Synthesis of robalzotan, ebalzotan, and rotigotine precursors via the stereoselective multienzymatic cascade reduction of α,β-unsaturated aldehydes

Article abstract of DOI:10.1021/jo4003097

A stereoselective synthesis of bicyclic primary or secondary amines, based on tetralin or chroman structural moieties, is reported. These amines are precursors of important active pharmaceutical ingredients such as rotigotine (Neupro), robalzotan, and eba

Full text of DOI:10.1021/jo4003097

Article
pubs.acs.org/joc
Synthesis of Robalzotan, Ebalzotan, and Rotigotine Precursors via
the Stereoselective Multienzymatic Cascade Reduction of α,β-
Unsaturated Aldehydes
Elisabetta Brenna,† Francesco G. Gatti,*,† Luciana Malpezzi,† Daniela Monti,‡ Fabio Parmeggiani,†
and Alessandro Sacchetti*,†
†Dipartimento di Chimica, Materiali ed Ingegneria Chimica ”G. Natta”, Politecnico di Milano, P.zza Leonardo da Vinci 32, 20133
Milano, Italy
‡Istituto di Chimica del Riconoscimento Molecolare, CNR, Via Mario Bianco 9, 20131 Milano, Italy
S
* Supporting Information
ABSTRACT: A stereoselective synthesis of bicyclic primary
or secondary amines, based on tetralin or chroman structural
moieties, is reported. These amines are precursors of
important active pharmaceutical ingredients such as rotigotine
(Neupro), robalzotan, and ebalzotan. The key step is based on
a multienzymatic reduction of an α,β-unsaturated aldehyde or
ketone to give the saturated primary or secondary alcohol, in a
high yield and with a high ee. The catalytic system consists of
the combination of an ene-reductase (ER; i.e., OYE2 or OYE3
belonging to the Old Yellow Enzyme family) with an alcohol
dehydrogenase (ADH), applying the in situ substrate feeding
product removal technology. By this system the formation of
the allylic alcohol side product and the racemization of the
chirally unstable α-substituted aldehyde intermediate are minimized. The primary alcohols were elaborated via a Curtius
rearrangement. The combination of OYE2 with a Prelog or an anti-Prelog ADH allowed the preparation of the secondary
alcohols with ee > 99% and de > 87%. The absolute configuration of the primary amines was unambiguously assigned by
comparison with authentic samples. The stereochemistry of secondary alcohols was assigned by X-ray crystal structure and NMR
analysis of Mosher esters.
well, because it is used for the production of rotigotine
(Neupro), a drug indicated for the treatment of Parkinson’s
disease.5 In addition to these relevant targets, also the 1R,3′S
diastereoisomer of amine 4 (key precursor of chromanyl
analogues of NK-1 tachykinin receptor antagonist6 recently
developed by Eli Lilly) was taken into consideration as a part of
our ongoing research program, which is devoted to the
syntheses of biologically active molecules by means of
biocatalytic methodologies.7,8
So far, amine 2 and its derivatives have been prepared by
starting from the chiral pool, using Garner’s aldehyde9 or by
means of a radical cyclization on an L-serine derivative.10
However, although they ensure a high optical purity of
products, both routes suffer from a long synthetic sequence
and a low overall yield. Recently we described the synthesis of 2
by means of an enzymatic resolution of its 3-aryl-2-nitro-
propanol precursor, but the ee remained unsatisfactory; another
biocatalytic approach based on the microbial reduction of an
α,β-unsaturated nitrostyrene was disappointing as well.11
INTRODUCTION
■
In the last few decades, the stereochemically well-defined chiral
bicyclic amines I and II, based on tetralin or chroman carbon
backbones, have become very popular in the pharmaceutical
industry. Indeed, they are precious precursors of several
biologically active molecules with a very large spectrum of
possible applications, spanning from the treatment of
depression and anxiety up to Parkinson’s disease and as
antibacterial agents (Figure 1). Despite the fact that a large
number of biological assays have proven a strong relationship
between their activity and a preferential stereochemistry, most
of the stereoselective syntheses of I and II are based on
chemical resolutions. This kind of strategy, even if it is
extremely reliable, is intrinsically inefficient because at least half
of the starting material has to be discarded,1 in contrast with the
more efficient catalytic asymmetric methodologies.
Among all the derivatives of I and II, the (R)-aminochro-
mans 1 and 2 (Figure 2) have gained a particular interest in
medicinal chemistry, since they are key intermediates of many
antidepressant drug candidates such as robalzotan2 and
ebalzotan3 and the DBH inhibitors chromanylimidazole-
thiones.4 The (S)-tetralin 3 is another important synthon as
Received: February 13, 2013
Published: April 23, 2013
© 2013 American Chemical Society
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Figure 1. Biologically active compounds based on amines I or II.
was achieved, whereas the opposite enantiomer of 1 was
prepared with an excellent ee = 96%.
For amine 3 just one preparation has been reported; it
consists of the elaboration of (1R,2R)-1-hydroxy-2-carboxye-
thyltetralin, which was obtained by microbial reduction of the
2-carboxyethyl-1-tetralone precursor with an excellent de and
optical purity.13 However, with the substantial improvement of
asymmetric catalytic methodologies, synthetic strategies based
on the removal of stereogenic centers are progressively
becoming less appealing.
To the best of our knowledge, no asymmetric synthesis of
amine 4 has ever been reported. Conversely, several tetralin
analogues have been prepared with quite good ee’s and de’s,
but exclusively by chemical resolutions of racemic amines by
means of resolving agents such as (+)-tartaric or (+)-mandelic
acids.6 The efficient enzymatic resolution of N-hydroxyl-O-
methylamines was explored as well, but that for the tetralin
analogue of 4 proceeded with an unacceptably low con-
version.14
Therefore, current synthetic methods for the preparation of
optically pure amines I and II show significant limitations for
different reasons. In the search for alternative strategies, the
introduction of biocatalysis into asymmetric synthesis,15
especially in the pharmaceutical chemistry, would be highly
desirable and should be encouraged.16
With this background, in the following we report a new
stereoselective synthesis of amines 1−4 by means of
biocatalytic methodologies.
Figure 2. Retrosynthesis of amines 1−4.
The asymmetric hydrogenation of chroman-3-enamides in
the presence of a Ru−Synphos catalyst has been exploited for
the preparation of several chiral 3-aminochromans.12 However,
despite the fact that an excellent enantioselection was obtained
with several substrates, for the acetamide of 2 a poor ee of 34%
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quenched using DMF as electrophile to give 8 together with
its regioisomer 10 (4/6 ratio by GC-MS) in a modest yield of
32%. Unfortunately, both purification and separation of the two
regioisomers by chromatography failed. The use of form-
aldehyde as electrophile and other reaction conditions also did
not provide any significant improvement.21
RESULTS AND DISCUSSION
■
Our retrosynthetic plan relies on the enantiospecific reduction
of the CC double bond of α,β-unsaturated aldehyde/ketone
III to give primary or secondary alcohols IV (Figure 2).
Concerning targets 1−3, the amino group is meant to be
introduced by Curtius rearrangement of the carboxylic acid
precursor V via isocyanate intermediate. Instead, for the more
complex amine 4 the second stereocenter is created after an
enantioselective reduction of the carbonyl group of III to give
the corresponding secondary alcohol IV; the latter should be
easily transformed into 4 after few simple functional group
manipulations.
Accordingly, the CC double bond biocatalyzed reduction
of activated olefins seems to fit particularly well with our
retrosynthetic plan; indeed, this biotransformation has been
largely exploited for the synthesis of many valuable chiral
products,8b,c,17 quite often with a high stereoselectivity, under
mild reaction conditions and with minimal generation of waste.
However, the nature of the products is strictly related to the
type of biocatalyst applied, so that the following compounds are
obtained: (i) chiral aldehydes or ketones with overexpressed
and/or purified recombinant ene-reductases (ERs),18 in which
the CC double bond reduction is dominant over other
catalytic activities (Figure 3), and (ii) more elaborated and
Alternatively, the strategy based on the elimination of the
mesylate leaving group of the cyanohydrin derivative 12
produced the α,β-unsaturated nitrile 13 with a high
regioselectivity. Thus, sodium cyanide addition to the tetralone,
in the presence of TMSCl, gave the corresponding O-protected
cyanohydrin 11 in a quantitative yield.22 The latter was
converted into 12 after acid cleavage of the O-silyl group
followed by treatment with MsCl in pyridine in an overall yield
of 78%. The elimination reaction needed some optimization;
indeed, when 12 was treated with pyridine at 90 °C, together
with α,β-unsaturated nitrile 12, also its regioisomer (83/17
ratio by 1H NMR) and the naphthalene derivative were
produced. However, if the reaction was carried out at 60 °C,
both the regioselectivity and chemoselectivity improved
substantially, affording 13 together with a lower amount of its
regioisomer (93/7). The pure isomer 13 was obtained by
crystallization in cold n-hexane/i-Pr2O (9/1). Lastly, the cyano
group was reduced with DIBAL-H affording the desired
aldehyde 8a in a 75% yield after column chromatography
purification.
If on one hand the use of purified ERs has allowed significant
improvements, among which the complete chemoselectivity
and the high conversions are undoubtedly the most important,
on the other hand it has introduced the problem of the
relatively low chemical stability of the products. This is
particularly relevant in the case of the optically pure α-
substituted aldehydes, which can spontaneously racemize under
the typical biotransformation conditions,23 therefore, even at
neutral pH (Figure 3).
It has been demonstrated that the loss of optical purity by
racemization during the reaction can be minimized using
aqueous−organic biphasic solvent systems17b,c or applying the
in situ substrate feeding product removal (SFPR) technol-
ogy.23,24 However, when the substrate 6a was submitted to our
standard reduction protocol25b in the presence of OYE2 and
using the SFPR technology, aldehyde (S)-6b was obtained in an
excellent yield but with an unacceptably low optical purity (ee =
32%, by chiral GC). The progress curve (Figure 4) clearly
shows that the ee observed at the end of the biotransformation
is not due to poor stereoselectivity of OYE2 but instead to a
concomitant racemization process.
