WATER ELECTROLYSIS

A split in the camp

PREBIOTIC CHEMISTRY
Making the right connections
PHASE TRANSITIONS
Nanocrystals under the microscope
Weighing up
chemistry
inside containers
MAY 2013 VOL 5 NO 5
www.nature.com/naturechemistry
Chemistry inside molecular containers in the
gas phase
Tung-Chun Lee
1
, Elina Kalenius
2
*
, Alexandra I. Lazar
3
, Khaleel I. Assaf
3
, Nikolai Kuhnert
3
,
Christian H. Grun
4
, Janne Janis
5
, Oren A. Scherman
1
and Werner M. Nau
3
*
Inner-phase chemical reactions of guest molecules encapsulated in a macromolecular cavity give fundamental insight into
the relative stabilization of transition states by the surrounding walls of the host, thereby modelling the situation of
substrates in enzymatic binding pockets. Although in solution several examples of inner-phase reactions are known, the
use of cucurbiturils as macrocyclic hosts and bicyclic azoalkanes as guests has now enabled a systematic mass
spectrometric investigation of inner-phase reactions in the gas phase, where typically the supply of thermal energy results
in dissociation of the supramolecular hostguest assembly. The results reveal a sensitive interplay in which attractive and
repulsive van der Waals interactions between the differently sized hosts and guests need to be balanced with a
constrictive binding to allow thermally activated chemical reactions to compete with dissociation. The results are
important for the understanding of supramolecular reactivity and have implications for catalysis.
I
t is well established that the reversible complexation of guests by
supramolecular hosts affects their chemical reactivity in sol-
ution
13
, and so affords impressive examples of supramolecular
catalysis and, in the case of reactions inside concave hosts,
enzyme-mimetic activity
410
. This situation is different in the gas
phase where the supply of thermal energy favours irreversible
dissociation of the hostguest complexes rather than chemical reac-
tions of the encapsulated guests
1113
. Although precedents for gas-
phase reactivity of hostguest complexes exist
1427
, inner-phase
reactions that involve chemical conversions of guests inside a
macrocyclic cavity have, until now, been limited to the solution
phase
28
. Irreversibly bound guests inside hemicarcerands serve as
particularly instructive examples of inner-phase reactions in sol-
ution
29,30
. For example, studies of diazirines in hemicarcerands
revealed the importance of dispersion interactions in stabilizing
transition states and accelerating deazatization
29,30
, although the
elimination of SO
2
from 3-sulfolene inside an asymmetric carcerand
was slower than that from the non-encapsulated guest
15
. We have
now investigated hostguest inclusion complexes between cucur-
bit[n]urils (CBn, n 68) and the bicyclic azoalkanes 13
(Fig. 1) by mass spectrometry, and document here several examples
of thermally activated, selective retro-DielsAlder reactions in the
gas phase. We observed an interesting reactivity pattern, in which
the cycloelimination inside the cavity became dominant when the
packing coefcients (PCs) of the complexes fell within a narrow
range of 3050%. A combination of constrictive binding and void
space emerged as an important structurereactivity relationship,
in which the irreversible dissociation generally prevailed over
cycloelimination when the packing was either too loose or too tight.
Among macrocyclic hosts, CBs stand out due to their high rigid-
ity, thermal stability and high-afnity binding
3134
. The structure of
their inclusion complexes is determined by a hydrophobic inner
cavity and two tighter portal regions with cation-receptor proper-
ties. Complementing examples of solution-phase reactions of
guests encapsulated in cyclodextrins and calixarenes, several
examples of chemoselective and, in part, catalytic reactions inside
CBs in water are already available
3438
. We now describe gas-phase
cycloelimination reactions of encapsulated guests, namely water-
soluble bicyclic azoalkanes. Their small size allows a complete
immersion inside the cavities of several CBs, which is accompanied
by large binding constants, on the order of 10
3
10
7
M
21
, for
the CB7 complexes in water
37,39,40
. Azoalkanes act as weak bases
41
and weak ligands
37,41
, which facilitates, in combination with
the cation-receptor properties of CBs themselves, the observation
of positively charged hostguest complexes by mass
spectrometric methods.
Results
Aqueous solutions that contained both CB7 and azoalkanes 13
showed mass spectrometry peaks of the corresponding
[CB7
.
azoalkane
.
H]

and [CB7
.
azoalkane
.
Na]

complexes.
When the latter complexes were isolated and investigated by col-
lision-induced dissociation (CID) experiments, thermal activation
led to the formation of [CB7
.
Na]

