Fruits of the glossy privet (Ligustrum lucidum-Oleaceae) as biocatalysts for producing chiral aromatic alcohols

Article abstract of DOI:10.3109/10242422.2014.976634

The screening of four invasive plant species for use as biocatalysts in the stereoselective reduction of ketones is reported. Our studies revealed that fruits of Ligustrum lucidum can be used for the bioreduction of acetophenone to (S)-1-phenylethanol (94% conversion, >99 enantiomeric excess [ee]%). Using this methodology, 13 substituted (S)-phenylethanols were synthesized with good ee values (>99.9 to 78%) using a technique which is more environmentally friendly than classical reduction of prochiral ketones. The results reveal the fruits of L. lucidum to be promising biocatalysts for the production of key intermediates.

Full text of DOI:10.3109/10242422.2014.976634

Biocatalysis and Biotransformation, 2014; 32: 348–357
SHORT COMMUNICATION
Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as
biocatalysts for producing chiral aromatic alcohols
1
1
1
MARIO L. AIMAR , DANIELA L. BORDÓN , STELLA M. FORMICA , JUAN
2
3
4
4
J. CANTERO , ANA M. VAZQUEZ , MANUEL I. VELASCO & LAURA I. ROSSI
1
Cathedra of Applied Chemistry, Faculty of Exact, Physical and Natural Sciences, National University of Córdoba,
2
Vélez Sársfield 1611, University City, Córdoba, Argentina, Department of Agricultural Biology, Faculty of
Agronomy andVeterinary, National University of River Fourth (UNRC), Córdoba, Argentina
3
Cathedra of Chemical Technology, Faculty of Chemical Sciences, Catholic University of Córdoba, Córdoba,
4
Argentina and Research Institute of Physical Chemistry of Cordoba, (INFIQC-CONICET), Department of
Organic Chemistry, Faculty of Chemical Sciences, National University of Córdoba, Córdoba, Argentina
Abstract
The screening of four invasive plant species for use as biocatalysts in the stereoselective reduction of ketones is reported.
Our studies revealed that fruits of Ligustrum lucidum can be used for the bioreduction of acetophenone to (S)-1-phenyleth-
anol (94% conversion, ꢀ99 enantiomeric excess [ee]%). Using this methodology, 13 substituted (S)-phenylethanols were
synthesized with good ee values (ꢀ99.9 to 78%) using a technique which is more environmentally friendly than classical
reduction of prochiral ketones. The results reveal the fruits of L. lucidum to be promising biocatalysts for the production
of key intermediates.
Keywords: Ligustrum lucidum, biocatalysts, bioreduction, substituted acetophenones, chiral aromatic alcohols, green chemistry
Introduction
their low productivity, their commercial application
is still limited. Consequently, increasing attention has
been paid for developing robust biocatalysts that can
carry out reduction processes at a high substrate
loading in order to achieve economic feasibility and
competitiveness for large-scale biotransformation
The asymmetric reduction of prochiral ketones to
produce chiral secondary alcohols is an important
reaction in organic chemistry (Singh 1992; Noyori
1994).These alcohols are required as key intermedi-
ates in the production of pharmaceuticals and
other important chemicals (Yadav 2002 and 2008;
Ishihara et al. 2003; Caron et al. 2005).
(Ni & Xu 2012).
During the last decade, chemical reactions using
plants as biocatalysts have been the subject of several
investigations carried out on the biotransformation
of foreign substrates (Cordell et al. 2007). Many
transformations of different substrates, such as
hydroxylation and oxidation reactions (Gynostemma
pentaphyllum, Sakamaki et al. 2005), hydrolysis of
esters (Solanum tuberosum and Helianthus tuberosus,
Mironowicz 1998), bioreduction of ketones and
aldehydes (Daucus carota, Foeniculum vulgare, Cucur-
bita pepo, Phaseolus aureus, Cocos nucifera, Saccharum
officinarum, Manihot dulcis, and M. esculenta, Cordell
Despite these reductive processes being of great
importance to the chemical industry, comparatively
few reduction methodologies have been developed,
which take into account the concept of green chem-
istry (environmentally friendly reaction systems) in
order to avoid the formation of toxic waste that
may pollute the environment (Wohlgemuth 2010).
Compared with conventional chemical processing,
biocatalytic asymmetric reductions can offer more
selective reactions, environmentally benign processes,
and energy-efficient operations. However, due to
Correspondence: Mario L. Aimar, Ph.D., Cátedra de Química Aplicada, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba,
Vélez Sársfield 1611, Ciudad Universitaria, (X5016GCA) Córdoba, Argentina. Tel/Fax: ꢁ54 351 4344983 int 7. E-mail: mlaimar@efn.uncor.edu
(
Received 20 March 2014; revised 26 May 2014; accepted 10 October 2014)
ISSN 1024-2422 print/ISSN 1029-2446 online © 2014 Informa UK, Ltd.
DOI: 10.3109/10242422.2014.976634

Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as biocatalysts 349
et al. 2007; Yadav et al. 2002; Bruni et al. 2002;
Methods
Villa et al. 1998; Kumaraswamy & Ramesh 2003;
Fonseca et al. 2009; Baldassarre et al. 2000; Maczka
General
&
Mironowicz 2004; Machado et al. 2006; Assunção
Ketones and NaBH were purchased from Sigma-
4
et al. 2008; Machado et al. 2008; Blanchard & van
de Weghe 2006), enzymatic lactonization (Malus
sylvestris and H. tuberosus, Olejniczak et al. 2003),
glycosylation (Ipomoea batatas and Eucalyptus
perriniana, Shimoda et al. 2008), etc., have been
performed, and have produced very good results.
These processes, in general, generate less waste than
conventional chemical reagents (Kumaraswamy &
Ramesh 2003).
The use of different plant species in the biore-
duction of ketones represents an interesting route for
the synthesis of useful compounds (Giri et al. 2001;
Longo & Sanromaín 2006).The main advantages of
using plants as reagents are derived from their easy
disposal after use (as they are biodegradable with
mild reaction conditions) as well as their wide
availability at very low costs. Moreover, the use of
functionallyintactcells(“wholeplantcells”)obtained
directly from cut portions of plants has emerged,
because whole cells can ensure the recycling of the
oxidized cofactors (Blanchard & van deWeghe 2006).
Additionally, these reaction systems do not need
laborious cultivation or development operations to
be performed, which are commonly employed in the
management of microorganisms (Bohman et al.
