Use of Saccharomyces cerevisiae yeasts in the chemoselective bioreduction of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one in biphasic system

Article abstract of DOI:10.5935/0103-5053.20130140

This work describes the chemoselective bioreduction of (1E,4E)-1,5-bis(4- methoxyphenyl)- 1,4-pentadien-3-one (1) mediated by baker's yeast (BY, Saccharomyces cerevisiae cells) in an aqueous/organic solvent biphasic system. The biotransformation of this c

Full text of DOI:10.5935/0103-5053.20130140

http://dx.doi.org/10.5935/0103-5053.20130140
J. Braz. Chem. Soc., Vol. 24, No. 7, 1116-1122, 2013.
Printed in Brazil - ©2013 Sociedade Brasileira de Química
0103 - 5053 $6.00+0.00
Article
A
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction of
(1E,4E)-1,5-Bis(4-Methoxyphenyl)-1,4-Pentadien-3-one in Biphasic System
César A. Schaefer,a Vanessa D. Silva,a Boris U. Stambukb and Maria da G. Nascimento*,a
aDepartamento de Química and bDepartamento de Bioquímica,
Universidade Federal de Santa Catarina, 88040-900 Florianópolis-SC, Brazil
Este trabalho descreve a biorredução quimiosseletiva da (1E,4E)-1,5-bis(4-metoxifenil)-
1,4-pentadien-3-ona (1) mediada por fermento de pão (BY, células de Saccharomyces cerevisiae) em
sistema bifásico contendo água e n-hexano.A biotransformação deste composto foi quimiosseletiva,
formando apenas a correspondente cetona saturada 1,5-bis(4-metoxifenil)-3-pentanona (2). Foi
verificada a influência de vários fatores reacionais que podem afetar a biorredução de 1 tais como,
tipo e porcentagem volumétrica do co-solvente, uso de seis diferentes leveduras S. cerevisiae (quatro
comerciais e duas industriais), variação da concentração de substrato e levedura, temperatura, pH
e volume das fases aquosa e orgânica. A melhor condição reacional foi obtida com 66,7 g L−1 de
BY da marca Fleischmann, 8,3 × 10−3 mol L−1 de substrato, pH de 6,5 a 35 oC na presença de 2,5%
(v/v) de N,N-dimetilsulfoxido (DMSO) como aditivo e razão V /V de 70/30. Nestas condições
aq
org
o produto 2 foi obtido com conversão de 82% em 5 h de reação.
This work describes the chemoselective bioreduction of (1E,4E)-1,5-bis(4-methoxyphenyl)-
1,4-pentadien-3-one (1) mediated by baker’s yeast (BY, Saccharomyces cerevisiae cells) in
an aqueous/organic solvent biphasic system. The biotransformation of this compound was
chemoselective and formed only the corresponding saturated ketone 1,5-bis(4-methoxyphenyl)-
3-pentanone (2). The influence of various factors which may alter the bioreduction of 1, such
as the type and percentage of co-solvents, use of six different S. cerevisiae yeast samples (four
commercial and two industrial), variations in the substrate and yeast concentrations, temperature,
pH and volume of aqueous and organic phases, was investigated. The best reaction conditions were
66.7 g L−1 of Fleischmann BY, 8.3 × 10−3 mol L−1 of substrate, pH 6.5 at 35 oC in the presence of
2.5% (v/v) of N,N-dimethyl sulfoxide (DMSO) as an additive and a V /Vorg ratio of 70/30. Under
aq
these conditions, the product 2 was recovered in conversions of 82% in 5 h reaction.
Keywords: Saccharomyces cerevisiae, α,β-unsaturated carbonyl compounds, bioreduction,
biphasic system
Introduction
of enones by biocatalysts have revealed that several
enone reductases can be used for the production of chiral
ketones.7,8
The use of whole cells or isolated enzymes in organic
chemistry to mediate the reduction of ketones, β-ketoesters,
imines and α,β-unsaturated systems with C=C activated
by strongly polarizing groups such as nitro, carbonyl or
hydroxyl groups is the subject of extensive study.1-6 It is
well known that biotransformation of an α,β-unsaturated
carbonyl compound in many cases gives a mixture of
saturated ketone or aldehyde, saturated alcohol or allylic
alcohol, indicating that several enzymes may catalyze the
reduction of C=C and C=O double bond competitively.