Very recently, we found a solution to this problem by
combining the in situ SFPR technology with a multienzymatic
catalytic system comprising an ER and an ADH.25 Indeed, by
this approach, if the unstable saturated aldehyde is immediately
reduced to the more stable alcohol, the detrimental
racemization should be suppressed. However, in this case, the
ADH should reduce exclusively the CO of saturated aldehyde;
otherwise, the undesired allylic alcohol would be produced as
well.
Figure 3. Typical chemical path of α,β-unsaturated aldehyde/ketone
reductions, with isolated enzymes or microorganisms.
complex products in all the cases where the ER activity is not
isolated but is combined with other enzymatic activities, often
in a cascade process. For instance, in the resting cells of baker’s
yeast (BY),19 ERs (specifically the old yellow enzymes OYE2
and OYE3) work in tandem with alcohol dehydrogenases
(ADHs),20 which catalyze the reduction of the carbonyl group
to give the saturated primary or secondary alcohols (Figure 3).
ADH-catalyzed competitive CO reduction, to give the side
product allylic alcohol, is more evident for enals than for
enones.
The synthetic plan adopted for the preparation of the α,β-
unsaturated substrates 5a−8a is depicted in Scheme 1. The
domino oxa-Michael aldol condensation between salicylalde-
hyde or 6-methoxysalicylaldehyde11 and acrolein or methyl
vinyl ketone followed by an in situ elimination reaction gave a
straightforward and very efficient access to the α,β-unsaturated
aldehydes 5a and 6a and to the enone 7a on a multigram scale
(Scheme 1a). In contrast, the synthesis of aldehyde 8a was
more troublesome (Scheme 1b). A first approach was based on
the Shapiro reaction: tosyl hydrazone 9, easily prepared from
the commercially available 5-methoxy-2-tetralone, was treated
with n-BuLi in the presence of TMEDA to give the
corresponding alkenyllithium intermediate that was then
In this work the concept was further investigated by testing a
set of commercially available ADHs (recombinant and/or
purified) from different sources. The reductions were
performed on an approximately equimolar mixture of the
unsaturated carbonyl compound and its corresponding
saturated compound, i.e., 5a/5b, 6a/6b, 7a/7b, and 8a/8b,
under the same reaction conditions usually adopted for OYE-
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Scheme 1. Synthesis of Substrates 5a−8a and Reference Racemic Materials 5b−8b and 5c−8c
with DRADH and 8a/8b with HLADH. Moreover, the enzyme
concentration can play a crucial role in the selectivity, as shown
for instance in the reduction of 8a/8b: with READH the
saturated aldehyde 8a was completely consumed but also a
small amount of allylic alcohol was formed (92/8), whereas
with one-fifth of the same ADH almost no allylic alcohol was
detected (99/1), but at the expense of a lower conversion of 8a
(61%).
Concerning ketones 7a/7b, in addition to the selectivity
toward the formation of the saturated alcohol, i.e. 7d, in certain
cases the CO reduction of 7b was also influenced by the
configuration of the stereocenter C(3′), giving rise to a
nonequimolar mixture of two possible diastereoisomers. For
instance, with HLADH the alcohols syn-7c and anti-7c were
produced in a ratio of 61/39. In any event, none of the
screened ADHs reached a diastereoselectivity level sufficiently
high to foresee a possible enzymatic resolution of racemic 7b.
Finally, we tested the combination of selected ADHs and
ERs on substrates 5a−8a in coupled reactions (Table 2). The
choice of the ADH was based on the best compromise between
selectivity and conversion, to achieve not only an exclusive but
also a fast reduction of the saturated aldehyde, thus preventing
undesired racemization.
Figure 4. Progress curve of the biotransformation of 6a into (S)-6b by
OYE2 (conversion ( ), ee ( )). Experimental conditions: 10 g L−1 of
substrate (loaded on 10 g L−1 resin), 150 μg mL−1 of OYE2, 4 U mL−1
of GDH, 4 equiv of glucose, 0.1 mM NADP+, 50 mM KPi buffer pH
7.0, 30 °C, 160 rpm.
●
○
catalyzed reductions. The saturated carbonyl compounds 5b−
8b and the racemic alcohols 5c−8c were prepared according to
Scheme 1c. For the regeneration of the NAD(P)H cofactors, a
glucose dehydrogenase (GDH) was employed, adding glucose
as the sacrificial substrate. In Table 1 we report the conversion
of the saturated aldehyde and the selectivity of ADHs evaluated
as ratios between the amount of saturated and allylic alcohols.
First, we observed that all the screened ADHs reduce the
unconjugated carbonyl group better than the conjugated one.
In certain cases, this preferential outcome leads to the exclusive
formation of the saturated alcohol, as for the mixtures 6a/6b
A clear example of this concept is offered by the reduction of
6a: in the presence of OYE2 and HLADH, (S)-6c was obtained
with an excellent ee of 99% and in a high yield of 88%,25b
whereas by replacing HLADH with DRADH, which, with
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a
Table 1. Selectivity of a Panel of Commercially Available ADHs
substrate 5a/5b
conversn 5b
substrate 6a/6b
substrate 8a/8b
conversn 8b
substrate 7a/7b
d
conversn 6b
conversn 7b
de
b
c
b
c
b
c
b
c
alcohol dehydrogenase
(%)
selec
(%)
selec
(%)
selec
(%)
selec
(%)
C. parapsilosis (CPADH)
R. erythropolis (READH)
horse liver (HLADH)
baker’s yeast (BYADH)
P. lavamentivorans (PLADH)
D. radiodurans (DRADH)
L. kefir (LKADH)
98
67/33
69/31
98/2
83/17
91/9
80
64
81
5
83/17
80/20
95/5
94
99
84
2
92/8
92/8
99/1
99/1
49
>99
39
94/6
24
−3
22
0
>99
>99
41
69/31
94/6
42/58
92/8
16
74/26
59/41
92/8
74
85
48
58
56
>99
47
86
65
76/24
96/4
>99
87
4
66
97/3
99/1
−9
2
58
94/6
74/26
60/40
62/38
93/7
>99
4
61/39
86/14
58/42
T. brockii (TBADH)
6
75/25
3
ketoreductase (KRED)
>99
0
a
Experimental conditions: 5 g L−1 of substrate (∼1:1 mixture, loaded on 5 g L−1 of resin), 100 μg mL−1 of ADH, 4 U mL−1 of GDH, 4 equiv of
b
c
glucose, 0.1 mM NAD(P)+, 50 mM KPi buffer pH 7.0, 30 °C, 160 rpm, reaction time 12 h. Conversion of b to c by GC. Defined as the ratio
between saturated alcohol c and allylic alcohol d, by GC. Defined as the ratio between syn-7c and anti-7c, by H NMR.
d
1
respect to the former, ensured a better selectivity on this
substrate (99/1 vs. 95/5, Table 1), the ee of the alcohol
decreased to 86%. This is likely due to the fact that DRADH
reduces 6b more slowly than HLADH (conversion 48% vs
81%; see Table 1) and thus the racemization becomes a
competitive process. This type of behavior was also observed in
the case of the reduction of 8a, in which between READH and
HLADH the latter provided the best results in terms of
conversion and selectivity (ee = 99%).
The reduction of 5a with OYE2 and HLADH gave the
alcohol (S)-5c with a good ee of 91%, and by substituting
OYE2 with OYE3 the ee was slightly worse (ee = 88%).
Assuming that the observed ee’s are the fruit of the
stereoselectivity of OYEs, these results confirm a trend already
seen for the reduction of 6a, in which OYE2 affords an ee
better than that achieved with OYE3.25b
saturated ketone (R)-7b, and the two diastereoisomers of
alcohol 7c, i.e. (1S,3R′)-7c and (1R,3R′)-7c, in a ratio of 77/23,
together with other side products, among which the allylic
alcohol was present in a negligible amount.
As shown for the reduction of aldehydes, the integration of
both biocatalytic transformations in a cascade system26 would
be particularly appealing. Thus, we tested the reduction of 7a
by simultaneous addition of OYE2 and DRADH, which
previously provided the best selectivity toward the formation
of 7c. However, in this case the alcohol 7c was obtained with a
lower diastereoselectivity (de = 76%). This might be explained
by the fact that DRADH reduces slightly more the ketone 7b
with an S absolute configuration (de = −9%, Table 1), whereas
OYE2 generates mainly (R)-7b; therefore, their combination is
stereochemically mismatched, giving rise to a lower diaster-
eoselectivity.
The ER-catalyzed reduction of α,β-unsaturated ketones such
as 7a is definitely less problematic, since it is well-known that
enantiomerically enriched ketones are less prone to racemize
than their aldehyde counterparts. This implies that the ADH
does not have to be added simultaneously with the ER, and
therefore, provided that a quantitative conversion of the α,β-
unsaturated ketone into the saturated one is ensured, the
chemoselectivity is no longer a critical parameter.
Both OYE2 and OYE3 afforded (R)-7b in almost
quantitative yields, but the best enantioselectivity was achieved
with OYE2 (98% vs 94% ee). Thus, the reduction of 7a was
carried out by a one-pot sequential addition of OYE2 and a
pro-(S) or a pro-(R) ADH under the same experimental
conditions adopted for the aldehydes. In the first case, either
CPADH or HLADH produced mainly the syn diastereoisomer,
i.e. (1S,3′R)-7c, with an excellent ee = 99% and discrete de’s of
87% and 80%, respectively, but the reduction with HLADH was
not quantitative (61% vs 97%, Table 2). Instead, KRED
furnished the anti diastereoisomer (1R,3′R)-7c with de = 89%
and once again with a high optical purity (99% ee). To stress
the efficiency of this multienzymatic catalytic system, we also
tested the reduction with baker’s yeast, which afforded a
complex mixture composed of the starting material 7a, the
Finally, the alcohols 5c−8c were produced in high yields on a
preparative scale using the best combination of ER and ADH
and applying always the SFPR technology: for the reduction of
aldehydes 5a, 6a, and 8a the cascade catalytic system was
adopted, whereas for the ketone 7a the one-pot sequential
approach was chosen.
