Unusual reversal of regioselectivity in antibody-mediated aldol additions with unsymmetrical methyl ketones

Article abstract of DOI:10.1016/j.tet.2003.10.115

A catalytic regio- and enantioselective aldol reaction of various unsymmetrical methyl ketones with para-nitrobenzaldehyde has been developed using aldolase antibodies as the catalysts. It has been found that the sense and level of regioselectivity for the reactions catalysed by antibody 38C2 and 33F12 are highly dependent on the structure of both the donor and the acceptor but in contrast, antibodies 84G3 and 93F3 catalyse the exclusive formation of the linear regioisomer independent of the structure of the reactants examined. The level of enantiocontrol is very high for most reactions. Both linear aldol enantiomers could be accessed through aldol or retro-aldol reactions using the same antibody. Theoretical studies on regioisomeric α- and β-heteroatom substituted enamines derived from unsymmetrical ketones suggest that most of the linear aldol products formed in the presence of antibodies 84G3 and 93F3 must be formed from intermediate enamines which are not the thermodynamically most favourable.

Full text of DOI:10.1016/j.tet.2003.10.115

Tetrahedron 60 (2004) 619–632
Unusual reversal of regioselectivity in antibody-mediated aldol
additions with unsymmetrical methyl ketones
V. Maggiotti,a S. Bahmanyar,b M. Reiter,a M. Resmini,c K. N. Houkb and V. Gouverneura
,
*
aThe Dyson Perrins Laboratory, University of Oxford, South Parks Road, Oxford OX1 3QY, UK
bDepartment of Chemistry and Biochemistry, University of California, Los Angeles, CA, 90095-1569, USA
cDepartment of Chemistry, Queen Mary, University of London, Mile End Road, London E1 4NS, UK
Received 29 August 2003; revised 30 September 2003; accepted 17 October 2003
Dedicated to Professor R.A. Lerner (The Scripps Research Institute, La Jolla, USA)
Abstract—A catalytic regio- and enantioselective aldol reaction of various unsymmetrical methyl ketones with para-nitrobenzaldehyde has
been developed using aldolase antibodies as the catalysts. It has been found that the sense and level of regioselectivity for the reactions
catalysed by antibody 38C2 and 33F12 are highly dependent on the structure of both the donor and the acceptor but in contrast, antibodies
84G3 and 93F3 catalyse the exclusive formation of the linear regioisomer independent of the structure of the reactants examined. The level of
enantiocontrol is very high for most reactions. Both linear aldol enantiomers could be accessed through aldol or retro-aldol reactions using
the same antibody. Theoretical studies on regioisomeric a- and b-heteroatom substituted enamines derived from unsymmetrical ketones
suggest that most of the linear aldol products formed in the presence of antibodies 84G3 and 93F3 must be formed from intermediate
enamines which are not the thermodynamically most favourable.
q 2003 Elsevier Ltd. All rights reserved.
1. Introduction
few direct catalytic aldol reactions with simple unsymme-
trical ketones possessing two sites of enolisation were
reported in the literature but examples of high levels of
regioselectivity are rare when the reaction pathways leading
to the regioisomers are too close in energy. We anticipate
that in more demanding cases, enzyme or antibody catalysts
can accentuate the energy difference in the regioisomeric
transition states through multiple interactions and possibly
favour the formation of the otherwise minor regioisomer.
Unfortunately, the use of natural aldolases is hindered by the
fact that enzymes usually show a strict requirement for the
donor substrate. Unlike natural enzymes, aldolase anti-
bodies were found to accept a wide range of ketone donor
substrates including small aliphatic ketones. We recently
described aldolase antibody 84G3, as a highly efficient
catalyst for the regio- and enantioselective aldol reaction
between para-nitrobenzaldehyde and four unmodified
unsymmetrical methyl ketones with the new C–C bond
formed on the less substituted a-side of these ketones. We
also demonstrated the use of 84G3 as an efficient catalyst for
the retro-aldol reaction, allowing for the kinetic resolution
of the corresponding racemic secondary aldols.9 With
antibodies 84G3, 38C2, 33F12 and 93F3, we have now
addressed four issues: (a) a study of the regioselectivity with
2-pentanone and 2-hexanone in the presence of these four
antibodies; (b) for antibodies 84G3 and 38C2, the effect on
the regioselectivity of the presence of an a- or b-heteroatom
such as oxygen, sulfur, chlorine and fluorine on the ketone;
(c) the enantioselectivity for both the forward aldol and
The development of strategies for the preparation of
enantiomerically pure aldol products remains a very
important area of research.1 An extensive number of
enantioselective aldol reactions of B, Ti, Si and Zr enolates
using stoichiometric amounts of chiral sources have been
reported.2 In addition, catalytic strategies involving pre-
formed enolate equivalents have been very promising.3
However, the development of direct catalytic asymmetric
aldol reactions starting from aldehydes and unmodified
ketones remains an exciting challenge. The first reports of
chemical catalysts for this process from the groups of
Shibasaki,4 List5 and Trost6 have recently appeared.
Complementing traditional synthetic strategies, many
diverse aldolase enzymes7 and antibodies8 have also been
involved in direct aldol reactions and have provided
efficient routes to elaborated targets including natural
products. General methods for the preparation of enantio-
merically enriched aldol products derived from unsymme-
trical ketones that could lead to two regioisomeric products
have not been developed. The simultaneous control of the
regio-, diastereo- and enantioselectivity of direct aldol
reactions involving unsymmetrical ketones constitutes one
of the most demanding challenges in synthetic chemistry. A
Keywords: Regioselectivity; Aldol additions; Methyl ketones.
*
Corresponding author. Tel./fax: þ44-1865-275-644;
e-mail address: veronique.gouverneur@chem.ox.ac.uk
0040–4020/$ - see front matter q 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2003.10.115

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V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
reverse aldol reactions as well as the determination of kinetic
parameters for these reactions; (d) theoretical calculations on
the tautomeric equilibrium of ‘imine–enamines’ of imines
derived from unsymmetrical methyl ketones.
38C2 and 33F12 are highly dependent on the structure of
both the donor and the acceptor. In contrast, antibodies
84G3 and 93F3 catalyse the exclusive formation of the
linear regioisomer independent of the structure of the
reactants examined herein (Table 1).
2.2. Effects of a- and b-heteroatom substituents on
ketone aldolisation
2. Results and discussion
2.1. Regioselectivity of the aldol reaction of 2-pentanone
and 2-hexanone with para-nitrobenzaldehyde or
3-(40-acetamidophenyl)propanal (Table 1)
Next, we focused on defining the regioselectivity in
reactions involving unsymmetrical a- or b-heteroatom
substituted ketones in the presence of antibodies 84G3 and
39C2. We examined first the aldol reactions between para-
nitrobenzaldehyde and a series of unsymmetrical ketones
possessing an a-heteroatom such as oxygen, sulfur, chlorine
or fluorine (Table 2). The presence of the heteroatom a to
the ketone greatly accelerates the spontaneous aldol
additions when compared to the corresponding all carbon
ketones. When performed under our standard conditions
(defined as above), these reactions give preferentially the
aldol products resulting from an addition at the more
substituted carbon. These results suggest that under these
conditions, thermodynamic control prevails but, for some of
these reactions, the equilibrium constants are probably not
sufficiently high to achieve a high degree of regioselectivity.
The reactions leading to the products resulting from an
addition of the aldehyde to the less substituted carbon of the
donor are the disfavoured processes under these conditions.
In order to define unambiguously the regioselectivity of
these same reactions in the presence of the antibodies, it was
critical to find screening conditions that suppress or at least
minimize any spontaneous background reaction. This was
achieved by lowering the temperature, adjusting the
concentration of the different reactants and when necessary
increasing the catalyst load. Under optimised conditions, it
was found that, in the presence of antibody 84G3,
methoxyacetone undergoes the aldol condensation with
para-nitrobenzaldehyde to afford exclusively the linear
regioisomer with a conversion of 76% after 41 h (entry 2).
Similarly, the antibody-catalysed reactions of thiomethoxy-
and chloroacetone are highly regioselective with the
preferential formation of the otherwise disfavoured linear
regioisomer resulting from an addition at the less substituted
carbon (entries 5 and 8). The conversion for chloroacetone
never exceeded 11% suggesting substrate or product
inhibition as expected from the presence of a highly reactive
electrophilic site that can chemically modify the active site
lysine residue essential to the activity of the antibody. The
reaction of fluoroacetone is much more efficient with 90%
conversion after only 30 min in the presence of 25 mol%
ab84G3 but less selective leading to a mixture of
regioisomers with nevertheless preferential formation of
the linear isomer (entry 11). Finally, hydroxyacetone proved
to be the poorest substrate for antibody 84G3 as reflected by
both the low conversion and the product distribution. After
15 h in the presence of a stoichiometric amount of ab84G3,
only 10% conversion was observed with the formation of a
1/1 mixture of the two regioisomers (entry 14). The
regioselectivity of these antibody-catalysed transformations
contrast sharply with the results obtained for the uncatalysed
reactions or in the presence of antibody 38C2. The product
distribution observed for the uncatalysed reactions is in
accordance with the reactivity expected from the presence
We set out to study the degree of regiocontrol that
antibodies 38C2, 33F12, 84G3 and 93F3 could exercise
on these reactions. For all experiments, the product
assignment and the product distribution were proven
unambiguously by comparison of the retention times with
independently chemically synthesised standards using high
performance liquid chromatography (HPLC). To screen for
catalytic activity, we performed the reactions under the
following defined conditions: 90 mM of donor, 130 mM of
acceptor and 9 mol% antibody in PBS (pH¼7.4) at room
temperature. Control experiments revealed that, in the
absence of antibody, no product formation could be detected
under these conditions (entries 1, 6, 11). When higher donor
and acceptor concentrations were used, the products were
the syn and anti stereoisomers resulting from an addition of
the more substituted carbon of 2-pentanone on the aldehyde.
These results suggest that under these conditions, the
reactions are under thermodynamic control. The antibody-
catalysed reactions were carried out using experimental
conditions under which no spontaneous reaction occurs.
Under these conditions, we found that the product
distribution is a function of the catalyst, the acceptor and
the donor. Both antibodies 38C2 and 33F12 are moderate
catalysts and afforded a mixture of regioisomeric products.
The reaction of 2-pentanone with 3-(40-acetamidophenyl)-
propanal produced predominantly the branched isomer
(entries 2 and 3) but in reaction with para-nitrobenzalde-
hyde, 2-pentanone led preferentially to the linear products
(entries 7 and 8). The reaction of 2-hexanone with para-
nitrobenzaldehyde did not produce any detectable amount
of aldol products after 69 h in the presence of up to 25 mol%
ab38C2 or ab33F12 suggesting that 2-hexanone is not a
suitable donor for these two antibodies (entries 12 and 13).
Previous reports in the literature revealed that in the
presence of ab38C2, butanone reacts with 3-(40-acetamido-
phenyl)propanal to give a 94:6 ratio of the two regioisomers
with the branched aldol product as the major compound.10
We found that in contrast to antibodies 38C2 and 33F12,
antibodies 84G3 and 93F3 catalysed the exclusive for-
mation of the linear regioisomer independent of the
structure of the aldehyde or the all carbon methyl ketone.
Indeed, all reactions inv0olving 2-pentanone with para-
nitrobenzaldehyde or 3-(4 -acetamidophenyl)propanal, and
2-hexanone with para-nitrobenzaldehyde afforded the
corresponding aldol products resulting from a reaction on
the less substituted carbon (entries 4–5, 9–10 and 14–15).
In addition, antibodies 84G3 and 93F3 are more efficient
catalysts as reflected by the reduced reaction times and the
higher conversions. These results suggest that for the all
carbon ketones that we have examined, the sense and level
of regioselectivity for the reactions catalysed by antibodies

