Experiment 20
Chemoselective Reduction with Sodium Borohydride
O
O
H
H
HO
C
C
O
O
+
H3C
O
OCH3
CH2
CH3
NaBH 4
EtOH
NaOH
OCH3
acetic anhydride
O
OH
4-hydroxy-3-methoxybenzaldehyde
(p-vanillin)
OCH3
O
O
CH3
O
CH3
4-acetoxy-3-methoxybenzaldehyde
(vanillin acetate)
4-acetoxy-3-methoxybenzyl alcohol
(vanillin alcohol)
Figure 20.1 Acetylation of p-vanillin followed by chemoselective reduction.
Introduction
Chemoselective reductions of carbonyl compounds to the corresponding alcohols are
very important functional group conversions in organic synthesis, yet there are very few
laboratory experiments which focus on this particular concept. In this experiment, the flavoring
additive vanillin with be acylated in order to produce a compound with two different carbonyl
functional groups. Once acylated, the product will be reduced with sodium borohydride. The
resulting product will illustrate the difference in selectivity of NaBH4 towards different carbonyl
containing functional groups. The reaction will be followed using TLC analysis, and the purity
of the product will be determined using HPLC analysis. Reactants and products will be
characterized using 1H-NMR and IR spectroscopy.
Reducing Agents
The most common laboratory reagents for the reduction of a carbonyl group of an
aldehyde or ketone to an alcohol are sodium borohydride (NaBH4) and lithium aluminum
hydride (LAH), shown in Figure 20.2. Both of these compounds behave as sources of a hydride
ion, which is a very strong nucleophile (see McMurry text, pages 609 and 709).
H
Na +
H
B
H
Li+
H
H
Al
H
H
H
Sodium
borohydride
H
Hydride
ion
Lithium
aluminum
hydride
Figure 20.2 Common reducing agents.
Lithium aluminum hydride is a very powerful reducing agent and reacts not only with
aldehydes and ketones, but also with many other carbonyl containing compounds such as esters,
carboxylic acids, and amides. One disadvantage of LAH is that it reacts violently with protic
solvents such as water and methanol, to produce metal hydroxides or alkoxides, and hydrogen
gas, which could result in an explosion or fire.
169
Sodium borohydride is much milder, much more selective reducing agent than lithium
aluminum hydride. Unlike lithium aluminum hydride, sodium borohydride will not reduce
carbonyl-containing compounds such as esters, carboxylic acids, or amides, therefore an
aldehyde or ketone can be reduced with sodium borohydride in the presence of these other types
of functional groups. These reductions can be performed in a wide variety of solvents, such as
aqueous methanol or ethanol, with a good to excellent yield.
The key step in the metal hydride reduction of an aldehyde or ketone is transfer of a
hydride ion from the boron atom of the reducing agent to the electropositive carbon of the
carbonyl group to form a tetrahedral intermediate (Figure 20.3) called an alkoxide ion. The
resulting alkoxide ion is protonated by the protic solvent to form a neutral alcohol. One mole of
NaBH4 is a source of four hydride ions, therefore can react with four carbonyl groups to produce
four moles of the product. Thus, one mole of the reducing agent reduces four moles of the
carbonyl compound.
4
R
O
C
O
4
H
R
C
OH
CH3CH2O H
4
R
H
H
C
H
H
H
H
B
H
H
Figure 20.3 Mechanism for the sodium borohydride reduction of an aldehyde.
Chromatographic Analysis
All reactants and products in this experiment contain aromatic rings which are
chromophores easily visualized using a UV lamp. It is also optional to stain the TLC plate with
certain compounds that not only aid in the visualization of compounds, but also provide a
method for determining which functional groups are contained within a molecule. A solution of
2,4-Dinitrophenylhydrazine (2,4-DNP) is a stain mainly used to detect aldehydes and ketones.
When these types of carbonyl compounds react with 2.4-DNP, they form the corresponding
hydrazone, which is usually yellow or orange, thus resulting in the spot on the TLC plate to
appear colored. Interestingly, 2,4-DNP does not react with other carbonyl containing functional
groups such as esters, carboxylic acids, and amides, therefore can be useful to distinguish
between compounds containing more than one type of carbonyl functional group.
