Steric and Electronic Control of 1,3-Dipolar Cycloaddition Reactions in Carbon Nanotube Nanoreactors

The use of single-walled carbon nanotubes as effective nanoreactors for preparative bimolecular reactions has been demonstrated for the first time. We show that the extreme spatial confinement of guest reactant molecules inside host carbon nanotubes increases the regioselectivity for the 1,4-triazole in thermally-initiated azide-alkyne cycloaddition reactions. Through comparison of the internal dimensions of the nanotube and the steric bulk of the reactants, we demonstrate that the formation of the more linear 1,4-regioisomer can be enhanced by up to 55% depending on the extent of spatial restrictions imposed within the nanoreactors. Furthermore, through systematic variation of the substituents in the para -position of the alkyne reactants, we reveal the unexpected influence of the electronic properties of the reactants on the regioselectivity of reactions within nanoreactors, which act to either oppose or promote the preferential formation of the 1,4-regioisomer induced by steric effects, reflecting the unique ability of carbon nanotubes to stabilise the dipole moment of confined reactants. Thus, we show that the observed regioselectivity of azide-alkyne cycloaddition reactions confined within carbon nanotube nanoreactors reflects a subtle interplay between both steric and electronic factors.


INTRODUCTION
The confinement of guest molecules inside host nanoscale containers offers an effective methodology for controlling the properties of the guest and thus the pathway and outcome of confined chemical reactions. 1 Inspired by the remarkable efficiency exhibited by enzymatic systems in nature, significant efforts of the synthetic chemistry community over the past two decades have focussed on developing a range of artificial nanoreactors, including cavitands, [2][3][4] calixarenes, 5,6 concurbiturils, 7,8 cyclodextrins, 9,10 zeolites 11,12 and supramolecular co-ordination cages, 13,14 to host chemical reactions and thus exploit the effects of extreme spatial confinement at the nanoscale on the yields and distribution of products. However, these custom-built structures often involve complex design and multi-step syntheses, typically relying upon specific host-guest interactions to facilitate high levels of encapsulation of the reactant molecules into the respective nanoscale volumes and are, therefore, generally only suitable for a limited range of reactions.
Moreover, the thermal and chemical stability of the host nanoreactor limits the range of reaction conditions available for examination. In contrast, carbon nanotubes utilise ubiquitous van der Waals forces 15 to drive the encapsulation of the widest range of guest molecules into the host internal channel while offering superior thermal, 16 mechanical 17 and chemical stability. 18 Coupled 3 with the ability to precisely tune the diameter of carbon nanotubes through control of synthesis procedures 19 and thus provide the optimum level of confinement for a given chemical reaction, these factors suggest that carbon nanotubes potentially represent the most versatile nanoscale container for chemical reactions. 20 Carbon nanotubes have been extensively studied as hosts for a broad range of catalytic chemical transformations whereby confinement within carbon nanotubes has been shown to influence the activity, selectivity and recyclability of molecular and nanoparticle catalysts; [21][22][23] yet, the direct effect of extreme nanoscale confinement in carbon nanotubes on non-catalytic chemical reactions remains relatively underexplored. The reported formation of unusual oligomers and polymers, [24][25][26] graphene [27][28][29] and inorganic nanoribbons, 30 nanotubes 31,32 and molecular nanodiamonds 33 inside carbon nanotubes clearly demonstrates the potential of narrow nanotubes to act as nanoscale reaction vessels, in these instances controlling the formation of one-dimensional macromolecular products inaccessible by other means. However, the resulting strong interactions between the host nanotube and the formed guest nanostructures prohibits the extraction and subsequent bulk characterisation of the afforded products, essential in preparative synthesis (the so-called product inhibition effect).
To the best of our knowledge and despite an increasing number of excellent theoretical contributions, [34][35][36][37][38][39] the only reported preparative, non-catalysed, organic chemical reaction performed experimentally within narrow carbon nanotubes is the aromatic halogenation reaction, 40 where it was observed that extreme spatial confinement in narrow single-walled carbon nanotubes (SWNT) directed the site selective electrophilic attack of the incarcerated aromatic guest reactant 4 and resulted in an enhancement in the formation of the more linear para-regioisomer from 68 to 97%. In this study, we explore the effect of nanoscale confinement in SWNT on the formation of the products of 1,3-dipolar cycloaddition reactions, a class of bimolecular reactions with two reactants typically of commensurate size and complexity. Through careful and systematic variation of the internal dimensions of the host nanotube and the steric and electronic properties of the guest reactant molecules, we demonstrate the enhanced formation of 1,4-triazoles in the confined cycloaddition of azides to alkynes, the magnitude of which depends on both the geometric matching of the specific transition state with the inner diameter of the host nanotube and the electron-withdrawing or -donating nature of the encapsulated reactant.

