Studies of the deracemization of (±)-2-hydroxy-1-tetralone by trichosporon cutaneum

Article abstract of DOI:10.1002/adsc.200600331

The diastereo- and enantioselective bioreduction of (±)-2-hydroxy-1- tetralone (6) to the corresponding enantiopure (1S,2R)-cis-1,2-dihydroxy-1,2,3, 4-tetrahydronaphthalene (1) (83% isolated yield, >99% ee), mediated by resting cells of the yeast Trichosporon cutaneum CCT 1903 through dynamic kinetic resolution is reported. Deracemization of (±)-6 was observed in kinetic studies on the biotransformation of the enantiomers (R)-6 and (S)-6.

Full text of DOI:10.1002/adsc.200600331

FULL PAPERS
DOI: 10.1002/adsc.200600331
Studies of the Deracemization of (Æ)-2-Hydroxy-1-tetralone by
Trichosporon cutaneum
InÞs Lunardi,a Tarcila Cazetta,a Gelson J. A. Conceiżo,b Paulo J. S. Moran,a
and J. Augusto R. Rodriguesa,*
a
Institute of Chemistry, State University of Campinas, CP 6154, 13084-971 Campinas, SP, Brazil
Fax : (+55)-19-3788-3023; e-mail: jaugusto@iqm.unicamp.br
Department of Basic Sciences, College of Animal Science and Food Engineering, University of S¼o Paulo, CP 23,
b
13635-900 Pirassununga, SP, Brazil
Received: July 4, 2006; Revised: December 13, 2006
Supporting information for this article is available on the WWW under http://asc.wiley-vch.de/home/.
Abstract: The diastereo- and enantioselective biore- of (Æ)-6 was observed in kinetic studies on the bio-
duction of (Æ)-2-hydroxy-1-tetralone (6) to the cor- transformation of the enantiomers (R)-6 and (S)-6.
responding enantiopure (1S,2R)-cis-1,2-dihydroxy-
1,2,3,4-tetrahydronaphthalene (1) (83% isolated
yield, >99% ee), mediated by resting cells of the Keywords: amino alcohols; biotransformations; hy-
yeast Trichosporon cutaneum CCT 1903 through dy- droxytetralone; isomerases; oxidoredutases; Tricho-
namic kinetic resolution is reported. Deracemization sporon cutaneum
Introduction
whereas others combine chemical and biocatalytic
steps.[6] In the latter case, enzymes or whole cells con-
Enantiomerically pure compounds are potentially tribute either to racemization[7] or to kinetic resolu-
useful intermediates for the synthesis of a wide varie- tion.[4]
ty of bioactive products in the pharmaceutical and
Following our research on DKR,[8] we now report
agrochemical industries.[1] Although asymmetric catal- the diastereo- and enantioselective preparation of
ysis has undergone an impressive development in the (1S,2R)-1,2-dihydroxy-1,2,3,4-tetrahydronaphthalene
last twenty years, as of today the most common way (1). The chiral cis-(1S,2R)-diol 1 can be a valuable in-
to obtain enantiomerically pure compounds in indus- termediate as a building block in asymmetric organic
try is still via resolution of racemic mixtures.[2] To synthesis. Furthermore, diols 1 and 2 are excellent
obtain a single enantiomer from such a mixture, one precursors to the cis-amino alcohol 3, which can be
can resolve it either by conventional separation tech- easily obtained through a Ritter reaction or by the
niques or by using an existing difference in reactivity methodology of Resnick et al. (Figure 1).[9] The amino
(kinetic resolution).[3] However, a major limitation of alcohol 3 has been used as a chiral ligand in the cata-
these techniques is that the maximum product yield is
only 50%. The unwanted enantiomer must be sepa-
rated, racemized and resubmitted to resolution in
order to increase this yield. These disadvantages can
be overcome by employing dynamic kinetic resolution
(DKR). DKR has recently become not only an alter-
native to traditional kinetic resolution, but also a new
procedure for asymmetric synthesis.[3] It is one of the
most useful and reliable methods to prepare a single
chiral compound bearing two or more stereocenters
starting from a racemate, with a theoretical yield of
100%. A number of successful examples for DKR
methods have been reviewed recently.[4] Some of
them involve solely conventional chemical methods,[5] Figure 1.
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InÞs Lunardi et al.
