Chemoenzymatic one-pot reaction of noncompatible catalysts: Combining enzymatic ester hydrolysis with Cu(i)/bipyridine catalyzed oxidation in aqueous medium

Article abstract of DOI:10.1039/c7ra05451c

The combination of chemical catalysts and biocatalysts in a one-pot reaction has attracted considerable interest in the past years. However, since each catalyst requires very different reaction conditions, chemoenzymatic one-pot reactions in aqueous media remain challenging and are limited today to metal-catalysts that display high activity in aqueous media. Here, we report the first combination of two incompatible catalytic systems, a lipase based ester hydrolysis with a water-sensitive Cu/bipyridine catalyzed oxidation reaction, in a one-pot reaction in aqueous medium (PBS buffer). Key to the solution was the compartmentalization of the Cu/bipyridine catalyst in a core-shell like nanoparticle. We show the synthesis and characterization of the Cu/bipyridine functionalized nanoparticles and the application in the oxidation of allylic and benzylic alcohols in aqueous media. Furthermore, the work demonstrates the implementation of a one-pot reaction process with optimized reaction conditions involving a lipase (CAL-B) to hydrolyze various acetate ester substrates in the first step, followed by oxidation of the resulting alcohols to the corresponding aldehydes under aerobic conditions in aqueous media.

Full text of DOI:10.1039/c7ra05451c

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Chemoenzymatic one-pot reaction of
noncompatible catalysts: combining enzymatic
ester hydrolysis with Cu(I)/bipyridine catalyzed
oxidation in aqueous medium†
Cite this: RSC Adv., 2017, 7, 33614
*
Henning Sand and Ralf Weberskirch
The combination of chemical catalysts and biocatalysts in a one-pot reaction has attracted considerable
interest in the past years. However, since each catalyst requires very different reaction conditions,
chemoenzymatic one-pot reactions in aqueous media remain challenging and are limited today to
metal-catalysts that display high activity in aqueous media. Here, we report the first combination of two
incompatible catalytic systems, a lipase based ester hydrolysis with a water-sensitive Cu/bipyridine
catalyzed oxidation reaction, in a one-pot reaction in aqueous medium (PBS buffer). Key to the solution
was the compartmentalization of the Cu/bipyridine catalyst in a core–shell like nanoparticle. We show
the synthesis and characterization of the Cu/bipyridine functionalized nanoparticles and the application
in the oxidation of allylic and benzylic alcohols in aqueous media. Furthermore, the work demonstrates
the implementation of a one-pot reaction process with optimized reaction conditions involving a lipase
(CAL-B) to hydrolyze various acetate ester substrates in the first step, followed by oxidation of the
resulting alcohols to the corresponding aldehydes under aerobic conditions in aqueous media.
Received 14th May 2017
Accepted 23rd June 2017
DOI: 10.1039/c7ra05451c
rsc.li/rsc-advances
composition and pH aer the Pd-catalyzed Suzuki reaction before
the enzymatic reduction was carried out.7 In a subsequent work,
Introduction
the conditions for the rst step were modied and both reactions
could be conducted at r.t. in a 2-PrOH/H2O solvent mixture that
required only a pH adjustment before the enzymatic reaction
step.8 An very impressive example has been recently reported by
The design and implementation of one-pot tandem reactions
has received signicant attention in organic chemistry in the
past years as an important tool to carry out multi-step reactions
without the need to isolate and purify intermediates.1–3 Of
particular interest is the combination of enzymes with metal–
organic catalysts.4,5 However, owing to the fact that the majority
of enzymes are only active in aqueous medium and most metal–
organic catalysts prefer organic solvents, only a few examples of
such reactions have been reported so far.6–12
One of the rst examples is the dynamic kinetic resolution
(DKR) reported by Williams and co-workers based on a lipase
catalyzed resolution of alcohols under substrate-racemizing
conditions catalyzed by a rhodium catalyst [Rh2(OAc)4].6 The
chemo-enzymatic reaction was carried out by using the enzyme
Pseudomonas uorescens lipase and 2 mol% of the rhodium
catalyst [Rh2(OAc)4] simultaneously in an organic solvent. The Pd-
catalyzed Suzuki coupling followed by an enzyme catalyzed
asymmetric reduction in aqueous medium was one of the rst
¨
Groger and coworker who described the one-pot reaction of the
CuCl/PdCl2 catalyzed Wacker oxidation followed by an enzymatic
reduction. To avoid any enzyme inhibition by Cu-ions, the Wacker
oxidation with CuCl/PdCl2 was conducted in the interior of
a polydimethylsiloxane thimble, which enables diffusion of
organic substrate and product into the exterior aqueous enzyme
phase while Cu ions are withheld.9
In most cases, the chemoenzymatic one-pot reaction of an
enzyme with a metal organic catalyst was only possible aer
extensive optimization of the reaction conditions, by using
aqueous solvent mixtures,7–10 adjusting reaction temperature or
pH,7–9 addition of scavengers,12 by consecutive addition of the two
catalysts to the one-pot system10,12,13 or by using a fed-batch
continuous-ow process with compartmentalized combination
of catalysts.14
¨
examples reported by Groger and coworker. Initial attempts
As a result, each catalytic reaction is conducted under non-
optimal conditions in favor of the one-pot reaction and over-
all activity is oen quite low. Another challenge is that all che-
moenzymatic one-pot reactions described so far require
a metal- or organo catalyst that displays activity in aqueous
media, water-sensitive reactions such as the L-proline catalyzed
required adjustment of the reaction temperature, solvent
Faculty of Chemistry and Chemical Biology, TU Dortmund, Otto-Hahn Str. 6, D 44227
Dortmund, Germany. E-mail: Ralf.Weberskirch@tu-dortmund.de
† Electronic supplementary information (ESI) available. See DOI:
10.1039/c7ra05451c
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aldol reaction in combination with an asymmetric reduction with a Cu(I)/bipyridine catalyzed oxidation reaction in a one-pot
catalyzed by the alcohol dehydrogenase (ADH) had to be con- two-step reaction in aqueous medium (PBS buffer). The key to
ducted in organic media.15
the combination of these two catalysts in a reaction sequence is
A reaction that attracted our attention recently, is the oxidation the compartmentalization of the Cu(I)/bipyridine catalyst in the
of a broad range of primary alcohols to the corresponding alde- hydrophobic core of a polymeric nanoparticle thus avoiding any
hydes by using a Cu(I)/bipyridine/nitroxyl co-catalyst system by mutual inhibition with the commercially available enzyme
Stahl and coworker.16,17 Compared to other oxidation procedure, lipase B from Candida antarctica (Immobead 150). At the same
the catalyst systems works at room temperature under aerobic time, the amphiphilic nanoparticle structure makes the Cu(I)/
conditions. Furthermore, catalyst activity can be signicantly bipyridine catalyst compatible with the aqueous reaction
enhanced by using ABNO instead of TEMPO as the N-oxyl source medium that is preferred by the enzyme (Scheme 1).
enabling also the oxidation of secondary alcohols under mild
reaction conditions.18 However, extensive testing of different
solvents showed that the best activity results were achieved using
Results and discussion
acetonitrile as a solvent whereas the presence of water reduced the
The idea to combine a lipase based ester hydrolysis with the
Cu(I)/bipyridine aerobic oxidation was based on several
considerations. Firstly, the lipase B from Candida antarctica
(CAL-B) is a well-known biocatalyst for ester hydrolysis and
product yield signicantly.16
An important reaction in organic chemistry is the selective
synthesis of an aldehyde from the corresponding ester.19 The
transesterication reactions.25–27 It exists in various commercial
forms, such as the Immobead 150, where the lipase is immo-
bilized on the surface of polymer microbeads as a carrier.28
Secondly, the very mild reaction conditions under which the
enzyme operates makes it an ideal candidate to be combined
with the Cu(I)/bipyridine based aerobic oxidation since both
reactions work at moderate temperatures under aerobic
conditions but differ in the best solvent for each reaction,
acetonitrile for the aerobic oxidation reaction and water or PBS
buffer for the enzymatic ester hydrolysis.
