Thymol Bromination - A Comparison between Enzymatic and Chemical Catalysis

Article abstract of DOI:10.1002/ejic.201500086

The catalytic activity of the vanadium-dependent bromoperoxidase isolated from the brown alga Ascophyllum nodosum is compared with the activity of a cheap, commercially available V-catalyst precursor in the bromination of thymol. Organic solvents have been avoided to make the system appealing from a sustainable chemistry point of view. It is noteworthy that, notwithstanding the low solubility of the substrate, the thymol bromination reactions were performed in water, with a safe brominating source, under mild conditions, and with relatively inexpensive reagents. In this regard, the greenness of the systems was evaluated by the estimation of the E-factor value; the result is that the chemical reaction has a lower environmental impact than the enzymatic process, with an E-factor in the range of eco-friendly processes. Catalysis of thymol bromination by vanadium derivatives is directly compared to catalysis by a V-dependent bromoperoxidase. All reactions were performed under mild and sustainable conditions with relatively inexpensive reagents. Appealing results were obtained in terms of selectivity and sustainability.

Full text of DOI:10.1002/ejic.201500086

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DOI:10.1002/ejic.201500086
Thymol Bromination – A Comparison between Enzymatic
and Chemical Catalysis
[
a]
[b]
[a]
Federica Sabuzi, Ekaterina Churakova, Pierluca Galloni,
[
c]
[b]
[a]
Ron Wever, Frank Hollmann, Barbara Floris, and
Valeria Conte*[
a]
Keywords: Vanadium / Enzyme catalysis / V-bromoperoxidases / Sustainable chemistry / Bromination / Thymol
The catalytic activity of the vanadium-dependent bromoper-
in water, with a safe brominating source, under mild condi-
oxidase isolated from the brown alga Ascophyllum nodosum tions, and with relatively inexpensive reagents. In this
is compared with the activity of a cheap, commercially avail- regard, the greenness of the systems was evaluated by the
able V-catalyst precursor in the bromination of thymol. Or-
ganic solvents have been avoided to make the system
appealing from a sustainable chemistry point of view. It is
noteworthy that, notwithstanding the low solubility of the
substrate, the thymol bromination reactions were performed
estimation of the E-factor value; the result is that the
chemical reaction has a lower environmental impact than the
enzymatic process, with an E-factor in the range of eco-
friendly processes.
Introduction
Haloperoxidases are able to oxidize halide ions to
halogenating species, whose nature may vary as a function
of the reaction conditions, in the presence of hydrogen per-
oxide. A synthetically useful HalPO for bromo-functionali-
zation of various substrates is the vanadium-dependent
Brominated compounds are the most abundant organo-
[
1]
halides in nature. This class of molecules finds applica-
tions in different fields such as agrochemicals and pharma-
ceuticals, mainly because of their interesting antifungal,
antibacterial, antiviral, and anti-inflammatory activities.
Furthermore, brominated derivatives are considered impor-
tant synthetic intermediates for several selective and ef-
ficient transformations. This applies likewise for brominated
thymol derivatives.
bromoperoxidase (V PO) isolated from the brown alga
Br
Ascophyllum nodosum. V PO-catalyzed oxidation of brom-
Br
ide in the presence of hydrogen peroxide has proved to be
a sustainable methodology to synthesize various bromo de-
rivatives. Edge and X-ray absorption near-edge structures
of this bromoperoxidase were obtained, and they substanti-
For these reasons, bromination of organic compounds
is a very important reaction in organic synthesis. Classical
methods usually involve the use of molecular bromine,
which is corrosive and toxic; therefore, the development of
safer and greener protocols for halogenation reactions is
mandatory. Haloperoxidase enzymes (HalPOs), isolated
mainly from marine algae and fungi, have received much
attention, because they are known to play a major role in
the biosynthesis of brominated compounds, such as
halogenated indoles, terpenes, acetogenins, phenols, and
[5]
ate its mechanism of action.
