Active-metal template synthesis of a molecular trefoil knot

Article abstract of DOI:10.1002/anie.201105012

Tying the knot: The marriage of catalysis and coordination chemistry enables two Cu^I ions (red; see picture) to work in partnership for the synthesis of a molecular trefoil knot. One ion entangles an acyclic building block to create a loop in t

Full text of DOI:10.1002/anie.201105012

Communications
DOI: 10.1002/anie.201105012
Chemical Topology
Active-Metal Template Synthesis of a Molecular Trefoil Knot**
Perdita E. Barran, Harriet L. Cole, Stephen M. Goldup, David A. Leigh,* Paul R. McGonigal,
Mark D. Symes, Jhenyi Wu, and Michael Zengerle
Although many different approaches to catenanes and
rotaxanes have been introduced,[1] few strategies have been
successfully developed for the synthesis of molecular knots.[2]
Trefoil knots, the simplest prime knot other than the
topologically trivial unknot (i.e., any ring or simple macro-
cycle),[3] have been found in DNA,[4] proteins,[5] and in
synthetic polymers.[6] Sauvage and co-workers prepared the
first synthetic molecular knot by using the preorganization of
two ligand strands around two tetrahedral CuI centers as the
key template interaction to generate the three crossing points
required for a trefoil knot.[7] Subsequently, donor–acceptor
interactions,[8] Watson–Crick base pairing,[9] amide hydrogen
bonding,[10] and ligand folding around an octahedral metal
ion[11] have all been used to template the formation of
molecular trefoil knots.[12]
A few years ago a strategy for the synthesis of rotaxanes
and catenanes was introduced in which metal ions play a dual
role, acting as a template to entwine or thread the building
blocks while also actively catalyzing the bond-forming
reaction that covalently traps the interlocked structure.[13]
This “active-template” approach has proven to be an effective
route to various types of mechanically interlocked molecules
and can be applied by using an increasing number of different
transition-metal-catalyzed reactions.[13j] Herein we report an
active-template reaction that occurs through a loop generated
through classical “passive-template” coordination to synthe-
size the smallest trefoil knot reported to date. The trefoil knot
was characterized by H and 13C NMR spectroscopy, mass
1
Figure 1. Schematic representation of the active-template synthesis of
spectrometry, and by drift tube ion mobility mass spectrom-
etry (DT IM-MS) experiments that show that the molecular
knot has a significantly smaller cross-sectional area (with a
narrower distribution) than the corresponding open-chain
and unknot-macrocycle isomers.
To apply active-template synthesis to a trefoil knot
architecture, we envisaged a system (Figure 1) in which a
single molecular strand with reactive functional groups at
each terminus (X and Y) could be geometrically manipulated
and knotted through multiple interactions with metal ions
a molecular trefoil knot. A single-strand ligand with one monodentate
and two bidentate binding sites (blue) and two functional end groups
(X and Y) is knotted by the action of metal ions (red, M). Step 1: One
metal ion creates a loop by coordination to the bidentate binding
sites. Step 2: The other metal ion binds to the functional end groups
and, through its preferred coordination geometry, threads the loop.
Step 3: The knotted architecture is captured by metal-catalyzed cova-
lent bond formation between X and Y.
(M). First, a loop in the strand would be formed by
coordination of two bidentate binding sites in the strand to
a tetrahedral metal ion (Figure 1, step 1). A second metal ion,
bound endotopically within the loop by a monodentate
ligating site, would then perform the twofold tasks of
1) gathering both functional end groups in a specific orienta-
tion that is dictated by the metalꢀs preferred coordination
geometry and places them on opposite sides of the loop
(Figure 1, step 2), and 2) catalyzing a covalent-bond-forming
reaction between the end groups to generate the molecular
trefoil knot (Figure 1, step 3).
[*] Dr. P. E. Barran, H. L. Cole, Dr. S. M. Goldup, Prof. D. A. Leigh,
Dr. P. R. McGonigal, Dr. M. D. Symes, J. Wu, M. Zengerle
School of Chemistry, The University of Edinburgh
The King’s Buildings, West Mains Road, Edinburgh EH9 3JJ (UK)
E-mail: david.leigh@ed.ac.uk
Homepage: http://www.catenane.net
[**] We thank the EPSRC National Mass Spectrometry Service Centre
(Swansea (UK)) for high-resolution mass spectrometry. This work
was supported by the EPSRC.