Aldrich S.A. (Argentina). Sterile deionized water
was used to prepare phosphate buffer, and ethyl
acetate and hexane were purified by a simple distil-
lation prior to use.The crude reaction products were
extracted with ethyl acetate, the organic solutions
evaporated, and the products were filtered on a short
column with silica gel (70–230 mesh) using hexane–
ethyl acetate 90:10 as the eluent. Gas chromatogra-
phy (GC) analyses were made on a Buck Scientific
Model 910/310 instrument with a flame ionization
detector (FID) detector, and GC–mass spectrome-
try (MS) analyses were carried out on a Hewlett
Packard HP 5890 Series II gas chromatograph
equipped with a Mass Detector HP 5970. The
1
13
H nuclear magnetic resonance (NMR) and
C
NMR spectra were recorded on a Bruker Avance
II 400 MHz, and optical rotations were measured
in a JASCO P-1010 polarimeter. All products were
1
13
identified by comparing their MS, and H and
C
NMR spectra with literature data.
GC-FID and GC–MS analyses
To establish the chromatographic conditions, all
substrates used were reduced with sodium borohy-
dride in ethanol to obtain a racemic mixture of the
corresponding alcohols with good yields. The GC
separations were performed on a fused silica capil-
lary column Supelco β-Dex120 (phenyl polysiloxane
with 20% of permethylated β-cyclodextrin, 30 m,
0.25 mm, 0.25 μm) with GC general conditions of
split, 145 mL/min; injector, 220°C; detector FID,
2009).
On the other hand, invasive plants have been
considered as one of the most important modern
ecological problems, with proliferation affecting
conservation of biodiversity (Cohen 2002; Pauchard
et al. 2004). One of the most widespread invasive
trees in central Argentina is the glossy privet (Ligus-
trum lucidum,W.T. Aiton (Oleaceae)), which is native
to China and imported for use as an ornamental
plant. Glossy privet has now become a very success-
ful invader of forested sites, resulting in dense stands
that eliminate most native vegetation (Hoyos et al.
220°C; carrier gas, N ; and head pressure, 29 psi
2
with the conversion percentages of the reactions
being determined by GC using normalized peak
areas without a correction factor. The GC–MS (70
eV) analyses were performed using the same condi-
tions as those in the GC analysis, but using a Hewl-
ett Packard HP-5 (Crosslinked 5% PhMe Siloxane,
30 m, 0.32 mm, 0.25 μm film thickness) capillary
column.
2
010). A similar situation has been observed with
Pyracantha angustifolia (Franchet) C. K. Schneider
Rosaceae), Phyracantha atalantioides (Hance) Stapf
Rosaceae), and Cotoneaster franchetii Bois (Rosaceae).
(
(
Given these considerations, a project to study the
fruits of foreign plants that have become very inva-
sive in the ecosystems of our province was started in
order to find some practical use for them. As part of
our research into novel biocatalytic systems, the
fruits of glossy privet, yellow firehorn (P. angustifolia),
Gigg’s firethorn (P. atalantioides), and orange cotoneas-
ter (C. franchetii) were selected for studies on the
reduction of prochiral phenyl ketones to produce
chiral phenyl alcohols.
Biocatalysts
Healthy and mature fruits of L. lucidum, P. angustifolia,
P. atalantioides, and C. franchetii were collected
between April and August in Córdoba City (Argen-
tina), and identified by a botanist. To carry out this
study, the fruits were washed with tap water to
remove traces of dirt and then disinfected by immer-
sion in a 5% sodium hypochlorite solution for 15–20

350 M. L. Aimar et al.
min. These were then washed again, but with sterile
deionized water. The fruits were used immediately
after collection to assure the integrity of the enzy-
matic system.
silica gel using hexane/ethyl acetate 95/5 as eluent;
conversion (by GC analysis using the same condi-
tions described in GC-FID and GC–MS analyses):
75%; ee, ꢀ99%; yield, 241 mg (36%).
GC and spectroscopic data
Bioreductions
All products were identified by comparing their GC
These reactions were conducted immediately after
acquisition of the fruits. Using a coffee bean grinder
that had been previously disinfected, the fruits were
ground to form a slurry.This slurry (20 g) was added
to a sterile Erlenmeyer flask (250 mL) with phos-
phate buffer (150 mL, pH: 6.5), and then the ketone
1
13
retention times, MS, and H and C NMR spectra
with literature data (Salvi & Chattopadhyay 2001
and 2008; Yu et al. 2011; Cheemala et al. 2007;
Matsugi et al. 2004).
(
ꢃ)-(S)-1-Phenylethanol: GC conditions: T ꢄ80°C
1
(
1 min), ΔTꢄ 2.5°C/min, T ꢄ 140°C. GC Rt ace-
(
(
100 mg) was dissolved in dimethyl sulfoxide
DMSO) (1.25% v/v) and added to this suspension.
2
tophenone: 15.15 min, Rt (ꢁ)-(R)-1-phenylethanol:
0.88 min, and Rt (ꢃ)-(S)-1-phenylethanol: 21.58
2
The reaction was carried out by stirring on an orbital
shaker (120 rpm) for six days at 20°C, with the flask
being closed. Finally, the crude reaction was filtered
through cotton in order to remove particles in
suspension, and the solution was extracted with
2
3
min. [α] ꢄꢃ44.7, cꢄ0.51 MeOH (Yadav et al.
D
2
5
1
2
(
2
5
(
002, [α] ꢄ ꢃ39.1, cꢄ3.5 MeOH). H NMR
D
400 MHz, CDCl ) δ (ppm)ꢄ1.49 (d, 3H, CH ),
3
3
.03 (s, 1H, OH), 4.89 (q, 1H, CH), 7.25–7.39 (m,
1
3
H, ArH).
C NMR (100 MHz, CDCl ) δ
40 mL of ethyl acetate, with very gentle stirring to
3
ppm)ꢄ25.16, 70.44, 125.39, 127.50, 128.52,
avoid formation of an emulsion. Similarly, a control
experiment was conducted without the addition of
the corresponding phenylketone, and the crude reac-
tion was analyzed by chiral GC-FID and GC–MS
analyses.
1
%
45.81. Electron ionization (EI) MS m/z (rel. int.
)ꢄ123 (M ꢁ1, 4), 122(M , 29), 121 (M ꢃ1, 6),
ꢁ
ꢁ
ꢁ
1
5
07 (74), 105 (13), 79 (100), 78 (26), 77 (68),
7 (19), 51 (30).
(
ꢃ)-(S)-1-(2´-Chlorophenyl)ethanol: GC condi-
tions: T ꢄ 110°C, ΔTꢄ 3°C/min, T ꢄ 210°C. GC
1
2
Rt 2´-chloroacetophenone: 10.88 min, Rt (ꢁ)-(R)-
-(2´-chlorophenyl)ethanol: 17.04 min, and Rt
ꢃ)-(S)-1-(2´-chlorophenyl)ethanol: 18.30 min.
Kinetic study of the bioreduction of acetophenone
1
In order to establish the optimal reaction time, a
kinetic study was made using acetophenone as the
model substrate. The reaction progress was moni-
tored by taking samples (5 mL) every 24 h, which
were first extracted by shaking with ethyl acetate
(
[
23
α] Dꢄꢃ60.3, cꢄ0.5 MeOH (Nakamura
&
25
Matsuda 1998, [α] ꢄꢃ62.7, cꢄ0,894 CHCl ).