FULL PAPERS
lytic reduction of ketones and as a toolbox to probe cesses (92–99% ee) and reasonable isolated yields
ligand conformations in transition metal-catalyzed re- (38–58%), through the deracemization of (Æ)-3-hy-
actions (Figure 2, compound 5).[10,11] The cis-(1R,2S)- droxy-4-chromanone mediated by resting cells of Tri-
diol 1 has been utilized as chiral intermediate for the chosporon cutaneum CCT 1903.[18] We report herein
synthesis of the chiral crown ether (S,R,R,S)-4 the diastereo- and enantioselective preparation of
(Figure 2).[12]
(1S,2R)-1,2-dihydroxy-1,2,3,4-tetrahydronaphthalene
(1) in high enantiomeric excess (>99% ee) and excel-
lent chemical yield (83%) by the asymmetric reduc-
tion of racemic 2-hydroxy-1-tetralone (6) mediated by
fresh resting cells of the yeast Trichosporon cutaneum
through a DKR process.
Results and Discussion
The substrate (Æ)-6 was prepared according to the
methodology proposed by Moriarty, which is well es-
tablished in the literature.[19] Racemic 2-hydroxy-1-tet-
ralone (6) was added to a slurry of resting cells of T.
cutaneum (in distilled water). The yeast cells were
previously cultivated in Sabouraud dextrose broth
(SDB) for 3 days and then harvested by centrifuga-
tion. The reaction was monitored by periodic sam-
pling of aliquots (1 mL), which were extracted with
ethyl acetate and analyzed by GC/MS (Table 1).
After incubation on a shaker (170 rpm) at 308C for
12 days, (Æ)-6 was converted into a mixture of cis-
Figure 2.
(1S,2R)-diol
1
and (1R,2R)-trans-1,2-dihydroxy-
A few biocatalytic methods have been reported for 1,2,3,4-tetrahydronaphthalene (2). The major product,
preparation of the optically active diol 1. Boyd and cis-(1S,2R)-diol 1, was obtained in 83% isolated yield
co-workers have found that Pseudomonas putida UV4 and high enantiomeric excess (>99%) whilst the
and NCIMB 8859 oxidize 1,2-dihydronaphthalene to minor product, trans-(1R,2R)-diol 2, was obtained in
cis-(1S,2R)-diol 1 as a minor metabolite (ca. 21% rel- 3% isolated yield and >99% ee (Scheme 1).
ative yield, >98% ee) and to the cis-(1R,2S)-diol 1
antipode as a major metabolite (ca. 30% relative
yield, >98% ee).[13,14] Boyd also reported the forma-
tion of trans-(1R,2R)-diol 2 as the major biotransfor-
mation product of 1,2-dihydronaphthalene by rat liver
microsomes in 75% (54% ee) and 60% (4% ee) rela-
tive yields.[15]
Gibson et al. have shown that m-xylene-induced
cells of Sphingomonas yanoikuyae strains B1 and B8/
36 were able to oxidize 1,2-dihydronaphthalene after
20–24 h to cis-(1R,2S)-diol 1 as the major products in
73% and 44% relative yields (>95% ee), respective-
ly.[16] Also, Gibson found that the toluene dioxygenase
Scheme 1.
of Pseudomonas putida F39/D oxidizes 1,2-dihydro-
naphthalene to cis-(1S,2R)-diol 1 and, in contrast, that
the naphthalene dioxygenase of Pseudomonas sp.
During the course of the reaction, which was moni-
strain NCIB 9816/11 oxidizes the same substrate to tored by GC/MS analysis with a chiral column, the
the opposite enantiomer, cis-(1R,2S)-diol 1.[17] The enantiomeric excess of the (S)-6 increased during the
cis-(1S,2R)-diol 1 was obtained in ca. 15% relative first 68 h (Table 1, entry 4). This increase was due to
yield while cis-(1R,2S)-diol 1 was obtained with ca. the consumption of (R)-6 of the (Æ)-2-hydroxy-1-tet-
70% relative yield. Both compounds presented ralone (6), which was diastereo- and enantioselective-
>98% ee.
ly reduced to produce cis-(1S,2R)-diol 1. The transfor-
We have recently reported the preparation of mation is diastereoselective since the cis-1 and trans-
(3R,4S)-3,4-chromanediol in high enantiomeric ex- diols 2 were potentially expected products, but the
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Deracemization of (Æ)-2-Hydroxy-1-tetralone by Trichosporon cutaneum
Table 1. Biotransformation of (Æ)-6 by resting cells of T. cutaneum at 308C.
