3 Determination of Mechanism
Philosophy of mechanistic studies:
• No reaction could be determined with 100% certainty.
• One can only disproof a hypothetical mechanism, not
proof.
• As the result, an approved, last mechanism is said to be
“reasonable”, not “correct”.
• More than one method would be needed to confirm, and
their results must all be consistent.
• Gather information from many experiments until enough
to induce or extrapolate to a general conclusion.
• Occam’s razor: In the event that several hypotheses are
found to fit the facts, the simplest one is given preference.
1
3. Determination of Mechanism
3.1 Identification of products
3.2 Determination of the
presence of intermediates
3.2.1 Isolation of intermediates
3.2.2 Detection of intermediates
3.2.3 Trapping of intermediates
3.2.4 Addition of a suspected
intermediate
3.3 Study of catalysis
3.3.1 General acid catalysis
3.3.2 Specific acid catalysis
3.4 Labeling study
3.4.1 Group labeling
3.4.2 Isotope labeling
3.4.3 Crossover experiments
3.5 Isomeric selectivity study
3.5.1 Regiochemical evidences
3.5.2 Stereochemical evidences
3.6 Kinetic studies
3.6.1 Measurement of rate
3.6.2 Mechanistic information
obtained from kinetic
studies
3.6.3 Rate law
3.7 Kinetic isotope effects
3.7.1 Deuterium isotope effects
3.7.2 Primary isotope effects
3.7.3 Secondary isotope effects
3.7.4 Solvent isotope effects
2
3. Determination of Mechanism
3.1 Identification of products
Mechanism must be compatible with its products including
the by-product.
e.g. von Richter Rearrangement
COO - At the first glance, the
NO 2
mechanism was though as a
simple nucleophilic substitution
of NO2 by CN- followed by the
hydrolysis of CN- to CO2H
CN-
NO2
COOCN-
However,
Br
Br
3
Early proposed mechanism
Br
Br
Br
CN -
H
NH
CN
O
N
O
O
Br
N
O
O
Br
Br
H2O
COOH
N O
H2O
-NO 2, -NH 3
NH
O
N
O
-H +
NH
O
N O
But, from its product study, none of the NO2 or NH3 gas
was found, instead, the N2 gas was detected.
4
The mechanism was then fixed as follow:
Br
Br
Br
CN-
H
NH
CN
O
N
O
O
Br
N
O
O
Br
Br
H2 O
H 2O
-NO2 , -NH 3
COOH
N O
NH
O
N
-H +
NH
O
O
Br
Br
Br
N O
-H 2O
H 2O
-N 2
O
O
COOH
N N
O
N
NH 2
5
3.2 Determination of the presence of intermediates
3.2.1 Isolation of intermediates
Isolate the intermediate which can give the same products
when subjected to the same reaction conditions at a rate no
slower than the starting compound
e.g. Hofmann rearrangement
CH3CH2
C
NH2
O
NaOH, Br2
CH3CH2NH2
H2O
CH3CH2
N
C
H
C
C
R'
O
Neber rearrangement
R
H2
C C
N
R'
OTs
EtO
-
R
H
C
C
NH2O
R'
R
N
6
3.2 Determination of the presence of
intermediates
3.2.2 Detection of an intermediate
• In many cases, intermediate cannot be isolated but can
be detected by IR, NMR, UV-Vis or other spectra.
• Radical and triplet species can be detected by ESR and
by Chemically Induced Dynamic Nuclear Polarization
(CIDNP).
• Radicals can also be detected by cis-trans isomerization
of stilbene.
Caution: Beware of non-intermediate species and
impurities which may give interference signals.
7
3.2 Determination of the presence of
intermediates
3.2.3 Trapping of an intermediate
• In some cases, the suspected intermediate is known
to be one that reacts in a given way with a certain
compound.
• Benzynes react with dienes in the Diels-Alder
reaction
O
Br
Li
(trap)
F
O
benzyne
8
3.2 Determination of the presence of
intermediates
• Trapping an anion to determine if the elimination of
alkenes is E2 or E1cb.
ClHC CCl 2
OH E2
E1cb
ClC CCl 2
ClC CCl
D2O
(trap)
D
ClC CCl 2
9
3.2 Determination of the presence of
intermediates
• Examples of free radical trapping agents are DPPH, oxygen
(O2), triphenylmethylradical (Ph3C), nitric oxide (NO), imine
oxide, iodine, hydroquinone and dinitrobenzene.
O2N
Ph2NN
NO 2
O
PhHC N
O2N
DPPH
Imine Oxide
• A radical reaction may proceed slower in the presence of air
if the free radical intermediate can be trapped by O2.
10
3.2 Determination of the presence of
intermediates
• Kinetic requirement of intermediate trapping
A
k1
B
k2[x]
C
k2'[x']
k'[x']?
D
- The intermediate B can be efficiently trapped by X
when k2  k2.
- The detection of D does not always guarantee the
formation of B intermediate as A may directly react
11
with X to form D.
3.2 Determination of the presence of
intermediates
3.2.4 Addition of a suspected intermediate
• Perform a reaction by using a suspected intermediate
obtained by other means can be used for a negative
evidence.
e.g. von Ritcher reaction:
NO2
CO2H
von Ritcher
condition
CN
von Ritcher
condition
CO2H
12
3. Determination of Mechanism
3.3 Study of catalysis
• Mechanism must be compatible with its catalysts ,
initiator and inhibitors.
• Utilization of catalytic amount of peroxide, AIBN and
iodine usually suggests a radical mechanism.
• Kinetic study of acid-base catalyzed reaction can reveal
the rate determination step (rds.) if it is involved with the
proton transfer process
3.3.1 General acid (or base) catalysis usually indicates
that the proton transfer process is the rds.
3.3.2 Specific acid (or base) catalysis usually indicates
that the proton transfer process is not the rds.
13
3.3.1 General acid (or base) catalysis
• In general acid catalysis all species capable of donating
protons contribute to reaction rate acceleration.
• The strongest acids (SH+) are most effective (k1 is the