D
3
1
H NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.50 (d,
H, CH ), 1.92 (s,1H, OH), 5.30 (c, 1H, CH),
.18–7.24 (m, 1H, ArH), 7.28–7.34 (m, 2H, ArH),
.59–7.61 (m, 1H, ArH). C NMR (100 MHz,
3
3
7
7
3
(2 mL) and the organic layer was collected. Then,
anhydrous calcium chloride was added to remove
the dissolved water, and the organic solution was
filtered and analyzed (1 μL) by GC using the same
general conditions described in GC-FID and GC–MS
analyses. The reactions were made in triplicate.
13
CDCl ) δ (ppm)ꢄ23.52, 67.00, 126.41, 127.22,
3
1
%
28.42, 129.42, 131.67, 143.04. EI MS m/z (rel. int.
)ꢄ158 (M ꢁ2, 5), 156(M , 15), 143 (24), 141
ꢁ
ꢁ
(
82), 113 (26), 77 (100), 51 (28).
ꢃ)-(S)-1-(3´-Chlorophenyl)ethanol: GC condi-
(
tions:T ꢄ110°C(1min),ΔTꢄ3°C/min,T ꢄ210°C.
1
2
Scale reduction
GC Rt 3´-chloroacetophenone: 12.94 min, Rt (ꢁ)-
(R)-1-(3´-chlorophenyl) ethanol: 19.60 min, and Rt
(ꢃ)-(S)-1-(3´-chlorophenyl)ethanol: 20.14 min.
Ground ripe fruits of L. lucidum, 132 g and phos-
phate buffer, pH: 6.5 (1000 mL) were placed into a
conical flask. Acetophenone (0.670 g, 5.5 mmoles)
dissolved in 12.5 ml of DMSO was added to the
slurry of L. lucidum in the above buffer, covered and
allowed to shake for 6 days at 20°C.Then the slurry
of glossy privet was filtered off and washed with
deionized water (4ꢂ250 ml). The combined aque-
ous filtrate was extracted with ethyl acetate (500 ml).
The organic layer was dried and the crude product
was obtained (0.802 g). Pure (S)-1-phenylethanol
was recovered after column chromatography with
2
3
[α] ꢄꢃ41.1, cꢄ0.48 MeOH (Nakamura & Mat-
D
2
5
1
suda 1998, [α] ꢄꢃ43.5, cꢄ1.08, CHCl ). H
D
3
NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.48 (d, 3H,
3
CH ), 1.89 (d, 1H, OH), 4.85–4.90 (m, 1H, CH),
3
1
3
7.22–7.30 (m, 3 H), 7.36–7.39 (m, 1 H). C NMR
(100 MHz, CDCl ) δ (ppm)ꢄ25.25, 69.83, 123.54,
3
125.64, 127.55, 129.80, 134.39, 147.86. EI MS m/z
ꢁ
ꢁ
(rel. int. %)ꢄ158 (M ꢁ2, 5), 156 (M , 16), 143
(16), 141 (54), 121 (11), 115 (11), 113 (38), 77
(100), 75(17), 51 (23).

Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as biocatalysts 351
ꢁ
(
ꢃ)-(S)-1-(4´-Chlorophenyl)ethanol: GC condi-
142.25, 153.04. EI MS m/z (rel. int. %)ꢄ167 (M ,
ꢁ
tions: T ꢄ110°C, ΔTꢄ 3°C/min, T ꢄ210°C (20
3), 166 (M ꢃ1, 1), 152 (100), 122 (13), 107 (89),
106 (22), 105 (22), 94 (31), 78(35), 77 (74), 66
(13), 51 (33).
1
2
min). GC Rt 4´-chloroacetophenone: 13.26 min, Rt
(
ꢁ)-(R)-1-(4´-chlorophenyl)ethanol: 18.09 min, and
Rt (ꢃ)-(S)-1-(4´-chlorophenyl)ethanol: 18.65 min.
(ꢃ)-(S)-1-(4´-Fluorophenyl)ethanol: GC condi-
tions: T ꢄ 100°C, ΔTꢄ2°C/min, T ꢄ200°C.
23
[
α] ꢄꢃ 45.9, cꢄ0.38 MeOH (Nakamura &
D
1
2
25
1
Matsuda 1998, [α] ꢄꢃ49.0, cꢄ1.84, Ether). H
GC Rt 4´-fluoroacetophenone: 9.52 min, Rt (ꢁ)-
(R)-1-(4´-fluorophenyl)ethanol: 15.65 min and
Rt (ꢃ)-(S)-1-(4´-fluorophenyl)ethanol: 16.54 min.
D
NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.47 (d, 3H,
3
CH ), 1.83 (s, 1H, OH), 4.87 (q, 1H, CH), 7.26–
3
13
23
7
.31 (m, 4H, ArH). C NMR (100 MHz, CDCl3)
[α] ꢄꢃ35.2, cꢄ0.045 (Nakamura & Matsuda
D
2
5
1
δ (ppm)ꢄ25.29, 69.77, 126.80, 128,62, 133.09,
1998, [α] ꢄꢃ37.7, cꢄ0.931, MeOH). H NMR
D
ꢁ
1
1
7
44.25. EI MS m/z (rel. int. %)ꢄ158 (M ꢁ2, 4),
(400 MHz, CDCl ) δ (ppm)ꢄ1.47 (d, 3H, CH ),
3
3
ꢁ
56 (M , 16), 143 (20), 141 (70), 121 (9), 113 (28),
2.39 (b, 1H, OH), 4.87 (c, 1H, CH), 7.00–7.05
1
3
7 (100), 75 (19), 51 (24).
(m, 2H, ArH), 7.31–7.35 (m, 2H, ArH). C NMR
(
ꢁ)-(S)-1-(2´-Nitrophenyl)ethanol: GC condi-
(100 MHz, CDCl ) δ (ppm)ꢄ25.27, 69.79, 115.25
3
tions: T ꢄ100°C, ΔTꢄ 2°C/min, T ꢄ210°C (20
(d, Jꢄ21.2 Hz), 127.04 (d, Jꢄ8 Hz), 141.49 (d,
Jꢄ3.1 Hz), 163.34 (d, Jꢄ243.8 Hz). EI MS m/z
1
2
min). GC Rt 2´-nitroacetophenone: 31.15 min, Rt
ꢃ)-(R)-1-(2´-nitrophenyl)ethanol: 40.48 min, and
Rt (ꢃ)-(S)-1-(2´-nitrophenyl)ethanol: 42.23 min.
ꢁ
(
(rel. int. %)ꢄ140 (M , 27), 125 (100), 123
(13), 97 (87), 96 (24), 95 (26), 77 (29), 75 (20),
51 (12).
23
[
2
(
2
7
1
(
1
α] ꢄꢁ18.0, cꢄ0.022 MeOH (Comasseto et al.