V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
Table 1. Aldol reactions with 2-pentanone and 2-hexanone
621
Entry
1
Acceptor and donor
Conditions
Conversion (%)
Ratio A/B
PBS, 11 days
0
0/0
2
3
4
5
ab38C2, 11 days
ab33F12, 11 days
ab84G3, 11 days
ab93F3, 11 days
33
37
63
79
31/69 (lit.: 27/73)10
29/71
100/0
100/0
6
PBS, 135 h
0
0/0
7
8
9
10
ab38C2, 135 h
ab33F12, 135 h
ab84G3, 3 h
59
75
88
87
62/38
60/40
100/0
100/0
ab93F3, 2.5 h
11
PBS, 26 h
0
0/0
12
13
14
15
ab38C2, 69ha
ab33F12, 69ha
ab84G3, 26 h
ab93F3, 12 h
0
0
66
35
0/0
0/0
99/1
100/0
a
25 mol% antibody for this experiment.
of the a-heteroatom on the donor ketone (entries 1, 4, 7, 10
and 13). Indeed, for chloroacetone and thiomethoxyacetone,
it has been reported that the rates of enolisation at the
methylene carbon are about 135 times faster than at the
methyl carbon.11 Aldol reactions with hydroxyacetone have
also been reported to be highly regioselective providing for
an easy access to the synthesis of vicinal diols. In contrast,
methoxyacetone and fluoroacetone can react on both the C-1
and C-3 carbon depending on the reaction conditions.12
Previous reports in the literature have shown that in contrast
to antibody 84G3, reactions involving hydroxyacetone as
the donor were the most efficient for antibody 38C2 yielding
exclusively the syn-aldol product (entry 15).13 Fluoroace-
tone is also a substrate for antibody 38C2 and combined to
3-(40-acetamidophenyl)propanal afforded a mixture of three
products, 72% of the syn branched isomer, 21% of the trans-
branched isomer and only 7% of the linear regioisomer.10
We found that ab38C2 afforded exclusively the branched
isomer for the aldol reactions of para-nitrobenzaldehyde with
methoxyacetone and with thiomethoxyacetone (entries 3 and
6). Therefore, for these two donors, it is possible to selectively
access the branched and the linear regioisomer by using
ab38C2 and ab84G3, respectively. Antibody 84G3 is a unique
catalyst as this antibody promotes the preferential or exclusive
formation of the less substituted regioisomer except for
hydroxyacetone. This unusual reactivity could be regarded as
an example, hitherto unknown, of the controlled generation of
the less-substituted regioisomer independent of the presence
of an a-heteroatom such as O, S, Cl or F under reaction
conditions that would normally favour the formation of the
more-substituted regioisomer.
For antibody 84G3, we also studied the regioselectivity of
the reaction of para-nitrobenzaldehyde with three different
b-heteroatom substituted ketones (Table 3). Under our
standard conditions, the uncatalysed reactions proceeded
slowly with the formation of the more substituted aldol
product. We found that the presence of antibody 84G3
accelerated all three reactions and afforded exclusively the
linear regioisomer. However, the reactions are substantially
slower than the reactions involving ketones possessing an a-
heteroatom with only poor conversions after extended
reaction times despite the use of large amount of antibody.
Nevertheless, the regioselectivity is excellent.
2.3. Enantioselectivity of the aldol and retro-aldol
products and catalytic efficiency
To probe further the synthetic scope of these antibodies, we
studied the enantioselectivity of several forward aldol
reactions. The enantiomeric excesses of the products as
determined by chiral-phase HPLC (Table 4) were all superior
to 94%. All aldol products possess the (R)-configuration. To
assign the absolute configuration unambiguously, products 5,

622
V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
Table 2. Aldol reactions of para-nitrobenzaldehyde and a-heteroatom substituted methyl ketones
Entry
R
Conditions
Conversion (%)
Ratio A/B
1
2
3
OMe
OMe
OMe
PBS, 0 8C, 41 h
25% ab84G3, 0 8C, 41 h
25% ab38C2, 0 8C, 35 h
0
76
35
0/0
100/0
0/100
4
5
6
SMe
SMe
SMe
PBS, 0 8C, 1h40
25% ab84G3, 0 8C, 1h40
25% ab38C2, 0 8C, 29 h
9
56
53
10/90
98/2
1/99
7
8
Cl
Cl
PBS, 0 8C, 40 h
25% ab84G3, 0 8C, 40 h
11
11
1/99
95/5
10
11
F
F
PBS, 0 8C, 0.5 h.
25% ab84G3, 0 8C, 0.5 h
4
90
30/70
70/30
13
14
15
OH
OH
OH
PBS, rt 15 h
100% ab84G3, 0 8C, 15 h
ab38C2, rt
2
10
nda
0/100
50/50
0/100a
a
Conversion not reported, see Ref. [10].
7, 9, 19 and 21 were prepared independently by asymmetric
synthesis.14 For compounds 3, 13 and 17, the absolute
configuration has been attributed by analogy. We then studied
theutilityofantibody84G3inthekineticresolutionofracemic
linear regioisomers. The racemic aldols were treated with
ab84G3 (4 mol%) in aqueous buffer. Analysis by HPLC
indicated that in each case the retro-aldolisation reactions
halted at approximately 50% conversion. The catalyst was
highly enantioselective and provided the recovered (S)-aldols
with ee values typically greater than 91%. Both aldol
enantiomers could, therefore, be accessed through aldol or
retro-aldol reactions using the same antibody 84G3.
reactions and four retro-aldol reactions are provided
(Table 5). The kinetic parameters are reported per antibody
site assuming that both sites of the antibody are active. All
the forward aldol reactions with para-nitrobenzaldehyde
and ketones performed under pseudo-first-order conditions
showed Michaelis–Menten kinetics. The determination of
kcat/kuncat was not possible because in the uncatalysed
reactions, the formation of the linear regioisomer was
negligible under our assay conditions. The data revealed
that the introduction of a heteroatom within the ketone leads
to an increase in the KM values (entry 1 versus entries 2 and
3). Addition of an extra methylene group increases even
further the KM and results in a slower reaction rate (entry 4).
The catalytic turnovers achieved by antibody 84G3 for the
The results of the kinetic studies of four aldolisation
Table 3. Aldol reactions of para-nitrobenzaldehyde and b-heteroatom substituted methyl ketones
Entry
R
Conditions
Conversion (%)
Ratio A/B
1
2
OH
OH
PBS, 0 8C, 77 h
25% ab84G3, 0 8C, 41 h
7
40
0/100
100/0
3
4
OMe
OMe
PBS, rt, 20 h
100% ab84G3, 0 8C, 20 h
1
11
0/100
100/0
5
6
SMe
SMe
PBS, rt, 22 h
100% ab84G3, 0 8C, 22 h
9
41
0/100
95/5

V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
623
Table 4. ee Values for antibody-catalysed aldol and retro-aldol reactions
with R¼NO2Ph–
2.4. Theoretical studies on regioisomeric a- and
b-heteroatom substituted enamines derived from
unsymmetrical ketones
Product aldol
ee
(%)
Recovered product
Retro-aldol
Conversion (%)
ee (%)
To gain a better understanding of the unique reactivity of
ab84G3, we carried out a computational study to assess the
effect of various heteroatoms on the stability of regioiso-
meric enamines of unsymmetrical ketones. Lien et al.
previously carried out similar computational studies on the
substituent effects on imine–enamine tautomers of a-sub-
stituted acetaldimines XCH2CHvNH (X¼H, BH2, CH3,
NH2, OH, F, Cl, CN, NO).15 The geometries of a- and
b-heteroatom substituted regioisomeric enamines derived
from a series of unsymmetrical ketones (Scheme 1) were
computed using hybrid density functional theory (B3LYP)16
and the 6–31G* basis set17 as implemented in Gaussian
98.18 All gas phase minima were characterised by frequency
analysis. Reported electronic energies include zero point
energy corrections scaled by 0.9806.19 The optimised
geometries of four enamines derived from hydroxy,
thiomethoxy, chloro, and fluoro acetones plus methylamine
are shown in Figure 1. These include four conformations of
each enamine with the substituent on the double bond, and
four conformations with the substituent on the allylic
carbon. The intramolecular hydrogen-bonded species are
generally more stable than those lacking hydrogen-bonds,
and the more substituted enamines are more stable than
those with allylic substituents except for the enamine
derived from fluoroacetone. The more substituted enamine
is more stable by 1.1 kcal/mol for the hydroxy, 0.9 kcal/mol
for the thiomethoxy and 1.1 kcal/mol for the chloro group
when compared to the other regioisomer. In contrast, the
allylic fluoride is more stable by only 0.2 kcal/mol. The
optimised geometries of four enamines of butanone plus
methylamine and five enamines derived from hydroxy
butanones plus methylamine are shown in Figure 2. These
include four conformations of each enamine with the
substituent on the double bond, and four conformations
with the substituent on the allylic carbon. The substituted
enamine of butanone is more stable by 0.9 kcal/mol and the
hydrogen bonded allylic enamine of hydroxy butanone is
more stable by 1.5 kcal/mol when compared to the
corresponding less substituted regioisomer. Antibody
55
97
94
98
98
97
99
95
nd
96
(R)-3
(R)-5
(S)-3
(S)-5
50
94
47
97
(R)-7
(S)-7
50
96
(R)-9
(S)-9
50
94
(R)-13
(R)-17
(R)-19
(R)-21
(S)-13
(S)-17
(S)-19
(S)-21
55
91
49
95
54
99
retro-aldol reactions were higher than those of the
corresponding aldol reactions by 2.5 to 10-fold depending
on the substrate (entries 5–8). The catalytic proficiency
compares favourably with the efficiency of aldolase
antibodies with other retro-aldol reactions.
Scheme 1. (A) Various conformations of regioisomeric enamines derived from the reaction of a-heteroatom substituted propanone with methyl amine. (B)
Various conformations of regioisomeric enamines derived from the reaction of b-heteroatom substituted butanone with methyl amine.