RESULTS AND DISCUSSION
The confined cycloaddition reaction in SWNT The 1,3-dipolar cycloaddition of azides to alkynes in solution under thermal initiation and in the absence of a metal catalyst are expected to produce a mixture of both 1,4-and 1,5-substituted triazoles. 41 Indeed, our control experiments exploring the reactions of benzyl azide (2) with various terminal alkynes (1a-h) afforded a mixture of both the 1,4-and 1,5-substituted triazoles (3 and 4 respectively), with the selectivity for the 1,4-triazole ranging from 8 to 62% (Table 1 and S1, Supporting Information).   Our preliminary investigations, whereby one of the reactants was selectively confined in the internal channel of SWNT (dNT = 1.3±0.1 nm) using the liquid-phase filling and fractional distillation procedure (Figure 2) developed in our previous study, 40 followed by the addition of the second reactant in solution, confirmed the validity of our hypothesis (S1.4, Supporting Information).

Figure 2.
Schematic representation of the liquid-phase encapsulation and fractional distillation procedure used to remove excess external molecules resulting (benzyl azide)@SWNT. As the azide-alkyne cycloaddition is known to occur in the absence of nanotubes, this procedure ensures that the reaction locus is exclusively defined as the hollow interior of SWNT and thus allows us to probe the influence of spatial restrictions on the selectivity of the reaction.
Using 1 H NMR spectroscopy analysis of the reaction mixtures extracted from SWNT nanoreactors into solution, we observed that confinement in carbon nanotubes improves the selectivity for the 1,4-regioisomer (S1,4) in the reaction of phenylacetylene (1a) with benzyl azide (2) by 7% (entry 2, Table 1). It is interesting to note that encapsulation of either the azide or alkyne reactant prior to initiation of the reaction results in similar changes in the observed regioselectivity (S1.4, Supporting Information), reflecting the comparable physical properties (e.g. melting point, boiling point, vapour pressure) of these two reactants. However, the efficacy of encapsulation for the range of organic reactants explored in this study is highly variable and therefore, for ease of methodology and comparison, all subsequent reactions were carried out by prior encapsulation of benzyl azide.
Whilst not as dramatic as the regioselectivity changes observed inside SWNT nanoreactors for the 9 aromatic halogenation reactions observed in our previous study, 40 it is important to note that the SWNT used in these initial experiments possess a mean internal diameter of 1.3 nm, which, based on consideration of the geometry of the most stable conformations of the 1,4-and 1,5-triazoles formed from the cycloaddition of benzyl azide (2) to phenylacetylene (1a), does not represent the optimum nanoscale reaction environment to promote the formation of the more linear 1,4regioisomer, i.e. through exclusion of the 1,5-regioisomer (Figure 3). Indeed, such an effect should theoretically be most prevalent inside narrow SWNT of mean internal diameter 1.0 nm; however, only a 3% increase in S1,4 was observed in the reaction confined in this narrower SWNT container relative to the control reaction in bulk solution (entry 1, Table   1). Moreover, the corresponding reaction conducted in a wider SWNT of a mean internal diameter of 2.0 nm -a reaction volume considerably larger than the dimensions of both regioisomers, where no spatial restriction is expected -showed a 6% increase in S1,4 compared to control reactions in the absence of nanotubes (entry 3, Table 1). Although these observations support the anticipated phenomenon that nanoscale confinement within SWNT results in an increase in the selectivity for the 1,4-triazole, the magnitude of this observed effect cannot be rationalised simply by consideration of the size and shape of the products alone as was noted in our previous studies of the aromatic halogenation reaction. 40 It seems logical that the minimum volume required for a reaction to occur depends most crucially on the critical dimension of the largest species along the reaction pathway. In many simple reactions, such as aromatic halogenation, the critical dimension may not vary significantly throughout the reaction profile and as such, consideration of the product geometry alone will suffice. However, for bimolecular reactions such as cycloaddition, which involve considerable geometric rearrangements while traversing the reaction coordinate, consideration of the geometry of the transition state, rather than the products, is more critical when attempting to control the extent of confinement and thus the reaction selectivity in confined preparative reactions.