D
006, [α]25 ꢄꢁ30.6, cꢄ3.06 CHCl ). H NMR
1
(ꢃ)-(S)-1-(4´-Bromophenyl)ethanol: GC condi-
tions: T ꢄ100°C, ΔTꢄ 2°C/min, T ꢄ200°C (20
D
3
400 MHz, CDCl ) δ (ppm)ꢄ1.56 (d, 3H, CH ),
3
3
1
2
.41 (b, 1H, OH), 5.40 (c, CH), 7.40 (dt, 1H, ArH),
.63 (dt, 1H, ArH), 7.82 (dd, 1H, ArH), 7.88 (dd,
min). GC Rt 4´-bromoacetophenone: 25.76 min, Rt
(ꢁ)-(R)-1-(4´-bromophenyl)ethanol: 33.44 min,
and Rt (ꢃ)-(S)-1-(4´-bromophenyl)ethanol: 34.18
1
3
H, ArH).
C NMR (100 MHz, CDCl ) δ
3
2
3
ppm)ꢄ24.20, 65.59, 124.30, 127.58, 128.11,
min. [α] ꢄꢃ35.1, cꢄ0.46 MeOH (Nakamura &
D
2
5
33.59,140.90,147.89.EI MS m/z (rel.int.%)ꢄ165
Matsuda 1998, [α] ꢄꢃ37.9, cꢄ1.13, CHCl ).
D 3
ꢁ
1
(
7
M ꢃ2, 1), 150 (100), 123 (12), 104 (7), 91 (11),
7 (14), 76 (35), 74 (14), 63 (11), 51 (50).
ꢃ)-(S)-1-(3´-Nitrophenyl)ethanol: GC condi-
tions: T ꢄ100°C, ΔTꢄ 2°C/min, T ꢄ210°C (20
H NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.47 (d,
3
3H, CH ), 1.92 (b, 1H, OH), 4.86 (c, 1H, CH),
3
(
7.23–7.26(m, 2H, ArH), 7.46–7.48 (m, 2H, ArH).
1
3
C NMR (100 MHz, CDCl ) δ (ppm)ꢄ25.27,
1
2
3
min). GC Rt 3´-nitroacetophenone: 34.18 min, Rt
ꢁ)-(R)-1-(3´-nitrophenyl)ethanol: 45.97 min, and
Rt (ꢃ)-(S)-1-(3´-nitrophenyl)ethanol: 46.35 min.
69.80, 121.18, 127.16, 131.57, 144.78. EI MS m/z
(rel. int. %)ꢄ202 (M ꢁ1, 27), 200 (25), 187 (81)
ꢁ
(
185 (86), 183 (9), 159 (25), 157 (31), 156 (9), 121
(23), 103 (12), 102 (8), 78 (55), 77 (100), 76 (20),
75 (21), 51 (42).
23
[
2
(
2
1
1
α] ꢄꢃ27.9, cꢄ0.44 MeOH (Comasseto et al.
D
006, [α]25 ꢄꢃ30.5, cꢄ2.99 CHCl ). H NMR
1
D
3
400 MHz, CDCl ) δ (ppm)ꢄ1.54 (d, 3H, CH ),
(ꢃ)-(S)-1-(4´-Trifluorophenyl)ethanol: GC condi-
tions: T ꢄ100°C, ΔTꢄ 2°C/min, T ꢄ200°C. GC
3
3
.17 (b, 1H, OH), 5.02 (c, 1H, CH), 7.50–7.54 (t,
H, ArH), 7.71–7.73 (d, 1H, ArH), 8.11–8.14 (dm,
H, ArH), 8.27 (t, 1H, ArH). 1 C NMR (100 MHz,
1
2
Rt 4´-trifluoroacetophenone: 8.87 min, Rt (ꢁ)-(R)-
1-(4´-trifluorophenyl)ethanol: 16.93 min, and Rt
(ꢃ)-(S)-1-(4´-trifluorophenyl)ethanol: 18.13 min.
3
CDCl ) δ (ppm)ꢄ25.49, 69.39, 120.45, 122.37,
29.44, 131.57, 147.91, 148.42. EI MS m/z (rel. int.
)ꢄ167 (M , 1), 166 (M ꢃ1, 1), 152 (100), 121
3
2
3
1
%
(
7
[α] ꢄꢃ26.3, cꢄ0.066 MeOH (Nakamura &
D
ꢁ
ꢁ
25
Matsuda 1998, [α] Dꢄꢃ28.1, cꢄ1.13, MeOH).
1
12), 107 (60), 106 (20), 105 (38),78 (38), 77 (82),
6(19), 51 (32).
ꢃ)-(S)-1-(4´-Nitrophenyl)ethanol: GC condi-
tions: T ꢄ100°C, ΔTꢄ 2°C/min, T ꢄ210°C (20
H NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.51 (d,
3
3H, CH ), 1.92 (b, 1H, OH), 4.97 (c, 1H, CH),
3
(
7.48–7.51(d, 2H, ArH), 7.60–7.62 (d, 2H, ArH).
1
3
C NMR (100 MHz, CDCl ) δ (ppm)ꢄ25.41,
1
2
3
min). GC Rt 4´-nitroacetophenone: 37.18 min, Rt
ꢁ)-(R)-1-(4´-nitrophenyl)ethanol: 48.66 min, and
Rt (ꢃ)-(S)-1-(4´-nitrophenyl)ethanol: 49.96 min.
69.83, 125.44, 125.47, 125.65, 149.71. EI MS m/z
(rel. int. %)ꢄ190 (M , 13), 175 (84), 171 (11), 145
ꢁ
(
(14), 127 (100), 95 (6), 77 (12), 69 (9), 51 (10).
(ꢃ)-(S)-1-(4´-Cyanophenyl)ethanol: GC condi-
tions: T ꢄ 100°C, ΔTꢄ 2°C/min, T ꢄ200°C.
23
[
[
α] ꢄꢃ30.0, cꢄ0.55 MeOH (Yadav et al. 2002,
D
25
1
α] ꢄꢃ30.5, cꢄ4 CHCl ). H NMR (400 MHz,
D
3
1
2
CDCl ) δ (ppm)ꢄ1.52 (d, 3H, CH ), 2.05 (b, 1H,
GC Rt 4´-cyanoacetophenone: 31.11 min, Rt (ꢁ)-
(R)-1-(4´-cyanophenyl)ethanol: 41.87 min, and Rt
(ꢃ)-(S)-1-(4´-cyanophenyl)ethanol: 43.04 min.
3
3
OH), 5.02 (c, 1H, CH), 7.53–7,56 (d, 2H, ArH),
.20–8.22 (d, 2H, ArH). 13C NMR (100 MHz,
CDCl ) δ (ppm)ꢄ25.53, 50.88, 123.78, 126.12,
8
2
3
[α] Dꢄꢃ40.0, cꢄ0.44 MeOH (Mathre et al. 1993,
3

352 M. L. Aimar et al.
R enantiomer [α]25Dꢄ ꢁ41.7, cꢄ1.063, MeOH,).