highest).
• Reactions in which proton transfer is rate-determining exhibit
general acid catalysis, for example diazonium coupling
reactions.
• When keeping the pH at a constant level but changing the
buffer concentration a change in rate signals a general acid
catalysis. (A constant rate is evidence for a specific acid
catalyst.)
14
3.3.2 Specific acid (or base) catalysis
• In specific acid catalysis taking place in solvent S , the reaction rate is
proportional to the concentration of the protonated solvent molecules
SH+.
• The acid catalyst itself (AH) only contributes to the rate acceleration
by shifting the chemical equilibrium between solvent S and AH in
favor of the SH+ species. S + AH  SH+ + A• For example, in an aqueous buffer solution the reaction rate for
reactants R depends on the pH of the system but not on the
concentrations of different acids.
• This type of chemical kinetics is observed when reactant R1 in a fast
equilibrium with its conjugate acid R1H+ which proceeds to react
slowly with R2 to the reaction product for example in the acid
catalyzed aldol reaction.
15
3.3 Study of catalysis
• Diazonium coupling shows general base catalysis.
Which step is the rds.?
• Aldol reaction shows specific acid catalysis.
Which step is the rds.?
16
3. Determination of Mechanism
3.4 Labeling study
3.4.1 Group labeling: Easy to obtain starting
materials but the group change may alter the
mechanism.
3.4.2 Isotope labeling: Difficult to obtain the starting
materials but no group alteration to affect the
mechanism. (Isotopic scrambling can
complicate the interpretation of the results.)
3.4.3 Crossover experiments: The experiments are
closely related to either group or isotope
labeling.
17
3.4.1 Group labeling
• Is Claisen rearrangement a [1,3] or [3,3] sigmatropic
process?
O
O
OH
Ph
OH
Ph
O
OH
Ph
Ph
18
3.4.2 Isotope labeling
• D can be detected by NMR, IR and MS
• 13C can be detected by 13C-NMR and MS
• 14C can be traced by its radio activity
• 15N can be detected by 15N-NMR
• 18O can be detected by MS
e.g.
*
*
RCN
RCOO + BrCN
O-
O
R
C
O-
N
C
R
Br
C
O
N
C
R
Br
C
O
N C O
isolated
intermediate
R
C
O
N
C
O
R
O
C + C
N
O
19
3.4.2 Isotope labeling
• Does the hydrolysis of ester proceed through
“alkyl” or “acyl” cleavage?
R
O
R'
H218O
R
O
R
OH
+ R'OH
O
18
O
O
18
R'
H2O
R
OH
+ R'18OH
Labeled water is
easier to find than
the labeled ester.
O
In these cases, the products can be easily
identified by MS.
20
Exercises
• Do the following ethanolyses of -lactone involve “alkyl” or
“acyl” cleavage?
EtOH
O
+
O
-
H or OH
EtOH
O
O
neutral
HO
OEt
O
EtO
OH
O
• Do the following hydrolyses of -lactone involve “alkyl” or
“acyl” cleavage?
H218O
O
+
O
H or OH
H218O
O
O
-
neutral
HO
OH
18
O
H18O
OH
O
21
3.4.3 Crossover Experiments
• Use for distinguishing between intra- and intermolecular
reaction
• Crossover products
+
+
indicate intermolecular
reaction.
A'
B'
B'
A'
• The method requires
No crossover product
identification of products
in the mixture.
• The method cannot
+
B
A
B
A'
A
B
distinguish between an
+
+
+
intramolecular and
“solvent cage” reactions.
A'
B'
B'
A' + B'
A
A
B
B
A
crossover
products
22
3.4.3 Crossover Experiments
• Is benzidine rearrangement an inter- or intramolecular
process?
H H
N N
H2N
OR
H H
N N
OR
NH2
RO
H 2N
OR'
H H
N N
OR'
R'O
H2N
OR
NH 2
OR'
NH 2
No crossover product indicates an intramolecular rearrangement
23
3.4.3 Crossover Experiments
• Is 1,2 rearrangement of alkyl lithium an inter- or
intramolecular process?
PhH2C
Ph
Ph
Li
CH2Li
Ph
Ph
CH2Ph
CH2
Upon an addition of14C-labeled
benzyl lithium (Ph*CH2-Li+), the
14C-labeled product was
detected, indicating an
intermolecular process.
Ph
Ph
Ph
Li
CH2Li
Ph
Ph
Ph
CH2
Upon an addition of14Clabeled phenyl lithium (Ph*Li+), no 14C-labeled product
was detected, indicating no
intermolecular process
involved.
This is called labeled fragment
addition technique
24
3. Determination of Mechanism
3.5 Isomeric selectivity study
• Selectivity = Non-statistical distribution of
products
• Specificity = Correspondence between isomeric
ratios of starting materials and products
• Level of isomeric selectivity: chemoselectivity
 regioselectivity  diastereoselectivity 
enantioselectivity
3.5.1 Regiochemical study
3.5.2 Stereochemical study
25
3.5.1 Regiochemical evidences
• HX addition on alkenes
Br
+ HBr
• Regioselectivity suggests
cationic mechanism.
• Polar solvents increase the
reaction rate supporting
the polar mechanism.
+ HBr
Br
H2O2
• Regioselectivity suggests
radical mechanism.
• Solvent polarity has no
effect on the reaction rate
supporting the radical
mechanism.
26
3.5.1 Regiochemical evidences
• Aromatic substitution by strong basic nucleophiles
Cl
NH2
NaNH 2
Possible mechanisms: SNAr or benzyne
Cl
Cl
NH 2-
NH 2
-Cl-
NH 2
(NH 2-)
-HCl
NH 2-
27
3.5.1 Regiochemical evidences
• The benzyne mechanism was supported by regiochemical
evidences obtained from group and isotope lebeling
OMe
SNAr
NH 2
OMe
+ NH2 Cl
OMe
benzyne
NH 2
14
C label
NH2
NH2
Cl
NH2-
+
1:1 ratio
28
3.5.1 Stereochemical evidences
• SN2 reaction
OTs
KOAc
and
OTs
• The reaction is stereospecific
OAc
with 100% inversion indicating
that the reaction is concerted
and the nucleophile attacks
from the back side of the
leaving group.
• The proposed transition state
OAc
is a trigonal bipyramid.
KOAc
Ph