, which indicates the convention-

al dissociation of the hostguest complex (Supplementary Fig. S7).
Surprisingly, however, CID experiments with the former complexes
not only affected dissociation, but also the formation of new com-
plexes that consisted of CB7 and a reaction product from the azo-
alkane (Supplementary Figs S8S13). This provided evidence of a
fragmentation reaction of the guest inside the cavity while maintain-
ing the supramolecular assembly. When the initially produced
[CB7
.
fragment
.
H]

complexes were further subjected to CID, sec-

ondary reactions through multiple reaction intermediates occurred
for the complexes of azoalkane 2, which led to [CB7
.
H]

only at
the end of a reaction cascade.
The combined results for the different azoalkanes 13 are sum-
marized in Fig. 2. Evidently, but unexpectedly, in several cases intra-
molecular (covalent) bond cleavage competes very efciently with
the intermolecular (non-covalent) one. For the protonated CB7
complex of azoalkane 1, thermal activation led to elimination of
1
Melville Laboratory for Polymer Synthesis, Department of Chemistry, University of Cambridge, Lenseld Road, Cambridge CB2 1EW, UK,
2
Department of
Chemistry, University of Jyvaskyla, Survontie 9, 40500 Jyvaskyla, Finland,
3
School of Engineering and Science, Jacobs University Bremen, Campus Ring 1,
D-28759 Bremen, Germany,
4
Unilever R&D Vlaardingen, Olivier van Noortlaan 120, 3133 AT Vlaardingen, The Netherlands,
5
Department of Chemistry,
University of Eastern Finland, Joensuu Campus, Yliopistokatu 7, 80100 Joensuu, Finland. *e-mail: elina.o.kalenius@jyu.; w.nau@jacobs-university.de
ARTICLES
PUBLISHED ONLINE: 7 APRIL 2013 | DOI: 10.1038/NCHEM.1618
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ethylene and subsequent dissociation of the intermediary pyrazole
complex. This reaction became a minor pathway, however, when
either a smaller host with a more tight packing (CB6) or a larger
one with a more loose packing (CB8) was selected. Specically,
the inner-phase reaction efciencies at a particular collision
energy (f
r
values near centre-of-mass collision energies (E
com
) of
5 eV, Table 1) for the complexes of azoalkane 1 with CB6 (0.04)
and CB8 (0.37) were signicantly smaller than that with the
medium-sized CB7 (0.62) (Supplementary Figs S8, S12 and S13).
To ease discussion, we assign reaction efciencies that are larger
and smaller than 0.5 to be predominantly inner-phase reactions
and dissociations, respectively. Strictly speaking, the f
r
values are
lower limits, because a fraction of the observed [CBn
.
H]

stems
from dissociation of the product rather than the reactant complexes,
even at low collision energies.
For azoalkane 2, CID of the protonated CB7 complex led to the
richest gas-phase chemistry, namely a sequential elimination of rst
ethylene and then hydrogen cyanide, the latter presumably after a
1,3-H shift. Following this two-step reaction cascade, the encapsu-
lated secondary product (assigned as propenimine) did not
further fragment, but its complex with CB7 dissociated. Although
protonated CB7 was detected as a dissociation product from the
corresponding complex of 2, its abundance was substantially less
than that of both fragment complexes at low collision energies,
which resulted in a large f
r
value of 0.69 (Supplementary
Figs S8S11). Azoalkane 2 is too large to form an inclusion
complex with CB6
31,39
and no complexes were observed in this
case, which furbishes an important piece of evidence that the obser-
vable complexes are not of the exclusion type (see below). The
complex of 2 with the larger CB8 required, however, a much
larger collision energy (7.5 eV E
com
) to afford sizable amounts
of fragmentation and dissociation products (f
r
0.63) than that
with CB7 (Supplementary Fig. S13 and Section S5.2).
For azoalkane 3, no complex was formed with the smallest host,
CB6, either, but the protonated CB7 complex underwent predomi-
nantly direct dissociation, with only a small amount of elimination
of a C
4
H
7
N fragment (f
r
0.19), which very probably also entailed
an initial ethylene extrusion (Supplementary Section S4). In the
complex with the largest host, CB8, azoalkane 3 suffered elimin-
ation of ethylene only at a larger energy (f
r
0.17 at 7.5 eV
E
com
), followed by dissociation of the intermediary complex.
Numerous control experiments were performed to verify the
reaction pathways presented in Fig. 2. The loss of ethylene as
opposed to molecular dinitrogen, which is commonly encountered
in the thermal deazatization of unprotonated azoalkanes
42,43
, was
veried by Fourier transform ion cyclotron resonance (FT-ICR)
mass spectrometry accurate mass determination experiments
(Supplementary Tables S9 and S10). The same method also con-
rmed that, on CID, the [CB7
.
2
.
H]

