Metathesis in water conducted by tailor-made encapsulated Grubbs' catalyst

Article abstract of DOI:10.1039/c8gc02164c

Metathesis in water represents a current challenge in green chemistry, since hydrophobic catalysts are non-soluble in this medium. Although this issue has been addressed by modification of the hydrophobic ligand structure, alternative methods for conducting metathesis in water are of interest, such as catalyst encapsulation. In this contribution we report successful encapsulation of the Grubbs' second-generation catalyst in alginate hydrogels, representing a renewable resource, to perform ring-closing metathesis (RCM) in water. We initially investigated the influence of different solvents on the reaction rate and confirmed that water is a preferred solvent. A comparison of non-encapsulated and encapsulated catalyst in calcium alginate revealed that the reaction rate for non-encapsulated catalyst was notably higher, which can be explained by "on water" conditions in this case. Inside the beads the encapsulated catalyst remained heterogeneous, thus allowing to switch the catalysis mode between "in/on water" through encapsulation. To overcome diffusion limitation and enhance reaction rate, we prepared a tailor-made bead material by introducing hydrophobic octyl-grafted alginate amide. Using such a hydrogel, diffusion limitation was positively influenced by hydrophobisation of the matrix, resulting in up to quadrupled reaction rates compared to calcium alginate as a standard encapsulation material. In terms of recycling, this encapsulated catalyst revealed promising performance, retaining 80% of its activity after ten runs. This is the first reported application of hydrophobised alginate derivatives in catalysed organic synthesis, achieving excellent encapsulation efficiency, no measurable leaching and yields of up to 87%.

Full text of DOI:10.1039/c8gc02164c

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Metathesis in water conducted by tailor-made
encapsulated Grubbs’ catalyst†
Jan Pauly, a,b Harald Gröger b and Anant V. Patel
Cite this: Green Chem., 2018, 20,
5179
a
*
Metathesis in water represents a current challenge in green chemistry, since hydrophobic catalysts are
non-soluble in this medium. Although this issue has been addressed by modification of the hydrophobic
ligand structure, alternative methods for conducting metathesis in water are of interest, such as catalyst
encapsulation. In this contribution we report successful encapsulation of the Grubbs’ second-generation
catalyst in alginate hydrogels, representing a renewable resource, to perform ring-closing metathesis
(RCM) in water. We initially investigated the influence of different solvents on the reaction rate and
confirmed that water is a preferred solvent. A comparison of non-encapsulated and encapsulated catalyst
in calcium alginate revealed that the reaction rate for non-encapsulated catalyst was notably higher,
which can be explained by “on water” conditions in this case. Inside the beads the encapsulated catalyst
remained heterogeneous, thus allowing to switch the catalysis mode between “in/on water” through
encapsulation. To overcome diffusion limitation and enhance reaction rate, we prepared a tailor-made
bead material by introducing hydrophobic octyl-grafted alginate amide. Using such a hydrogel, diffusion
limitation was positively influenced by hydrophobisation of the matrix, resulting in up to quadrupled reac-
tion rates compared to calcium alginate as a standard encapsulation material. In terms of recycling, this
encapsulated catalyst revealed promising performance, retaining 80% of its activity after ten runs. This is
the first reported application of hydrophobised alginate derivatives in catalysed organic synthesis, achiev-
ing excellent encapsulation efficiency, no measurable leaching and yields of up to 87%.
