Tandem enzyme-catalysed oxidations of alkyl phenyl sulfides and alkyl benzenes: enantiocomplementary routes to chiral phenols.

Article abstract of DOI:10.1039/b205903g

Dioxygenase-catalysed trioxygenation of alkyl phenyl sulfides and alkyl benzenes yields enantiopure cis-dihydrodiol sulfoxides and triols respectively; naphthalene cis-dihydrodiol dehydrogenase-catalysed aromatisation of these diastereoisomers gives enant

Full text of DOI:10.1039/b205903g

Tandem enzyme-catalysed oxidations of alkyl phenyl sulfides and alkyl
benzenes: enantiocomplementary routes to chiral phenols
Derek R. Boyd,*ab Narain D. Sharma,a Vera Ljubez,a Breige E. Byrne,a Steven D. Shepherd,a
Christopher C. R. Allen,b Leonid A. Kulakov,bc Michael J. Larkinb and Howard Daltond
a School of Chemistry, The QueenAs University of Belfast, Belfast, UK BT9 5AG. E-mail: dr.boyd@qub.ac.uk
b QUESTOR Centre, The QueenAs University of Belfast, Belfast, UK BT9 5AG
c
School of Biology and Biochemistry, The QueenAs University of Belfast, Belfast, UK BT9 5AG
d Department of Biological Sciences, The University of Warwick, Coventry, UK CV4 7AL
Received (in Cambridge, UK) 19th June 2002, Accepted 9th July 2002
First published as an Advance Article on the web 29th July 2002
Dioxygenase-catalysed trioxygenation of alkyl phenyl sul-
fides and alkyl benzenes yields enantiopure cis-dihydrodiol
sulfoxides and triols respectively; naphthalene cis-dihy-
drodiol dehydrogenase-catalysed aromatisation of these
diastereoisomers gives enantiopure catechols of either con-
figuration.
3BS were separable by HPLC. The absolute configurations of
the enantiopure cis-diol sulfoxide metabolites 3AR, 3BR, 3ER,
3ES, 3FR and 3FS were determined by circular dichroism (CD)
spectral comparison (and stereochemical correlations) with the
corresponding catechols 4 and their dimethoxy derivatives 5
with alkyl aryl sulfoxides of established configuration (Table
1).1 Absolute configurations and enantiopurity values were also
Toluene dioxygenase (TDO), from Pseudomonas putida UV4,
has been found to catalyse stereoselective monooxygenation
(benzylic hydroxylation or sulfoxidation),1–3 dioxygenation
(cis-dihydroxylation or bis-benzylic hydroxylation),4–6 and
trioxygenation (benzylic hydroxylation/cis-dihydroxylation).7,8
Two new approaches to trioxygenated arenes using P. putida
UV4 i.e. monosulfoxidation/cis-dihydroxylation e.g. 1A ? 2A
? 3A and ketone reduction/cis-dihydroxylation e.g. 9D ? 10D
? 11D, are presented in this report. The trioxygenated products
3 and 11 were all found to be substrates for naphthalene cis-diol
dehydrogenase (NDD) present in Escherichia coli DH5a
(pUC129::narB), a recombinant strain (E. coli narB) con-
structed using the NDD gene expressed by Rhodococcus sp.
NCIMB 12038.9
Using whole cells of P. putida UV4, a source of TDO,
stereoselective sulfoxidation is the preferred biotransformation
pathway of alkyl, aryl and diaryl sulfides.1 Conversely, cis-
dihydroxylation of a phenyl group was strongly favoured over
sulfoxidation of a benzyl alkyl sulfide by this strain.3 It has now
been found that using the standard method for triol formation,8
allied to an extended period of biotransformation ( > 18 h) and
the reported improved isolation procedure (involving complete
removal of the water under mild conditions),3 with alkyl aryl
sulfides as substrates, e.g. 1A, 1B, 1E, TDO-catalysed tandem
trioxygenation (1? 2 ? 3) produces the cis-dihydrodiol
sulfoxide diastereoisomers 3AR, 3BR, and 3ER in ca.: 80% (30
mmol), 80% (32 mmol) and 20% (5 mmol) isolated yields
respectively (Scheme 1).10 Formation of the enantiopure
metabolite 3FS from the diaryl sulfide 1F provides a novel
example of enzyme-catalysed stereodifferentiation between
prochiral phenyl groups i.e. preferential cis-dihydroxylation of
the pro-S group. TDO-catalysed oxygenation reactions in P.
putida UV4 show a marked preference for one of the two
prochiral lone pairs,1 sulfur atoms,11 hydrogen atoms,8 methy-
lene groups5 and now phenyl groups. The formation of a
remarkably stable cis-dihydrodiol sulfone bioproduct 7B of
sulfide 1B (15% isolated yield) using P. putida UV4, brings to
light a new type of TDO-catalysed polyoxygenation, i.e.
tetraoxygenation (sulfoxidation/cis-dihydroxylation/sulfonida-
tion).
Scheme 1 Major bioproducts formed from arenes 1, 8 and 9.
Mixtures of the cis-dihydrodiol sulfoxide diastereoisomers
3AR/3AS, 3BR/3BS, 3ER/3ES or 3FR/3FS10 were prepared from:
(i) biotransformation of the corresponding racemic, 2A, 2B and
2E, or meso, 2F, sulfoxide substrates and (ii) dimethyl
dioxirane oxidation of the corresponding cis-dihydrodiol sul-
fides 6A, 6B, 6E and 6F obtained by a chemoenzymatic
method.3,12 Separation of the cis-dihydrodiol sulfoxide diaster-
eoisomeric pairs 3ER/3ES or 3FR/3FS was achieved by PLC
(6% MeOH in CHCl3). The diastereoisomers 3AR/3AS or 3BR/
Table 1 Optical rotations, ([a]D, CHCl3), and absolute configurations (R/S)
of compounds (Cpd) 5A, 5B, 5E, 5F and 13A-D
Cpd
5A
5B
5E
5F
13A
13B
13C
13D
(+)-[a]D 220 (R) 256 (R) 230 (R) 159 (S) 23 (R) 9 (R) 15 (R) 6 (R)
(2)-[a]D 231 (S) 161 (R) 25 (S) 9 (S) 16 (S) 5 (S)
1914
CHEM. COMMUN., 2002, 1914–1915
This journal is © The Royal Society of Chemistry 2002