complex did not yield

[2
.
H]

(and neutral CB7) as the alternative dissociation pathway

(Supplementary Fig. S8), a result that is fully in line with the
larger calculated proton afnities of CBs versus those of the azoalk-
anes (Supplementary Table S11). CID experiments with the com-
plexed and free azoalkanes 13 demonstrated further that N
2
H

is not a fragmentation product (Supplementary Figs S9 and S16).

Most of the observed reactions in Fig. 2 are retro-DielsAlder
reactions, which are common fragmentation pathways in mass spec-
trometry
44
. DielsAlder reactions to produce or revert bicyclic
azoalkanes are known to require the protonation of the azo
group
42,43
. The importance of the proton to catalyse the observed
cycloelimination reactions could be deduced from CID experiments
with the corresponding sodium adducts of the azoalkane complexes
(see above), which did not afford any evidence for inner-phase reac-
tions. We therefore assign the chemical reactions of the complexes
between CBn and 13 to the encapsulated protonated azoalkane
inside the CBn rather than to the encapsulated azoalkane inside
the protonated CBn. Indeed, although the proton afnities of free
CBs are higher than those of free azoalkanes 13, the complexes
with the protonated azoalkanes are energetically much more
favoured (for example, 283.1 kcal mol
21
for CB7
.
[2
.
H]

versus
232.6 kcal mol
21
for [CB7
.
H]

.
2, Supplementary Table S4)
because of additional iondipole interactions with the CBn
portals and the protonated guest. The gas-phase reactions deter-
mined for the protonated CBn complexes of azoalkanes 13 were
also observed for the uncomplexed protonated azoalkanes
(Supplementary Information), which demonstrates that the same
reactive molecular species are involved.
To conclude rigorously that the reactions took place in the inner
phase, we needed to rule out the involvement of exclusion com-
plexes, in which the protonated azo group could be associated elec-
trostatically with one carbonyl rim and the bicyclic organic residue
would remain outside the inner cavity (Fig. 3b)
45
. Although
inclusion complexes of azoalkanes 13 with CBs were established
rigorously in aqueous solution, for example, by characteristic
upeld
1
H NMR shifts
37,39
, the hydrophobic driving force for
inclusion
31,32,40
is absent in the gas phase. The relative complex stab-
ility, inclusion versus exclusion, in the gas phase would conse-
quently need to derive from differential dispersion interactions,
which, although much smaller than those for hemicarcerands
2931
,
would be maximized for the inclusion complex. Indeed, when the
geometries of the protonated inclusion and exclusion complexes
were optimized at the HF/6-31G* level of theory (Fig. 3), energy cal-
culations with ab initio (for example, HF/6-31G*) or density func-
tional theory (DFT, for example, B3LYP/6-311G**) methods
O
N
N
N
N
O
O
N
N
N
N
O
O
N
N
N
N
O
O
N
N
N
N
N
N
O
O
N
N
N
N
O
N
N
O
O
n
Cucurbit[n]uril
CB6 (n = 1)
CB7 (n = 2)
CB8 (n = 3)
N
N
N
N
N
N
1 2 3
Figure 1 | Chemical structures of the hosts (CBn, n 568) and guests
(azoalkanes 13) investigated in this study.
Table 1 | Packing coefcients (PCs), calculated activation
energies and inner-phase reaction efciencies for the
uncomplexed protonated guests.
Guest E
A
(kcal
mol
21
)
Host
CB6 CB7 CB8
PC (%) f
r
PC (%) f
r
PC (%) f
r
1 27.6 68% 0.04 40% 0.62 26% 0.37
2 35.8 78% n/a 46% 0.69 30% 0.0
3 38.1 89% n/a 52% 0.19 34% 0.0
PCs were obtained by dividing the volume of the neutral guests (Supplementary Table S10) by the
inner cavity volume of the host cavity (142
3
for CB6, 242
3
for CB7 and 367
3
for CB8),
neglecting the minor inuence (+1
3
) of the proton. The f
r
values (at approximately 5 eV E
com
(Supplementary Section S5.2)) were calculated as the ratio of the sum of the m/z peak
intensities of all fragmented complexes and the sum of the m/z peak intensities of both the
fragmented and dissociated complexes observed. The activation energies of the uncomplexed
guests (E
A
) were determined at the B3LYP/6-311G** level of theory. For the [CB8
.
2
.
H]