Received 12th July 2018,
Accepted 16th October 2018
DOI: 10.1039/c8gc02164c
rsc.li/greenchem
catalysis by heterogenisation methods, such as encapsulation,
is highly desirable to enable industrial application of these
Introduction
In industrial chemistry, more than 80% of chemical products types of reactions. Encapsulation represents one of such
are generated in processes requiring catalysts. Syntheses for methods and results in stable, separable and selective hetero-
bulk products are often catalysed heterogeneously, whereas geneous catalysts.1
homogeneous catalysis remains to be a sub-branch. Although
Besides developing new catalysts, new catalytic processes
some bulk processes such as olefin polymerisation are per- featuring both bio- and chemocatalysis conducted either
formed by homogeneous catalysis, for many processes the sequentially or in tandem, have been gaining increasing atten-
advantages of heterogeneous catalysts are required. The latter tion in recent years.2 Combinations of metal catalysts and
are often highly temperature stable, easy to handle and suit- enzymes as well as the use of organocatalysts along with
able for application in both batch and continuous processes. enzymes have been reported.3 Despite numerous reports on
Therein, they can be fixed in packed bed reactors or used in chemoenzymatic syntheses, there are few on using immobi-
fluidised-bed-reactors to separate the catalyst from the lised catalysts.4 Besides the abovementioned advantages of
product. However, selectivities are often limited and certain heterogenisation, encapsulation can contribute to compart-
reaction types are restricted to homogeneous catalysis. mentation of different catalysts.5 Therefore, developing a suit-
Consequently, eliminating the disadvantages of homogeneous able immobilisate for the chemocatalyst as well as for the
enzyme is required. An important reaction type is olefin meta-
thesis that has already been successfully applied in chemo-
aFermentation and Formulation of Biologicals and Chemicals, Faculty of Engineering
and Mathematics, Bielefeld University of Applied Sciences, Interaktion 1,
33619 Bielefeld, Germany. E-mail: anant.patel@fh-bielefeld.de
bChair of Organic Chemistry I, Faculty of Chemistry, Bielefeld University,
Universitätsstrasse 25, 33615 Bielefeld, Germany
enzymatic synthesis before. For example, in a previous report
pig liver esterase was combined with Grubbs’ second-generation
catalyst to create cyclic malonic acid monoesters in aqueous
conditions.6 While we have already reported the encapsulation
of pig liver esterase in biopolymer hydrogels,7 encapsulation of
Grubbs-type catalysts in such hydrogels has not been reported
†Electronic supplementary information (ESI) available. See DOI: 10.1039/
c8gc02164c
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until now. Grubbs’ first-generation catalyst is very oxygen- the experiments with 0.5 mol% catalyst and 20 mM substrate
sensitive, whereas Grubbs’ second-generation catalyst is stable concentration at room temperature. After we started the reac-
in oxygen and both catalysts barely react with water, thus being tions, we took samples in defined time periods and deter-
stable in aqueous medium.8 Accordingly, several applications mined the conversions via HPLC analysis. These data were
of Grubbs’ catalysts in aqueous media have been reported to used to calculate initial reaction rates within the first
meet the requirements of green chemistry.9–16 There are 60 minutes. This procedure determining (initial) reaction rates
different strategies to perform olefin metathesis reaction in was applied for all following experiments as well. For all experi-
water: On the one hand catalysts are used homogeneously ments in the entire study, high conversions (≥90%) were
without immobilisation, e.g., commercial Grubbs’ catalysts or measured.
new water-soluble catalysts via ligand design.10–17 Limitations
We demonstrated that DCM remained the best solvent for
of this approach exist in terms of complex and effortful ligand our model reaction, providing a reaction rate of 3.58
synthesis, catalyst separation and recycling. These challenges 0.17 µmol min−1, while water still maintained 74% of this
have been addressed by heterogenisation. For example, among activity (Fig. 1). When we added tBuOH to water, which has
miscellaneous techniques,18 Grubbs-type catalysts have been been used in a previous study for chemoenzymatic one-pot
immobilised on and in different supports such as inorganic reactions,6 the reaction rate dropped drastically to 0.456
silica or alumina,19 and synthetic polymers including silicone 0.054 µmol min−1. This drop was further increased by using
matrices.20 In other approaches, supported ionic liquid phase pure tBuOH, indicating that tBuOH is not suitable for this
(SILP) catalysts have been designed.21–23 Those heterogenisa- RCM. Even acetone–water (2 : 1), which was reported to act as
tion techniques, however, typically share one major dis- suitable solvent for RCM in water, however at a higher catalyst
advantage: The matrices are not perfectly designed to be loading of 3%, was not suitable for our reaction.10
applied in water, since they provide either hydrophobic pro-
As this solvent study revealed that water acts as highly suit-
perties or consist of ionic liquids that could, at least in part, be able solvent for the RCM of substrate 1, this experiment in
miscible with water, thus making the use of water as solvent aqueous medium built the base and set the benchmark for the
difficult.