The catalytic effect of V PO is related to the formation
Br
of a vanadium–peroxido complex, in the active site, in the
[6]
presence of hydrogen peroxide. This species, which is a
stronger oxidant than H O , is able to catalyze the oxid-
2
2
ation of bromide ion to a brominating intermediate, which
may then react with an appropriate organic substrate to
form products or with another oxidant molecule to cause
[7]
its decomposition with formation of singlet dioxygen
(Scheme 1).
[
2–4]
hydrocarbons.
[
a] Dipartimento di Scienze e Tecnologie Chimiche,
Università di Roma “Tor Vergata”,
Via della Ricerca Scientifica, 00133, Rome, Italy
E-mail: valeria.conte@uniroma2.it
http://www.stc.uniroma2.it/
[
[
b] Department of Biotechnology, Delft University of Technology,
Julianalaan 136, 2628 BL, Delft, The Netherlands
c] Van ’t Hoff Institute for Molecular Sciences (HIMS),
Faculty of Science, University of Amsterdam,
Science Park 904, 1098 XH Amsterdam, The Netherlands
Supporting information for this article is available on the
WWW under http://dx.doi.org/10.1002/ejic.201500086.
Scheme 1. VBrPO-catalyzed bromide oxidation by hydrogen perox-
ide.
These processes likely occur in different regions of the
enzyme. In particular, the first one may take place in a
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V
Scheme 2. V -based two-phase system for bromination reactions.
hydrophilic portion close to the active site, considering that sive method to mimic the mode of action of V PO, provid-
Br
at least two molecules of water and a proton are required ing the opportunity to study the chemistry of such enzymes.
[
8]
to oxidize bromide, while the functionalization of the sub- Additionally, the oxidative bromination of toluene has been
strate may occur in a more hydrophobic location.
performed with the system described above, achieving the
Several reactions with different substrates have been per- functionalization at the benzylic position.[28] The reaction
formed by using V PO and, under appropriate conditions, has also been carried out without solvent, thus avoiding the
Br
bromocyclization reactions, indole functionalizations, and use of chlorinated co-solvents to obtain products derived
bromohydrin synthesis have been achieved.[ Also oxidative from the electrophilic aromatic substitution.
3]
bromination reactions of phenol and substituted phenols
The aim of the present work is to directly compare the
have been performed, and these reactions have usually led oxidative bromination reaction catalyzed by a cheap, com-
to selective para-bromination with respect to the hydroxy mercially available vanadium catalyst precursor, such as
group together with formation of the corresponding NH VO , with that of the V PO from Ascophyllum
4
3
Br
ortho,para-diBr derivatives, both resulting from the classical nodosum by using thymol, a monoterpenoid phenolic com-
[8,9]
electrophilic aromatic substitution.
No products of side- pound, as model substrate. To the best of our knowledge,
chain bromination were detected. The role of vanadium only few examples of thymol bromination reaction are re-
haloperoxidases in the formation of volatile brominated ported in the literature.[4,29,30] To be able to demonstrate
compounds from natural sources has been recently dis- the sustainability of our procedures, the chemical and the
cussed in a Perspective Article, evidencing their impact on enzyme-catalyzed bromination of thymol were performed
[
10]
the environment.
Bromoperoxidases as catalysts for oxi- without solvent.
[4]
dative bromination have been reviewed.
A wide variety of vanadium complexes have been synthe-
sized and used as functional models for V-haloperoxidases Results and Discussion
in order to have a better understanding of the mechanism
V
The V catalyst was used to perform reactions on a pre-
of action of such enzymes, and these complexes are effective
catalyst precursors in various oxidation reactions such as
epoxidation of alkenes, oxidation of sulfides, alkanes,
arenes and alcohols, as well as in oxidative bromination
parative scale in order to isolate and characterize the reac-
tion products (Scheme 3). The reactions proceeded hetero-
geneously, yielding high amounts of product mixtures.