Figure 1. Optimized geometries of the various conformations of regioisomeric enamines derived from substituted methyl ketone. Relative energies are reported in kcal/mol and distances are reported in Angstroms.

Figure 2. Optimized geometries of the various conformations of regioisomeric enamines derived from butanone and hydroxy substituted butanone. Relative energies are reported in kcal/mol and distances are reported
in Angstroms.

626
V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
Table 5. Kinetic parameters for selected aldol and retro-aldol reactions
Entry
Substrate
2-Pentanonea
K
cat (min21
)
KM (mM)
Kcat/Kuncat
(Kcat/KM)Kuncat (M21
)
1
2
3
4
5
6
7
8
0.030
0.040
0.089
0.008
0.18
0.099
0.810
0.035
28
390
217
383
79
170
69
263
–
–
–
–
–
–
Methoxyacetonea
Thiomethoxyacetonea
4-Thiomethoxybutanonea
(^)-3
–
–
1.2£105
5.5£104
4.3£105
1.9£104
1.5£109
3.2£108
6.2£109
7.4£107
(^)-7
(^)-9
(^)-21
a
With p-nitrobenzaldehyde.
84G3 is able to overcome the thermodynamic preferences
shown by these calculations. That is, many of the observed
products must be formed from intermediate enamines,
which are not the thermodynamically most favourable.
calcium hydride. Yields refer to chromatographically and
spectroscopically (1H NMR) homogeneous materials.
Commercially available reagents were used without further
purification, unless otherwise stated. Reactions were
monitored by thin-layer chromatography (TLC) carried
out on Merck aluminum foil backed sheets precoated with
Kieselgel 60F-254 using UV light as visualizing agent and
an ethanolic solution of potassium permanganate and heat as
developing agent. Merck Silica gel C60 (40–60 mM) was
used for flash column chromatography. NMR spectra were
recorded on a Bruker DPX-400 or Bruker AMX-500
spectrometer and calibrated using residual undeuterated
solvent as an internal reference. The following abbrevi-
ations were used to explain multiplicities: s, singlet; d,
doublet; t, triplet; q, quadruplet; m, multiplet; b, broad. The
coupling constants J are given in hertz. IR spectra were
recorded on a Perkin–Elmer Paragon 1000 FT-IR spec-
trometer. Mass spectra (m/z) and HRMS were recorded on
Micromass GCT using Chemical Ionisation (NH3, CI),
Electronic Impact (EIþ) or Field Ionization (FI). Micro-
analyses were performed by ‘Elemental Microanalysis
Limited’, Devon. Melting points were determined in a
capillary and are uncorrected.
3. Conclusion
Two major issues were under consideration. First was the
critical issue as to whether aldolase antibodies 84G3 and
38C2 could differentiate the two reactive sites of unsymme-
trical methyl ketones. Secondly, the possibility of control-
ling simultaneously the regio- and enantioselectivity of
aldol reactions with a variety of unsymmetrical methyl
ketones had to be determined. We have identified important
differences between antibodies 38C2 and 84G3. As an
overall trend, antibody 38C2 favours the formation of the
branched regioisomer resulting from a reaction on the more
substituted carbon whereas antibody 84G3 leads preferen-
tially to the formation of the linear isomer. Therefore, the
ab84G3-catalysed aldols and retro-aldolisations provide a
rapid entry to enantioenriched linear aldol products that are
difficult to access with other catalysts. Both enantiomers are
accessible by using both the forward aldol and retro-aldol
reactions. The retro-aldolisations are more efficient than the
synthetic aldols as reflected by the higher rate constants. The
unique reactivity of antibody 84G3 is quite remarkable
since, to our knowledge, no known catalysts display similar
level of regio- and enantiocontrol in the presence of the
ketones used for this study. This unusual reactivity further
demonstrates that, through a combination of immunological
diversity and basic chemical principles, efficient catalysts
that display new reactivity can be created. Further work is in
progress to address the question of whether L-proline is
capable of catalysing the aldol reactions of para-nitrobenz-
aldehyde and various unsymmetrical methyl ketones and
assesses the degree of regio- and stereocontrol exercised by
this catalyst. This study aims to compare the synthetic value
of the antibody-with the L-proline-catalysed reactions.
4.2. Synthesis of racemic aldol products. Path A: direct
cross aldolizations with LDA
To a solution of LDA, prepared by dropwise addition of
n-butyllithium (1.1 mmol) to a solution of diisopropylamine
(1.2 mmol) in dry THF (2 mL) at 0 8C, was added a solution
of ketone (1 mmol) in THF (2 mL) at 278 8C. After the
resulting mixture was stirred at 278 8C for 30 min, the
aldehyde (1 mmol) in THF (1 mL) was added. The reaction
was quenched after 3 h at 278 8C by addition of saturated
NH4Cl. The mixture was extracted three times with AcOEt,
the combined organic layers were dried over MgSO4 and
solvents were removed in vacuo. Product ratios were
determined by crude 1H NMR spectroscopy before
purification by flash column chromatography.
4.2.1. 1-(40-Nitrophenyl)-1-hydroxy-3-hexanone 3. Crude
ratio regio/syn/anti¼78/13/9. Purification by column chro-
matography over silica (hexane/AcOEt: 80/20), followed by
recrystallization from AcOEt yielded 3 as a white solid
(38% pure and 18% in mixture with 4 (syn and anti). Rf 0.16
(hexane/AcOEt, 80/20); mp 59 8C; 1H NMR (400 MHz,
CDCl3) d 8.18 (d, J¼8.6 Hz, 2H), 7.52 (d, J¼8.6 Hz, 2H),
5.25 (m, 1H), 3.50 (bs, 1), 2.80 (m, 2H), 2.43 (t, J¼7.4 Hz,
2H), 1.6 (tq, J¼7.4, 7.4 Hz, 2H), 0.90 (t, J¼7.4 Hz, 3H); 13C
NMR (100 MHz, CDCl3) d 211.0, 150.2, 147.2, 126.4,
4. Experimental
4.1. General
All reactions were carried out under an argon atmosphere
with dry solvents under anhydrous conditions, unless
otherwise noted. Dry tetrahydrofuran (THF) was obtained
by distillation over sodium and benzophenone, dry methyl-
ene chloride (CH2Cl2) was obtained by distillation over