The de of each diastereoisomer of alcohol 7c was then
improved by crystallization in cold n-hexane (de = 99% for
either (1S,3′R)-7c or (1R,3′R)-7c). It must be mentioned that
the same procedure performed on a mixture of diaster-
eoisomers of 7c obtained from BY (de = 54%) was
unsuccessful, and any attempt to enrich the de by column
chromatography failed as well.
The analysis of the X-ray structure of (1S,3′R)-7c allowed us
to assign the relative stereochemistry (see the Supporting
Information, Figure S6), whereas, in the absence of a significant
anomalous scattering, the attempt to determine the absolute
configuration by refinement of the Flack parameter27 led to an
inconclusive value. The absolute stereochemistry was initially
predicted on the basis of the following considerations: (i)
substrate 7a likely binds to the catalytically active site of OYE2
with the same orientation of the aldehydes, and consequently
the newly created C(3′) stereocenter should be R; (ii) it is
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g
Finally, the synthetic sequence for the preparation of amines
1−4, from the optically pure precursors 5c−8c, is illustrated in
Scheme 2. First, alcohol (S)-5c was oxidized30 with BAIB and
catalytic TEMPO in a biphasic solvent system (CH2Cl2/H2O,
3/1), to give the carboxylic acid (R)-5e in a yield of 94%
without any significant loss of optical purity (vide infra). Then,
by treatment with diphenylphosphoryl azide (DPPA) and Et3N
in refluxing toluene, followed by addition of benzyl alcohol,31
acid (R)-5e was converted to the N-Cbz-protected amine (R)-
5f in 84% yield after column chromatography.
Table 2. Cascade Synthesis of Alcohols 5c−8c
As an alternative to the Curtius rearrangement, we also
considered the Hoffman degradation route via the primary
amide of (R)-5e; however, using the standard conditions (Br2,
NaOH in MeOH) together with the formation of 1 a
considerable amount of bromo derivatives were produced as
well. This result was ascribed to the presence of electron-
donating groups on the phenyl ring, which favor the aromatic
electrophilic substitution reaction.
Finally, deprotection of the amino function by catalytic
hydrogenolysis of the Cbz group afforded the target compound
(R)-1 in 96% yield after a simple acid−base workup. Moreover,
since the specific optical rotatory power value of the
hydrochloride salt (R)-1·HCl matched quite well with that
reported in the literature ([α]D25 = +60.2° (c 1.6, MeOH), lit.4b
25
[α]D = +61.3° (c 0.5, MeOH, ee > 99%)), the initial
stereochemistry was completely retained during the entire
sequence.
The same route was applied to alcohols 6c and 8c, affording
amines (R)-2 ([α]D = −13.3° (c 1.5, CHCl3) lit.9 [α]D
=
25
25
−13.7° (c 1.0, CHCl3)) and (S)-3 ((S)-3·HCl [α]D25 = −60.3°
(c 1.3, MeOH) lit.9 [α]D = −61.0° (c 2.3, MeOH)) with
25
similar yields.
The amine (1R,3′S)-4 was prepared by a three-step
sequence: formation of the tosyl derivative of alcohol
(1S,3′R)-7c, which after nucleophilic substitution with NaN3
in DMF gave the azide (1R,3′S)-14 in an overall yield of 65%;
then the latter was cleanly reduced to amine (1R,3′S)-4 in a
quantitative yield by hydrogenation with a catalytic amount of
Pd/C, without any significant loss of optical purity, since the
initial de = 99% was preserved.
a
A plus sign indicates a cascade system, and a semicolon indicates a
b
one-pot sequential system. By GC on the crude reaction mixture,
c
except for BY where the products have been isolated. By chiral HPLC.
d
e
f
g
Reference 25b. 20 μg mL−1 of READH. By 1H NMR. Experimental
conditions: 5 g L−1 of substrate (loaded on 15 g L−1 of resin), 150 μg
mL−1 of OYE, 100 μg mL−1 of ADH, 4 U mL−1 of GDH, 4 equiv of
glucose, 0.1 mM NAD(P)+, 50 mM KPi buffer pH 7.0, 30 °C, 160
rpm, reaction time 12 h.
CONCLUSION
■
In summary, we have described a new synthetic route for the
preparation of amines 1−4, including a key biocatalytic step
based on selected ERs and ADHs. In the case of α-substituted
aldehydes, the addition of an ADH allows us to minimize the
detrimental racemization process, preserving the optical purity
of the chirally unstable aldehyde intermediate generated during
the highly enantioselective reductive step of the CC double
bond.
known that the stereochemistry of baker’s yeast mediated
reduction of ketones bearing a methyl group and a large
substituent, such as 7b, very often satisfies the Prelog rule,20,28
according to which the absolute configuration of the C(1)
stereocenter of the alcohol would be S. Altogether, these
configurations are in agreement with the relative stereo-
chemistry. Then, H and 13C NMR spectroscopic analysis of
1
Moreover, for the reduction of ketone 7a the selection of a
Prelog or an anti-Prelog ADH together with OYE2 gave access
to (1S,3′R)-7c and (1R,3′R)-7c, respectively, with a discrete
diastereoselectivity and with an excellent optical purity (ee =
99%). However, in this case, the one-pot sequential addition of
the enzymes was preferred to the cascade system, since the
intermediate ketones are typically less unstable. Finally, due to
the different nature of all substrates taken into consideration in
this work, we are quite confident that our multienzymatic
catalytic system together with the synthetic strategy adopted
should be broadly applicable to the enantioselective synthesis of
other important amines I and II.
the (S)-MTPA esters29 of rac-7c and of the optically pure 7c
obtained from the sequential reduction with OYE2 and
CPADH confirmed our initial assignment of the absolute
configuration: i.e., (1S,3′R)-7c. Indeed, both 1H and 13C
chemical shifts of the methyl group of (S)-MTPA-(1S,3′R)-7c
are upfield with respect to those of the Mosher derivative of its
antipode alcohol (Δδ = −0.07 and −0.28 ppm for H and 13C
1
NMR, respectively; see the Supporting Information, Figure S9),
whereas the CH2O signals resonated downfield (Δδ = +0.10
and +0.14 ppm for H and 13C NMR, respectively; see the
1
Supporting Information, Figure S10).
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Scheme 2. Synthesis of Amines 1−4 from Alcohols 5c−8c
TBADH from Thermoanaerobium brockii, and KRED (ketoreductase)
from an unspecified source were purchased from Sigma-Aldrich. Fresh
baker’s yeast from Lesaffre Italia was employed.
EXPERIMENTAL SECTION
■
General Methods. Chemicals and solvents were purchased from
suppliers and used without further purification. 1H and 13C NMR
spectra were recorded on a 250, 400, or 500 MHz spectrometer at
General Procedure for the Synthesis of 5a−7a. To a
mechanically stirred solution of salicylaldehyde or 6-methoxysalicy-
laldehyde (0.133 mol), in 1,4-dioxane (200 mL), was added
portionwise K2CO3 (22.0 g, 0.160 mol) and then acrolein (10.5 mL,
0.160 mol) or methyl vinyl ketone (13.3 mL, 0.160 mol); the reaction
mixture was refluxed for 4 h. After cooling, water was slowly added (70
mL). The solid was removed by vacuum filtration, and the solution
was concentrated under reduced pressure and then extracted with
EtOAc (3 × 200 mL). The organic phase was washed with an aqueous
solution of H3PO4 (5%, 200 mL) and then with brine (200 mL). The
organic phase was dried over Na2SO4, and the solvent was removed
under reduced pressure. Purification by column chromatography
afforded the products (EtOAc/n-hexane, 10/90).
1
room temperature, using TMS as an internal standard for H and
CDCl3 for 13C; chemical shifts δ are expressed in ppm relative to TMS.
The GC-MS analyses of the α,β-unsaturated 5a, 6a, and 8a, saturated
aldehydes 5b, 6b, and 8b, saturated alcohols 5c, 6c, and 8c, and allylic
alcohols 5d, 6d, and 8d were performed on a HP-5MS column (30 m
× 0.25 mm × 0.25 μm). Method: 60 °C (1 min)/6 °C min−1/150 °C
(1 min)/12 °C min−1/280 °C (5 min). The GC-MS analyses of
compounds 7a−d were performed on a DB-5ht column (15 m × 0.25
mm × 0.10 μm) and flame ionization detector. Method: 80 °C (1
min)/0.8 °C min−1/105 °C (0 min)/30 °C min−1/220 °C (3 min).
The enantiomeric excess values of alcohols 5c, 6c, and 8c were
determined by chiral HPLC analysis with a Chiralcel OD column and
UV detector (210 nm): mobile phase n-hexane/i-PrOH 97/3, flow
rate 0.6 mL min−1. The enantiomeric excess values of ketone 7b and of
alcohol 7c were determined by chiral HPLC equipped with a Lux 5u
Cellulose-3 column and UV detector (210 nm): mobile phase n-
hexane/i-PrOH 97/3; flow rate 0.6 mL min−1. High-resolution MS
spectra were recorded with an FT-ICR (Fourier transform ion
cyclotron resonance) instrument, equipped with an ESI source.
Optical rotations were measured at 589 nm and are given in deg cm3
g−1 dm−1. X-ray diffraction data were collected on a diffractometer
using Mo Kα radiation (λ = 0.71073 Å). CIF files containing
crystallographic data can be obtained free of charge from the
Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/
data_request (CCDC reference file 918584). TLC analyses were
performed on precoated silica gel 60 F254 plates, and spots were
visualized either by UV light (254 nm) or by spraying with
phosphomolybdic acid reagent. All chromatographic separations
were carried out on silica gel columns (230−400 mesh). Protein
concentrations were determined according to Bradford, using bovine
serum albumine (BSA) as a standard.