[11–25]
reactions in the presence of hydrogen peroxide
hydroperoxides.
or alkyl
[
26]
Inspired by the mechanism of action of V PO, a bi-
Br
phasic procedure for bromination reactions was proposed
[
27]
in 1994. According to this procedure, in order to mimic
the hydrophobic and hydrophilic regions of the enzyme, a
double-phase system is required (Scheme 2).
Scheme 3. Product analysis of the V-catalyzed oxidative thymol
bromination.
With this system, good results in terms of yields and
Thymol bromination accomplished with an equimolar
selectivity have been achieved for the bromination of aro- amount of bromide with respect to the substrate occurred
matic substrates, alkenes and alkynes[ and, according to on the activated position for electrophilic aromatic substitu-
these results, the system seems to be a simple and inexpen- tion in the phenol ring. Specifically, the bromination reac-
15]
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tion led to the formation of two isomers, 4-bromothymol oxybromination of alkenes and alkynes;[27,32] recently good
being, as expected, the main product because of steric results have also been obtained in the benzylic bromination
[
28]
hindrance at the ortho position. A further bromination of of toluene.
At higher reaction temperature (Entry 7),
the para isomer gave 2,4-dibromothymol in good yield. quantitative conversion of the substrate was observed, and
Subsequently, the reaction was explored by changing pa- the bromination reaction proceeded faster. However, it was
rameters that may influence the rate and the overall yield accompanied by the oxidation of the substrate with
of the chemical bromination of thymol. In particular, as hydrogen peroxide, which also led to the formation of 2-
shown in Table 1, KBr and H O concentrations, together isopropyl-5-methyl-1,4-benzoquinone as byproduct.
2
2
with reaction temperature and pH, were changed systemati-
cally.
The kinetic behavior of the model bromination is shown
in Figure 1. It can be seen that the initial part of the process
is quite fast; in fact, substrate conversion and product
formation are almost complete only after 3 h, while the cat-
alyst remains active even after 24 h, indicating that, under
these experimental conditions, the decomposition of
hydrogen peroxide is not predominant.
Table 1. Oxidative bromination of thymol: chemical catalysis. Reac-
tion conditions: 100 mm thymol, 100 mm KBr, 200 mm H O , un-
2 2
less otherwise stated.
[
a] Reaction volume: 1 mL, at 30 °C, pH = 1 for 24 h. [b] Calculated
with respect to substrate conversion. [c] 100 mm H . [d] 200 mm
2 2
O
KBr. [e] Reaction volume: 1 mL, at 30 °C, pH = 2.5 for 24 h. [f]
Reaction volume: 1 mL, at 45 °C, pH = 1 for 24 h. [g] 2-Isopropyl-
5-methyl-1,4-benzoquinone detected as further product (7%).
Reactions performed with about 5% of the catalyst pre-
cursor (Entries 1–3) showed high yields and good selectivity
toward the formation of 4-bromothymol. Conversions and
product distributions did not vary significantly when either
the initial bromide or H O concentrations were changed.
With lower precatalyst concentration, lower substrate Figure 1. V -catalyzed oxidative thymol bromination as a function
conversion with similar product distribution was observed of time. Reaction conditions: volume 1 mL, 100 mm thymol,
2
2
V
1
2 2 4 3
00 mm KBr, 200 mm H O , 5 mm NH VO , pH = 1, 30 °C.
(Entry 4). On the other hand, in the presence of 0.5 equiv.
of NH VO (Entry 5), a dibromination reaction occurred:
under these conditions the relatively high catalyst concen-
tration likely accelerates successive bromination.