R
H
C
HN
NH 2
HN
O
NO 2
N
NO 2
+
H
R
aldehyde or ketone
NO 2
NO 2
product hydrazone
(yellow-orange)
2,4-DNP
(red-orange)
Figure 20.4 Reaction between 2,4-DNP and an aldehyde or ketone.
170
IR Spectroscopy
The products can be easily distinguished from the reactants using IR spectroscopy. The
first step of the reaction results in the conversion of a hydroxyl group to an acetyl group. The
phenolic O-H stretch typically appears as a broad absorption in the range of 3200-3600 cm-1.
Also present is the C-O bond absorptions, which typically appear between 1000-1300 cm-1. It is
interesting to note that a characteristic of esters is the presence of two of these absorption bands,
one stronger and broader than the other. Aldehydes exhibit two C-H stretches around 2700 and
2800 cm-1, however these may be difficult to distinguish when broad O-H absorptions are
present. Finally, a C=O stretching absorption would appear between 1680-1740 cm-1. Make
note that if an acetyl group was successfully added onto the reactant, a second C=O stretching
absorption could appear in this region.
The second step of the reaction results in the conversion of a carbonyl group to a
hydroxyl group. IR spectroscopy can certainly be used to determine whether the reducing agent
reduces one, or both, of the carbonyl substituents based on the appearance or disappearance of
certain types of absorptions characteristic of those types of carbonyl groups. The IR spectra of
the reactants and products are shown in Figure 20.5.
NMR Spectroscopy
The 1H NMR spectra of the reactant and products are shown in Figure 20.6 The chemical
shift of a phenolic hydrogen in proton NMR varies depending on the purity of the sample, the
temperature, and the sample solvent. It typically appears in the range of 5.0-10.0 ppm,
depending on the experimental conditions. Hydrogens on the carbon bearing the hydroxyl group
are deshielded by the oxygen atom, and typically appear in the range of 3.5-4.5 ppm. Hydrogens
attached to carbons to the carbonyl typically resonate between 2.0-3.0 ppm. The aldehyde
proton signal is very distinctive, appearing as a singlet 10.0 ppm. Very few signals appear that
far downfield in a 1H NMR spectrum.
Objectives
In this experiment you will synthesize acetylate p-vanillin using acetic anhydride. Once
acylated, the product will be reduced using NaBH4 in ethanol. The identity and purity of the
product will be analyzed using TLC and HPLC analysis. Finally, reactants and products will be
characterized using IR and 1H NMR spectroscopy.
Experimental
PART 1:
Synthesis: Preparation of Acylation product
Mix ~0.76 g of p-vanillin and 12.5 mL 10% NaOH in a 125 mL Erlenmeyer flask containing
a stir bar. Stir for 2-3 minutes.
Add 15.0 g of ice to flask, then add 2.0 mL acetic anhydride while stirring. Place a large
cork loosely in the top of the flask, and allow reaction to stir for 15 minutes. Record any
observations in your laboratory notebook.
171
Purification of Acylation product:
If a white precipitate forms:
o Suction filter to isolate the crude solid. Use ~25 mL ice cold water to help
transfer solid from reaction flask to funnel, and to rinse solid once filtered.
o Transfer crude solid to a 50 mL Erlenmeyer flask. Recrystallize the solid in ~3
mL hot ethanol. Once dissolved, cool to room temperature, and then place in an
ice bath for 15 minutes to induce crystallization.
o Suction filter to isolate the pure solid. Use ice cold ethanol to help transfer the
solid to the funnel, and to rinse the solid once filtered. If any solid appears in the
filtrate, repeat filtration process.
o Prepare an HPLC sample by transferring 2-3 crystals to an autosampler vial and
dissolving completely in the provided HPLC solvent (2:3 ethyl acetate/hexane).
Submit this sample for analysis.
o Prepare a TLC sample by transferring 2-3 crystals to a small sample vial and
dissolving in ½ mL reagent acetone. Label this vial clearly ACYLATION
PRODUCT and use during Product Analysis below.
o Transfer the solid from the funnel to a preweighed large filter paper. Secure with
a small amount of tape and submit to instructor to dry until next lab period.
o Once dry, reweigh solid to obtain a final product mass, calculate % yield, and
obtain an experimental melting point of this solid. Complete Tables 20.1 and 20.3
on the final lab report.