and
[CB8
.
3
.
H]

complexes, f
r
could not be determined at 5 eV because of negligible fragment
abundances. The f
r
values of [CB8
.
2
.
H]

and [CB8
.
3
.
H]

near 7.5 eV E
com
were 0.63 and
0.17, respectively. n/a, not applicable.
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predicted the exclusion complex between CB7 and [2
.
H]

(selected
as a representative case) to be 18 kcal mol
21
more stable than the
inclusion complex; only advanced methods that encode empirically
(wB97XD/6-31G**) or explicitly (MP2/6-31G*) for dispersion
interactions predict the CB7 inclusion complex of 2 to be distinctly
more stable than the exclusion counterpart (by 12 kcal mol
21
, see
Supplementary Table S3).
Experimentally, exclusion complexes, as they would need to be
postulated for azoalkanes 2 and 3 with CB6 on the basis of size argu-
ments
31,39
, did not afford any detectable mass spectrometry signals,
which suggests that these are too labile to be observable under our
electrospray ionization (ESI) conditions. More denitively, ion-
mobility experiments (Fig. 4) revealed that the [CB7
.
2
.
H]

complex has the same drift time (11.25+0.07 ms) and consequently
the same collision cross-section (185
2
) as CB7 itself. This result
can only be accounted for by assuming a deep immersion of 2 inside
the cavity. Exclusion complexes between guests and CBs would
display larger collision cross-sections with signicantly longer
drift times (for example, 195
2
for [CB7
.
2
.
H]