Therefore, initial experiments focused on studying enzyme
activity with a variety of ester substrates. We synthesized various
acetyl esters, including aromatic, allylic, aliphatic and heterocyclic
esters (ESI†) and studied their enzymatic hydrolysis by using the
commercially available lipase B from Candida antarctica (CAL-B)
immobilized on polyacrylate beads (Immobead 150). In a control
most common reducing agent involve the use of diisobutyla-
luminum hydride (DIBALH), which is commercially available
but gives moderate to high yields (48–90%) and requires very
low temperature (ꢀ70 ꢁC) in organic solvents, such as hexane.20
A second method is based on two steps via reduction to the
alcohol with LiAlH4 followed by oxidation to the aldehyde with
Pyridinium Chlorochromate (PCC) or Pyridinium Dichromate
(PDC) in THF (see Scheme 1) giving high yields with >90%.21
However, the latter reaction uses chromium compounds that
are category 1 heavy metals according to the International
agency for Research on Cancer.22 Therefore, the application of
catalytic reaction steps under environmentally friendly reaction
conditions (e.g. aqueous medium and moderate temperature) is
of great interest for the sustainable production of chemicals.23,24
Herein, we report the rst example to prepare aldehydes
from esters by a combination of a lipase catalyzed ester cleavage
Scheme 1 (A) Most common methods in organic chemistry to synthesize aldehydes from the corresponding aliphatic or benzylic esters. (B)
Alternative approach by the combination of enzymatic hydrolysis and Cu(I)/bipyridine mediated oxidation in a one-pot process in PBS buffer.
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Table 1 CAL-B catalysed cleavage of representative acetyl esters
Entry
1
Substrate
Conversiona (%)
Yieldb (%)
97 (ꢂ3)
93 (ꢂ2)
2
93 (ꢂ2)
87 (ꢂ2)
3
4
5
96 (ꢂ2)
95 (ꢂ1)
88 (ꢂ2)
87 (ꢂ2)
89 (ꢂ3)
79 (ꢂ2)
6
96 (ꢂ3)
71 (ꢂ4)
89 (ꢂ3)
67 (ꢂ4)
7c
8c
67 (ꢂ5)
59 (ꢂ4)
a
Determined by 1H-NMR-spectroscopy. b Isolated yield. c Reaction time of 40 min leads to quantitative conversion.
experiment, we checked the hydrolytic stability of benzyl acetate in
Although in the classical Cu(I)/bipyridine mediated oxidation
PBS buffer at 40 ꢁC for 20 min and no hydrolysis was observed. In of alcohols small amounts of water as co-solvent led to a drastic
the presence of the enzyme CALB, benzylic esters were converted reduction of product yield,16 we were recently able to transfer this
ꢁ
efficiently aer 20 min at 40 C to the corresponding alcohols in reaction to an aqueous medium by site-isolation of the Cu(I)/
87–93% yield (entry 1–4, Table 1). The allylic ester showed similar bipyridine catalyst in micelles prepared from polymeric amphi-
enzymatic activities with 67–89% yield (entry 5–7, Table 1), philes.29 Oxidation of benzyl alcohol as a model compound was
whereas lowest enzyme activity was observed for the aliphatic shown to be quantitative at room temperature aer 3 h (with
substrate with 59% yield (entry 8, Table 1). However, when the 5 mol% Cu(I)Br; 5 mol% polymer ligand, 5 mol% TEMPO and
reaction time was extended to 40 min the latter also gave quanti- 10 mol% NMI as a base) in water as reaction medium, suggesting
tative yield. Furthermore, the immobilized enzyme could be iso- that the micellar core–shell like structure is able to protect the
lated and separated from the reactants by simple ltration (ESI†). catalyst very efficiently from the aqueous phase. Therefore, our
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Scheme 2 Schematic structure of the core-crosslinked nanoparticle NP1 prepared from the amphiphilic block copolymer P1.
Scheme 3 Synthesis scheme of the 2-oxazoline monomer M1.
thought was to combine the enzymatic ester cleavage with the
To synthesize the desired nanoparticles we needed to intro-
micellar catalytic oxidation in a one-pot two-step system by using duce a new 2-oxazoline monomer M1 with a styrene function-
benzyl acetate as a model substrate. Initial studies revealed, ality for the nanoparticle crosslinking reaction. The monomer
however, that the enzymatic ester cleavage was quantitative aer synthesis is depicted in Scheme 3 and is based on two frag-
product isolation, whereas the subsequent oxidation step of the ments that are coupled via Click chemistry to yield M1. The 2-(5-
benzyl alcohol showed only 9% yield. The low product yield azidopentyl)-2-oxazoline fragment 2 was synthesized from 6-
indicated inhibition of the Cu(I)/bipyridine catalyst in the pres- bromohexanoic acid 1 according to Lav et al.38 in four steps with
ence of the enzyme most likely due to the dynamic nature of an overall yield of 43%. In the rst step 1 was reacted with NaN3
micellar aggregates.
in a nucleophilic substitution reaction. The a,u-azido hexanoic
A strategy to overcome this limitation of micelles is to stabilize acid was then converted to the corresponding active ester with
them by crosslinking the micellar core or shell and thus prevent N-hydroxyl succinimide (NHS) and 1-ethyl-3-(3-dimethylamino-
the dynamic exchange of single polymer amphiphiles.30–33 The propyl)carbodiimide (EDC) as a coupling agent. The corre-
main focus to apply shell crosslinked nanoparticles has been sponding NHS-ester was then reacted with 2-chlorethylamine
drug delivery whereas few reports used them also for catalysis before the 2-oxazoline ring was formed in the presence of a KOH
application.34,35 In contrast to medical application, where the as a base. The second fragment 1-(prop-2-yn-1-yloxy)methyl)-4-
focus is mainly the stabilization of micelles to prevent any vinylbenzene 4 was obtained by etherication of p-vinyl benzyl
premature drug release, the nanoparticles design for catalysis chloride 3 with propargyl alcohol PA in the presence of tetra-
application should enable high accessibility of the catalyst and butylammonium iodide (TBAI) and sodium hydride (NaH) in
easy diffusion of the substrate(s)/product(s) in and out the 98% yield. Both fragments 2 and 4 were nally coupled by Cu-
nanoparticle. Therefore, we decided to use the microemulsion mediated click chemistry to give the desired monomer M1.
approach for the preparation of catalytically active nano- The structure and purity of the monomer M1 were conrmed by
particles.36,37 These nanoparticles are composed of a crosslinked 1H NMR (ESI†).
core to prevent any interaction between the catalyst and the
The synthesis of the amphiphilic bipyridine functionalized
enzyme and an amphiphilic polymer shell which is compatible block copolymers was then carried out by a cationic ring-
with the aqueous phase and provides an hydrophobic layer that opening polymerisation mechanism (Scheme 4). The living
contains the Cu(I)/bipyridine catalyst (Scheme 2).
nature of the polymerisation allows the preparation of tailor-
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Scheme 4 Synthesis of the amphiphilic, bipyridine-functionalized block copolymer P1.
