9535 N/A
DOE 6
0.9818 0.9810 0.9136 0.9213 0.9885 0.9280 0.9684
May DOE 7
3rd 0.9709 0.9802 0.9792 0.9643 N/A 0.8929 1.0000
DOE 8
0.9804 0.9691 0.9785 0.9762 0.9756 N/A 0.9753
DOE 9
0.9706 0.9804 0.8866 0.9620 0.9885 0.9870 0.9882
May Make-
4th Up 0.8182
0.9765
0.9875 0.9762 0.9412
Average
0.9781 0.9789 0.9445 0.9304 0.9643 0.9597 0.9498
Table 3 displays every Rf value calculated. All of the data points with a
line through them were corrupted data and redone on May 4th.
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Table 4
Observations
Observations
April DOE 1 - A standard 1 trial was removed from the usable data
28th due to it falling off the beaker edge
- The (+,+) dots were unevenly placed, but should not
have an inconsequential impact
- Other trials were normal
DOE 3 - The (+,+) dots were unevenly placed, but should not
have an inconsequential impact
- Other trials were normal
May 1st DOE 4 - A standard 1 trial was removed from the usable data
due to it falling off the beaker edge
- Other trials were normal
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May 3rd DOE 7 - A standard 1 trial was removed from the usable data
due to it falling off the beaker edge
- Other trials were normal
May 4th Make-Ups - All previous corrupt trials were repeated with success
the standards were corrupted and had to be redone as a result. Notice it says
that most trials were normal. This means that the chromatograms were
successfully completed and had no contact with the beaker wall. These trials
were not contaminated in any way. The trials that were contaminated were
redone later.
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Table 5
Table of Factors
Factors (+) Values Standards (-) Values
Table above shows the different factors used in the experiment. Porosity
and Polarity each had a high and a low level used in the D.O.E.
Table 6
Average Rf Value For Each Factor
Porosity Porosity
(+) (-)
The table above shows the average retention factor for each level of the
D.O.E. Notice that the level with the highest average retention factor was the
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Table 7
Rf Values Of All 27 Standards
27 Standards
Standard 0.8182 0.9775 0.9882 0.9765 0.9778 0.9885 0.9875 0.9756 0.9885
1
Standard 0.9778 0.9780 0.9775 0.9667 0.9535 0.9280 0.8929 0.9762 0.9870
2
Standard 1.0000 0.7542 0.9794 0.9412 0.9412 0.9684 1.0000 0.9753 0.9882
3
calculated by subtracting the highest Rf value from the lowest. This number was
calculated to be 0.2458. This number is relatively high due to some outliers. The
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Rf value for each standard is relatively constant except for a few outliers.
Standard 20, is extremely low with a Rf value of 0.7542. Another standard that
seems a bit low is the first standard with a Rf value of 0.8182. These low
numbers could have occurred for a multitude of different reasons. The most
plausible reason for this could be that the paper touched the side of the beaker
during the trial resulting in a lower Rf value. Standard 20 could significantly play a
role in whether the effects will be statistically significant or not by making the
Table 8
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Grade 1 Grade 3
(-) (+)
0.9304 0.9781
0.9445 0.9789
Table 8 above, shows the average data for the corresponding factor,
porosity of the paper. The averages for the high and low of porosity were taken
and then subtracted from each other to calculate the effect value. This number
will be used later on to determine if the porosity of the paper was significant to
Figure 9 above, shows low and high averages on the effect of paper
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0.0410. As previously stated, this number was calculated by subtracting the high
Table 9
Rf Averages of Solvent Polarity
Polarity Of the Solvent
Isopropyl Water
Alcohol (+)
(-)
0.9304 0.9781
0.9789 0.9445
Table 9 shows the average data for the corresponding factor, polarity of
the solvent. The averages for the high and low of porosity were taken and then
subtracted from each other to calculate the effect value. This number will be used
later on to determine if the polarity of the solvent was significant to the results
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Figure 10 above, shows low and high averages on the effect of solvent
0.0066. As previously stated, this number was calculated by subtracting the high
Table 10
Interaction Effect of Porosity and Polarity
Porosity
- +
- 0.9304 0.9789
Table 10 above, shows both the corresponding Rf values with each factor
in this experiment.