FULL PAPERS
Entry
Time [h]
Yield [%][a]
cis-diol 1
ee% [Configuration][b]
6[d] cis-diol[e]
6
trans-diol 2[c]
1
2
3
4
5
6
7
8
9
0
5
20
68
116
143
169
193
288
100
89
70
55
42
24
14
5
-
-
racemate
26 (S)
69 (S)
94 (S)
45 (S)
9 (S)
46 (R)
73 (R)
96 (R)
-
10
29
42
57
75
85
94
93
1
1
3
1
1
1
1
1
92 (1S,2R)
83 (1S,2R)
81 (1S,2R)
88 (1S,2R)
90 (1S,2R)
96 (1S,2R)
98 (1S,2R)
>99 (1S,2R)
6
[a]
Chromatographic yield.
Absolute configuration.
trans-(1R,2R)-diol 2.
[b]
[c]
[d]
[e]
The ee of (Æ)-6 was determined by GC/MS analysis using a Hydrodex-b 3P fused capillary chiral column.
The ee of diol 1 was determined by HPLC using a chiral column Chiracel OJ-H, eluent hexane:2-propanol (90:10).
former was by far the major product. The reaction is
To explain the experimental data obtained for dera-
considered to be enantioselective since although two cemization of (Æ)-2-hydroxy-1-tetralone (6), we rea-
stereoisomers are expected for the major product soned that an isomerase is required to perform the
(cis-diol), only the cis-(1S,2R)-diol 1 was isolated as isomerization of unreactive (S)-6 during the biotrans-
the sole product.[20]
formation reaction.[21] Thus, only (R)-6 is reduced to
After the first 68 h of the reaction, (R)-6 was the cis-diol 1. In order to confirm these hypotheses,
almost entirely depleted and selectively converted more detailed studies were conducted to elucidate the
into the cis-diol 1. The remaining (S)-6 was slowly reaction mechanism proposed here. Experiments
converted into (R)-6 until the end of the reaction. It were carried out under the same experimental condi-
is worth mentioning that after 143 h of reaction, the tions described above, but this time racemic 6 was re-
GC analysis indicated the formation of a racemic mix- placed by (S)- and (R)-2-hydroxy-1-tetralone (6)
ture of 6 and thereafter the enantiomeric excess of (96% and 99% ee, respectively) as substrates. The en-
(S)-6 decreased (Table 1, entry 6).
antiomer (R)-6 was prepared by the oxidation of the
The formation of the products was followed by cis-(1S,2R)-diol 1 through a standard Jonesꢁ protocol.
chiral HPLC analysis, which gave a high enantiomeric The antipode (S)-6 was readily available through bio-
excess for the major cis-(1S,2R)-diol 1 during the catalysis using the yeast Pichia stipitis (see Experi-
course of the reaction (Table 1, entries 1–9). After 12 mental Section).
days of incubation, the experiment was stopped, the
For the experiments performed with (R)-6, the
yeast cells were removed by centrifugation and the analysis of the chromatograms obtained after 24 h of
diols were recovered from the aqueous phase by ex- reaction showed 76% conversion to the cis-(1S,2R)-
traction with ethyl acetate. After purification of the diol 1 and 18% conversion to the trans-(1R,2R)-diol
crude extract by flash column chromatography, cis- 2. Total consumption of the substrate (R)-6 was ach-
(1S,2R)-diol 1 was isolated in 83% yield and >99% ieved after 30 h of incubation and the diols (1S,2R)-1
ee (Scheme 1). Then, (R)-6 was recovered in 6% and (1R,2R)-2 were isolated in 60% and 10% yield,
yield with 96% ee.
respectively, both with>99% ee. It is worth mention-
A yield greater than 50% was achieved for diol 1, ing that no intermediates could be detected through-
and it is clear that T. cutaneum was able to promote out the reaction; neither the isomerization of (R)-6 to
deracemization of (Æ)-6 through a DKR process. (S)-6 nor the oxidation of (R)-6 to 3,4-dihydro-1,2-
DKR effectively combines the resolution step of ki- naphthalenedione was observed.
netic resolution with an in situ equilibration, or race-
On the other hand, the experiment performed with
mization, of the chirally labile substrate. As outlined the (S)-6 isomer demonstrated that its biotransforma-
previously,[3b,c] DKR needs two supplementary steps tion is a sluggish process (Table 2). Therefore, after
to take place: racemization with a consecutive asym- 24 h of incubation, only 23% of (S)-6 was converted
metric transformation. Consequently, the faster react- to the cis-(1S,2R)-diol 1, with 19% ee (Table 2,
ing (R)-6 is depleted during the course of the enantio- entry 5) and the enantiomeric excess of (S)-6 de-
selective reaction and the equilibrium of R/S-enantio- creased to 81%. In the course of the reaction, (S)-6
mers is constantly re-adjusted by racemization of the was isomerized to the (R)-6 isomer and then (R)-6
slow-reacting counterpart (S)-6.
was further reduced to cis-diol 1 until almost com-
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Table 2. Biotransformation of (S)-6 (96% ee) by resting cells of T. cutaneum at 308C.
