Tandem peroxidase-glucose oxidase catalysed enantioselective sulfoxidation of thioanisoles

Article abstract of DOI:10.1039/b000529k

The synthesis of chiral sulfoxides from sulfides with high level of chemical conversion and asymmetric induction was discussed. The method used the oxygen transfer propensity of a peroxidase-glucose oxidase bioenzymatic system. The degree of conversion of

Full text of DOI:10.1039/b000529k

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Tandem peroxidase–glucose oxidase catalysed enantioselective
sulfoxidation of thioanisoles
a,c
b
c
Krzysztof Okrasa, Eryka Guibé-Jampel* and Michel Therisod*
1
a
Division of Drugs Technology and Biotechnology, Warsaw University of Technology,
Noakowskiego 3, 00-662 Warsaw, Poland
UMR 8615 Réactivité et Synthèses Sélectives and Laboratoire de Chimie Bioorganique
et Bioinorganique, I. C. M. O. - Bat. 410–420, Université Paris-Sud, F-91405 Orsay cedex,
France. E-mail: therisod@icmo.u-psud.fr; eriguibe@icmo.u-psud.fr
b
c
Received (in Cambridge, UK) 18th January 2000, Accepted 6th March 2000
Chiral sulfoxides were synthesised from sulfides with high
levels of chemical conversion and asymmetric induction
using the oxygen transfer propensity of a peroxidase–
glucose oxidase bienzymatic system.
Peroxidases (POD) are potentially attractive biocatalysts for
selective oxidative transformations which meet industrial and
ecological preferences. Indeed, numerous peroxidases have been
shown to catalyse enantioselective sulfoxidation, epoxidation1,
benzylic hydroxylation and oxidation of alcohols and indole.
The mechanism of peroxidase oxygenation of organic com-
pounds involves the transfer of two electrons at the expense of
H O . However, the synthetic utility of this reaction is limited
2
2
by the fact that haem-POD shows low stability towards added
H O or other hydroperoxides (oxidative haem inactivation,
2
2
1,2
oxidation of the porphyrin ring). Furthermore, the enantio-
selective oxidation of various substrates by POD is often in
competition with their spontaneous oxidation by H O . Slow
2
2
addition of the oxidant is then essential for successful stereo-
selective biotransformation. Numerous processes for keeping
a low hydroperoxide concentration through the continuous
Fig. 1 GOD catalysed production of gluconic acid from glucose (2
mmol/10 ml, pH 7) at 25 (A) and 40 ЊC (B), monitored by pH-stat
controlled addition of 2 M NaOH. Concentration of hydrogen
peroxide in the medium was estimated spectrophotometrically at
240 nm (C).
2
3
addition of oxidant (use of a syringe pump, of an H O -stat;
2
2
4
5
use of t-BuOOH or other alkyl hydroperoxides or urea–
H O ) have been proposed. Recently, chloroperoxidase (CPO)
6
2
2
catalysed oxidations assisted by dihydroxyfumaric acid or
7,8
ascorbic acid and O have been published. However, form-
2
ation of H O as an intermediate in these transformations was
2
2
titration through oxidation of a chromogenic substrate by a
peroxidase (available as commercial kits). However, this pro-
considered doubtful by the authors. Furthermore, only non-
enzymatic oxidation was observed in the presence of horse-
9
cedure has not been applied to preparative peroxidase mediated
oxidations (only one patent—disclaimed—reports the prepar-
ation of propylene oxide via a halohydrin formed by the com-
8
radish peroxidase (HRP). We have investigated the oxygen
transfer propensity of a glucose oxidase–peroxidase system
in which H O is progressively generated in situ at the expense
10
2
2
bined action of a chloroperoxidase and an oxidase ).
of glucose, H O and O (Scheme 1).
2
2
The well documented transformation of sulfides to sulf-
1,4,11–13
oxides
was taken as a model in our work. A plant
peroxidase from Coprinus cinereus (Cip: EC 1.11.1.7), recently
studied in the oxidation of methyl phenyl sulfide, was
2,5
selected. Cip is a robust enzyme employed in a mixture with
lipases (BioCip Novo Nordisk) as a cleaning agent and is now
available from Roche Molecular Biochemicals (Mannheim).
Glucose oxidase (GOD: EC 1.1.3.4) from Aspergilius niger
was chosen as the standard catalyst for the oxidation of glucose.
It is a commercially available, inexpensive enzyme which is
9
produced for food or diagnostic use. Commercial GOD (here
Gluzyme BG from Novo Nordisk) can be used without any
purification. The kinetics of the formation of gluconic acid by
GOD were monitored at 20 and 40 ЊC in an aqueous solution
of glucose maintained at pH 7 by addition of NaOH using a
pH-stat. Simultaneous H O formation was checked by UV-
2
2
2
Scheme 1
spectrophotometry (Fig. 1). A mixture of substrate and Cip
was added at the same time to GOD in glucose solution. The
2
A standard method for determination of glucose concen-
tration in biological fluids is based on glucose oxidase (GOD)
catalysed formation of hydrogen peroxide and its concomitant
pH of the reaction medium was held at 7. The sulfide oxidation
was monitored by GC (Fig. 2) and the enantiomeric excess of
the sulfoxide was measured by HPLC on a chiral column.
DOI: 10.1039/b000529k
J. Chem. Soc., Perkin Trans. 1, 2000, 1077–1079
This journal is © The Royal Society of Chemistry 2000
1077