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Figure 11 ,above, shows the interaction effect between the porosity of the
paper used and the polarity of the solvent used. Judging from the graph, there
intersect. Another graph was also created to represent this data but it served no
The above figure shows how the interaction effect was calculated. This
number was calculated by finding the slopes of both lines and subtracting the low
slope from the high slope. The high slope was calculated to be 0.0168. This was
done by dividing the difference in y’s over the difference of x’s. The low slope
was calculated using the same method. This was calculated to be 0.0243. The
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Figure 13, above, displays all the effect values on a line. The boundaries
shown, depict whether these effects are statistically significant. These fences
shown above, none of the effects were statistically significant as they all fell
inside of the fences. Porosity had the most effect on the Rf values as it had a
The above figure shows the prediction equation for this experiment. Y was
calculated to be 0.9781. This means that if this experiment were to be ran again,
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Y = 0.9580 + “noise”
Figure 15. Parsimonious Prediction Equation
The above figure shows the parsimonious prediction equation. All of the
Conclusion:
The objective of this experiment was to find the effect of solvent polarity
and the porosity of qualitative paper had on the resulting retention factor in paper
chromatographic fields of science. Using the highest polarity solvent and the
qualitative paper, was expected to yield the highest and most significant retention
factor results, but was concluded to be rejected as the interaction between the
hypothesis was also denied because the highest grade of paper and the lowest
The effect value of the porosity of the qualitative paper, 0.0410, the
polarity of the solvent, 0.0066, and their interaction, -0.0075, were all found to be
statistically insignificant because all of the effect values laid inside of the
very high probability that the effects occurred purely by chance alone.
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Ciaramitaro-Doerr-A.Weidemann
The effect value of the porosity was expected to yield the highest effect
value because of the paper used. The type of paper used include Grade 1, 2,
and 3 Whatman Qualitative paper. Grade 3 is significantly more porous and rigid
than the other grades, which makes it more absorbent. When the porosity of the
paper is increased, the solvent has more holes to latch on to therefore resulting
in a speedier climb. The data collected supports the fact that the grade 3
and compound front. Grade 1 Whatman paper is very weak at absorbing solvent
Likewise with the porosity of paper, the polarity of the solvents will also
greatly affect the retention factor value. Non-polar molecules will tend to have
little attraction towards the cellulose of the qualitative paper and as a result, lose
its speed which causes the solvent front to retain its distance with the compound
front. The reasoning of this has to do with the adhesion of the solvent. Water has
very strong negative and positive molecular ends. As result, water likes to bond
to itself, also known as cohesiveness. When decreasing the polarity, the solvent
loses its “stickiness” and doesn’t adhere to the paper as well. These findings do
not support the proposed hypothesis, but do support the understanding of why
the highest porosity and lowest polarity had the highest retention factor.
chromatograms came into contact with the beaker wall. This caused the solvent
front to rush up the paper leaving the compound front behind. The
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Ciaramitaro-Doerr-A.Weidemann
chromatograms that came into contact with the wall were not included in the data
because they are outliers. These trials were repeated later in the week. Another
oversight that may have altered the results would be the amount of dye placed
on the strip of qualitative paper. A capillary tube was used to place two to three
drops of dye on the paper. The capillary tube was very inconsistent at measuring
this research could be tested to see if it will impact the efficiency in similar
chromatography such as its use in the medical field for the identification of
material and should be seen as a point of study for the future of every scientific
field.
Appendix A
Pre-Lab:
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1. To prepare the 100 mL of dye solution, add 0.5 g of the FD&C red 40 solid
dye to a separate beaker with 100mL of distilled or deionized water. Mix
thoroughly.
2. To cut the 150mm by 20mm paper strips out of the 150mm paper circles,
first take a properly cut grade 3 strip and place it ontop of the circle so that
it fits inside of it. Once in position, trace the paper strip onto the circle of
choice. Carefully cut out the paper strip using the guidelines that were
drawn. Repeat this until the sufficient amount of paper strips have been
acquired.
are the distance travelled by the solvent front, and the distance travelled by the
compound front. The equation for calculating the retention factor is shown below.
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Ciaramitaro-Doerr-A.Weidemann
=0.9567
Works Cited
"Chemistry for Kids." Ducksters Educational Site. N.p., n.d. Web. 19 Mar. 2017
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Piety, Nathaniel Z., Xiaoxi Yang, Julie Kanter, Seth M. Vignes, Alex George, and
Sergey S. Shevkoplyas. "Validation of a Low-Cost Paper-Based Screening
Test for Sickle Cell Anemia." PLOS ONE. Public Library of Science, 6 Jan.
2016. Web. 14 Mar. 2017.
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