However many studies on the enantioselective reduction
Isolated enzymes frequently lead to higher enantiomeric
excesses (e.e.) since they avoid problems associated
with competing catalysts of different stereoselectivity.
Unfortunately, reduced cofactors must be regenerated
in situ in a second catalytic cycle or provided in
stoichiometric amounts to sustain the catalytic activity.
Thus, the use of whole living cells is attractive since they do
not require the addition of exogenous cofactors because of
their own cofactor regeneration systems.9-11As an example,
Rhodotorula sp AS2.2241 has been used as a biocatalyst
for the asymmetric reduction of 4’-methoxyacetophenone
to enantiopure (S)-1-(4-methoxyphenyl)ethanol, and
*e-mail: maria.nascimento@ufsc.br

Vol. 24, No. 7, 2013
Schaefer et al.
1117
several water-immiscible ionic liquids (ILs) have been
employed as green solvents.9 Also, biphasic whole-
cell biotransformations have been carried out for the
asymmetric reduction of 4-chloroacetophenone to
(R)-1-(4-chlorophenyl)ethanol and of 2-octanone to
(R)-2-octanol. The evaluation of different hydrophobic
ionic liquids has also been carried out.10
Different strategies to improve the conversion into product,
varying the substrate and yeast concentrations, temperature,
pH, and volume of aqueous and organic phases, were also
considered.
Experimental
Among the various possible biocatalysts, baker’s
yeast (S. cerevisiae) is perhaps the most well-known
whole-cell biocatalyst in the scientific and industrial
world and is certainly the most commonly used yeast by
organic chemists to mediate enantioselective reductions
due to its high chemo-, regio- and enantioselectivity, high
bioavailability, and ease of treatment and because it acts
under mild reaction conditions. Microbial reactions are
usually carried out in aqueous media where the cells are
most active. Recently, the kinetics of the biotransformation
of geraniol into citronellol by resting cells of S. cerevisiae
(baker’s yeast type II) was investigated in a continuous-
closed-gas-loop bioreactor (CCGLB).12 Unfortunately,
many organic compounds of commercial interest which can
potentially be transformed by enzymes or microorganisms
are poorly soluble in this medium and display toxic effects
on the biocatalyst. Thus, aqueous/organic solvent biphasic
systems have been developed to overcome these limitations.
These biphasic systems consist of an aqueous phase and a
water immiscible organic phase where the microbial cells
stay in the aqueous phase while the substrate and product(s)
remain mainly in the organic phase.13-14 The biphasic system
offers numerous advantages, such as increased solubility
of hydrophobic substrates, smaller reaction volumes,
increased volumetric productivity and minimization of
substrate/product inhibition of the biocatalyst. Enzyme
inhibition/inactivation by the solvents is minimized due
to their low concentration in the aqueous phase, where
the reaction takes place, and the recovery of the product
from the reaction medium is facilitated.11,15 Modifying the
reaction medium through the addition of water-miscible
organic co-solvents is a method commonly used to improve
the activity, selectivity and the stability of enzymatic
reactions.16-18 These co-solvents have the potential to
increase the activity by increasing the dissolution of poorly
soluble substrates; however, in some cases they may
inhibit the enzyme by changing its conformation through
denaturation.13,19
Yeasts
Six S. cerevisiae yeast strains were used in their active
dry yeast form. Four samples were commercial baker’s yeast
(BY), named Fleischmann (AB Brasil Industria e Comércio
deAlimentos LTDA, Brazil), Dona Benta (Pak Gida Uretim
ve Pazarlam A. S., Turkey), Nordeste (Moinho do Nordeste
S.A., Brazil) and Brugguemann (Algist Bruggeman NV,
Belgium), and two were industrial yeast strains (CAT-1
and PE-2) which are currently used for ethanol production
in Brazil.20,21 These two industrial fuel ethanol yeasts have
been commercially available since the late 1990s, and are
distributed by LNF Latino Americana Ltda. from Brazil.22
The pellets of dehydrated yeast cells were used directly in
the reduction reactions as described below.