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Table 1 Oxidation of aryl methyl sulfides 1a–e to aryl methyl sulfoxides 2a–e catalysed by GOD–POD systems
Substrate
0.5 mmol)
Conversion
a
b
(
Time/h
Temp./ЊC
Volume/ml
GOD/mg
POD
(%)
Ee (%)
1
1
1
1
1
1
1
1
1
a
a
a
a
b
c
d
d
e
3
4
4
25
25
25
25
25
40
40
25
40
20
5
4
2
2
2
2
2
4
2
4
—
Cip
Cip
HRP
90
95
85
80
90
91
65
95
0
0
75
79
64
88
90
57
58
—
c
d
10
10
20
10
20
10
20
e
4
c
3.5
12
4
15
20
Cipc
Cipc
Cipc
Cip
c
Cip
a
b
c
d
e
Estimated by GC. Estimated by chiral HPLC. NOVO SP 676 (20 mg). From Roche (5 ml). Sigma, type II (40 mg).
H O from the cheap and environmentally friendly precursors
2
2
glucose and oxygen. The substrates and the enzymes can be
mixed at once, which avoids the cumbersome, slow continuous
addition of the oxygen donor. Chemical yields and enantio-
meric excesses are in keeping with the highest values reported in
the literature for the tested enzymes.
Experimental
Cip was obtained from Novo Nordisk (Novo SP 676) as a
freeze-dried powder, containing 639 mg of protein per g of
Ϫ1
solid, with a specific activity of 2675 kPODU g (one PODU
converts 1 µmol of hydrogen peroxide per min in a system
where 2,2Ј-azinobis(3-ethyl-1,3-benzothiazoline-6-sulfonate) is
oxidised), and from Roche Molecular Biochemicals (Mann-
heim) as an aqueous solution with a specific activity of 10 kU
per g of solution. HRP (type II) was from Sigma. GOD was
obtained from Novo Nordisk (Gluzyme 10000 BG) as a solid,
Ϫ1
with a specific activity of 10 kU g . Sulfides 1a, 1b and 1e
are commercially available. Sulfides 1c and 1d were prepared
14,15
Fig. 2 Simultaneous production of gluconic acid from glucose
according to described methods.
(
1 mmol) (A) and of phenyl methyl sulfoxide (estimated by GC) from
phenyl methyl sulfide (0.5 mmol) (B) catalysed by a GOD–Cip system at
Typical procedure
2
5 ЊC, pH 7 (GOD: 2 mg; Cip: 20 mg).
Thioanisole (0.5 mmol) and Cip (20 mg, 53 000 units) were
added to a solution of GOD (2 mg, 20 units) and glucose
(1 mmol) in 10 ml of water. The mixture was gently stirred, and
the pH was maintained at 7 by continuous addition of 2 M
sodium hydroxide using a pH-stat (Metrohm). After 4 hours,
the reaction was quenched by addition of sodium sulfite, and
the product was extracted with ethyl acetate. The formed sulfox-
ides are all known in optically active form. The physical and
spectroscopic properties of our specimen were in agreement
with those reported. Enantiomeric excesses were determined by
HPLC on Chiralcel OD, using pentane–propan-2-ol 95:5 as
the eluent. The degree of conversion of the sulfides and the
chemical purity of the sulfoxide (>98%) were determined by