Table 2 Analytical data of the polymers P1 and P2
Polymer compositiona
Mn (g molꢀ1
)
Mn (g molꢀ1
)
Đc
cmcd (mol lꢀ1
)
dhe (in H2O) (nm)
dhe (in MeOH) (nm)
b
c
No.
P1
P2
Me40(Hep5BiPy4)Sty1
Me53(Hep4BiPy3)Hep3Sty1
6070
7170
6860
8940
1.18
1.50
1.2 ꢃ 10ꢀ6
15.2 (ꢂ0.51)
18.1 (ꢂ0.43)
2.8 (ꢂ0.16)
3.8 (ꢂ0.27)
5.0 ꢃ 10ꢀ7
a
Polymer composition as determined by 1H-NMR analysis; Me ¼ 2-methyl-2-oxazolin; Hep ¼ 2-n-heptyl-2-oxazolin; BiPy ¼ 4-methoxy-4-pentoxy-2,2-
bipyiridine units; Sty ¼ styrene-oxazoline M1. b Determined by 1H-NMR end group analysis in CDCl3. c Determined by SEC analysis in DMF/5 g lꢀ1
LiBr with linear PS standards. With pyrene (0.1 mM in MeOH). e By DLS measurements in water (1 mM) at room temperature.
d
made block copolymers of dened composition and molecular In the last step, P1 and P2 were employed in a microemulsion
weight.39–41 Based on this approach, we now extended the procedure using hexanediol dimethacrylate (HDDMA) as a cross-
amphiphilic diblock copolymer structure that is suitable for linking monomer and AIBN as the radical initiator. This resulted
micellar catalysis by another polymer block composed of the in stable core-crosslinked nanoparticles NP1 and NP2 with sizes
styrene-functionalized 2-oxazoline M1 without an additional in a range of 23–27 nm in water, which is slightly larger than the
spacer (three block model, P1) or with a short poly(2-n-heptyl-2- pure micelles. This is in good agreement with recent data.36,37
oxazoline) block (four block model, P2). The bipyridine ligand Successful core-crosslinking and nanoparticle formation was
was introduced into the polymers P1 and P2 in a polymer
analogous reaction aer the polymerisation has been nished.
The structure, composition and molecular weight of the
resulting polymers P1 and P2 were analysed by H NMR spec-
Table 3 Size of the nanoparticles NP1 and NP2 after crosslinking
1
a
No
dh (H2O) (nm) PDIa
dha (MeOH) (nm) PDIa
troscopy and size exclusion chromatography. Table 2 summa-
rizes the analytical data.
NP1 23.27 (ꢂ0.43)
NP2 25.38 (ꢂ0.40)
0.34 (ꢂ0.01) 23.46 (ꢂ1.26)
0.23 (ꢂ0.01) 27.70 (ꢂ0.48)
0.37 (ꢂ0.01)
0.23 (ꢂ0.01)
The critical micelle concentrations (cmc ¼ 0.5–1.2 mmol lꢀ1
)
and the hydrodynamic radii of the micelles with 15.2 to 18.1 nm
matched the typical range for amphiphilic poly(2-oxazoline)s.42–44
a
By DLS measurements (1 mM) at room temperature.
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Fig. 1 TEM images of NP1 (a, b) and NP2 (c, d) in water on a 100 nm and 50 nm scale.
Table 4 Aerobic oxidation of benzyl alcohol with P1/NP1 and P2/NP2 as polymeric liganda
Entry
Polymer
N-Oxyl
t (h)
Conversionb (%)
1a
1b
2a
2b
4a
4b
5a
5b
P1
P1
P2
P2
NP1
NP1
NP2
NP2
ABNO
TEMPO
ABNO
TEMPO
ABNO
TEMPO
ABNO
2
3
2
3
2
3
2
3
95 (ꢂ3)
99 (ꢂ0)
98 (ꢂ2)
96 (ꢂ2)
96 (ꢂ1)
95 (ꢂ1)
96 (ꢂ0)
95 (ꢂ1)
TEMPO
40 mol of CuBr, 5 mM solution of P1/NP1 and P2/NP2. Average conversion (2 runs) aer work-up and isolation determined by 1H-NMR-
a
b
spectroscopy.
conrmed in methanol as a non-selective solvent, showing similar substrate scope of the polymeric nanoparticles. Table 5
values for NP1 and NP2 compared to water (see Table 3). summarizes the results of representative alcohols in the Cu(I)/N-
Furthermore, nanoparticle formation was conrmed by TEM oxyl catalysed aerobic oxidation with NP1 as polymeric ligand.
measurements, which showed spherical particles for both nano-
particles NP1 and NP2 (Fig. 1).
As can be seen from Table 5, aromatic alcohols gave excellent
results with high yields (entry 1–5, Table 5), only the allylic and
aliphatic substrates led to lower product yield due to their lower
reactivity (entry 6, 7 Table 5). Importantly, for all tested alcohols
there was no signicant difference in activity between the
micellar catalytic approach and the nanoparticle system (see
ESI, Table S1†).
In the next set of experiments, we compared the recycling
properties of the polymeric nanoparticles with the micellar
catalytic approach using either TEMPO or ABNO as the N-oxyl
source. Therefore, the oxidation of benzyl alcohol was examined
in ve consecutive runs for the amphiphilic polymer P1 and the
nanoparticle system NP1 (Fig. 2).
As can be seen in Fig. 2 there is no signicant difference in
catalyst activity when comparing the micellar catalytic system
based on P1 with the corresponding nanoparticle system NP1.
The P1-system showed still around 90% conversion aer the 5th
run for both ABNO and TEMPO whereas the NP1-system ach-
ieved only around 80 to 85% aer the 5th cycle for TEMPO and
ABNO.