Materials
The following chemicals were used as received:
potassium phosphate buffer purchased from Vetec; citric
acid from Acros; sodium hydroxide from Grupo Química;
acetone (99.5%) from Merck; 4-methoxybenzaldehyde
(98%) and silica gel (70-230 mesh) from Sigma Aldrich;
sodium borohydride (98%) from Merck; and deuterium
chloroform (99.8%) from Cambridge Isotope Laboratories.
All organic solvents were obtained from commercial
sources and were of analytical grade.
General procedure for the bioreduction reactions employing
yeasts
In a typical experiment, a mixture of 30 mL of n-hexane
containing 290 mg of 1, 4 g of BY and 30 mL potassium
phosphate/citric acid buffer was incubated at 35 ºC in
250 mL Erlenmeyer flasks with magnetic stirring.Aliquots
were periodically withdrawn from the aqueous and organic
phases and extracted with dichloromethane (2 × 10 mL).
The organic phase was combined, dried with MgSO4 and
the solvent was removed under vacuum. The residue was
analyzed by GC to evaluate the percentage of conversion to
product. Substrate and product peak areas were compared,
and the sum of the two was considered as 100%. After the
bioreduction of 1 with BY, the product 2 was isolated and
characterized by 1H NMR and GC.
This paper reports a study on the baker’s yeast-catalyzed
bioreduction of an α,β-unsaturated carbonyl compound
(1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one.
Firstly, the addition of various co-solvents to the buffer/n-
hexane biphasic systems was investigated to find the most
appropriate organic solvent to solubilize the substrate.

1118
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction
J. Braz. Chem. Soc.
Details regarding the co-solvents, co-solvent percentage
and this finding was confirmed by comparison of the
retention times (Rt) obtained from the analysis of 1, 2
and 3 by gas chromatography (GC), which were 12.5, 4.1
and 5.1 min, respectively. The structure of compound 2,
obtained in the bioreduction reaction, was confirmed by
(%), substrate and yeast concentrations, buffer pH,
temperature, and volumetric percentage of organic phase
are specified for each assay in the results and discussion
section.
1
IR and H NMR spectral analysis, as described in the
Analytical methods
experimental section.
After the spectroscopic identification of the product
obtained in the bioreduction of 1 with S. cerevisiae yeast
in the buffer/n-hexane biphasic system, the influence of
various factors which may influence this reaction was
investigated, as shown in the following sections.
Proton nuclear magnetic resonance (1H NMR) 400 MHz
was carried out on a Varian 400 Mercury Plus spectrometer
using CDCl3 as the solvent and tetramethyl silane (TMS)
as the internal standard. Infrared spectra (IR) were acquired
with an ABB Boomer FTLA 2000-100 spectrometer using
KBr pellets (range 4000-400 cm−1) at the Center ofAnalysis,
Department of Chemistry, UFSC (Florianópolis-SC,
Brazil). Gas chromatography (GC) analysis was carried
out with a Agilent 78202A GC-instrument equipped
with an FID detector, using hydrogen as the carrier gas
(19,84 psi), equipped with a Shimadzu CBP5 column
(25m × 0.22 mm × 0.25 mm). The injector and detector
temperatures were 280 and 290 ºC, respectively, and the
column temperature was 250 ºC. The retention times observed
for 1, 2 and 3 were 12.5, 4.1 and 5.1 min, respectively. In
the thin-layer chromatography (TLC) n-hexane:ethyl acetate
(8:2 v/v) was used as the eluent.
Effect of addition of co-solvents and DMSO percentage
Since the substrate is not totally soluble in the media
used to study the bioreduction of 1 catalyzed by BY in
buffer/n-hexane biphasic systems, the influence of the
addition of several co-solvents in the reaction medium,
such as dichloromethane, ethanol, ethyl acetate, DMSO
and N,N-dimethylformamide (DMF), was evaluated. The
conversions to 2, as a function of the co-solvents added and
the reaction time, are presented in Table 1.