Entry
Time [h]
Yield [%][c]
cis-diol 1
ee%[b] [Configuration][d]
6
trans-diol 2
6[a]
cis-diol 1
trans-diol 2
1
2
3
4
5
6
7
8
9
10
0
2
6
18
24
48
72
96
144
288
100
100
95
77
73
62
45
29
9
-
-
5
19
23
34
50
68
86
89
-
-
-
4
4
5
5
5
5
6
96 (S)
91 (S)
87 (S)
83 (S)
81 (S)
79 (S)
73 (S)
61 (S)
28 (S)
16 (S)
-
-
-
-
-
4 (1S,2R)
10 (1S,2R)
19 (1S,2R)
38 (1S,2R)
56 (1S,2R)
67 (1S,2R)
83 (1S,2R)
>99 (1S,2R)
17 (1R,2R)
22 (1R,2R)
26 (1R,2R)
30 (1R,2R)
44 (1R,2R)
52 (1R,2R)
60 (1R,2R)
5
[a]
The ee of (S)-6, determined by GC/MS analysis through a Hydrodex-b 3P fused capillary chiral column.
The ee of diol 1 and 2 was determined by HPLC using a chiral column Chiracel OJ-H, eluent hexane:2-propanol (90:10).
Chromatographic yield.
Absolute configuration.
[b]
[c]
[d]
plete consumption of the initial substrate. It is inter- absence of yeast cells were carried out under the
esting to note that, although the ee of cis-(1S,2R)-diol same experimental conditions as already described
1 initially formed was low (4% ee, Table 2, entry 3), for the deracemization of (Æ)-ketone 6. After 10 days
as the reaction progressed, it increased drastically.
of incubation, no isomerization of either substrate
It should be emphasized as already stated for the was observed and starting materials were recovered
(R)-6, that no trace of side-products was found for in high yield. Thus, these data strongly suggest that
the biotransformation of the (S)-6 within the detec- the isomerization step is not “spontaneous” but
tion limits. Since intermediates like diones are absent, should be attributed to specialized enzymes of T. cuta-
it is reasonable to believe that the isomerization of neum, such as isomerases, a class of enzymes able to
(S)-6 is performed by enzymes from T. cutaneum.
perform biological inversions of stereochemistry.[2a]
After 12 days of incubation, the reaction was stop-
To elucidate the unusual formation of cis-(1S,2R)-
ped and products were extracted and purified by flash diol 1 as the major product of the reduction of (S)-6
column chromatography on silica gel. The diols cis- and to understand the conversion of the (1R,2S)-diol
(1S,2R)-1 and trans-(1R,2R)-2 were isolated in 47% 1 to the (1S,2R)-diol 1 (since the enantiomeric excess
(>99% ee) and 3% (60% ee) yields, respectively. In of the latter increases to >99%), further experiments
addition, substrate (S)-6 was recovered in 20% yield using the cis-(1R,2S)-diol 1 and cis-(1S,2R)-diol 1 as
and 16% ee.
substrates (both >99% ee), and T. cutaneum as the
To confirm that the isomerization of ketone (S)-6 biocatalyst were carried out under the same standard
into the (R)-isomer was a complete enzymatic trans- experimental conditions as already described here.
formation and not a “spontaneous” process, experi- The results for the cis-(1R,2S)-diol 1 (see Experimen-
ments with (S)-6 (96% ee) and (R)-6 (99% ee) in the tal Section for its preparation) are shown in Table 3.
Table 3. Biotransformation of cis-(1R,2S)-diol 1 (>99% ee) by resting cells of T. cutaneum at 308C.
Entry
Time (days)
Yield [%][c]
cis-diol 1
ee%[b] [Configuration][d]
6
trans-diol 2
6[a]
cis-diol 1
trans-diol 2
1
2
3
4
5
6
7
8
4 h
1
2
3
4
7
10
14
3
93
76
64
56
38
77
66
57
1
1
1
<2
<2
1
<1
0
-
>99 (1R,2S)
96 (1R,2S)
86 (1R,2S)
72 (1R,2S)
38 (1R,2S)
64 (1S,2R)
91 (1S,2R)
99 (1S,2R)
>99 (1R,2R)
>99 (1R,2R)
>99 (1R,2R)
>99 (1R,2R)
>99 (1R,2R)
>99 (1R,2R)
>99 (1R,2R)
20
32
42
43
19
33
42
93 (S)
93 (S)
89 (S)
64 (S)
1 (R)
96 (R)
>99 (R)
-
[a]
The ee of (S)-6, determined by GC/MS analysis through a Hydrodex-b 3P fused capillary chiral column.
