Abstract
Synthetic self-assembled systems combine responsiveness and reversibility with the ability to perform chemical tasks such as molecular recognition and catalysis. An unmet challenge is the construction of polymeric materials that, like nature's tubulin, are simultaneously reversible and capable of useful physical tasks. We report here a class of reversibly formed polymers that show covalent-polymer mechanical integrity in solution and in the solid state. Non-Newtonian, polymeric behavior is observed despite the low molecular weight of the individual subunits and the seemingly weak forces holding the assemblies together. These polymers assemble through self-complementary hydrogen bonding and by physical encapsulation of small molecules; accordingly, the emergent macroscopic structure and function can be controlled by appropriate chemical signals.
Reversible polymers offer many advantages over their traditional covalent counterparts because of their benign processing conditions and ability to organize rapidly into ordered systems with few imperfections (1–3). A range of noncovalent polymeric systems based on hydrogen bonding has appeared in recent years, and ordered films and fibers have been produced that display a rich array of structures (4–8). Stadler and coworkers (5, 9) were among the first to examine the solid-state bulk dynamic properties of polymers in which hydrogen bonds partly define the polymer main chain. Likewise, Meijer and coworkers (6) showed that reversible polymers constructed from strongly associating subunits (Ka > 106 M−1) can exhibit increased viscosities in solution and covalent polymer viscoelasticity in the bulk. We have previously used NMR spectroscopy (10) and light scattering to verify the polymeric nature of assemblies composed of reversibly formed encapsulation complexes. All of these polymers are formed from specific interactions that are well-characterized in solution. Although weak, noncovalent forces are known to contribute to polymeric viscoelasticity in other systems, e.g., micelles (11) and organometallic assemblies (12), little is known about the mechanical properties of linear, hydrogen-bonded, main-chain polymers in solution (13). We now report that synthetic reversible polymeric capsules—the polycaps—evince mechanical properties and viscoelastic behavior that are characteristic of covalent polymers.
Materials and Methods
Proton 1H NMR spectra were recorded on a Bruker (Billerica, MA) DRX-600 (600 MHz) spectrometer. IR spectra were recorded on a Perkin–Elmer Paragon 1000PC Fourier-transform infrared spectrometer. Electrospray ionization mass spectrometry experiments were performed on an API III Perkin–Elmer SCIEX triple quadrupole mass spectrometer. Rheometry measurements were obtained with both parallel-plate and cone-and-plate geometries on a Rheometric Scientific (Piscataway, NJ) SR-5000 dynamic stress rheometer. Rheology studies were performed in o-dichlorobenzene, which is both high boiling (180°C) and an excellent guest for the calixarene capsules. Temperature control was provided by a Peltier heating stage. All reagents were purchased from Aldrich or Fluka and used without further purification. The PAMAM tetraamine (generation 0) was purchased from Aldrich as a 20 wt % solution in methanol. Before coupling, the methanol was removed in vacuo and the reagent was dried under high vacuum overnight.
Synthesis of the linear polycap monomer 1 has been previously reported (10). The tetravalent cross-linker 2 was achieved as follows. To a solution of calixarene monoacid (0.26 g, 0.17 mmol; ref. 10) in dimethylformamide (10 ml) was added PyBOP (0.091 g, 0.17 mmol), NEt3 (39 μl, 0.28 mmol), and the PAMAM tetraamine (0.018 g, 0.035 mmol) as a solution in dimethylformamide (2 ml). The solution was heated to 40°C for 2.5 h and overnight at room temperature. After aqueous work-up, the crude material was purified by preparative TLC (10:1 CH2Cl2/MeOH) and methanol precipitation to give 0.055 g (25%) of the pure material as a white powder. 1H NMR (40°C, 600 MHz, dimethylformamide-d7) δ 8.47–8.11 (m, 40H), 7.37–7.35 (m, 16H), 7.30 (d, 16H, J = 13.4 Hz), 7.07–6.99 (m, 48H), 6.79 (m, 16H), 4.56 (s, 8H), 4.49 (d, 8H, J = 13.4 Hz), 4.45 (d, 8H, J = 13.1 Hz), 4.01 (m, 8H), 3.93 (m, 8H), 3.88 (m, 8H), 3.53 (m, 8H), 3.45 (m, 8H), 3.22 (d, 8H, J = 13.6 Hz), 3.15 (d, 8H, J = 13.3 Hz), 2.25 (s, 4H), 2.23 (s, 48H), 1.89 (m, 24H), 1.42–1.25 (m, 184H), 0.92–0.88 (m, 36H). IR (thin film) 3280, 2909, 2842, 1656, 1627, 1594, 1544, 1511, 1468, and 1202 cm−1. Low resolution mass spectrum (electrospray ionization; Mavg) calculated for C340H512N42O40 6428, found 6428.