)
22
. Gratifying
to observe, on dissociation of the nal fragment (propenimine),
the empty CB7 cage recovered back to its original collision cross-
section, as expected (Fig. 4e).
C
2
H
4
[CB73H]
+
[CB83H]
+
CB6 [3H]
+
Complex not
observed
C
4
H
7
N
N
NH
1,3H-shift
C
2
H
4
CB6 [2H]
+
Complex not
observed
HCN
1,3H-shift
[[[[222H] H] H] ] H]
++++++
C
[[[2222 HHH] H] ]]
+++++++
N
NH
Dissociation
Direct
dissociation
Direct
dissociation
C
2
H
4
C
2
H
4
Dissociation
C
2
H
4
[CB61H]
+
[CB71H]
+
[CB81H]
+
c
b
a
Dissociation
Dissociation
HCN Dissociation
Dissociation
Dissociation
Direct
dissociation
C
2
H
4 Direct
dissociation
Direct
dissociation
Direct
dissociation
Direct
dissociation
+
+
[CB72H]
+
[CB82H]
+
Figure 2 | Reaction and dissociation pathways of the inclusion complexes of the bicyclic azoalkanes 13 with CBn (n 568) in CID experiments. a, For
azoalkane 1, the CB7 complex eliminates ethylene as the major pathway, followed by the dissociation of the retro-DielsAlder reaction product (pyrazole)
complex. For the more tightly packed complex with CB6 and the more loosely packed complex with CB8, dissociation becomes the major pathway (dashed
arrows indicate the minor of the two competing pathways). b, For azoalkane 2, no complex was observed with CB6. The major pathway for the CB7 complex
is elimination of ethylene and then hydrogen cyanide, followed by dissociation of the complex with the secondary product (propenimine). The CB8 complex
reacts similarly, with a slightly lower preference for inner-phase reaction than that observed for CB7 (see Table 1). c, For azoalkane 3, no complex was
observed with CB6 either, and the CB7 and CB8 complexes predominantly undergo direct dissociation. As a minor pathway, the CB7 complex eliminates a
C
4
H
7
N fragment and the CB8 complex eliminates ethylene (at a larger collision energy, see Table 1), in both cases followed by dissociation of the product
complexes. For CID mass spectra, structural assignments and fragmentation mechanisms, see Supplementary Figs S8S13, Table S15 and Section S4.
ARTICLES
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Discussion
The observation of efcient inner-phase reactions for some (but not
for all) homologous hostguest complexes discloses fundamental
insights into the factors that govern chemical reactivity in isolated
conned environments. Although unprecedented reaction pathways
observed in mass spectrometry for macromolecules are frequently
attributed to the rate and efciency by which internal energy is dis-
tributed
4648
, such RiceRamspergerKasselMarcus effects cannot
account comprehensively for the non-systematic trends of the reac-
tion efciencies in Table 1. We interpret the observed trends in
terms of a balance between three contributing factors: (1) the intrin-
sic activation energies for chemical reactions of the guest, (2) the
constrictive binding displayed by the particular host and (3) the
void space inside the hostguest complex. For example, based on
the results obtained by DFT calculations, the experimentally
observed elimination of ethylene from the protonated azoalkanes
13 shows a typical ring-strain effect in the sense that the activation
barrier increases signicantly from the smallest to the largest bicycle
(see Table 1). Some trends of the reaction efciencies for the host
guest complexes, such as that azoalkane 3 shows invariably lower
reaction efciencies than those of the smaller homologues, are unques-
tionably related to its intrinsically lower retro-DielsAlder reactivity.
When the reaction efciencies for the same azoalkane are com-
pared at the same low collision energy (f
r
values at 5 eV E
com
,
Table 1) for the differently sized hosts another trend becomes
apparent, namely that the largest host, CB8, shows smaller f
r
values than CB7 does. This can be rationalized in terms of a different
degree of constrictive binding. Constrictive binding, that is, the
observation of an activation barrier towards dissociation of encapsu-
lated guests, is a phenomenon originally described for hemicarcer-
ands
1,29,30
. For CBs, the constrictive binding originates from the
carbonyl portals, which are signicantly tighter than the inner
cavity (for example, 3.9 versus 5.8 for CB6)
38
, such that
guests need to squeeze through the portals to ingress or
egress
37,39
. Accordingly, the degree of constrictive binding increases
as the portal diameter of the host becomes narrower (from 6.9 for
CB8, to 5.4 for CB7, to 3.9 for CB6)
38
. This peculiarity retards
both the association of guests and the dissociation of their CBn
inclusion complexes, and may render them sufciently kinetically
stable to undergo (in the gas phase) chemical reactions in compe-
tition with dissociation. The idea that constrictive binding is an
important factor in triggering inner-phase reactions receives
twofold experimental support. First, fragmentation reactions occur
only for the inclusion complexes, but exclusion complexes of
azoalkanes 13 may be too unstable because they are not observed
under our conditions (see Results). Second, the protonated host
guest complex of azoalkane 2 with b-cyclodextrin, a exible macro-
cycle that does not exhibit constrictive binding, did not undergo any
detectable inner-phase reactions (Supplementary Fig. S19). The
constrictive binding in the inclusion complexes of CB8 is apparently
sufcient to allow chemical reactions to compete, but to a signi-
cantly smaller extent than for CB7, or only at a higher collision
energy (7.5 eV E
com
).
Strikingly, the reaction efciency for the inclusion complex of the
intrinsically most-reactive azoalkane 1 with the most constrictive
CB6 fell below expectation; indeed, only trace amounts of
retro-DielsAlder product complexes were detected (Table 1).
This reveals the importance of the third factor in determining
a b
Figure 3 | Geometry-optimized molecular structures (HF/6-31G* level of
theory) of [CB7
.
2
.
H]
1
complexes. a,b, Side views (top) and top views
(bottom) of the inclusion (a) and the exclusion (b) complex between CB7
and the azoalkane 2. Ab initio (for example, HF/6-31G*) and DFT (for
example, B3LYP/6-311G**) calculations predict the exclusion complex to
be up to 18 kcal mol
21
more stable than the inclusion one. However,
advanced methods, which consider dispersion interactions as well, namely
wB97XD/6-31G** and MP2/6-31G*, predict the inclusion complex to be
more stable by 12 kcal mol
21
(Supplementary Table S3). The two types of
complexes can be discriminated experimentally through their cross-sections.
As becomes obvious from the side view of (a), the [CB7
.
2
.
H]

