Cyclotrimerization of phenylacetylene catalyzed by a cobalt half-sandwich complex embedded in an engineered variant of transmembrane protein FhuA

Article abstract of DOI:10.1039/c8ob01369a

An (η⁵-cyclopentadienyl)cobalt(i) complex was covalently incorporated into an engineered variant of the transmembrane protein ferric hydroxamate uptake protein component: A, FhuA ΔCVF^tev, using a thiol-ene reaction. A CD spectrum shows the structural integrity of the biohybrid catalyst. MALDI-TOF of the segment containing the anchoring site for the cobalt complex Cys545 confirmed successful conjugation. This biohybrid catalyst catalyzed the cyclotrimerization of phenylacetylene to give a mixture of regioisomeric 1,2,4- and 1,3,5-triphenylbenzene in aqueous medium.

Full text of DOI:10.1039/c8ob01369a

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Takeharu Haino et al.
Solvent-induced emission of organogels based on tris(phenylisoxazolyl)
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Cyclotrimerization of phenylacetylene catalyzed by a cobalt half‐
sandwich complex embedded in an engineered variant of
transmembrane protein FhuA
Received 00th January 20xx,
Accepted 00th January 20xx
A. Thiel,a D. F. Sauer,a S. Mertens,b T. Polen,c H.‐H. Chen,d U. Schwaneberg,b and J. Okudaa,*
DOI: 10.1039/x0xx00000x
An (5‐cyclopentadienyl)cobalt(I) complex was covalently incorporated in an engineered variant of the transmembrane
protein Ferric hydroxamate uptake protein component: A, FhuA ΔCVFtev, using the thiol‐ene reaction. CD spectrum shows
the structural integrity of the biohybrid catalyst. MALDI‐TOF of the segment containing the anchoring site for the cobalt
complex Cys545 confirmed sucessful conjugation. This biohybrid catalyst catalyzed the cyclotrimerization of
phenylacetylene to give a mixture of regiosomeric 1,2,4‐ and 1,3,5‐triphenylbenzene in aqueous medium.
www.rsc.org/
The metal‐catalyzed cyclotrimerization of acetylene derivatives
produces benzene derivatives as two regioisomers. No
Introduction
trimerization
products
were
observed
using the
Artificial metalloproteins consisting of a synthetic metal catalyst
embedded in a protein scaffold expand the reaction scope of
natural metalloenzymes.1‐6 Whitesides et al.7, 8 introduced an
achiral biotin‐modified rhodium bis(phosphine) complex in the
protein avidin and performed asymmetric hydrogenation of
α‐acetamidoacrylic acid to obtain the product with 44% ee.
Later, Ward et al. improved the selectivity by protein
engineering2 and used this biotin‐(strept)avidin method to
generate a variety of artificial metalloproteins.3, 8‐12
Watanabe et al. incorporated a rhodium complex in the
spherical ferritin cage and utilized its discrete reaction space to
polymerize phenylacetylene with narrow molecular weight
distributions.13 Hayashi et al. prepared an artificial metallo‐
protein consisting of a variant of the β‐barrel protein
metalloproteins mentioned above. This might be due to high
temperatures required for rhodium catalysts to catalyze the
cyclotrimerization. Catalysts based on cobalt promise to
perform this reaction under milder conditions.16, 17 Low‐valent
(

5‐cyclopentadienyl)cobalt(I)
cyclotrimerization of terminal alkynes in aqueous media include
catalysts
for
the
dicarbonyl(

5‐cyclopentadienyl)‐cobalt(I)
in
supercritical
water18, in ethanol/water mixture (3:2),19 and in
methanol/water mixture (1:4) at room temperature.17
Here we report on the covalent anchoring of a modified
(
5‐cyclopentadienyl)cobalt(I) complex bearing a maleimide
nitrobindin (10
β‐strands) and a rhodium half‐sandwich
complex (Figure 1) that is covalently anchored via the
1
maleimide function at the cyclopentadienyl (Cp) ligand.