Results and discussion
1
H NMR (400 MHz, CDCl ) δ (ppm)ꢄ1.50 (d,
3
Reactions using acetophenone as a model substrate
3
H, CH ), 2.01 (b, 1H, OH), 4.97 (c, 1H, CH),
3
7
.48–7.50 (d, 2H, ArH), 7.64–7.66 (dd, 2H, ArH).
In our attempt to produce chiral aromatic alcohols
in high ee, acetophenone 1 was used as a test
substrate and it was found that ground fresh fruits
of L. lucidum could be used for enantioselective
bioreduction of 1 to give (ꢃ)-(S)-1-phenylethanol
13
C NMR (100 MHz, CDCl ) δ (ppm)ꢄ25.46,
3
69.70, 111.16, 118.86, 126.06, 132.37, 151.05.
ꢁ
EI MS m/z (rel. int. %)ꢄ147 (M , 7), 132 (64),
1
7
30 (10), 105 (12), 104 (100), 102 (22), 77 (21),
5 (13), 63 (6), 51 (12), 50 (10).
2
in 94% of conversion with an excellent enantiose-
lectivity (ee% ꢀ99, Scheme 1).
(ꢃ)-(S)-1-(4´-Methylphenyl)ethanol: GC condi-
To assure that the fruit did not contain the chiral
aromatic alcohols or their substrates, a control
experiment with fruits alone, without adding aceto-
phenone (or any other substrate) was carried out.
The crude of this reaction was analyzed, and neither
the presence of acetophenone nor the corresponding
product of its reduction was observed.
In order to establish the optimal reaction time, a
kinetic study using ground ripe fruits of L. lucidum
as the biocatalyst was made (Figure 1).
It can be seen in Figure 1 that the reaction
reached optimal performance after 6 days (94% of
conversion), while the enantioselectivity remained
very high throughout the study (ꢀ99% ee), reveal-
ing the high efficiency and stereoselectivity of the
enzymatic complex present in the fruits of L.lucidum.
While we have not conducted studies to establish the
group of enzymes that are responsible for carrying
out this transformation,it is probable that the enzyme
alcohol dehydrogenase—ketoreductase, dependent
on the nicotinamide cofactors as redox equivalents
(Xie et al. 2009; Blanchard & van deWeghe 2006)—
is involved in this transformation. However, further
studies are needed to support this hypothesis.
To demonstrate the viability of this bioreduction
protocol, a major scale reduction of acetophenone
tions: T ꢄ 100°C, ΔTꢄ 2°C/min, T ꢄ 200°C. GC
1
2
Rt 4´-methylacetophenone: 14.83 min, Rt (ꢁ)-(R)-
-(4´-methylphenyl)ethanol: 18.13 min, and Rt (ꢃ)-
1
2
3
(
S)-1-(4´-methylphenyl)ethanol:19.10 min.[α] Dꢄ
ꢃ 40.4, cꢄ0.049 MeOH (Nakamura & Matsuda
998, [α]25 ꢄ ꢃ43.5, cꢄ0.998, MeOH). H NMR
1
1
D
(
1
400 MHz, CDCl ) δ (ppm)ꢄ1.47 (d, 3H, CH ),
3
3
.89 (b, 1H, OH), 2.33 (s, 3H, CH ), 4.86 (c, 1H,
3
1
3
CH), 7.15 (d, 2H, ArH), 7.25 (d, 2H, ArH).
C
NMR (100 MHz, CDCl ) δ (ppm)ꢄ21.10, 25.09,
3
7
0.27, 125.37, 129.18, 137.17, 142.89. EI MS
ꢁ
m/z (rel. int. %)ꢄ136 (M , 39), 121 (98), 119 (9),
1
5
17 (5), 93 (100), 92 (21), 91 (77), 77 (45), 65 (27),
1 (15).
(ꢃ)-(S)-1-(Thiophen-2-yl)ethanol: GC condi-
tions: T ꢄ 100°C, ΔTꢄ 1°C/min, T ꢄ 200°C. GC
1
2
Rt 2-acetylthiophene: 13.86 min, Rt (ꢁ)-(R)-1-
(
1
thiophen-2-yl)ethanol: 19.33 min, and Rt (ꢃ)-(S)-
-(thiophen-2-yl)ethanol: 20.39 min. [α] Dꢄ
2
3
ꢃ22.3, cꢄ0.051 MeOH (Ohkuma et al. 2000,
24
1
[
α] ꢄ ꢃ26.0, cꢄ1.02, CHCl ). H NMR (400
D 3
MHz, CDCl ) δ (ppm)ꢄ1.61 (d, 3 H, CH ), 1.99
(
3
3
b, 1 H, OH), 5.14 (c, 1 H, CH), 6.95–6.99 (m, 2
13
H), 7.23–7.26 (dd, 1 H). C NMR (100 MHz,
CDCl ) δ (ppm)ꢄ25.27, 66.30, 123.20, 124.46,
3
126.67, 149.87. EI MS m/z (rel. int. %)ꢄ128 (Mꢁ,
29), 113 (55), 111 (15), 95 (16), 85 (100), 84 (19),
69 (7), 58 (11), 57 (9), 51 (7).
(
≈ 0.7 g batch) was conducted. Although, the extent
of chemical conversion of 1 decreased (94% to 75%),
the product 2 was obtained with an excellent ee%
(ꢃ)-(S)-1-(Pyridin-2-yl)ethanol: GC conditions:
(
ꢀ99%).
T ꢄ40°C (15 min), ΔTꢄ 1°C/min, T ꢄ160°C (20
1
2
A reaction using unripe fruits of L. lucidum was
min). GC Rt 2´-acetylpyridine: 37.19 min, Rt (ꢁ)-
also performed and the outcome was similar, but
with slightly lower values (84% conversion, 98% ee)
than those achieved using ripe fruits. Moreover,
other reactions using ripe fruits of L. lucidum were
performed using water as solvent. In this case the
(
(
R)-1-(pyridin-2-yl)ethanol: 43,52 min, and Rt (ꢃ)-
2
3
S)-1-(pyridin-2-yl)ethanol: 43.94 min. [α] Dꢄꢃ
3.9, cꢄ0.056 MeOH (Ohkuma et al. 2000,
5
25
[
α] Dꢄꢃ58.3, cꢄ0.51 EtOH). 1H NMR (400
MHz, CDCl ) δ (ppm)ꢄ1.51–1.52 (d, 3H, CH ),
3
3
3
7
7
.64 (b, OH), 4.76 (b, 1H, OH) 4.91 (c, 1H, CH),
.20–7.23 (m, 1H, ArH), 7.30–7.32 (m, 1H, ArH),
.69–7.73 (m, 1H, ArH), 8.53–8.54 (d, 1H, ArH).