The ee of diol 1 and 2 was determined by HPLC using a chiral column Chiracel OJ-H, eluent hexane:2-propanol (90:10).
Chromatographic yield.
Absolute configuration.
[b]
[c]
[d]
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Deracemization of (Æ)-2-Hydroxy-1-tetralone by Trichosporon cutaneum
FULL PAPERS
With respect to the formation of (S)-ketone 6, it is Conclusions
evident that the cis-(1R,2S)-diol 1 is enzymatically
oxidized to (S)-ketone 6, which is slowly isomerized A biocatalytic process to deracemize (Æ)-2-hydroxy-
to (R)-ketone 6. Thereafter, the formation of the cis- 1-tetralone (6) efficiently has been successfully de-
(1S,2R)-diol 1 is a consequence of a further and faster vised using the versatile, easily-cultivated, non-con-
reduction of (R)-ketone 6, rather than (S)-ketone 6, ventional yeast Trichosporon cutaneum CCT 1903 to
by the yeast cells. After 14 days of incubation, the prepare the (1S,2R)-diol 1 in enantiomerically pure
stereoinversion of cis-(1R,2S)-diol 1 resulted in cis- form (>99% ee) and in high yield (83%). In order to
(1S,2R)-diol 1 (40% yield) and (R)-6 (22% yield), understand the mechanism of the complete dynamic
both in high enantiomeric excess (Table 3, entry 8). kinetic resolution of the substrate, a rationale based
Furthermore, the experiment performed with the cis- on a comprehensive study with chiral substrates was
(1S,2R)-diol 1 revealed that (R)-ketone 6 was the sole proposed. In this mechanism, the intervention of iso-
product observed, even after 14 days of incubation merase and the oxidation-reduction equilibrium are
(6% yield, >99% ee).
key features to be highlighted. Moreover, the enzy-
The deracemization of ketone 6 by T. cutaneum matic DKR of (Æ)-6 is unprecedented in the litera-
was scaled up to a 0.5–1.0 g scale of the substrate ture and presents a promising environmentally friend-
without loss of the high isolated yield or of the high ly method to obtain enantiopure (1S,2R)-diol 1 in
enantiomeric excess of the product. Hence, after 12 larger scales.
days of incubation, the diol (1S,2R)-1 was isolated in
82% and 80% yield (0.5 and 1.0 g scales, respective-
ly) and the unreacted substrate was typically recov- Experimental Section
ered in 6% and 12% yield (0.5 g and 1.0 g scales, re-
spectively).
General Remarks
The experiments performed with the chiral sub-
strates show that the deracemization of the title com-
pound is not a simple process. The formation of cis-
(1S,2R)-diol 1 seems to require both the intervention
of an isomerase to convert (S)-ketone 6 into (R)-
ketone 6 and an equilibrium-controlled oxidation-re-
duction sequence to convert the cis-(1R,2S)-diol 1
back to the (S)-ketone 6 and keep the balance be-
tween diols and hydroxy ketones (Scheme 2). Con-
All reagents and solvents were obtained from commercial
sources. Ethyl acetate, hexane, methanol and chloroform
were distilled before use.1H and 13C NMR spectra were re-
corded in CDCl3 and DMSO using an INOVA-500 or
Varian Gemini 300P spectrometer. Optical rotations were
measured on a Perkin–Elmer Polarimeter 341. Melting
points were measured on a Microquimica MQ APF-301.
HPLC analyses were carried out using a Shimadzu LC-
20 AT prominence instrument and a chiral column Chiralcel
OJ-H from Daicel. GC/MS analysis and mass spectra were
obtained on a QP 5000-Shimadzu or an Agilent CG 6890/
Hewlett Packard 5973 equipped with HP-5MS (5% phenyl-
methylpolysiloxane, 30 m”250 mm”0.25 mm) and Macherey
212117/91 Hydrodex-b 3P fused capillary columns with
either helium or hydrogen as carrier gas (1.0 mLminÀ1). The
strains of T. cutaneum (CCT 1903) and Pichia stipitis (CCT
2617) were purchased from the “AndrØ Tosello” Research
and Technology Tropical Foundation (Brazil). Bactopeptone
was purchased from Biobrµs (Brazil). Thin layer chromatog-
raphy (TLC) analyses were performed with pre-coated alu-
minum sheets (silica gel 60 Merck) and flash column chro-
matography was carried out on silica (200–400 mesh,
Merck).