Ligand 1 (Scheme 1) was synthesized in nine steps from
commercially available starting materials (for experimental
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.201105012.
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Angew. Chem. Int. Ed. 2011, 50, 12280 –12284

4:1 chloroform-nitromethane was found to maintain the
reactants and products in solution during the course of the
reaction. An optimized concentration (1.5 mm) of 1, with
1.5 molar equivalents[18] of [Cu(CH3CN)4]PF6 at 608C, led to
complete consumption of 1 after 4 days, as evidenced by the
1H NMR spectrum of the crude reaction mixture.
After demetalation by washing with a basic ethylene-
diaminetetraacetic acid disodium salt/ammonia (Na2EDTA-
NH3) solution,[13g] the products were purified using a combi-
nation of size-exclusion (SEC) and high-performance liquid
(HPLC) chromatographies. SEC enabled facile removal of
the oligomeric by-products and the resulting mixture was
separated into its individual components by reverse-phase
preparative HPLC. Two products were isolated, in 24% and
10% yields, both of which were shown to be isomers of the
acyclic starting material 1 by high-resolution electrospray
ionization-mass spectrometry (HRESI-MS; see Figure 2 cap-
tion and the Supporting Information). The isomer formed in
lower yield (10%) was identified as the simple macrocycle 3
(i.e., a cyclic structure with unknot topology) by comparison
of its 1H NMR spectrum (Figure 2b) with that of the starting
Scheme 1. Active-metal-template synthesis of trefoil knot 2: a) CHCl3/
CH3NO2 (4:1), [(CH3CN)4Cu]PF6 (1.5 mol per mol of 1), 608C, 96 h;
b) Na2EDTA, NH3. 2 24%, 3 10%. Trefoil knots are topologically
chiral;[2,3] only one enantiomer of 2 is shown.
details see the Supporting Information). The single-strand
molecule has three potential metal binding sites: two
bipyridyl groups to create the loop (and one of the three
required crossing points) by chelation to a tetrahedral CuI ion,
and a 2,6-pyridine unit to bind the catalytically active metal
center. The CuI-catalyzed azide–alkyne cycloaddition[14]
(CuAAC) “click” reaction was chosen for the covalent-
capture reaction that forms the remaining two crossing points,
as it utilizes CuI ions (thus avoiding the complication of
having different types of metal ions in the reaction) and
because previous studies have shown this reaction to be highly
effective in rotaxane- and catenane-forming active-template
reactions.[13a,d,i,k,n,15] Molecular modeling[16] was used to esti-
mate an appropriate length for the alkyl chain spacers
between the functional end groups and the CuI binding sites.
We initially investigated reaction conditions for the
active-metal-template knotting reaction of ligand 1 (Sche-
me 1) using dichloromethane, chloroform, and 1,2-dichloro-
ethane, as these solvents had been employed in previously
reported CuAAC active-template reactions.[13a,d,i,k,n] How-
ever, upon addition of [Cu(CH3CN)4]PF6 to a dilute solution
of 1 in any of these halogenated solvents, a precipitate formed
immediately.[17] After screening a number of solvent mixtures,
Figure 2. Partial 1H NMR spectra (400 MHz, CDCl3, 300 K) of three
isomers (1, 2, and 3) of molecular formula C88H84N8O6: building block
(1) and the two products (2 and 3) isolated from the reaction of 1
shown in Scheme 1. The signals of the Hs’ protons, associated with the
-NCH2- group (adjacent to the azide in 1 and the triazole ring in 2 and
3), are shown in orange. The signals of the Hs protons, associated
with the -CH- of the triazole ring in 2 and 3, are shown in green.
a) Open-chain building block 1, HRESI-MS: m/z 1361.6613 [M+H]+
(calcd for C88H85N8O6 1361.6597). b) Unknot-macrocycle 3, HRESI-MS:
m/z 1361.6603 [M+H]+. c) Trefoil knot 2, HRESI-MS: m/z 1361.6601
[M+H]+. Expansion of the region between d=5.11 and 5.02 ppm of
1
the d) 500 MHz and e) 400 MHz H NMR spectra of 2 showing a
propargylic methylene (Hl,l’,morm’) AB system (JAB =13.2 Hz). The pro-
tons are diastereotopic as a consequence of the chirality of the trefoil
knot. The lettering corresponds to that shown in Scheme 1.