Table 1. Effect of co-solvent on the bioreduction of 1 catalyzed by baker’s
yeast in buffer/n-hexane biphasic systema
Results and Discussion
Conversionb to 2 / %
Entry
Co-solvent (Log P)
5 h
ndc
10
10 h
ndc
14
24 h
ndc
14
39
3
In the first step of this study, (1E,4E)-1,5-bis(4-
methoxyphenyl)-1,4-pentadien-3-one (1) was synthesized
by the aldol condensation reaction between the
4-methoxybenzaldehyde and acetone under basic
conditions. This compound was used as a substrate in the
bioreduction reaction using S. cerevisiae cells in aqueous/
organic solvent biphasic systems formed with n-hexane
and potassium phosphate/citric acid buffer. The selection
of n-hexane as the organic solvent was based on previous
results obtained in the biohydrogenation of (2E)-1,3-
diaryl-2-propen-1-ones catalyzed by S. cerevisiae yeast.23
The biocatalytic reduction of 1 by BY was completely
chemoselective, producing only the corresponding saturated
ketone 1,5-bis(4-methoxyphenyl)-3-pentanone (2)
(Scheme 1). The alcohol (1E,4E)-1,5-bis(4-metoxyphenyl)-
1,4-pentadien-3-ol (3) was not detected in any experiments,
1
2
3
4
5
Dichloromethane (0.93)
Ethyl acetate (0.68)
Ethanol (−0.24)
29
29
DMF (−1.00)
ndc
42
ndc
90
DMSO (−1.30)
90
aReaction conditions: dry Fleischmann BY (4 g), substrate (290 mg),
0.2 mol L-1 potassium phosphate/0.1 mol L-1 citric acid buffer (pH 5.5)
(30 mL), n-hexane (24.6 mL), co-solvent (5.4 mL, equivalent to 9 % v/v),
35 ºC. bThe conversion values were determined by GC. cnot detected.
As observed in Table 1, when ethyl acetate and ethanol
were used as co-solvents the conversions were between
10-39% and were influenced by the reaction time (Table 1,
entries 2 and 3). On using dichloromethane or DMF no
product was detected in 10 h of reaction, and with the use
Scheme 1. Biotransformation of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one catalyzed by S. cerevisiae yeasts in a biphasic system.

Vol. 24, No. 7, 2013
Schaefer et al.
1119
Table 2. Screening of yeast strains for the bioreduction of 1 in buffer/n-
hexane biphasic system with addition of DMSO
of DMF the conversion was only < 5% in 24 h of reaction
(Table 1, entries 1 and 4). Better results were achieved with
DMSO as the co-solvent, presenting conversions of 42, 90
and 90% in 5, 10 and 24 h of reaction, respectively (Table 1,
entry 5). However, no direct relationship with the organic
solvent polarity (expressed by Log P values) was observed.
These results indicate that DMSO was the most suitable
co-solvent for the biosynthesis of 2 in a buffer/n-hexane
biphasic system. Thus, the effect of varying the percentage
of this co-solvent from 2.5% to 15% (v/v) was examined
in 5 h of reaction. Higher percentages of DMSO were not
feasible due to an increase in the reaction viscosity and
experimental problems in the reaction work up, related to
the reactant and product isolation. With the use of DMSO
percentages of 2.5, 5, 9 and 15% the conversions were
38, 47, 42 and 38%, respectively, showing no significant
changes as different quantities of DMSO were added to
the reaction medium.
In several studies reported in the literature it was found
that the addition of DMSO to the reaction medium leads
to an increase in the conversion into products, as well as
in the selectivity of reaction.24-26 In addition, the solvent
toxicity must be considered and DMSO is well recognized
as being non-toxic toward certain cells and can be used as
a cryoprotectant, depending on its final concentration.27-29
For example, Wang et al.26 reported that the reduction of
acenaphthenequinone using baker’s yeast in a monophasic
system provides an improvement in the conversion to
the mono-alcohol 2-hydroxyacenaphthenones with the
addition of 10% (v/v) DMF and DMSO as co-solvents,
with conversions increasing from 72% to 97% and 98%,
respectively. Thus, considering the above results, 2.5%
(v/v) DMSO was used as the solvent in the subsequent
experiments.