In this study, we addressed these challenges by designing
encapsulation of Grubbs’ catalyst in biopolymer hydrogels.
Influence of encapsulation and substrate
tailor-made hydrogel beads for the encapsulation of meta-
thesis catalysts; using the biopolymer calcium alginate as
renewable resource, we further addressed the demands of
green chemistry. Although biopolymer hydrogels have been
used as support for ionic liquids in SILP catalysts,22,23 they
have not been used for immobilisation of metathesis catalysts
until now. We encapsulated Grubbs’ second-generation cata-
lyst in biopolymer hydrogels to perform ring-closing meta-
thesis (RCM) and evaluate reaction rates, catalyst leaching,
encapsulation efficiency and recyclability. Furthermore, we
gained a deeper understanding of how catalyst, matrix, sub-
strate and product interact and influence the reaction, in par-
ticular via diffusion limitation. Besides encapsulation, since
RCM with non-modified Grubbs’ catalysts in water has been
reported,9–12 we wanted to further investigate whether the reac-
tion runs “in”, or “on” water.
With this efficient model reaction in water in hand, we evalu-
ated how encapsulating the catalyst affects the reaction rate.
We expected that diffusion limitation and hydrophobicity of
the substrate lead to significantly lower reaction rates than the
ones when using a non-encapsulated catalyst. As matrix for
our catalyst we chose calcium alginate, which has been applied
widely for the encapsulation of biocatalysts.25 After adding the
Results and discussion
Solvent influence on non-encapsulated catalyst
In our initial study, we evaluated the influence of different sol-
vents on the reaction rate of the non-encapsulated Grubbs’
catalyst by performing the RCM of diethyl diallylmalonate (1)
as model substrate. Since RCM reactions are usually per-
formed in dichloromethane (DCM),24 we wanted to clarify
whether the reaction rate is influenced by the choice of solvent
and whether the reaction can be performed efficiently in
water. Besides these two solvents, we also evaluated two
solvent mixtures with different ratios, having already been
Fig. 1 Solvents influenced the reaction rate of RCM with non-encapsu-
lated catalyst (means standard deviations; n = 3; different letters (a–d)
above bars indicate significant differences according to ANOVA with
used successfully for metathesis reactions before.6,10 We ran Tukey’s post hoc test at p < 0.05).
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catalyst to sodium alginate solution, it was mixed and thinly dis- using the encapsulated catalyst. With substrate 3 showing a
persed with a spatula. Dropping this mixture into calcium chlor- higher solubility in water and, thus, better diffusion into the
ide led to uniform beads with a diameter of 2.68 0.17 mm. alginate matrix containing the catalyst, a higher reaction rate
After the reaction, these beads shrunk minimally to 2.46
of 0.095 0.035 µmol min−1 was found. Besides water solubi-
0.13 mm. To evaluate catalyst retention during the encapsula- lity as one factor influencing the reaction rate, intrinsic reactiv-
tion process (encapsulation efficiency) and after the reaction ity differences of the two different substrate types have to be
(leaching), we extracted the encapsulation solution and the reac- considered as well. As without encapsulation the reaction
tion solution. Catalyst content in solution was determined by (with high reaction rates) occurs via an “on water” mode, the
spectrophotometric measurement. Every experiment revealed method “encapsulation” is complementary leading to a switch
excellent encapsulation efficiency and no measurable leaching. to an “in/on water” fashion, in which water-solubility of the
Reaction conditions were the same as for the reaction with non- substrate now plays an important role for the mass transport
encapsulated catalyst to enable comparability.
into the beads.