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Communications
material 1 (Figure 2a). The spectrum of the flexible 76-
Further insight into the structure of trefoil knot 2 was
provided by drift tube ion mobility mass spectrometry (DT
IM-MS). In these experiments, the velocity with which an ion
travels through a cell containing a buffer gas (commonly
helium), under the influence of a weak electric field, depends
on the collision cross section (CCS) of the ion with the buffer
membered-ring macrocycle (3) is very similar to that of the
open-chain isomer (1), the only significant differences being
the shift of the Hs’ (-NCH2-) and Hs protons (the latter from
d = 2.0 ppm in 1 to d = 7.25 ppm in 3) following conversion of
the azide and terminal alkyne to the triazole ring.
The isomer isolated in greater yield (24%) was confirmed
as the trefoil knot 2 through a series of NMR, mass
spectrometry, and DT IM-MS experiments. The 1H NMR
spectrum of 2 (Figure 2c) is very different to those of its open-
chain 1 and unknot-macrocycle 3 isomers (Figure 2a and b).
Many of the resonances in the aromatic region of 2 are
separated into two sets of inequivalent signals. The increase in
the overall number of resonances observed is a result of the
loss of the pseudosymmetry of 1 (and 3) upon formation of
the conformationally contorted trefoil knot structure—the
76-atom entwined loop is the smallest knot reported to
date[11c]—and molecular modeling[16] and DT IM-MS results
(see below) indicate it to be tightly wound. Two of the
propargylic methylene resonances (two of Hl/l’/m/m’) are shifted
significantly downfield in 2 compared to the unknot-macro-
cycle 3 (from d = 4.59 ppm to d = 5.33 and 5.07 ppm). It
appears that these protons spend a significant amount of time
edge-on to an aromatic ring in low-energy conformations of
the knot, and are deshielded through ring current effects.
Notably, the propargylic methylene resonance at d =
5.07 ppm appears as an AB system (Figure 3d,e), thus
indicating that the protons are diastereotopic.[19] This behav-
ior is a consequence of the inherent chirality of a trefoil
knot.[11d,20]
gas (averaged over all possible orientations of the ion).[21,22]
A
larger ion with few conformational restrictions takes longer to
traverse the drift cell, undergoing more collisions with the
buffer gas, than a smaller, more compact, structure. This
behavior has previously been demonstrated for naturally
occurring antimicrobial peptides[21] and for synthetic cyclic
and linear peptides.[22] With low charge numbers, flexible
molecules may wrap tightly around the charged regions in
order to solvate them with heteroatoms and aromatic rings.
However, as the number of charges on a molecular ion
increases, the size of the adopted conformations increases as
electrostatic repulsions try to force the largest distance
between the charges that the molecule will allow. In general,
the observed CCS increases with the amount of charge that a
flexible structure carries. In addition to the magnitude of the
CCS, which gives information about the size of the molecular
ion, the broadness of the distribution indicates the flexibility
(the number of differently sized and shaped conformations
adopted) of the molecular structure.
Following nanoelectrospray ionization, DT IM-MS
showed significant differences in the rotationally averaged
CCS areas of ions of the three isomers 1–3 (Figure 3).[23] The
largest average CCS areas of the open-chain isomer 1
([1+3H]3+ ion) and unknot-macrocycle 3 ([3+3H]3+ ion)
were (395 ꢀ 3.5) ꢁ2 and (368 ꢀ 5.3) ꢁ2, respectively. The
largest average CCS area observed for the trefoil knot 2
([2+2H]2+ ion) was (292 ꢀ 1) ꢁ2. Therefore for the highest
charge state observed for each species, open-chain isomer 1
has a larger molecular cross-section than unknot-macrocycle
3 which, in turn, has a much larger cross-sectional area than
trefoil knot 2. Furthermore, the open-chain isomer 1 has the
broadest CCS distribution, followed by the unknot-macro-
cycle 3, with the trefoil knot 2 having the narrowest range.
These results indicate that the trefoil knot has a much more
compact and inflexible structure than the unknot-macrocycle,
which is more compact and less flexible than the acyclic
strand. Calculations of the expected CCS values from the
Spartan-minimized[16] structures of the most extended form of
each molecular species support the observed experimental
trend (see the Supporting Information).