Conversionb to 2 / %
Entry
S. cerevisiae strain
5 h
38
28
31
34
19
5
24 h
94
66
68
80
27
7
1
2
3
4
5
6
Fleischmann
Nordeste
Dona Benta
Brugguemann
CAT-1
PE-2
aReaction conditions: dry yeast (4 g), substrate (290 mg), 0.2 mol L-1
potassium phosphate/0.1 mol L-1 citric acid buffer (pH 5.5) (30 mL),
n-hexane (28.5 mL), 2.5% (v/v) DMSO (1.5 mL), 35 ºC. bThe conversion
values were determined by GC.
However, when the bioreduction was carried with 24 h of
reaction, the commercial BY from Fleischmann was the
most efficient biocatalyst and the product 2 was obtained
in a conversion of 94%. Low conversions to 2 (5 to 27%)
were obtained when the industrial yeast samples CAT-1
and PE-2 were employed in 5 and 24 h of reaction. These
were unexpected results considering that when these
yeasts were used in the biohydrogenation of chalcones
the conversions were similar to those obtained with BY
from Fleischmann.23 Based on these data, the commercial
BY from Fleischmann was selected as the biocatalyst for
the subsequent experiments.
Effect of BY concentration
An optimum yeast concentration is an important
experimental parameter in terms of shortening the reaction
time and also increasing the product yield, making
the process more efficient. Thus, the effect of yeast
concentrations from 0 to 100.0 g L−1 on the bioreduction
of 1 catalyzed by Fleischmann BY was evaluated in order
to determine the minimum yeast concentration required
to obtain the 1,5-bis(4-methoxyphenyl)-3-pentanone with
high conversions. The results are presented in Figure 1.
The conversions increased linearly with an increase in the
yeast concentration and that the highest conversion to 2
(61%) was obtained when the yeast concentration was
100.0 g L−1. Although higher conversions into product
could be obtained using more biomass, when the cell
concentration was higher than 66.7 g L−1 the mixture
became too thick to be effectively stirred well and the
reaction medium was not homogeneous. Thus, BY
concentrations higher than 100.0 g L−1 were not used, and
a yeast concentration of 66.7 g L−1 was employed in the
subsequent experiments.
Screening of yeasts
In the next study, six different S. cerevisiae strains,
four commercial (Fleischmann, Nordeste, Dona Benta
and Brugguemann) and two industrial (CAT-1 and PE-2)
were screened in order to identify which yeast strain was
the most efficient in the bioreduction of (1E,4E)-1,5-bis(4-
methoxyphenyl)-1,4-pentadien-3-one in a biphasic system
(Table 2).
As observed in Table 2, the conversion to product
was strongly influenced by the use of different BY
and reaction times. When the reaction was carried out
for 5 h, no significant differences in the conversions
were observed using the commercial yeasts named
Fleischmann, Nordeste, Dona Benta and Brugguemann,
the values obtained being 38, 28, 31 and 34%, respectively.

1120
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction
J. Braz. Chem. Soc.
Figure 1. Effect of the yeast concentration on the bioreduction of
1 catalyzed by baker’s yeast in water/n-hexane biphasic system with
addiction of DMSO. Reaction conditions: BY (0-6 g), 1 (290 mg), 30 mL
of citrate/phosphate/borate buffer (0.1 mol L-1, pH 5.5), 28.5 mL n-hexane,
1.5 mL DMSO (2.5% v/v), 35 ºC, 5 h, magnetic stirring.
Figure 2. Effect of the substrate concentration on the bioreduction of
1 catalyzed by baker’s yeast in water/n-hexane biphasic system with
addiction of DMSO. Reaction conditions: BY (4 g), 1 (140-720 mg),
30 mL of citrate/phosphate/borate buffer (0.1 mol L-1, pH 5.5), 28.5 mL
n-hexane, 1.5 mL DMSO (2.5% v/v), 35 ºC, 5 h, magnetic stirring.