Application of the nanoparticles NP1/NP2 in catalysis
In the rst set of experiments, the catalytic activity of the
nanoparticles NP1/NP2 in the aerobic oxidation of benzyl
alcohol as a test substrate was investigated and compared with
the micellar catalytic approach based on P1/P2. As can be seen
from Table 4, no difference in catalytic activity was observed
when using polymer micelles based on P1/P2 or polymer
nanoparticles NP1/NP2 with a crosslinked core. This result
supported our hypothesis that core-crosslinked nanoparticles
do not show a reduced catalytic activity compared to polymer
micelles. Larger differences are observed when using different
N-oxyl sources, TEMPO versus ABNO, which is in agreement
with literature data.18
Since we did not observe any difference in catalytic activity
between NP1 (three block copolymer) and NP2 (four block
copolymer), we continued to work with NP1 to investigate the
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Table 5 Representative examples of functionalized alcohols in the Cu(I)/N-oxyl catalysed aerobic oxidation with NP1 as polymeric liganda
Entry
Substrate
N-Oxyl
t (h)
Conversionb (%)
Isolated yieldc (%)
1a
1b
ABNO
TEMPO
2
3
96 (ꢂ1)
95 (ꢂ1)
94
91 (ꢂ3)
2a
2b
ABNO
TEMPO
2
3
98 (ꢂ1)
98 (ꢂ1)
95 (ꢂ1)
95 (ꢂ2)
3a
3b
ABNO
TEMPO
2
3
98 (ꢂ1)
99 (ꢂ0)
93 (ꢂ2)
96 (ꢂ1)
4a
4b
ABNO
TEMPO
2
3
86 (ꢂ2)
64 (ꢂ1)
78 (ꢂ3)
58 (ꢂ1)
5a
5b
ABNO
TEMPO
2
3
98 (ꢂ1)
99 (ꢂ0)
96 (ꢂ2)
96 (ꢂ2)
6a
6b
ABNO
TEMPO
2
3
84 (ꢂ1)
57 (ꢂ1)
79 (ꢂ3)
49 (ꢂ3)
7a
7b
ABNO
TEMPO
2
3
46 (ꢂ2)
39 (ꢂ3)
2 (ꢂ0)
n. d.
a
40 mol of CuBr, 5 mM solution of NP1. b Average conversion (3 runs) aer work-up and isolation determined by 1H-NMR-spectroscopy. c Average
isolated yield (3 runs) aer purication.
Fig. 2 Recycling experiments in 5 consecutive runs with P1 (micellar catalytic approach) and NP1 (nanoparticle approach); conversions were
1
determined by H-NMR-spectroscopy.
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Table 6 Optimizing the reaction conditions for the one-pot two-step CALB/Cu-bipyridine reaction systema
Entry
Solvent
T (ꢁC)
t (h)
DIPEA (eq.)
Conversionb step 1 (%)
Conversionb step 2 (%)
1
2
H2O
PBS
PBS
PBS
PBS
rt
rt
rt
rt
40
2
2
24
4
4
0
0
0
1
1
81 (ꢂ2)
97 (ꢂ3)
97 (ꢂ3)
$99
4 (ꢂ1)
20 (ꢂ3)
26 (ꢂ2)
82 (ꢂ3)
96 (ꢂ1)
3
4c
5c
$99
a
First reaction step: CAL-B, 40 ꢁC, 20 min; second step: 5 mol% CuBr, 5 mol% NP1-ligand, 5 mol% ABNO, 10 mol% NMI. b Average conversion (3
1
c
runs) aer work-up and isolation determined by H-NMR-spectroscopy. DIPEA (1 eq.), 40 ꢁC, 20 min.
reaction with yields of 73 to 93% of the aldehydes. By using 20
Multi-step-catalysis
times the amount of catalysts (NP1, CALB) and reagents two
upscale experiments were carried out for the two substrates
benzyl acetate and p-methoxy benzyl acetate (Table 7, entry 6, 7).
The results for the isolated yields of the corresponding alde-
hydes were 93 and 95%, respectively and were in excellent
agreement with the previous reported product yields of the
small scale experiments (Table 7, entry 1, 2).
In the last set of experiments we studied the combination of the
enzymatic ester hydrolysis which resulted in the formation of
alcohols followed by the Cu(I)/bipyridine based oxidation of
these alcohols to their corresponding aldehydes. To realize such
a two-step-reaction, various aspects had to be considered.
In the initial experiments, we conducted the tandem reac-
tion as a one-pot two-step reaction and isolated the interme-
diary product benzyl alcohol. Aer 2 h at room temperature, we
obtained a 97% conversion in PBS buffer versus 81% in water,
clearly favouring the PBS buffer for the enzymatic step (Table 6,
entry 1–2). However, the second step, involving the aerobic
oxidation of benzyl alcohol to the corresponding benzaldehyde
gave only unsatisfactory conversion of 20–26% in PBS buffer,
even aer extending the reaction time from 2 h to 24 h (Table 6,
entry 3).
No catalyst inhibition was expected by the side product
acetic acid, due to its hydrophilicity which should prevent it
from being solubilized in the Cu(I)/bipyridine nanoparticle.
However, the acetic acid led to a shi in pH of the aqueous
medium to around 5 that was assumed to be the reason for the
signicantly lower Cu(I)/bipyridine activity. A possibility to
overcome the problem was to add a base such as N,N-diiso-
propyl ethylamine (DIPEA) to the aqueous medium aer the
ester hydrolysis and thus neutralize the acetic acid. Conse-
quently, the protocol for the one-pot two-step reaction was
changed and 1 eq. DIPEA was added aer the enzymatic
hydrolysis and allowed to stir for 20 min before the aerobic
oxidation for the benzyl alcohol was carried out in the presence
of NP1. The results obtained now showed quantitative conver-
sion of the ester hydrolysis step aer 20 min at 40 ꢁC and a 82 to
96% conversion for the aerobic oxidation step at r.t. and 40 ꢁC,
respectively (Table 6, entry 4, 5). In an attempt to assess the
scope and utility of this one-pot two-step system, a small series
of aromatic/allylic alcohols with various functional groups was
examined (Table 6). The results highlight the efficiency of the
consecutive enzymatic ester hydrolysis and aerobic oxidation
Conclusions
Herein, we presented the rst successful one-pot two-stage
reaction of an enzymatic ester hydrolysis followed by Cu(I)/
bipyridine catalysed aerobic oxidation of the alcohol interme-
diate to the corresponding aldehyde. The key to the successful
one-pot reaction in an aqueous medium was the site-isolation of
the Cu(I)/bipyridine catalyst in core-crosslinked nanoparticles
with a hydrophilic shell and a hydrophobic core which were
prepared by a microemulsion process. The nanoparticle design
efficiently prevents any interaction of the Cu/bipyridine catalyst
with the enzyme that has been shown to lead to catalyst inhi-
bition. Furthermore, it provides
a hydrophobic nano-
environment for the Cu/bipyridine system as a prerequisite
for catalyst activity in the aerobic oxidation. The nanoparticles
displayed no signicant loss in activity compared to the
micellar catalytic approach and could also be used in ve
consecutive runs still displaying a 80% substrate conversion in
the 5th run. The one-pot tandem reaction was nally conducted
as a two-step procedure, where the acetyl ester hydrolysis pro-
ceeded quantitatively aer 20 min at 40 ꢁC in the presence of the
enzyme CAL-B followed by addition of DIPEA to neutralize the
acetic acid side product. Then, the Cu(I)/bipyridine functional-
ized nanoparticle NP1 was added to the reaction mixture to
carry out the aerobic oxidation of the intermediate alcohols to
the corresponding aldehydes in high yields of 73–93% for
various aromatic and allylic acetic esters to their corresponding
aldehydes. We believe that this approach of metal-catalyst
compartmentalization within core–shell nanoparticles should
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Table 7 Tandem catalysis of representative acetyl esters to the corresponding aldehydesa
Entry
1
Substrate
Conversionb step 1 (%)
Conversionb step 2 (%)
Isolated yieldc (%)
$99
96 (ꢂ1)
93
2
$99
97 (ꢂ1)
93
3d
4d
5
$99
$99
$99
84 (ꢂ2)
78 (ꢂ4)
96 (ꢂ1)
80
73
91
6e
$99
$99
93 (ꢂ2)
96 (ꢂ1)
93
95
7e
a
Reactions were carried out in PBS-buffer (pH 7.4) at 40 ꢁC; (1) CAL-B, 20 min; (2) DIPEA (1 eq.) 20 min; (3) 5 mol% CuBr, 5 mol% NP1-ligand
(12.14 mg, 2 mmol, 0.0125 eq.), 5 mol% ABNO, 10 mol% NMI, 4 h. Average conversion (3 runs) aer work-up and isolation determined by 1H-
NMR-spectroscopy. Average isolated yield of the aldehyde (3 runs). 5 h reaction time lead to nearly quantitative conversion. Scale-up-
experiment: CuBr (86 mg, 600 mmol, 5 mol%), NP1-ligand (243 mg, 40 mmol, 5 mol%), ABNO (22.4 mg, 160 mmol, 5 mol%), NMI (25.6 ml, 320
mmol, 10 mol%), acetate (3.2 mmol, benzyl: 480.6 mg; 4-methoxybenzyl: 576.6 mg, 1 eq.), same reaction conditions as described in a.
