The objective of this experiment is to inspection the stereoselectivity of miscellaneous reducing agents in the palliation of a carbonyl group. The very first part that this experiment will emphasis on the stereoselectivity of sodium borohydride, lithium aluminum hydride, and also lithium tri-sec-butylborohydride (L-selectride) in the reduction of 4-tert-butylcyclohexanone.1 The second part of this experiment will focus on the stereoselectivity of saccharomyces cerevisiae (baker’s yeast) in the reduction of ketone in ethyl acetoacetate.2 The stereoselectivity of every reducing agent will be examined via 1H-NMR spectroscopy, and the product will certainly be further defined using infrared (IR) spectroscopy and melting point analysis. Figure 1 below shows the structures of the steel hydride reduce agents.

You are watching: Reduction of 4-tert-butylcyclohexanone

Figure 1: framework of salt borohydride, lithium aluminum hydride, and also L-selectride, respectively.

Metal hydride palliation of a carbonyl team to one alcohol involves activation that the carbonyl carbon by the metal followed by the nucleophilic strike of hydride come the carbonyl carbon. The job-related up is frequently performed in H­2O or H3O+ come protonate the oxygen atom and kind the alcohol oh group.3 A reaction scheme for the reduction of 4-tert-butylcyclohexanone is presented below.

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Figure 2: palliation of 4-tert-butylcyclohexanone.

The cis and also trans assets in figure 2 show the resulting alcohol OH team in the axial and also equatorial positions, respectively. Because of the sp2 hybridization of the carbonyl carbon, it is planar, and thus, can be attacked from either the top or bottom challenge of the molecule. By attacking from the top face of the molecule, the resulting alcohol OH group is driven to the equatorial position, as presented in the infectious diseases world fashion product above. Top top the contrary, attack from the bottom confront of the molecule pushes the alcohol OH team into the axial position, as shown in the cis product above. The infectious diseases worldwide product is the thermodynamically an ext stable product because of the tert-butyl and also alcohol OH teams both being in the equatorial positions. The equatorial position results in much less steric crowding, and thus, the is favorable for larger substituents come occupy that position.3 However, the cis product forms more rapidly because of the bulky tert-butyl group impeding the ability of the hydride ion to assault the top confront of the molecule.

Being the the infectious diseases world fashion product is an ext thermodynamically stable, the is stated to be under thermodynamic control. Thermodynamic control method the product that is much more energetically stable will form preferential to other, less stable products.3 However, the formation of the thermodynamic product involves a better activation energy.4 The cis product entails a smaller activation energy, and forms much more rapidly than the thermodynamic product.4 Thus, the cis product is pertained to as being under kinetic control. Kinetic control means the product that creates the most rapidly is favored, despite not being the most stable.3 The formation of a thermodynamically favored product is reversible, and the formation of a kinetically favored product is irreversible.4 By assessing the 1H-NMR spectra that the hydride palliation products, the proportion of trans come cis product developed in each reduction reaction can be determined.

Understanding each steel hydride reducing agent enables for an educated guess regarding whether the hydride will strike from the top or bottom face of the 4-tert-butylcyclohexanone. Both sodium borohydride and also lithium aluminum hydride are smaller, less bulky metal hydride reduce agents, and also thus that is intended that lock will have the ability to deliver the hydride from the top confront of the molecule (see “Figure 1” and also “Figure 2”).4 The bulky tert-butyl group will not hinder the hydride attacking from the optimal face. However, L-selectride is a lot larger and bulkier metal hydride reagent, and thus, would most most likely not be able to strike from the top confront with the presence of the bulky tert-butyl team (see “Figure 1” and “Figure 2”).4 Accordingly, the hypothesis is the the salt borohydride and also lithium aluminum hydride will form the infectious diseases worldwide product, and also the L-selectride will kind the cis product.

Baker’s yeast consists of naturally occurring biochemical enzymes v reductive capabilities. This experiment focuses on the yeast reduction of ethyl acetoacetate.2 The reaction system for the yeast reduction of ethyl acetoacetate is shown below.