AcO
OTs
H
CH3
29
3.5.1 Stereochemical evidences
• Neighboring group participation (NGP)
Cl
HO
HCl
Cl
+
The reaction is not stereospecific but diastereoselective. Both
diastereomers give the same major product. The results
suggest a common intermediate for all diastereomers.
The stereochemistry is
controlled by the intermediate
not by the starting material.
Ph
Which one is the major product?
30
3.5.1 Stereochemical evidences
• Neighboring group participation (NGP)
Ph
KOAc
Ph
only product
OAc
OTs
Ph
KOAc
Ph
+ enantiomer
OAc
OTs
C2

homotopic
enantiotopic
Each reaction involves
NGP in which an
intermediate with 2
reactive sites is formed.
31
3.5.1 Stereochemical evidences
• Addition
Br2
Br
Br
Anti addition in which a bromonium ion was proposed as an
intermediate.
Br
32
3.5.1 Stereochemical evidences
• Photorearrangement of spirofuran
COOMe
COOMe
h
O
OH
Possible mechanisms: pericyclic or biradical
COOMe
[1,3]
sigmatropic
COOMe
homo [1,5]
sigmatropic
OH
OH
Stereospecific product
COOMe
Racemic product
O
biradical
COOMe
COOMe
O
radical
recombination
33
OH
3. Determination of Mechanism
3.6 Kinetic studies
3.6.1 Measurements of rate
3.6.2 Mechanistic information obtained from
kinetic studies
3.6.3 Rate law
34
3.6.2 Measurement of rate
• Real Time Analysis by Periodic or Continuous Spectral
Readings
• Quenching and Analyzing
• Removal of Aliquots at Intervals
A+B
P
 1 d [ A]  1 d [ B ]
1 d [ P]