b
c
d
e
ꢁ
also be of interest to other catalyst combinations to prevent measurements were performed on a Thermo Electro at 160 C
mutual catalyst inhibition on the one side and provide and 70 eV, using peruorokerosene as reference. Gel perme-
a hydrophobic reaction environment for the metal-catalyst in ation chromatography (SEC) was carried out on a DMF (5 g lꢀ1
aqueous medium.
LiBr) based SEC at 60 ꢁC with PSS GRAM analytical 1000 A and
30 A columns equipped with a Knauer RI detector Smartline
ꢁ
2300 using linear polystyrene standards for the poly(2-
oxazoline). Dynamic light scattering was carried out on a -
Malvern Zetasizer Nano-Z5 with a HeNe-laser (l ¼ 633 nm).
Fluorescence spectroscopy was recorded on a Hitachi F2700
Experimental
Measurements and materials
1H (500.13 MHz) and 13C NMR (100.63 MHz) spectra were
recorded on a Bruker DRX 500 spectrometer. GC-EI-HRMS
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using a wavelength of l ¼ 334 nm and a 0.1 mM solution of stirred for 12 h under reux. The solvent was removed under
pyrene in methanol. The analyses were carried out with the reduced pressure following which dichloromethane and water
program FLWinLab. TEM measurements were performed on was added to the residue. The aqueous phase was extracted with
a Philips CM200 using an Orius SC200 camera of Gatan. All dichloromethane (3 ꢃ 30 ml) and the organic layers were dried
commercially available chemicals were purchased from Sigma with magnesium sulfate. The solvent was removed under
Aldrich, TCI Chemicals, ABCR and Acros Organics and used reduced pressure and ethyl acetate was added to the residue.
without further purication. CALB immobilized on Immobead Aer ltration the solvent was removed under reduced pressure
150 was also purchased from Sigma Aldrich.
and the residue was puried via column chromatography
(cyclohexane/ethyl acetate 6/1). 4 (2.19 g, 12.72 mmol) was ob-
tained as light yellow oil in 98% yield.
Monomer synthesis
1H-NMR (500 MHz, CDCl3): d ¼ 2.49 ppm (t, J ¼ 2.2 Hz, CHC),
Synthesis of 2-heptyl-2-oxazoline (M2). The synthesis was 4.19 (d, J ¼ 2.4, CHCCH2), 4.63 (s, OCH2Ar), 5.27 (d, J ¼ 10.8 Hz,
carried out according to a modied procedure of Seeliger et al.45 CH2CH), 5.77 (d, J ¼ 17.6 Hz, CH2CH), 6.74 (dd, J ¼ 17.6 Hz/
starting from octanonitrile (24.57 ml, 154.94 mmol, 1.0 eq.). 2-n- 11.2 Hz, CH2CH), 7.31–7.45 (m, Ar). 13C-NMR (125 MHz, CDCl3):
Heptyl-2-oxazoline was obtained as a colorless liquid with d ¼ 56.9 (CH2CCH), 71.1 (OCH2Ar), 74.6 (CH2CCH), 79.5
a yield of 60%.
(CH2CCH), 113.9 (CH2CH), 126.2 (2 ꢃ CHCCH), 128.3 (2 ꢃ
1H-NMR (400 MHz, CDCl3): d ¼ 0.86 ppm (m, CH3), 1.30 (m, 4 CH2CCH), 136.4 (CH2CH), 136.7 (CH2C(Ar)), 137.2 (CHC(Ar)).
3
3
ꢃ CH2), 1.61 (quin, J ¼ 7.4 Hz, CH3CH2), 2.25 (t, J ¼ 7.7 Hz,
Synthesis of 2-(5-(1-(((4-vinylbenzyl)oxy)methyl)-1H-1,2,3-
3
3
CCH2), 3.80 (t, J ¼ 9.4 Hz, CH2N), 4.20 (t, J ¼ 9.5 Hz, CH2O). triazol-4-yl)pentyl)-2-oxazoline (M1). 2 (500 mg, 2.74 mmol, 1.0
13C-NMR (100 MHz, CDCl3): d ¼ 14.0 ppm (CH3), 22.6 (CH3CH2), eq.), 4 (519.82 mg, 3.02 mmol, 1.1 eq.) and diisopropylethyl-
25.9 (CH2CH2CO), 27.9 (CH2CH2CH2CH3), 28.9 (CH2CN), 29.2 amine (93.32 ml, 548.77 mmol, 0.2 eq.) were added to a mixture of
(CH2CH2CH2CN), 31.6 (CH2CH2CH3), 54.3 (CH2N), 67.1 (CH2O), tetrahydrofuran and water (1/1). Aer stirring for 5 min at room
168.6 (CO). HR-ESI-MS: Mcalculated ¼ 169.1467 [M ¼ C10H19NO]; temperature copper-sulfate-pentahydrate (34.25 mg, 137.19
Mmeasured ¼ 170.1538 [M + H]+.
mmol, 0.05 eq.) and sodium ascorbate (54.36 mg, 274.38 mmol,
Synthesis of (2-(5-chloropentyl)-2-oxazoline) (M3). The 0.1 eq.) were added to the mixture and it was stirred for 18 h at
synthesis was carried out according to Litt et al.46 Starting from room temperature. Following this, an excess of saturated EDTA-
3-caprolactone (10 g, 87.61 mmol, 1.0 eq.) M3 was obtained aer solution was added to stop the reaction. The water layer was
three steps as a colorless liquid with an overall yield of 66%.