Performing the RCM with substrate 1 and encapsulated
catalyst revealed an extremely low reaction rate of 0.047
0.002 µmol min−1, in fact the reaction rate with non-encapsu-
lated catalyst was 56 times higher (Fig. 2). This high difference
may be explained by two effects. In the case of non-encapsula-
tion, the catalyst dissolves in poorly water-soluble substrate 1,
leading to very high reaction rates. In this scenario, the reac-
tion runs “on water”, corresponding to a reaction under neat
conditions instead of a reaction running “in water” with a dis-
solved substrate. The other effect is caused by encapsulation of
the catalyst, requiring that the substrate dissolves in water and
diffuses through the hydrogel to the catalyst. In addition, in
this scenario product diffusion has to be considered as well.
Consequently, the high reaction rates with non-encapsulated
catalyst cannot be achieved with an encapsulated catalyst for
this substrate.
To support these hypotheses, we introduced two new sub-
strates in our study. On the one hand, we chose diallyltrifluoro-
acetamide (3) as a more water-soluble substrate and 1,6-hepta-
diene (5) as an almost water-immiscible substrate. After we
conducted the experiments with substrate 3, the difference
between reaction rates for non-encapsulated and encapsulated
catalyst was remarkably reduced (ten-fold factor). In contrast,
when using substrate 5, no activity was measured for the
encapsulated catalyst. These results indicate that the sub-
strate’s water solubility is essential for the RCM in water when
Solvent influence on encapsulated catalyst
For the non-encapsulated catalyst, we demonstrated that water
acts as more beneficial solvent compared to tBuOH, acetone
and methanol, which lead to lower reaction rates. Since hydro-
phobic substrate diffusion into hydrophilic alginate hydrogel
turned out to be limited (as described above), we examined
how these solvents affect the reaction rate with the encapsu-
lated catalysts and in particular if the diffusion limitation of
substrate 1 is reduced when using an organic solvent com-
ponent (Fig. 3). The experiments revealed that all mixtures led
to decreased reaction rates compared to water. Since the
results of Fig. 1 agree with this, pure water, among those sol-
vents tested, remains to act as optimal solvent for our appli-
cation. However, for the encapsulated catalyst the difference
especially between water and acetone–water was notably
decreased (Fig. 3) compared to the non-encapsulated catalyst
(Fig. 1). We suggest that this effect is due to enhanced sub-
strate and product diffusion. Despite methanol–water has suc-
cessfully been introduced as solvent for ligand-modified water-
soluble RCM catalysts,13 here this solvent inhibited the
reaction.
Next, we investigated whether an emulsifier (Tween® 20)
increases the reaction rate by highly dispersing the substrate.
However, the reaction rate dropped significantly to 0.006
0.002 µmol min−1, which is even below the rate in acetone–
Fig. 2 Encapsulating the catalyst in calcium alginate and selecting the substrate influenced the reaction rate of RCM (means standard deviations;
n = 3; different letters (a, b) above bars indicate significant differences according to ANOVA with Tukey’s post hoc test at p < 0.05).
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to calcium alginate, it has been concluded that the grafted
hydrophobic carbon chain improved the hydrophobicity. In
the same study it has been shown that crosslinking at the shell
is occurring at a high density while crosslinking in the core is
lower, and thus lead to softer bead material structure inside,
possibly enhancing diffusion properties. Therefore, we
regarded this material to be promising for Grubbs’ catalyst
encapsulation. Moreover, it has been applied for the release of
hydrophobic substances in medical use before.29–32 This
derivative is partially hydrophobised (27% degree of amida-
tion),29 retaining its capability to form a hydrogel, which is
necessary for applying it in water. Since there has been no
report using alginate amide for the encapsulation of catalysts
until now, we investigated the encapsulation of Grubbs’ cata-
lyst in this material to reduce substrate and product diffusion
limitation for our system (Fig. 4). Therefore, we synthesised
octyl-grafted alginate amide adapting a reported procedure.29
After we created alginate amide beads with calcium chloride,
we found that they were not perfect spherically shaped like
alginate beads. Nevertheless, the bead diameter was uniform
with 2.59 0.27 mm. Also, the bead size after reaction was
Fig. 3 Solvents influenced the reaction rate of RCM with catalyst
encapsulated in calcium alginate (means standard deviations; n = 3;
different letters (a–e) above bars indicate significant differences accord-
ing to ANOVA with Games-Howell post hoc test at p < 0.05).
similar to calcium alginate beads with 2.41
0.23 mm. In
accordance to the results with calcium alginate, no leaching
after reaction was detected when using alginate amide.
water, even though we observed that the substrate was highly
dispersed. Revealing that the emulsifier had no beneficial
effect, the results suggest that the diffusion limitation through
a hydrophilic network of the beads might be the limiting step
which has to be addressed.