Synthesis of (Æ)-2-Hydroxy-1-tetralone (6)
Scheme 2.
The substrate (Æ)-6 was prepared through hypervalent
iodine oxidation according to Moriartyꢁs protocol.[19] Then,
the commercially available a-tetralone (1.0 g; 6.85 mmol)
was converted to 2-hydroxy-1-tetralone dimethyl ketal, iso-
lated as a dark red oil; GC/MS: m/z (%)=210 [M+] (9), 178
(24), 166 (100), 150 (36) 134 (48), 120 (82), 119 (88), 107
(26), 91 (89), 77 (28), 63 (15), 51 (10).
cerning the former transformations (from the point of
view from the substrates) a typical DKR could be
considered. A closer look at the latter transformations
(from the point of view from the products) shows that
the product diol cis-(1R,2S)-diol 1 is being ultimately
converted to the cis-(1S,2R)-diol 1 through a stereoin-
version-like process.
The ketal was then submitted to hydrolysis with 3 molLÀ1
aqueous HCl in ethanol for 1 hour. After neutralization
with NaHCO3 and extraction with ethyl acetate, purification
of (Æ)-2-hydroxy-1-tetralone (6) was achieved by Kugelrohr
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929

InÞs Lunardi et al.
FULL PAPERS
distillation utilizing a preheated oven (1308C) at 15 mmHg.
Compound (Æ)-6 was obtained as a colorless oil; yield:
0.98 g (6.05 mmol; 89%); IR (KBr): n=3474, 1686, 1603,
1434, 1282, 1090, 994, 931, 757 cmÀ1; GC/MS: m/z (%)=162
[M+] (42), 144 (20), 133 (12), 118 (100), 116 (36), 105 (8), 90
(65), 77 (11), 63 (9), 51 (8); 1H NMR (300 MHz, CDCl3):
d=2.17–2.32 (m, 1H), 2.69–2.78 (m, 1H), 3.18–3.42 (m,
2H), 4.14 (s, 1H, OH), 4.59 (dd, J=13.5 and 5.7 Hz, 1H),
7.47 (d, J=7.8 Hz, 1H), 7.54 (t, J=7.5 Hz, H-1), 7.72 (td,
J=7.5 and 1.5 Hz, 1H), 8.23 (d, J=8.1 Hz, 1H); 13C NMR
(75.5 MHz, CDCl3): d=27.7, 31.8, 73.8, 126.8, 127.5, 128.8,
130.4, 134.1, 144.3, 199.6.
(53%),120 (95%), 119 (100%), 91 (45%), 77 (11%), 65
(11%), 51 (6%).
The biotransformation of (Æ)-6 was carried out for 12
days on 0.5 g (3.08 mmol) and 1.0 g (6.17 mmol) scales,
under the same experimental conditions as already de-
scribed above. The product was extracted into ethyl acetate,
dried and concentrated. After purification by flash column
chromatography on silica gel (hexane:ethyl acetate, 2:1 and
1:1), cis-(1S,2R)-diol 1 (>99% ee) was isolated in 82%
(415 mg, 2.53 mmol) and 80% (810 mg, 4.94 mmol) yields,
respectively. The trans-(1R,2R)-diol 2 (>99% ee) was also
obtained in 3% isolated yield for both experiments. The
substrate 6 was recovered in 6% and 12% yields (0.5 g and
1.0 g scale, respectively).
Biotransformation of (R)-2-hydroxy-1-tetralone (6): The
biotransformation of (R)-6 (45 mg, 0.28 mmol, 99% ee) was
carried out under the same experimental conditions as al-
ready described above. After 30 h the product was extracted
into ethyl acetate, dried and evapored. Purification by flash
column chromatography on silica gel (hexane:ethyl acetate,
1:1) gave cis-(1S,2R)-diol 1 ([a]20: +30.68 (c 2.0, CHCl3),
>99% ee) and trans-(1R,2R)-diDol 2 (>99% ee) in 60%
(27 mg, 0.17 mmol) and 10% (5 mg, 0.028 mmol) yields, re-
spectively.