extracted with dichloromethane (3 ꢃ 50 ml) and the organic
1H-NMR (400 MHz, CDCl3): d ¼ 1.47 pp (m, CH2CH2CH2Cl), layer was washed again with a saturated EDTA-solution. The
1.63 (quin, 3J ¼ 7.6 Hz, CH2CH2C(O)N), 1.76 (quin, 3J ¼ 7.2 Hz, organic layer was dried with magnesium sulfate and the solvent
3
3
CH2CH2O), 2.25 (t, J ¼ 7.5 Hz, CH2C(O)N), 3.5 (t, J ¼ 6.7 Hz, was removed under reduced pressure. Ethyl acetate was added
3
3
NCH2CH2O), 3.78 (t, J ¼ 9.7 Hz, CH2Cl), 4.19 (t, J ¼ 9.5 Hz, to the residue and the suspension was ltered. The solvent was
NCH2CH2O). 13C-NMR (100 MHz, CDCl3): d ¼ 25.1 ppm (CH2- removed under reduced pressure and the residue was treated
CH2C(O)N), 26.3 (CH2CH2CH2C(O)N), 27.6 (CH2CH2CH2Cl), with acetonitrile. Aer ltration the solvent was removed under
32.1 (CH2C(O)N), 44.7 (CH2N), 54.3 (CH2CH2CH2Cl), 67.1 reduced pressure M1 (931.7 mg, 2.63 mmol) was obtained as
(NCH2CH2O), 168.1 (C(N)O). HR-ESI-MS: Mcalculated ¼ 175.0764 yellow viscous oil in 96% yield (storage at 0 ꢁC resulted in a light
[M ¼ C8H14ClNO]; Mmeasured ¼ 176.0838 [M + H]+.
yellow solid).
Synthesis of 2-(5-azidopentyl)-2-oxazoline (2). The synthesis
1H-NMR (500 MHz, CDCl3): d ¼ 1.32–1.42 (m, CCH2CH2CH2),
was carried out according to Lav et al.38 Starting from 6-bro- 1.62–1.71 (m, CCH2CH2CH2CH2), 1.86–1.96 (m, CCH2CH2), 2.26
mohexanoic acid (25 g, 128.17 mmol, 1 eq.) 2 was obtained aer (t, J ¼ 7.48 Hz, CCH2CH2), 3.79 (t, J ¼ 9.46 Hz, OCH2CH2N), 4.18
four steps as a colorless liquid with an overall yield of 43%.
(td, J ¼ 9.42/1.71 Hz, OCH2CH2N), 4.28–4.38 (m, CH2NN), 4.58
1H-NMR (500 MHz, CDCl3): d ¼ 1.28–1.42 ppm (m, CCH2- (s, CHCCH2), 4.66 (s, OCH2Ar), 5.23 (d, J ¼ 10.8 Hz, CH2CH),
CH2CH2), 1.48–1.64 (m, CCH2CH2CH2CH2CH2N3), 2.20 (td, J ¼ 5.74 (dd, J ¼ 17.70, 0.61 Hz, CH2CH), 6.70 (dd, J ¼ 17.6 Hz/
7.46/1.22 Hz, CCH2), 3.19 (td, J ¼ 6.85/1.47 Hz, CH2N3), 3.66– 11.2 Hz, CH2CH), 7.28–7.42 (m, Ar), 7.53 (s, CHN). 13C-NMR (125
3.82 (m, CH2O), 4.14 (td, J ¼ 9.42/1.71 Hz, CH2N). 13C-NMR (125 MHz, CDCl3): d ¼ 25.1 ppm (CH2CH2C(O)N), 25.9 (CH2CH2-
MHz, CDCl3): d ¼ 25.3 ppm (CH2CH2C(O)N), 26.1 (CH2CH2- CH2C(O)N), 27.5 (CH2CH2CH2N), 29.9 (CH2C(O)N), 50.0 (CH2N),
CH2C(O)N), 27.6 (CH2CH2CH2N3), 28.4 (CH2C(O)N), 51.1 54.2 (CH2CH2CH2N), 63.6 (OCH2CN), 67.1 (NCH2CH2O), 72.2
(CH2N), 54.2 (CH2CH2CH2N3), 67.0 (NCH2CH2O), 168.1 (C(O)N). (OCH2Ar), 113.8 (CH2CH), 122.3 (CHN), 126.2 (2 ꢃ CHCCH),
HR-ESI-MS: Mcalculated ¼ 182.1168 [M ¼ C8H14N4O]; Mmeasured
¼
128.1 (2 ꢃ CH2CCH), 136.4 (CH2CH), 137.0 (CH2C(Ar)), 137.3
183.1246 [M + H]+.
(CHC(Ar)), 145.0 (OCH2CN), 168.0 (C(O)N). HR-ESI-MS:
Synthesis of 1-((prop-2-yn-1-yloxy)methyl)-4-vinylbenzene Mcalculated
¼
354.2056 [M
¼
C20H26N4O2]; Mmeasured
¼
(4). Sodium hydrate (786.19 mg, 19.66 mmol, 1.5 eq.) was 355.2136 [M + H]+.
added to a solution of propargylꢁalcohol (833.15 ml, 14.41 mmol,
Synthesis of 40-methoxy-[2,20-bipyridin]-4-ol (L1). The
1.1 eq.) in tetrahydrofuran at 0 C. The mixture was stirred for synthesis was carried out according to Weberskirch et al.29
45 min at room temperature. Aerwards 4-vinylbenzylchloride starting from 4,40-dimethoxy-2,20-bipyridine (500 mg,
(1.85 ml, 13.10 mmol, 1.0 eq.) and tetrabutylammonium iodide 2.31 mmol, 1.0 eq.). L1 was obtained as a white powder with
(2.42 g, 6.55 mmol, 0.5 eq.) were added and the mixture was 70% yield.
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1H-NMR (400 MHz, CDCl3): d ¼ 3.91 ppm (s, CH3O), 6.53 (dd, (m, 4 ꢃ CHAr(StyOx)), 7.49–7.65 (s(br), CHTriazol(StyOx)), 7.94 (s(br),
1
1J ¼ 7.2 Hz, CHCHCOCH3), 6.88 (dd, J ¼ 5.8 Hz, CHCHCOH), 2 ꢃ CH(BiPyOx)), 8.44 (s, 2 ꢃ CH(BiPyOx)).
1
7.11 (s, br, CCHCOH), 7.39 (s, CCHCOCH3), 7.71 (d, J ¼ 7 Hz,
P2: The synthesis of the precursor copolymer PP2 was similar
CHCHCOH), 8.42 (d, 1J ¼ 5.8 Hz, CHCHCOCH3). 13C-NMR (100 to the one described for PP1 starting with the addition of
MHz, CDCl3): d ¼ 55.5 ppm (CH3O), 106.1 (2 ꢃ CCHCOR), 111.2 methyltriate (37.99 ml, 335.72 mmol, 1 eq.) to 2-methyl-2-
(2 ꢃ CHCHCOR), 150.2 (2 ꢃ CHCHCOR), 167.0 (2 ꢃ NCCH, 2 ꢃ oxazoline (990.10 ml, 11.75 mmol, 35 eq.) in 10 ml acetonitrile
ꢁ
ꢁ
COR). HR-ESI-MS: Mcalculated ¼ 202.0742 [M ¼ C11H10N2O2]; at 0 C. Aer stirring the mixture for 3 h at 120 C, 2-heptyl-2-
Mmeasured ¼ 203.0816 [M + H]+.
oxazoline (295.97 ml, 1.68 mmol, 5 eq.) and (2-(5-chloropentyl)-
2-oxazoline) (294.85 mg, 1.68 mmol, 5 eq.) were added at
room temperature to form the second block for 24 h at 100 ꢁC.
