Novel unusual microbial dehalogenation during enantioselective reduction of ethyl 4,4,4-trifluoro acetoacetate with baker's yeast

Article abstract of DOI:10.1016/S0040-4039(00)02240-1

In the course of investigating microbial syntheses for chiral pharmaceutical intermediates, ethyl 4,4,4-trifluoro acetoacetate (1) was submitted to baker's yeast reduction. With the purpose of obtaining the D-carbinol in high enantiopurity, several additives were tested for L-reductase inhibitor activity. Allyl alcohol proved to be not only a suitable additive, but also an inducer for effective defluorination of the substrate.

Full text of DOI:10.1016/S0040-4039(00)02240-1

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 1267–1268
Novel unusual microbial dehalogenation during enantioselective
reduction of ethyl 4,4,4-trifluoro acetoacetate with baker’s yeast
Martin Bertau*
Institut fu¨r Biochemie, Technische Universita¨t Dresden, 01062 Dresden, Germany
Received 6 November 2000; accepted 30 November 2000
Abstract—In the course of investigating microbial syntheses for chiral pharmaceutical intermediates, ethyl 4,4,4-trifluoro
acetoacetate (1) was submitted to baker’s yeast reduction. With the purpose of obtaining the
D-carbinol in high enantiopurity,
L-reductase inhibitor activity. Allyl alcohol proved to be not only a suitable additive, but also an
several additives were tested for
inducer for effective defluorination of the substrate. © 2001 Elsevier Science Ltd. All rights reserved.
The baker’s yeast reduction of ethyl 4,4,4-trifluoro ace-
toacetate (1) is an established synthetic method for the
remarkable example for dehalogenation at C-4 was
observed.
production of
L
-carbinol 2 as a precursor of novel
fluorinated pharmaceuticals, such as the antidepressant
The presence of 1.0 g/l allyl alcohol afforded ent-2 in
28% ee with total conversion of 1, which is somewhat
lower than the enantiopurity reported by Davoli et al.7
Upon addition of 2.0 g/l allyl alcohol the ee increased
to 76%. However, ent-2 was present in the reaction
mixture in merely 33%.13
befloxatone.1–6 Baker’s yeast produces 2 in 62% ee,
while the respective
D
-carbinol ent-2 is accessible by
adding the
L-reductase inhibitor allyl alcohol to the
culture.7
In order to obtain the
purity, the -reductase inhibitor activities of unsatu-
D
-isomer ent-2 in high enantio-
L
The main product formed (62%) was ethyl D-3-hydroxy-
rated thioethers were compared with that of allyl alco-
hol. Additionally, a blank experiment without additive
was conducted (Table 1). None of these additives
butanoate (4). Remarkably, this defluorination occurs
at C-4. Several examples for dehalogenations of b-keto
esters are hitherto reported, but they refer to a-halo-
genated b-keto esters and hereof mostly to 2-iodo spe-
cies.8,9 The competing dehalogenation of 1 at C-4 takes
place solely on the stage of keto ester 1 and furnishes 3
to give 4. This is due to a three-center resonance
stabilization of an intermediate sp2-center at C-4 by the
carbonyl sp2-centers. Carbinol ent-2 is not dehalo-
showed stereoinduction towards the
the exception of allyl alcohol.
D-carbinol, with
In the course of investigating the influence of the
concentration of allyl alcohol on the enantioselectivity
of the baker’s yeast reduction, an outstandingly
Table 1.
L-Reductase inhibitor activities of additives tested for synthesis of ent-2
Entry
Additive
Conc. (g/l)
Isomer
ee
[h]2D0 (°)
Yield (%)
1
2
3
None
–
L
L
L
D
D
62
62
62
28
76
+20.1a
+20.1
+20.1
−5.9
36.5
44.3
48.0
39.5
35.3
Phenyl allyl sulfide
Phenyl vinyl sulfide
Allyl alcohol
Allyl alcohol
1.0
1.0
1.0
2.0
4
5b
−16.2
a Pure 2: +20.1° (Ref. 2).
b Substrate 1 was defluorinated to give 4.
* Corresponding author. Fax: +49 351 463-5506; e-mail: martin.bertau@chemie.tu-dresden.de
0040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(00)02240-1

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M. Bertau / Tetrahedron Letters 42 (2001) 1267–1268
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Scheme 1. Concurrence between enantioselective reduction
and dehalogenation of 1.
13. Experimental procedure:
Materials. Fala baker’s yeast was the product of Societe´
Industrielle de Levure FALA, Strasbourg, France. Ethyl
4,4,4-trifluoro acetoacetate (1), additives and organic sol-
vents were purchased from Fluka and Aldrich.
genated as for these reasons the b-halohydrin CꢀF
bond is not activated. Thus, the dehalo reduction must
occur before keto reduction (Scheme 1).
Baker’s yeast reduction of ethyl 4,4,4-trifluoro acetoac-
etate (1). The fermentations were carried out in a flask
containing 2 L of tap water, 20 mL of 1, 250 g Fala
baker’s yeast, and the respective amount of allyl alcohol.
After 20 h the products were recovered by extraction with
tert-butyl methylether, dried over sodium sulfate, and
then evaporated to dryness under reduced pressure.
Determination of the extent of conversion. The conversions
of the yeast reduction were measured using a J & W
Scientific DB-5 column (30 m, 0.25 mm i.d.) at 50°C
(isothermal). The pressure of N2 gas was 80 kPa; the
temperatures of the injector and the detector were 210
and 260°C, respectively. The keto-substrate 1 and prod-
ucts 2 and ent-2 were observed at retention times of 3.7
and 5.5 min, respectively. The % conversions were deter-
mined using an integrator.
Determination of enantiomeric excess (ee). After the con-
version and rates had been analyzed, the crude mixture
was distilled in vacuo at 5 mbar. The purified carbinolic
fractions were converted into the corresponding trifluoro
acetates by reacting with a 1.2 molar amount of trifluoro
acetic acid anhydride in dry CH2Cl2 at 65°C. After the
reaction was complete, the volatile components were
evaporated. GC analysis of the resulting trifluoro acetate
was conducted using a Macherey & Nagel Lipodex A
column (50 m, 0.25 mm i.d.) running a ramp from 60 to
120°C with 10°C/min. The trifluoro acetates of NaBH4
reduced 1 were used to find the suitable conditions for
chiral GC analysis. The pressure of N2 gas was 130 kPa;
the temperatures of the injector and the detector were 250
This defluorination is especially noteworthy, as reduc-
tive dehalogenations of b-keto esters have been
observed with activated CꢀHal bonds at C-2. More-
over, it is mostly iodo species that are dehalogenated
microbially.8 The enzymic dehalo reductions of 2-halo-
b-keto esters have been investigated by several groups
who assign these reactions to NADH dependent
monoelectronic reduction mechanisms, but the chem-
istry of these processes is not yet fully understood.10–12
To the author’s knowledge, this type of reductive
microbial dehalogenation of fluorinated b-keto esters
has not yet been reported. These results demonstrate
impressively, how the effect of additives on the out-
come of the microbial enantioselective reductions of
halo-b-keto ester 1 is the consequence of a complex
interplay of enzymatic activities and competing path-
ways. The elucidation and understanding of this novel
defluorination reaction offers a great potential for
dehalogenative microbial transformations and bioreme-
diative processes.
The reasons for this remarkable reaction are currently
under investigation.
References
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-carbinol ent-2 was
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L
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Determination of absolute configuration. The absolute
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