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Figure 3: palliation of ethyl acetoacetate using baker’s yeast.

The enzymes present in baker’s yeast selectively alleviate the ketone end the ester. The ester is more difficult to reduce due to the visibility of the pi bond and also resonance stabilization.3 The result alcohol OH team can adopt either the R or S configuration. However, it is not feasible to recognize the relative amount of R and also S product via 1H-NMR spectroscopy4. The R and S configurations are enantiomers, and enantiomers cannot be identified in an achiral environment, such together 1H-NMR. Consequently, the product must undergo additional functionalization in order to be distinguishable via 1H-NMR spectroscopy.2 S-(+)-a-methoxyphenylacetic mountain is added to the enzyme palliation product, creating a diastereomeric pair. The enzyme palliation product exists together both R and also S, and also the chiral acid is strict S configuration. Diastereomers have the right to be identified using 1H-NMR spectroscopy, and thus, the relative amounts of SR and SS isomer created can it is in determined.4 The reaction system for the enhancement of S-(+)-a-methoxyphenylacetic acid to the enzyme reduction product is presented below.

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Figure 4: enhancement of S-(+)-a-methoxyphenylacetic mountain to the enzyme reduction product.

 

The reduction of b-ketoesters using baker’s yeast has been extensively studied and is recognized to present enantioselectivity.5 literary works precedence support the hypothesis that the S configuration will certainly be favored end the R configuration.6

In regards to a “real world” application, reduction reactions entailing baker’s yeast open up a window of chance for “green chemistry.” Baker’s yeast is neither toxic nor pathogenic, and also is a really economical catalyst.5 Additionally, by analyzing the varying staminas of the steel hydride to reduce agents, selective reduce of carboxylic acid derivatives room possible.3

Experimental Procedure—Sodium Borohydride Reduction

The experimental procedure was very similar to the one outlined in the activities handout, v some young modifications.1

4-tert-butylcyclohexanone (202 mg, 1.31 mmol, 1 equiv) was added to a 5 mL conical vial equipped with an waiting condenser, spinvane, and also stir bar. Methanol (CH3OH) (200 mL, 4.94 mmol, 3.77 equiv) was included through the waiting condenser and also the systems was stirred continuously. Salt borohydride (NaBH4) (400 uL, 11.31 mmol, 8.56 equiv) to be slowly included to the conical vial and the reaction to be stirred for 20 minutes. Thin-layer chromatography was supplied to screen the reaction. Thin-layer chromatography shown the presence of starting material in the reaction mixture, and also thus, the reaction to be stirred for second 30 minutes. An infrared (IR) spectrum the the starting material was got while stirring.

Cold hydrochloric mountain (HCl) (4 mL) was included drop wise to the reaction. The reaction was moved to a 15 mL centrifuge tube. The aqueous layer to be extracted via three 2 mL washes that methylene chloride (CH2Cl­2). The centrifuge tube was easy shaken to help in the separation the the layers. The CH2Cl2 layer was extracted utilizing a Pasteur pipette, and also it was transferred to a drying tower containing sodium sulfate anhydrous (Na2SO­4). A tared 25 mL round-bottom flask was provided to collection the dried eluates from the dry column. CH2Cl2 was supplied to rinse the continuing to be product native the dry column into the flask. A boiling stone was added to the flask. A sand bathtub was provided to warm the flask and remove the CH2Cl2 from the reaction mixture. The reaction was concentrated to dryness, and the product was weighed. 1H-NMR and IR spectra as well as a melting point measurement that the product to be acquired.

See attached page for calculations of mmol used, theoretical yield, and also a flow chart for the “Work-up and Isolation that the crude oil Product.”