Then 2-heptyl-2-oxazoline (295.97 ml, 1.68 mmol, 5 eq.) was
Polymer synthesis
All polymerizations were carried out in Schlenk tubes under added at room temperature and polymerization proceeded at
inert atmosphere using freshly distilled and dried solvents. A 100 ꢁC for another 24 h (3rd block) before M2 (356.99 mg,
typical procedure for P1 was as follows:
1.01 mmol, 3 eq.) was added at room temperature to form the
To 2-methyl-2-oxazoline (990.10 ml, 11.75 mmol, 35 eq.) in fourth block aer 18 h at 100 ꢁC. The polymerisation was
10 ml acetonitrile was given methyltriate (37.99 ml, 335.72 terminated at room temperature by addition of piperidine
mmol, 1 eq.) at 0 ꢁC. Aer stirring the mixture for 3 h at 120 ꢁC, 2- (36.54 ml, 369.28 mmol, 1.1 eq.) for 12 h. Aer precipitation in
heptyl-2-oxazoline (295.97 ml, 1.68 mmol, 5 eq.) and (2-(5- diethyl ether PP2 as a white powder. Coupling reaction with the
chloropentyl)-2-oxazoline) (294.85 mg, 1.68 mmol, 5 eq.) were bipyridine ligand L1 was carried out as described before by
added at room temperature. Aer stirring the mixture for 24 h at using (1 g, 1 eq. related to Cl-functionality) in 20 ml N,N-
ꢁ
100 C M2 (356.99 mg, 1.01 mmol, 3 eq.) was added at room dimethylformamide and addition of L1 (18.23 mg, 0.90 mmol,
temperature. The mixture was stirred for 18 h at 100 ꢁC and the 1.2 eq.) and K2CO3 (12.5 mg, 0.90 mmol, 1.2 eq.). Drying yielded
reaction terminated at room temperature by addition of piper- 0.87 g polymer P1 as a white powder in 86% yield. The 1H-NMR
idine (36.54 ml, 369.28 mmol, 1.1 eq.) for 12 h. Aer removal of (500 MHz, CDCl3) for P2 shows the same peak signals as P1 (see
the solvent, the residue was dissolved in 15 ml of chloroform above). Further analytical data of P1 and P2 are summarized in
and stirred with K2CO3 for 3 h. Aer ltration, PP1 was puried Table 1.
by precipitation in ice cold diethyl ether and dried under
reduced pressure.
Nanoparticle synthesis
1H-NMR (500 MHz, CDCl3): d ¼ 0.85 ppm (s(br), CH3(HepOx)),
1.26 (s(br), 4 ꢃ CH2(HepOx)), 1.45–1.90 (m, CH2(HepOx), 3 ꢃ
CH2(ClOx), 2 ꢃ CH2(StyOx), 3 ꢃ CH2(Pip)), 1.90–2.06 (s(br),
CH2(StyOx)), 2.06–2.19 (m, CH3(MeOx), 2 ꢃ CH2(Pip)), 2.19–2.50 (m,
CH2(HepOx), CH2(ClOx), CH2(StyOx)), 2.94/3.01 (CH3(In)), 3.19–3.75
(m, CH2–CH2(backbone), CH2(ClOx)), 4.33 (s(br), CH2(StyOx)), 4.58–
4.70 (m, 2 ꢃ CH2(StyOx)), 5.20–5.26 (m, CH(StyOx)), 5.69–5.79 (m,
CH(StyOx)), 6.64–6.74 (m, CH(StyOx)), 7.28–7.42 (m, 4 ꢃ CHAr(StyOx)),
7.49–7.65 (s(br), CHTriazol(StyOx)).
A typical procedure is described for NP1. P1 (110 mg, 0.02
mmol) was dissolved in 2 ml of milli-q-water (10 mM solution)
in
a test tube. 1,6-Hexanediol dimethacrylate (55.3 ml,
0.22 mmol, 50 wt%) and 1 ml heptadecane were added to the
solution. Aer adding a stem solution of azobis(isobutyroni-
trile) in dioxane (4 ml, 1.22 mmol, 0.18 wt%), the solution was
vented with argon for 30 min to remove oxygen. The tube was
sealed with a septum and kept in an ultrasonic bath for 5 min.
Aerwards the solution was heated for 24 h at 85 ꢁC. The
solution was centrifuged several times until no more solid was
precipitated. The water was evaporated via lyophilisation and
the residue was precipitated in ice cold diethyl ether. Drying
yielded 133 mg of nanoparticle NP1 as a white powder (yield:
81%). NP2 was prepared in a similar fashion by using P2
(140 mg, 0.02 mmol) in 2 ml of milli-q-water (10 mM solution)
in a test tube and 1,6-hexanediol dimethacrylate (70 ml,
0.28 mmol, 50 wt%). Drying yielded 175 mg of nanoparticle NP2
as a white powder (yield: 83%).
L1 (18.23 mg, 0.90 mmol, 1.2 eq.) and K2CO3 (12.5 mg,
0.90 mmol, 1.2 eq.) were added to a solution of PP1 (1 g, 1 eq.
related to Cl-functionality) in 20 ml N,N-dimethylformamide.
Aer stirring the mixture for 24 h at 100 ꢁC, the solvent was
removed. The residue was dissolved in dichloromethane,
ltered and the solvent was removed. The polymer was puried
by precipitation in ice cold diethyl ether and dialyzed against
ethanol (MWCO 1000) for 24 h. Aer removal of the solvent, the
polymer P1 was again precipitated in ice cold diethyl ether.
Drying yielded 0.83 g polymer P1 as a white powder in 82%
yield.