Rate 

N B dt
N p dt
N A dt
Rate  k[ A] [ B]
nA
nB
N = stoichiometric number
nA = order of reaction for reactant A
ni = order of overall reaction
(Rate Expression)
k = rate constant
kobs = rate constant directly obtained
experimentally
molecularity = number of molecules
35
come together in a single step
3.6.2 Measurement of rate
Zeroth order
A
d [ A]

 k0
dt
First order
A
k0
B
A  A0  k 0t
[B]
A0
[A]
k1
B
d [ A]

 k1[ A]
dt
A  A0 e  k1t ln A  ln A0  k1t
ln A0
ln A
slope = -k1
slope = -k0
t
36
t
3.6.2 Measurement of rate
• Second order
2A
k2
P
d [ A]

 2k 2 [ A]2
dt
1
1

 2k 2 t
[ A] [ A]0
A+B
k2
P
d [ A]

 k 2 [ A][ B ]
dt
Use pseudo first order: B0>>A0
[B] constant = B0
Treat like first order
d [ A]

 (k 2 B0 )[ A]
dt
37
3.6.3 Mechanistic information obtained
from kinetic studies
• Order of reaction can give information about which
molecules take part in rate determining step and the
previous steps.
• Changes in rate constants upon structural and condition
changes can give much information about mechanisms.
(Linear free energy relationships)
• From transition state theory, rate constants measured at
various temperature can lead to important energetic
parameters.
kr = A e-Ea/RT ;
A = kT eDS
h
/R ;
Ea = D H
+ RT
38
3.6.3 Rate law
• First order: Rate = k[A] (rds. is unimolecular process)
• Second order: Rate = k[A]2 or Rate = k[A][B]
• Order is for the whole reaction while molecularity is the
order for each step.
• Rate law depends on the rate-determining step.
– The first step is the rate-determining step:
A + B
I + B
slow
f ast
I
C
Rate = k[A][B]
39
3.6.1 Rate Law
– The first step is a rapid equilibrium:
k1
A + B
I + B
k -1
k2
I
C
Rate = -d[A]/dt = k1[A][B] - k-1[I]
d[I]/dt = k1[A][B] - k-1[I] - k2[I][B] = k1[A][B] - (k-1 + k2[B])[I]
Steady state assumption: d[I]/dt = 0
[I] = k1[A][B]/(k-1 + k2[B])
Therefore
Rate = k1[A][B] - k1k-1[A][B]/(k-1 + k2[B])
Rate = k1k2[A][B]2/(k-1 + k2[B])
For rapid equilibrium in the first step k-1[I] » k2[I][B] or k-1 » k2[B]
Thus
Rate = K1k2[A][B]2
40
Exercise
• Using the steady state assumption, derive a rate
expression for the following reaction if (a) the first step
is a rate determining step, (b) the first step is a fast
equilibrium.
A
k1
k-1
B
k2
Rate = -d[A]/dt = k1[A] - k-1[B]
d[B]/dt = k1[A] - k-1[B] - k2[B]
Steady state assumption d[B]/dt = 0
[B] = k1[A]/(k-1 + k2)
Rate = k1[A] - k-1k1[A]/(k-1 + k2)
Rate = k1k2[A ]/(k-1 + k2)
a) k-1 << k2: Rate ~ k1[A]
b) k2 << k-1: Rate ~ Kk2[A]
C
41
Exercise
• Condensations
a) CH2(COOEt)2 + CH2O
OH-
HOCH2CH(COOEt)2
Rate = k[CH2(COOEt)2] [CH2O] [OH-]
The reaction between the enolate and formaldehyde is the rds.
b) 2CH 3CHO
OH -
CH 3CH(OH)CH 2CHO
Rate = k[CH3CHO] [OH-]
The formation of the enolate is the rds.
Write a reasonable mechanism and specify the rds. of each
reaction.
42
3.7 Kinetic Isotope effects
3.7.1 Deuterium isotope effects (kH/kD) is the ratio between
the rate of reaction of the protonic substrate and that of
the corresponding deutero substrate.
• A normal isotope effect has kH/kD > 1 indicating that
the reaction of the protonic substrate is faster than the
reaction of the corresponding deutero substrate.
• An inverse isotope effect has kH/kD < 1 indicating that
the reaction of the protonic substrate is slower than
the reaction of the corresponding deutero substrate.
3.7.2 Primary isotope effect is observed in the reaction that its
rate determining step involves the breaking of the bond
connecting to the isotopic H.
• The primary isotope effects usually have 2 ≤ kH/kD ≤ 7.
43
3.7.2 Primary isotope effects
• Origin of the primary isotope effects
H X kH/kD < 7 (late T.S)
R
R
R
H
H
X maximum kH/kD ~ 7
X kH/kD < 7 (early T.S)
E0 
R H
R D
1
4
 AB 
k