2H-Chromene-3-carbaldehyde (5a): 17.61 g, 83% yield, yellow
wax; 1H NMR (500 MHz, CDCl3) δ 9.51 (s, 1H), 7.25 (dt, J = 7.5, 1.9
Hz, 1H), 7.18 (d, J = 0.9 Hz, 1H), 7.16 (dd, J = 7.5, 1.9 Hz, 1H), 6.92
(dt, J = 7.5, 0.9 Hz, 1H), 6.83 (d, J = 8.5 Hz, 1H), 4.98 (d, J = 1.4 Hz,
2H); 13C NMR (101 MHz, CDCl3) δ 189.1, 155.5, 140.4, 132.6,
131.1, 128.8, 121.4, 120.0, 115.9, 62.7; HRMS (ESI) m/z (M + H)+
+
calcd for C10H9O2 161.0597, found 161.0599; GC tr 18.01 min.
5-Methoxy-2H-chromene-3-carbaldehyde (6a): 21.45 g, 85%
1
yield, yellow solid, mp 51−58 °C; H NMR (500 MHz, CDCl3) δ
9.48 (s, 1H), 7.56 (s, 1H), 7.18 (t, J = 8.3 Hz, 1H), 6.44 (d, J = 8.3 Hz,
1H), 6.41 (d, J = 8.3 Hz, 1H), 4.92 (s, 2H), 3.83 (s, 3H); 13C NMR
(101 MHz, CDCl3) δ 189.4, 157.0, 156.6, 136.4, 133.4, 129.4, 110.4,
108.8, 103.3, 62.5, 55.5; HRMS (ESI) m/z (M + H)+ calcd for
+
C11H11O3 191.0703, found 191.0706; GC tr 22.33 min.
1-(2H-Chromen-3-yl)ethanone (7a): 16.42 g, 71% yield, colorless
1
solid crystals, mp 48−50 °C (Et2O); H NMR (400 MHz, CDCl3) δ
7.18 (s, 1H), 7.14 (td, J = 7.7, 1.5 Hz, 1H), 7.06 (dd, J = 7.7, 1.8 Hz,
1H), 6.81−6.86 (m, 1H), 6.75 (d, J = 8.1 Hz, 1H), 4.89 (s, 2H), 2.27
(s, 3H); 13C NMR (101 MHz, CDCl3) δ 195.5, 155.5, 133.7, 132.3,
130.6, 129.2, 121.7, 120.7, 116.2, 64.2, 24.8; HRMS (ESI) m/z (M +
H)+ calcd for C11H11O2+ 175.0754, found 175.0747; GC tr 26.40 min.
Synthesis of 5-Methoxy-3,4-dihydronaphthalene-2-carbal-
dehyde (8a). Method A (via Shapiro Reaction). N′-(5-Methoxy-3,4-
dihydronaphthalen-2(1H)-ylidene)-4-methylbenzenesulfonohydra-
zide (9). To a slurry of tosylhydrazine (3.8 g, 20.6 mmol) in EtOH (10
mL) was added a solution of 5-methoxy-2-tetralone (4.0 g, 22.7 mmol)
in EtOH (45 mL). The mixture was stirred for 24 h and then filtered.
The solid was washed with EtOH (2 × 20 mL) and dried to afford 9 as
Enzymes and Strains. OYE2-3 from Saccharomyces cerevisiae and
GDH from Bacillus megaterium were overexpressed in E. coli BL21
(DE3) strains harboring a specific plasmid, according to standard
molecular biology techniques as described in ref 23. CPADH from
Candida parapsilosis and READH from Rhodococcus erythropolis were
purchased from Julich. HLADH from horse liver, BYADH from
̈
baker’s yeast, PLADH from Parvibaculum lavamentivorans, DRADH
from Deinococcus radiodurans, LKADH from Lactobacillus kefir,
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a mixture of two isomers: 6.5 g, 91% yield, yellow powder, mp 155−
then was diluted with CH2Cl2 (40 mL). The solution was washed with
aqueous HCl solution (1 N, 40 mL), and the organic phase was
separated and dried over Na2SO4 and concentrated under reduced
pressure. Purification by column chromatography (EtOAc/n-hexane,
10/90) of crude afforded compound 13: 10.95 g, 78% yield after two
steps; 1H NMR (400 MHz, CDCl3) δ 7.16 (t, J = 7.8 Hz, 1H), 6.73 (d,
J = 7.8 Hz, 1H), 6.69 (d, J = 7.8 Hz, 1H), 3.82 (s, 3H), 3.58 (d, J =
17.1 Hz, 1H), 3.49 (d, J = 17.1 Hz, 1H), 3.17 (s, 3H), 2.91−3.06 (m,
2H), 2.45−2.57 (m, 2H); 13C NMR (101 MHz, CDCl3) δ 157.0,
130.4, 127.5, 126.9, 122.1, 121.0, 120.9, 108.5, 55.3, 40.3, 40.2, 33.1,
20.3; HRMS (ESI) m/z (M + H)+ calcd for C13H16NO4S+ 282.0795,
found 282.0791.
1
162 °C; H NMR (400 MHz, CDCl3, 1/1 mixture of two isomers) δ
7.83 (d, J = 8.0 Hz, 2 H, isomer A), 7.79 (d, J = 8.0 Hz, 2H, isomer B),
7.24−7.27 (m, 4H, isomer A + B), 7.07−7.01 (m, J = 8,4 Hz, 2H
isomer A + B), 6.58−6.68 (m, 4H, isomer A + B), 3.72 (s, 3H, isomer
A), 3.71 (s, 3H, isomer B), 3.45 (s, 2 H, isomer A), 3.42 (s, 2 H,
isomer B), 2.80 (t, J = 7.0 Hz, 2H, isomer A), 2.74 (t, J = 7.0 Hz, 2H,
isomer B), 2.43 (t, J = 7.0 Hz, 2H, isomer A), 2.35 (s, 3H, isomer A),
2.34 (s, 3H, isomer B), 2.29 (t, J = 7.0 Hz, 3H, isomer B); 13C NMR
(101 MHz, CDCl3, 1/1 mixture of two isomers) δ 159.6 and 159.4
(isomer A + B), 156.5 and 155.9 (isomer A + B), 144.1 and 144.0
(isomer A + B), 136.2 (isomer A), 135.4 and 135.3 (isomer A + B),
133.1 (isomer B), 129.6 (2C, isomer A + B), 128.0 and 127.9 (2C,
isomer A + B), 127.2 and 127.1 (isomer A + B), 125.6 and 125.4
(isomer A + B), 120.8 and 119.9 (isomer A + B), 108.4 and 108.0
(isomer A + B), 55.4 and 55.3 (isomer A + B), 38.0 (isomer A), 31.5
(isomer B), 30.5 (isomer A), 25.3 (isomer B), 21.7 and 21.6 (isomer A
+ B), 21.5 (isomer A), 19.5 (isomer B); HRMS (ESI) m/z (M + H)+
calcd for C18H21N2O3S+ 345.1267, found 345.1271.
5-Methoxy-3,4-dihydronaphthalene-2-carbaldehyde (8a). Hydra-
zone 3b (2.41 g, 6.97 mmol) in THF (50 mL) was cooled to −40 °C.
TMEDA (2.2 mL) and n-BuLi (6.2 mL, 2.5 M hexane solution, 15.15
mmol) were added dropwise to the reaction mixture, which was stirred
at −40 °C for 1 h. Then the mixture was warmed to 0 °C and N2
evolution took place. After 30 min DMF (1.6 mL, 21 mmol) was
added. The reaction mixture was stirred for 12 h at 0 °C. The reaction
mixture was then treated with water (20 mL) and warmed to room
temperature. The reaction mixture was diluted with Et2O (30 mL),
and the layers were separated. The aqueous layer was extracted with
Et2O (3 × 30 mL). The combined organic layers were dried with
Na2SO4 and concentrated under reduced pressure. The residue was
purified by chromatography (EtOAc/n-hexane, 10/90) to afford an
inseparable mixture of 8a and its regioisomer 10 (4;6): 420 mg, 32%
yield.
5-Methoxy-3,4-dihydronaphthalene-2-carbonitrile (13). Com-
pound 12 (10.0 g, 0.035 mol) was dissolved in pyridine (20 mL)
and then heated at 60 °C for 12 h. After completion of the reaction,
the solvent was removed under reduced pressure. The crude product
was then dissolved in EtOAc (30 mL) and washed with aqueous HCl
solution (1 N, 3 × 20 mL). The organic phase was separated and dried
with Na2SO4. After evaporation of the solvent the crude product was
purified by column chromatography (EtOAc/n-hexane, 10/90) to
afford the product 13 and its regioisomer (93/7). Further
crystallization from cold n-hexane/Et2O (9/1) afforded 13 as a single
isomer: 4.7 g, 72% yield, yellow-brown powder, mp 62−65 °C
1
(hexane/Et2O, 9/1); H NMR (400 MHz, CDCl3) δ 7.20 (t, J = 8.2
Hz, 1 H) 7.15 (m, 1H), 6.90 (d, J = 8.2 Hz, 1 H), 6.79 (d, J = 7.4 Hz, 1
H), 3.84−3.87 (m, 3 H), 2.91 (t, J = 8.6 Hz, 2 H), 2.52 (td, J = 8.6, 1.5
Hz, 2 H); 13C NMR (101 MHz, CDCl3) δ 156.3, 141.6, 132.0, 127.4,
123.3, 120.6, 119.7, 112.8, 109.8, 55.6, 24.2, 19.1; HRMS (ESI) m/z
(M + H)+ calcd for C12H12NO+ 186.0913, found 186.0907.