Exchanging amino acids in this metalloprotein by site‐directed
mutagenesis the stereoselectivity during phenylacetlyene
polymerization was shifted from 93% cis to 82% trans content.8,
9 By duplicating two
β‐strands the cavity size in nitrobindin was
engineered to generate sufficient cavity space for the
incorporation of bulky metal catalysts.14 A cell‐surface display
whole‐cell biohybrid system based on nitrobindin polymerized
phenylacetylene with up to 80% trans content.15
a. Institute of Inorganic Chemistry, RWTH Aachen University, Landoltweg 1, 52074
Aachen, Germany.
b. Institute of Biotechnology, RWTH Aachen University, Worringer Weg 3, 52074
Aachen, Germany.
c. Institute of Bio‐ and Geoscience, IGB‐1: Biotechnology, Forschungszentrum Jülich
GmbH, 52425 Jülich, Germany.
d. National Kaohsiung Normal University, 62 Shenjhong Rd., Yanchao, Kaohsiung,
Taiwan 82444
Figure 1: Polymerization of phenylacetylene using catalyst 1 to give cis‐PPA and using
nitrobindin‐supported catalyst to give trans‐PPA (PDB:3wjc).9
† Footnotes relaꢀng to the ꢀtle and/or authors should appear here.
Electronic Supplementary Information (ESI) available: [details of any supplementary
information available should be included here]. See DOI: 10.1039/x0xx00000x
This journal is © The Royal Society of Chemistry 20xx
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moiety in a variant of the membrane protein “Ferric
hydroxamate uptake protein component: A” (FhuA) and its
catalysis of the cyclotrimerization of phenylacetylene.
View Article Online
DOI: 10.1039/C8OB01369A
Results and discussion
5‐cyclopentadienyl)cobalt(I) complex
The water‐stable ( 2 was
reported by Eaton et al. to catalyze the cyclotrimerization of
terminal alkynes in aqueous media.19 The maleimide moiety
was attached via esterification of the alcohol residue in
the acetyl chloride group of in the presence of triethylamine
(Scheme 1). Complex
was characterized by 1H NMR, 13C NMR
and IR spectroscopy as well as elemental analysis (ESI, Figure S1
and S2). Comparing the NMR spectra of complex and
revealed a downfield shift of the protons Hd and He and
additional signals for the proton Hf, Hg and Hh (Scheme 1, ESI,
Figure S1). IR spectroscopy showed additional CO vibrational
bands at νCO = 1707 cm‐1 (ESI, Figure S2).
2 and
3
4
Scheme 1: Synthesis of 4 and 6. i) NEt3, THF, 23 °C, 14 h; ii) SDS 1 (w/w)%, H2O, 20 (v/v)%
THF, 0.1 equiv. FhuA ΔCVFTEV, 23 °C, 24 h; iii) NaPi buffer (100 mM, pH = 8), PE‐PEG (0.125
mM), EDTA (1 mM), H2O, 23 °C, 3 d.
2
4
FhuA is named FhuA_
FhuA ΔCVFtev
Conjugation of
proceeded in an anaerobic environment by addition of
1‐159_C545V548_F501_tev or in short
.
4
to cysteine residue 545 (FhuA ΔCVFtev
(10
)
Covalent conjugation of the maleimide group with the thiol
group of cysteine 545 within FhuA proceeds by Michael‐type
4
equiv.) in degassed THF to FhuA ΔCVFtev dissolved in degassed
H2O (containing 1(w/w)% sodium dodecyl sulfate, SDS).
Refolding of the β‐barrel structure was achieved by dialysis
against a sodium phosphate buffer solution (NaPi, 100 mM, pH
= 8) containing either 2‐methyl‐2,4‐pentanediol (MPD, 50 mM)
or polyethylene‐block‐poly(ethylene glycol) (PE‐PEG, 0.125
mM) as detergent (ESI, Scheme S2).
addition reaction. Selective reaction of complex
4
under basic
conditions was shown using protected
L
‐cysteine as a model for
the protein. The reaction was monitored by 1H NMR
spectroscopy following the olefinic signal of the maleimide
group at δ = 6.79 ppm (Scheme 1; Hh).20
The ferric hydroxamate uptake protein component: A (FhuA),
originally located in the outer membrane of Escherichia coli (E.
coli)21, 22 was engineered by removing the cork domain (amino
Coupling efficiency of up to 91% was determined with the
fluorescence titration method of the cysteine using ThioGlo®.20,
acids

1‐159).23 A cysteine was introduced in position 545
24‐27
(K545C) as single accessible cysteine for catalyst conjugation
within the barrel structure. The surrounding of C545 was
engineered to improve accessibility to the thiol and to avoid
catalyst binding to coordinating residues (mutations N548V and
E501F).24 This FhuA variant has a molecular weight of ca. 63 kDa.
