Substitution-lab

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Substitution Lab
Karolinska Institutet Department of Medical
Biochemistry and Biophysics Biomedical candidate
program, H08

• October 31st, 2008
• Craig Wheelock
• craig.wheelock_at_ki.se
• http//www.metabolomics.se/
• (slides can be downloaded from my homepage)

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Outline

• Theory
• Understand substitution reactions (SN1 vs SN2)
• Experimental equipment
• Familiarity with the necessary equipment
• Specific tips on each experiment
• Tips for conducting each experiment
• Safety issues
• Potential hazards associated with this lab
• Lab reports
• What do you need to include in your lab report??

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NUCLEOPHILIC SUBSTITUTION
NUCLEOPHILIC DISPLACEMENT
leaving group
substrate
product
nucleophile
The nucleophile displaces the leaving group.
This is a substitution reaction
Nu substitutes for X (takes its place).
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IMPORTANT
This is a reaction at sp3 (tetrahedral) carbon
atoms.
sp3
sp2
sp
yes
no
Compounds that have sp2 or sp carbons
generally do not give nucleophilic substitution
reactions.
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Nucleophilicsubstitution-reaction

• A displacement reaction of one chemical group
  to another
• R X Nu- ? R Nu X-
• Nucleophilic substitution can occur by two
  mechanisms SN1 and SN2
• Substitution Nucleophilic uni / bimolecular
• 4 main factors
• Leaving group weak bases are better (X)
• Attacking group strong bases are better (Nu-)
• Solvent protic vs. aprotic
• Sterics steric interactions affect reaction
  mechanism

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NUCLEOPHILIC SUBSTITUTION
MANY FACTORS INFLUENCE SN1 AND SN2 REACTIONS
SOME PARAMETERS
a) solvent
b) temp.
c) pH
d) DH
a) structure
a) structure of R, stereochemistry
a) nature of X
a) bond strength
b) atom used
b) atom used
b) concentration
c) concentration
c) base strength
c) bond strength
d) base strength
e) solubility
f) size
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Alkyl halides
R X

• Halides (X-) are electronegative groups that
  pull electrons through the C-X bond
• good leaving groups for substitution rxns
• C X

reactivity of halides
I gt Br gt Cl gt F
basicity
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SN2 Reaction

• Bimolecular substitution
• 2 molecules in the transition state
• - 2nd order reaction both reactants affect the
  reaction rate
• v k Nu R-X ,
  where v rate of reaction
• 
  k reaction constant
• Nu, RX concentration
  of
• 
  nucleophile, alkyl
  halide
• 
  
• Single step reaction bond breaking/forming
  simultaneously

Transition state
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SN2 Reaction

• Reactivity of alkyl halides
• Methyl gt primary gt secondary gtgt
  tertiary

large groups introduce steric hindrance
easy access no steric hindrance
R
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SN2 Reaction

• Results in inversion of configuration
• if there is a chiral center, then R S
• Supported by polar solvents that do not
  solvate the nucleophile (aprotic solvents), e.g.,
  DMSO

H
C
H
H
C
C
H
H
3
3
3
H


X
O
H
C
X
O
H
C
H
O
C
X
R
R
R
(
S
)
(
R
)
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CONCEPTUAL ANALOGY
INVERSION OF AN UMBRELLA IN THE WIND
Inversion of the umbrella is similar in
concept to the inversion of an SN2 atom.
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EFFECT OF DEGREE OF SUBSTITUTION - SN2
acetone
R
B
r

N
a
OH
N
a
B
r
R
OH

H
O
2
methyl primary secondary
tertiary
decreasing rate
EFFECT OF SUBSTRATE ON RATE
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Example SN2
CH3CH2Br NaOH
d-
d
Transition state
bromoethane
ethanol
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SN1 Reaction

• unimolecular one molecule in the transition
  state
• 1st order only concentration of the alkyl halide
  affects the rate of reaction v k R3CX
• occurs via an unstable carbocation intermediate
  R3C
• reaction occurs in several steps
• two substitution reactions and an acid-base
  reaction, deprotonation

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1st step cleavage of alkyl halide in polar
solvent
RATE LIMITING!