The reaction performed at higher pH value (Entry 6) has
a very low yield, probably due to the formation of a diper-
oxido–metal complex. In fact, it is known that vanadate
dissolved in aqueous solutions in the presence of hydrogen
peroxide forms several species in equilibrium. The preva-
4
3
The results obtained with the V PO enzyme are
Br
collected in Table 2. Interestingly, under experimental con-
ditions similar to those used for the chemical model, a
lower conversion of the substrate together with a different
product distribution were observed (Table 2, Entry 1). In
this case, the amounts of both 2- and 4-bromothymol were
similar, and formation of dibrominated 2,4-dibromothymol
was also observed. Possible explanations of this behavior
are either a permanence of the monobrominated derivatives
near the active site of the enzyme, which makes a second
functionalization possible, or different reactivity due to the
lence of the different peroxido complexes is strongly in-
fluenced by pH.[
31,32]
In particular, at pH = 2.5 species with
more than one peroxido group bound to the metal are
favored, while at pH = 1 the monoperoxido complex is the
most abundant species in solution (Scheme 4).
[
6]
higher pH.
As far as the enzymatic process is concerned, the kinetic
plot (Figure 2) shows that the reaction does not proceed
any further after 3 h. The low substrate conversion (27%)
might be due to inactivation of the enzyme by hydrogen
peroxide. To substantiate this hypothesis, the stability of the
enzyme in the presence of high concentrations of H O was
2
2
Scheme 4. Mono- and diperoxido vanadium(V) complexes in equi-
librium at pH = 2.5.
investigated, and indeed after 5 h of incubation, the enzyme
lost 85% of its initial activity, in agreement with the work
Furthermore, the monoperoxido vanadium complex was by Soedjak and Butler,[33] who showed inactivation and
more reactive than the diperoxido complex toward the inhibition at high peroxide concentrations (Figure 3).
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Table 2. Oxidative bromination of thymol: enzymatic catalysis. Re- ation was measured as a function of KBr concentration
action conditions: 100 mm thymol, 100 mm KBr.
with 1 mm H O (Figure 4).
2
2
[
a] Reactions were carried out in 50 mm citrate buffer pH = 6.3
final volume 1 mL), at 30 °C for 24 h using VBrPO 1 mg/mL. [b]
Calculated with respect to the substrate conversion. [c] 10 μL ali-
quots of 1 m H were added every 30 min for 10 times (final
(
2
O
2
concentration 100 mm).
Figure 4. VBrPO activity at different KBr concentrations.[34] Reac-
tion conditions: phosphate buffer 50 mm, pH = 8, 1 mm H
00 μm TB, VBrPO 0.002 mg/mL, 25 °C.
O ,
2 2
1
A saturation kinetic behavior was observed. In addition,
with bromide concentrations equal to those used to perform
chemical bromination (between 100 and 200 mm), the en-
zyme showed the highest activity while bromide inhibition
was not detected.
In an attempt to reduce the inactivation o the enzyme
tion of time. Reaction conditions: volume 1 mL, 100 mm thymol, due to the presence of high hydrogen peroxide concentra-
Figure 2. VBrPO-catalyzed oxidative thymol bromination as func-
2 2
100 mm KBr, 100 mm H O , VBrPO 1 mg/mL, 50 mm citrate buffer
tions particularly at the beginning of the reaction, the
bromination was performed by adding the oxidant stepwise.
As expected, a higher yield was observed (Table 2, Entry 2),
accompanied by a different product distribution.
pH = 6.3, 30 °C.
To further improve the enzyme performance, it is advis-
[
35–37]
able to generate H O in situ.
Preliminary experiments
2
2
using light-driven H O generation with flavin mononucleo-
2
2
tide (FMN) as photocatalyst and sodium formate as sacrifi-
cial electron donor have been performed (Scheme 5).
2 2
Scheme 5. Light-driven in situ H O generation.
As shown in Figure 5, by using in situ H O generation,
thymol consumption was observed for 70 h, and a conver-
sion of about 80% was obtained. This shows that the en-
2
2
Figure 3. VBrPO remaining activity after incubation in 100 mm of
[
34]
H
H
2
O
O
2
.
Incubation conditions: [VBrPO] = 1 mg/mL; 100 mm
2
2
; 50 mm citrate buffer pH = 6.3, 30 °C; activity assay: 50 mm
phosphate buffer, pH = 8; 100 mm KBr, 1 mm H
.01 mg/mL VBrPO.