O
H
OH
ground fruits of Glossy privet
buffer pH 6.5
CH3
CH3
13
C NMR (100 MHz, CDCl ) δ (ppm)ꢄ24.16,
3
2
0°C.
9
4%,
68,87, 119.96, 122.32, 137.08, 147.93, 163.04. EI
>99 e.e.%( S)
2
1
MS m/z (rel. int. %)ꢄ123 (Mꢁ, 3), 122 (8), 119
1), 108 (100), 106 (40), 80 (52), 79 (43), 78 (47),
3 (20), 52 (38), 51 (27).
(
5
Scheme 1.Conversion of acetophenone to (-)-(S)-1-phenylethanol
using fruits of L. lucidum as the biocatalyst.

Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as biocatalysts 353
Figure 1. Kinetic study of the bioreduction of acetophenone catalyzed by L. lucidum.
outcome was similar, but with slightly lower values
82% conversion, 95% ee) than those achieved using
a solution of phosphate buffer at pH: 6.5 as sol-
vent.
Also, other potential tree biocatalysts also were
screened under the same conditions for the above
reduction. The ground fresh fruits of P. angustifolia,
P. atalantioides, and C. franchetii were also tested and
they yielded 59%, 56%, and ꢀ99 ee%, respectively,
but with a conversion to alcohol 2 of 76%, 26%, and
5%, respectively, after 6 days of reaction.
(
One of the major disadvantages of using fruits of
plants as sources of enzymes capable of producing
chemical reactions is that plants only produce fruits
for limited and specific seasons of the year.To address
this situation, fruits of L. lucidum were frozen and
kept for 1–2 weeks at ꢃ20° C.Then, the fruits were
ground and a reaction was performed using aceto-
phenone as the substrate. However, in this case it
was observed that the reductive capacity of the fro-
zen fruit decreased significantly (9% conversion,
Bioreduction of substituted prochiral phenyl ketones
by L. lucidum
The results observed using L. lucidum as biocatalyst
in the acetophenone reductions encouraged us to
study the reduction of different substituted aceto-
phenones in the same system (Table I).This showed
(Table I) that fruits of glossy privet were efficient
bioreducers of a number of prochiral aromatic
ketones to their corresponding alcohols, with con-
versions ranging from 79 to 98%, except for 2´-
nitroacetophenone, 4´-methylacetophenone, and
2-acethylthiophene (conversions from 7 to 48%;
Table I, entries 5, 12, and 13). All other substrates
tested could be effectively reduced by the biocatalyst,
and most of their products had satisfactory optical
purities. It is interesting to note that the biocatalyst
reduced all substrates observing Prelog’s rule.
81% ee).
It should be noted that, although there are several
reports in the literature on the use of plant parts as
biocatalysts in the reduction of prochiral ketones,
few studies have been conducted using fruits. In the
results reported in this communication, conversion
(%) and ee(%) achieved in the reduction of aceto-
phenone are better than those described using M.
pumila (40.9%, 81.5 ee%(R), Yang et al. 2008), C.
nucifera (79%, 95 ee%(S), Fonseca et al. 2009), Pas-
siflora edulis (74%, 23 ee%(S), Machado et al. 2008),
and Lycopersicum esculentum (80%, 95 e.e%(S), Phu-
kan & Devi 2012). Moreover, it is interesting to note
that, unlike the above fruits, L. lucidum is a highly
invasive weed whose fruit has not been previously
reported for practical use. Thus, the results detailed
here have identified a potential practical use for the
fruits of this plant.
An influence of electron-donating substituents
on the reduction of the carbonyl group was evident,
producing the corresponding alcohol at moderate-
to-low conversions (Table I, entries 12 and 13).
Thus, 4´-methylacetophenone showed moderate
conversion (48%) and 2-acetylthiophene showed low

354 M. L. Aimar et al.
Table I. Ability of L. lucidum to reduce different substituted prochiral ketones.
Ground fruits of
Glossy privet
H
OH
O
HO
Ar
H
+
Buffer pH 6.5
.25% DMSO
20°C.
CH3
Ar
CH3
Ar
CH3
1
6 days
(
R)
(S)
Entry
Ar
Conversion %a
94
ee%b
Isolated yield %c
79
1
ꢀ99 (S)
2
90
93
ꢀ99.9 (S)
97 (S)
20
26
Cl
3
Cl
4
5
92
7
91 (S)
88 (S)
38
Cl
traces
NO
2
6
7
79
98
83 (S)
87 (S)
45
49
NO
2
2
O N
8
9
90
82
97 (S)
81 (S)
57
13
F
Br
1
1
1
0
1
2
84
97
48
78 (S)
95 (S)
88 (S)
5
30
14
F3C
NC
CH
3
1
1
3
4
S
17
82
79 (S)
81 (S)
7
N
52
a
Measured by GC analysis.
ee values were determined by chiral GC analysis. Absolute configurations were assigned by comparing
b
the retention time of GC data with the reports in the literature and by comparison of the sign of optical
rotations with the literature values.
c
Isolated by column chromatography.
conversion to the corresponding alcohol (17%). In
contrast, electron-withdrawing groups increased the
rate of conversion, with practically quantitative con-
version being observed with 4´-cyanoacetophenone
(97% conversion, Table I, entry 11) and 4´-
nitroacetophenone (98% of conversion, Table I,
entry 17) as substrates. A similar situation has been
observed using D. carota, Apium graveolens, and Lens

Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as biocatalysts 355
culinaris as the biocatalysts (Yadav et al. 2002; Liu
Conclusions
et al. 2010; Ferreira et al. 2012).
In conclusion, several chiral (S) alcohols with excel-
lentorverygoodenantiomericexcesses(ꢀ99.9-78%)
were synthesized, thus revealing the fruits of Ligus-
trum lucidum to be a promising biocatalyst for the
production of key intermediates. In addition, the
presence of substituents (electron-withdrawing or
electron-donating) was observed in the aromatic
ring, which could have steric and/or electronic influ-
ences on the course of reduction.
The results obtained here using fruits of glossy
privet as the biocatalyst may offer new strategies for
the reduction of selected prochiral phenyl ketones as
a critical step in the synthetic organic pathway,
thereby avoiding the use of costly and non-renewable
metal reducing agents and organic solvents that are
commonly utilized in organic synthesis. As a result
of this study with wild foreign plants, it is clear that
an unexpected opportunity has arisen to establish
new applications for the foreign flora, especially for
those species which do not have any other reported
practical utility and are considered to be highly inva-
sive weeds.The bioreduction method presented here
allows chiral phenyl alcohols to be obtained with a
very good enantioselectivity by using a methodology
which is more environmentally friendly than classical
reductions of prochiral ketones.
For 2´-, 3´-, and 4´-nitroacetophenones (Table I,
entries 5, 6, and 7), very good stereo- and chemose-
lective bioreduction was also observed, without
reduction of the nitro group.There are reports in the
literature which describe not only the production of
alcohols, but also the reduction of the nitro group to
amines from nitroacetophenones using plants as bio-
catalysts (Ferreira et al. 2012). However, in this
study, nitroacetophenones showed a very variable
percentage of conversion, and in the case of 2´-
nitroacetophenone there was possible steric influ-
ence on the reduction of the carbonyl group, giving
very low conversion rates (7%, Table I, entry 5). A
similar situation was observed by Ferreira et al.
(2012) in the reduction of 2´-nitroacetophenone
using L. culinaris as the biocatalyst. On the other
hand, Liu et al. (2010) using A. graveolens as bio-
catalyst showed that the presence of a substituent on
the aromatic ring, especially an o-substituent, caused
steric hindrance, reducing the rate of reaction. It is
noteworthy that (S)-1-(4´-nitrophenyl) ethanol,
which can be produced in this reaction, is the
pharmacophore found in important drugs such as
nifenalol and sotalol (Pedragosa-Moreau et al. 1997;
Blay et al. 2010).
The ee observed for all reactions ranged from
very good (4´-trifluoromethylacetophenone 78%;
Table I, entry 10) to excellent (2´-chloroacetophenone
Further investigations are currently ongoing, trying
to expand the specificity and exploring new catalytic
activities of this new inexpensive biocatalytic agent.
ꢀ
99.9%;Table I, entry 2). A reactivity similar to that
of acetophenone was observed with chloroacetophe-
nones, with the conversion to their corresponding
alcohols ranging from 90 to 93%, but with the
enantioselectivity decreasing in the sequence 2´-, 3´-,
and 4´-chloroacetophenone (ꢀ99.9, 97, and 91
ee%, respectively; Table I, entries 2, 3, and 4). It
is interesting to note the importance of these
results, since the three chlorophenyl ethanols
obtained are pharmacophores in drugs such as
chlorprenaline, solabegron, and in the other β3-
adrenergic receptor agonists currently in develop-
ment (Lu et al. 2011).
The moderate-to-low yields obtained during
purification of the synthesized compounds was due
to the fact that during the extraction procedures, it
was difficult to break the emulsions formed, which
significantly complicated the extraction procedure.
Currently, we are conducting studies to find a
good technique for preserving the activity of the
fruits of glossy privet, and thereby eliminate the
dependence on their seasonal availability. A green
and biodegradable biocatalyst, such as this, can
be used as manure after service in the synthetic
reaction, thus minimizing the waste of chemicals
Acknowledgements
M.L. Aimar, L.I. Rossi, and M.I. Velasco gratefully
acknowledge financial support from Secretaría de
Ciencia y Tecnología de la Universidad Nacional de
Córdoba, Consejo Nacional de Investigaciones
Científicas y Técnicas (CONICET), and Agencia
Nacional de Promoción Científica y Técnica. L.I.R
is a researcher from CONICET. M.I.V. is a grateful
recipient of a fellowship from CONICET. We thank
Dr Paul Hobson, native speaker, for the revision of
this manuscript and Dr Gloria M. Bonetto for per-
forming NMR spectra.This work is dedicated to the
teaching career of Dr. Juan Carlos Maria Oberti.
Declaration of interest: The authors report no
conflict of interest.The authors alone are responsible
for the content and writing of the paper.
References
Assunção JCC, Machado LL, Lemos TLG, Cordell GA,
Monte FJQ. 2008. Sugar cane juice for the bioreduction of
carbonyl compouds. J Mol Catal B: Enzym 52–53:194–198.
(Kumaraswamy & Ramesh 2003).

356 M. L. Aimar et al.
Baldassarre F, Bertoni G, Chiappe C, Marioni F. 2000. Prepara-
tive synthesis of chiral alcohols by enantioselective reduction
with Daucus carota root as biocatalyst. J Mol Catal B: Enzym
Machado LL, Monte FJQ, de Oliveira MCF, de Mattos MC,
Lemos TLG. 2008. Bioreduction of aromatic aldehydes and
ketones by fruits barks of Passiflora edulis. J Mol Catal B: Enzym
54:130–133.
1
1:55–58.
Blanchard N, van de Weghe P. 2006. Daucus carota L. mediated
bioreduction of prochiral ketones. Org Biomol Chem 4:
Machado LL, Souza JSN, de Mattos MC, Sakata SK,
Cordell GA, Lemos TLG. 2006. Bioreduction of aldehydes
and ketones using Manihot species. Phytochemistry 67:
1637–1643.
Maczka WK, Mironowicz A. 2004. Enantioselective reduction of
bromo- and methoxy-acetophenone derivatives using carrot
and celeriac enzymatic system. Tetrahedron: Asymmetry
15:1965–1967.
2
348–2353.
Blay G, Hernández-Olmos V, Pedro JR. 2010. Synthesis of (S)-
ꢁ)-sotalol and (R)-(–)-isoproterenol via a catalytic enanti-
oselective Henry reaction. Tetrahedron: Asymmetry 21:
78–581.
(
5
Bohman B, Cavonius LR, Unelius CR. 2009. Vegetables as
biocatalysts in stereoselective hydrolysis of labile organic
compounds. Green Chem 11:1900–1905.
Bruni R, Fantin G, Medici A, Pedrine P, Sachetti G. 2002. Plants
in organic synthesis: an alternative to baker’s yeast.Tetrahedron
Lett 43:3377–3379.
Caron D, Coughlan AP, Simard M, Bernier J, Piché Y,
Chênevert R. 2005. Stereoslective reduction of ketones by
Daucus carota hairy root cultures. Biotechnol Lett 27:713–716.
Cheemala N, Gayral M, Brown JM, Rossen K, Knochel P. 2007.
New paracyclophane phosphine for highly enantioselective
ruthenium-catalyzed hydrogenation of prochiral ketones.
Synthesis 24:3877–3885.
Cohen AN. 2002. Success factors in the establishment of
human-dispersed organisms. In: Bullock JM, Kenward RE,
Hails RS, eds. Dispersal Ecology. Oxford: Blackwell Science
Ltd. pp. 374–394.
Mathre DJ, Thompson AS, Douglas AW, Hoogsteen K,
Carroll JD, Corley EG, Grabowski EJJ. 1993. A Practical Proc-
ess for the Preparation of Tetrahydro-l-methyl-3,3-diphenyl-
lH,3H-pyrrolo1[,2-c]-[1,3,2]oxazaborole-Borane.
A
Highly
Enantioselective Stoichiometric and Catalytic Reducing Agent.
J Org Chem 58:2880–2888.
Matsugi M, Itoh K, Nojima M, Hagimoto Y, Kita Y. 2004.
A novel separation technique of diastereomeric esters of pyri-
dylethanols by extraction: formal total synthesis of PNU-
142721, HIV-1 reverse transcriptase inhibitor. Tetrahedron
60:3311–3317.
Mironowicz A. 1998. Biotransformation of racemic acetates by
potato and topinambur tubers. Phytochemistry 47:1531–1534.
