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Experiment 3.7 Solvent and Polarity Effects in Thin-layer Chromatography






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Experiment 3.7 Solvent and Polarity Effects in Thin-layer Chromatography. Purpose of Experiment. Determine factors that affect rate of elution (R f ) of organic compounds. Relationship between polarities of compounds and their rates of elution (R f )
Experiment 3.7 Solvent and Polarity Effects in Thin-layer Chromatography

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Experiment 3 7 solvent and polarity effects in thin layer chromatography l.jpgSlide 1

Experiment 3.7Solvent and Polarity Effects in Thin-layer Chromatography

Purpose of experiment l.jpgSlide 2

Purpose of Experiment

  • Determine factors that affect rate of elution (Rf) of organic compounds.

    • Relationship between polarities of compounds and their rates of elution (Rf)

    • Relationship between solvent polarity and rate of elution (Rf)

  • Select an appropriate solvent system to separate and identify components of a mixture.

Purpose of experiment3 l.jpgSlide 3

Purpose of Experiment

  • Possible components of unknown mixture

trans-stillbene

9-fluorenone

benzoic acid

Chromatography color writing l.jpgSlide 4

Chromatography “Color Writing”

Chromatography l.jpgSlide 5

Chromatography

Stationary PhaseMobile Phase

What does it do? stays put! flows around SP

What is it? fine solid with fluid (liquid or gas)

(physical form) lots of surface

How does it effect retains sample by moves sample along

sample movement? surface interaction

Analogous to… an obstacle course “motivating force”

silica gel (SiO2)x ethyl acetate

Thin layer chromatography l.jpgSlide 6

These exposed OH units

give silica gel a

relatively polar surface.

surface

Thin Layer Chromatography

TLC plate

silica gel - silicon dioxide (SiO2)x

(a common, inexpensive stationary phase)

5 x 10 cm

250 mm silica gel layer

impregnated with a

fluorescent indicator,

on a foil backing

bulk (SiO2)x

Thin layer chromatography7 l.jpgSlide 7

Thin Layer Chromatography

  • Five Stages in TLC

  • Prepare Developing Chamber Saturate with solvent vapor.

  • Apply Samples Capillary used to spot solution of each sample.

  • Develop Plate This is when the separation actually occurs.

  • Visualize Developed Plate View under UV light.

  • Interpret Results Determine Rf values & identify components.

Slide8 l.jpgSlide 8

1. Prepare Developing Chamber

  • 400mL beaker.

  • Place a piece of filter paper against side of beaker.

  • Pour 10mL of developing solvent (mobile phase) onto filter paper.

  • Cover beaker with a watch glass and allow to stand undisturbed.

2 apply samples spot the plate l.jpgSlide 9

2. Apply Samples (spot the plate)

1 cm.

“starting line”

1 cm.

TLC plate

Process

A. Draw “guide lines”

lightly with pencil

B.Dissolve solid

sample in CH2Cl2

C. Use TLC capillary

to transfer and spot

dissolved sample

T-stillbene benzoic acid

9-fluorenone unknown

3 develop tlc plate l.jpgSlide 10

TLC plate

3. Develop TLC Plate

  • Place spotted TLC plate in developing chamber.

  • Developing solution is drawn up the plate by capillary action.

  • Remove TLC plate when solvent is ~ 1cm from top.

  • Mark solvent front position with a pencil

NOTE: During this ~20 min.

developing stage, compounds

in the original spots are being

pulled through the silica gel.

Developing Chamber

(400 mL beaker with 10mL solvent)

4 visualize results l.jpgSlide 11

B. View results under UV light.

Look for colored spots on the

fluorescent green background

4. Visualize Results

A. Allow solvent to evaporate

from surface of TLC plate.

UV

C. Mark spots with a pencil

while viewing under UV light.

5 interpret results r f values l.jpgSlide 12

X

Y

T

Y

Z

Y

distance spot has moved

distance solvent has moved

distance spot has moved

distance solvent has moved

distance spot has moved

distance solvent has moved

____________ ___________

_______________________

_______________________

Rf=

Rf=

Rf=

=

=

=

5. Interpret Results (Rf Values)

T

Y

Z

Z

X

5 interpret results identify components l.jpgSlide 13

UNK 9-f ts ba

std std std

5. Interpret Results (identify components)

  • Compare Rf values of components in sample to Rf values of standards.

Prelab preparation l.jpgSlide 14

Prelab Preparation

  • Read Technique K (pp. 92-100)

  • Read Experiment 3.7 A & B (pp. 210 – 216)

  • Prepare Notebook

    • Title, purpose, procedural reference, procedural changes

    • Table of reagents:

      • Structures and hazards of solvents (hexane, ethyl acetate and acetone) and compounds to be separated

    • Rank the solvents in order of increasing polarity.

    • Rank the compounds in order of increasing polarity.

    • Prepare a data table like Table 3.7-1 to record distances and Rf data.

Procedural changes l.jpgSlide 15

Procedural Changes

  • Ethanol will not be evaluated as a solvent.

  • Do part A in groups of 3.

  • Do part B individually.

In lab l.jpgSlide 16

In Lab

  • Part A

    • Determine Rf of trans-stilbene, 9-fluorenone and benzoic acid in hexane, ethyl acetate and acetone.

      • Each person in group does 1 solvent.

    • Determine the best solvent to use to separate a mixture of trans-stilbene, 9-fluorenone and benzoic acid.

      • Review with lab instructor before doing part B.

Data collection l.jpgSlide 17

Data Collection

  • Part A

    • Record the solvent used for each TLC plate.

    • Measure and record the distances that each compound and the solvent front traveled for each solvent used.

    • Calculate and record the Rf values for each compound in each solvent.

In lab18 l.jpgSlide 18

In Lab

  • Part B

    • Use the best solvent determined in part A to detemine components in an unknown mixture.

Data collection19 l.jpgSlide 19

Data Collection

  • Part B

    • Record the unknown number.

    • Record the solvent used for the analysis of the unknown.

    • Measure and record the distances that each known compound, each spot in the unknown and the solvent front traveled.

    • Calculate and record the Rf for each known compound and each spot in the unknown.

    • Identify components in unknown.

Thin layer chromatography20 l.jpgSlide 20

Thin Layer Chromatography

  • TLC lab technique hints

    • Slide watch glass off beaker instead of lifting it off to maintain solvent vapor saturation in beaker.

    • Keep spots small.

    • Never double dip spotting pipette.

    • Put sample(s) to be analyzed near center of plate.

    • Separate spotting points evenly.

    • Do not disturb beaker during development.

Thin layer chromatography21 l.jpgSlide 21

Thin Layer Chromatography

How does solvent polarity affect Rf?

How does solvent polarity affect relative Rf of 2 or more components?

How does component polarity affect Rf?


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