2
O
2
, 100 μm TB; zyme activity is preserved over many hours, and conver-
sions similar to those with the chemical system can be
achieved. Further optimization is currently ongoing in our
0
Moreover, in order to evaluate V PO activity in pres- laboratories to turn this system into a practical and synthet-
Br
ence of KBr, the initial rate of thymol blue (TB) bromin- ically relevant one.
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in principle, solvents can be recycled, and this should
significantly decrease the E-factor.
For these reasons, when solvent contributions are not
taken into account, E-factor values decrease drastically for
both reactions. In particular, a value of 3.7 kg
ϫ
waste
kgproduct–1 has been obtained for the chemical reaction and
1
6 kgwaste ϫ kgproduct–1 for the enzymatic one (Figure 7).
V
Hence, these estimations indicate that at present V -
catalyzed thymol bromination is more sustainable than the
enzymatic reaction in terms of produced wastes, having an
E-factor in the range of eco-friendly processes.
Figure 5. VBrPO-catalyzed oxidative thymol bromination in a light-
driven reaction as a function of time. Reaction conditions: volume
1
mL, 100 mm thymol, 200 mm KBr, 0.5 mg/mL VBrPO, 200 mm
sodium formate, 5 mm FMN, 50 mm citrate buffer, pH = 6.3, 30 °C.
Enzymatic reactions are often claimed as environmen-
tally friendly in comparison to conventional catalytic reac-
tions. For this reason, the environmental impact of the
enzymatic bromination of thymol was evaluated and com-
pared to that of the catalytic process. To this end, the
environmental factor (E-factor = kgwaste ϫ kgproduct–1) can
[
37–40]
be used to firstly evaluate the greenness of a reaction.
Obviously, the ideal E-factor is zero.
In the present work, the E-factor value for the optimized Figure 7. E-factor values for chemical and enzymatic thymol
bromination (solvents contributions not considered).
chemical reaction is 280, while for the non-optimized enzy-
matic reaction it is around 1000. It should be noted that
the two main contributions for both systems derive from
the solvent used for the reaction (i.e. water) and from the
solvent (i.e. diethyl ether) used to extract the products (see
Conclusions
It is fairly difficult to tell which brominating method is
more sustainable”. Table 3 compares some important
Figure 6).
“
features of both methods.
Table 3. Key parameters to compare chemical and enzymatic
bromination of thymol.
Parameter
NH
4
VO
3
VBrPO
Conversion (%)
100
92
15
42
TON (molprod ϫmolcat–1)
1804
1481
0.0031
–
1
TFmax (TONϫh )
Catalyst consumption (gcat ϫgprod–1)
0.25
The chemical method yields full conversion, whereas the
enzyme catalyst enables only 42%. The major reason for
this discrepancy was shown to be the rather poor robustness
of the biocatalyst towards the oxidant (H O ). Already por-
2
2
tionwise addition of H O resulted in significantly higher
2
2
product concentrations. Further improvements can be ex-
pected from in situ generation of H O to supply the bio-
Figure 6. E-factor values for chemical and enzymatic thymol
bromination.
2
2
catalyst with sufficient but not detrimental amounts of
[
30,35–37]
Nevertheless, water is usually considered a green solvent H O for generation of the brominating species.
2
2
and it is generally excluded from the E-factor calculation.
A simple comparison of the environmental impact of the
However, this assumption is valid only for pure water, two methods by using the E-factor[
39–41]
demonstrates that
whereas when the water is contaminated it needs treatment biocatalysis is not per se environmentally more benign than
and purification, which constitutes a negative contribution chemical catalysis. In both cases, the relatively high solvent
to the eco-compatibility of the system. As for diethyl ether, contribution (due to the low product concentrations used)
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4 2 4
again demonstrates the importance of high reactant con- another centrifugation step (2800 rpm, 15 min). (NH ) SO was
added to the supernatant to a final saturation of 80%. The precipi-
tated bromoperoxidase was separated from the supernatant by cen-
trifugation (13000 rpm, 30 min). The precipitate was homogenized
centrations to improve not only the economic attractiveness
but also the environmental impact.