5-Methoxy-3,4-dihydronaphthalene-2-carbaldehyde (8a). A sol-
ution of DIBAL-H in toluene (1.5 M, 20 mL) was added dropwise to a
solution of 13 (4.50 g, 24 mmol) in THF (60 mL) at 0 °C. The
reaction mixture was slowly warmed to room temperature and stirred
for an additional 5 h before being quenched with saturated aqueous
Rochelle salt (20 mL). The resulting mixture was partitioned between
aqueous HCl (2 N, 20 mL) and EtOAc (40 mL). The organic layer
was washed with aqueous saturated NaHCO3 (2 × 40 mL). The
organic phase was dried over Na2SO4, and the solvent was removed
under reduced pressure. Purification by column chromatography
(EtOAc/n-hexane, 10/90) afforded the product 8a: 3.38 g, 75% yield,
Method B (via Cyanohydrin Formation). 5-Methoxy-2-(trime-
thylsilyloxy)-1,2,3,4-tetrahydronaphthalene-2-carbonitrile (11). To
a solution of NaCN (4.92 g, 0.1 mol) in dry DMSO (50 mL) at 60 °C
was added 5-methoxy-2-tetralone (8.81 g, 0.05 mol). The mixture was
stirred for 15 min at 60 °C, and then TMSCl (7.5 mL, 0.06 mol) was
added dropwise. After 15 min the reaction mixture was cooled to room
temperature and diluted with water. The solution was extracted with
ether (3 × 30 mL). The collected organic phases were washed with
brine (2 × 30 mL). After the extract was dried with Na2SO4, the
solvent was removed under reduced pressure and the residue was
purified by chromatography (EtOAc/n-hexane, 5/95) to give the TMS
1
oil which after prolonged standing turned into a solid wax; H NMR
(250 MHz, CDCl3) δ 9.68 (s, 1H), 7.19−7.27 (m, 2H), 6.95−6.92 (m,
2H), 3.87 (s, 3H), 2.89 (t, J = 8.9 Hz, 2H), 2.55 (t, J = 8.9 Hz, 2H);
13C NMR (63 MHz, CDCl3) δ 192.8, 156.5, 145.5, 139.3, 133.1, 127.2,
126.1, 121.5, 113.0, 55.6, 19.4, 18.7; HRMS (ESI) m/z (M + H)+ calcd
1
derivative 11 (8.8 g, 64% yield, oil): H NMR (250 MHz, CDCl3) δ
+
for C12H13O2 189.0910, found 189.0916; GC tr 22.67 min.
7.15 (t, J = 7.9 Hz, 1 H), 6.71 (t, J = 8.4 Hz, 2 H), 3.84 (s, 3 H), 3.32
(d, J = 16.5 Hz, 1 H), 3.11 (d, J = 15.6 Hz, 1 H), 2.80−3.04 (m, 2 H),
2.22−2.34 (m, 1 H), 2.03−2.17 (m, 1 H), 0.29 (s, 9 H); 13C NMR (63
MHz, CDCl3) δ 157.0, 132.6, 126.9, 123.0, 121.6, 121.1, 107.9, 68.4,
55.2, 42.9, 35.4, 20.8, 1.4; HRMS (ESI) m/z (M + H)+ calcd for
C15H22NO2Si+ 276.1414, found 276.1418.
General Procedure for the Synthesis of Compounds 5c−8c.
To a solution of the unsaturated carbonyl precursor 5a−8a (10.0
mmol) in MeOH (50 mL) was added NaBH4 (850 mg) in one portion
at 0 °C. After completion of the reaction (by TLC and GC
monitoring: tr 20.25 min for 5d, tr 23.25 min for 6d, tr 29.28 min for
7d, tr 23.15 min for 8d) the solution was acidified with aqueous HCl
(1 N, 30 mL), the solvent was evaporated under reduced pressure, and
the residue was neutralized with aqueous saturated NaHCO3. The
solution was extracted with EtOAc (3 × 30 mL), and the organic
phases were dried over Na2SO4. The solvent was evaporated under
reduced pressure to give the allylic alcohols 5d−8d, which were
submitted to the next step without further purification. Thus, 5d−8d
were dissolved in EtOH (10 mL) and 10% Pd/C (100 mg) was added.
The mixture was stirred under a H2 atmosphere overnight. The
solution was filtered and the solvent evaporated under reduced
pressure. The crude product was purified by column chromatography
(EtOAc/n-hexane, 10/90) to afford the alcohols 5c−8c.
2-Cyano-5-methoxy-1,2,3,4-tetrahydronaphthalen-2-yl metha-
nesulfonate (12). To a solution of 11 (8.0 g, 0.029 mol) in THF
(50 mL), an aqueous HCl solution (3 N, 20 mL) was added. The
mixture was stirred for 3 h and then diluted with water (30 mL). The
two layers were separated, and the aqueous layer was extracted with
EtOAc (3 × 50 mL). The organic layers were then dried with
anhydrous Na2SO4 and filtered. The solvent was removed under
reduced pressure to yield 2-hydroxy-5-methoxy-1,2,3,4-tetrahydro-
naphthalene-2-carbonitrile, which was used in the next step without
further purification: 1H NMR (400 MHz, CDCl3) δ 7.14 (t, J = 7.8 Hz,
1H), 6.72 (d, J = 7.8 Hz, 1H), 6.69 (d, J = 7.8 Hz, 1H), 3.82 (s, 3H),
3.33 (d, J = 16.4 Hz, 1H), 3.10 (d, J = 16.4 Hz, 1H), 2.83−3.00 (m,
2H), 2.59 (br. s., 1H), 2.23−2.31 (m, 1H), 2.11−2.20 (m, 1H); 13C
NMR (101 MHz, CDCl3) δ 157.1, 131.9, 127.1, 122.8, 121.6, 121.2,
Chroman-3-ylmethanol (5c): 1.22 g, 74% yield, yellow oil; 1H
NMR (400 MHz, CDCl3) δ 6.93 (t, J = 2.2 Hz, 1H), 6.88 (dd, J = 1.9,
0.2 Hz, 1H), 6.64−6.73 (m, 2H), 4.18 (dt, J = 2.6, 0.7 Hz, 1H), 3.86
(dd, J = 2.6, 1.9 Hz, 1H), 3.55−3.41 (m, 3H), 2.76 (dd, J = 3.9, 1.4 Hz,
1H), 2.49 (dd, J = 4.0, 2.0 Hz, 1H), 2.10−2.23 (m, 1H); 13C NMR
(101 MHz, CDCl3) δ 154.2, 129.7, 126.9, 120.8, 120.2, 116.2, 67.3,
+
108.2, 67.3, 55.3, 41.3, 33.8, 20.3; HRMS (EI) calcd for C12H14NO2
204.1019, found 204.1014). To a stirred solution of the crude
cyanohydrin in CH2Cl2 (50 mL) were added pyridine (10 mL) and
MsCl (2.47 mL, 0.032 mol). The mixture was stirred overnight and
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+
62.7, 34.5, 27.1; HRMS (ESI) m/z (M + H)+ calcd for C10H13O2
165.0910, found 165.0906; GC tr 19.36 min.
(0.1 mM), GDH (4 U mL−1), and an OYE (150 μg mL−1). The
mixture was stirred for 12 h in an orbital shaker (160 rpm, 30 °C). The
solution was decanted, and both the resin and the aqueous phase were
extracted with EtOAc (500 μL each), centrifuging after each extraction
(15000g, 1.5 min). The combined organic solutions were dried over
Na2SO4 for further analysis.
General Procedure for the Screening-Scale ADH-Mediated
Reductions. The same procedure as for OYE-mediated reduction was
followed, adding an ADH (100 μg mL−1) instead of the OYE and
adding also NAD+ (0.1 mM) to the reaction mixture. In this case,
instead of a single starting material, a ∼1/1 mixture of the unsaturated
(5a−8a) and the racemic saturated (5b−8b) substrates was loaded on
the resin (according to Table 1). The crude mixtures were analyzed by
GC.
General Procedure for the Screening-Scale Multienzymatic
Cascade Reductions. The same procedure as for OYE-mediated
reduction was followed, adding also an ADH (100 μg mL−1) and
NAD+ (0.1 mM) to the reaction mixture (according to Table 2). The
crude mixtures were analyzed by GC.
General Procedure for the Preparative Scale Biotransforma-
tions. For all of the substrates a similar protocol was followed (either
as a cascade system or as a one-pot sequential system) on a larger
scale, employing the OYE and the ADH which provided the best
conversion and/or ee.
(5-Methoxychroman-3-yl)methanol (6c): 1.51 g, 78% yield, oil; 1H
NMR (400 MHz, CDCl3) δ 7.05 (t, J = 8.2, Hz, 1H), 6.48 (d, J = 8.2
Hz, 1H), 6.42 (d, J = 8.2 Hz, 1H), 4.27 (ddd, J = 10.7, 3.1, 1.4 Hz,
1H), 3.95 (dd, J = 10.7, 7.7 Hz, 1H), 3.81 (s, 3H), 3.73 (dd, J = 10.8,
5.8 Hz, 1H), 3.64 (dd, J = 10.8, 7.8 Hz, 1H), 2.79 (dd, J = 17.1, 6.0 Hz,
1H), 2.38 (dd, J = 17.1, 7.8 Hz, 1H), 2.29−2.19 (m, 1H), 1.62 (br s,
1H); 13C NMR (101 MHz, CDCl3) δ 158.2, 155.4, 126.9, 110.2,
109.4, 102.0, 67.3, 63.6, 55.4, 34.4, 21.9; HRMS (ESI) m/z (M + H)+
calcd for 195.1016, found 195.1020; GC tr 22.74 min.
1-(Chroman-3-yl)ethanol (7c): 1.22 g, 69% yield, wax; the product
was obtained as an inseparable mixture of syn/anti (4/6) isomers (vide
infra for spectra of isolated isomers); GC tr 26.77 min for anti-7c and
27.06 min for syn-7c.