V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
627
123.7, 69.0, 50.5, 45.4, 17.0, 13.6; IR (neat) nmax 3516,
1702, 1515, 1346, 1088; Anal. calcd for C12H15NO4: C,
60.75; H, 6.37; N, 5.90. Found: C, 60.82; H, 6.36; N, 5.98;
HRMS (EIþ): Mþ 237.1001 (expected), 239.1009
(observed).
CDCl3) d 208.5, 149.9, 147.2, 126.3, 123.7, 77.7, 68.7, 59.3,
47.2; IR (neat) nmax 3436, 2936, 1726, 1519, 1348, 1088;
Anal. calcd for C11H13NO5: C, 55.23; H, 5.48; N, 5.86.
Found: C, 55.01; H, 5.57; N, 5.83; HRMS (FI): Mz 239.0794
(expected), 239.0789 (observed).
4.2.2. 1-(40-Nitrophenyl)-1-hydroxy-3-heptanone 5.
Crude ratio regio/syn/anti¼78/13/9. Purification by column
chromatography over silica (AcOEt/hexane: 30/70)
afforded 5 as a colorless oil (26% pure and 23% in mixture
Analytical data for syn-8. Rf 0.25 (AcOEt/hexane: 40/60);
1H NMR (500 MHz, CDCl3) d 8.23 (d, J¼8.7 Hz, 2H), 7.57
(d, J¼8.7 Hz, 2H), 5.06 (dd, J¼6.8 and 4.1 Hz, 1H), 3.77 (d,
J¼4.1 Hz, 1H), 3.37 (s, 3H), 3.15 (d, J¼6.8 Hz, 1H), 2.20
(s, 3H); 13C NMR (125 MHz, CDCl3) d 209.8, 147.5, 147.2,
127.1, 123.5, 89.8, 73.1, 59.6, 27.5; IR (neat) nmax 3511,
1722, 1518, 1350, 1111; HRMS (FI): Mz 239.0794
(expected), 239.0793 (observed).
1
with 6 (syn and anti). Rf 0.28 (AcOEt/hexane: 30/70); H
NMR (400 MHz, CDCl3) d 8.16 (d, J¼8.8 Hz, 2H), 7.52 (d,
J¼8.8 Hz, 2H), 5.25 (m, 1H), 3.80 (d, J¼2.4 Hz, 1H), 2.81
(m, 2H), 2.44 (t, J¼7.6 Hz, 2H), 1.55 (m, 2H), 1.28 (m, 2H),
0.88 (t, J¼7.6 Hz, 3H); 13C NMR (100 MHz, CDCl3) d
211.1, 150.2, 147.2, 126.4, 123.7, 69.0, 50.5, 43.3, 25.5,
22.2, 13.8; IR (neat) nmax 3466, 2959, 1709, 1520, 1347,
1081; HRMS (EIþ): Mþ 251.1158 (expected), 251.1162
(observed).
Analytical data for anti-8. Rf 0.29 (AcOEt/hexane: 40/60);
1H NMR (500 MHz, CDCl3) d 8.23 (d, J¼8.7 Hz, 2H), 7.57
(d, J¼8.7 Hz, 2H), 5.03 (dd, J¼6.2, 3.7 Hz, 1H), 3.71 (d,
J¼6.2 Hz, 1H), 3.33 (s, 3H), 3.14 (d, J¼3.7 Hz, 1H), 2.17
(s, 3H); 13C NMR (125 MHz, CDCl3) d 210.4, 147.1, 128.1,
123.9, 90.0, 73.7, 60.1, 27.9; IR (neat) nmax 3398, 1708,
1517, 1347, 1108; HRMS (FI): Mz 239.0794 (expected),
239.0792 (observed).
4.2.3. 8-(40-Acetamidophenyl)-6-hydroxy-4-octanone 1.
Crude ratio regio/syn/anti¼85/8/7. Purification by column
chromatography over silica (Et2O/AcOEt: 80/20), followed
by recrystallization from hexane/DCM afforded 1 as a white
solid (48%); mp 96 8C. Rf 0.22 (Et2O/AcOEt: 80/20); H
4.2.6. 4-(40-Nitrophenyl)-4-hydroxy-1-methylsulfanyl-2-
butanone 9. Crude ratio regio/syn/anti¼35/20/45. Purifi-
cation by column chromatography over silica (CH2Cl2/
Et2O: 98/2), followed by recrystallization from AcOEt
afforded 9 as a pale yellow solid (10%). Rf 0.19 (DCM/
Et2O: 98/2); mp 86 8C; 1H NMR (400 MHz, CDCl3) d 8.21
(d, J¼8.6 Hz, 2H), 7.57 (d, J¼8.6 Hz, 2H), 5.28 (t,
J¼6.4 Hz, 1H), 3.57 (s, 1H), 3.19 (s, 2H), 3.05 (d,
J¼6.4 Hz, 2H), 2.05 (s, 3H); 13C NMR (100 MHz,
CDCl3) d 204.7, 149.9, 147.3, 126.5, 123.7, 69.3, 48.0,
43.4, 15.6; IR (neat) nmax 3468, 2921, 1706, 1519, 1348,
1063; Anal. calcd for C11H13NO4S: C, 51.75; H, 5.13; N,
5.49. Found: C, 51.96; H, 5.19; N, 5.49.
1
NMR (400 MHz, CDCl3) d 7.56 (bs, 1H), 7.39 (d,
J¼8.0 Hz, 2H), 7.12 (d, J¼8.0 Hz, 2H), 4.02 (m, 1H),
3.30 (d, J¼2.8 Hz, 1H), 2.76 (m, 1H), 2.63 (m, 1H), 2.55 (m,
2H), 2.39 (t, J¼7.2 Hz, 2H), 2.14 (s, 3H), 1.77 (m, 1H), 1.59
(m, 1H), 1.59 (tq, J¼7.2, 7.2 Hz, 2H), 0.91 (t, J¼7.2 Hz,
3H); 13C NMR (100 MHz, CDCl3) d 212.5, 168.5, 137.9,
135.7, 128.9, 120.0, 66.7, 48.9, 45.5, 38.0, 31.1, 24.4, 17.0,
13.6; IR (neat) nmax 3304, 2962 and 2932, 1705, 1667, 1603,
1538 and 1515; HRMS (EIþ): Mþ 277.1678 (expected),
277.1672 (observed).
4.2.4. 4-(40-Nitrophenyl)-1,4-dihydroxy-2-butanone 15.
2.2 equiv. of diisopropylamine and 2.1 equivalent of
n-BuLi were used. Purification by column chromatography
over silica (DCM/AcOEt: 80/20) afforded 15 as a colourless
oil (25%). Rf 0.12 (DCM/AcOEt: 80/20); 1H NMR
(400 MHz, CD3OD) d 8.21 (d, J¼8.4 Hz, 2H), 7.63 (d,
J¼8.4 Hz, 2H), 5.28 (dd, J¼8.9, 4.2 Hz, 1H), 4.26 (d,
J¼18.7 Hz, 1H), 4.22 (d, J¼18.7 Hz, 1H), 2.88 (dd, J¼15.8,
8.9 Hz, 1H), 2.75 (dd, J¼15.8, 4.2 Hz, 1H); 13C NMR
(100 MHz, CD3OD) d 208.5, 152.2, 147.5, 126.8, 123.4,
68.9, 68.5, 47.7; IR (neat) nmax 3413, 2913, 1703, 1512,
1348, 1035; HRMS (FI): Mz 225.0637 (expected), 225.0638
(observed).
4.2.7. 1-(40-Nitrophenyl)-1,5-dihydroxy-3-pentanone 17
and 4-(40-nitrophenyl)-4-hydroxy-3-hydroxymethyl-3-
butanone 18. Crude ratio regio/syn/anti¼40/23/37. Purifi-
cation by column chromatography over silica (AcOEt)
followed by (toluene/AcOEt: 40/60) or recrystallization
from AcOEt afforded 17 and 18-D2 as pale yellow solids
(22 and 9%, respectively) and 18-D1 as a pale yellow oil
(7%).
Analytical data for 17: mp 63 8C. Rf 0.35 (AcOEt); 1H NMR
(250 MHz, CDCl3) d 8.22 (d, J¼8.9 Hz, 2H), 7.55 (d,
J¼8.9 Hz, 2H), 5.32 (m, 1H), 3.91 (dt, J¼6.0, 5.4 Hz, 2H),
3.56 (d, J¼3.4 Hz, 1H), 2.89 (m, 2H), 2.74 (t, J¼5.4 Hz,
2H), 2.33 (t, J¼6.0 Hz, 1H); 13C NMR (100 MHz, CDCl3) d
210.6, 149.8, 147.3, 126.4, 123.8, 68.9, 57.6, 51.4, 45.4; IR
(neat) nmax 3388, 2897, 1710, 1605, 1518, 1348, 1056; Anal.
calcd for C11H13NO5: C, 55.23; H, 5.48; N, 5.86. Found: C,
55.24; H, 5.51; N, 5.82; HRMS (FI): Mz 239.0794
(expected), 239.0800 (observed).
4.2.5. 4-(40-Nitrophenyl)-4-hydroxy-1-methoxy-2-buta-
none 7 and 4-(40-nitrophenyl)-4-hydroxy-3-methoxy-2-
butanone 8 (syn and anti). Crude ratio regio/syn/anti¼25/
50/25. Purification by column chromatography over silica
(AcOEt/hexane: 40/60) and (DCM/Et2O: 90/10) afforded 7
as a white solid (8%) and syn-8 and anti-8 as pale yellow
oils (6 and 23%, respectively).
Analytical data for 7. Rf 0.20 (AcOEt/hexane: 40/60); mp
1
Analytical data for 18-D1. Rf 0.31 (toluene/AcOEt: 40/60);
1H NMR (400 MHz, CDCl3) d 8.25 (d, J¼8.8 Hz, 2H), 7.64
(d, J¼8.8 Hz, 2H), 4.98 (d, J¼8.6 Hz, 1H), 3.62 (dd, 1H,
J¼10.8, 8.6 Hz), 3.38 (dd, J¼10.8, 4.4 Hz, 1H), 3.07 (ddd,
J¼8.6, 8.6, 4.4 Hz, 1H), 2.28 (s, 3H); 13C NMR (125 MHz,
78 8C; H NMR (500 MHz, CDCl3) d 8.21 (d, J¼7.0 Hz,
2H), 7.55 (d, J¼7.0 Hz, 2H), 5.31 (t, J¼5.0 Hz, 1H), 4.05
(d, J¼13.8 Hz, 1H), 4.01 (d, J¼13.8 Hz, 1H), 3.48 (s, 1H),
3.42 (s, 3H), 2.90 (d, J¼5.0 Hz, 2H); 13C NMR (125 MHz

628
V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
1
CDCl3) d 211.6, 151.0, 147.9, 127.8, 123.5, 72.0, 62.3, 60.7,
30.9; IR (neat) nmax 3426, 2897, 1710, 1606, 1520, 1348,
1063, 1004; HRMS (CIþ): MþNHþ4 257.1137 (expected),
257.1132 (observed).
47 8C; H NMR (500 MHz, CDCl3) d 8.23 (d, J¼7.9 Hz,
2H), 7.56 (d, J¼7.9 Hz, 2H), 5.31 (m, 1H), 3.48 (d,
J¼3.3 Hz, 1H), 2.87 (d, J¼6.5 Hz, 2H), 2.77 (m, 4H), 2.12
(s, 3H); 13C NMR (125 MHz, CDCl3) d 208.7, 149.7, 147.3,
126.3, 123.7, 68.8, 51.0, 43.0, 27.6, 15.7; IR (neat) nmax
3423, 2919, 1710, 1603, 1519, 1347, 1077; HRMS (FI): Mz
269.0722 (expected), 269.0719 (observed).
Analytical data for 18-D2: deduced from a 13/87 mixture of
D1/D2. Rf¼0.21 (toluene/AcOEt: 40/60); 1H NMR
(400 MHz, CDCl3) d 8.22 (d, J¼8.4 Hz, 2H), 7.60 (d,
J¼8.4 Hz, 2H), 5.04 (d, J¼7.6 Hz, 1H), 3.97 (dd, J¼11.2,
7.6 Hz, 1H), 3.88 (dd, J¼11.2, 4.4 Hz, 1H), 3.17 (ddd,
J¼7.6, 7.6, 4.4 Hz, 1H), 2.07 (s, 3H); 13C NMR (125 MHz,
CDCl3) d 210.7, 151.1, 147.7, 127.8, 123.4, 71.6, 62.2, 60.4,
30.9; IR (neat) nmax 3425, 2897, 1710, 1606, 1521, 1348,
1063, 1004; HRMS (CIþ): MþNHþ4 257.1137 (expected),
257.1132 (observed).
Analytical data for syn-22. Rf 0.29 (hexane/Et2O: 40/60);
1H NMR (400 MHz, CDCl3) d 8.22 (d, J¼8.7 Hz, 2H),
7.55 (d, J¼8.7 Hz, 2H), 5.14 (d, J¼4.4 Hz, 1H), 3.31 (s,
1H), 3.13 (ddd, J¼9.6, 4.4, 4.4 Hz, 1H), 2.81 (dd,
J¼13.4, 9.6 Hz, 1H), 2.66 (dd, J¼13.4, 4.4 Hz, 1H), 2.24
(s, 3H), 2.00 (s, 3H); 13C NMR (125 MHz, CDCl3) d
211.8, 148.8, 147.9, 127.5, 124.2, 73.1, 58.1, 32.9, 31.5,
16.9; IR (neat) nmax 3429, 2919, 1708, 1519, 1347;
HRMS (CIþ): MþNHþ4 287.1066 (expected), 287.1064
(observed).
4.2.8. 1-(40-Nitrophenyl)-1-hydroxy-5-methoxy-3-penta-
none 19 and 4-(40-nitrophenyl)-4-hydroxy-3-methoxy-
methyl-2-butanone 20 (syn and anti). Crude ratio regio/
syn/anti¼34/33/33. Purification by column chromatography
over silica (DCM/AcOEt: 80/20) followed by (DCM)
afforded 19 as a colourless oil (20%) and 20-syn and
20-anti as pale yellow oils (20 and 23%, respectively).
Analytical data for anti-22: Analyses are deduced from a 68/
1
32 mixture of syn/anti. Rf 0.25 (hexane/Et2O: 40/60); H
NMR (400 MHz, CDCl3) d 8.23 (d, J¼9.0 Hz, 2H), 7.53 (d,
J¼9.0 Hz, 2H), 5.06 (d, J¼6.0 Hz, 1H), 3.45 (s, 1H), 3.14
(m, 1H), 2.72 (dd, J¼13.0, 8.0 Hz, 1H), 2.63 (dd, J¼13.0,
6.4 Hz, 1H), 2.17 (s, 3H), 2.10 (s, 3H); 13C NMR (125 MHz,
CDCl3) d 212.3, 149.7, 148.8, 127.4, 124.3, 74.2, 58.0, 33.9,
33.2, 16.9; IR (neat) nmax 3430, 2919, 1709, 1520, 1348;
HRMS (CIþ): MþNHþ4 287.1066 (expected), 287.1062
(observed).
1
Analytical data for 19. Rf 0.27 (DCM/AcOEt: 80/20); H
NMR (500 MHz, CDCl3) d 8.22 (d, J¼8.8 Hz, 2H), 7.55 (d,
J¼8.8 Hz, 2H), 5.29 (m, 1H), 3.67 (m, 3H), 3.35 (s, 3H),
2.88 (d, J¼7.2 Hz, 2H), 2.71 and 2.72 (2t, J¼6.0 Hz, 2H);
13C NMR (125 MHz, CDCl3) d 209.7, 150.4, 147.8, 126.9,
124.2, 69.4, 67.9, 59.4, 52.1, 43.9; IR (neat) nmax 3470,
2857, 1710, 1516, 1345, 1106; HRMS (FI): Mz 253.0950
(expected), 253.0955 (observed).
4.3. Synthesis of racemic aldols. Path B: direct cross
aldolisations with NaOH
Analytical data for anti-20. Rf 0.16 (DCM); 1H NMR
(400 MHz, CDCl3) d 8.20 (d, J¼8.6, 2H), 7.52 (d,
J¼8.6 Hz, 2H), 5.24 (dd, J¼4.4, 3.0 Hz, 1H), 3.94 (d,
J¼3.0 Hz, 1H), 3.66 (dd, 1H, J¼9.5, 5.2 Hz), 3.59 (dd,
J¼9.5, 6.2 Hz, 1H), 3.29 (s, 3H), 3.06 (m, 1H), 2.15 (s, 3H);
13C NMR (100 MHz, CDCl3) d 209.4, 149.1, 147.3, 126.9,
123.6, 72.6, 70.3, 59.3, 58.3, 31.2; IR (neat) nmax 3424,
2928, 1711, 1521, 1343, 1111; HRMS (FI): Mz 253.0950
(expected), 253.0956 (observed).
A 1% solution of sodium hydroxide in water (5.5 mL,
1.4 mmol) was added to the ketone (132 mmol) at RT. The
aldehyde (6.6 mmol) was then added. After 3 h at RT, the
layers were separated and the aqueous layer was extracted
three times with AcOEt, the combined organic layers were
washed with saturated NH4Cl, dried over MgSO4 and
solvents were removed in vacuo. Product ratios were
determined by crude 1H NMR spectroscopy before
purification by flash column chromatography.
Analytical data for anti-20: Analyses are deduced from a 10/
90 mixture of syn/anti. Rf 0.16 (DCM); 1H NMR (400 MHz,
CDCl3) d 8.20 (d, J¼8.6 Hz, 2H), 7.51 (d, J¼8.6 Hz, 2H),
5.12 (d, J¼7.2 Hz, 1H), 3.72 (bs, 1H), 3.49 (dd, J¼9.4,
4.2 Hz, 1H), 3.34 (dd, J¼9.4, 6.4 Hz, 1H), 3.24 (s, 3H), 3.04
(ddd, J¼7.2, 6.4, 4.2 Hz, 1H), 2.20 (s, 3H); 13C NMR
(100 MHz, CDCl3) d 211.0, 149.2, 147.4, 127.1, 123.7,
71.9, 70.9, 59.2, 59.0, 31.4; IR (neat) nmax 3420, 2929, 2897,
1711, 1521, 1348, 1118; HRMS (CIþ): MþNHþ4 271.1290
(expected), 271.1294 (observed).
4.3.1. 3-[Hydroxy-(40-nitrophenyl)-methyl]-2-pentanone
4 (syn and anti). Crude ratio regio/syn/anti¼16/35/49.
Purification by column chromatography over silica (hexane/
AcOEt: 80/20) followed by semi-preparative HPLC (C8
column) afforded syn-4 as a colourless oil and anti-4 as a
white solid (total yield as a mixture with anti-4, 60%).
Analytical data for syn-4. Rf 0.16 (hexane/AcOEt: 80/20);
1H NMR (500 MHz, CDCl3) d 8.22 (d, J¼8.9 Hz, 2H), 7.54
(d, J¼8.9 Hz, 2H), 5.11 (d, J¼4.5 Hz, 1H), 3.25 (bs, 1H),
2.84 (ddd, J¼9.0, 9.0, 4.5 Hz, 1H), 2.19 (s, 3H), 1.74
(m, 1H), 1.59 (m, 1H), 0.86 (t, J¼7.0 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 213.4, 149.4, 147.5, 127.2, 123.8,
72.6, 59.8, 32.0, 19.7, 12.4; IR (neat) nmax 3409, 2967, 1702,
1521, 1348, 1044; HRMS (CIþ): MþNHþ4 255.1345
(expected), 255.1347 (observed).
4.2.9. 1-(40-Nitrophenyl)-1-hydroxy-5-methylsulfanyl-3-
pentanone 21 and 4-(40-nitrophenyl)-4-hydroxy-3-
methylsulfanyl-3-butanone 22 (syn and anti). Crude
ratio regio/syn/anti¼74/13/13. Purification by column
chromatography over silica (hexane/Et2O: 40/60) afforded
21 as a yellow solid (40%) and 22-syn and 22-anti as pale
yellow oils (3.4 and 5%, respectively).
Analytical data for anti-4. Rf 0.16 (hexane/AcOEt: 80/20);
1H NMR (400 MHz, CDCl3) d 8.23 (d, J¼8.8 Hz, 2H), 7.52
(d, J¼8.8 Hz, 2H), 4.93 (d, J¼6.0 Hz, 1H), 3.26 (bs, 1H),
Analytical data for 21. Rf 0.21 (hexane/Et2O: 40/60); mp

V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
629
2.88 (ddd, J¼8.0, 6.0, 6.0 Hz, 1H), 2.14 (s, 3H), 1.63 (m,
1H), 1.52 (m, 1H), 0.93 (t, J¼7.0 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 213.7, 150.1, 147.7, 127.3, 124.0,
74.3, 60.1, 32.4, 22.7, 11.7; IR (neat) nmax 3401, 2922, 1702,
1522, 1347; HRMS (CIþ): MþNHþ4 255.1345 (expected),
255.1343 (observed).
4.4.2. 4-(40-Nitrophenyl)-4-hydroxy-3-methylsulfanyl-2-
butanone 10 (syn and anti). Crude ratio regio/syn/anti¼5/
55/40. Purification by column chromatography over silica
(CH2Cl2/Et2O: 98/2), followed by recrystallization from
AcOEt afforded syn-10 and anti-10 as pale yellow solids (8
and 16%, respectively).
4.3.2. 3-(Hydroxy-(40-nitrophenyl)-methyl)-2-hexanone
6 (syn and anti). Crude ratio regio/syn/anti¼20/33/47.
Purification by column chromatography over silica (toluene/
AcOEt: 95/5) afforded syn-6 and anti-6 as colourless oils
(22 and 49%, respectively).
Analytical data for syn-10. Rf 0.35 (CH2Cl2/Et2O: 98/2); mp
73 8C; 1H NMR (400 MHz, CDCl3) d 8.19 (d, J¼8.7 Hz, 2
H), 7.58 (d, J¼8.7 Hz, 2 H), 5.16 (d, J¼7.8 Hz, 1 H), 3.51
(bs, 1 H), 3.45 (d, J¼7.8 Hz, 1 H), 2.22 (s, 3 H), 2.06 (s, 3
H); 13C NMR (100 MHz, CDCl3) d 202.8, 147.8, 147.6,
127.9, 123.5, 69.1, 59.9, 29.1, 12.3; IR (neat) nmax
3478, 1701, 1520, 1348, 1064; Anal. calcd for
C11H13NO4S: C, 51.75; H, 5.13; N, 5.49. Found: C, 51.70;
H, 5.13; N, 5.47.
Analytical data for syn-6. Rf 0.22 (toluene/AcOEt: 95/5); 1H
NMR (400 MHz, CDCl3) d 8.21 (d, J¼8.8 Hz, 2H), 7.52
(d, J¼8.8 Hz, 2H), 5.09 (d, J¼4.2 Hz, 1H), 3.32 (bs, 1H),
2.88 (ddd, J¼8.8, 4.2 and 4.2, 1H), 2.17 (s, 3H), 1.70–
1.60 (m, 1H), 1.51–1.42 (m, 1H), 1.34–1.20 (m, 1H),
1.17–1.04 (m, 1H), 0.82 (t, J¼7.3 Hz, 3H); 13C NMR
(100 MHz, CDCl3) d 213.3, 149.2, 147.3, 126.9, 123.5,
72.5, 58.1, 31.6, 28.4, 21.1, 14.2; IR (neat) nmax 3456,
1703, 1521, 1348; HRMS (CIþ): MþNHþ4 269.1501
(expected), 269.1498 (observed).
Analytical data for anti-10. Rf 0.16 (CH2Cl2/Et2O: 98/2);
mp 129 8C; 1H NMR (400 MHz, CDCl3) d 8.23 (d,
J¼8.8 Hz, 2 H), 7.59 (d, J¼8.8 Hz, 2 H), 5.12 (d,
J¼9.2 Hz, 1 H), 3.42 (d, J¼9.2 Hz, 1 H), 3.20 (bs, 1 H),
2.41 (s, 3 H), 1.93 (s, 3 H); 13C NMR (100 MHz, CDCl3) d
204.5, 147.7, 147.6, 127.8, 123.5, 72.1, 58.7, 28.6, 13.0; IR
(neat) nmax 3386, 1698, 1510, 1343, 1034; Anal. calcd for
C11H13NO4S: C, 51.75; H, 5.13; N, 5.49; O, 25.07. Found:
C, 51.84; H, 5.14; N, 5.36; O, 25.13.
Analytical data for anti-6. Rf 0.16 (toluene/AcOEt: 95/5);
1H NMR (400 MHz, CDCl3) d 8.21 (d, J¼8.7 Hz, 2H), 7.50
(d, J¼8.7 Hz, 2H), 4.88 (d, J¼6.8 Hz, 1H), 3.43 (bs, 1H),
2.92 (ddd, J¼8.7, 6.8, 5.3 Hz, 1H), 2.13 (s, 3H), 1.56 (m,
1H), 1.28 (m, 3H), 0.86 (t, J¼8.1 Hz, 3H); 13C NMR
(100 MHz, CDCl3) d 213.5, 150.0, 147.4, 127.1, 123.7,
74.4, 58.3, 32.1, 31.4, 20.4, 14.0; IR (neat) nmax 3444, 1711,
1523, 1348; HRMS (FI): Mz 251.1158 (expected), 251.1164
(observed).
4.4.3. 4-(40-Nitrophenyl)-3-chloro-4-hydroxy-2-butanone
12 (syn and anti). Crude ratio regio/syn/anti¼1/59/40.
Purification by column chromatography over silica (DCM/
Et2O: 98/2) afforded 12 as a colourless oil (43%) in 3:2 ratio of
syn:anti. Analyses deduced from a mixture of 60% of syn and
40% of anti. Rf 0.20 (DCM/Et2O: 98/2); 1H NMR (400 MHz,
CDCl3) syn d8.25 (d, J¼8.3 Hz, 2H), 7.59 (d, J¼8.3 Hz, 2H),
5.45 (dd, J¼4.0, 4.0 Hz, 1H), 4.44 (d, J¼4.0 Hz, 1H), 3.20 (d,
J¼4.0 Hz, 1H), 2.40 (s, 3H); anti d 8.25 (d, J¼8.4 Hz, 2H),
7.61 (d, J¼8.4 Hz, 2H), 5.16 (dd, J¼8.0, 4.2 Hz, 1H), 4.28 (d,
J¼8.0 Hz, 1H), 3.35 (d, J¼4.2 Hz, 1H), 2.41 (s, 3H); 13C
NMR (100 MHz, CDCl3) syn d 203.3, 147.8, 146.1, 127.4,
123.6, 72.0, 67.1, 28.3; anti d 203.3, 148.0, 145.8, 128.1,
123.6, 73.8, 63.4, 28.0; IR (neat) nmax 3441, 1718, 1606, 1521,
1349; HRMS of the mixture (CIþ): MþNHþ4 261.0642
(expected), 261.0638 (observed).
4.4. Synthesis of racemic aldols. Path C: direct cross
aldolisations in presence of PBS pH57.4
The ketone (1.45 mmol) was added to a solution of
4-nitrobenzaldehyde (140 mg, 0.95 mmol) in DMF
(15 mL). PBS pH¼7.4 (20 mL) was then added and the
resulting solution was stirred at RT or at reflux for 48 h. The
mixture was extracted three times with AcOEt, dried
over MgSO4 and solvents were removed in vacuo.
Product ratios were determined, when possible, by crude
1H NMR spectroscopy before purification by flash
chromatography.
4.4.4. 4-(40-Nitrophenyl)-3-fluoro-4-hydroxy-2-butanone
14 (syn and anti). Purification by column chromatography
(two fold) over silica (hexane/Et2O: 60/40, followed by
pentane/Et2O: 50/50) afforded 14 as a pale yellow solid in
3:4 ratio of syn:anti (15%). Analyses deduced from a
mixture of 43% of syn and 57% of anti. Rf 0.24 (pentane/
4.4.1. 4-(40-Nitrophenyl)-3,4-dihydroxy-2-butanone 16
(syn and anti). Crude ratio regio/syn/anti¼3/48/49. Purifi-
cation by column chromatography over silica (DCM/AcOEt:
80/20) and recrystallization from AcOEt afforded a mixture of
syn-16 and anti-16 as a yellow solid (40%). Analyses deduced
from a mixture of 79% of syn and 21% of anti). Rf 0.25 (DCM/
AcOEt: 80/20); 1H NMR (400 MHz, CD3OD) Syn d 8.23 (d,
J¼8.5 Hz, 2H), 7.71 (d, J¼8.5 Hz, 2H), 5.26 (d, J¼2.6 Hz,
1H), 4.30 (d, J¼2.6 Hz, 1H), 2.34 (s, 3H); Anti d 8.22 (d,
J¼8.5 Hz, 2H), 7.66 (d, J¼8.4 Hz, 2H), 4.94 (d, J¼6.0 Hz,
1H), 4.23 (d, J¼6.0 Hz, 1H), 2.18 (s, 3H); 13C NMR
(100 MHz, CD3OD) Syn d 211.9, 151.4, 149.0, 129.1, 124.5,
82.4, 74.9, 27.4; Anti d 212.1, 150.8, 149.2, 129.7, 124.4, 82.3,
75.8, 28.1; IR (neat) nmax 3422, 1715, 1606, 1518, 1348, 1108,
1064; HRMS of the mixture (CIþ): MþNHþ4 243.0981
(expected), 243.0985 (observed).
1
Et2O: 50/50); mp 55 8C; H NMR (400 MHz, CDCl3) syn
d8.26 (d, J¼8.8 Hz, 2H), 7.61 (d, J¼8.8 Hz, 2H), 5.31 (ddd,
J¼24.8, 6.2, 2.2 Hz, 1H), 4.86 (dd, J¼48, 2.2 Hz, 1H), 2.84
(d, J¼6.2 Hz, 1H), 2.34 (d, J¼4.9 Hz, 3H); anti d 8.24 (d,
J¼8.8 Hz, 2H), 7.57 (d, J¼8.8 Hz, 2H), 5.18 (ddd, J¼12.7,
5.4, 2.4 Hz, 1H), 4.82 (dd, J¼48.9, 5.4 Hz, 1H), 3.30 (d,
J¼2.4 Hz, 1H), 2.23 (d, J¼6.2 Hz, 3H); 13C NMR
(100 MHz, CDCl3) syn d 207.4, 148.0, 144.7, 127.3,
123.8, 96.3 (d, J¼195.0 Hz), 72.9 (d, J¼18.0 Hz), 27.6;
anti d 207.1, 147.9, 145.8, 127.9, 123.6, 95.6 (d,
J¼192.0 Hz), 72.6 (d, J¼21.3 Hz), 27.4; 19F NMR
(235 MHz, CDCl3) syn d 2195.3 (ddq, J¼48, 24.8,
4.9 Hz); anti d 2204.7 (ddq, J¼48.9, 12.7, 6.2 Hz); IR

630
V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
(neat) nmax 3438, 1720, 1607, 1520, 1348; HRMS of the
mixture (CIþ): MþNHþ4 245.0938 (expected), 245.0932
(observed).
CH3CN (3 mL) was added at RT Selectfluor (397 mg,
1.1 mmol) in dry acetonitrile (15 mL). The reaction mixture
was stirred at RT for 1 h. The mixture was then diluted with
EtOAc, washed with NaHCO3 and extracted with EtOAc. The
combined organic layers were washed with brine and dried
over Na2SO4. Removal of the solvents in vacuo and
purification by column chromatography over silica
(DCM/hexane: 60/40, followed by hexane/Et2O: 80/20)
afforded a white solid (35% overall yield). Rf 0.15 (hexane/
Et2O: 80/20); mp 64 8C; 1H NMR (400 MHz, CDCl3) d 8.22
(d, J¼8.6 Hz, 2H), 7.54 (d, J¼8.6 Hz, 2H), 5.34 (dd, J¼8.3,
4.1 Hz, 1H),4.86(dd,J¼47.2, 16.0 Hz, 1H), 4.81(dd,J¼47.2,
16.0 Hz, 1H), 3.02 (ddd, J¼15.8, 4.1, 2.5 Hz, 1H), 2.65 (ddd,
J¼15.8, 8.3, 2.4 Hz, 1H), 0.86 (s, 9H), 0.05 (s, 3H), 20.14
(s, 3H); 13C NMR (100 MHz, CDCl3) d 203.7 (d, J¼19 Hz),
151.3, 147.4, 126.6, 123.8, 85.4 (d, J¼185 Hz), 70.2, 48.6,
25.6, 18.0, 24.8, 25.3; 19F NMR (235 MHz, CDCl3) d
2227.0 (tt, J¼47.2, 2.4 Hz); IR (neat) nmax 2956, 1734, 1642,
1517, 1344, 1258; HRMS (CIþ): MþNHþ4 359.1824
(expected), 359.1811 (observed).
4.4.5. 6-(40-Acetamidophenyl)-3-ethyl-4-hydroxy-2-hex-
anone 2 (syn and anti). A DCM solution (2 mL) of a
mixture of trimethyl-(1-propyl-vinyloxy)-silane and tri-
methyl-(1-methyl-but-1-enyloxy)-silane (75 mg, 0.47 mmol)
was added dropwise into a mixture of aldehyde (100 mg,
0.52 mmol) and TiCl4 (57 mL, 0.52 mmol) in 4 mL of DCM
at 278 8C. After stirring the mixture for 1.5 h at 278 8C,
sat. NH4Cl solution was added. The aqueous phase was
extracted three times with AcOEt, the combined organic
layers were washed with brine and dried over Na2SO4.
Purification by column chromatography (Et2O/AcOEt: 80/
20), afforded 2 as a D1/D2: 50/50 mixture. Analytical data
of the mixture. Rf 0.20 (Et2O/AcOEt: 80/20); 1H NMR
(400 MHz, CDCl3) d 7.40 (d, J¼8.2 Hz, 2H), 7.32 (bs, 1H),
7.13 (d, J¼8.2 Hz, 2H), 3.80 (dt, J¼9.2, 4 Hz, 0.5H, D1),
3.73 (ddd, J¼8.8, 5.2, 3.6 Hz, 0.5H, D2), 2.81 (m, 1H), 2.63
(m, 1H), 2.52 (m, 1H), 2.19 (s, 1.5H, D2), 2.18 (s, 1.5H,
D1), 2.16 (s, 3H,), 1.70 (m, 4H,), 0.92 (t, J¼7.2 Hz, 1.5H,
D1), 0.90 (t, J¼7.2 Hz, 1.5H, D2); 13C NMR (100 MHz,
CDCl3) d 214.7 (D2), 213.7 (D1), 168.3, 137.8 (D1), 137.7
(D2), 135.7, 128.9, 120.0, 71.2 (D2), 70.6 (D1), 59.0 (D1),
58.8 (D2), 37.8 (D2), 36.2 (D2), 31.7 (D2), 31.6 (D1), 31.5
(D1), 31.4 (D2), 24.5, 22.2 (D2), 19.8 (D1), 12.4 (D1), 11.7
(D2); IR (neat) nmax 3308, 2953, 2939, 1708, 1665, 1601,
1543, 1522; HRMS of the mixture (FI): Mz 277.1678
(expected), 277.1679 (observed).
4.4.8. 1-Fluoro-4-hydroxy-4-(4-nitrophenyl)-butan-2-
one 13. To a solution of 1-fluoro-4-(tert-butyl-dimethyl-
silanyloxy)-4-(40-nitrophenyl)-2-butanone (69 mg, 0.2 mmol)
in dry DCM (2 mL) was added at 278 8C BF3.Et2O (56 mL,
0.44 mmol). The mixture was stirred at 278 8C for 1.5 h
and at 0 8C for 5 h. The reaction was quenched by
addition of water and extracted with DCM. The combined
organic layers were dried over Na2SO4, filtered and the
solvent was removed in vacuo. Purification by column
chromatography over silica (hexane/EtOAc: 65:35)
afforded 13 as a white solid (66%). Rf0.16 (hexane/
4.4.6. 4-(tert-Butyl-dimethyl-silanyloxy)-4-(40-nitro-
phenyl)-2-butanone. Imidazole (2.9 g, 43.4 mmol) and
TBSCl (3.2 g, 21.2 mmol) was added to a solution of
4-(40-nitrophenyl)-4-hydroxy-2-butanone (1.8 g, 8.8 mmol)
in 40 mL of DMF at room temperature. The reaction was
stirred overnight, quenched with water, extracted with DCM
and the combined organic layers were washed with brine
and dried over MgSO4. Removal of solvents and purifi-
cation by chromatography over silica (hexane/DCM: 50/50)
afforded a white solid (92%). Rf 0.26 (hexane/DCM: 50/50);
mp 36 8C; 1H NMR (500 MHz, CDCl3) d 8.19 (d, J¼9.0 Hz,
2H), 7.52 (d, J¼9.0 Hz, 2H), 5.28 (dd, J¼8.3, 4.3 Hz, 1H),
2.94 (dd, J¼15.8, 8.3 Hz, 1H), 2.58 (dd, J¼15.8, 4.3 Hz,
1H), 2.16 (s, 3H), 0.86 (s, 9H), 0.04 (s, 3H), 20.14 (s, 3H);
13C NMR (100 MHz, CDCl3) d 206.0, 151.9, 147.2, 126.6,
123.7, 70.7, 53.8, 31.7, 25.6, 18.0, 24.8, 25.2; IR (neat)
1
EtOAc: 65/35); mp 76 8C; H NMR (400 MHz, CDCl3) d
8.24 (d, J¼8.4 Hz, 2H), 7.57 (d, J¼8.4 Hz, 2H), 5.37 (dd,
J¼8.5, 3.8 Hz, 1H), 4.86 (d, J¼47.1 Hz, 2H), 3.22 (bs,
1H), 3.02 (ddd, J¼18.0, 8.5, 2.4 Hz, 1H), 2.98 (ddd,
J¼18.0, 3.8, 2.4 Hz, 1H); 13C NMR (100 MHz, CDCl3) d
206.4 (d, J¼20 Hz), 149.6, 147.5, 126.4, 124.0, 85.0 (d,
J¼184 Hz), 68.4, 47.0; 19F NMR (235 MHz, CDCl3) d
2228.4 (tt, J¼47.1 Hz, 2.4); IR (neat) nmax 3423, 2921,
1733, 1642, 1520, 1343; HRMS (CIþ): MþNH4þ
245.0938 (expected), 245.0933 (observed).
4.4.9. 1-Chloro-4-(tert-butyl-dimethyl-silanyloxy)-4-(40-
nitrophenyl)-butan-2-one.
Trimethylsilyl
triflate
(336 mL, 1.9 mmol) was added0 to a solution of 4-(tert-
butyl-dimethyl-silanyloxy)-4-(4 -nitrophenyl)-2-butanone
(300 mg, 0.95 mmol) and lutidine (432 mL, 3.8 mmol) in
DCM (3 mL) at 278 8C. The reaction mixture was stirred
2 h at 278 8C, 30 min at 0 8C and then diluted with cold
DCM and washed with cold sat. NaHCO3. The organic layer
was separated and the aqueous layer was extracted three
times with cold DCM. The combined organic layer was
washed with cold sat. NaCl, dried over Na2SO4 and
concentrated to give the corresponding silyl enol ether
which was used in the next step without purification. To a
solution of the residue in CH3CN (3 mL) was successively
added at 0 8C, N-chlorosuccinimide (120 mg, 0.9 mmol
160 mg) and water (0.5 mL). The mixture was warmed to
RT and stirred for 3 h. Then water was added and the
solution was extracted with AcOEt, washed with brine and
dried over Na2SO4. Removal of solvents and purification by
chromatography over silica (hexane/Et2O: 80/20) afforded a
n
324.1631 (expected), 324.1643 (observed).
max 2930, 1720, 1523, 1347, 1090; HRMS (CIþ): MþHþ
4.4.7. 1-Fluoro-4-(tert-butyl-dimethyl-silanyloxy)-4-(40-
nitrophenyl)-2-butanone. Trimethylsilyl triflate (436 mL,
1.9 mmol) was added to a solution of 4-(tert-butyl-dimethyl-
silanyloxy)-4-(40-nitrophenyl)-2-butanone (300 mg, 0.95
mmol) and lutidine (432 mL, 3.7 mmol) in DCM (3 mL) at
278 8C. The reaction mixture was stirred for 2 h at 278 8C,
30 min at 0 8C and then diluted with cold DCM and washed
with cold sat. NaHCO3. The organic layer was separated and
the aqueous layer was extracted three times with cold DCM.
The combined organic layers were washed with cold sat.
NaCl, dried over Na2SO4 and concentrated to give the
corresponding silyl enol ether which was used in the next
step without purification. To a solution of the residue in

V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
631
white solid (150 mg, 35% for both steps). Rf 0.13
(hexane/Et2O: 80/20); mp 54 8C; 1H NMR (400 MHz,
CDCl3) d8.22 (d, J¼8.6 Hz, 2H), 7.54 (d, J¼8.6 Hz, 2H)
5.30 (dd, J¼8.5, 4.1 Hz, 1H), 4.15 (d, J¼15.8 Hz, 1H), 4.11
(d, J¼15.8 Hz, 1H), 3.06 (dd, J¼15.6, 8.5 Hz, 1H), 2.70 (dd,
J¼15.6, 4.1 Hz, 1H), 0.86 (s, 9H), 0.04 (s, 3H), 20.15 (s,
3H); 13C NMR (125 MHz, CDCl3) d 199.5, 151.0, 147.4,
126.5, 123.8, 70.8, 50.0, 49.4, 25.6, 17.9, 24.8, 25.4; IR
(neat) nmax 2990, 1738, 1523, 1348, 1254, 1093; HRMS
(CIþ): MþHþ 358.1241 (expected), 358.1235 (observed).
Reactions were initiated by addition of 100 mL of ketone
(65 mM) in PBS (10 mM phosphate, 150 mM NaCl, pH 7.4)
containing 10% of acetonitrile. The reactions were mon-
itored by RP-HPLC and, after a minimum of 8% conversion
was reached, the unreacted aldol was isolated by semi-
preparative reversed-phase HPLC, Hypersil ODS column,
5 mm particle size, 7£250 mm, flow rate 2.0 mL/min. The
fractions were freeze-dried, the residue was redissolved in
500 mL of dichloromethane/hexane (50/50) and the ee was
determined by normal-phase HPLC using a Chiralcel OD or
OJ Daicel column.
4.4.10. 1-Chloro-4-hydroxy-4-(4-nitrophenyl)-butan-2-
one 11. To a solution of 1-chloro-4-(tert-butyl-dimethyl-
silanyloxy)-4-(40-nitrophenyl)-butan-2-one (79 mg, 0.22
mmol) in dry DCM (3 mL) was added at 278 8C
BF3.Et2O (61 mL, 0.48 mmol). The mixture was stirred at
278 8C for 1.5 h, at 0 8C for 1.5 h and at RT for 4 h. The
reaction was quenched by addition of water and extracted
with DCM. The combined organic layers were dried over
Na2SO4, filtered and the solvent was removed in vacuo.
Purification by column chromatography over silica (hexane/
EtOAc: 65/35) afforded 11 as a white solid (67%). Rf 0.21
(hexane/EtOAc: 65/35); mp 94 8C; 1H NMR (400 MHz,
CDCl3) d 8.24 (d, J¼8.4 Hz, 2H), 7.57 (d, J¼8.4 Hz, 2H),
5.34 (m, 1H), 4.14 (s, 2H), 3.15 (d, J¼3.6 Hz, 1H), 3.03 (m,
2H); 13C NMR (100 MHz, CDCl3): d 202.1, 149.4, 147.5,
126.4, 123.9, 68.9, 48.4, 48.2; IR (neat) nmax 3390, 2912,
1726, 1601, 1509, 1344; HRMS (CIþ): MþHþ 261.0642
(expected), 261.0645 (observed).
4.8. Determination of enantiomeric excesses: retro-aldol
reaction
10 mL of a racemic stock solution of aldol (80 mM in
acetonitrile) was added to 1 mL of antibody (32 mM in PBS,
pH 7.4). The reactions were monitored by RP-HPLC. After
reaching 50% conversion, the unreacted aldol was isolated
by semi-preparative reverse-phase HPLC, Hypersil ODS
column, 5 mm particle size, 7£250 mm, flow rate
2.0 mL/min. The fractions were freeze-dried, the residue
was redissolved in 200 mL of dichloromethane/hexane
(50/50) and the ee was determined by normal-phase
HPLC using a Chiralcel OD or OJ Daicel column.
Acknowledgements
We are most grateful to Professor Richard Lerner for his
generous support and for providing all the antibodies for this
study. This work was supported by the University of Oxford
(grant B(RE)9862 to V. G. and grant to V. M.), the Royal
Society of Chemistry (RSRG 20750 grant to V. G.) and the
EPSRC (grant to M.R.). We thank the National Institute of
General Medical Sciences, National Institute of Health (GM
36700) for financial support of this research, and the
National Computational Science alliance (MCA93S015N),
the National Science Foundation (CHE-9610080), and
UCLA Academic Technology Services for computer
resources at UCLA.
4.5. Antibody assays
All antibody-catalyzed reactions were performed in
phosphate buffered saline (10 mM phosphate, 150 mM
NaCl, pH 7.4). All antibody-catalyzed reactions and
background reactions were monitored by high-pressure
liquid chromatography (HPLC; Waters HPLC system
(626 Pump, 600S Controller, 996 Photodiode Array
Detector, Millenium32 Software) using a Nova-pak
˚
Waters column (C-18, 60 A pore size, 4 mm particle
size, 3.9£150 mm) and acetonitrile/water or methanol/-
water mixtures (containing 0.1% trifluoroacetic acid) as
eluents at a flow rate of 1.0 mL/min. For antibody assays
run for significant periods of time, a control assay was
performed to rule out the possibility of catalysis due to
the presence of microflora.
References and notes
1. For general reviews on the catalytic enantioselective aldol
reaction: (a) Carreira, E. M. Comprehensive Asymmetric
Catalysis; Jacobsen, E. N., Pfaltz, A., Yamamoto, H., Eds.;
Springer: Heidelberg, 1999; Vol. 3. Chapter 29–1. (b) Nelson,
4.6. Michaelis–Menten kinetics
Product formation or percent conversion of antibody-
catalyzed reaction mixtures was monitored by HPLC. The
points were determined experimentally and the best fit value
of Vmax and Km were obtained by fitting the vi versus [S]0
data to hyperbolic saturation curves by weighted non-linear
regression. All data are reported per antibody active site. An
IgG antibody possesses 2 active sites per MW of
,150,000 g/mol.
¨
S. G. Tetrahedron: Asymmetry 1998, 9, 357–389. (c) Groger,
H.; Vogl, E. M.; Shibasaki, M. Chem. Eur. J. 1998, 4,
1137–1141. (d) Machajewski, T. D.; Wong, C.-H. Angew.
Chem., Int. Ed. Engl. 2000, 39, 1352–1375.
2. (a) Mukaiyama, T. Org. React. 1982, 28, 203–331. (b)
Heathcock, C. H. Asymmet. Synth.; Morrison, J. D., Ed.;
Academic: New York, 1984; Vol. 3. Chapter 2. (c) Evans,
D. A.; Nelson, J. V.; Taber, T. R. Topics Stereochem. 1982, 13,
1–115. (d) Masamune, S.; Choy, W.; Petersen, J. S.; Sita, L. R.
Angew. Chem., Int. Engl. 1985, 97, 1–31. (e) Evans, D. A.;
Bartoli, J.; Shih, T. L. J. Am. Chem. Soc. 1981, 103,
2127–2129.
4.7. Determination of enantiomeric excesses: forward
aldol reaction
10 mL of a 66 mM solution of p-nitrobenzaldehyde in
acetonitrile was added to 2 mL of antibody (32 mM in PBS).
3. See for example: (a) Keck, G. E.; Krishnamurthy, D. J. Am.

632
V. Maggiotti et al. / Tetrahedron 60 (2004) 619–632
Chem. Soc. 1995, 117, 2363–2365. (b) Mikami, K.;
Barbas, C. F., III. Angew. Chem., Int. Ed. 1999, 38,
3738–3741.
Matsukawa, S.; Nagashima, M.; Funabashi, H.; Morishima,
H. Tetrahedron Lett. 1997, 38, 579–582. (c) Singer, R. A.;
Carreira, E. M. Tetrahedron Lett. 1997, 38, 927–930. (d)
Carreira, E. M.; Singer, R. A.; Lee, W. J. Am. Chem. Soc.
1994, 116, 8837–8838. (e) Mikami, K.; Matsukawa, S. J. Am.
Chem. Soc. 1994, 116, 4077–4078. (f) Kim, Y.; Singer, R. A.;
Carreira, E. M. Angew. Chem., Int. Ed. 1998, 37, 1261–1263.
(g) Evans, D. A.; Murry, J. A.; Kozlowski, M. C.; Burgey,
C. S.; MacMillan, D. W. C. J. Am. Chem. Soc. 1997, 119,
7893–7894. (h) Kobayashi, S.; Nagayama, S.; Busujima, T.
Chem. Lett. 1999, 71–72.
9. Maggiotti, V.; Resmini, M.; Gouverneur, V. Angew. Chem.,
Int. Ed. 2002, 41, 1012–1014.
10. (a) Hoffmann, T.; Zhong, G.; List, B.; Shabat, D.; Anderson,
J.; Gramatikova, S.; Lerner, R. A.; Barbas, C. F., III. J. Am.
Chem. Soc. 1998, 120, 2768. (b) Lerner, R. A.; Barbas, C. F.,
III. Acta Chem. Scand. 1996, 36, 672–678.
11. (a) Guth, J. J.; Gross, R. L.; Carson, F. W. J. Org. Chem. 1982,
47, 2666–2669. (b) Guthrie, J. P.; Cossar, J. Can. J. Chem.
1986, 64, 1250–1266.
12. (a) Adolph, H. G.; Kamlet, M. J. J. Am. Chem. Soc. 1966, 88,
4761–4763. (b) Hine, J.; Mahone, L. G.; Liotta, C. L. J. Am.
Chem. Soc. 1967, 89, 5911–5920. (c) Hine, J.; Hampton,
G. K.; Menon, B. C. J. Am. Chem. Soc. 1967, 89, 2664–2668.
(d) Hine, J.; Sinha, A. J. Org. Chem. 1984, 49, 2186–2190.
13. (a) List, B.; Shabat, D.; Barbas, C. F., III.; Lerner, R. A. Chem.
Eur. J. 1998, 4, 881–885. (b) Shabat, D.; List, B.; Lerner,
R. A.; Barbas, C. F., III. Tetrahedron Lett. 1999, 40,
1437–1440.
4. (a) Yamada, Y. M. A.; Yoshikawa, N.; Sasai, H.; Shibasaki,
M. Angew. Chem., Int. Ed. Engl. 1997, 36, 1871–1873. (b)
Arai, T.; Yamada, Y. M. A.; Yamamoto, H.; Sasai, H.;
Shibasaki, M. Chem. Eur. J. 1996, 2, 1368–1372. (c)
Yoshikawa, N.; Yamada, Y. M. A.; Das, J.; Sasai, H.;
Shibasaki, M. J. Am. Chem. Soc. 1999, 121, 4168–4178. (d)
Yamada, Y. M. A.; Shibasaki, M. Tetrahedron Lett. 1988, 39,
5561–5564. (e) Kumagai, N.; Matsunaga, S.; Yoshikawa, N.;
Ohshima, T.; Shibasaki, M. Org. Lett. 2001, 3, 1539–1542.
5. (a) List, B.; Lerner, R. A.; Barbas, C. F., III. J. Am. Chem. Soc.
2000, 122, 2395–2396. (b) Notz, W.; List, B. J. Am. Chem.
Soc. 2000, 122, 7386–7387. (c) List, B. Synlett 2001, 11,
1675–1686. (d) List, B.; Pojarliev, P.; Castello, C. Org. Lett.
2001, 3, 573–575. (e) List, B. Tetrahedron 2002, 58,
5573–5590. (f) Hoang, L.; Bahmanyar, S.; List, B. J. Am.
Chem. Soc. 2003, 125, 16–17. (g) Bahmanyar, S.; Houk, K. N.;
Martin, H. J.; List, B. J. Am. Chem. Soc. 2003, 125,
2475–2479.
14. Maggiotti, V.; Wong, J.-B.; Razet, R.; Cowley, A. R.;
Gouverneur, V. Tetrahedron: Asymmetry 2002, 13,
1789–1798.
15. Lin, J.-F.; Wu, C.-C.; Lien, M.-H. J. Phys. Chem. 1995, 99,
16903–16908.
16. (a) Becke, A. D. J. Chem. Phys. 1993, 98, 1372–1377, see also
5648–5652. (b) Lee, C.; Yang, W.; Parr, R. G. Phys. Rev. B
1988, 37, 785–789.
17. (a) Ditchfield, R.; Hehre, W. J.; Pople, J. A. J. Chem. Phys.
1971, 54, 724–728. (b) Hehre, W. J.; Ditchfield, R.; Pople,
J. A. J. Chem. Phys. 1972, 56, 2257–2261. (c) Hariharan, P. C.;
Pople, J. A. Theor. Chim. Acta 1973, 28, 213–222.
18. Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.;
Robb, M. A.; Cheeseman, J. R.; Zakrzewski, V. G.;
Montgomery, J. A., Jr.; Stratmann, R. E.; Burant, J. C.;
Dapprich, S.; Millam, J. M.; Daniels, A. D.; Kudin, K. N.;
Strain, M. C.; Farkas, O.; Tomasi, J.; Barone, V.; Cossi, M.;
Cammi, R.; Mennucci, B.; Pomelli, C.; Adamo, C.; Clifford,
S.; Ochterski, J.; Petersson, G. A.; Ayala, P. Y.; Cui, Q.;
Morokuma, K.; Malick, D. K.; Rabuck, A. D.; Raghavachari,
K.; Foresman, J. B.; Cioslowski, J.; Ortiz, J. V.; Stefanov,
B. B.; Liu, G.; Liashenko, A.; Piskorz, P.; Komaromi, I.R.;
Gomperts, R.; Martin, L.; Fox, D. J.; Keith, T.; Al-Laham,
M. A.;Peng, C. Y.; Nanayakkara, A.; Gonzalez, C.;
Challacombe, M. P.; Gill, M.W.; Johnson, B.; Chen, W.;
Wong, M. W.; Andres, J. L.; Gonzalez, C.;Head-Gordon, M.;
Replogle, E. S.; Pople, J. A. Gaussian 98 (Revision A.11);
Gaussian, Inc.: Pittsburgh, PA, 1998.
6. (a) Trost, B. M.; Ito, H. J. Am. Chem. Soc. 2004, 122,
12003–12004. (b) Trost, B. M.; Silcoff, E. R.; Ito, H. Org.
Lett. 2001, 3, 2497–2500. (c) Trost, B. M.; Ito, H.; Silcoff,
E. R. J. Am. Chem. Soc. 2001, 123, 3367–3368.
7. (a) Wong, C.-H.; Whitesides, G. M. Enzymes in Synthetic
Organic Chemistry. Pergamon: Oxford, 1994. (b) Drauz, K.;
Waldmann, H. Enzyme Catalysis in Organic Synthesis. VCH:
Weinheim, 1995. (c) Gijsen, H. J. M.; Qiao, L.; Fitz, W.;
Wong, C.-H. Chem. Rev. 1996, 96, 443–476.
8. (a) Wagner, J.; Lerner, R. A.; Barbas, C. F., III. Science 1995,
¨
270, 1797–1800. (b) Bjornestedt, R.; Zhong, G.; Lerner, R. A.;
Barbas, C. F., III. J. Am. Chem. Soc. 1996, 118, 11720–11724.
(c) Zhong, G.; Hoffmann, T.; Lerner, R. A.; Danishefsky, S.;
Barbas, C. F., III. J. Am. Chem. Soc. 1997, 119, 8131–8132.
(d) Hoffmann, T.; Zhong, G.; List, B.; Shabat, D.; Anderson,
J.; Gramatikova, S.; Lerner, R. A.; Barbas, C. F., III. J. Am.
Chem. Soc. 1998, 120, 2768–2779. (e) Zhong, G.; Shabat, D.;
List, B.; Anderson, J.; Sinha, S. C.; Lerner, R. A.; Barbas, C. F.,
III. Angew. Chem., Int. Ed. 1998, 37, 2481–2484. (f) List, B.;
Shabat, D.; Zhong, G.; Turner, J. M.; Li, A.; Bui, T.;
Anderson, J.; Lerner, R. A.; Barbas, C. F., III. J. Am. Chem.
Soc. 1999, 121, 7283–7291. (g) Zhong, G.; Lerner, R. A.;
19. Reported energies include zero point energy corrections and
were scaled by 0.9806 Scott, A. P.; Radom, L. J. Phys. Chem.
1996, 100, 16502–16513.