Freeze-fracture transmission electron microscopy samples were prepared as follows. A mixture of 1 and 2 was sonicated in CH2Cl2/MeOH (10:1) until homogeneous. The solvent was removed in vacuo and the resulting film was sonicated, with mild heating, in chloroform (≈1 g). Freeze-fracture replicas were prepared by first placing a small portion of the gel on a gold specimen carrier. The sample was cooled in a bath of liquid nitrogen (−196°C) and transferred to a Balzers BAF 400T freeze-fracture machine. The sample was fractured at −150°C and a pressure of 10−7 torr (1 torr = 133 Pa). Platinum was shadowed at a 45° angle to a depth of ≈20 Å followed by a 250- to 300-Å carbon support layer at normal incidence. The replicas were washed with CH2Cl2/MeOH (10:1), transferred to copper grids, and examined by using a Philips CM100 transmission electron microscope (Philips Electronic Instruments, Mahwah, NJ).
Results and Discussion
The polycaps are constructed from calixarenes that bear ureas on their upper, wider rims. The calixarenes dimerize by hydrogen bonding to form capsular host complexes (Kd = 106−108 M−1; ref. 14). When two such molecules are linked at their lower rims, as in 1 (Fig. 1a), the recognition elements diverge, and assembly results in a polymeric string of capsules (Fig. 1b). Unlike other main-chain, hydrogen-bonded polymers, the polycaps are func tional host units and form only when guests of proper size, shape, and chemical surface are present. For these studies, o-dichlorobenzene and chloroform have the dual role of solvent and guest for the calixarene capsule hosts.
The polymeric nature of 1 in o-dichlorobenzene is evident from its solution viscosity. As shown in Fig. 2, the viscosity increases slowly with concentration of 1 (log–log slope = 0.4) until a critical concentration of 0.6% by weight is attained, at which point the viscosity begins to rise much more significantly (slope = 3.3). This behavior is characteristic of a transition from the dilute to semidilute concentration regime (c*), where individual polymer chains overlap and entanglements influence the observed viscoelasticity (15). The value of c* observed here is in excellent agreement with the prediction based on independent light-scattering studies (0.6% in chloroform; A. Lomakin, G. B. Benedek, R.K.C., C.N., and J.R., personal communication), and the scaling laws (G′, ν ≈ 1; G′′, ν ≈ 1.8) agree with previous theoretical (16) and experimental (6) studies.‖
Unlike covalent polymers, the molecular weight distribution of the polycaps changes significantly with temperature, and this temperature dependence is reflected in their solution rheology. Typically, viscoelastic behavior from multiple temperatures may be superimposed by using a single shift factor describing the activation energy of the relaxation process (15). For the polycaps, a single parameter is not sufficient; two parameters are required, representing a concatenation of the viscoelastic activation energy and the change in molecular weight with temperature. The parameters for the TTS (time–temperature superposition) fit from 258–318 K and referenced to 298 K are: ΔH(α) = 19.8 kcal mol−1; ΔH(β) = 3.8 kcal mol−1.
The reversible nature of the polymerization is also dramatic in its chemical sensitivity. Protic solvents disrupt the hydrogen-bonded assembly, and the addition of just 5% methanol turns the polymerization “off,” as shown in Fig. 3 for a 2.8% solution of 1. With added methanol, the viscosity decreases by two orders of magnitude—from that of thin syrup back to nearly that of the pure solvent (Fig. 3). The methanol is removed easily by open heating at 50°C for a few minutes and the solution viscosity returns to its initial value, highlighting that these materials may easily be recycled between their polymeric and monomeric forms by using extremely gentle conditions.