Unstable carbocation intermediate
Transition state 1
2nd step attack by the nucleophile and formation
of the protonated product

Transition state 2
3rd step deprotonation of the product, an
acid-base reaction
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SN1 Reaction

• results in a racemic mixture
• nucleophile can attack from either side of the
  carbocation
• mixture of R / S configuration of products

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SN1 MECHANISM
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sp2

-
planar carbocation
attacks top and bottom equally
(R)
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RACEMIZATION
(S)
enantiomers
(R)
racemic mixture
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SN1 Reaction

• activity order of alkyl halides
• tertiary gt secondary gt primary gt methyl
• in practice only occurs with tertiary
  secondary
• more stable carbocation
• more atoms share the positive charge
• activated by solvating polar solvents (protic)
  e.g., water
• stabilizes the carbocation

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CARBOCATION STABILITY
HYPERCONJUGATION
H
electrons in an adjacent C-H s bond help to
stabilize the positive charge of the carbocation
by proximity (overlap)
..

R
C
C
H
R
H
lowest energy
highest energy
ltlt
lt
tertiary
secondary
primary
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EFFECT OF INCREASING SUBSTITUTION - SN1
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RBr H2O
ROH HBr
HCOOH
methyl primary secondary
tertiary
1.0
1.7
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Guess ?
relative rate
increasing rate
rate
rel rate
rate CH3Br
EFFECT OF SUBSTRATE ON RATE
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Example SN1
Tert-butylbromide methanol (MeOH)
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SUMMARY
Notice that benzyl and allyl are good for both
SN1 and SN2
SN1
SN2
(fastest)
(fastest)
tertiary
methyl
BEST
BEST
benzyl
benzyl
allyl
allyl
In SN2 reactions benzyl is actually better
than methyl, but allyl is not.
secondary
primary
primary
secondary
tertiary
bridgehead
WORST
(bicyclic)
(slowest)
neopentyl
WORST
bridgehead
APPROXIMATE RATE ORDERS
(slowest)
(bicyclic)
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Outline of the lab

1. Substitution reaction (1 of 3 reactions)
2. Reflux to increase reaction rate
3. Monitor progress by TLC (for ethyl phenyl ether)
4. Extract the product from the reaction mix
5. Wash and dry the organic phase
6. Remove the solvent by roto-evaporation
7. Purify the product by vacuum distillation and
   record its boiling point

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Reflux
Do NOT preheat the peg-bath Use CaCl2 in
the drying tube, torkrör Use gloves with
glass wool mix well, use large magnetic
stirrer Do not let stötkoka (bounce) Use
2 neck roundbottom flask, tvåhalsad kolv
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Separatory Funnel

• Dry with Na2SO4
• - 1-2 spoons
• cover the flask
• 15-30 min
• - filter

- organic phase on top - watch out for gas
formation
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Roto-evaporation
(rullindunstning)
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Distillation
- do not use vacuum grease - measure vacuum -
start at low vacuum to prevent bouncing - foil
around the neck improves heating - use magnetic
stirrer in oil bath - weigh the flasks to
determine yield!!!!!!
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1-Bromooctane

• HBr, H2SO4
• TLC not necessary
• long reflux time of 4h, so get going!!!
• watch for gas formation during extraction
• use syringe with HBr and octanol

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n-Butylmalonic acid diethyl ester

• fill 2 neck round bottom flask with N2
• use ice-bath to cool when mixing diethyl
  malonate, bromobutane, THF and NaH
• after gas evolution stops, then reflux for 3h
• mix well
• long experiment, 3h reflux, so get going!
• no TLC needed

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n-Butylmalonic acid diethyl ester