inclusion
complex should have a cross-section comparable to that of [CB7
.
H]

,
which is observed by ion-mobility measurements (see text and Fig. 4).
In contrast, the exclusion complex has a signicantly larger cross-section
(side view of (b)) and no experimental evidence was obtained for its
existence in the course of this study.
A
b
u
n
d
a
n
c
e
Drift time (ms)
8.0 9.0 10.0 11.0 12.0 13.0
[CB72H2]
+
[CB72HC
2
H
4
HCN]
+
[CB72HC
2
H
4
]
+
[CB72H]
+
[CB7H]
+
Cross-section (
2
)
170 180 190
11.29
11.29
11.18
11.04
11.29
9.84
e
d
c
b
a
Figure 4 | Ion mobilograms. a, [CB7
.
H]

. b, [CB7
.
2
.
H]

. ce, Product ions

after CID of the [CB7
.
2
.
H]

complex. The corresponding chemical

structures are shown on the right. The cross-section scale (top) was
approximated from the calculated cross-sections for regular and inverted
CB7, assuming a linear correlation with drift time (Supplementary
Information). The relative differences in drift times (maxima) are
experimentally signicant within+0.07 ms. The peak with a 9.84 ms drift
time in (a) and (e) was identied as being caused by inverted CB7, which
forms under the CID conditions from (regular) protonated CB7 in the
absence of an included guest. The propensity for inversion of CBs in the gas
phase will be investigated separately.
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inner-phase reactivity of the hostguest complexes in the gas phase:
the necessity of void space. We have assessed void-space effects by
building on rst principles of intermolecular interactions and con-
sidering empirical observations on the preferential binding of guests
inside concave hosts. The chemical reaction, that is, the extrusion of
ethylene, was modelled by constructing a Lennard-Jones 12-6
potential of a spherical guest expanding inside a rigid host cavity
that matches the cavity size of the different CBs (Fig. 5a and
Supplementary Section S2). This potential, which includes attractive
(dispersion) and repulsive van der Waals interactions, neglects
solvent effects, and thereby provides a good comparison with the
observed gas-phase results. It also neglects electrostatic interactions
of the cationic sites with the carbonyl rim; the latter were assumed
to remain constant in the course of the ethylene extrusion. The
potentials were plotted against the guest volume and adjusted (by
varying the effective collision diameter) to display their minimum
at a 55% PC to match Rebeks empirical solution for ideal host
guest inclusion
31,32,49
.
The model potential (energy versus guest volume, Fig. 5b) nicely
reveals an initially increasing stabilization of the hostguest complex
as the guest volume increases, followed by the expected minimum,
and a steep increase as the volume of the guest becomes too large.
Evidently, the synchronous rupture of two bonds and the incipient
formation of two fragments require a positive volume of activation.
As reference points, we inserted in Fig. 5c interval bars that bracket
the volumes of the reactant azoalkane and the sum of the volumes of
the products (fragment and ethylene). The volumes of the transition
states must fall in between; for example, calculations suggest
an increase from the reactant volumes by 56
3
or 5%
(Supplementary Table S10). The propensity towards cycloelimina-
tion inside a molecular container depends critically on whether
the intermolecular forces between the host walls and the guest are
van der Waals attractive or repulsive in nature, as indicated by the
slopes of the arrows in Fig. 5c. These reect the assistance (by
increasing dispersion interactions with the host) or resistance (by
increasing Pauli repulsion against the host) as the guest expands
during cycloelimination. As can be approximated from the DFT-
calculated differential binding energies of the reactants versus pro-
ducts (Supplementary Fig. S1) as well as those of the different
guests (these are also caused by dispersion or repulsion, see
Supplementary Tables S4 and S5), the additional supramolecular
stabilization or destabilization energy amounts to several kilocal-
ories per mole, which presents a highly signicant modulation of