Growth Conditions of Trichosporon cutaneum CCT
1903 and Pichia stipitis CCT 2617
Cells of T. cutaneum CCT 1903 were grown in sterile SDB
(Sabouraud dextrose broth, 1 L) for 3 days at 308C on an or-
bital shaker (170 rpm). The yeast Pichia stipitis CCT 2617
was cultivated in YM (yeast-malt extract) nutrient broth
(3 L) for 2 days at 288C on an orbital shaker (170 rpm). The
cells were harvested by centrifugation (5000 rpm, 10 min)
prior to use in the reactions.
Biotransformation of (S)-2-hydroxy-1-tetralone (6): (S)-6
(50 mg, 0.31 mmol, 96% ee) was biotransformed as describe
before. After 12 days of incubation, the products were ex-
tracted and purified by flash column chromatography on
silica gel. The cis-(1S,2R)-diol 1 {[a]2D0: +31.678 (c 2.25,
CHCl3), >99% ee} and trans-(1R,2R)-diol 2 (60% ee) were
isolated in 47% (24 mg, 0.145 mmol) and 3% (1.5 mg,
0.009 mmol) yields, respectively. The substrate (S)-6 was re-
covered in 20% yield and 16% ee.
Biotranformation of cis-(1S,2R)- and cis-(1R,2S)-1,2-dihy-
droxy-1,2,3,4-tetrahydronaphthalene (1): The cis-(1S,2R)-
and cis-(1R,2S)-diol 1 (50 mg each compound) were also
used as substrate in the same experimental conditions as de-
scribed above. The reaction mixture was incubated at 308C
in an orbital shaker (170 rpm) for 14 days. The cells were
then separated and the products extracted into ethyl acetate,
concentrated and purified by flash column chromatography
on silica gel. In the experiments, cis-(1S,2R)-diol 1 (99% ee)
was isolated in 76% and 40% yield, respectively. The oxida-
tion product (R)-6 (99% ee) was also obtained in 6% and
22% yield, respectively.
General Procedure for Biotransformation
Boitransformation of (Æ)-2-hydroxy-1-tetralone (6): A solu-
tion of (Æ)-2-hydroxy-1-tetralone (6) (250 mg) in ethanol
(2.5 mL) was added to a slurry of T. cutaneum CCT 1903
(15 g, wet weight) in glucose (5 g) and sterile distilled water
(250 mL). The resulting suspension was stirred on an orbital
shaker (170 rpm) at 308C until total consumption of 6 (ca.
12 days). After centrifugation (5000 rpm), the supernatant
and cell mass were thoroughly extracted with ethyl acetate.
The organic extracts were combined, dried over anhydrous
Na2SO4, and the solvent was removed under reduced pres-
sure. The residue was purified by flash column chromatogra-
phy on silica gel (hexane:ethyl acetate, 1:1) to give cis-
(1S,2R)-diol 1 (yield: 210 mg, 1.28 mmol, 83%, >99% ee)
and trans-(1R,2R)-diol 2 (yield: 7.6 mg, 0.046 mmol, 3%,
>99% ee). A small amount of substrate was recovered
(15 mg, 6%) and showed 96% ee for (R)-6. The absolute
configurations were assigned by comparison of the sign of
the specific rotation with that reported in the literature.[13]
Data for (1S,2R)-1: white crystals, mp 135–1368C (Lit.[13]
129–1308C); [a]D20: +33.68 (c 3.1, CHCl3) {Lit.[13] [a]2D0: +398
1
(c 3.1, CHCl3) for>98% ee}; H NMR (300 MHz, DMSO):
d=1.66–1.77 (m, 1H), 1.86–1.98 (m, 1H), 2.60–2.70 (m,
1H), 2.80–2.90 (m, 1H), 3.77–3.84 (m, 1H), 4.43–4.47 (m,
1H), 4.51 (d, J=4.8 Hz, 1H), 4.84 (d, J=5.7 Hz, 1H), 7.03–
7.08 (m, 1H), 7.12–7.15 (m, 2H), 7.32–7.35 (m, 1H);
13C NMR (75.5 MHz, DMSO): d=26.0, 26.3, 68.3, 69.2,
125.2, 126.6, 127.6, 129.1, 135.7, 138.3; IR (KBr): n=3259,
2935, 1459, 1071, 966; 775, 738 cmÀ1; GC/MS: m/z=164
(M+., 11%), 146 (54%), 120 (95%), 119 (100%), 91 (46%),
77 (11%), 65 (11%), 51 (6%).