5-Methoxy-1,2,3,4-tetrahydronaphthalen-2-yl)methanol (8c):
1.57 g, 82% yield, oil; H NMR (500 MHz, CDCl3) δ 7.10 (t, J =
1
8.0 Hz, 1 H), 6.74 (d, J = 8.0 Hz, 1 H), 6.68 (d, J = 8.0 Hz, 1 H), 3.84
(s, 3 H), 3.64 (d, J = 6.6 Hz, 2 H), 2.85−2.96 (m, 2 H), 2.48−2.61 (m,
2 H), 2.01−2.09 (m, 1 H), 1.90−2.00 (m, 1 H), 1.65−1.75 (br s, 1 H),
1.37−1.47 (m, 1 H); 13C NMR (126 MHz, CDCl3) δ 157.6, 137.6,
126.3, 125.9, 121.8, 107.3, 68.0, 55.6, 37.0, 32.9, 25.8, 22.9; HRMS
(ESI) m/z (M + H)+ calcd for C12H17O2+ 193.1223, found 193.1217;
GC tr 21.65 min.
Method A (Multienzymatic Cascade System). The substrate 5a,
6a, or 8a (1.25 mmol) adsorbed on resin (750 mg) was added to a KPi
buffer solution (100 mL, 50 mM, pH 7.0) containing OYE2 (150 μg
mL−1), HLADH (100 μg mL−1), GDH (400 U), glucose (5 mmol,
900 mg), NAD+ (10 μmol, 6.6 mg), and NADP+ (10 μmol, 7.4 mg).
The reaction mixture was stirred in an orbital shaker (160 rpm, 30 °C)
and monitored by GC until complete conversion, with further addition
of aliquots of OYE2, HLADH, and GDH if necessary. The solution
was decanted, and both the resin and the aqueous phase were
extracted with EtOAc (2 × 20 mL). The organic phase was dried over
Na2SO4 and concentrated under reduced pressure. The crude
materials were submitted to column chromatographic purification
(EtOAc/n-hexane 10/90).
General Procedure for the Preparation of Compounds 5b−
8b. To a solution of the alcohols 5c−8c (1.0 mmol) in CH2Cl2 (20
mL) was added Dess−Martin periodinane (508 mg, 1.2 mmol). After
completion of the reaction by TLC monitoring, saturated aqueous
Na2SO3 (8 mL) was added and the mixture stirred for 30 min. The
organic layer was separated, and the aqueous phase was extracted with
EtOAc (3 × 30 mL). The collected organic phases were then washed
with saturated aqueous NaHCO3, separated, and dried with Na2SO4.
The solvent was evaporated under reduced pressure, and the crude
residue was purified by column chromatography (EtOAc/n-hexane,
10/90) to afford the products 5b−8b.
1
Chroman-3-carbaldehyde (5b): 134 mg, 83% yield, oil; H NMR
(400 MHz, CDCl3) δ 9.83 (d, J = 0.8 Hz, 1H), 7.08−7.15 (m, 2H),
6.90 (td, J = 7.4, 1.2 Hz, 1H), 6.83 (d, J = 8.6 Hz, 1H), 4.41 (ddd, J =
11.3, 3.5, 1.2 Hz, 1H), 4.34 (ddd, J = 11.3, 6.7, 0.8 Hz, 1H), 3.12 (dd, J
= 16.4, 7.4 Hz, 1H), 3.02 (dd, J = 16.4, 5.9 Hz, 1H), 2.90−2.98 (m,
1H); 13C NMR (101 MHz, CDCl3) δ 200.6, 154.4, 129.8, 127.6,
121.0, 119.6, 116.9, 64.6, 45.2, 24.5; HRMS (ESI) m/z (M + H)+ calcd
Bioreduction of 5a with OYE2 and HLADH gave (S)-5c: 184 mg,
90% isolated yield, 91% ee by HPLC (tR(R) = 34.6 min, tR(S) = 37.5
min); [α]25 = −19.6 (c 1.2, CHCl3), lit.32 [α]25 = −19.0 (c 0.3,
D
D
CHCl3).
Bioreduction of 6a with OYE2 and HLADH gave (S)-6c: 201 mg,
83% isolated yield, 99% ee by HPLC (tR(S) = 39.2 min, tR(R) = 42.9
min); [α]D = −6.9 (c 1.2, CHCl3), lit.25b [α]D = −6.2 (c 1.17, CHCl3).
Bioreduction of 8a with OYE2 and HLADH gave (S)-8c: 214 mg,
89% isolated yield, 99% ee by HPLC (tR(R) = 22.8 min, tR(S) = 23.6
min); [α]D = −42.2 (c 1.5, CHCl3).
Method B (Multienzymatic One-Pot Sequential System). Sub-
strate 7a (1.25 mmol, 220 mg) adsorbed on resin (750 mg) was added
to a KPi buffer solution (100 mL, 50 mM, pH 7.0) containing OYE2
(150 μg mL−1), GDH (400 U), glucose (5 mmol, 900 mg), and
NADP+ (10 μmol, 7.4 mg). The reaction mixture was stirred in an
orbital shaker (160 rpm, 30 °C) and monitored by GC until complete
conversion, and then the required ADH (100 μg mL−1) and NAD+
(10 μmol, 6.6 mg) were added in one portion. The stirring was
continued until complete conversion, with further addition of aliquots
of ADH and GDH if necessary. The mixture was then processed as
described in Method A. The products 7c were then recrystallized from
cold n-hexane in order to improve the de.
+
for C10H11O2 163.0754, found 163.0751; GC tr 17.02 min.
5-Methoxychroman-3-carbaldehyde (6b): 165 mg, 86% yield, oil;
1H NMR (500 MHz, CDCl3) δ 9.83 (s, 1H), 7.07 (t, J = 8.5 Hz, 1H),
6.48 (d, J = 8.5 Hz, 1H), 6.46 (d, J = 8.5 Hz, 1H), 4.36 (dd, J = 10.8,
2.8 Hz, 1H), 4.30 (dd, J = 10.8, 6.1 Hz, 1H), 3.84 (s, 3 H), 3.82 (m,
1H), 2.88−2.99 (m, 2H); 13C NMR (126 MHz, CDCl3) δ 201.5,
158.4, 155.5, 127.7, 109.8, 109.3, 102.7, 64.6, 55.8, 45.0, 19.7; HRMS
(ESI) m/z (M + H)+ calcd for C11H13O3+ 193.0859, found 193.0855;
GC tr 21.25 min.
1-(Chroman-3-yl)ethanone (7b): 142 mg, 81% yield, oil; 1H NMR
(500 MHz, CDCl3) δ 7.06−7.12 (m, 2H), 6.87 (td, J = 7.4, 1.2 Hz,
1H), 6.82 (d, J = 8.0 Hz, 1H), 4.40 (ddd, J = 10.8, 2.8, 2.6 Hz, 1H),
4.04 (ddd, J = 10.8, 7.5, 1.4 Hz, 1H), 2.88−3.04 (m, 3H), 2.25 (s, 3H);
13C NMR (126 MHz, CDCl3) δ 207.9, 154.5, 130.1, 127.8, 121.1,
120.8, 117.0, 66.7, 46.1, 28.9, 27.5; HRMS (ESI) m/z (M + H)+ calcd
+
for C11H13O2 177.0910, found 177.0907; GC tr 22.02 min.
5-Methoxy-1,2,3,4-tetrahydronaphthalene-2-carbaldehyde (8b):
175 mg, 92% yield, oil; 1H NMR (400 MHz, CDCl3) δ 9.72 (d, J = 1.2
Hz, 1H), 7.03 (t, J = 7.6 Hz, 1H), 6.69 (d, J = 7.4 Hz, 1H), 6.60 (d, J =
7.8 Hz, 1H), 3.74 (s, 3H), 2.78−2.92 (m, 3H), 2.47−2.64 (m, 2H),
2.11−2.22 (m, 1H), 1.60−1.73 (m, 1H); 13C NMR (101 MHz,
CDCl3) δ 203.9, 157.2, 135.6, 126.3, 124.9, 121.4, 107.3, 55.2, 46.6,
Bioreduction of 7a with OYE2 and no ADH gave (R)-7b: 211 mg,
96% isolated yield, 99% ee by HPLC (tR(S) = 29.1 min, tR(R) = 30.7
min), [α]D = −39.1 (c 1.5, CHCl3).
Bioreduction of 7a with OYE2 and CPADH gave (1S,3′R)-7c: 158
mg, 71% isolated yield after recrystallization, colorless solid, mp 48−51
°C, 99% ee by HPLC (tR(1S,3′R) = 28.5 min, tR(1R,3′S) = 30.2 min,
tR(1R,3′R) = 31.3 min, tR(1S,3′S) = 36.0 min); [α]D = −50.1 (c 1.1,
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.06 (t, J = 8.1 Hz, 1H), 7.02
(d, J = 7.3 Hz, 1H), 6.77−6.85 (m, 2H), 4.42 (ddd, J = 10.8, 3.1, 1.8
Hz, 1H), 3.95 (dd, J = 10.8, 8.8 Hz, 1H), 3.72 (quin, J = 6.6 Hz, 1H),
2.78 (dd, J = 15.9, 5.1 Hz, 1H), 2.57 (dd, J = 15.9, 9.4 Hz, 1H), 1.94−
+
28.7, 22.7, 22.0; HRMS (ESI) m/z (M + H)+ calcd for C12H15O2
191.1067, found 191.1073; GC tr 21.65 min.
General Procedure for the Screening-Scale OYE-Mediated
Reductions. The substrates 5a−8a (5 mg) adsorbed on XAD 1180
resin (15 mg) were added to a KPi buffer solution (1.0 mL, 50 mM,
pH 7.0) containing glucose (4 equiv with respect to 5a−8a), NADP+
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Article
+
2.04 (m, 1H), 1.47 (s, 1H), 1.30 (d, J = 6.2 Hz, 3H); 13C NMR (126
MHz, CDCl3) δ 155.1, 130.2, 127.6, 121.5, 120.7, 116.8, 68.8, 68.0,
CHCl3); HRMS (ESI) m/z (M + H)+ calcd for C12H15O3 207.1016,
found 207.1010.