Therefore, in consideration of these results, we had to look
for other options to enhance diffusion and by thus, reaction
rate.
Influence of the bead material and design of a tailor-made
bead
To overcome diffusion limitation and, thus, to increase reac-
tion rates, we focused on the design of an alternative bead
material, since the used substrates are quite hydrophobic.
Although various hydrophobic synthetic polymers are avail-
able, they are not suitable for using them in water due to their
hydrophobicity. We assume that in contrast to the substrate,
easily diffusing into the hydrophobic matrix, formed product
would stay inside the beads, avoiding the hydrophilic environ-
ment outside. Since we wanted to use water as solvent and bio-
polymer hydrogels, we chose the pathway of modifying the
bead material. One promising approach is the hydrophobic
modification of alginate, which has been reported in literature
before.26 It is possible to modify the hydroxyl or the carboxyl
groups to create materials with altered properties. We focused
on the modification of the carboxylic group by amidation,
since those derivatives have been well characterised, possess
hydrophobic properties and are available by straightforward
syntheses.27–31 As representative for hydrophobic alginate
derivatives, we chose octyl-grafted alginate amide. This algi-
nate derivative has been tested for hydrophobicity before,
using the wettability as parameter.28 Measuring a notably
Fig. 4 Top: Hydrophilic calcium alginate limits diffusion (thin arrows) of
hydrophobic substrate and product inside beads. Bottom: More hydro-
phobic octyl-grafted alginate amide enhances diffusion (thick arrows) of
higher contact angle of octyl-grafted alginate amide compared hydrophobic substrate and product.
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When we started the reaction with substrate
1
and summary, catalyst encapsulated in alginate amide provided
remarkably higher increases in reaction rates along with raised
0.5 mol% catalyst loading, the reaction rate doubled to 0.108
0.041 µmol min−1, compared to calcium alginate (Fig. 5a). We catalyst loading (Fig. 5c).
suggest that enhanced hydrophobicity led to this improve-
Subsequently, we evaluated conversions and yields for pro-
ment, though this did not elucidate whether diffusional or ducts of reactions with 2.5 mol% catalyst loading in alginate
kinetic limitation inside the beads took place. Therefore, we amide. Conversions of 92% (for product 2) and 99% (for
raised the catalyst loading to 2.5 and 5.0 mol% and evaluated product 4), respectively, were measured by means of HPLC
the differences in reaction rates between calcium alginate and analysis after 24 h reaction time. After we purified the raw pro-
alginate amide. Once the reaction rate cannot be enhanced by ducts by column chromatography, we characterised them by
raised catalyst loading, this would prove diffusion limitation. means of NMR analysis achieving yields of 87% (for 2) and
Following this, diffusion limitation occurred at 2.5 mol% cata- 82% (for 4), respectively (Fig. 6).
lyst for calcium alginate. In contrast, for alginate amide the
reaction rate was still enhanced with 5.0 mol% catalyst, indi-
cating that the diffusion limitation was strongly reduced when
using this modified and more hydrophobic alginate matrix.
We repeated the experimental series with the more hydro-
philic substrate 3 and found out that with this substrate
diffusion limitation already occurred at 0.5 mol% catalyst for
calcium alginate (Fig. 5b). For alginate amide, diffusion limit-
ation set in at 2.5 mol% catalyst, and reaction rate was quad-
rupled. Although we revealed that diffusion limitation was
lowered using alginate amide, this appears to be not the single
reason for the enhanced reaction rates. While no diffusion
limitation for substrate 1 at 0.5 mol% catalyst for both
matrices occurred, a difference between reaction rates was
demonstrated. This result indicates that hydrophobisation of
alginate influences the reaction rate, possibly by providing a
hydrophobic environment for the catalyst besides enhancing
Fig. 6 Preparative runs with catalyst (2.5 mol%) encapsulated in algi-
nate amide gave very good yields for 2 and 4. Products were cleaned up
diffusion, which should be addressed in further studies. In
via column chromatography.