Preparation of (R)-2-Hydroxy-1-tetralone (6)
To a stirred solution of (1S,2R)-diol 1 (2.0 g, 0.012 mol) in
acetone (40 mL) at 08C, Jonesꢁ reagent (10 mL) was added
dropwise. After complete addition of the oxidizing agent,
the mixture was stirred until the reaction reached comple-
tion. Methanol (20 mL) was added to quench excess of
Jonesꢁ reagent. The reaction mixture was treated with a sa-
turated aqueous solution of NaHCO3 and extracted with
ethyl acetate (4”30 mL). The organic phase was washed
with a saturated aqueous solution of NaCl (3”50 mL), dried
over Na2SO4 and concentrated at reduced pressure. The res-
idue was purified by flash column chromatography on silica
gel (hexane:ethyl acetate, 2:1) to give (R)-2-hydroxy-1-tetra-
lone (6) as a yellow oil; yield: 395 mg (2.44 mmol, 20%);
Data for (1R,2R)-2: white crystals, [a]2D0: +70.58 (c 0.31,
CHCl3) {Lit.[22] data for enantiomer (1S,2S)-2; [a]2D0: À1108
(CHCl3)}; 1H NMR (300 MHz, DMSO): d=1.61–1.74 (m,
1H), 1.90–2.10 (m, 1H), 2.64–2.86 (m, 2H), 3.62–3.72 (m,
1H), 4.27 (t, J=5.85, 1H), 4.82 (d, J=3.9 Hz, 1H), 5.25 (d,
J=5.4 Hz, 1H), 7.02–7.06 (m, 1H), 7.11–7.16 (m, 2H), 7.37–
7.41 (m, 1H); GC/MS: m/z=164 (M+, 11%), 146
930
www.asc.wiley-vch.de
ꢀ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2007, 349, 925 – 932

Deracemization of (Æ)-2-Hydroxy-1-tetralone by Trichosporon cutaneum
FULL PAPERS
[a]20: +44,58 (c 1.54, CHCl3) with 99% ee {Lit.[21a] [a]2D0: References
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Preparation of (S)-2-Hydroxy-1-tetralone (2) and cis-
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To a slurry of Pichia stipitis CCT 2617 (30 g, wet weight)
and glucose (5 g), in sterile distilled water (0.8 L), a solution
of racemic (Æ)-2-hydroxy-1-tetralone (6) (0.8 g) in ethanol
(5 mL) was added. The resulting suspension was stirred on
an orbital shaker (170 rpm) at 308C for 48 h. After centrifu-
gation (5000 rpm), the supernatant and cell mass were thor-
oughly extracted with ethyl acetate. The organic extracts
were combined, dried over anhydrous Na2SO4, and the sol-
vent was removed under reduced pressure. Purification was
achieved by flash column chromatography on silica gel (hex-
ane:ethyl acetate, 2:1 and 1:1) to furnished (S)-6 (yield:
0.54 g, 68%) and cis-(1R,2S)-diol 1 (yield: 0.15 g; 18%).
(S)-6: white crystals, mp 44.5–45.58C, [a]2D0: À43.68 (c 1.57,
CHCl3) with 96% ee.
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cis-(1R,2S)-diol 1: white crystals, mp 132–133.48C, (Lit.[23]
129–1308C); [a]2D0: À31.08 (c 0.89, CHCl3) with >99% ee
{Lit.[23] [a]2D0: À388 (c 0.87, CHCl3)}.
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Determination of Enantiomeric Excess of the
Substrate and Products Formed
The enantiomeric excesses of products 1 and 2 from the bio-
transformation reactions were analyzed on HPLC using a
chiral column Chiracel OJ-H from Daicel, Japan. The
mobile phase used was hexane:2-propanol (9:1), flow rate
0.7 mLminÀ1, monitored at 265 nm. For comparison, racemic
cis- and trans-diols standards were used. Retention times for
the two diols were: cis-diol 1 (14.5 and 21.4 min) and trans-
diol 2 (11.9 and 16.5 min). The ee of substrate (Æ)-6 was de-
termined by GC/MS analysis through a Hydrodex-b 3P
fused capillary chiral column (25 m”0.25 mm”0.25 mm)
using hydrogen as carrier gas. In this method the injector
and the detector were set a 2008C and 2308C, respectively,
initial column temperature was 508C (3.0 min) increasing
the temperature by 2.08C minÀ1 until 1808C (1.0 min); re-
tention times: (R)-6 at 32.2 min and (S)-6 at 32.4 min.
Acknowledgements
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This work was supported by grants from The State of S¼o
Paulo Research Foundation -FAPESP (05/00660–4) and
CNPq. FAPESP has also provided a grant to IL (03/05882–
0).
Adv. Synth. Catal. 2007, 349, 925 – 932
ꢀ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.asc.wiley-vch.de
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932
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ꢀ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2007, 349, 925 – 932