m A mB
 mA
m A  mB
E 0D  E 0H
44
3.7.2 Primary isotope effects
• Alcohol oxidation
R
R
OH
+ H2CrO4
R
O
H (D)
R
Gives kH/kD = 6.9
The transition state proposed for the rds. is as follow
R O CrO 3H
R
H
base
45
Exercise
• Write a reasonable mechanism and specify the rate
determining step for the following reaction which
shows kH/kD  7
O
O
H+
CH3CCH2Br
CH3CCH3 + Br2
46
3.7 Kinetic Isotope effects
3.7.3 Secondary isotope effect is observed in the reaction that its
rate determining step does not involve the breaking of any
bond connecting to the isotopic H.
• -secondary isotope effect usually has kH/kD in the range
0.7-1.5. It is the result of the greater vibration amplitude of
C-H bond comparing to C-D bond.
– A normal -secondary isotope effect (kH/kD > 1)
generally suggests a rehybridization of the carbon
connecting to the isotopic H from sp3 to sp2 in the rate
determining step.
– An inverse -secondary isotope effect (kH/kD < 1)
generally suggests a rehybridization of the carbon
connecting to the isotopic H from sp2 to sp3 in the rate
determining step.
• -secondary isotope effect has kH/kD > 1. It is mainly 47
attributed to hyperconjugation.
3.7.3 Secondary isotope effect
• Solvolysis of cyclopentyl tosylate
OTs
H (D)
H (D)
sp3C-H
sp2C-H
(kH/kD = 1.17)
normal
• Addition on aldehyde
O
Ph
O
CNH (D)
sp2C-H
Ph
H (D)
CN
sp3C-H
(kH/kD = 0.833)
inverse
48
Summary of primary and secondary
kinetic isotope effects
(CH3)3CD + X
(CH3)2CDX
(CH3)2C=CD2 + H+
(CD3)2CHX
(CH3)3C. + DBr
+
(CH3)2CD + X+
(CH3)2CCD2H
+
(CD3)2CH + X-
primary
-secondary (normal)
-secondary (inverse)
-secondary
49
3.7 Kinetic Isotope effects
3.7.4Solvent isotope effects
Generally observed when a protic solvent e.g. D2O
or ROD is used.
• kH/kD < 1 when the reaction involves a rapid
equilibrium protonation because the acidity of
D3O+ is greater than H3O+ (specific acid catalysis
can be used for confirmation)
• kH/kD > 1 when proton transfer is the rate
determining step (general acid catalysis can be
used for confirmation)
• Secondary solvent isotope effect can interfere the
interpretation. Solvent isotope effect is thus used
50
only as a supporting evidence.
Exercise
• Write a reasonable mechanism for hydration of styrene and
predict which step is the rate determining step. Suggest 3
experiments and the expected results that can support the
proposed mechanism and rate determining step.
51