If a colorless liquid or oil forms:
o Transfer reaction solution to a separatory funnel, using ethyl acetate to aid
transfer if necessary.
o Extract the solution 2X with 15 mL ethyl acetate (review Experiment 6 if
necessary).
o Wash the combined organic layers with 25 mL 1M NaOH. Check pH of aqueous
solution. Continue to extract with NaOH until the aqueous layer is strongly basic
(pH>9).
o Dry organic layer over MgSO4. Decant the solution into a preweighed 250 mL
beaker containing a few boiling chips, and evaporate solvent on a warm hotplate
(setting of 3, NO HIGHER!) to yield the solid product.
o Prepare an HPLC sample by transferring 2-3 crystals to an autosampler vial and
dissolving completely in the provided HPLC solvent (2:3 ethyl acetate/hexane).
Label this vial clearly ACYLATION PRODUCT and store in lab drawer until
next lab period.
o Prepare a TLC sample by transferring 2-3 crystals to a small sample vial and
dissolving completely in reagent acetone. Label this vial clearly ACYLATION
PRODUCT and store in lab drawer until next lab period.
o Cover the beaker containing the product with a Kim Wipe sheet and secure with a
rubber band. Allow the product to dry until the next lab period.
o Once dry, reweigh solid to obtain a final product mass, calculate % yield, and
obtain an experimental melting point of this solid. Complete Tables 20.1 and 20.3
on the final lab report.
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Product Analysis:
HPLC Analysis
Compare the HPLC results of the ACYLATION PRODUCT sample submitted to the
standard chromatogram. Identify any compounds present in samples, and quantify
compounds present. Complete Table 20.5 on the final lab report.
TLC Analysis
Perform a TLC experiment on the ACYLATION PRODUCT sample along with the
provided standard of p-vanillin. Develop this plate in 2:3 ethyl acetate/hexane and
visualize under UV lamp.
Using tweezers, dip this plate in 2,4-DNP TLC stain. Blot excess stain on a paper
towel, before transferring back to lab hood. Once completely dry, the 2,4-DNP
reagent will produce a dark orange spot with any compound containing an aldehyde
functional group. Sketch a diagram of this plate in your laboratory notebook, complete
with color descriptions of the spots which appear. Complete Table 20.6 on the final
lab report.
PART 2:
Synthesis: Reduction of Acylation product
Record product mass from previous experiment. Transfer solid to 100 mL beaker. Add 5
mL ethanol, and swirl to mix completely.
Hold the beaker and swirl directly on the surface of a warm hotplate (setting of 3) to dissolve
solid completely.
Lower the beaker into a shallow water bath, cooled with a few ice cubes.
Weigh ~0.30 g of NaBH4. Add a small portion of this solid to the beaker and stir with glass
rod (CAUTION! Do NOT add large amounts of NaBH4 all at once!). Continue adding the
NaBH4 in small portions while stirring over a period of 2 minutes. Stir the reaction solution
every few minutes for a total of 15 minutes in the cold water bath, then for an additional 15
minutes at room temperature.
After 30 minutes, remove 1-2 drops of the reaction solution with a pipet and transfer to a
small test tube. Add ½ mL reagent acetone to prepare a TLC sample of the CRUDE
REDUCTION PRODUCT (the solid will not completely dissolve).
Perform a TLC experiment on this reaction mixture along with the provided p-vanillin
standard, and the ACYLATION PRODUCT sample prepared during the previous lab period.
Develop the plate in 2:3 ethyl acetate/hexane, and visualize under a UV lamp. If there is any
evidence of reactant present, stir the reaction for an additional 15 minutes and repeat the TLC
analysis until there is no longer evidence of the starting material, thus indicating that the
reaction is complete.
Once complete, add 10 mL water to the reaction mixture. Stir this solution for an additional
5 minutes.
Purification of Reduction product:
Transfer the reaction solution to a separatory funnel. Extract the aqueous layer twice with 10
mL diethyl ether (review Experiment 6 if necessary).
Wash the combined organic layers with 10 mL water, and then 10 mL saturated NaCl.
173
Dry the organic layer over MgSO4. Decant the solution into a preweighed 50 mL flask, and
clamp to a ring stand.