Fig. 5 (a) Alginate amide for catalyst encapsulation enhanced reaction rates for substrate 1; for calcium alginate, diffusion limitation occurred at
2.5 mol% cat. loading. (b) Alginate amide for catalyst encapsulation enhanced reaction rates for substrate 3; for calcium alginate, diffusion limitation
occurred at 0.5 mol% cat. loading, whereas for alginate amide, diffusion limitation occurred at 2.5 mol% cat. loading. (c) In contrast to calcium algi-
nate, alginate amide strongly enhanced reaction rates with increased cat. loading for both substrates (means standard deviations; n = 3; different
letters (a, b) above bars indicate significant differences according to ANOVA with Tukey’s post hoc test at p < 0.05).
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In summary, the highest reaction rates for substrate 1 of phine ligand, since this issue has already been discussed and
0.502 0.011 µmol min−1 and 0.398 0.096 µmol min−1 for reported before.8 Without this interim storage, the drop of
substrate 3 were measured. We showed that the matrix miti- activity would probably be displayed more continuously during
gates internal diffusion limitation, in this instance by encapsu- the runs. However, activity remained stable at approximately
lating Grubbs’ catalyst in hydrophobic alginate amide. This led 80% after storage. For each recycling run, turn over numbers
to doubled up to quadrupled reaction rates and built a base (TON) from 36 to 39 were achieved. For all runs together, an
for further research on reasonable application of encapsulated overall TON of 372 was reached. In summary, the encapsulated
Grubbs’ catalyst in water.
catalyst revealed promising activity and stability.
Recycling study
Conclusions
To compete with its non-encapsulated analogue, the immobi-
lised catalyst has to show explicit advantages. In consequence
of its observed significantly lowered reaction rate, recycling the
catalyst for several runs is necessary to compensate this dis-
advantage. Recycling requires long-term stability and prefer-
ably minor activity losses for an extended period. To investi-
gate these recycling properties, we encapsulated 2.5 mol%
Grubbs’ catalyst in alginate amide and performed ten sub-
sequent runs with substrate 1 (Fig. 7). To provide sufficient
substrate conversion, we ran each cycle for 24 hours, reaching
≥90% conversion, before starting the next one. To avoid mis-
leading data, either through substrate from a previous run con-
verted in the subsequent run or product from previous run
leaching out of the beads during the subsequent run, we
washed the beads carefully with an excess of water prior to the
recycling. After we performed the first six runs straight in suc-
cession, we stored the beads for three days in the fridge,
before the last four runs were started. We could demonstrate
that the activity remained constant over the first six runs with
no decrease. For the last four runs, we observed a slight
reduction in activity; we suggest that above-mentioned storage
of the beads led to this effect. The key reason for the decrease
in activity in cycle seven might be the dissociation of the phos-
In summary, we reported the first encapsulation of Grubbs’
second-generation catalyst in alginate hydrogels to perform
ring-closing metathesis in water. Using diethyl diallylmalonate
as model substrate, we investigated the influence of different
solvents on the reaction rate and demonstrated that water
serves as a preferred solvent. A comparison of non-encapsu-
lated and encapsulated catalyst in calcium alginate revealed
that reaction rate for non-encapsulated catalyst was remarkably
higher due to “on water” conditions. At the same time, it
turned out that the method “encapsulation” is complementary
and acts as a switch for “in/on water” reaction conditions.
