Biocatalytic Reduction (Final)

Biocatalyst

Any living or dead cells, cell extracts, pure enzymes

Advantages of using a Biocatalyst

Inactivated C-H functionalized, stereoselective, conversion at preferred temp and pH, catalyst reused

Nobel Prize winner 2018 for Directed Evolution

Frances Arnold, Gregory Winter, and George Smith

Goal of the lab

Reduce ketone using carrot peelings as reagent. Compare results to reduction with NaBH4

Enzymes can ___ ___ reduction of prochiral ketone giving __ enantiomer.

Selectively catalyze, one enantiomer

What reagent was used in the reduction?

Carrot peelings, a biocatalyst

Enantiomers are ___ that are __ -__ mirror images.

Stereoisomers, non-superimposable

ALL physical properties of ___ are the ___.

Enantiomers, the same

What is different between enantiomers?

Interaction with another chiral material or with plane-polarized light (opposite signs)

In an achiral environment reactions with achiral or racemic material gives __ or __ product (no __).

Gives achiral or racemic product (no enantioselectivity)

How to get a pure enantiomer?

Asymmetric synthesis— chiral reagents/catalysts with chiral environment

Addition of metal hydride reagents to __ yields __ __.

Aldehydes yields primary alcohols.

Reaction with metal hydrides

Nucleophilic hydride “H-” reagent adds to polarized C=O

What was the solvent and proton source for the NaBH4 reaction?

Ethanol

What enzyme do the carrots contain?

Keto-reductase enzyme

Enzymes are proteins, or polymers of __ amino acids.

Chiral amino acids.

Enzymes act as __ and in the chiral environment __ __ occur.

Catalysts, enantioselective reactions occur.

Reduction with NaBH4 intermediate

Csp2 to a Csp3 with a negative oxygen

Product of NaBH4 reduction, with ethanol

Ketone to alcohol

What is Rf?

Retention factor for TLC

How to calculate Rf values

Distance of A / Distance solvent travelled
(measured from where the spots are)

TLC mobile phase

Nonpolar organic solvent, travels UP the plate

TLC stationary phase

Thin silica gel layer, polar solid

TLC versus column chromatography

TLC is small scale and Column Chromatography is larger scale

Column mobile phase

Nonpolar organic solvent, travels DOWN the column

Column stationary phase

Silica gel layer in tube, polar solid

Principles of separation (TLC and Column)

Polar substances associate with silica gel and move slower

Nonpolar compounds travel __ and polar compounds travel __.

Nonpolar travel further/faster, Polar travel less/slower.

Equilibrium constant for stationary and mobile phase

K = A_(stationary)/ A_(mobile)

What does the polarimeter measure?

Rotation of the angle of plane-polarized light by chiral, non-racemic compounds

What is resolution?

Separation of two enantiomers, but is wasteful because half the mass is undesired

What is a prochiral ketone?

A ketone with two different R groups that can become chiral when reduced

TLC is used to determine the __ of a compound, the satus of an ongoing reaction, or as a preliminary means of __.

Purity of a compound, means of identification

How does teh mobile phase travel through the stationary phase?

Capillary action

The more __ the compound, the lower its Rf value.

Polar

A pure compound should produce a __ spot in TLC. More than that in the same vertical line indicates impurity.

A single spot

Column chromatography works well when the compounds and its impurities have __ __ __ in TLC.

Different Rf values

Column chromatography set up

Cotton plug

Column chromatography set up

Sand

Column chromatography set up

Absorbent silica gel

Column chromatography set up

Sand to hold sample

Rf depends greatly on nature of compounds and solvent: no __ values.

No standard values

In aqueous or ethanol solution, NaBH4 behaves as a source of a __ __, adding H^- to aldehydes and ketones.

Hydride nucleophile

An addition of a hydride to an aldehyde or ketone makes an __ __ that can accept a proton from water or ethanol to make the product.

Alkoxide ion

TLC

Watch glass, prevents solvent evaporation

TLC

TLC plate

TLC

Solvent front