determined by 1H-NMR analysis of the corresponding boronate
derivatives formed using (R)- and (S)-2-(1-methoxyethyl)
benzeneboronic acids as reported for other trioxygenated
biopoducts.3,8
Aromatisation (dehydration) of the individual cis-diol sulf-
oxide diastereoisomers 3AR, 3BR, 3ER, 3ES, 3FR and 3FS,
either thermally or in the presence of acid, yielded a mixture of
ortho-and meta-phenols with evidence, in some cases, of partial
racemisation of the sulfoxide stereogenic centre. A milder
enzyme-catalysed approach to aromatisation was thus
adopted.
methylated, and aromatised/deprotected under acid conditions
to give enantiopure phenol 15.10 This phenol proved to be a
promising new reagent for the determination of enantiopurity of
chiral carboxylic acids e.g. the pharmaceutical intermediate
ketoprofen, by 1H-NMR (OMe signal) and 19F-NMR (CF3
signal) spectral analysis of the derived esters. It was also found
to be a good resolving agent (TLC or HPLC separation) when
tested on racemic samples of chiral acid e.g. 2-arylpropanoic
acids.
In conclusion, enantiopure cis-dihydrodiol sulfoxides 3 and
triols 11, produced in good yields by enzyme-catalysed and
chemoenzymatic reactions of sulfides 1, alkyl benzenes 8 and a
ketone 9D have been used to develop enantiocomplementary
routes to a series of new enantiopure phenols. These have
already shown potential as new chiral ligands, reagents for
diastereoisomeric resolution and determination of enantiopur-
ity.
Recently we have shown9 that the NDD enzyme present in
the recombinant strain, E. coli narB, accepts naphthalene cis-
dihydrodiol as substrate. This new strain has not however been
tested with other types of cis-dihydrodiol substrates. E. coli
narB was thus used with the cis-diol sulfoxide substrates 3AR,
3BR, 3ER and 3FS (from the corresponding sulfide precursors
1A, 1B, 1E and 1F), 3ES and 3FR (from the cis-dihydrodiol
sulfides 6E and 6F). The corresponding catechol enantiomers
4AR, 4BR, 4ER, 4ES, 4FR and 4FS were obtained as metabolites
(50–70% yield) using a general procedure.† Due to their
variable stability in solution, the catechols were characterised as
the stable dimethoxy or the diacetoxy derivatives. The absolute
configurations of all the catechol metabolites were determined
by stereochemical correlation. These assignments were con-
firmed for catechols 4AR, 4BR, 4ER, 4ES and 4FS by
comparison of the CD spectra of the dimethoxy derivatives
5AR, 5BR, 5ER and 5ES, with those of the corresponding
alkylphenyl sulfoxides of known configurations (Table 1).
The tandem conversion (TDO-catalysed oxidation using P.
putida UV4) of the alkylbenzene substrates 8A–8C via the
corresponding monol intermediates 10AR–10CR to the triol
metabolites 11AR–11CR was carried out using the reported
method8 and the improved isolation procedure.3 Addition of the
commercially available benzylic alcohol enantiomers 10AS–
10CS as substrates yielded the corresponding triol diastereoi-
somers 11AS (4 mmol, 79%), 11BS (5 mmol, 89%) and 11CS (1
mmol, 65%). A novel biotransformation pathway was observed
when ketone 9D was used as substrate with whole cells of P.
putida UV4; the only bioproducts observed were the separable