To facilitate the characterization with mass spectrometry, two
specific Tobacco Etch Virus (TEV)‐protease cleavage sites were
introduced within the loops 7 and 8 of FhuA. The digested
protein fragment containing the metal catalyst has a molecular
weight of approximately 6‐7 kDa. The engineered variant of
The structural integrity of the β‐barrel protein host was
confirmed by circular dichroism (CD) spectroscopy. CD spectra
showed absorptions characteristic for β‐barrel structure with a
minimum at λmin = 215 nm and a maximum at λmax = 192 nm
(Figure 2). Refolded metal‐free protein and metalloprotein
were compared displaying similar characteristics (ESI, Figure
S3). Similar spectra were observed for different metalloproteins
based on FhuA ΔCVFtev 20, 24‐27
Digestion of FhuA ΔCVFtev and biohybrid catalyst
.
6
was
performed with TEV‐protease resulting in three fragments with
O
O
O
N
Co
S
O
O
calc. : 6371 Da
found: 6377 Da
Figure 2: CD spectrum of metalloprotein 6 with MPD (left), MALDI‐TOF mass spectrum of smallest fragment after digestion with TEV protease (right)).
2 | J. Name., 2012, 00, 1‐3
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5.9, 17.3 and 40.3 kDa. The 5.9 kDa fragment contains the >99% conv. (504 h) (Table 1, Entry 2‐6). The regioselectivity was
View Article Online
DOI: 10.1039/C8OB01369A
cysteine C545 and the metal catalyst was analyzed by Matrix‐ between 63% and 70% favoring the isomer 1,2,4‐triphenyl‐
Assisted‐Laser‐Desorption/Ionisation Time‐of‐Flight mass benzene (Table 1, Entry 2‐6). When sodium phosphate buffer
spectroscopy (MALDI‐TOF MS). The mass spectrum obtained (100 mM, pH = 8) was added to the reaction mixture, neither
shows the metal‐free protein at m/z = 5917 Da (calc. 5902 Da) the conversion nor the regioselectivity was affected (Table 1,
(ESI, Figure S4). Metalloprotein
Da (calc. 6371 Da) along with fragmentation of the metal FhuA ΔCVFtev limits the reaction conditions such as temperature
catalyst (Figure 2; ESI Figure S5).
(up to 64 °C)27 and the choice and amount of organic co‐
Catalytic activity of
in the cyclotrimerization was solvent.27 60 °C were selected as reaction temperature together
demonstrated with phenylacetylene. Due to insolubility of
6 was identified at m/z = 6377 Entry 9 and 11).
4
4
in with 10 (v/v)% THF as co‐solvent. FhuA ΔCVFtev tolerates up to
water, an organic co‐solvents was necessary to achieve catalytic 40 (v/v)% THF without denaturation.27
activity (Tabel 1, Entry 1). THF, ethanol, isopropanol and Generally utilizing the biohybrid conjugate
6, no additional THF
tert‐butanol were probed (ESI, Table S1). Complex 4 turned out is necessary for the catalysis because the biohybrid catalyst is
to be insoluble in tert‐butanol and only moderately soluble in perfectly soluble in water (Table 1, Entry 16). Due to low water
ethanol and isopropanol (ESI, Table S1). THF showed good solubility of the substrate, the reaction is performed in an
solubility and thus the highest conversion (39%).
emulsion‐like fashion as for the polymerization of
Increasing catalyst loadings (1.0, 2.5, 5.0, 7.5 mol%) resulted in phenylacetylene.9 Addition of 10 (v/v)% co‐solvent increased
increase in phenylacetylene conversion to give the the solubility of phenylacetylene in the aqueous phase and the
triphenylbenzene regioisomers (ESI, Table S2). The variation of activity (Table 1, Entry 17). Using PE‐PEG as detergent, the
pH from 6 to 8) did not influence selectivity or activity (Table 1, metalloprotein precipitated and only low conversions were
Entry 9 and 11). Temperature, solvents, pH‐value and additives observed (Table 1, Entry 18 and 19). In contrast, MPD as the
were varied without complex
conversion of phenylacetylene was observed (Table 1, Entry 8, during catalysis and
10, 12‐14). These results indicate that is necessary. In all catalytic runs with
The cyclotrimerization of phenylacetylene in an aqueous polymerization products of phenylacetylene were observed by
solution (H2O/THF, 9:1) catalyzed by , only
resulted in 18% conv. (24 1H NMR spectroscopy and GC MS. Using catalyst
4
present. In these cases, no detergent did not lead to precipitation of the metalloprotein
showed increased activity (TON = 130).