Since water remained the best solvent for encapsulated cata-
lyst, we modified the bead material to overcome diffusion
limitation and enhance the reaction rate. By introducing
hydrophobic octyl-grafted alginate amide as tailor-made bead
material, we showed that diffusion limitation was positively
influenced by hydrophobisation of the matrix, resulting in a
maximum of four-fold increased reaction rate for alginate
amide. Finally, the encapsulated catalyst showed promising
recyclability and could be re-used for in total ten runs showing
no measurable leaching. Nevertheless, the remaining activity
after encapsulation and number of recycling steps should be
increased to be attractive for broad applicability. This is the
first reported application of hydrophobised alginate derivatives
in catalysed organic synthesis. With all its features, this encap-
sulation method is interesting for further optimisation of
metathesis catalysts as well as in terms of a generally appli-
cable technology platform for encapsulation of chemocatalysts
for organic synthesis in aqueous media. Furthermore, the
reported insights will contribute to better understanding and
application of encapsulated catalysts in biopolymer hydrogels
for organic syntheses.
Experimental
General
All commercially available chemicals were used without
further purification. Substrate 1 was obtained from Alfa Aesar,
substrate 3 was obtained from Sigma-Aldrich, substrate 5 and
Grubbs’ second-generation catalyst were obtained from abcr
GmbH. Octyl-grafted alginate amide was prepared according to
Fig. 7 Recycled beads (alginate amide; 2.5 mol% cat. loading) provided
a
reported procedure,29 and described in the ESI.†
constant reaction rates for the first six runs (means
ations; n = 3).
standard devi-
Determination of conversions for the calculation of the initial
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reaction rates was done by means of HPLC on a Macherey- Spectrophotometric analysis
Nagel NUCLEODUR® C18 Gravity-SB (5 µm) column. Further
The term “encapsulation efficiency” (in %) describes the ratio
of the amount of the catalyst in beads compared to the
amount that was used in the biopolymer solution prior to the
preparation of the beads. The term “leaching” (in %) describes
the ratio of the amount of leached enzyme after the reaction
compared to the amount that was encapsulated. The sample,
collected after encapsulation or reaction, was evaporated, then
solved in 5 ml dichloromethane and absorbance was measured
at 387 nm. Recording a calibration curve with standard con-
centrations revealed the amount of Grubbs’ catalyst in the
sample.
information on HPLC analysis is given in the ESI.†
Quantification of the catalyst content for calculation of the
encapsulation efficiencies and leaching were done by spectro-
photometric measurement with a Thermo Scientific Genesys
10S UV-Vis photometer. 1H and 13C NMR spectra were
recorded with a Bruker Avance III 500 HD (measuring fre-
quency: 1H NMR = 500.2 MHz, 13C NMR = 125.8 MHz)
spectrometer in CDCl3 or D2O solution. Chemical shifts are
referenced to the residual peaks of the solvent [CDCl3:
7.26 ppm (1H NMR), 77.16 ppm (13C NMR)].33 Diameters of
ten randomly chosen beads per experiment were measured
with a calliper ruler and the data were pooled for statistical
treatment. Each experiment was repeated three times for stat-
Analytical data of diethyl cyclopent-3-ene-1,1-dicarboxylate (2)
istical analyses, which were carried out with the software Synthesis of 2 was carried out according to ring-closing meta-
SPSS Statistics V22.0. Values are given as mean standard thesis reaction procedure with 6.25 µmol catalyst encapsulated
deviation. Means were tested for significant differences by in alginate amide and substrate 1. After 24 h reaction time, the
analysis of variances (ANOVA) followed by Tukey’s or Games- reaction mixture was extracted with CH2Cl2 (3 × 30 ml). The
Howell post hoc test at p < 0.05.
combined organic layers were dried over MgSO4, filtered and
the solvent was evaporated in vacuo. Product 2 was purified by
column chromatography on silica gel (n-hexane/ethyl acetate
9 : 1, Rf = 0.53) and provided as pale yellow liquid (46 mg; 87%
yield).