• NaH, bromobutane (butylbromide)
• NaH reacts strongly with water!!!!
• releases H2 gas
• be careful when using ice-bath
• dry equipment!!!
• quench with acetone
• use NH3 / 95 EtOH to quench bromobutane
• test ether for peroxides
• bromobutane and diethyl malonate in hood
• use syringe to transfer bromobutane

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Ethyl phenyl ether

• phenol, iodoethane (etyljodide)
• dry equipment!!!
• measure phenol in hood, no open containers
• fill 2 necked round bottom flask with N2
• make sure that sodium ethoxide is fully dissolved
  in abs EtOH before adding phenol (30 min)
• prepare brine (saturated solution of NaCl) (for
  500 ml, 36g/100ml)
• one bottle for the whole lab is sufficient

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Ethyl phenyl ether

• Follow reaction by TLC
• collect sample prior to refluxing!!
• run TLC after 30 min
• if reaction has gone to completion, stop
  refluxing
• TLC mobile phase
• heptaneethyl acetate 91

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• For some reagents need to calculate volume from
  density . . .
• s m / V ? V m / s where s density
• 
  V volume
• 
  m mass
• densities diethyl malonate 1.055 g/ml
• 1-bromobutane 1.276 g/ml
• HBr 1.49 g/ml
• 1-octanol 0.827 g/ml
• iodoethane 1.95 g/ml
  

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Safety Issues . . .

• Peroxide-test ether (with strips), mark bottle
  when tested
• Ether is explosive do not heat!!!
• Let ether evaporate in the hood (dragskåp), do
  not put in organic waste
• Do not preheat the PEG bath
• Be careful extracting gas formation
• Dry equipment (dry overnight in drying oven)

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Safety Issues...

• use gloves with alkyl halides
• do not put them in the sink, measure in the hood
• NH3/EtOH (11) as quenching solution (motmedel)
  for alkyl halides
• prepare your own solution in the lab
• rinse all glassware that has been in contact with
  RX
• reuse the same solution
• after rinsing wash with water

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Safety continued

• weigh chemicals in hood (dragskåp)
• rinse all glassware in the hood first!
• check for residual smell from previous lab
• do not carry around open containers with
  chemicals (stinks and is dangerous)!
• can use aluminum foil to cover containers
• weigh phenol in the hood

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Lab reports

• Abstract
• experiment aim, what did you do? what did you
  see?
• Introduction
• experimental theory, pertinant chemical
  reactions, reaction mechanisms, SN1 / SN2? Draw
  the transition state
• Materials and Methods
• what did you do? include an extraction scheme,
  include lots and lots of observations!
• Results and Discussion
• how did your experiment work? what went wrong?
  what went right? draw TLC-plates with Rf-values,
  boiling points, yield (include reactant amounts),
  demonstrate understanding of experiment
• YOU ARE NOT GRADED BASED UPON YIELD

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Calculation of yield

• calculate from the limiting compound
• least amount of compound in the reaction
• yield 100 x n(product) / n(limiting compound)
• where n amount in moles
• Example a b ? c

2 mol 1 mol 0.8 mol
yield 100 x 0.8 mol / 1 mol 80
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Day of the lab . . . .

• Come prepared
• Read laboratory protocol thoroughly
• Time-consuming, so important to be familiar with
  laboratory protocol
• Perform calculations in advance
• Must wear goggles (safety glasses)
• Dont even think of eating/drinking in the lab
• Have fun . . .

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Questions? Concerns? Comments?PLEASE ASK!Good
luck!!!
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R
THE INVERSION PROCESS
2p
HO C B
sp2
partial bonding
Br
HO
C
activated complex is trigonal planar (sp2 )
CH3
H
configuration is inverted
sp3
sp3
Ea
(R)-configuration
(S)-configuration
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BENZYL ( GOOD FOR SN1 ) IS ALSO A GOOD
SN2 SUBSTRATE
primary, but faster than other primary
overlap in the activated complex lowers
the activation energy
I
H
H
Br
critical overlap