Effect of substrate concentration
increased from 20 ºC to 40 ºC, but when the temperature
was further increased from 40 ºC to 45 ºC, a small decrease
in the conversion was observed (from 89% to 82%). These
results are interesting because they show that BY enzymes
responsible for the reduction of 1 maintained they catalytic
activity even at high temperatures. Although the highest
conversion was achieved at 40 ºC, the temperature of 35 ºC
(with a conversion of 65%) was chosen to evaluate the effect
of the buffer pH and the volume ratio of aqueous to organic
It is extensively reported in the literature that a high
substrate concentration may lead to a decrease in the
product formation owing to the possible inhibitory effects
on the cells.30,31 Thus, in order to verify this effect in the
bioreduction of 1, a substrate concentration range of
8.3 to 41.7 × 10−3 mol L−1 was assayed. The conversion
values obtained in 5 h of reaction are given in Figure 1. The
conversion to 2 decreases from 69 to 21% with an increase in
the substrate concentration from 8.3 to 41.7 × 10−3 mol L−1.
Similar results were obtained by Wang et al.9 in the
asymmetric reduction of 4’-methoxyacetophenone catalyzed
by immobilized Rhodotorula sp. AS2.2241 cells in a
phase (V /V ).At lower temperatures, the organic solvent
aq org
evaporation is minimized, important to keep the initial total
•
C4MIM PF6/buffer biphasic system. They observed a
decrease in the maximum substrate conversion when the
substrate concentration was above 40 × 10−3 mol L−1,
probably due to the inhibition of the reaction caused by the
product. In this study the optimal substrate concentration
was 8.3 × 10−3 mol L−1 and this value was chosen for the
subsequent experiments to investigate the effects of the
temperature, pH and volume ratio of aqueous to organic phase.
Effect of reaction temperature
The influence of temperature on bioreduction of 1
employing BY was also investigated. Earlier reports have
described that the enzyme activity and stability, as well
as the stereoselectivity, can be affected by the reaction
temperature.32,33 As observed in Figure 3, the conversions
into 2 after 5 h of reaction increased as the temperature
Figure 3. Effect of reaction temperature on the bioreduction of 1catalyzed
by baker’s yeast in water/n-hexane biphasic system with addition of
DMSO. Reaction conditions: BY (4 g), 1 (140 mg), 30 mL of citrate/
phosphate/borate buffer (0.1 mol L-1, pH 5.5), 28.5 mL n-hexane, 1.5 mL
DMSO (2.5% v/v), 20-45 ºC, 5 h, magnetic stirring.

Vol. 24, No. 7, 2013
Schaefer et al.
1121
reaction volume. Meitan et al.34 reported similar results in
relation to optimal reaction temperature, in the asymmetric
synthesis of (2S,5S)-2,5-hexanediol catalyzed by BY
number 6 in aqueous media with the addition of glucose,
where the highest yield into product was achieved at 40 ºC.
However, a decrease in the e.e. value from >95.7 to ca. 90%
was observed at this temperature.
2.5 % (v/v) DMSO was studied varying the organic phase
volumetric percentage from 0 to 100%, maintaining a
total reaction volume of 60 mL. The results are presented
in Figure 5.
Effect of buffer pH
The effect of reaction medium pH on the bioreduction
of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one
in a biphasic system mediated by BY was investigated for
an aqueous phase pH range from 3.5 to 8.5 at 35 ºC. The
conversion into 2 was found to be strongly dependent on
pH (Figure 4), and under the experimental conditions used,
the highest conversion (74%) to product was achieved
at the pH 6.5, while the decrease in the conversion was
more pronounced at more acidic. These results are in
agreement with data reported in the literature, where it is
well documented that all enzymes have an optimal pH at
which the reaction rate is maximized. Deviations in pH
from the optimal value can lead to decreased activity due
to changes in the ionization of amino acid residues at the
active site of the enzymes, while larger deviations in pH
lead to denaturation of the enzyme protein itself.35,36
Figure 5. Effect of V /Vorg on the bioreduction of 1 catalyzed by baker’s
aq
yeast in water/n-hexane biphasic system with addition of DMSO. Reaction
conditions: BY (4 g), 1 (140 mg), 0.0-58.5 mL of citrate/phosphate/
borate buffer (0.1 mol L-1, pH 6.5), 0.0-58.5 mL n-hexane, 1.5 mL DMSO
(2.5% v/v), 35 ºC, 5 h, magnetic stirring.