Micellar/nanoparticle catalysis
1H-NMR (500 MHz, CDCl3): d ¼ 0.85 ppm (s(br), CH3(HepOx)),
1.26 (s(br), 4 ꢃ CH2(HepOx)), 1.45–1.90 (m, CH2(HepOx), 3 ꢃ All catalytic oxidations wereꢁcarried out in open Schlenk tubes
CH2(BiPyOx), 2 ꢃ CH2(StyOx), 3 ꢃ CH2(Pip)), 1.90–2.06 (s(br), under air atmosphere at 20 C and constant stirring rates. For
CH2(StyOx)), 2.06–2.19 (m, CH3(MeOx), 2 ꢃ CH2(Pip)), 2.19–2.50 (m, preparation of the catalytically active polymer solution only
CH2(HepOx), CH2(BiPyOx), CH2(StyOx)), 2.94/3.01 (CH3(In)), 3.19–3.75 freshly distilled and dried acetonitrile was used. A typical
(m, CH2–CH2(backbone)), 3.93 (s(br), OCH3(BiPyOx)), 4.10 (s(br), procedure was as follows:
OCH2(BiPyOx)), 4.33 (s(br), CH2(StyOx)), 4.58–4.70 (m,
2
ꢃ
CuBr (4.30 mg, 30 mmol, 0.05 eq.) was added to polymer P1 or
CH2(StyOx)), 5.20–5.26 (m, CH(StyOx)), 5.69–5.79 (m, CH(StyOx)), nanoparticle NP1 (51.34 mg, 10 mmol, 0.017 eq.) in 2 ml
6.64–6.74 (m, CH(StyOx)), 6.82 (s(br), 2 ꢃ CH(BiPyOx)), 7.28–7.42 acetonitrile under an inert atmosphere. Aer stirring for 1 h at
33624 | RSC Adv., 2017, 7, 33614–33626
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room temperature, the solvent was removed and the polymer was dissolved in 20 ml PBS-buffer (1 mM solution). N-Methyl-
was dissolved in 2 ml water (5 mM solution). N-Methylimidazole imidazole (25.6 ml, 320 mmol, 0.1 eq.) and ABNO (22.4 mg, 160
(4.78 ml, 60 mmol, 0.1 eq.), N-oxyl-radical (ABNO 4.21 mg/TEMPO mmol, 0.05 eq.) were added to the solution and the mixture was
4.69 mg, 30 mmol, 0.05 eq.) and benzyl alcohol (62.38 mml, stirred at room temperature. Benzyl acetate (480.6 ml, 3.2 mmol,
0.6 mmol, 1.0 eq.) were added to the solution. The solution was 1.0 eq.) or 4-methoxybenzyl acetate (576.6 mg, 3.2 mmol, 1.0
stirred at room temperature with a constant rate in an open eq.) were added to 300 mg of the immobilized enzyme CALB
ask for 2 h (ABNO) or 3 h (TEMPO). To isolate the product, the (Immobead 150) in 20 ml of PBS-buffer. Aer stirring for 20 min
solution was extracted with diethyl ether (4 ꢃ 20 ml). The at 40 ꢁC N,N-diisopropyl ethylamine (557.4 ml, 3.2 mmol, 1.0 eq.)
organic layers were dried with MgSO4 and removed under was added to the solution and it was stirred for 20 min at 40 ꢁC.
reduced pressure. The product was puried via column chro- Aerwards the previously prepared nanoparticle solution was
matography (cyclohexane/ethyl acetate 5/1) and yielded with added and the mixture was stirred for 4 h at 40 ꢁC. To isolate the
94%. The catalyst solution in water could be used for further product, the aqueous solution was extracted with diethyl ether
runs by simply adding N-methylimidazole and N-oxyl radical (4 ꢃ 20 ml). The organic layers were dried with MgSO4 and
again.
removed under reduced pressure. The product was puried via
column chromatography (cyclohexane/ethyl acetate 5/1) and
yielded 93% benzaldehyde and 95% p-methoxy benzaldehyde.
Enzymatic ester cleavage
All enzymatic ester cleavages were carried out under air atmo-
sphere at 40 ꢁC and constant stirring rates. A typical procedure
was as follows: benzyl acetate (86.63 ml, 600 mmol) was added to
Acknowledgements
50 mg of the immobilized enzyme CALB (Immobead 150) in We thank Monika Meuris (Prof. J. C. Tiller, BCI, TU Dortmund)
ꢁ
1 ml of PBS-buffer. Aer stirring for 20 min at 40 C, the solu- for preparing the TEM-images. Furthermore, we thank the
tion was extracted with diethyl ether (3 ꢃ 10 ml). The organic working group of Prof. Tiller for providing some analytical
layers were dried with MgSO4 and the solvent removed under devices.
reduced pressure. The product was puried via column chro-
matography (cyclohexane/ethyl acetate 5/1). Benzyl alcohol
(60.3 mg, 557 mmol, 93%) was obtained as a colorless liquid.
Notes and references
1 L. F. Tietze, Chem. Rev., 1996, 96, 115–136.
2 J. C. Waslike, S. J. Obrey, R. T Baker and G. C. Bazan, Chem.
One-pot multi-step-reaction
All reactions were carried out in open Schlenk tubes under air
atmosphere at 40 C and constant stirring rates. For the prep-
aration of the catalytically active nanoparticle solution only
freshly distilled and dried acetonitrile was used. A typical
procedure is described below:
Rev., 2005, 105, 1001–1020.
ꢁ
3 Y. Hayashi, Chem. Sci., 2016, 7, 866–880.
4 C. A. Denard, J. F. Hartwig and H. Zhao, ACS Catal., 2013, 3,
2856–2864.
¨
5 H. Groger and W. Hummel, Curr. Opin. Chem. Biol., 2014, 19,
Small scale. CuBr (4.30 mg, 30 mmol, 0.05 eq.) was added to
nanoparticle NP1 (12.14 mg, 2 mmol, 0.0125 eq.) in 0.5 ml
acetonitrile under inert atmosphere. Aer stirring for 1 h at
room temperature, the solvent was removed and the polymer
was dissolved in 1 ml PBS-buffer (1 mM solution). N-Methyl-
imidazole (1.28 ml, 16 mmol, 0.1 eq.) and ABNO (1.12 mg, 8 mmol,
0.05 eq.) were added to the solution and the mixture was stirred
at room temperature. Benzyl acetate (22.6 ml, 160 mmol, 1.0 eq.)
was added to 15 mg of the immobilized enzyme CALB (Immo-
171–179.
6 P. M. Dinh, J. A Howarth, A. R Hudnott and J. M. J. Williams,
Tetrahedron Lett., 1997, 37, 7623–7626.
7 E. Burda, W. Hummel and H. Groger, Angew. Chem., Int. Ed.,
2008, 47, 9551–9554.
8 S. Borchert, E. Burda, J. Schatz, W. Hummel and H. Groger, J.
¨
¨
Mol. Catal. B: Enzym., 2012, 84, 89–93.
9 H. Sato, W. Hummel and H. Groger, Angew. Chem., Int. Ed.,
¨
2015, 54, 4488–4492.
bead 150) in 1 ml of PBS-buffer. Aer stirring for 20 min at 40 ꢁC 10 K. Tenbrink, M. Seßler, J. Schatz and H. Groger, Adv. Synth.
¨
N,N-diisopropyl ethylamine (27.87 ml, 160 mmol, 1.0 eq.) was
Catal., 2011, 353, 2363–2367.
added to the solution and it was stirred for 20 min at 40 ꢁC. 11 N. Rıos-Lombardıa, C. Vidal, E. Liardo, F. Morıs, J. Garcıa-
´
´
´
´
´
´
´
Aerwards the previously prepared nanoparticle solution was
added and the mixture was stirred for 4 h at 40 ꢁC. The product
Alvarez and J. Gonzalez-Sabın, Angew. Chem., Int. Ed., 2016,
55, 8691–8695.
´
´
was extracted with diethyl ether (3 ꢃ 20 ml). The organic layers 12 N. Rıos-Lombardı, C. Vidal, M. Cocina, F. Morıs, J. Garcıa-
´
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10937–10940.
scale and conversion and yield were determined by 1H-NMR- 13 J. S. Willemsen, R. P. Megens, G. Roelfes, J. C. M. van Hest
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10 ml acetonitrile under inert atmosphere. Aer stirring for 1 h 15 M. Heidlindemann, G. Rulli, A. Berkessel, W. Hummel and
and F. P. J. T. Rutjes, Eur. J. Org. Chem., 2014, 14, 2892–2898.
¨
H. Groger, ACS Catal., 2014, 4, 1099–1103.
at room temperature, the solvent was removed and the polymer
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