triol diastereoisomers 11DR/11DS (95+5, 230 mmol, 50%
yield).10 It was evident that a stereoselective dehydrogenase-
catalysed reduction of ketone 9D had occurred and that the
transient benzylic alcohol products 10DR/10DS were rapidly
oxidised to yield triols 11DR/11DS.
We thank the BBSRC for postdoctoral support (N. D. S.),
DENI for a CAST Award (B. B.) and a Distinction Award (S.
S.) and the QueenAs University of Belfast (B. B. and V. L.) for
postgraduate studentships and Mrs D. Markovich for technical
assistance with the biotransformations.
Notes and references
† E. coli narB was grown at 37 °C in Luria broth with ampicillin (0.1 mg/
cm3) and isopropyl-b- -thiogalactopyranoside (IPTG, 0.05 mg/cm3); cells
D
were harvested, in late exponential growth phase, by centrifugation, washed
and resuspended (shake flasks; OD600 = 5–10) in potassium phosphate
buffer (0.05 M, pH 7.2) for performing biotransformations at 30 °C.
Substrates were added at concentrations between 0.2–0.5 mg/cm3 and the
reactions terminated after 18 h. The catechols were isolated, from the
aqueous medium after saturating it with sodium chloride, by repeated
extractions with EtOAc.
1 D. R. Boyd, N. D. Sharma, S. A. Haughey, M. A. Kennedy, B. T.
McMurray, G. N. Sheldrake, C. C. R. Allen, H. Dalton and K. Sproule,
J. Chem. Soc., Perkin Trans. 1, 1998, 1929.
2 D. R. Boyd, N. D Sharma and C. C. R. Allen, Curr. Opin. Biotechnol.,
2001, 12, 564.
3 D. R. Boyd, N. D. Sharma, S. A. Haughey, J. F. Malone, A. King, B. T.
McMurray, R. Holt and H. Dalton, J. Chem. Soc., Perkin Trans.1, 2001,
3288.
4 D. R. Boyd and G. N. Sheldrake, Nat. Prod. Rep., 1998, 15, 309.
5 N. I. Bowers, D. R. Boyd, N. D. Sharma, P. A. Goodrich, M. R.
Groocock, A. J. Blacker, D. A. Clarke, P. Goode and H. Dalton, J.
Chem. Soc., Perkin Trans 1, 1999, 1461.
6 T. Hudlicky, D. Gonzalez and D. T. Gibson, Aldrichim. Acta, 1999, 32,
33.
7 D. T. Gibson, B. Gshwendt, W. K. Yeh and V. M. Kobal, Biochemistry,
1973, 12, 1520.
8 D. R. Boyd, N. D. Sharma, N. I. Bowers, J. Duffy, J. S. Harrison and H.
Dalton, J. Chem. Soc., Perkin Trans. 1, 2000, 1345.
9 L. A. Kulakov, C. C. R. Allen, D. A. Lipscomb and M. J. Larkin, FEMS
Microbiol. Lett.,, 2000, 182, 327.
Addition of the triols 11AR–11DR and 11AS–11DS to E. coli
nar B cultures,† gave the corresponding catechols 12AR–12DR
and 12AS–12DS (50–70% yield). These enantiopure catechols
were also characterised as their more stable dimethylethers
13AR–13DR and 13AS–13DS and as triacetates (Table 1).
Enantiopure catechols 4 and derivatives 14 are currently
10 cis-Diol sulfoxides ([a]D, MeOH): 3AR (+200), 3BR (+254), 3ER
(+141), 3ES (2188), 3FR (+297), 3Fs (+178), cis-diol sulfone 7B (27);
triols 11DR (+15) and 11DS (+16); phenol 15 (247, CHCl3).
11 J. R. Cashman, L. D. Olsen, D. R. Boyd, R. A. S. McMordie, R. Dunlop
and H. Dalton, J. Am. Chem. Soc., 1992, 114, 8772.
12 D. R. Boyd, N. D. Sharma, B. Byrne, M. V. Hand, J. F. Malone, G. N.
Sheldrake, J. Blacker and H. Dalton, J. Chem. Soc., Perkin Trans., 1,
1998, 1935.
being evaluated as chiral ligands for asymmetric alkylation and
other reactions. The acetonide derivative of triol 11DR was
CHEM. COMMUN., 2002, 1914–1915
1915