As can be observed, the volume ratio of aqueous to
organic phase influenced the conversion into product. The
conversion to 2 increased from 41 to 82% with an increase
in the organic phase up to 30%. A further increases in
the organic phase led to a decline in the conversion from
82 to 0%. These results show the importance of using a
biphasic system in reactions catalyzed by yeasts in order
to increase the substrate solubility and reduce the possible
toxic effects of organic solvents on the enzymes present
in this microorganism. The highest conversion (82%) was
achieved when a ratio ofV /V 70/30 (v/v) was used in 5 h
aq org
of reaction. A similar result was obtained by Bie et al.37 in
the bioconversion of methyltestosterone to methandienone
by Arthrobacter simplexAS.1.94 in a carbon tetrachloride-
phosphate buffer two-phase system with the addition of
menadione, where the highest biocatalytic rate was obtained
with a volumetric phase ratio of 70/30 V /V .
aq
org
Figure 4. Effect of buffer pH on the bioreduction of 1catalyzed by baker’s
yeast in water/n-hexane biphasic system with addition of DMSO. Reaction
conditions: BY (4 g), 1 (140 mg), 30 mL of citrate/phosphate/borate buffer
(0.1 mol L-1, pH 3.5-8.5), 28.5 mL n-hexane, 1.5 mL DMSO (2.5% v/v),
35 ºC, 5 h, magnetic stirring.
Conclusions
The 1,5-bis-(4-methoxyphenyl)-3-pentanone was
obtained in good conversion (82%) after the optimization
of the experimental conditions using baker’s yeast-mediated
bioreduction of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-
pentadien-3-one in a buffer/n-hexane biphasic system
with the addition of DMSO as a co-solvent. From the
Effect of volume ratio of aqueous to organic phase (Vaq/Vorg)
The effect ofV /V on the bioreduction of 1 employing
aq org
BY in a buffer/n-hexane biphasic system with addition of

1122
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction
J. Braz. Chem. Soc.
synthetic point of view, the use of BY in the bioreduction
of the C=C double bonds of an α,β-unsaturated carbonyl
compound showed considerable advantages since the
product was obtained with high chemoselectivity under
mild, eco-friendly reaction conditions.
12. Arifin, A. A.; Don, M. M.; Uzir, M. H.; Biochem. Eng. J. 2011,
56, 219.
13. León, R.; Fernandes, P.; Pinheiro, H. M.; Cabral, J. M. S.;
Enzyme Microb. Technol. 1998, 23, 483.
14. Khmelnitsky, Y. L.; Rich, J. O.; Curr. Opin. Chem. Biol. 1999,
3, 47.
Supplementary Information
15. Jurček, O.; Wimmerová, M.; Wimmer, Z.; Coord. Chem. Rev.
2008, 252, 767.
Supplementary data are available free of charge at
http://jbcs.sbq.org.br as PDF file.
16. Wen, S.; Tan, T.; Yu, M.; Process Biochem. 2008, 43, 1259.
17. Bencze, L. C.; Paizs, C.; Tosa, M. I.; Irimie, F. D.; Tetrahedron:
Asymm. 2010, 21, 356.
Acknowledgments
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This study was supported by the Universidade Federal
de Santa Catarina (UFSC–Brazil), the Coordenação de
Aperfeiçoamento de Pessoal de Nível Superior (CAPES),
the Conselho Nacional de Desenvolvimento Científico e
Tecnológico (CNPq), and Instituto Nacional de Ciência
e Tecnologia (INCT-Catalysis), which provided financial
support and scholarships (C.A.S.,V.D.S, B.U.S. and M.G.N).
The authors are also grateful to Fermentec Ltda (Brazil) for
the donation of the yeast strains CAT-1 and PE-2.
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Submitted: November 26, 2012
Published online: June 14, 2013

J. Braz. Chem. Soc., Vol. 24, No. 7, S1-S3, 2013.
Printed in Brazil - ©2013 Sociedade Brasileira de Química
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Supplementary Information
SI
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction of
(1E,4E)-1,5-Bis(4-Methoxyphenyl)-1,4-Pentadien-3-one in Biphasic System
César A. Schaefer,a Vanessa D. Silva,a Boris U. Stambukb and Maria da G. Nascimento*,a
aDepartamento de Química and bDepartamento de Bioquímica,
Universidade Federal de Santa Catarina, 88040-900 Florianópolis-SC, Brazil
Preparation of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-
pentadien-3-one (1)
procedure described in the literature.1 This compound was
obtained as a yellow solid in 60% yield after 4 h of reaction.