Nakamura K, MatsudaT. 1998. Asymmetric reduction of ketones
by the acetone powder of Geotrichum candidum. J Org Chem
63:8957–8964.
Comasseto JB, Assis LF, Andrade LH, Schoenlein-Crusius IH,
Porto ALM. 2006. Biotransformations of ortho-, meta and para-
aromatic nitrocompounds by strains of Aspergillus terreus:
Reduction of ketones and deracemization of alcohols. J Mol
Catal B: Enzym 39:24–30.
Ni Y, Xu J-H. 2012. Biocatalytic ketone reduction: A green and
efficient access to enantiopure alcohols. Biotechnol Adv
30:1279–1288.
Noyori R. 1994. Asymmetric Catalysis in Organic Synthesis. New
York. Wiley-Interscience.
Cordell GA, Lemos TLG, Monte FJQ, de Matos MC. 2007.
Vegetables as chemical reagents. J Nat Prod 70:478–492.
Ferreira DA, da Costa JC, Gomes de Lemos ATL, Queiroz Monte
FJ. 2012. Asymmetric reduction of acetophenone derivatives by
Lens culinaris. Biocatal Biotransform 30:469–475.
Fonseca AM, Monte FJQ, Oliveira MCF, Mattos MC,
Cordell GA, Braz-Filho R, Lemos TLG. 2009. Coconut
water (Cocos nucifera L.) - A new biocatalyst system for organic
synthesis. J Mol Catal B: Enzym 57:78–82.
Ohkuma T, Koizumi M, Yoshida M, Noyori R. 2000. General
asymmetric hydrogenation of hetero-aromatic ketones. Org
Lett 2:1749–1751.
Olejniczak T, Mironowicz A, Wawrzenczyk C. 2003. Enzymatic
lactonizatio of gamma-epoxy esters by the apple fruit and
Jerusalem artichoke bulb. Bioorg Chem 31:199–205.
Pauchard A, Cavieres L, Bustamante R, Becerra P, Rapoport E.
2004. Increasing the understanding of plant invasions in south-
ern South America: First symposium on Alien Plant Invasions
in Chile. Biological Invasions 6:255–257.
Pedragosa-Moreau S, Morisseau C, Baratti J, Zylber J,
Archelas A, Furstoss R. 1997. An enantioconvergent synthesis
of the p-blocker (R)-Nifenalol using a combined chemoenzy-
matic approach. Tetrahedron 53:9707–9714.
Phukan K, Devi N. 2012. Biocatalytic Preparative Method of
Asymmetric Alcohols Using Lycopersicum esculentum (Tomato).
Int J Chem Tech Research 4:203–207.
Giri A, DhingraV, Giri CC, Singh A,Ward OP, Narasu ML. 2001.
Biotransformations using plant cells, organ cultures and enzyme
systems: current trends and future prospects. Biotechnol Adv
1
9:175–199.
Hoyos L, Gavier Pizarro GI, Kuemmerle T, Bucher EH,
RadeloffVC,Tecco P. 2010. Invasion of glossy privet (Ligustrum
lucidum) and native forest loss in the Sierra Chicas of Córdoba,
Argentina. Biological Invasions 12:3261–3275.
Ishihara K, Hamada H, HirataT, Nakajima N. 2003. Biotransfor-
mation using plant cultured cells. J Mol Cat B: Enzym 23:
Sakamaki H, Itoh K, Taniai T, Kitanaka S, Takagi Y, Chai WC,
Horiuchi A. 2005. Biotransformation of valencene by cultured
cells of Gynostemma pentaphyllum. J Mol Catal B: Enzym
32:103–106.
Salvi NA, Chattopadhyay S. 2001. Studies on Rhizopus arrhizus
mediated enantioselective reduction of arylalkanones. Tetrahe-
dron 57:2833–2839.
Salvi NA, Chattopadhyay S. 2008. Asymmetric reduction of
halo-substituted arylalkanones with Rhizopus arrhizus.
Tetrahedron: Asymmetry 19:1992–1997.
Shimoda K, Hamada H, Hamada H. 2008. Glycosylation of
hesperetin by plant cell cultures. Phytochemistry 69:
1135–1140.
1
45–170.
Kumaraswamy G, Ramesh S. 2003. Soaked Phaseolus aureus L:
an efficient biocatalyst for asymmetric reduction of prochiral
aromatic ketones. Green Chem 5:306–308.
Liu X, Pan ZG, Xu JH, Li HX. 2010. Enantioselective reduction
of acetophenone analogues using carrot and celeriac enzymes
system. Chinese Chemical Letters 21:305–308.
Longo MA, Sanromaín MA. 2006. Production of food aroma
compounds: microbial and enzymatic methodologies. Food
Technol Biotechnol 44:335–353.
Lu C, Luo Z, Huang L, Li X. 2011. The Ru-catalyzed enantiose-
lective preparation of chiral halohydrins and their application
in the synthesis of (R)-clorprenaline and (S)-sotalol. Tetrahe-
dron: Asymmetry 22:722–727.
Singh VK. 1992. Practical and useful methods for the enantiose-
lective reduction of unsymmetrical ketones. Synthesis 7:
605–617.

Fruits of the glossy privet (Ligustrum lucidum—Oleaceae) as biocatalysts 357
Villa R, Molinari F, Levati M, Aragozzini F. 1998. Stereoselective
reduction of ketones by plant cell cultures. Biotechnol Lett
0:1105–1108.
Wohlgemuth R. 2010. Biocatalysis–key to sustainable industrial
chemistry. Curr Opin Chem Biol 21:713–724.
Xie Y, Xu J-H, Lu W-Y, Lin G-Q. 2009. Adzuki bean: A new
resource of biocatalyst for asymmetric reduction of aromatic
ketones with high stereoselectivity and substrate tolerance.
Bioresource Technology 100:2463–2468.
Yadav JS, Nanda S, Reddy PT, Rao AB. 2002. Efficient enanti-
oselective reduction of ketones with Daucus carota Root. J Org
Chem 67:3900–3903.
Yadav JS, Reddy BVS, Sreelakshmi GC, Kumar GGKSN,
Rao AB. 2008. Enatioselective reduction of 2-substituted
tetrahydropyran-4-ones using Daucus carota plant cells. Tetra-
hedron Lett 49:2768–2771.
Yang Z-H, Zeng R, Yang G, Wang Y, Li L-Z, Lv Z-SH, Yao M,
Lai B. 2008. Asymmetric reduction of prochiral ketones to
chiral alcohols catalyzed by plants tissue J Ind Microbiol
Biotechnol 35:1047–1051.
Yu F, Zhang X-CH,Wu F-F, Zhou J-N, FangW,Wu J, Chan ASC.
2011. Cobalt(II)-catalyzed asymmetric hydrosilylation of
simple ketones using dipyridylphosphine ligands in air. Org
Biomol Chem 9:5652–5654.
2