the intrinsic activation energies (Table 1).
Arguing in terms of the potentials (Fig. 5c) and focusing on the
variations in host size, the very small f
r
value of 1 in CB6 reects
that the reaction occurs in the repulsive region of the associated
potential (uphill arrow). In contrast, the reactions of 1 in CB7
and CB8 are efcient, because the cycloelimination occurs in the
attractive region of the potential, that is, at volumes smaller than
the minimum on the potential energy surface. Interestingly, accord-
ing to Fig. 5c, the elimination of 3 inside CB7 occurs very close to
the minimum (ideal packing (almost horizontal arrow)), which
could contribute to the lower f
r
value of the largest guest. The
increase in f
r
for CB7 on going from 1 to 2 cannot be explained
with the potentials nor with the intrinsic activation energies, such
that the lower degree of constrictive binding for the CB7
.
1
complex (facilitating dissociation) must be the important factor in
this case. Similarly, the lower f
r
values of all guests in CB8 versus
CB7 most probably result from a lower degree of constrictive
binding and cannot be judged with this simplistic Lennard-Jones
approach, which would predict similar (for 1 and 2) or more ef-
cient (for 3) reactions, as judged by the slopes of the arrows.
Arguing in terms of PC values (Table 1) and focusing on the
variation in guest size, the PC values of the CB7 complexes with
1 and 2 are sufciently small (4046%)
31,32
to accommodate com-
fortably for the required volume of activation, and for the
complex between the largest guest homologue 3 and CB7 the PC
is essentially ideal (52%) and gives little incentive for the guest to
react. In the latter case, dissociation predominates over the chemical
reaction. The complex between azoalkane 1 and the smaller host
homologue CB6 (PC 68%) is the most tightly packed, but, neverthe-
less, mainly dissociation is detected on CID. This emphasizes that a
connement of the guest is a necessary, but not a sufcient criterion
to observe predominant inner-phase reactions in hostguest com-
plexes. The competitive dissociation observed for the CB8 com-
plexes reveals, vice versa, that a sufcient amount of void space
alone is also insufcient to promote predominant inner-phase reac-
tions, but that a sufciently large connement of the guest is
another prerequisite, which must be met in parallel. Fully in line
with our train of thought, experiments in which the void space
in CB7 is articially reduced by blocking and tightening the
second portal of CB7 with either a proton or a sodium ion
(Supplementary Figs S4 and S14) show that the inner-phase reaction
pathway can be effectively shut down for azoalkanes 2 and 3 and
that dissociation then prevails. Only for the smallest azoalkane 1
does there appear to remain sufcient void space to allow the reac-
tion to occur even from doubly charged CB7 complexes. An alterna-
tive, consistent, line of argumentation involves the effect of internal
pressure on the reactions (Supplementary Section S3)
50
.
In conclusion, gas-phase reactions of guests included in molecu-
lar containers can become efcient when the hostguest complexes
full stringent boundary conditions. First, the host must display a
50 75 100 125 150
0.3
0.2
0.1
0.0
Guest volume (
3
)
1 2 3
c
0 50 100 150 200 250
0.4
0.2
0.0
0.2
0.4
Guest volume (
3
)
CB8
CB7
CB6
a
b
CB8
CB7
CB6
E
p
o
t
r
e
l
E
p
o
t
r
e
l
Figure 5 | Model potentials for the interaction of a spherical guest
positioned centrosymmetrically inside a host cavity versus guest volume.
a, Illustration of the hypothetical expansion of a spherical guest inside a CBn
cavity, as a mimic of a chemical reaction with a positive volume of
activation. b, Relative potential energies (E
rel
pot
) of the CB6 (solid), CB7
(dashed) and CB8 (dotted) complexes versus guest volume, modelled with
a 12-6 Lennard-Jones function (Supplementary Section S2). c, Magnied
area of (b) with interval bars for the volumes that correspond to guests 1
(red), 2 (blue) and 3 (orange) as they expand to the corresponding
fragment and ethylene; the arrows indicate the change in intermolecular