and neither dimerization nor
6
4
4
6
4
1
h), 39% conv. (48 h), 51% conv. (72 h), 82% conv. (168 h) and formation of poly(phenylacetylene) was observed without any
Table 1: Cyclotrimerization reaction of phenylacetylene
Entrya
Catalyst
(mol%)
4 (5)
4 (5)
4 (5)
4 (5)
4 (5)
4 (5)
4
Solvent
pH
Buffer, Detergent
Time
[h]
72
24
48
A/Bb
Conv.b
[%]
‐
18
39
51
82
>99
‐
‐
45
‐
48
‐
‐
TON
1c,e
2c
H2O
H2O/THF
H2O/THF
H2O/THF
H2O/THF
H2O/THF
‐
H2O/THF
H2O/THF
H2O/THF
H2O/THF
H2O
H2O
H2O
H2O
H2O
H2O/THF
H2O
H2O
‐
‐
‐
‐
‐
‐
‐
‐
‐
‐
‐
‐
4
8
10
16
20
‐
‐
9
‐
10
‐
70/30
67/33
64/36
67/33
63/37
69/31
‐
63/37
‐
62/38
‐
3c
4c
72
5c
168
504
72
72
72
72
72
72
72
72
6C
‐
‐
‐
‐
7f
8c
‐
6
6
8
8
8
8
8
8
8
8
8
8
NaPi
NaPi
NaPi
NaPi
9c
4 (5)
‐
4 (5)
‐
‐
‐
6 (0.1)
6 (0.1)
6 (0.1)
6 (0.1)
6 (0.1)
10c
11c
12c
13c
14c
15d
16d
17d
18d
19d
NaPi, SDS
NaPi, MPD
NaPi, PE‐PEG
NaPi, SDS
NaPi, MPD
NaPi, MPD
NaPi, PE‐PEG
NaPi, PE‐PEG
‐
‐
‐
‐
‐
5
5
13
<1
<1
24
24
24
24
69/31
70/30
69/31
‐
50
52
130
‐
48
‐
‐
a) V = 0,5 mL, 60 °C; b) determined by GC MS with a minimum of three runs, internal standard: mesitylene (1.0 mL, 15 mM in THF) selectivity = ±4%, conversion = ±4%; c)
31.5 mM phenylacetylene; d) 63 mM phenylacetylene; e) THF was removed before H2O and phenylacetylene were added; f) neat conditions: THF was removed in vacuum
before addition of phenylacetylene (70 µL), no conversion determined.
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Table 2: Cyclotrimerization of methyl propiolate
evidence for cyclotrimerization. Field et al. showed for a series
of Co(I), Rh(I) and Ir(I) catalysts that phenylacetylene can be
cyclotrimerized as well as dimerized.28 Steric and electronic
effects of the ligand system as well as the substrate influence
the regioselectivity of the cyclotrimerization. If steric and
electronic effects are minimized regioselectivity with the
statistical ratio of approximately 3:1 favoring the 1,2,4‐
substituted benzene derivative will be observed. The
cyclotrimerization of phenylacetylene showed similar regio‐
selectivities with approximately 2:1 favoring the 1,2,4‐
View Article Online
DOI: 10.1039/C8OB01369A
Entrya
1b
2c
Catalyst
4 (1)
A/Be
68/32
1/99
1/99
Conv.f [%]
19
40
39
FhuA (0.1)
6 (0.1)
3d
a) V = 0.5 mL, 60 °C; b) a stock solution of 4 (0.1 M in THF); c) 22 mM methyl propiolate;
d) 22 mM methyl propiolate; e) determined by GC MS with a minimum of three runs, int.
std.: mesitylene (1.0 mL, 15 mM in THF) errors are standard deviation: selectivity = ±4%;
f) determined by 1H NMR; int. std. acetone (4 µL), error: conversion = ±5%;
triphenylbenzene. Neither
regioselectivity. Catalyst
cobalt(I) catalyst used by Butenschön et al. in aqueous media
show similar regioselectivities.29 In contrast to catalyst
(ethanol/water mixture; 3:2)19 and the ( 5‐cyclopentadienyl)
4
2
nor
and the (
6
showed large effects on the
5‐cyclopentadienyl)‐

2

cobalt(I) catalyst (methanol/water mixture; 1:4) used by
Conclusion
Butenschön et al.29 we decreased the amount of organic co‐
A
modified
(
5‐cyclopentadienyl)cobalt(I) catalyst was
solvent necessary by incorporating catalyst
4 in the protein
anchored in the engineered transmembrane protein FhuA
ΔCVFtev with a cysteine anchoring site C545 and characterized
by MALDI‐TOF mass spectrometry and CD spectroscopy. The
cyclotrimerization of phenylacetylene with the water‐soluble
scaffold to obtain water‐soluble biohybrid catalyst
6
.