Evaporate the solvent by blowing air gently into the flask. The product will be a colorless,
viscous oil.
Reweigh the flask to obtain a final product mass, and calculate % yield. Complete Tables
20.2 and 20.4 on the final lab report.
Add 3 mL reagent acetone directly to the flask, and swirl to dissolve the oily product. Use
this solution as your TLC sample for the PURE REDUCTION PRODUCT.
Prepare an HPLC sample of the PURE REDUCTION PRODUCT by transferring 2 drops of
the liquid sample (in acetone from above) to an autosampler vial, and dissolving completely
in the provided HPLC solvent (2:3 ethyl acetate/hexane). Submit this sample for analysis.
Proceed to Product Analysis.
Product Analysis:
HPLC Analysis
Compare the HPLC results of the PURE REDUCTION PRODUCT sample to the
standard chromatogram. Identify any compounds present in the sample, and quantify
compounds present. Complete Table 20.5 on the final lab report.
Compare the HPLC results of the ACYLATION PRODUCT sample to the PURE
REDUCTION PRODUCT sample. Was the acylation product pure when you started the
reduction portion of the experiment?
TLC Analysis
Perform a final TLC experiment on the PURE REDUCTION PRODUCT sample
along with the provided standards of p-vanillin, vanillin acetate, vanillyl alcohol, and
vanillin alcohol. Develop this plate in 2:3 ethyl acetate/hexane and visualize under
UV lamp. Sketch a diagram of this plate in your laboratory notebook.
Using tweezers, dip this plate in 2,4-DNP TLC stain. Blot excess stain on a paper
towel, before transferring back to lab hood. Once completely dry, the 2,4-DNP
reagent with produce a dark orange spot with any compound containing an aldehyde
functional group. Sketch a diagram of this plate in your laboratory notebook, complete
with color descriptions of the spots which appear. Complete Table 20.7 on the final
lab report.
IR Analysis
Using the provided spectra in Figure 20.5, identify all characteristic absorptions of
reactants and products. Complete Table 20.8 on the final lab report.
NMR Analysis
Using the provided spectra in Figure 20.6, identify and tabulate all characteristic
resonances of reactants and products. Complete Table 20.9 on the final lab report.
174
WASTE MANAGEMENT
Place all solid waste into the container labeled “SOLID WASTE” located in the waste
hood. Place all liquid waste into the container labeled “LIQUID WASTE”. Place any used
melting point capillaries and TLC capillaries in the “BROKEN GLASS” container. TLC plates
and filter papers can be discarded in the yellow trashcan.
References
McMurry, John. (2008). Organic Chemistry, 7th ed. Belmont: Thomson/Brooks/Cole.
Baru, A. and Mohan, R. (2005). The Discovery-Oriented Approach to Organic Chemistry.
Selective Reduction in Organic Chemistry: Reduction of Aldehydes in the Presence of Esters
using Sodium Borohydride. Journal of Chemical Education. 82 (11): 1674-1675.
175
2843, 2751
3374
1673
1291
2845, 2753
1756
1126
1690
1207
3395
1150
1762
1197
Figure 20.5 IR spectra of reactants and products.
176
7.04
1H, d
CDCl3
NMR Solvent
H2,6
7.43
2H, m
6.21
1H, s
O
H1
C
9.83
1H, s
6
3.97
3H, s
2
5
OCH3
3
OH
H2,6
7.49
2H, m
CDCl3
NMR Solvent
4
7.22
1H, d
O
3.91
3H, s
2.35
3H, s
H1
C
6
2
5
OCH3
3
9.93
1H, s
O
CH3
4
C
CDCl3
NMR Solvent
O
H2,6
6.90
2H, m
7.01
1H, d
3.84
3H, s
CH2 OH
2.31
3H, s
7
1
2
6
5
4.62
2H, s
5.30
1H, s
Figure 20.6 1H-NMR spectra of reactants and products.
177
OCH3
3
O
C
O
CH3
4
178
Final Lab Report
Due Date_______
Names______________________________
______________________________
Exp. 20: Chemoselective Reduction with Sodium Borohydride
EXPERIMENTAL RESULTS
(Tables in INK only!)