Retention time on GC Rt of 12.5 min; m.p. 116-118 °C
(115-117 °C)2; (KBr) IV νmax/cm−1 3434, 1653, 1630,
1600, 1511, 1256, 1175, 823; 1H NMR (CDCl3, 400 MHz)
d (ppm) 7.7 (d, 2H, J 16 Hz), 7.5 (d, 4H, J 8 Hz), 6.9 (m,
6H), 3.8 (s,6H, –OCH3).
The α,β-unsaturated carbonyl compound 1was prepared
by aldol condensation in a medium with 50% (m/v) of KOH
using two equivalents of 4-methoxybenzaldehyde with one
equivalent of acetone according to a commonly applied
Figure S1. IR spectrum (KBr) of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one.
*e-mail: maria.nascimento@ufsc.br

S2
Use of Saccharomyces cerevisiae Yeasts in the Chemoselective Bioreduction
J. Braz. Chem. Soc.
Figure S2. 1H NMR spectrum (CDCl3, 400 MHz) of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one.
Preparation of 1,5-bis(4-methoxyphenyl)-3-pentanone (2)
and characterized by 1H NMR and GC. 1H NMR (CDCl3,
400 MHz) d (ppm) 7.1 (m, 4H), 6.8 (m, 4H), 3.8 (s, 6H,
–OCH3), 2.8 (t, 4H, CH2) 2.6 (t, 4H, CH2). Retention time
on GC Rt of 4.1 min.
The saturated ketone 2 was prepared by baker’s
yeast-catalyzed bioreduction of 1 in an aqueous/organic
solvent biphasic system. This compound was isolated
Figure S3. 1H NMR spectrum (CDCl3, 400 MHz) of 1,5-bis(4-methoxyphenyl)-3-pentanone.

Vol. 24, No. 7, 2013
Schaefer et al.
S3
Preparation of (1E,4E)-1,5-bis(4-metoxyphenyl)-1,4-
pentadien-3-ol (3)
J 8 Hz), 6.8 (d, 4H, J 8 Hz), 6.5 (d, 2H, J 16Hz), 6.1 (dd,
2H), 4.9 (m, 1H), 3.7 (s, 6H, –OCH3).
Compound 3 was prepared from (1E,4E)-1,5-bis(4-
methoxyphenyl)-1,4-pentadien-3-one by the reduction with
NaBH4/wet SiO2 as described in the literature with some
modifications, and was used as a standard in the 1H NMR
and GC analysis.3 The alcohol 3 was obtained as a pale
yellow solid in 49% yield after 1 h of reaction. Retention
time on GC Rt of 5.1 min; m.p. 77-80 °C (85-87 °C)4; IV
References
1. Vogel,A. I.; Química Orgânica 2 Análise Orgânica Qualitativa,
3th ed., Ao Livro Técnico S. A.: Rio de Janeiro, Brazil, 1971.
2. Youssef, K. M.; El-Sherbeny, M. A.; El-Shafie, F. S.; Farag, H.
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Med. Chem. 2004, 337, 42.
ν
max/cm−1(KBr): 3538, 2957, 1605, 1513, 1250, 1174, 1030,
971, 817; 1H NMR (CDCl3, 400 MHz) d (ppm) 7.3 (d, 4H,
3. Zeynizadeh, B.; Behyar, T.; J. Braz. Chem. Soc. 2005, 16, 1200.
4. Hesse, G. ; Thieme, P.; Liebigs Ann. Chem. 1965, 686, 64.
Figure S4. IR spectrum (KBr) of (1E,4E)-1,5-bis(4-metoxyphenyl)-1,4-pentadien-3-ol.
Figure S5. 1H NMR spectrum (CDCl3, 400 MHz) of (1E,4E)-1,5-bis(4-metoxyphenyl)-1,4-pentadien-3-ol.