Results/Data/Spectra

Table 1: 1H-NMR (CDCl3) key Peak Table because that 4-tert-butylcyclohexanone

Chemical transition (ppm)

Type the Signal

J worths (Hz)

Integration

Type that Proton

0.94

S

—-

9

CH3

1.76

M

7.0, 7.0

4

CH2

1.77

M

7.0, 7.1

1

CH

2.22

T

7.1

4

CH2

*Predicted making use of ChemBioDraw 13.0

 

Table 2: 1H-NMR (CDCl3) optimal Table because that Hydride palliation Product

Chemical transition (ppm)

Type the Signal

J values (Hz)

Integration

Type of Proton

1.85

S

—-

9

CH3

3.45

T the T

11.12, 5.16

2.39

CH2

4.15

Q

5.82*

0.99

CH2

*Average of all J-values, calculations presented on spectrum

Table 3: Infrared Spectroscopy height Table because that 4-tert-butylcyclohexanone

Absorption (cm-1)

Functional Group

2946.85, 2867.14

Alkane C-H Stretch

1720.59

Carbonyl C=O Stretch

 

Table 4: Infrared Spectroscopy top Table because that Hydride reduction Product

Absorption (cm-1)

Functional Group

3292.91

Alcohol O-H Stretch

2938.69, 2858.90

Alkane C-H Stretch

1719.99

Carbonyl C=O Stretch*

*Carbonyl peak transmittance: 81.36%

Melting point Measurement: 45-530 C

Table 5: class Data

Type the Hydride Reduction

Product Ratio

Sodium Borohydride

2.4:1.0 (Trans : Cis)

Lithium Aluminum Hydride

9.5:1.0 (Trans : Cis)

L-Selectride

20:1 (Cis : Trans)

Enzyme Reduction

10% ee S; 44% ee S

 

 

Discussion

The melting suggest measurement of the hydride reduction product was 45-530 C. The literary works value for the melting suggest of the mixed isomers is 62-700 C.7 The melting allude is a rather vast range, and occurs well listed below the literature value, indicating the product is not entirely pure. The product is a mixture that the cis and trans isomers as well as the beginning material. Thin-layer chromatography and IR spectroscopy additionally indicate the visibility of beginning material and will be disputed further.

IR spectroscopy is specifically useful in determining the functional groups in a compound, and also thus, is incredibly helpful in determining even if it is the reduction product formed. The IR spectrum the the starting material includes a characteristic optimal at 1720.59 cm-1, representing a carbonyl C=O.3 In the IR spectrum the the hydride palliation product, this top is quiet present, but not as intense (see “Table 4”). This shows that there is beginning material present in the product, and this is in commitment with the thin-layer chromatography evaluation (see “Thin-Layer Chromatography”). The thin-layer color layer analyzer plate indicated starting material was present in the reaction mixture, and thus, the reaction did not reach completion. The IR spectrum the the hydride palliation product exhibits a large peak at 3292.91 cm-1, describe the existence of one alcohol O-H.3 The appearance of this stretch supports the idea the the hydride reduction product to be formed. However, the existence of the carbonyl C=O stretch in this spectrum reflects that the product is not totally pure, as it has a far-reaching amount of starting material.

The significant product developed in this reaction was the trans-4-tert-butylcyclohexanol product. Upon analysis of the 1H-NMR spectrum, that is apparent that the trans product created over the cis product in a 2.4:1 proportion (see “Table 5”). The trans come cis ratio was figured out by examining the 1H-NMR spectrum in the 3.0 ppm to 5.0 ppm region. In this region, there is a triplet of triplets in ~ 3.45 ppm and also a quintet at 4.15 ppm (see “Hydride reduction Product 1H-NMR Spectrum). The integration values for every peak deserve to be offered to determine the relative ratio of trans to cis product. However, the two peaks must an initial be figured out as one of two people the trans optimal or cis top in order to recognize the ratio.