Table 20.1 Acylation Experimental Results
Table 20.2 Reduction Experimental Results
Theoretical yield (g)
Actual yield (g)
Percent yield
Product Appearance
Experimental Melting Point (oC)
Theoretical yield (g)
Actual yield (g)
Percent yield
Product Appearance
Table 20.3 Acylation Green Chemistry Results
Table 20.4 Reduction Green Chemistry Results
Atom Economy (%)
Experimental Atom
Economy (%)
“Eproduct”
Cost per synthesis ($)
Cost per gram ($/g)
Atom Economy (%)
Experimental Atom
Economy (%)
“Eproduct”
Cost per synthesis ($)
Cost per gram ($/g)
Table 20.5 HPLC Analysis
Sample
ACYLATION
PURE REDUCTION
PRODUCT
PRODUCT
Retention
Area
Retention
Area
Times
Percent
Times
Percent
(min)
(min)
Standard
Compound
Retention
Times
(min)
p-vanillin
Vanillin acetate
Vanillin alcohol
Vanillyl alcohol
Table 20.6 TLC Analysis—Part 1
Compound
Part 1
Standard Sample
Rf
Rf
TLC Diagram
Table 20.7 TLC Analysis—Part 2
Compound
p-vanillin
p-vanillin
Vanillin
acetate
Vanillin
alcohol
Vanillin
acetate
Vanillyl
alcohol
179
Part 2
Standard
Sample
Rf
Rf
TLC Diagram
Table 20.8 IR Analysis
Base
Values
p-vanillin
Vanillin acetate
Vanillin alcohol
Frequency
(cm-1)
Frequency
(cm-1)
Frequency
(cm-1)
Functional Group
Frequency
(cm-1)
3200-3600
1000-1300
1680-1740
2700-2800
OH stretch
C-O stretch
C=O stretch
Aldehyde C-H stretch (2)
Table 20.9 1H NMR Analysis
p-vanillin
O
Vanillin acetate
H1
C
O
C
H1
Vanillin alcohol
CH2 OH
H1
6
H1
6
2
7
1
H1
2
6
2
5
H2,6
OCH3
5
5
OCH3
OCH3
H2,6
3
3
O
OH
C
4
3
H2,6
O
C
CH3
CH3
4
4
O
H3
H3
H4
H5
H3
O
H4
H5
H4
H5
H7
DISCUSSION/CONCLUSIONS
(In the space provided, briefly answer the following questions. Use numerical values to support conclusions where applicable.)
O
O
H
C
O
O
C
H3C
OCH3
OH
4-hydroxy-3-methoxy
benzaldehyde
(p-vanillin)
$33.00/100g
1.
CH3
NaBH 4
sodium borohydride
$74.50/100g
C
O
H
C
CH3
NaOH
CH3CH2OH
ethanol
$20.60/1L
acetic anhydride
$39.20/500mL
OCH3
H3C
O
C
O
4-acetoxy-3-methoxy
benzaldehyde
(vanillin acetate)
$190.20/100g
OCH3
H3C
O
C
4-acetoxy-3-methoxy
benzyl alcohol
O
Based on your HPLC analysis, did you synthesize only the desired product during the first step of the
synthesis, or were other products formed? Did you synthesize only the desired product during the second
step of the synthesis? Identify any compounds present in your sample chromatograms and give actual area
percent values for each. If more than one product formed, briefly explain why this occurred.
180
2.
What is one type of IR absorption band that could be used to indicate that the conversion from the aldehyde
to the alcohol took place? What is the typical frequency for this type of absorption? What is the actual
frequency of the absorption in the provided spectrum?
3.
What is one signal in the 1H NMR spectrum that indicates that the reduction took place? What are the
typical chemical shifts that these types of protons may appear? What is the actual chemical shift of this
proton signal?
4.
Once stained, how was the TLC plate used to indicate conversion of the reactant to the product alcohol? In
your answer, briefly describe the Rf values and appearance of spots.
5.
Draw the product, and a complete mechanism for its formation, for the following reaction. In your
mechanism, use only the hydride ion as your nucleophile.
O
H
O
Li
H3C
C CH2 CH2 C
OCH3
H
Al
H
CH3CH2OH
H
**Attach sample chromatograms!**
181