potential energy as the intramolecular reaction proceeds.
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constrictive binding of the guest. Second, the intermolecular forces
between the guest and the cavity walls of the host must match the
volume of activation of the chemical reaction of interest. For the
cycloeliminations examined here, which have a positive volume of
activation, there must be sufcient void space in the cavity; for
cycloadditions with a negative volume of activation, tight packing
is favourable. If any of the conditions are not met, the chemical reac-
tion of the guest inside the host is readily switched off. For CBs as
macrocycles, the void space can be reduced by selecting a smaller
host (CB6) or by docking of cations to the portals, and the constric-
tive binding can be alleviated via the selection of a host with large
portals (CB8). The two supramolecular effects should directly mani-
fest themselves on the pre-exponential factors and activation ener-
gies of inner-phase reactions and, therefore, be transferable to
catalysis and biocatalysis inside conned reaction space.
Methods
Mass spectrometry. Mass spectrometric experiments were performed using ESI FT-
ICR, quadrupole time-of-ight and quadrupole ion trap mass spectrometers for the
accurate mass measurements, prole spectra measurements, and the CID and
energy-resolved CID experiments. Ion-mobility measurements were done by direct
infusion on a Waters Synapt G2 HDMS mass spectrometer equipped with an
ion-mobility cell (see Supplementary Section S5.1 for further details, including
computational details for the analysis of the cross-sections).
Model potential function. The model potential function for the interaction of a
spherical guest positioned centrosymmetrically inside a host cavity with the guest
volume was derived by setting up a 12-6 Lennard-Jones potential and considering
dispersion as the only attractive interaction. The polarizability of the guest was
assumed to increase linearly with its size, and dispersion interactions were
assumed to arise mainly from atoms in the host interacting with atoms near the
periphery of the guest. The nal functional relationship (Supplementary Section
S2, equation (S4)) allows one to construct the potential as a function of the
volume of the guest (see Supplementary Information for further details, including
computational details for the analysis of the packing coefcients).
Received 24 September 2012; accepted 27 February 2013;
published online 7 April 2013
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Acknowledgements
A.I.L., W.M.N. and K.I.A. thank the Deutsche Forschungsgemeinschaft (DFG, grant
number NA-686/5), the Deutsche Akademische Austauschdienst (DAAD) and the Center
for Functional Materials and Nanomolecular Science (NanoFun) for nancial support,
including graduate fellowships for A.I.L. and K.I.A. The Academy of Finland is
acknowledged by E.K. for nancial support (project number 127941). The FT-ICR facility
is supported by Biocenter Finland. The authors thank L. Isaacs for making a reference
sample of inverted CB7 available, M. Olivucci for donating computing time and
D. V. Dearden for helpful comments on the results.
Author contributions
T-C.L. initiated this project with E.K., O.A.S. and W.M.N. The manuscript was co-written
by T-C.L., A.I.L. and W.M.N. and commented on by all the authors. All mass spectrometry
experiments were conducted by E.K. and A.I.L. in the laboratories of J.J. and N.K., and
C.H.G. conducted the ion-mobility experiments. K.I.A. performed the quantum-chemical
and cross-section calculations. W.M.N. and A.I.L. analysed the data in terms of the model
potentials. The student authors T-C.L. and A.I.L. contributed equally.
Additional information
Supplementary information and chemical compound information are available in the
online version of the paper. Reprints and permissions information is available online at
www.nature.com/reprints. Correspondence and requests for materials should be addressed to
E.K. and W.M.N.
Competing nancial interests
The authors declare no competing nancial interests.
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