Several other terminal alkynes of varying water‐solubility were
tested in the cyclotrimerization under conditions shown in
Table 1 (ESI, Table S4). Propargyl alcohol, N,N‐dimethyl‐2‐
propynylamine, N,N,N‐trimethyl‐2‐propynyl ammonium iodide,
propiolamide and 5‐hexynenitile did not show any conversion
biohybrid catalyst
6 gave a 2:1 mixture of 1,2,4‐ and 1,3,5‐
triphenylbenzenes. The regioselectivity appears to be not
affected by the protein environment, as the formation of the
intermediate cobaltacyclopentadiene (“cobaltole”) seems to be
not influenced by the bioconjugation.
with catalyst
Regarding the activity, catalyst
cyclotrimerization of phenylacetylene.19 With respect to
phenylacetylene and did not show any difference in activity
or selectivity. Butenschön et al. observed TON’s up to 17 (5
mol%, 8 h, rt, TOF = 2.1 h‐1) for phenylacetylene.17 Catalyst
(5
mol%, 24 h, 60 °C, TOF = 0.2 h‐1) showed low activity even at
4 or 6.
2
was not used for the
2
4
Conflicts of interest
There are no conflicts to declare.

elevated temperatures. The metalloprotein
6 (0.1 mol%, 24 h,
60 °C, TOF = 5.4 h‐1) on the other hand were not active, but due
to low catalyst concentrations activity increased in aqueous
media with TON’s up to 130.
Acknowledgements
We gratefully acknowledge the financial support by the
Deutsche Forschungsgemeinschaft through the International
Research Training Group “Selectivity in Chemo‐ and
Biocatalysis”. We thank Profs. Hayashi and Onoda for various
discussions.
Acetylenes with a carbonyl group in α‐position to the C‐C triple
bond were converted with
4 as well. Methyl propiolate for
example showed a regioselectivity of 68:32 favoring the
1,2,4‐substituted benzene derivative (Table 2, Entry 1).
Biohybrid catalyst
6 selectively converted methyl propiolate
into the 1,3,5‐substituted benzene derivative (>99%) (Table 2,
Entry 3). Other acetylenes bearing a carbonyl group in the
α‐position such as ethyl propiolate or tert‐butyl propiolate
showed the same selectivity. Since FhuA without any metal co‐
References
1. J. C. Lewis, Curr. Opin. Chem. Biol., 2015, 25, 27‐35.
2. J. Collot, J. Gradinaru, N. Humbert, M. Skander, A. Zocchi and T.
R. Ward, J. Am. Chem. Soc., 2003, 125, 9030‐9031.
factor showed similar activity and selectivity as did catalyst
6
(Table 2, Entry 2‐3) it became apparent that the metal catalyst 3. F. Schwizer, Y. Okamoto, T. Heinisch, Y. Gu, M. M. Pellizzoni, V.
Lebrun, R. Reuter, V. Köhler, J. C. Lewis and T. R. Ward, Chem.
Rev., 2018, 118, 142‐231.
4. T. Matsuo and S. Hirota, Bioorganic Med. Chem., 2014, 22, 5638‐
may not be involved in the cyclotrimerization of methyl
propiolate. Previously, primary and secondary amines were
reported to act as organo catalysts.30‐32 Acetylenes bearing a
Michael‐acceptor in α‐position undergo triple Michael‐type
addition leading exclusively to the 1,3,5‐regioisomer. The
protein catalyzes these substrates and therefore the metal
complex does not participate in the reaction. Experiments with
5656.
5. J. Steinreiber and T. R. Ward, Coord. Chem. Rev., 2008, 252, 751‐
766.
6. F. Rosati and G. Roelfes, ChemCatChem, 2010, 2, 916‐927.
7. M. E. Wilson and G. M. Whitesides, J. Am. Chem. Soc., 1978, 100,
306‐307.
8. K. Fukumoto, A. Onoda, E. Mizohata, M. Bocola, T. Inoue, U.
Schwaneberg and T. Hayashi, ChemCatChem, 2014, 6, 1229‐
1235.
L
‐lysine and L‐histidine as c amino acids support this suggestion
by converting methyl propiolate exclusively to the 1,3,5‐
substituted regioisomer (ESI, Table S3).
