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PAPER
Ionic liquids for enhancing the enantioselectivity of isolated
BVMO-catalysed oxidations†‡
Cristina Rodr´ıguez,a Gonzalo de Gonzalo,b Marco W. Fraaijeb and Vicente Gotor*a
Received 16th September 2010, Accepted 22nd October 2010
DOI: 10.1039/c0gc00560f
The present study describes the first-time usage of an isolated thermostable Baeyer–Villiger
monooxygenase (phenylacetone monooxygenase, PAMO) in the presence of ionic liquids. The
stability, activity and selectivity of PAMO as an oxidative enzyme in the presence of different ionic
liquids were studied. This revealed that the addition of some specific ionic liquids, such
AmmoengTM 102 and [bmim]MeSO4, can significantly enhance the E-value in the oxidation of
racemic benzylketones. Moreover, the use of ionic liquids increases the optimal substrate
concentration for performing Baeyer–Villiger oxidation, thereby extending the biocatalytic
repertoire of PAMO for synthetic applications.
The Baeyer–Villiger reaction implies the oxidative cleavage
Introduction
of a carbon–carbon bond adjacent to a carbonyl group,11
representing a very useful tool for the preparation of high-added
value compounds. Some years ago, phenylacetone monooxy-
genase from Thermobifida fusca (PAMO, EC 1.14.13.92) was
purified and structurally characterized.12 From then on, several
studies have demonstrated the potential of this thermostable
flavoprotein as a biocatalyst in Baeyer–Villiger oxidations and
sulfoxidations.13 Recently it has been reported that PAMO
is also able to perform oxidations when working in non-
conventional media. In fact, the use of certain cosolvents such
as methanol or ethanol can improve and even reverse the
enantioselective behaviour of the biocatalysts.14–15 Moreover,
it has been discovered that PAMO can be used in Tris/HCl
or imidazole buffer with high ionic strength (unpublished)
and therefore tolerance towards ILs might be of practical
interest.
Biotransformations are considered as an effective methodology
in organic synthesis and pharmaceutical chemistry due to the
high selectivity and the eco-friendly conditions required for these
processes.1 Although water is the natural solvent for biocatalysis,
it is well-known that isolated enzymes can be used in the presence
of organic solvents.2 Despite the advantages provided by these
compounds, the search for alternative solvents to replace
conventional organic solvents is a major incentive for the op-
timization of biocatalytic processes in industry and for reducing
the environmental impact. In this regard, ionic liquids (ILs) are
attractive alternatives as they enable a green chemical approach.3
In the last decade ILs have been established as an alternative
reaction media for biocatalytic transformations due to their
properties such as non-volatility, non-flammability and high
thermal stability.4 Another advantage of using ILs instead of
conventional organic solvents is that the polarity, hydropho-
bicity, viscosity and solvent miscibility of IL can be tuned by
altering the cation and anion. This allows the design of media
for different purposes, e.g. to increase the substrate solubility or
to modify the enzyme activity and/or selectivity.
In this work, we report for the first time on the application
of a BVMO for catalysis of the oxidation of benzylketones in
media containing IL. The activity, regio- and enantioselectivity,
as well as the stability of PAMO has been studied in the presence
of different ILs.
For a number of enzymes it has been shown that they are cat-
alytically active in ILs or biphasic aqueous-IL systems. Although
the majority of these studies involved lipases5 or proteases,6 a few
reports on the use of redox enzymes in combination with ILs
have appeared recently.7–9 However, no such studies have ever
been reported on Baeyer–Villiger monooxygenases (BVMOs),
an interesting class of cofactor-dependent enzymes.10
Results and discussion
PAMO-catalyzed Baeyer–Villiger oxidation of phenylacetone in
aqueous-ionic liquid media
In order to investigate the efficiency of PAMO for better
catalysing Baeyer–Villiger (BV) reactions in mixtures of aqueous
buffer and ionic liquid, the oxidation of phenylacetone (1a) in
the presence of miscible or non-miscible ILs was firstly tested
(Fig. 1). ILs with different cations such as N-alkyl pyridinium
(bmp)+, N,N¢-alkyl imidazolium [(bmim)+, (bdmim)+, (hmim)+],
tetraalkylated ammonium [(thma)+, (tbma)+] or tetraalkylated
phosphonium (tbmp)+ derivatives and anions such as tetraflu-
aDepartamento de Qu´ımica Orga´nica e Inorga´nica, Instituto
Universitario de Biotecnolog´ıa de Asturias, Universidad de Oviedo,
C/Julia´n Claver´ıa 8, 33006, Oviedo, Spain. E-mail: vgs@fq.uniovi.es;
Fax: +34 985 103448; Tel: +34 985 103448
bLaboratory of Biochemistry, Groningen Biomolecular Sciences and
Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747
AG, Groningen, The Netherlands
-
-
† Electronic supplementary information (ESI) available: Complete
experimental procedures and chromatographic data. See DOI:
10.1039/c0gc00560f
‡ We would like to dedicate this paper to Prof. Antonio Garc´ıa Mart´ınez
on the occasion of his retirement.
oroborate (BF4 ), hexafluorofosfate (PF6 ), trifluoromethylsul-
-
fonate (OTf-) or methylsulfate (MeSO4 ) were used. Also the
effects of other quaternary ammonium salts, AmmoengTM 100,
AmmoengTM 101 and AmmoengTM 102, were studied. While
This journal is
The Royal Society of Chemistry 2010
Green Chem., 2010, 12, 2255–2260 | 2255
©