The effect of sterics on the confined cycloaddition reaction in SWNT
Harnessing specific non-covalent interactions in host-guest nanoscale architectures represents a powerful approach to manipulate the reactivity of confined molecules. Therefore, and to gain specific insight into the importance of the aromaticity of reactants on the mechanisms of organic chemical reactions confined within carbon nanotubes, the reactions of benzyl azide (2) with cyclohexylacetylene (1b), the aliphatic analogue of phenylacetylene (1a), confined within SWNT of 1.3 nm diameter were appraised. As expected, a less pronounced effect of confinement was found, with only a 2% increase in S1,4 in nanoreactors observed (entry 4, Table 1). This indicates that the extent of non-covalent interactions between the incoming reactant and the π-framework of the SWNT interior can be used to modulate the diffusion of the reactant into the SWNT inner cavity and thus the propensity for cycloaddition reactions to occur within SWNT nanoreactors. As  Table 1). In general, it was found that increasing the steric bulk of the alkyne reactant within the same diameter nanotube produced considerably larger increases in 1,4-selectivity (ΔS1,4), up to a 55% shift (entry 5, Table 1). As the steric bulk is increased within a SWNT of a given diameter, the transition state and products experience a greater degree of spatial confinement which results in a more pronounced increase in selectivity for the 1,4-triazole (e.g. 1,1'-biphenyl cf. phenyl). Conversely, when the reaction species are smaller in relation to the SWNT cavity and thus experience less extreme spatial confinement, there is a less pronounced change in selectivity as a consequence of confinement. In this way, by varying the steric bulk of reactant functional groups, the extent of confinement can be tuned to achieve optimum confinement and thus greater control over reaction selectivity. It must be acknowledged that, by increasing the steric bulk of the alkyne, the aromaticity has also been increased and thus, the non-covalent interaction of the alkyne with the SWNT will undoubtedly be enhanced, further facilitating the confined reaction. However, the influence of these two variables is not easily deconvoluted. 14 Electron withdrawing groups tend to increase the relative molecular orbital coefficients on the terminal alkyne carbon atom, which introduces a slightly more favoured orbital overlap with the azide benzylic nitrogen, thus stabilising the transition state leading to the 1,4-triazole. As parasubstituents can influence the regioselectivity by subtle distortion of molecular orbitals, the external influence of a carbon nanotube host on the confined molecular orbitals has the potential to offer additional influence through a similar mechanism. Interestingly, confinement within SWNT increased the 1,4-triazole selectivity in all but one case, up to an 18% shift in the most extreme instance. Furthermore, the trend in unconfined regioselectivity as a function of alkyne electron-withdrawing nature appears to be amplified by confinement within SWNT (Figure 4a, blue line). The greatest enhancement in S1,4 upon confinement was observed for the alkyne with the most electron-withdrawing para-substituent (18% increase for 4-nitrophenylacetylene). It is important to note that the overall steric bulk of each of the appraised functional groups is similar (LFG, Table 1 The high polarisability of carbon nanotubes allows stabilisation of guest species with high charge-15 separation (i.e. high dipole moment) which effectively increases the dipole moment. 34 The high dipole moment of the guest molecule induces this response from the polarisable host carbon nanotube and therefore the magnitude of this stabilisation varies depending on the magnitude of the intrinsic dipole moment of the molecule. In this way, encapsulated molecules with a high dipole moment will exhibit an even greater dipole moment due to confinement, whereas guest molecules with a lower dipole moment will be influenced to a lesser extent (Figure 5). confined within a SWNT. The highly polarisable host carbon nanotube stabilises the polar species, effectively increasing the guest alkyne dipole moment (red) in comparison to the same alkyne in bulk solution (black). In this way, the extent to which the carbon nanotube influences the electronic properties of the confined molecule, and thus the regioselectivity of the reaction, depends on the initial dipole moment (i.e. extent to which the para-substituent is electron-withdrawing or -donating).