4 | J. Name., 2012, 00, 1‐3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 5 of 5
Organic & Biomolecular Chemistry
Please do not adjust margins
Journal Name
ARTICLE
9. A. Onoda, K. Fukumoto, M. Arlt, M. Bocola, U. Schwaneberg and
T. Hayashi, Chem. Commun., 2012, 48, 9756‐9758.
10. Y. Okamoto, V. Köhler and T. R. Ward, J. Am. Chem. Soc., 2016,
138, 5781‐5784.
View Article Online
DOI: 10.1039/C8OB01369A
11. F. Schwizer, V. Köhler, M. Dürrenberger, L. Knörr and T. R. Ward,
ACS Catal., 2013, 3, 1752‐1755.
12. T. R. Ward, Acc. Chem. Res., 2011, 44, 47‐57.
13. S. Abe, K. Hirata, T. Ueno, K. Morino, N. Shimizu, M. Yamamoto,
M. Takata, E. Yashima and Y. Watanabe, J. Am. Chem. Soc., 2009,
131, 6958‐6960.
14. A. R. Grimm, D. F. Sauer, M. D. Davari, L. Zhu, M. Bocola, S. Kato,
A. Onoda, T. Hayashi, J. Okuda and U. Schwaneberg, ACS
Catalysis, 2018, 8, 3358‐3364.
15. A. R. Grimm, D. F. Sauer, T. Polen, L. Zhu, T. Hayashi, J. Okuda and
U. Schwaneberg, ACS Catalysis, 2018, 8, 2611‐2614.
16. N. Weding, R. Jackstell, H. Jiao, A. Spannenberg and M. Hapke,
Adv. Synth. Catal., 2011, 353, 3423‐3433.
17. L. Yong and H. Butenschon, Chem. Commun., 2002, 2852‐2853.
18. K. S. Jerome and E. J. Parsons, Organometallics, 1993, 12, 2991‐
2993.
19. M. S. Sigman, A. W. Fatland and B. E. Eaton, J. Am. Chem. Soc.,
1998, 120, 5130‐5131.
20. D. F. Sauer, M. Bocola, C. Broglia, M. Arlt, L.‐L. Zhu, M. Brocker,
U. Schwaneberg and J. Okuda, Chem. Asian J., 2015, 10, 177‐182.
21. J. W. Coulton, P. Mason, D. R. Cameron, G. Carmel, R. Jean and
H. N. Rode, J. Bacteriol., 1986, 165, 181‐192.
22. G. Carmel and J. W. Coulton, J. Bacteriol., 1991, 173, 4394‐4403.
23. M. Nallani, S. Benito, O. Onaca, A. Graff, M. Lindemann, M.
Winterhalter, W. Meier and U. Schwaneberg, J. Biotechnol.,
2006, 123, 50‐59.
24. F. Philippart, M. Arlt, S. Gotzen, S.‐J. Tenne, M. Bocola, H.‐H.
Chen, L. Zhu, U. Schwaneberg and J. Okuda, Chem. ‐ Eur. J., 2013,
19, 13865‐13871.
25. J. Kinzel, D. F. Sauer, M. Bocola, M. Arlt, T. Mirzaei Garakani, A.
Thiel, K. Beckerle, T. Polen, J. Okuda and U. Schwaneberg,
Beilstein J. Org. Chem., 2017, 13, 1498‐1506.
26. H. Osseili, D. F. Sauer, K. Beckerle, M. Arlt, T. Himiyama, T. Polen,
A. Onoda, U. Schwaneberg, T. Hayashi and J. Okuda, Beilstein J.
Org. Chem., 2016, 12, 1314‐1321.
27. S.‐J. Tenne and U. Schwaneberg, Int. J. Mol. Sci., 2012, 13, 2459‐
2471.
28. L. D. Field, A. J. Ward and P. Turner, Aust. J. Chem., 1999, 52,
1085‐1092.
29. R. T. Kettenbach, W. Bonrath and H. Butenschoen, Chem. Ber.,
1993, 126, 1657‐1669.
30. M. Tanaka, K. Nakamura, T. Iwado, T. Sato, M. Okada, K. Sue, H.
Iwamura and T. Hiaki, Chem. ‐ Eur. J., 2011, 17, 606‐612.
31. K. K. Balasubramanian, S. Selvaraj and P. S. Venkataramani,
Synthesis, 1980, 1, 29‐30.
32. T. Iwado, K. Hasegawa, T. Sato, M. Okada, K. Sue, H. Iwamura and
T. Hiaki, J. Org. Chem., 2013, 78, 1949‐1954.
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