Preparation of calcium alginate and alginate amide beads
Grubbs’ second-generation catalyst (1.25, 6.25 or 12.5 µmol;
1.06, 5.31 or 10.61 mg) was immobilised using the method of
encapsulation. After the catalyst was suspended in sodium
alginate or alginate amide solution (2.5 ml; 2% w/w in H2O),
catalyst particle aggregates were crushed with a spatula to
reduce the particle size. Further details on particle size are
given in the ESI.† To enhance homogeneous distribution of
catalyst particles in the hydrogel, the suspension was stirred at
500 rpm for 10 min at room temperature. Then, the solution
was dropped in CaCl2 solution (30 ml; 0.2 M) through a hypo-
dermic needle (0.90 mm external diameter; 40 mm length).
The beads were stirred to cross-link for 10 min before they
were washed for 10 min with H2O to remove excess CaCl2. For
determining encapsulation efficiency, the CaCl2 solution was
extracted with dichloromethane (3 × 30 ml) and spectrophoto-
metric analysis was performed.
3
1H-NMR (500.2 MHz, CDCl3, 296 K): δ = 1.24 (t, JH,H = 7.1
Hz, 6H, 2 × CH2CH3), 3.00 (s, 4H, CH2–CHvCH–CH2), 4.18 (q,
3JH,H = 7.1 Hz, 4H, 2 × CH2CH3), 5.59 (s, 2H, CH2–CHvCH–
CH2) ppm.
13C-NMR (125.8 MHz, CDCl3, 296 K): δ = 14.2 (CH2CH3),
41.0 (CH2–CHvCH–CH2), 59.0 (CH2CCH2), 61.6 (CH2CH3),
127.9 (CH2–CHvCH–CH2), 172.4 (COOCH2) ppm.
The analytical data are in accordance to those reported.34
Analytical data of 1-(trifluoroacetyl)-2,5-dihydro-1H-pyrrole (4)
Synthesis of 4 was carried out according to ring-closing
metathesis reaction procedure with 6.25 µmol catalyst
encapsulated in alginate amide and substrate 3. After 24 h
reaction time, the reaction mixture was extracted with
CH2Cl2 (3 × 30 ml). The combined organic layers were dried
over MgSO4, filtered and the solvent was evaporated in
vacuo. Product 4 was purified by column chromatography
on silica gel (ethyl acetate, Rf = 0.29) and provided as brown
liquid (34 mg; 82% yield).
Ring-closing metathesis reaction procedure
In a 15 ml glass beaker, a batch of beads (2.5 g) or non-encap-
sulated catalyst (1.25, 6.25 or 12.5 µmol; 1.06, 5.31 or
10.61 mg) was suspended in 12.5 ml solvent and crushed with
a spatula. The reaction was started by adding 0.25 mmol of
either diethyl diallylmalonate (substrate 1; 60 mg), N,N-diallyl-
2,2,2-trifluoroacetamide (substrate 3; 48 mg) or 1,6-heptadiene
(substrate 5; 24 mg) under stirring at room temperature. For
determining conversion, samples were taken at specific times
(5, 10, 20, 40, 60, 120, 240, 360, 1440 min) and analysed via
HPLC. For recycling experiments, the beads were washed with
water for 30 min before the subsequent run to remove
unreacted substrate and product within the beads. For deter-
mining leaching, the reaction solution was extracted with di-
chloromethane (3 × 30 ml) and spectrophotometric analysis
was performed.
1H-NMR (500.2 MHz, CDCl3, 296 K): δ = 4.35–4.46 (m, 4H,
2 × CHCH2), 5.82–5.90 (m, 2H, CHvCH) ppm.
4
13C-NMR (125.8 MHz, CDCl3, 296 K): δ = 53.2 (q, JC,F
=
1
3.8 Hz, CHCH2), 54.8 (CHCH2), 116.4 (q, JC,F = 287.3 Hz,
2
CF3), 124.8, 125.0 (CHCH2), 155.6 (q, JC,F = 37.3 Hz, CvO)
ppm.
The analytical data are in accordance to those reported.34
Conflicts of interest
There are no conflicts to declare.
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Paper
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(q) S. Suljić, J. Pietruszka and D. Worgull, Adv. Synth.
Catal., 2015, 357, 1822.
Acknowledgements
We would like to thank Carmen Plass for assistance in NMR
measurements.
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