The effect of electronics on the confined cycloaddition reaction in SWNT
In the case of an electron-deficient alkyne (e.g. 4-nitrophenylacetylene), an increase to the dipole moment due to confinement within SWNT will further reduce electron density on the alkyne, thus promoting the formation of the 1,4-triazole to a greater extent. Whereas a less electron-deficient alkyne with a lower dipole moment (e.g. 4-bromophenylacetylene) will experience a less pronounced increase in dipole moment upon confinement, and thus will exhibit a smaller increase in selectivity for the 1,4-triazole by comparison. Conversely, for alkynes with electron-donating substituents, exhibiting a dipole moment in the opposite direction, an increase in dipole moment due to confinement will further increase the electron density on the alkyne, thus, reducing selectivity for the 1,4-triazole. Therefore, in this way, confinement within carbon nanotubes effectively amplifies the underlying substituent electronic effects, resulting in an increase to the gradient of the trend demonstrated (Figure 4a).
Within this dipole-dependent postulation, it might be expected that the reaction of 4-aminophenylacetylene within carbon nanotubes would afford a lower S1,4 value than that of the bulk solution. However, in addition to the postulated electronic effect, the subtle effect of steric confinement will act to increase the selectivity for the 1,4-triazole equally for each of the appraised alkynes. The observed effect of confinement on selectivity is likely to reflect a balance between both steric and electronic effects. Therefore, the net zero change in selectivity upon confinement for the reaction of 4-aminophenylacetylene may be a result of competition between electronic effects, acting to subtly reduce S1,4, and steric effects, acting to subtly increase S1,4 (Figure 4b).
However, it is important to note that the maximum change in S1,4 observed as a consequence of electronic effects (up to 18%) remains much smaller than that observed when the extent of steric confinement is optimised (up to 55%). The conclusion that, even within narrow carbon nanotubes, steric effects are typically dominant over electronic effects is supported by the solution phase studies of Huisgen. 48

The selective retention of products from the confined cycloaddition reaction in SWNT
When analysing the effect of confinement on the selectivity of cycloaddition reactions, it is of utmost importance that the product mixture analysed is a true representation of the confined reaction. Typical reaction work up procedures must include the removal of solid nanotube from suspension in order to allow solution-phase analysis of the product mixture. However, previous studies into reactions confined within molecular nanoreactors have highlighted the potential problems of product inhibition, whereby the product molecule can become irreversibly bound within the host cavity and are thus difficult to remove. 42  imperative that all organic material (i.e. reactant and product) is removed from the nanotube prior to analysis in order to be confident that spectroscopy measurements accurately represent the outcome of the confined reaction.
To quantitatively extract the triazole products in this case, the carbon nanotube solid required consecutive washing, up to three times, with sonication in a small amount of organic solvent. It was shown that additional washing of the nanotube beyond this did not extract any additional organic products within the detection limits of NMR spectroscopy and sensitivity of thin layer chromatography. Furthermore, energy dispersive X-ray (EDX) analysis of the SWNT remaining after this procedure showed no remaining encapsulated products within the limits of detection ( Figure 6). The complete extraction of products was found to be particularly important when studying the selectivity of confined cycloaddition reactions because the measured distribution of products in the extracted solution varied when comparing product mixtures from consecutive extraction stages ( Table 2). Table 2. Selectivity for the 1,4-triazole (S1,4) after initial ( i S1,4) and total ( T S1,4) product extraction, consistently showing deceptively high values for ΔS1,4 in the initial extraction, potentially due to a selective retention of the 1,5triazole by the nanotube. It was shown that the apparent 1,4-triazole selectivity values for the initial extraction aliquots were, in many cases, positively skewed when compared to the overall product mixtures after complete extraction. In almost all cases, the selectivity for the 1,4-triazole is greater in the initial extraction,