in a medium containing ethanol (60%) and TRIS–H
pH 8.3). After centrifugation at 10000 rpm for 30 min, the pellet
obtained was resuspended in TRIS–H SO buffer (0.2 m, pH 8.3)
and loaded onto a DEAE–Sephacel column. The protein was
eluted with TRIS–H SO buffer (1 m, pH 8.3) containing NaCl
1 m). Fractions containing bromoperoxidase activity were pooled,
concentrated on an Amicon ultrafiltration cell (Filter, PM 30), and
dialyzed against sodium acetate (10 mm, pH 5). Portions of this
enzyme preparation were stored at –20 °C.
2 4
SO (0.2 m,
Experimental Section
2
4
Instrumentation: Gas chromatographic analyses were carried out
with a Varian 3900 instrument equipped with an FID 1770 detector
and a 30 m Supelco SPB-5 column (0.25 mm diameter and 0.25 μm
2
4
(
internal film). GC–MS spectra were recorded with a Shimadzu
GC–MS-QP2010 Ultra spectrometer. 1H NMR spectra were re-
3
corded with a Bruker Avance 300 MHz instrument, with CDCl as
solvent. UV assays were performed with use of a Shimadzu UV/Vis
spectrophotometer UV-2401 PC, with disposable plastic cuvettes of
V
BrPO Activity Assay: VBrPO activity was evaluated by using a
colorimetric assay based on thymolsulfonphthalein (Thymol Blue,
1.5 mL volume.
[34]
TB) bromination at 25 °C.
00 μL) dissolved in H O/dimethyl sulfoxide (4:1) was added to
phosphate buffer (880 μL, 100 mm, pH 8) containing KBr
100 mm). Then, H (10 μL, 100 mm) and VBrPO (10 μL) from
A stock solution of TB (1 mm,
1
2
Materials: All commercial reagents and solvents were used as re-
ceived, without further purification. Thymol bromination products
were synthesized according to the catalytic procedure described be-
(
2 2
O
the 50ϫ diluted stock solution were added (final VBrPO concentra-
tion 2 μg/mL). The absorbance change of the resulting mixture at
low and purified chromatographically (SiO
chloromethane 3:1).
2
; eluent: hexane/di-
620 nm was recorded every minute, and it was converted to di-
4-Bromo-2-isopropyl-5-methylphenol (4-Bromothymol): The com-
bromothymolsulfonphthalein (TBBr ) formed per minute (in mm),
2
pound was isolated as a white powder. MS (EI): m/z (%) = 228 (30)
–
1
–1
by using ε = 37.2 mm cm . The specific activity in thymol blue
bromination was (11.2Ϯ0.6) U/mg.
+
+
+
[
M] , 230 (30) [M + 2] , 213 (85) [M – CH
3
] , 134 (100) [M –
+
1
CH
3
Br] . H NMR (CDCl
3
): δ = 7.31 (s, 1 H), 6.66 (s, 1 H), 4.65
(s, 1 H), 3.19–3.07 (m, 1 H), 2.32 (s, 3 H), 1.26–1.23 (d, J = 7 Hz,
General Procedure for VBrPO Catalyzed Reactions: Thymol (15 mg,
6
H) ppm.