In bespeak to determine which peak corresponds to i m sorry isomer, the dihedral angle of the surrounding carbon-six and carbon-two protons in relation to the carbon-one proton should be investigated. Drawing a Newman estimate of the infectious diseases world fashion product looking under carbon-two come carbon-one mirrors dihedral angles of 600 in between H1 and H2 and 1800 in between H1 and also H2’. Therefore, the H2 and H2’ protons would certainly both split the H1 proton, creating a double of doublets. Drawing a Newman estimate of the infectious diseases world fashion product looking down carbon-one come carbon-six mirrors dihedral angles 600 in between H1 and also H6 and also 1800 between H1 and also H6’ Consequently, the H6 and also H6’ protons would both break-up the H1 proton, producing a double of doublets. The H1 is split into a doublet of doublets that doublets that doublets, or more simply put, a triplet the triplets.8 The top at 3.45 ppm is plainly identified as a triplet of triplets with a small amount of height overlap. The Newman projections are presented in number 5 below.

Figure 5: Newman projections for the trans sodium borohydride palliation product. Looking down carbon two-carbon one (left) and carbon one-carbon six (right).

Drawing a Newman forecast of the cis product looking under carbon-two come carbon-one reflects dihedral angles of 600 in between H1 and H2 and also 600 in between H1 and also H2’. Drawing a Newman forecast of the cis product looking under carbon-one come carbon-six mirrors dihedral angles 600 between H1 and also H6 and also 600 in between H1 and H6’. Being the all four neighboring protons have the exact same dihedral angle to the H1 proton, they are equivalent. Consequently, these 4 protons produce a quintet, and also this is determined as the optimal at 4.15 ppm.8 The Newman projections are shown in number 6 below.

Figure 6: Newman projections because that the cis sodium borohydride palliation product. Looking down carbon two-carbon one (left) and carbon one-carbon 6 (right).

After identifying the optimal at 3.45 ppm as the trans product and also the top at 4.15 ppm together the cis product, the relative proportion of trans come cis product deserve to be calculated using the integration worths of the peaks. By dividing the integration because that the trans optimal by the integration because that the cis peak, the relative ratio of trans come cis product was established to it is in 2.4:1.0.

By evaluating the chair conformations that the cis and trans products, it is apparent that the infectious diseases world fashion product is thermodynamically favored and the cis product is kinetically favored. It is clear the the infectious diseases world fashion product was formed in the sodium borohydride reduction of 4-tert-butylcyclohexanone. The thermodynamically favored product was formed due to the fact that sodium borohydride is not a sterically hindered reducing agent, and also is as such able to attack the 4-tert-butylcyclohexanone from the top face of the molecule regardless of the existence of the large tert-butyl group. The enhancement of the hydride native the top face places the alcohol OH group in the equatorial position, creating the an ext energetically steady product.3 If the salt borohydride assaulted from the bottom face, the alcohol OH team would be forced into one energetically unfavorable axial position. In moving to this axial position, the oxygen should past v a extremely unstable overshadowed conformation.4 Thus, the thermodynamically favored product is developed when utilizing sodium borohydride in the palliation of 4-tert-butylcyclohexanone.

Lithium aluminum hydride selectively decreased the 4-tert-butylcyclohexanone come the trans product over the cis product in a 9.5:1.0 ratio. Lithium aluminum hydride is comparable to salt borohydride in the it is no sterically hindered and also has the capability to the assault the carbonyl indigenous the peak face.4 Thus, as soon as the hydride assaults from the peak face, the alcohol OH group is moved to the equatorial position. This develops the thermodynamically favored product. ~ above the contrary, L-selectride selectively formed the cis product end the trans product in a 20:1.0 ratio. L-Selectride is a bulky, sterically hindered reducing agent, and thus, does not have the ability to assault from the top confront of the starting material.4 that is required to strike the 4-tert-butylcyclohexanone from the bottom confront due to the presence of the bulky tert-butyl group. The bottom-ide strike pushes the alcohol OH team into an energetically unfavorable axial position. Back this product is not energetically favorable, that forms an ext quickly, and also thus, is the kinetic product. Thus, the L-selectride palliation of 4-tert-butylcyclohexanone is under kinetic control.

