TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 791–793
Benzylic biooxidation of various toluenes to aldehydes
by peroxidase
a,
b
a
Rainer Russ, * Thomas Zelinski and Timm Anke
a
LB Biotechnologie, Universit a¨ t Kaiserslautern, Paul-Ehrlich-Str. 23, D-67663 Kaiserslautern, Germany
b
BASF-AG, Forschung Feinchemie, D-67056 Ludwigshafen, Germany
Received 4 November 2001; revised 28 November 2001; accepted 29 November 2001
Abstract—A catalytic method is described for the oxidation of toluene and substituted derivatives to the corresponding
benzaldehydes by hydrogen peroxide, using peroxidase. In most cases the respective benzoic acid was produced as a byproduct.
The reaction proceeds under mild conditions in an aqueous medium. © 2002 Elsevier Science Ltd. All rights reserved.
Substituted benzaldehydes are often used as feedstock
in industrial chemistry. The selective oxidation of aro-
matic methyl groups to the respective aldehyde is,
of our strain collection were able to catalyze the trans-
formation of toluene to benzaldehyde and, only to a
minor extent, benzoic acid (Table 1).
1
however, difficult. The chemical oxidation of the
methyl group commonly proceeds directly to the car-
boxylic acid. We therefore chose to investigate enzy-
matic methods, because enzymes can be chemoselective.
We started with the laccase/2,2-azino-bis(3-ethylbenz-
thiazoline-6-sulfonic acid) (ABTS) system of Potthast et
The transformation of 21 different substituted methyl
aromatics by hydrogen peroxide and Coprinus perox-
idase was tested. Only three of the compounds tested,
p-cymene (4-isopropyl-toluene), m-cresol, and p-cresol,
were not at all transformed into the corresponding
benzaldehydes. All other 18 compounds were trans-
formed into the respective benzaldehydes whereby the
efficiency of the reaction varied (Table 1). Suitable
substituents comprised methyl, halogen, methoxy, and
nitro groups. It seems that the position of the sub-
stituent was more important than its composition.
Ortho or para positions of the substituent to the methyl
group were preferred against meta, except for the nitro-
toluenes. o-Nitrobenzaldehyde was obtained in a low
yield, whereas m-nitrobenzaldehyde was formed with
yields comparable to the p-isomer. In the case of o-
nitrotoluene, an interaction of the intermediate methyl
cation radical with the nitro group perhaps prevented
the formation of the aldehyde. o-Nitrotoluene was the
only substrate that produced the alcohol derivative.
The cresols were found not suitable for catalytic con-
version by Coprinus peroxidase and hydrogen peroxide,
probably due to polymerization reactions as described
2
al. However, with toluene and laccase from different
fungi, such as Bjerkandera adusta, Coriolus sp., Phelli-
nus sp., and Pleurotus ostreatus, we found no transfor-
mation at all, which is in accordance with the findings
3
of Fritz-Langhals and Kunath. Subsequently, we tried
several peroxidases with hydrogen peroxide as the oxi-
dant. Using lignin peroxidase from Phanerochaete
4
5
chrysosporium or Coprinus cinereus, we found no
transformation of toluene. Chloroperoxidase from Cal-
dariomyces fumago gave a slight transformation to ben-
6
,7
zyl alcohol and benzaldehyde, as reported earlier.
†
Finally, peroxidases isolated from a Coprinus species
Keywords: oxidation; chemoselectivity; biotransformation; biocataly-
sis; toluene; benzaldehyde.
*
Corresponding authors. Tel.: +49-6221-4038-430; fax: +49-6221-
4
Coprinus sp., a member of the ‘ink-cap’ mushroom family, was
fermented in 20 l soy meal medium. Mycelia and soy meal were
removed by centrifugation. The resulting supernatants were cleared
by ultrafiltration (pore size 0.16 mm). The filtrates were then concen-
trated by ultrafiltration using a membrane with a 10 kDa cutoff.
The partial purification of the peroxidase was performed by FPLC.
Two subsequent runs on Q-Sepharose at different pH-values (5 and
†
4
for lignin peroxidase. The preparation with o-cresol
immediately turned yellow after addition of the enzyme.
A weaker discoloration to yellow was observed for m-
and p-cresol, as well as toluene, 3-chloro-, 4-methoxy-,
and 4-fluorotoluene. p-Cymene was transformed into
two compounds that were not identified. A molecular
mass of 132 and 136 inferred that none of the com-
pounds was either an aldehyde- or carboxyl-derivative
of p-cymene.
7
.3, respectively) resulted in a 12 fold activity enrichment (specific
activity with ABTS 7.6 U/mg of protein) and the total removal of
laccase activity. Manuscript in preparation.
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