Entry
indicating that the 1,5-triazole is preferentially held within the internal cavity due to stronger interactions with the host nanotube (relative to the 1,4-regioisomer). This leads to a reduced rate of diffusion of the 1,5-triazole into the bulk solution, a consequence of the greater energy required to extract (EEXT) the 1,5-regioisomer from the nanotube cavity (Figure 7), and deceptively high values for 1,4-triazole selectivity in the initial extract, although it is important to note that these tend to decrease slightly after total extraction. SWNT is expected to be smaller than that for the 1,5-triazole due to the combined confinement effects discussed.
However, when the 1,5-triazole forms, it seemingly exhibits stronger interactions with the host SWNT and thus, the energy required to extract (EEXT) the 1,5-triazole is greater than that required for the 1,4-triazole.
This complication must always be considered for reactions confined within any nanoscale host, because the additional stage of product extraction may involve an energy barrier that must be overcome. However, selective interactions between the host nanotube and isomeric products may, if strong enough, provide a novel means of in situ product separation, affording further control over the resulting products but not by directly influencing the reaction pathway.

CONCLUSION 21
The 1,3-dipolar cycloaddition of various alkynes to benzyl azide within the internal cavity of narrow carbon nanotubes has been studied, showing consistent increases in selectivity for the 1,4triazole (up to 55%) due to extreme spatial confinement within SWNT. As hypothesised, the more linear 1,4-triazole is favoured within the SWNT cavity in almost all cases. It was shown that the extent of confinement can be tuned by varying the diameter of the SWNT host or the steric bulk of guest reactants in order to maximise the effect of confinement on selectivity. In many examples, measurable increases in 1,4-triazole selectivity were observed, despite the host nanotube cavity providing sufficient volume for both products to be encapsulated freely. This suggests that, to understand the mechanism of size or shape selectivity induced by the nanotube, the geometry of the regioisomeric transition states is the most important consideration.
A clear correlation between the electron-withdrawing nature of alkyne substituents and the increase in 1,4-triazole selectivity within SWNT suggests that the polarisability of carbon nanotubes provides an additional subtle influence on selectivity. It is postulated that the highly polarisable host SWNT stabilises the alkyne dipole moment, effectively increasing the partial charge separation. This phenomenon acts to amplify the underlying trend in regioselectivity by decreasing the alkyne electron density for electron-deficient alkynes and increasing the electron density for electron-rich alkynes. Therefore, confined cycloaddition reactions are influenced by a balance of both steric and electronic factors induced by confinement within carbon nanotube nanoreactors. However, from these results, it appears that steric effects can induce the greatest change in selectivity; the most sterically constricted reaction showing an increase in selectivity for the 1,4-triazole of 55%, compared with an 18% increase for the most electron-withdrawing.

22
It was found that, although the formation of the 1,5-triazole is less favoured within SWNT, once formed, this isomer can be selectively withheld from solution by stronger interactions with the SWNT cavity. This demonstrates that efficient extraction of products is critical, but this also highlights an additional potential method offering further control over product distribution if this effect could be harnessed.
Whilst the selectivity results obtained do not compare with those obtained in the copper-catalysed analogue, 49,50 this study clearly demonstrates that, in the absence of a catalyst, the intrinsic effects of the nanoreactor, such as spatial confinement and electronic interactions between the reactants and nanotube, can be harnessed to modulate molecular reactivity. Coupled with recent advances in operando spectroscopic analysis, 51

Author Contributions
The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript.