Chemoenzymatic synthesis of vitamin B5-intermediate (R)-pantolactone via combined asymmetric organo- and biocatalysis

Article abstract of DOI:10.1021/jo502667x

The combination of an asymmetric organocatalytic aldol reaction with a subsequent biotransformation toward a "one-pot-like" process for the synthesis of (R)-pantolactone, which to date is industrially produced by a resolution process, is demonstrated. This process consists of an initial aldol reaction catalyzed by readily available l-histidine followed by biotransformation of the aldol adduct by an alcohol dehydrogenase without the need for intermediate isolation. Employing the industrially attractive starting material isobutanal, a chemoenzymatic three-step process without intermediate purification is established allowing the synthesis of (R)-pantolactone in an overall yield of 55% (three steps) and high enantiomeric excess of 95%.

Full text of DOI:10.1021/jo502667x

Article
pubs.acs.org/joc
Chemoenzymatic Synthesis of Vitamin B5-Intermediate
(R)‑Pantolactone via Combined Asymmetric Organo- and Biocatalysis
Marcel Heidlindemann,† Matthias Hammel,‡ Ulf Scheffler,†,§ Rainer Mahrwald,§ Werner Hummel,∥
Albrecht Berkessel,*,‡ and Harald Groger*,†
̈
†Faculty of Chemistry, Bielefeld University, Universitatsstrasse 25, 33615 Bielefeld, Germany
̈
‡Department of Chemistry, University of Cologne, Greinstrasse 4, 50939 Cologne, Germany
§Institute of Chemistry, Humboldt University, Brook-Taylor-Straße 2, 12489 Berlin, Germany
∥Institute of Molecular Enzyme Technology, Heinrich Heine University of Dusseldorf, Research Centre Julich, Stetternicher Forst,
̈
̈
52426 Julich, Germany
̈
S
* Supporting Information
ABSTRACT: The combination of an asymmetric organocatalytic aldol
reaction with a subsequent biotransformation toward a “one-pot-like” process
for the synthesis of (R)-pantolactone, which to date is industrially produced by
a resolution process, is demonstrated. This process consists of an initial aldol
reaction catalyzed by readily available L-histidine followed by biotransformation
of the aldol adduct by an alcohol dehydrogenase without the need for
intermediate isolation. Employing the industrially attractive starting material
isobutanal, a chemoenzymatic three-step process without intermediate
purification is established allowing the synthesis of (R)-pantolactone in an
overall yield of 55% (three steps) and high enantiomeric excess of 95%.
(R)-panthenol,9 (R)-pantetheine,10 and pantoyl taurine, which
has been shown to inhibit the growth of certain strains of
diphtheria bacilli,11 among others. Furthermore, (R)-1 is a key
building block for natural products like 4′-phosphopantetheine,
the prosthetic group of fatty acid synthase,12 and pantothenic
acid (vitamin B5, Figure 1), which is needed for the
biosynthesis of coenzyme A (CoA).12,13
INTRODUCTION
■
The asymmetric synthesis of (R)-pantolactone ((R)-1, Figure
1) continues to be of interest to organic chemists due to its
wide use as a chiral auxiliary1−4 and key industrial intermediate
for the production of pantothenate.5,6 Besides the inhibitory
effects of optically pure (R)-1 toward lactic acid bacteria and
malaria,7,8 it serves as a starting material for the preparation of
For industrial production of (R)-pantothenic acid and its
commercial form calcium (R)-pantothenate, first racemic
pantolactone (rac-1) is synthesized via aldol condensation
using isobutanal and formaldehyde, followed by addition of
hydrogen cyanide and saponification under acidic condi-
tions.13,14 Resolution of rac-1 is conducted through diastereo-
meric crystallization using chiral auxiliaries such as (+)-3-
aminoethylpinane,15 cinchona alkaloids16 or quinine,17 with the
latter method being applied on an industrial scale.6,18
Enzymatic approaches for the kinetic resolution of rac-1
make use of lipase-catalyzed esterification of the unwanted S-
enantiomer19 or hydrolysis of acylated pantoyl lactone,20
whereas microbial methods focus on either S-21 or R-specific
lactonase-catalyzed hydrolysis of the racemate.22,23 Notably, the
latter biocatalytic resolution has been in commercial operation
since 1995.24,25 Apart from chiral resolution, different concepts
for the direct synthesis of (R)-1 are known, relying on either
Figure 1. Industrial production of (R)-pantolactone ((R)-1) and (R)-
pantothenic acid.
Received: November 22, 2014
© XXXX American Chemical Society
A
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
“pure” chemical procedures, such as Sharpless asymmetric
epoxidation26 and Sharpless dihydroxylation,27 or an enzymatic
approach that uses oxynitrilase-catalyzed synthesis of cyanohy-
drin, starting from pantolactone precursors.28,29
50% w/w toluene) and 10 mol % of L-histidine as an
organocatalyst in water (Scheme 2).
Scheme 2. Organocatalytic Synthesis of (R)-2 Using
To the best of our knowledge, there is to date only one
method that describes combining both chemo- and biocatalysis
using the industrially well-established starting material iso-
butanal.30 However, in this process, chemical preparation of the
precursor ketopantoyl lactone suffers from side product
formation, and it therefore has to be purified via vacuum
distillation (65% yield). The subsequent microbial reduction
effectively provides (R)-1 in high optical purity (99%
conversion, 94% enantiomeric excess (ee)) but is limited to
low substrate concentrations of <80 mM, which makes this
route less attractive industrially.
In this study, we present a chemoenzymatic “one-pot-like”
process for the two-step production of (R)-1 with high
enantioselectivity starting from readily available isobutanal
and glyoxylate (which is in an equilibrium with its hydrate as
the predominant form in aqueous medium) as starting
materials (Scheme 1; in the following schemes, only glyoxylate
Histidine
Although proline and its derivatives are widely applied
catalysts for asymmetric aldol reactions,33,34 α-branched
aldehydes only react as electrophiles in proline-catalyzed
cross-aldol additions, ruling out their use for optimizing this
synthetic route.35 Accordingly, similar to an attempt made by
Lu,36 we investigated natural amino acids other than histidine
and proline with respect to their ability to act as organocatalysts
in the aldol reaction using isobutanal as an aldol donor.
However, none of the tested amino acids gave better results
than histidine (data not shown). As histidine remained the
organocatalyst of choice, we next conducted kinetic studies to
gain a better insight into the reaction course of this aldol
reaction (Figure 2). Within 24 h, product formation already
Scheme 1. Chemoenzymatic Synthesis of (R)-Pantolactone
((R)-1)
Figure 2. Reaction course of the histidine-catalyzed aldol reaction of
isobutanal and ethyl glyoxylate.
reached 75% of its final value (60%), which did not increase
further even over a prolonged reaction time. The enantiomeric
excess of (R)-2 interestingly increases initially, reaching its
maximum after 24 h before slightly dropping afterward. For
further optimization, a reaction time of 24 h was chosen.
Encouraged by analogous results to that of Singh et al.,37 we
then tried to enhance the reaction performance by substituting
water for brine, which is known to have a positive effect on
conversion rate as well as enantiomeric excess of aldol addition
products in proline-catalyzed reactions. However, due to the
low solubility of L-histidine in water, adding brine to the
reaction mixture led to partial precipitation of the catalyst,
resulting in lower conversions compared to the benchmark
system (see also Scheme 2).
In general, the conversion data (in %) given in this
manuscript are defined as the percentage of substrate
consumption leading to the formation of the corresponding
product, and determination of the conversion was done by
measuring the absolute amount of product in the crude product
by means of a calibration line.
form 3 is shown, and the structure of the corresponding
hydrate is omitted for simplicity). This process is based on a
combination of asymmetric organo- and biocatalytic trans-
formations, employing high substrate concentrations (e.g., up
to 500 mM) for the biotransformation step. Notably, in
contrast to today’s existing industrialized syntheses, this
chemoenzymatic route does not require the use of highly
toxic cyanide.
RESULTS AND DISCUSSION
■
Organocatalytic Aldol Reaction. As a starting point for
the synthesis of pantolactone precursor (R)-2, we chose a
procedure recently published by some authors of this study,31,32
which is based on an asymmetric histidine-catalyzed cross-aldol
reaction yielding aldehyde (R)-2 in 60% with 65% ee. Our first
goal was to improve this asymmetric aldol reaction in terms of
conversion, yield, and ee of (R)-2. Analogous to the previous
procedure,31,32 we initially used equimolar amounts of
isobutanal and ethyl glyoxylate 3 (technical grade, containing
B
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
To clarify the role of water in the organocatalytic synthesis of
(R)-2, we varied the water-to-substrate molar ratio (Figure 3).
tions,32,42 the importance of the formation of an enamine
adduct (as an intermediate) for the stereoselective reaction
course is well known. In addition, reaction kinetics are
dominated by the formation and disaggregation of the enamine
intermediate adduct consisting of the organocatalyst and both
carbonyl compounds. As disaggregation is controlled by proton
transfer, it can be assumed that (i) media that support proton
transfer and (ii) the addition of Brønsted acids can boost the
reaction. Accordingly, adding acid cocatalysts to a mixture
comprised of a polar protic solvent (e.g., 2-propanol) and water
resulted in both higher conversion and enantiomeric excess of
the aldol addition product (R)-2 (Table 1).
Table 1. Acid Cocatalysts Improve Conversion and
Selectivity
Figure 3. Influence of water on the reaction rate.
Notably, the absence of water nearly completely stopped the
reaction, most likely due to insufficient solubility of the catalyst.
Whereas maximum conversion was found at a water/substrate
ratio of 1.9, the amount of water had no effect on the
enantiomeric excess of the aldol reaction product (R)-2.
We also evaluated the impact of the reaction temperature on
the reaction course. In agreement with data in the
literature,37−39 lowering the temperature resulted in lower
conversions. Interestingly, enantioselectivity was not a linear
correlation and reached a maximum at 10 °C (Figure 4).
entry
acid
TFA
pKa
conv. [%]
ee of (R)-2 [%]
1
0.26
2.75
3.77
3.87
4.17
4.28
4.44
4.76
5.02
5.78
5.82
65
75
84
86
90
85
84
83
85
82
74
65
72
67
71
75
75
75
75
75
72
74
2
salicylic
formic
L-lactic
benzoic
m-toluic
cinnamic
acetic
3
4
5
6
7
8
9
pivalic
lauric
10
11
uric
When applying equimolar amounts of organocatalyst and
acid additive, the best results in terms of accelerating the
reaction as well as improving optical purity of (R)-2 are found
with cocatalysts that have pKa-values between 4 and 5 (Table 1,
entries 5−9). These findings are in good agreement with results
published by Jia et al. regarding improved activities and
enantioselectivities of amino acid-derived primary amine
organocatalysts in asymmetric aldol reactions upon the addition
of acid additives.43
In parallel to amino acids as organocatalysts leading to up to
75% ee when using L-histidine, a second screening of various
types of other organocatalysts was carried out (e.g.,
sulfonamides, amino acids, alkaloids, and squareamides),
revealing bifunctional primary-amine thiourea organocatalysts
as further suitable catalysts for the desired transformation
(Table 2).
Figure 4. Effect of reaction temperature on conversion.
Catalysts 4, 6, and 7, which gave the best results, are based
on (R,R)-diaminocyclohexane (trans-DACH). The presence of
an aromatic substituent on the thiourea moiety proved crucial
for achieving good activity and enantioselectivity. Catalysts 4, 6,
and 7 can be prepared in one step from (R,R)-DACH-
monohydrochloride by reaction with the appropriate isocya-
nate, yielding the desired organocatalyst after basic workup.44
In general, urea-type organocatalysts gave enantioselectivities
that were higher than their thiourea analogues as exemplified
by, for example, reactions with compounds 445 and 7 (Table 2,
Varying the stoichiometry of the starting materials showed
no significant increase when switching the ratio of isobutanal/3
from 3:1 to 4:1 or to 5:1 (for details, see the Supporting
Information). With respect to atom economy, a molar ratio of
2:1 donor-to-acceptor aldehyde 3 was chosen for final
optimizations.
From mechanistic studies on proline-catalyzed asymmetric
aldol addition40,41 and histidine-catalyzed analogue reac-
C
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
conversion is observed at an alcohol/water ratio in the range of
0.7−1.3, whereas enantioselectivity was almost unaffected.
In summary, for the organocatalytic asymmetric aldol
addition step starting from isobutanal and ethyl glyoxylate 3,
optimized reaction conditions were found, which consist of the
use of 2 equiv of the donor aldehyde isobutanal, 1 equiv of 3
(technical grade, containing 50% w/w toluene), 10 mol % of L-
histidine, 1 equiv of Brønsted acid additive relative to the
amount of organocatalyst, 0.2−0.4 mL/mmol 3 mixture of a
monovalent alcohol (<C4) with water in a molar ratio of 1:1,
reacted for 24 h at 10 °C (Scheme 3). Applying such optimized
conditions enabled the synthesis of (R)-2 with up to 95%
conversion, 85% yield, and 79% ee.
Table 2. Screening of Amino-Urea and Amino-Thiourea
Organocatalysts
a
a
entry
catalyst
time [h]
conv. [%]
ee of (R)-2 [%]
b
1
(R,R)-4
(S,R)-5
(R,R)-6
(R,R)-7
(R,R)-7
(R,R)-7
1
85
74
86
94
77
95
55
75
75
79
81
81
b
2
1
3
4
3
2.5
3
c
5
Scheme 3. Optimized Conditions for the L-Histidine-
Catalyzed Asymmetric Aldol Reaction Step
d
6
4
a
Experiments were run on a 0.4 mmol scale. Yield and ee were
determined via chiral gas chromatography with dodecane as the
b
c
internal standard. Used 10 mol % catalyst. Used 25 mol % acetic
Biocatalytic Reduction. With respect to the second step of
this chemoenzymatic synthesis, we used an alcohol dehydro-
genase (ADH) for reduction of the (R)-pantolactone precursor
(R)-2 with formation of α-hydroxy ester (R)-4, which should
then undergo spontaneous cyclization to desired product (R)-1
(Scheme 4).
d
acid. Used 50 mol % acetic acid.
entries 1 and 4). Urea-type catalyst 5,46 derived from cis-(S,R)-
DACH, gave an enantioselectivity of 75% ee (entry 2), and
when using the simple phenyl-substituted (R,R)-DACH-based
urea 647 as an organocatalyst, a similar enantioselectivity and
improved yield was obtained (entry 3).
Scheme 4. Enzymatic Reduction of (R)-2 to (R)-
Pantolactone ((R)-1)
Amino-urea 7, which was introduced by Jacobsen in 2007,44
showed the best result with an enantioselectivity of 79% ee. By
increasing the amount of cocatalyst acid to 50 mol %, a product
yield of 95% and an enantiomeric excess of 81% were finally
achieved (Table 2, entries 4−6). It is noteworthy that these
results were achieved at catalyst loading of only 5 mol % and
within a 4 h reaction time.
Although preparation of organocatalyst 7 is just one step
from commercially available starting materials, after detailed
evaluation of the potential of all the applied organocatalysts, we
decided to use the natural amino acid L-histidine as the
organocatalyst of choice for further process optimization due to
its direct commercial availability, attractive price, and similar
enantioselectivities (75% ee, see Table 1, entries 5−9), which
were obtained in the asymmetric aldol reaction for synthesis of
(R)-2 when using this natural L-amino acid.
Next, with L-histidine as the prioritized organocatalyst
component, we investigated the influence of solvent properties
on the aldol reaction in a solvent engineering study. Although
polar solvents like DMF or DMSO gave better results than
unpolar cosolvents (data not shown), the best results were
gained when applying polar protic solvents, such as alcohols, in
the aldol reaction (for details, see the Supporting Information).
Low molecular-weight alcohols, such as methanol, ethanol, and
2-propanol, were favorable because of their positive effects on
catalyst activity and selectivity and for their attractiveness from
economic and sustainability perspectives.48 Furthermore, when
studying the impact of the molar ratio of water and a selected
alcohol (e.g., 2-propanol; for details, see Supporting Informa-
tion) on the L-histidine-catalyzed aldol addition, maximum
It has to be emphasized that the natural cofactor
nicotinamide adenine dinucleotide (NADH in its reduced
form) does not have to be added in stoichiometric amounts
when following this strategy. Instead, the reducing agent
NADH can be applied in catalytic amounts and is regenerated
via in situ oxidation of the cosubstrate 2-propanol to acetone
(as a process catalyzed by the same ADH), which makes cheap
2-propanol the reducing equivalent used in a stoichiometric
amount (Scheme 4). Additionally, stereorecognition of the
precursor aldehyde (R)-2 by the ADH would lead to
enantiomeric enrichment of (R)-1. Accordingly, the first
challenge was to find an appropriate ADH that would (i)
have a high activity for the reduction of (R)-2, thus efficiently
convert this precursor to pantolactone, (ii) possess a stereo-
preference for (R)-2 over (S)-2, and (iii) ensure effective
substrate-coupled cofactor regeneration.
D
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
When we screened a limited set of commercially available,
robust recombinant alcohol dehydrogenases, such an enzyme
was found in ADH-20049 from evocatal (evo-1.1.200), which in
an initial preliminary experiment converted 51% of aldehyde
rac-2 and gave (R)-1 in 40% ee (Scheme 5). In contrast, under
buffer to 2-propanol can be applied without causing significant
stability or activity problems, even up to a substrate
concentration of 800 mM (R)-2.
Chemoenzymatic One-Pot Synthesis of (R)-1. With
these optimized reactions in hand, we next evaluated a possible
“one-pot-like” reaction concept, which is based on direct use of
the reaction mixture resulting from the organocatalytic step for
the subsequent enzymatic reduction, thus avoiding the need to
isolate intermediate (R)-2. Compared to the benchmark
enzymatic reduction system using purified aldol product (R)-
2 (Table 4, entry 1), this process with the two coupled
Scheme 5. Enzymatic Reduction of rac-2 to (R)-
Pantolactone ((R)-1)
Table 4. Preliminary Tests for “One-Pot-Like” Synthesis
the same reaction conditions, the ADH from Rhodococcus sp.49
gave somewhat lower conversion (36%) and 36% ee, whereas
the ADH from L. kefir,50 as a widely applied ADH, led to <5%
conversion (reactions with these two enzymes not shown in
detail).
conv.
[%]
ee of (R)-2
a
entry conditions
additive
[%]
The following pH-optimization study is shown in Table 3.
Utilizing the enantiomerically enriched precursor aldehyde (R)-
1
2
3
4
5
6
7
8
A
B
C
A
A
A
A
A
69
33
64
63
67
71
42
48
95
94
96
95
95
94
96
97
Table 3. pH-Optimization Tests
b
b
b
toluene
acetic acid
L-His
isobutyraldehyde (1 equiv)
acetone (1 equiv)
a
Reaction conditions: A, purified aldehyde (R)-2; B, reaction mixture
derived according to Scheme 3 using 2-propanol as cosolvent and
acetic acid as additive; C, similar to B but with all volatile components
removed in vacuo prior to introducing the residual mixture (crude
entry
pH
conv. [%]
ee of (R)-2 [%]
b
product) to the enzymatic reduction. Additive amounts as described
1
2
3
4
5
6
7
8
9
3.8
4.6
6.3
7.0
7.6
8.0
8.5
9.0
9.6
11
30
54
49
65
67
67
63
58
n.d.
96
93
93
93
95
95
96
94
in Scheme 3.
reactions unfortunately drops the conversion from 69 to 33%,
while enantioselectivity remains unchanged. Obviously, com-
ponents remaining from the initial organocatalytic step
suppress efficient enzymatic reduction of (R)-2, thus making
a “standard” tandem-type one-pot synthesis a less-favorable
option in further process development.
However, we were pleased to find that when removing all
volatile materials from the reaction mixture of the aldol reaction
prior to its use for biotransformation, conversion is in the same
range as that of the benchmark reaction (entry 3). This
illustrates that a workup and purification of intermediate (R)-2
is not required and that simple evaporation of the volatile
materials after conducting the organocatalytic transformation is
sufficient for a subsequent efficient biocatalytic reduction
process.
Next, we evaluated which component was responsible for the
negative impact on the biotransformation with the observed
strong drop of conversion. Besides residual acid additive, L-
histidine, toluene, and the aldol product, at least one equiv of
isobutanal remains in the reaction mixture, even at a maximum
conversion of 95% to (R)-2 when using benzoic acid as the
cocatalyst. Interestingly, when adding toluene, acid additive, or
histidine to purified (R)-2, no effect on biotransformation was
detected (entries 4−6). However, when isobutanal (entry 7) or
acetone (entry 8) is present in the mixture, a significant drop of
conversion after 24 h to <50% occurs, possibly due to the
2 (78% ee) then led to desired pantolactone (R)-1 in high
enantiomeric excess when slighty basic buffer solutions of pH
(8.0−8.5) were used (entries 6 and 7).
Having achieved high enantiomeric excess of (R)-1 via
enzymatic reduction, we next focused on overcoming the
thermodynamic limitations of the system due to the reverse
reaction, in which acetone is reduced by oxidizing the product,
to improve the overall conversion and thus the process
economy. First, we changed the reaction temperature or used 2-
propanol in high excess to shift the equilibrium toward the
direction of the product. Raising the temperature led to higher
conversion (e.g., 77% at 45 °C vs 67% at room temperature
(rt); see Table 3, entry 6) but at the expense of selectivity (89%
ee at 45 °C vs 95% ee at rt). Applying a higher amount of 2-
propanol can shift the equilibrium toward the formation of
alcohol 4, however, only to a certain extent because high
concentrations of organic solvents tend to have a negative
impact on enzyme stability. A volumetric ratio of 3:1 (v/v)
E
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
ability of these additives to act as competitive substrates for
aldehyde (R)-2.
Table 5. Biotransformation Using ISPR at Low Catalyst
Loadings
a
Although isobutanal (as well as acetone from the aldol
reaction) can easily be removed in vacuo before starting the
enzymatic reduction, acetone is also formed in situ as a
stoichiometric side-product during cofactor regeneration with
isopropanol as the reducing agent (Scheme 3). On the basis of
the low boiling point of acetone, we thus became interested in
applying an “in situ product removal” (ISPR) of acetone to
effectively withdraw this ketone byproduct during the reaction
and thus shift the equilibrium in the direction of the desired
product.51,52
In the chosen setting, air saturated with water/2-propanol (5
vol %) was continuously passed through the biotransformation
reaction mixture during the reaction, thus effectively removing
the acetone formed during the course of the reaction. As the 2-
propanol concentration is kept stable, applying 1.1 equiv of the
alcohol is sufficient for effective substrate ((R)-2)-coupled in
situ cofactor regeneration. We were pleased to find that by
means of this methodology, the constant removal of acetone
resulted in the reaction equilibrium shifting significantly to the
product side. Notably, this enabled us to further decrease
catalyst loading to an excellent value of only 10 U of alcohol
dehydrogenase per mmol of substrate (R)-2, while conversion
was further improved to 85% (Figure 5).
entry
time [h]
conv. [%]
ee of (R)-2 [%]
1
2
3
4
5
6
0.5
1.0
58
75
81
86
87
85
98
98
96
95
88
80
1.5
2.0
3.0
24.0
a
Graph shows analytical total ion current (TIC) gas chromatograms
for ee control at different reaction times normalized to the (R)-1 peak
(for details, see “General Information” in the Experimental Section).
2), thus suggesting a typical kinetically controlled reaction
course of biotransformation.
Finally, with respect to the overall process economy, we also
addressed the issue of enzyme recycling. Notably, when
applying a molecular weight cutoff (MWCO) membrane of
10 kDa in our ISPR-type enzymatic reduction setup (Table 5)
at 2 h of reaction time, no biocatalyst leaching, deactivation, or
destabilization was observed (Figure 6). Furthermore, con-
version (77
2%) to and optical purity (95% ee) of (R)-1
remained unchanged over 5 cycles, thus further decreasing the
required overall catalyst loading to a very low value of only 2
U/mmol of substrate (R)-2. As no negative effects on
conversion or selectivity were observed during 5 cycles of the
enzymatic reduction of (R)-2, it can be assumed that catalyst
loading can be further decreased by extension of the recycling
cycles.
Figure 5. ISPR-settings of catalyst loading (24 h reaction time) and in
situ removal of acetone.
Combining both reaction steps led to a chemoenzymatic
“one-pot-like” process (Scheme 6). As it concerns the
organocatalytic aldol reaction, we chose a setup with acetic
acid as cocatalyst because it can be removed in vacuo together
with all of the other volatile compounds that are present in the
crude product. Conducting the organocatalytic aldol reaction
under these conditions, followed by subsequent biotransforma-
tion under ISPR conditions and spontaneous ring-cyclization
then afforded (R)-1 with 79% conversion, 70% yield, and high
enantiomeric excess of 95%. The overall yield of this
chemoenzymatic synthesis of (R)-pantolactone, (R)-1, based
on the economically attractive starting material ethyl glyoxylate
3 was 55% over three steps (organocatalytic aldol reaction,
enzymatic reduction, and spontaneous cyclization; Scheme 6).
Although to date the chemoenzymatic one-pot synthesis
It has to be mentioned that the increase in conversion,
however, was initially at the expense of enantiomeric excess of
reduction product (R)-1, which dropped from 95% (Table 4,
entry 1; reaction without ISPR of acetone) to 80% with acetone
being constantly removed. This value resembles the ee of
starting material (R)-2; thus, in this case, there is no further
enrichment of the enantiomeric excess via enantioselective
enzymatic reduction.
Interestingly, however, reducing the reaction time upon
applying 10 U/mmol ADH led to the formation of (R)-
pantolactone with high conversion and in excellent enantio-
meric excess (Table 5, entry 4). Furthermore, the highest ee
values were seen at the beginning of the process (entries 1 and
F
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
the use of highly toxic cyanide is not required in the process we
developed. With considerably high substrate concentrations
(2.5 M ethyl glyoxylate 3 for step 1, 500 mM intermediate 2 for
the biotransformation step; Scheme 5), the chemoenzymatic
“one-pot-like” process for the synthesis of (R)-pantolactone,
(R)-1, gave an overall yield of 55% of (R)-1 with a high
enantioselectivity of 95%.
EXPERIMENTAL SECTION
■
General Information. Reagents were purchased from commercial
sources and used without further purification. Isobutanal was freshly
distilled before use (75 °C at atmospheric pressure). Ethyl glyoxylate
(3) was used in technical grade containing 50% (w/w) toluene. All
buffers used were 100 mM (acetate buffer for pH 3.8 and 4.6;
phosphate buffer for pH 6.3, 7.0, 7.6, and 8.0; tris(hydroxymethyl)-
aminomethane buffer for pH 8.5 and 9.0; ethanolamine buffer for pH
1
9.6). All products were characterized by their NMR spectra. H and
13C NMR spectra were recorded in deuterochloroform (CDCl3) on a
500 MHz instrument. Chemical shifts were reported in parts per
million (ppm, δ) downfield from residual CHCl3 signal (δ(1H), 7.26
ppm; δ(13C), 77.16 ppm). Proton coupling patterns are described as
singlet (s), doublet (d), triplet (t), quartet (q), and multiplet (m).
Enzyme recycling experiments were performed in a stirred cell with 10
mL of overall volume using Ultracel PL-10 ultrafiltration discs.
Analytical HPLC was carried out with a supercritical fluid
chromatography setup consisting of a sampler/injector (AS-2059-SF
Plus), pressure regulator (BP-2080 Plus), detector (MD-2010 Plus),
cryostat (F250), pumps (PU-2080 Plus), line degasser (DG-2080-53),
net box (LC-Net II/ADC), and column oven (CO-2060 Plus). A
mixture of carbon dioxide, hexane, and 2-propanol was used as the
solvent. The enantiomeric excess (ee) of (R)-2 was determined using a
Daicel Chiralpak column OD-H with the above-mentioned HPLC
settings. Quantitative gas chromatography (GC) for conversion
determination was carried out on an Rxi-5 ms column (25 m, 0.2
mm inner diameter) at 140 °C (isotherm, 3 min) with N2 as a carrier
gas using a straight calibration line. Conversion was determined before
crude products were purified. In general, the conversion data (in %)
given in this study are defined as the percentage of substrate
consumption leading to the formation of the corresponding product,
and the conversion was determined measuring the absolute amount of
product in the crude product by means of a calibration line. The
enantiomeric excess of (R)-2 could also be determined using chiral GC
analysis (HP 6890 Series GC System; Lipodex E (25 m, 0.25 mm
inner diameter) column; N2 as carrier gas; isothermic at 85 °C). Both
yield and ee given in Table 2 were determined from crude reaction
mixtures with the former calibrated against n-dodecane as an internal
standard. Enantiomeric excess of (R)-1 was determined by chiral GC
using an Rt-βDEXm column (30 m, 0.25 mm inner diameter). The
temperature was regulated at 80 °C for 10 min, heated to 120 °C at 10
°C/min, and held at 120 °C for 10 min.
Procedure for the Analytical Method to Determine Product
Formation of (R)-1 and (R)-2 by Means of a Calibration Curve.
In 100 mL volumetric flasks, stock solutions in ethyl acetate were
prepared containing the aldol addition product (R)-2 and the
biotransformation product (R)-1, each in mass concentrations of
0.25, 0.5, 1.0, 2.0, 3.0, 4.0, and 5.0 mg/mL. Each analyte solution was
measured in a 6-fold determination via GC using an Rxi-5 ms column
(25 m, 0.2 mm inner diameter) at 140 °C (isotherm, 3 min). For each
mass concentration, a leveled value based on the area peaks (total
integration) was calculated, and a linear regression line (correlation
factor R2 > 0.999) was applied to all monitored data points. From this
straight calibration line, the unknown mass concentrations of (R)-1
and (R)-2 were identified by their area integrals. In all of the
experiments, stock solutions of crude reaction mixtures were
examined.
Figure 6. ISPR-type enzymatic reduction with recycling and reuse of
the alcohol dehydrogenase.
Scheme 6. Chemoenzymatic “One-Pot-Like” Process for the
Synthesis of (R)-Pantolactone (R)-1
(according to Scheme 6) has been performed on a smaller lab
scale (1.28 mmol), the feasibility of scaling up this process is
expected because this process does not contain unit operation
steps that are known to be critical when applied on a larger
scale.
CONCLUSION
■
In summary, we have developed an alternative route for the
synthesis of (R)-pantolactone, (R)-1, using the combination of
an organocatalytic asymmetric aldol reaction with subsequent
stereoselective biocatalytic reduction of the intermediate 2 and
spontaneous ring-cyclization to the desired (R)-pantolactone,
(R)-1. This chemoenzymatic three-step sequence, which is
based on industrially attractive isobutanal as the starting
material, can be carried out without intermediate purification,
leading to a “one-pot-like” process in which the biocatalyst
ADH can be fully reused during 5 consecutive reaction cycles
without leaching or deactivation effects. Notably, in contrast to
the existing multistep industrial process based on resolution,
General Procedure for the L-Histidine-Catalyzed Synthesis
of (R)-2 Using Equimolar Amounts of the Starting Materials
Ethyl Glyoxylate 3 and Isobutanal. To a solution of 155 mg (1
mmol) of L-histidine in 3 mL of water were added 2.05 g (10 mmol) of
G
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
ethyl glyoxylate 3 and 0.9 mL (10 mmol) of isobutanal. The reaction
mixture was stirred at room temperature (rt) for 0.5−360h (see
Scheme 2 and Figure 2), extracted with ethyl acetate, and dried over
anhydrous magnesium sulfate. From the crude product mixture, a
stock solution in ethyl acetate was prepared and measured via GC to
determine conversion (see Scheme 2 and Figure 2).
Influence of Water on the Reaction Rate of the L-Histidine-
Catalyzed Synthesis of (R)-2 Using Equimolar Amounts of the
Starting Materials Ethyl Glyoxylate 3 and Isobutanal. To 155
mg (1 mmol) of L-histidine and 0−10 mL (0−55 equiv relative to
ethyl glyoxylate 3) of water were added 2.05 g (10 mmol) of ethyl
glyoxylate 3 and 0.9 mL (10 mmol) of isobutanal. The reaction
mixture was stirred at rt for 24 h (see Figure 3), extracted with ethyl
acetate, and dried over anhydrous magnesium sulfate. From the crude
product mixture, a stock solution in ethyl acetate was prepared and
measured via GC to determine conversion (see Figure 3).
Effect of Reaction Temperature on Conversion in the L-
Histidine-Catalyzed Synthesis of (R)-2 Using Equimolar
Amounts of the Starting Materials Ethyl Glyoxylate (3) and
Isobutanal. To a solution of 155 mg (1 mmol) of L-histidine in 0.333
mL of water were added 2.05 g (10 mmol) of ethyl glyoxylate 3 and
0.9 mL (10 mmol) of isobutanal. The reaction mixture was stirred at
0−50 °C for 24 h (see Figure 4), extracted with ethyl acetate, and
dried over anhydrous magnesium sulfate. From the crude product
mixture, a stock solution in ethyl acetate was prepared and measured
via GC to determine conversion (see Figure 4).
Influence of Donor Aldehyde Equivalents on the Con-
version to (R)-2. To a solution of 155 mg (1 mmol) of L-histidine in
0.333 mL of water were added 2.05 g (10 mmol) of ethyl glyoxylate 3
and 721−3605 mg (10−50 mmol, 1−5 equiv with reference to ethyl
glyoxylate 3) of isobutanal. The reaction mixture was stirred at rt for
24 h (see Figure S1 in the Supporting Information), extracted with
ethyl acetate, and dried over anhydrous magnesium sulfate. From the
crude product mixture, a stock solution in ethyl acetate was prepared
and measured via GC to determine conversion (see Figure S1 in the
Supporting Information).
chiral GC (see Table 2) in which tr (n-dodecane) = 8.41 min, tr ((S)-
2) = 45.35 min, and tr ((R)-2) = 45.52 min.
Organocatalytic Synthesis of (R)-2: General Procedure for
Solvent Engineering Experiments. To a solution of 1.02 g (5
mmol) of ethyl glyoxylate 3 and 0.9 mL (10 mmol) of isobutanal in 20
mmol organic solvent (with reference to ethyl glyoxylate 3, see Figure
S2 in the Supporting Information) was added 0.4 mL (corresponding
to 20 mmol of water) of a 1.25 M catalyst solution containing L-
histidine and acetic acid (10 mol % each with reference to ethyl
glyoxylate 3). The reaction mixture was stirred at rt for 24 h, extracted
with ethyl acetate, and dried over anhydrous magnesium sulfate. From
the crude product mixture, a stock solution in ethyl acetate was
prepared and measured via GC to determine conversion (see Figure
S2 in the Supporting Information).
Effect of Differing Water to Alcohol Ratios on the Synthesis
of (R)-2. To 1.02 g (5 mmol) of ethyl glyoxylate 3 and 0.9 mL (10
mmol) of isobutanal was added 2.0 mL of a mixture of 2-propanol and
water in volumetric ratios of 90:10 to 70:30 (see Figure S3 in the
Supporting Information). After the addition of 77.6 mg (0.5 mmol, 10
mol % with reference to ethyl glyoxylate 3) of L-histidine and 30 mg
(0.5 mmol, 10 mol % with reference to ethyl glyoxylate 3) of acetic
acid (10 mol % each with reference to ethyl glyoxylate 3), the reaction
mixture was stirred at rt for 18 h, extracted with ethyl acetate, and
dried over anhydrous magnesium sulfate. From the crude product
mixture, a stock solution in ethyl acetate was prepared and measured
via GC to determine conversion (see Figure S3 in the Supporting
Information).
Optimized Procedure for the Organocatalytic Synthesis of
(R)-2. To a solution of 2.05 g (10 mmol) of ethyl glyoxylate 3 and 1.44
g (20 mmol) of isobutanal in 40 mmol alcohol (see Figure 4) was
added 0.8 mL of a 1.25 M catalyst solution containing L-histidine and
acid additive (see Table 1). The reaction mixture was stirred at 10 °C
for 24 h, extracted with ethyl acetate, and dried over anhydrous
magnesium sulfate. The solvent was removed, and the crude product
was purified by distillation (0.2 mbar, 73 °C). Yield: 95% (MeOH,
benzoic acid), 79% (2-propanol, acetic acid); ee: 79%.
General Procedure for the Screening of Acid Cocatalysts for
the Organocatalytic Synthesis of (R)-2. To a solution of 1.02 g (5
mmol) of ethyl glyoxylate 3 and 0.9 mL (10 mmol) of isobutanal in
1.6 mL of 2-propanol were added 0.4 mL of a 1.25 M L-histidine
solution and 10 mol % (with reference to ethyl glyoxylate 3) of an acid
additive (see Table 1). The reaction mixture was stirred at rt for 24 h,
extracted with ethyl acetate, and dried over anhydrous magnesium
sulfate. From the crude product mixture, a stock solution in ethyl
acetate was prepared and measured via GC to determine conversion
(see Table 1).
1H NMR (500 MHz, CDCl3): δ 9.57 (s, 1H), 4.33 (s, 1H), 4.31−
4.18 (m, 2H), 3.05 (s, 1H), 1.27 (t, J = 7.2 Hz, 3H), 1.14 (s, 3H), 1.06
(s, 3H). 13C NMR (126 MHz, CDCl3): δ 202.7, 172.8, 73.6, 62.2, 50.4,
18.2, 16.9, 14.1. Spectral data are in accordance with literature
values.31,32 GC (quantitative, GC 2010 Plus): tr = 2.4 min. HPLC
(Chiralpak column OD-H at 20 °C, CO2/hexane/2-propanol 90:9:1,
flow 1.0 mL/min, 212 nm): tr = 8.9 min for (S)-2, 9.7 min for (R)-2.
General Procedure for the Enzymatic Synthesis of (R)-1. To
an emulsion of 175 mg (1 mmol) of precursor (R)-2 (79% ee; used in
purified form (condition A), used as reaction mixture prepared
according to the optimized procedure for the organocatalytic synthesis
utilizing 2-propanol as alcohol and acetic acid as additive components
(condition B) or used as crude product obtained from such a reaction
mixture after subsequent removal of volatile components in vacuo
(condition C)) in 4 mL of buffer (100 mM, pH 3.8−9.6; see Table 3)
were added 1 mL of 2-propanol, 14 mg (2 mol %) of NAD+, and 100
U/mmol ADH-200. The reaction mixture was stirred for 24 h at rt
prior to the addition of 1 mL of diluted hydrochloric acid. The mixture
was extracted with ethyl acetate and separated via centrifugation
(10000 rpm, 5 min, 15 °C) three times. A stock solution in ethyl
acetate was prepared in a 100 mL volumetric flask, and conversion was
determined by quantitative GC (see Table 3). The solvent was
removed, and the crude product was purified by column
chromatography (chloroform/acetone 9:1 (v/v), 3.5 cm column
diameter, 100 mL silica gel). Determination of ee values was done by a
direct measurement of pure product-containing fractions via chiral GC
(see Table 3).
Synthesis of the Amino-Thiourea Organocatalysts 4−7.
Organocatalyst 4 was prepared according to the protocol in ref 45.
Amino-urea 5 was synthesized according to methods in ref 46, and
catalysts 6 and 7 were assembled following methods in ref 44. The
analytical data were in accordance with the literature.47 Compound 5,
which was not described in the literature, was prepared in the same
manner as its thiourea analogue.46
1
Characterization of cis-(S,R)-Amino-Urea 5. Colorless oil. H
NMR (300 MHz, MeOD-d4): δ 7.99 (s, 2H), 7.47 (s, 1H), 3.94−3.90
(m, 1H), 2.99−2.96 (m, 1H), 1.77−1.60 (m, 4H), 1.55−1.37 (m, 4H).
13C NMR (300 MHz, MeOD-d4): δ 157.1, 143.4, 133.1 (q, J = 32.9
Hz, CCF3), 124.8 (q, J = 271.9 Hz, CCF3), 118.9, 115.4, 52.0, 51.9,
31.5, 29.8, 23.4, 22.9. FT-IR (ATR) ν [cm−1]: 3312 (w), 2934 (w),
2860 (w), 1655 (w), 1560 (w), 1474 (w), 1443 (w), 1387 (m), 1275
(s), 1245 (w), 1171 (m), 1125 (s), 1043 (w), 999 (w). ESI-MS M+
[MeOH] = 370.1 g/mol (M+H+).
General Procedure for Thiourea Catalyzed Synthesis of (R)-
2. Stock solutions in dry dichloromethane were prepared from
isobutanal, ethyl glyoxylate, n-dodecane, acetic acid, and catalysts 4−7
(see Table 2). In the order given, to a GC-vial were added 0.2 mmol n-
dodecane, 0.8 mmol isobutyraldehyde, the respective amount of acetic
acid, 0.4 mmol ethyl glyoxylate, and finally the respective amount of
catalyst. The stock solutions were prepared in such a manner that a
total volume of 1 mL resulted. The GC vial was shaken at 500 rpm
under an inert atmosphere. The reaction progress was followed by
Preliminary Tests for “One-Pot-Like” Synthesis: Influence of
Additives on the Biocatalytic Reduction of (R)-2. To an emulsion
of 175 mg (1 mmol) of precursor (R)-2 (78% ee) in 1.875 mL of
buffer (100 mM, pH 8) were added 0.625 mL of 2-propanol, 14 mg (2
mol %) of NAD+, and 100 U/mmol ADH-200. To this mixture no
additive or (under condition A only) one of the following additives
was added: 10−20 vol% toluene (additive 1) or 0.1 mmol (10 mol %
H
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
with reference to (R)-2) of acetic acid (additive 2) or 15.5 mg (10 mol
% with reference to (R)-2) of L-histidine (additive 3) or 72 mg (1
mmol) of isobutanal (additive 4) or 58 mg (1 mmol) of acetone
(additive 5) (see Table 4). The reaction mixture was stirred for 24 h at
rt prior to the addition of 1 mL of diluted hydrochloric acid. The
mixture was extracted with ethyl acetate and separated via
centrifugation (10000 rpm, 5 min, 15 °C) three times. A stock
solution in ethyl acetate was prepared in a 100 mL volumetric flask,
and the conversion was determined by quantitative GC (see Table 4).
The solvent was removed, and the crude product was purified by
column chromatography (chloroform/acetone 9:1 (v/v), 3.5 cm
column diameter, 100 mL silica gel). Determination of ee values was
done by direct measurement of pure product-containing fractions via
chiral GC (see Table 4).
Evaluation of Catalyst Loading for the Enzymatic Synthesis
of (R)-1 using ISPR of Acetone. To an emulsion of 175 mg (1
mmol) of precursor (R)-2 (79% ee) in 1.875 mL of buffer (100 mM,
pH 8) were added 85 μL of 2-propanol, 14 mg (2 mol %) of NAD+,
and 10−100 U/mmol ADH-200 (see Figure 5). A water/2-propanol
(5 vol %) saturated air sparge was attached, and the reaction mixture is
stirred for 24 h at rt prior to adding 1 mL diluted hydrochloric acid.
The mixture is extracted with ethyl acetate and separated via
centrifugation (10000 rpm, 5 min, 15 °C) three times. A stock
solution in ethyl acetate is prepared in a 100 mL volumetric flask and
the conversion is determined via quantitative GC (see Figure 5). The
solvent is removed and the crude product is purified by column
chromatography (chloroform/acetone 9/1 (v/v), 3.5 cm column
diameter, 100 mL silica gel). Determination of ee-values is done by
direct measurement of pure product containing fractions via chiral GC
(see Figure 5).
General Procedure for the Enzymatic Synthesis of (R)-1
Using ISPR of Acetone. To an emulsion of 175 mg (1 mmol) of
precursor (R)-2 (79% ee) in 1.875 mL of buffer (100 mM, pH 8) were
added 85 μL of 2-propanol, 14 mg (2 mol %) of NAD+, and 10 U/
mmol ADH-200. A water/2-propanol (5 vol %) saturated air sparge
was attached, and the reaction mixture was stirred for 0.5−24 h at rt
prior to the addition of 1 mL of diluted hydrochloric acid. The mixture
was extracted with ethyl acetate and separated via centrifugation
(10000 rpm, 5 min, 15 °C) three times. A stock solution in ethyl
acetate was prepared in a 100 mL volumetric flask, and conversion was
determined by quantitative GC. The solvent was removed, and the
crude product was purified by column chromatography (chloroform/
acetone 9:1 (v/v), 3.5 cm column diameter, 100 mL silica gel) to give
(R)-1 as a colorless solid. Conversion: 58−85% (see Table 5). Yield:
76% (2 h); ee: 80−98% (see Table 5).
(R)-1 as a colorless solid. Conversion per cycle: 77 2% (see Figure
6). Yield: 70%; ee: 95%.
For the analytical data, please refer to the section “General
Procedure for the Enzymatic Synthesis of (R)-1 Using ISPR of
Acetone”.
Chemoenzymatic Synthesis of (R)-1. To a solution of 263 mg
(1.28 mmol) of ethyl glyoxylate 3 and 184 mg (2.56 mmol) of
isobutanal in 410 μL (5.1 mmol) 2-propanol were added 103 μL of
1.25 M catalyst solution containing L-histidine and acetic acid. The
reaction mixture was stirred at 10 °C for 24 h prior to removing all
volatile materials in vacuo. The highly viscous, yellow residue was
emulsified in 1.875 mL of buffer (100 mM, pH 8) and 85 μL of 2-
propanol. Then, 14 mg (2 mol %) of NAD+ and 10 U/mmol ADH-
200 were added, and a water/2-propanol (5 vol %) saturated air sparge
was attached. The reaction mixture was stirred for 2 h prior to adding
1 mL of diluted hydrochloric acid. The mixture was extracted with
ethyl acetate and separated via centrifugation (10000 rpm, 5 min, 15
°C) three times. The solvent was removed, and the crude product was
purified by column chromatography (chloroform/acetone 9:1 (v/v),
3.5 cm column diameter, 100 mL silica gel) to give (R)-1 as colorless
solid with an overall yield of 55% (95% ee) with reference to ethyl
glyoxylate 3 (see Scheme 6).
For the analytical data, please refer to the section “General
Procedure for the Enzymatic Synthesis of (R)-1 Using ISPR of
Acetone”.
ASSOCIATED CONTENT
* Supporting Information
■
S
1H and 13C spectra, gas and HPL chromatograms of
synthesized compounds, GC straight calibration lines used for
quantitative analysis, scheme for ISPR settings, and figures
illustrating the influence of several reaction parameters on
conversion and enantioselectivity of the organocatalytic aldol
reaction. This material is available free of charge via the Internet
at http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Authors
*E-mail: berkessel@uni-koeln.de.
*E-mail: harald.groeger@uni-bielefeld.de.
■
Notes
The authors declare no competing financial interest.
1H NMR (500 MHz, CDCl3): δ 4.11 (d, J = 3.4 Hz, 1H), 4.03 (d, J
= 8.9 Hz, 1H), 3.95 (d, J = 9.0 Hz, 1H), 2.50 (d, J = 3.4 Hz, 1H), 1.24
(s, 3H), 1.08 (s, 3H). 13C NMR (126 MHz, CDCl3): δ 177.8, 76.6,
75.9, 41.0, 23.0, 18.9. Spectral data are in accordance with literature
values.27,31,32 GC (quantitative, GC 2010 Plus): tr = 1.7 min. GC
(analytical, GC 2010): tr = 17.9 min for (S)-1, 18.2 min for (R)-1.
General Procedure for the Synthesis of (R)-1 using ISPR of
Acetone under Enzyme Recycling. A 10 mL stirred cell, equipped
with a 10 kDa MWCO membrane, was equilibrated by flushing with
buffer (100 mM, pH 8). The cell was loaded with 350 mg (2 mmol) of
precursor (R)-2 (79% ee), 4.7 mL of buffer (100 mM, pH 8), and 190
μL of 2-propanol. To it were added 28 mg (2 mol %) of NAD+ and 10
U/mmol ADH-200. A water/2-propanol (5 vol %) saturated air sparge
was attached, and the reaction mixture was stirred for 2 h at rt. The
solution was flushed from the cell by applying 35 psi of argon pressure
and mixed with 1 mL of diluted hydrochloric acid. The mixture was
extracted with ethyl acetate and separated via centrifugation (10000
rpm, 5 min, 15 °C) three times. The solvent was removed in vacuo.
For each following cycle, the cell was again loaded with 2 mmol
aldehyde 1, 4.7 mL of buffer, 190 μL of 2-propanol, and 28 mg of
NAD+. No additional ADH was added. The reaction times, conditions,
and workup remained unchanged as described above. The crude
products were purified by column chromatography (chloroform/
acetone 9:1 (v/v), 3.5 cm column diameter, 100 mL silica gel) to give
ACKNOWLEDGMENTS
■
We thank the Deutsche Forschungsgemeinschaft (DFG) for
generous support within the priority programme SPP1179
“Organokatalyse” (BE 998/11-1, GR 3461/2-1). Membership
in the European Cooperation in Science and Technology
(COST) Action no. CM0905 Organocatalysis (ORCA) is
gratefully acknowledged. We also thank Ms. Anna Schwartzen
and Mr. Christoph Butenuth for technical assistance.
REFERENCES
■
(1) Evans, D. A.; Wu, J.; Masse, C. E.; MacMillan, D. W. C. Org. Lett.
2002, 4, 3379−3382.
(2) O’Meara, J. A.; Gardee, N.; Jung, M.; Ben, R. N.; Durst, T. J. J.
Org. Chem. 1998, 63, 3117−3119.
(3) Davies, H. M. L.; Ahmed, G.; Calvo, R. L.; Churchill, M. R.;
Churchill, D. G. J. Org. Chem. 1998, 63, 2641−2645.
(4) Hansen, M. M.; Bertsch, C. F.; Harkness, A. R.; Huff, B. E.;
Hutchison, D. R.; Khau, V. V.; LeTourneau, M. E.; Martinelli, M. J.;
Misner, J. W.; Peterson, B. C.; Rieck, J. A.; Sullivan, K. A.; Wright, I. G.
J. Org. Chem. 1998, 63, 775−785.
(5) Shimizu, S.; Kataoka, M. Encyclopedia of Bioprocess Technology:
Fermentation, Biocatalysis, and Bioseparation; Wiley VCH, 1999, pp
1923−1934.
I
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
(6) Kaiser, K.; de Potzolli, B. Ullmann’s Encyclopedia of Industrial
Chemistry; Wiley-VCH, 1996; pp 559−572.
(7) Drell, W.; Dunn, M. S. J. Am. Chem. Soc. 1948, 70, 2057−2063.
(8) Drell, W.; Dunn, M. S. J. Am. Chem. Soc. 1954, 76, 2804−2808.
(9) Shimizu, S.; Tani, Y.; Ogata, K. Agric. Biol. Chem. 1974, 38,
1989−1996.
(10) King, T. F.; Stewart, C. J.; Cheldelin, V. H. J. Am. Chem. Soc.
1953, 75, 1290−1292.
(11) Roblin, R. O., Jr. Chem. Rev. 1946, 38, 255−367.
(12) Leonardi, R.; Zhang, Y. M.; Rock, C. O.; Jackowski, S. Prog.
Lipid Res. 2005, 44, 125−153.
(13) Bonrath, W.; Netscher, T. Appl. Catal., A 2005, 280, 55−73.
(14) Glaser, E. Monatsh. Chem. 1904, 25, 46−54.
(15) Paust, J.; Pechl, S.; Reif, W.; Schmidt, W. Liebigs Ann. Chem.
1978, 1024−1029.
(16) Major, R. T.; Finkelstein, J. J. Am. Chem. Soc. 1941, 63, 1368−
1371.
(17) Stiller, E. T.; Harris, S. A.; Finkelstein, J.; Keresztesy, J. C.;
Folkers, K. J. Am. Chem. Soc. 1940, 62, 1785−1795.
(18) Eggersdorfer, M.; Laudert, D.; Letinois, U.; McClymont, T.;
́
Medlock, J.; Netscher, T.; Bonrath, W. Angew. Chem. 2012, 124,
13134−13165; Angew. Chem., Int. Ed. 2012, 51, 12960−12990.
(19) Haughton, L.; Williams, J. M. J.; Zimmermann, J. A.
Tetrahedron: Asymmetry 2000, 11, 1697−1701.
(20) Baumann, M.; Hauer, B. H.; Bornscheuer, U. T. Tetrahedron:
Asymmetry 2000, 11, 4781−4790.
(43) Da, C.-S.; Che, L.-P.; Guo, Q.-P.; Wu, F.-C.; Ma, X.; Jia, Y.-N. J.
Org. Chem. 2009, 74, 2541−2546.
(44) Tan, K. L.; Jacobsen, E. N. Angew. Chem., Int. Ed. 2007, 46,
1315−1317.
(45) Brown, A. R.; Kuo, W.-H.; Jacobsen, E. N. J. Am. Chem. Soc.
2010, 132, 9286−9288.
(46) Berkessel, A.; Ong, M.-C.; Nachi, M.; Neudorfl, J.-M.
̈
ChemCatChem. 2010, 2, 1215−1218.
(47) Bied, C.; Moreau, J. J. E.; Man, M. W. C. Tetrahedron:
Asymmetry 2001, 12, 329−336.
(48) Alfonsi, K.; Colberg, J.; Dunn, P. J.; Fevig, T.; Jennings, S.;
Johnson, T. A.; Kleine, H. P.; Knight, C.; Nagy, M. A.; Perry, D. A.;
Stefaniak, M. Green Chem. 2008, 10, 31−36.
(49) Both the (S)-alcohol dehydrogenase from Rhodococcus sp.
(developed by the group of Prof. Hummel) and (S)-alcohol
dehydrogenase ADH-200 (evo-1.1.200) enzymes are commercially
available from evocatal GmbH, Merowinger Platz 1a, 40225
Dusseldorf, Germany. http://www.evocatal.com.
̈
(50) (R)-Alcohol dehydrogenase from Lactobacillus kefir:
Weckbecker, A.; Hummel, W. Biocatal. Biotransform. 2006, 24, 380−
389.
(51) Stillger, T.; Bonitz, M.; Filho, M. V.; Liese, A. Chem. Eng.
̈
Technol. 2002, 74, 1035−1039.
(52) Goldberg, K.; Edegger, K.; Kroutil, W.; Liese, A. Biotechnol.
Bioeng. 2006, 95, 192−198.
(21) Kataoka, M.; Nomura, J.-i.; Shinohara, M.; Nose, K.; Sakamoto,
K.; Shimizu, S. Biosci., Biotechnol., Biochem. 2000, 64, 1255−1262.
(22) Shimizu, S.; Kataoka, M.; Honda, K.; Sakamoto, K. J. Biotechnol.
2001, 92, 187−194.
(23) Sakamoto, K.; Hondab, K.; Wada, K.; Kita, S.; Tsuzaki, K.;
Nosea, H.; Kataoka, M.; Shimizu, S. J. Biotechnol. 2005, 118, 99−106.
(24) Sakamoto, K.; Yamada, H.; Shimizu, S. (Fuji Yakuhin Kogyo
Kabushiki Kaisha). EP 436730B1, 1995.
(25) Sakamoto, K.; Yamada, H.; Shimizu, S. (Fuji Yakuhin Kogyo
Kabushiki Kaisha). EP 504421 B1, 1995.
(26) Rao, A. V. R.; Rao, S. M.; Sharma, G. V. M Tetrahedron Lett.
1994, 35, 5735−5738.
(27) Upadhya, T. T.; Gurunath, S.; Sudalai, A. Tetrahedron:
Asymmetry 1999, 10, 2899−2904.
(28) Effenberger, F.; Eichhorn, J.; Roos, J. (Degussa AG). DE
19506728, 1996.
(29) Effenberger, F.; Eichhorn, J.; Roos, J. Tetrahedron: Asymmetry
1995, 6, 271−282.
(30) Shimizu, S.; Yamada, H.; Hata, H.; Morishita, T.; Akutsu, S.;
Kawamura, M. Agric. Biol. Chem. 1987, 51, 289−290.
(31) Markert, M.; Scheffler, U.; Mahrwald, R. J. Am. Chem. Soc. 2009,
131, 16642−16643.
(32) Scheffler, U.; Mahrwald, R. J. Org. Chem. 2012, 77, 2310−2330.
(33) Bisai, V.; Bisai, A.; Singh, V. K. Tetrahedron 2012, 68, 4541−
4580.
(34) Trost, B. M.; Brindle, C. S. Chem. Soc. Rev. 2010, 39, 1600−
1632.
(35) Guillena, G.; Najera, C.; Ramon, D. J. Tetrahedron: Asymmetry
2007, 18, 2249−2293.
(36) Jiang, Z.; Yang, H.; Han, X.; Luo, J.; Wong, M. W.; Lu, Y. Org.
Biomol. Chem. 2010, 8, 1368−1377.
(37) Maya, V.; Raj, M.; Singh, V. K. Org. Lett. 2007, 9, 2593−2595.
(38) Zhou, Y.; Shan, Z. Tetrahedron: Asymmetry 2006, 17, 1671−
1677.
(39) Raj, M.; Maya, V.; Ginotra, S. K.; Singh, V. K. Org. Lett. 2006, 8,
4097−4099.
̈
(40) Cheong, P. H.-Y.; Legault, C. Y.; Um, J. M.; Çelebi-Olcu̧ m, N.;
̈
Houk, K. N. Chem. Rev. 2011, 111, 5042−5137.
(41) Allemann, C.; Gordillo, R.; Clemente, F.; Cheong, P.; Houk, K.
N. Acc. Chem. Res. 2004, 37, 558−569.
(42) Lam, Y-h.; Houk, K. N.; Scheffler, U.; Mahrwald, R. J. Am.
Chem. Soc. 2012, 134, 6286−6295.
J
DOI: 10.1021/jo502667x
J. Org. Chem. XXXX, XXX, XXX−XXX