0.1 mmol) and KBr (11.9 mg, 0.1 mmol) were dissolved in citrate
2 2
buffer (890 μL, 50 mm, pH 6.3). Then, H O (10 μL, 10.4 m) and
2
-Bromo-6-isopropyl-3-methylphenol (2-Bromothymol): The com-
the stock solution of VBrPO (100 μL, 10.8 mg/mL) were added, and
the mixture was stirred at 30 °C. Afterwards, the reaction products
were extracted with three portions of diethyl ether, dried with
pound was isolated as a colorless oil. MS (EI): m/z (%) = 228 (30)
+
+
+
[M] , 230 (30) [M + 2] , 213 (85) [M – CH
3
] , 134 (100) [M –
+
1
CH
6
3
Br] . H NMR (CDCl
3
): δ = 7.08–7.05 (d, J = 7 Hz, 1 H),
anhydrous Na
reaction products were dissolved in CDCl
solution (50 μL) was diluted with CDCl
,4-Dibromo-6-isopropyl-3-methylphenol (2,4-Dibromothymol): The known amount of CH NO and analyzed by 1H NMR spec-
2
SO
4
, and filtered. The solvent was evaporated. The
(500 μL). Then, this
(450 μL) containing a
.83–6.79 (d, J = 8 Hz, 1 H), 5.72 (s, 1 H), 3.39–3.25 (m, 1 H), 2.39
3
(s, 3 H), 1.28–1.23 (d, J = 7 Hz, 6 H) ppm.
3
2
3
2
compound was synthesized with the same procedure but with the
troscopy.
previously synthesized 4-bromothymol as substrate. MS (EI): m/z
+
+
+
=
306 (20) [M] , 308 (40) [M + 2] , 310 (20) [M + 4] , 291 (55)
+
+
]+, Acknowledgments
[M – CH
3
] , 293 (100) [M + 2 – CH
12 (50) [M – CH
3
] , 295 (55) [M + 4 – CH
3
+
+ 1
2
3
Br] , 214 (50) [M + 2 – CH
3
Br] . H NMR
(
(
CDCl
3
): δ = 7.33 (s, 1 H), 5.70 (s, 1 H), 3.35–3.21 (m, 1 H), 2.54
COST Action CM1003 “Biological Oxidation Reactions – Mecha-
nisms and Design of New Catalysts” is acknowledged for having
allowed a stimulating European Research Environment and a Short
Term Scientific Mission for F. S. Dr. Heiko Lange is gratefully ac-
knowledged for GC–MS analysis and Dr. Martina Tiravia for help-
ful discussions.
s, 3 H), 1.26–1.22 (d, J = 7 Hz, 6 H) ppm.
Catalytic Procedure: Thymol (15 mg ,0.1 mmol) and KBr (11.9 mg
0.1 mmol) were dissolved in an aqueous solution of NH VO
(20 μL, 10.4 m) and perchloric acid
20 μL of 5 m) were added to obtain a pH value of 1. The mixture
,
(
(
4
3
2 2
960 μL, 5 mm). Then, H O
was stirred at 30 °C for 24 h. Afterwards, the reaction products
were extracted with three portions of diethyl ether, dried with
[1] See, for example: a) G. W. Gribble, Chem. Soc. Rev. 1999, 28,
335–346; b) M. Liu, P. E. Hansen, X. Lin, Mar. Drugs 2011, 9,
anhydrous Na
reaction products were dissolved in CDCl
aliquot (50 μL) of this solution was diluted with CDCl
containing CH NO as internal standard and analyzed by
NMR spectroscopy.
2
SO
4
, and filtered. The solvent was evaporated. The
(500 μL). Then, an
(450 μL)
1
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© 0000 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

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Received: January 27, 2015
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FULL PAPER
V-Catalyzed Oxidative Bromination
F. Sabuzi, E. Churakova, P. Galloni,
R. Wever, F. Hollmann, B. Floris,
V. Conte* .......................................... 1–8
Catalysis of thymol bromination by vana-
dium derivatives is directly compared to
catalysis by a V-dependent bromoperox-id-
ase. All reactions were performed under
mild and sustainable conditions with rela-
tively inexpensive reagents. Appealing re-
sults were obtained in terms of selectivity
and sustainability.
Thymol Bromination – A Comparison be-
tween Enzymatic and Chemical Catalysis
Keywords: Vanadium / Enzyme catalysis /
V-bromoperoxidases / Sustainable chemis-
try / Bromination / Thymol
Eur. J. Inorg. Chem. 0000, 0–0
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