+
40.2, 28.3, 21.8; HRMS (ESI) m/z (M + H)+ calcd for C11H15O2
General Procedure for the Preparation of (R)-5f, (R)-6f, and
(S)-8f. To solutions of the carboxylic acids (R)-5e, (R)-6e, and (S)-8e
(0.87 mmol) in toluene (3 mL) were added DPPA (234 μL, 1.1
mmol) and Et3N (151 μL, 1.1 mmol). The reaction mixture was
refluxed for 3 h, and then benzyl alcohol (224 μL, 2.2 mmol) was
added. After 2 h the reaction mixture was cooled to room temperature
and the solvent was removed under reduced pressure. The residual oil
was purified by chromatography (EtOAc/n-hexane, 5/95) to yield the
N-Cbz-protected amine.
179.1067, found 179.1072.
Bioreduction of 7a with OYE2 and KRED gave (1R,3′R)-7c: 163
mg, 73% isolated yield after recrystallization, colorless powder, mp <45
°C, 99% ee by HPLC (tR reported above); [α]D = −50.6 (c 0.7,
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.08−7.18 (m, 2H), 6.90 (td,
J = 7.6, 1.4 Hz, 1H), 6.85 (d, J = 8.6 Hz, 1H), 4.23 (ddd, J = 11.0, 3.4,
1.3 Hz, 1H), 3.94 (dd, J = 11.0, 9.2 Hz, 1H), 3.85 (quin, J = 6.4, 1H),
2.85 (dd, J = 16.2, 6.4, 1H), 2.76 (dd, J = 16.2, 11.0 Hz, 1H), 1.90−
2.02 (m, 1H), 1.22 (d, J = 6.4 Hz, 3H); 13C NMR (101 MHz, CDCl3)
δ 154.7, 130.1, 127.2, 121.6, 120.4, 116.4, 68.2, 68.1, 39.6, 26.4, 21.3;
(R)-Benzyl chroman-3-ylcarbamate ((R)-5f): 207 mg, 84% yield,
oil; H NMR (400 MHz, CDCl3) δ 7.24−7.38 (m, 5H), 7.10 (td, J =
1
+
HRMS (ESI) m/z (M + H)+ calcd for C11H15O2 179.1067, found
7.7, 0.8 Hz, 1H), 7.01 (d, J = 7.3 Hz, 1H), 6.87 (dt, J = 7.6, 1.4 Hz,
1H), 6.82 (dd, J = 8.1, 1.1 Hz, 1H), 5.09 (br. s., 3H), 4.23 (br s, 1H),
4.05−4.14 (m, 2H), 3.10 (dd, J = 16.5, 5.0 Hz, 1H), 2.75 (d, J = 16.5
Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 155.7, 153.9, 136.3, 130.4,
128.5, 128.2, 128.1, 127.8, 121.2, 119.1, 116.8, 68.2, 66.9, 44.0, 31.1;
179.1070.
Baker’s Yeast Mediated Reduction of Ketone 7a. To a
mechanically stirred mixture of commercial baker’s yeast (250 g) in tap
water (2.0 L) at 30 °C was added a solution of glucose (100 g) in
water (200 mL). After 1 h the substrate 7a (3.0 g, 17.2 mmol)
adsorbed on XAD 1180 resin (60 g) was added in one portion.
Vigorous stirring was continued for 5 days. After 24 h more baker’s
yeast (250 g) and glucose (100 g) were added. Then, the mixture was
filtered on a sintered glass funnel (porosity 0, >165 μm) and the
aqueous phase was extracted again with more resin (50 g). The
combined resin crops were washed with acetone (20 mL) and EtOAc
(3 × 50 mL). The organic solution was washed with brine (2 × 20
mL), dried over Na2SO4, and concentrated under reduced pressure to
give a brownish oil, which was submitted to column chromatographic
purification (EtOAc/n-hexane, 10/90), affording residual 7a (180 mg,
6% yield), ketone (R)-7b (1.27 g, 42% yield, colorless oil), and a
mixture of saturated alcohol 7c (800 mg, 26% yield, diastereomeric
ratio 77/23 syn-7c/anti-7c).
General Procedure for the Preparation of Acids (R)-5e, (R)-
6e, and (S)-8e. To stirred solutions of alcohols (S)-5c, (S)-6c, and
(R)-8c (1.0 mmol) in CH2Cl2/H2O (2/1, 6 mL) were added TEMPO
(46 mg, 0.3 mmol) and BAIB (640 mg, 2.0 mmol). After completion
of the reaction, the mixture was quenched with saturated aqueous
Na2SO3 solution (5 mL). Then, the mixture was washed with saturated
aqueous NaHCO3 (2 × 10 mL). The combined aqueous phase was
washed with EtOAc (5 mL) and acidified with HCl (1 M, 10 mL). The
aqueous solution was washed with EtOAc (3 × 20 mL). The
combined organic phase was dried over Na2SO4 and concentrated
under reduced pressure to give the corresponding acid without further
purification.
[α]25 +16.3 (c 1.2, CHCl3); HRMS (ESI) m/z (M + H)+ calcd for
D
+
C17H18NO3 284.1281, found 284.1289.
(R)-Benzyl 5-methoxychroman-3-ylcarbamate ((R)-6f): 248 mg,
91% yield, oil; H NMR (400 MHz, CDCl3) δ 7.26−7.36 (m, 5 H),
1
7.06 (t, J = 8.3 Hz, 1 H), 6.48 (d, J = 8.4 Hz, 1 H), 6.43 (d, J = 8.1 Hz,
1 H), 5.02−5.16 (m, 3 H), 4.23 (br. s., 1 H), 4.02−4.11 (m, 2 H), 3.78
(s, 3 H), 2.88 (dd, J = 17.4, 5.6 Hz, 1 H), 2.71 (br. d, J = 17.4 Hz, 1
H); 13C NMR (101 MHz, CDCl3) δ 153.2, 149.5, 149.4, 146.4, 131.1,
123.2, 122.8, 122.1, 104.1, 103.1, 97.3, 62.5, 61.5, 50.1, 38.3, 20.7;
[α]25 +46.4 (c 1, CHCl3); HRMS (ESI) m/z (M + H)+ calcd for
D
+
C18H20NO4 314.1387, found 314.1381.
(S)-Benzyl 5-methoxy-1,2,3,4-tetrahydronaphthalen-2-ylcarba-
mate ((S)-8f): 246 mg, 91% yield, oil; H NMR (400 MHz, CDCl3)
1
δ 7.30−7.40 (m, 5H), 7.10 (t, J = 8.0 Hz, 1H), 6.68 (d, J = 8.0 Hz,
2H), 5.11 (br s, 2H), 4.75−4.89 (m, 1H), 4.01−4.11 (m, 1H), 3.79−
3.86 (m, 3H), 3.11 (dd, J = 16.2, 4.4 Hz, 1H), 2.59−2.88 (m, 3H),
2.12−2.00 (m, 1H), 1.85−1.73 (m, 1H); 13C NMR (101 MHz,
CDCl3) δ 153.4, 151.0, 136.3, 134.9, 128.2, 127.8, 126.3, 126.1, 124.0,
121.3, 107.0, 66.3, 54.9, 46.0, 35.6, 28.0, 20.6; [α]25 +27.2 (c 1,
D
+
CHCl3); HRMS (ESI) m/z (M + H)+ calcd for C19H22NO3
312.1594, found 312.1600.
General Procedure for the Synthesis of Amines (R)-1, (R)-2,
and (S)-3. Compounds (R)-5f, (R)-6f, or (S)-8f (0.7 mmol) were
dissolved in MeOH (2 mL), and then 10% Pd/C (5 mg) and a few
drops of concentrated aqueous HCl were added. The mixture was
stirred under an H2 atmosphere for 12 h. Then the mixture was filtered
and the solvent was removed under reduced pressure to give the amine
without further purification.
(R)-Chroman-3-carboxylic acid ((R)-5e): 165 mg, 94% yield, yellow
1
1
solid, mp 125−128 °C; H NMR (400 MHz, CDCl3) δ 9.53 (br s,
(R)-Chroman-3-amine ((R)-1): 127 mg, 98% yield, oil; H NMR
1H), 6.98−7.07 (m, 2H), 6.82 (td, J = 8.1, 0.8 Hz, 1H), 6.77 (d, J = 8.1
Hz, 1H), 4.36 (d, J = 10.6 Hz, 1H), 4.06−4.15 (m, 1H), 2.93−3.07
(m, 3H); 13C NMR (126 MHz, CDCl3) δ 178.1, 154.3, 130.0, 128.0,
(HCl salt, 400 MHz, DMSO-d6) δ 8.74 (s, 3H), 7.08−7.16 (m, 2H),
6.89 (t, J = 7.6 Hz, 1H), 6.81 (d, J = 8.4 Hz, 1H), 4.27 (d, J = 11.2 Hz,
1H), 4.12−4.21 (m, 1H), 3.68 (br s, 1H), 3.17 (dd, J = 16.9, 4.1 Hz,
1H), 2.96 (dd, J = 16.9, 6.1 Hz, 1H); 13C NMR (HCl salt, 101 MHz,
DMSO-d6) δ 153.4, 130.0, 127.5, 121.1, 118.7, 116.5, 65.4, 43.3, 28.0;
121.2, 120.3, 117.1, 66.4, 38.6, 27.5; [α]25 +29.5 (c 1.3, CHCl3);
D
+
HRMS (ESI) m/z (M + H)+ calcd for C10H11O3 179.0703, found
[α]25 +60.2 (c 1.6, MeOH); HRMS (ESI) m/z (M + H)+ calcd for
179.0707.
D
C9H12NO+ 150.0913, found 150.0916.
