CHML211
Exp. 4 TLC and HPLC Analysis of Nitroanilines
IN-LAB ASSIGNMENT: (EACH STUDENT will record the following data in his/her laboratory notebook in a well organized
manner during lab period. Yellow carbon copies will be submitted for grading along with Post-Lab assignment.) …..max
score = 20 pts.
6. Data/Calculations (The following information should be recorded directly into laboratory notebook.)
a.
b.
c.
Names of TLC developing solvent systems (3)
Sketch of all TLC plates. Each spot detected should be labeled with the appropriate cm measurement, as well as
the cm measurement for the solvent front on each plate
All TLC Rf calculations
7. In-lab Questions (The following questions should be answered in laboratory notebook.)
a.
Complete the following table in your laboratory notebook:
TLC
HPLC
Stationary phase
Mobile phase
Indication of
Purity
Indication of
Identity
Polar
analyte
Nonpolar
analyte
Effect of increasing
solvent system
polarity…
b.
List the interm0lecular forces present in each of the compounds below. Which compound would have a
greater affinity for the stationary phase in liquid chromatography? How would this affect the TLC R f value?
How would this affect the HPLC retention time? Which compound would have a greater affinity for the
mobile phase in liquid chromatography? How would this affect the TLC R f value? How would this affect the
HPLC retention time of this compound?
O
OH
C
benzoic acid
naphthalene
CHML211
Exp. 4 TLC and HPLC Analysis of Nitroanilines
POST-LAB ASSIGNMENT: (EACH LAB GROUP will submit one copy of a typewritten, paragraph style report addressing
all of the points listed below. Must be written using PAST TENSE, PASSIVE VOICE. ) …..max score = 50 pts.
8. Experimental (Write 1-2 paragraphs including all of the following. Do NOT present a bulleted outline.)




Describe the process used to prepare the TLC plates, including:
o How the plate itself was prepared
o How the compounds were applied to the plate
o Which compounds were applied to the plate
Describe the process used to prepare the TLC developing chamber, including:
o Which solvents were added to the chambers
o The volume of each solvent used
Describe how the plates were developed, and how the compounds were identified
Describe the process of analyzing the HPLC results, including:
o How were the HPLC results presented?
o How were the compounds in each HPLC chromatogram identified?
9. Results (Copy and paste the completed tables into your document.)
Table 4.1 TLC Results
(This table can be completed by hand, once copy/pasted into document!)
Rf values in
Solvent System 1
80:20
hexane/
ethyl acetate
ANALYTE
TLC Rf Data
Rf values in
Solvent System 2
50:50
hexane/
ethyl acetate
Rf values in
Solvent System 3
100%
ethyl acetate
o-nitroaniline
p-nitroaniline
mixture
TLC
Diagrams
(Show measurements
for solvent front
and all spots in cm)
X
o
X
mix
X
p
X
o
X
mix
X
p
X
o
X
mix
Table 4.2 HPLC Results
(This table can be completed by hand or computer, once copy/pasted into document!)
ANALYTE
o-nitroaniline
p-nitroaniline
STANDARD
Rt (min)
HPLC Rt Data
SAMPLE
SAMPLE
Rt (min)
Rt (min)
Solvent System
Solvent System
1
2
SAMPLE
Rt (min)
Solvent System
3
X
p
10. Discussion (Write 1-2 paragraphs including the following.)




What effect did increasing the TLC developing solvent polarity have on the R f values of the individual compounds?
How would this affect your ability to separate these compounds from one another? Support your conclusion
using actual numerical TLC Rf data from Table 4.1.
What effect did increasing the HPLC eluting solvent polarity have on the Rt values of the individual compounds?
How would this affect your ability to separate these compounds from one another? Support your conclusion
using actual numerical HPLC Rt data from Table 4.2.
If your compounds were contaminated with trace amounts of ethyl acetate, hexane, or water, would this
contaminant be detected and appear as a peak using HPLC analysis? Briefly explain considering the type of
detector used in HPLC analysis.
Include a short comment addressing what could be done differently to improve the experimental results, if
repeated.