The enzyme reduction reaction favored the S construction product in 10% enantiomeric excess and 44% enantiomeric excess. The ethyl acetoacetate contains both a ketone and an ester practical group. The baker’s yeast selectively reduce the ketone come an alcohol rather of reducing the ester. The ester is more an overwhelming to reduce because of the visibility of the pi bond and also resonance stabilization.3 Baker’s yeast is a naturally emerging chiral fixing agent and also is stereoselective in the reduction of the ketone come an alcohol.2 it favors the S configuration, together demonstrated in the data. However, that is not feasible to distinguish in between enantiomers in an achiral environment, such as in 1H-NMR spectroscopy, and also thus, the enzyme reduction product necessary to be more functionalized to determine whether the R or S configuration was favored in the ketone reduction.4 A chiral acid, S-(+)-a-methoxyphenylacetic acid, was included to the enzyme reduction product. The enzyme palliation product exists together either R or S, and the chiral mountain is strictly S. Thus, a diastereomeric pair was generated, which to be easily differentiated via 1H-NMR spectroscopy. By examining the 1H-NMR spectrum, it was clear that the SS diastereomer developed over the SR diastereomer, and thus, the S construction of the alcohol was favored in the enzyme palliation of ethyl acetoacetate.

 

Conclusion

The purpose of this experiment to be to investigate the stereoselectivity of assorted hydride reducing agents.1,2 The reduction of 4-tert-butylcyclohexanone making use of sodium borohydride, lithium aluminum hydride, and also L-selectride to be studied in detail. ~ extensive evaluation of 1H-NMR spectra, IR spectra, and melting point measurements, it is clean that sodium borohydride favored the infectious diseases worldwide product end cis product in a ratio 2.4:1.0, lithium aluminum hydride favored the infectious diseases worldwide product end cis product in a proportion of 9.5:1.0, and the L-selectride favored the cis product over trans product in a proportion of 20:1.0. The smaller, much less bulky sodium borohydride and also lithium aluminum hydride attacked the carbonyl carbon indigenous the top face of the molecule, forming the trans and thermodynamically-controlled product.3,4 ~ above the contrary, the large, bulky L-selectride attacked the bottom face of the molecule, forming the cis and kinetically-controlled product.3,4 The steric hindrance produced by the bulky tert-butyl team did not affect the top-side strike of salt borohydride and lithium aluminum hydride, yet forced a bottom-side assault with the L-selectride. Thus, the theory was correct.

The reduction of ethyl acetoacetate utilizing baker’s yeast favored the S construction in 10% enantiomeric excess and 44% enantiomeric excess. Therefore, that is obvious that baker’s yeast favored the S configuration, and also thus, the hypothesis was correct.

Future work could include 13C-NMR spectroscopy to further characterize the product, and also proton-decoupling or COSY experiments to more analyze the 1H-NMR spectra.

References

 

“Hydride to reduce of 4-tert-butylcyclohexanone,” Dr. Lynn Bradley, The college of brand-new Jersey.Patterson, J.; Sigurdsson, S. “Use of enzyme in organic Synthesis: palliation of Ketones by Baker’s Yeast Revisited,” J. Chem. Educ. 2005, 1049-1050.Brown, William, Christopher Foote, Brent Iverson, and also Eric Anslyn. Essential Chemistry. 5th ed. Brooks/Cole, 2008. Print.“Lecture notes on palliation Reactions,” Dr. Lynn Bradley, The university of brand-new Jersey.Mahmoodi, N. O.; Navrood, M. N. “Enantio-, regio-, chemoselective of fragrant a-diketones by baker’s yeast in varied organic-water solvent systems,” Arkivoc. 2007, 37-45.

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“Experiment 8, Synthesis and Purification that Diastereomers: sodium Borohydride reduction of a Chiral Ketone,” Dr. Alan Shusterman, Reed College.Mayo, Pike, and Trumper. Microscale organic Laboratory. 3rd ed. New York: man Wiley and Sons, 1994. 165. Print.“Lecture notes on 1H-NMR Splitting,” Dr. Lynn Bradley, The college of brand-new Jersey.