To what extent do the assemblies withstand physical forces, for example those imposed by shear? Viscosity decreased with increasing oscillatory shear rate (Fig. 4a), but even large, steady shear (500 s−1) did not completely destroy the polymeric behavior, and instantaneous (<1 sec) recovery of viscosity was observed (Fig. 4b). A more stringent test of the mechanical integrity of the polycaps is to measure physical forces such as normal stresses in solution. When polymers deform in a flow field, they may relax in directions perpendicular to the applied shear, creating normal forces in that direction (17). In noncovalent systems, an alternative response is possible; namely, the assembly can simply break into monomers if the noncovalent junction is not strong enough to withstand the forces. Despite the reversible nature of the polycap assembly, we observed normal forces in excess of 100 g (≈1,000 Pa) from a 3.7% solution of 1 in o-dichlorobenzene (Fig. 5); the reversible polymerization is strong enough to withstand these disruptive conditions. This behavior is, to our knowledge, the first demonstration of significant physical integrity in hydrogen-bonded main-chain polymers in solution.
The polycaps are even stronger in the solid state. Polycap 1 forms liquid crystalline phases in concentrated chloroform solutions, and fibers may be drawn from those solutions (8). Despite the crude drawing conditions, the fibers are highly ordered and have tensile strengths on the order of 108 Pa, or 1 g/denier as measured by the load at break (Fig. 6). By comparison, commercial nylon fibers, which are less ordered, typically have strengths of ≈5 g/denier (18). Although covalent polymer fibers are difficult to recycle, however, the polycap fibers can easily be dissolved and redrawn on the benchtop.
Introduction of the tetravalent cross-linker 2 creates three-dimensional networks. These networks are reminiscent of physical gels, in which covalent polymers are cross-linked through weakly associative interactions (19, 20). In our system, however, the structural components are reversed: The noncovalent chains have interspersed covalent cross-links, leading to a greater number of relaxation and rearrangement pathways in the network.
As with the linear polycaps, the networks have significant mechanical integrity in o-dichlorobenzene solution, and the cross-linker imparts a greater elastic component to the viscoelasticity than observed in the solutions of linear polycaps. Because of the reversible connectivity, gels formed from 4.5% 1 and 0.5% 2 are rigid on short timescales but flow on longer timescales. Like many physical gels, they are dilatant and thicken under shear to the point that a discrete ball may be formed and manipulated (Fig. 7; ref. 21). The relaxation pathway is complex and characterized by a nonexponential decay in viscoelastic properties once the shear is halted.
For a 5% solution (4:1 1:2) in chloroform, the associations are such that >99% of the polycap material should exist in effectively “infinite” networks (19). In covalent polymers, the high cross-linking density would permanently fix the structure of the network, but the polycaps retain the ability to rearrange into well ordered mesoscopic and macroscopic structures. Fig. 8 shows a transmission electron microscope surface image of the gel after a few seconds of solvent evaporation. Phase separation at the surface is apparent, and the polycap network responds to form an array of tendrils possessing a remarkably regular diameter of only a few molecules (≈10 nm).
In conclusion, the polycaps combine the chemical and thermal control of assembly with desirable properties such as elasticity and steric repulsion that depend on persistent polymeric structure. Although the hydrogen bonds holding the assembly together are weak enough to dissipate on relatively short timescales (from seconds to hours), the polymeric chains have measurable and meaningful mechanical integrity—even in solution. The solid-state properties of oriented fibers approach those made from traditional, covalent polymers. Finally, because assembly depends on the encapsulation of a suitable guest, the reversible polymerization can be triggered by molecular recognition, providing macroscopic structural changes in response to chemical signals.
Acknowledgments
We thank Dr. Frank Würthner and Dr. Uwe Beginn (Universität Ulm, Ulm, Germany), and their students, Christoph Thalacker and Jürgen Ellman, for initial viscosity measurements and insightful discussions, Dr. Malcolm Wood and Theresa Fassell (The Scripps Research Institute) for assistance with the freeze-fracture experiments, and Dr. Tom Wilson (Rohm and Haas, Bristol, PA) for helpful discussions. We thank the Skaggs Research Foundation and the National Institutes of Health for financial support. C.N. and S.L.C. thank the National Institutes of Health for postdoctoral fellowships. R.K.C. thanks the American Chemical Society, Division of Organic Chemistry, and Schering Plough, Inc., for a graduate fellowship.
Footnotes
The c* transition has recently been observed by Meijer and colleagues in their study of polymers formed by quadruple hydrogen-bonds. (S. H. M. Sontjens, R. P. Sijbesma, and E. W. Meijer, personal communication.)
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