(R)-5-Methoxychroman-3-carboxylic acid ((R)-6e): 172 mg, 92%
1
yield, yellow solid, mp 171−173 °C; H NMR (400 MHz, CDCl3) δ
(R)-5-Methoxy-3-aminochroman ((R)-2): 118 mg, 94% yield, oil;
1H NMR (400 MHz, CDCl3) δ 6.97 (t, J = 8.3 Hz, 1H), 6.40 (d, J =
8.4 Hz, 1H), 6.34 (dd, J = 8.1, 0.8 Hz, 1H), 3.99 (ddd, J = 10.4, 2.9, 1.5
Hz, 1H), 3.72 (s, 3H), 3.67 (ddd, J = 10.4, 7.1, 1.3 Hz, 1H), 3.22 (tdd,
J = 7.0, 7.0, 5.6, 3.1 Hz, 1H), 2.86 (ddd, J = 16.8, 5.6, 1.4 Hz, 1H), 2.32
(dd, J = 16.9, 6.9 Hz, 1H), 1.59 (br s, 2H); 13C NMR (101 MHz,
CDCl3) δ 158.3, 154.7, 127.0, 109.4, 109.1, 102.1, 70.9, 55.3, 43.7,
29.2; [α]25D −13.3 (c 1.5, CHCl3); HRMS (ESI) m/z (M + H)+ calcd
10.1−9.3 (s br, 1H), 7.08 (t, J = 8.2, Hz, 1H), 6.50 (dd, J = 8.2, 0.8 Hz,
1H), 6.45 (dd, J = 8.2, 0.8 Hz, 1H), 4.41 (ddd, J = 10.7, 3.2, 1.7 Hz,
1H), 4.13 (dd, J = 10.7, 8.2 Hz, 1H), 3.83 (s, 3H), 3.04 (m, 2H), 2.89
(dd, J = 18.5, 10.7 Hz, 1H); 13C NMR (101 MHz, CDCl3) δ 178.5,
158.0, 154.8, 127.2, 109.4, 102.3, 65.7, 55.4, 38.0, 29.6, 22.0; [α]25
D
−14.9 (c 1.2, CHCl3); HRMS (ESI) m/z (M + H)+ calcd for
+
C11H13O4 209.0808, found 209.0811.
+
(S)-5-Methoxy-1,2,3,4-tetrahydronaphthalene-2-carboxylic acid
((S)-8e): 181 mg, 89% yield, colorless solid, mp 149−151 °C; 1H
NMR (400 MHz, CDCl3) δ 9.14−10.14 (s, br, 1H), 7.08 (t, J = 7.9
Hz, 1H), 6.72 (d, J = 7.8 Hz, 1H), 6.67 (d, J = 8.1 Hz, 1H), 3.80 (s,
3H), 2.98−3.05 (m, 1H), 2.89−2.98 (m, 1H), 2.68−2.77 (m, 1H),
2.59 (ddd, J = 17.6, 11.1, 6.2 Hz, 1H), 2.22−2.31 (m, 1H), 1.76−1.90
(m, 2H); 13C NMR (101 MHz, CDCl3) δ 180.9, 157.1, 135.9, 126.1,
for C10H14NO2 180.1019, found 180.1022.
(S)-5-Methoxy-2-amino-1,2,3,4-tetrahydronaphthalene ((S)-3):
143 mg, 96% yield, oil; H NMR (HCl salt, 400 MHz, DMSO-d6) δ
1
8.48 (br s, 3H), 7.10 (t, J = 8.1 Hz, 1H), 6.77 (d, J = 8.1 Hz, 1H), 6.69
(d, J = 7.3 Hz, 1H), 3.75 (s, 3H), 3.32 (br s, 1H), 3.05 (dd, J = 16.1,
3.7 Hz, 1H), 2.79−2.85 (m, 2H), 2.41−2.60 (m, 2H), 2.15 (br d, J =
10.3 Hz, 1H), 1.64−1.79 (m, 1H); 13C NMR (HCl salt, 101 MHz,
DMSO-d6) δ 157.2, 134.5, 127.2, 123.7, 121.4, 108.3, 55.7, 46.8, 33.4,
124.5, 121.1, 107.1, 55.0, 39.4, 31.4, 25.3, 22.3; [α]25 −19.3 (c 1.6,
D
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Article
26.7, 21.8; [α]25D −60.3 (c 1.3, MeOH); HRMS (ESI) m/z (M + H)+
(1S,3′R)-7c. This material is available free of charge via the
calcd for C11H16NO+ 178.1226, found 178.1220.
Internet at http://pubs.acs.org.
(S)-3-((R)-1-Azidoethyl)chroman ((1R,3′S)-14). To a stirred
solution of (1S,3′R)-7c (85 mg, 0.48 mmol) in CH2Cl2 (5 mL) and
pyridine (2 mL) was added TsCl (91 mg, 0.48 mmol). The mixture
was stirred overnight and then was diluted with CH2Cl2 (10 mL). The
solution was washed with aqueous HCl (1 N, 10 mL), and the organic
phase was dried with Na2SO4 and concentrated under reduced
pressure. Purification by column chromatography (EtOAc/n-hexane,
10/90) afforded the intermediate tosylate: 130 mg, 82% yield, yellow
oil; 1H NMR (400 MHz, CDCl3) δ 7.81 (d, J = 8.3 Hz, 2H), 7.36 (d, J
= 7.8 Hz, 2H), 7.08 (td, J = 8.3, 1.5 Hz, 1H), 7.01 (d, J = 7.3 Hz, 1H),
6.84 (td, J = 7.3, 1.5 Hz, 1H), 6.75 (dd, J = 8.3, 1.0 Hz, 1H), 4.60 (dq,
J = 7.2, 6.2 Hz, 1H), 4.22 (ddd, J = 10.8, 2.9, 1.9 Hz, 1H), 3.67 (dd, J =
10.8, 9.3 Hz, 1H), 2.76 (dd, J = 16.6, 1.5 Hz, 1H), 2.57 (dd, J = 16.1,
9.8 Hz, 1H), 2.47 (s, 3H), 2.18−2.27 (m, 1H), 1.41 (d, J = 6.4 Hz,
3H); 13C NMR (101 MHz, CDCl3) δ 154.3, 144.8, 134.0, 129.8 (2C),
129.7, 127.7 (2C), 127.5, 120.5, 120.1, 116.5, 79.6, 66.5, 37.9, 27.3,
21.6, 18.8. The tosylate (110 mg, 0.33 mmol) was dissolved in DMF
(5 mL), and NaN3 (33 mg, 0.5 mmol) was added. The reaction
mixture was stirred at 60 °C for 8 h, and then after cooling water (10
mL) was added. The mixture was extracted with Et2O (3 × 20 mL).
The combined organic phases were washed with brine (2 × 10 mL).
The organic phase was dried over Na2SO4, filtered, and concentrated
under reduced pressure. Purification of the crude product by column
chromatography (EtOAc/n-hexane, 5/95) afforded (1R,3′S)-14: 53
AUTHOR INFORMATION
Corresponding Author
*E-mail: francesco.gatti@polimi.it (F.G.G.); alessandro.
sacchetti@polimi.it (A.S.).
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
Prof. Claudio Fuganti (Politecnico di Milano) is gratefully
acknowledged for fruitful discussions and for providing a
sample of (S)-3·HCl. We thank Dr. Martin Hedberg
(AstraZeneca, UK) for the gift of a sample of (R)-2. Prof.
Sven Panke and Christian Femmer (ETH-Zurich, Department
̈
of Biosystems Science and Engineering, Basel, CH) are kindly
acknowledged for the help and advice given in the preparation
of plasmids and strains. We are particularly grateful to the
undergraduate students Marco Rigamonti, Francesco Rossi, and
Ernesto Mandarino, who contributed to the preliminary phase
of this work. This research has been was supported by EU
project INTENANT (Grant FP7-NMP2-SL2008-214129).
1
mg, 79% yield, yellow oil; H NMR (400 MHz, CDCl3) δ 0.01−7.14
(m, 2H), 6.85 (t, J = 7.2 Hz, 1H), 6.79 (d, J = 8.0 Hz, 1H), 4.19 (dt, J
= 10.8, 2.4 Hz, 1H), 3.94 (dd, J = 10.8, 8.4 Hz, 1H), 3.55 (quint, J =
6.4 Hz, 1H), 2.89 (dd, J = 16.5, 5.2 Hz, 1H), 2.77 (dd, J = 16.5, 8.4 Hz,
1H), 2.05 (dtq, J = 12.4, 6.0, 2.8 Hz, 1H), 1.40 (d, J = 6.0 Hz, 3H); 13C
NMR (101 MHz, CDCl3) δ 154.5, 130.1, 127.3, 120.8, 120.6, 116.5,
67.7, 58.3, 37.9, 27.3, 16.9; HRMS (ESI) m/z (M + H)+ calcd for
C11H14N3O+ 204.1131, found 204.1136; [α]25D −48.2 (c 2.3, CHCl3).
(R)-1-((S)-Chroman-3-yl)ethanamine ((1R,3′S)-4). (1R,3′S)-14
(30 mg, 0.15 mmol) was dissolved in EtOAc (5 mL), and then 10%
Pd/C (5 mg) was added. The mixture was stirred under an H2
atmosphere for 12 h. The mixture was filtered, and the solvent was
removed under reduced pressure to give (1R,3′S)-4 without further
purification: 25 mg, 97% yield, yellow oil; 1H NMR (400 MHz,
CDCl3) δ 7.04−7.08 (m, 2H), 6.84 (t, J = 7.2 Hz, 1H), 6.79 (d, J = 8.0
Hz, 1H), 4.27 (ddd, J = 10.8, 3.2, 1.2 Hz, 1H), 3.89 (dd, J = 10.8, 8.8
Hz, 1H), 2.97 (quin, J = 6.4 Hz, 1H), 2.89 (ddd, J = 16.4, 5.6, 2.0 Hz,
1H), 2.73 (dd, J = 16.4, 9.6 Hz, 1H), 1.86−1.96 (m, 1H), 1.20 (d, J =
6.4 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ 155.9, 130.3, 127.5,
122.7, 120.7, 116.7, 68.9, 48.3, 40.0, 27.9, 21.9; HRMS (ESI) m/z (M
+ H)+ calcd for C11H16NO+ 178.1226, found 178.1222; [α]25D −7.1 (c
1, CHCl3).
REFERENCES
■
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