gas-chromatography on a CP.SIL 19 CB capillary column. The
concentration of hydrogen peroxide during the GOD catalysed
Aryl methyl sulfides were oxidised as a suspension in water
Ϫ1
at a concentration of 0.05–0.1 mol l . The optimal Cip to
substrate molar ratio was approximately 1:1400. At lower
ratios, spontaneous (non-stereoselective) thioanisole oxidation
by H O could be observed. Two mole equivalents of glucose
2
2
ensured the continuous formation of H O at a flux of 100 µmol
2
2
Ϫ1
h . The activity of the Cip–GOD tandem stays stable at 20, 40
and 50 ЊC in terms of the yield and enantioselectivity of the
reaction.
After consumption of 1.2 eq. of NaOH the reaction mixture
was quenched with sodium sulfite, extracted with ethyl acetate
and the product was isolated with a yield >90% (Table 1).
Under these conditions, thioanisole 1a gave the corresponding
sulfoxide 2a with an ee of 75%, In the absence of peroxidase,
oxidation was complete, but as expected, was without asym-
metric induction.
2
oxidation of glucose was monitored spectrophotometrically at
Ϫ1
Ϫ1
2
40 nm (ε = 43.6 M cm ).
Methyl p-tolyl sulfide 1b gave 2b with 88% ee and 2-naphthyl
methyl sulfide 1c gave 2c (at 40 ЊC) with 90% ee. p-Chlorophenyl
methyl sulfide 1d gave 2d with only 57% ee. This modest value
could not be improved by doing the reaction at various temper-
atures (4–40 ЊC). p-Nitrophenyl methyl sulfide 1e was not
oxidised at all, probably as a result of the deactivation of the
sulfide toward oxidation due to the electron withdrawing
substituent. These last two results are in keeping with the
Acknowledgements
We are grateful to Novo Nordisk and to Roche Molecular
Biochemical (Mannheim) for the kind supply of enzymes,
and to the Laboratoire des Carbocycles, Orsay, for helpful
assistance.
5,13
literature.
Under the conditions described, a GOD–HRP
system oxidised 1a to 2a with an ee of 64% at 80% conversion.
Notes and references
2,12
For both Cip and HRP, in accordance with the literature,
the (S)-sulfoxides were formed in contrast to the sulfoxidation
1
M. P. J. van Deurzen, F. van Rantwijk and R. A. Sheldon, Tetra-
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A. Tuynman , M. K. S. Vink, H. L. Dekker, H. E. Schoemaker and
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1,11
catalysed by CPO, which gives the (R)-enantiomer.
2
In conclusion, we have demonstrated that GOD–POD medi-
ated preparative oxidation of sulfides provides an improved
process that takes advantage of the progressive generation of
3 M. P. J. van Deurzen, K. Seelbach, F. van Rantwijk, L. U. Krag and
R. A. Sheldon, Biocatal. Biotransform., 1997, 15, 1.
1
078
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