In conclusion, we have demonstrated an active-template
approach to the synthesis of molecular knots based upon the
cooperative manipulation of a ligand with reactive end groups
by two metal ions. One of the metal centers creates a loop in
the ligand whilst the other catalyzes a covalent-bond-forming
reaction that links the end groups through the cavity. The
resulting trefoil knot and its unknot and acyclic isomers were
characterized by NMR spectroscopy, mass spectrometry, and
DT IM-MS experiments. The latter technique is able to
discriminate between the isomers through both the size and
relative flexibility of their multiply charged molecular ions.
Active-template strategies for entangling molecular chains,
and novel methods for the structural characterization of the
Figure 3. DT IM-MS spectra of building block 1 (green, [M+3H]3+),
and products 2 (blue, [M+2H]2+) and 3 (orange, [M+3H]3+) isolated
from the reaction of 1 shown in Scheme 1. The CCS area distributions
were calculated from the measured arrival times for the highest
observed charge state of each isomer. Data shows the arrival times at
a drift voltage of 50 V. Intensities (I) are normalized to the peak areas.
Ligand strand 1, which is expected to have a large degree of flexibility,
exhibits the broadest distribution and largest CCS. Unknot-macrocycle
3 has a smaller CCS and narrower distribution and trefoil knot 2
displays the smallest CCS and the narrowest distribution, reflecting its
compact structure and persistent size and shape.
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Active-template trefoil knot synthesis: In a typical procedure, a
solution of [(CH3CN)4Cu]PF6 (14.7 mg, 39.4 mmol) in CH3NO2
(2.5 mL) was added to a solution of 1 (35.8 mg, 26.3 mmol) in
CHCl3 (14 mL) and CH3NO2 (1.0 mL), and the reaction mixture was
heated at 608C for 96 h. The solution was allowed to cool to RT and
diluted with CH2Cl2 (30 mL). A 17.5% aqueous solution of NH3
saturated with Na2EDTA (30 mL) was added and the mixture stirred
vigorously for 30 min. The phases were separated and the organic
phase was further extracted with a 17.5% aqueous solution of NH3
saturated with Na2EDTA (30 mL), H2O (30 mL), and brine (30 mL),
then dried (MgSO4) and concentrated under reduced pressure. The
resulting residue was purified by size-exclusion chromatography
(CH2Cl2 mobile phase) followed by preparative HPLC (reverse-phase
column, gradient elution: 1) MeOH with 5!0% H2O, 2) MeOH with
0!10% CH2Cl2 to give trefoil knot 2 as a colorless film (8.6 mg,
24%) and macrocycle 3 as a yellow film (3.5 mg, 10%). Full details of
experimental procedures and compound characterization are given in
the Supporting Information.
Received: July 18, 2011
Published online: September 14, 2011
Keywords: chemical topology · click chemistry · self-assembly ·
.
supramolecular chemistry · template synthesis
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MacMillan, K. Taylor, J. Dorin, P. E. Barran, Anal. Chem. 2008,
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[17] Upon adding [(CH3CN)4Cu]PF6 to a solution of 1 in chloroform
a red, gummy solid formed instantaneously. The solid was
insoluble in all common laboratory solvents and was thus
assumed to be a result of rapid polymerization.
[18] The use of less than a stoichiometric amount of copper is to try to
minimize the amount of CuI ions not coordinated to 1. “Free”
CuI ions could catalyze the CuAAC reaction without the end
groups passing through the ligand strand loop, thus generating
unknot macrocycle 3. CuI ions can turn over during active-
template CuAAC reactions [Ref 13a,d,i,l] and therefore stoi-
chiometric amounts of the metal are unnecessary.
[22] D. Macmillan, M. De Cecco, N. L. Reynolds, L. F. A. Santos,
P. E. Barran, J. R. Dorin, ChemBioChem 2011, DOI: 10.1002/
cbic.201100364.
[23] For all three isomers (1–3) the [M+2H]2+ ion was the dominant
charge state observed in the DT IM-MS experiments (see the
Supporting Information). The absence of the [M+3H]3+ species
for the trefoil knot may be indicative of a compact conformation
unable to support the electrostatic repulsions between three
protons.
[24] E. R. Kay, D. A. Leigh, F. Zerbetto, Angew. Chem. 2007, 119,
72 – 196; Angew. Chem. Int. Ed. 2007, 46, 72 – 191.
[19] The assignment is confirmed as an AB system by a coupling
constant value (JAB = 13.2 Hz) that is unchanged when measured
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