Virus-like Cage Hybrid: Covalent Organic Cages Attached to Metal Organic Cage

Abstract

A well-defined virus-like cage hybrid (VCH) with 24 covalent organic cages (COCs) attached to one metal organic cage (MOC) is presented here. The quantitative assembly of VCH was completed through coordination between soluble anisotropic COC bearing one bipyridine moiety and Pd(II) ions. The obtained VCH exhibited discrete, uniform and stable structures with good solubility and was well characterized by NMR, FT-IR, TEM, AFM, DLS, TGA, and so on. This designable cage hybrid promotes a new strategy to expand the structural and functional complexities of porous molecular cages.

1. Introduction

For billions of years, the evolution of nature has given birth to huge amounts of precise objects with various structures and functions, ranging from atoms, molecules, biomacromolecules, bacteria, viruses, cells, organs, to lives [1]. Inspired by these precise objects that nature has brought to perfection, the last few decades have also seen an upsurge in research related to well-defined nanoobjects, based on nanoscience and nanotechnology. Moreover, this leads to deeper insight into biological evolution, as well as the exploration of practical functionality and applicability.

The typical well-defined nanoobjects include nanocarbons [2,3,4], dendrimers [5,6], metal nanoclusters [7,8], and porous molecular cages (PMCs) [9,10,11]. By virtue of their high specific surface area, tunable porosity and excellent chemical stability, PMCs are gaining more and more recent attention [10]. A growing number of PMCs are developed by linking various building blocks with different bending angles and decoration of functional groups through metal coordinative or strong covalent bonds [12,13]. Based on the interaction modes between building blocks, PMCs are mainly divided into two categories: covalent organic cages (COCs) and metal organic cages (MOCs). Different from insoluble extended porous frameworks, these cages have good solubility, which allows processing options. The permanent intrinsic cavities inside a rigid structure and windows as the passages make the binding of guests possible. Therefore, this relatively new type of porous material has proven to be a versatile functional material platform in catalysis, controlled drug release, gas adsorption and compound separation. Recent work in this area encompasses a wide range of sophisticated architectures, such as interlocked cages [14], cage oligomers [15,16], porous polymers [17,18], and so on. However, compared to their natural paradigms, the complexity of PMCs is still at the infancy stage [19]. With these structures limited to single kinds of PMCs, it is encouraging to realize the hybridization of different cage species, because this undoubtedly gives full play to their respective advantages.

The main challenge to effectively construct cage hybrids is that most of the PMCs are structurally isotropic with high symmetries, and tend to form infinite frameworks instead of discrete cage hybrids. Until recently, we achieved the combination of two types of cages for the first time by designing anisotropic COCs bearing one isophthalate moiety, which coordinate with Cu(II) ions, with or without the help of extra dicarboxylic ligands, to generate different topological MOCs [20]. We obtained discrete and uniform planet-satellite cage hybrids with excellent solubility in common solvents and with film-forming properties.

Among a series of giant MOCs with the general formula Pd_nL_2n, developed by the Fujita group, the highly-stable Pd₁₂L₂₄ complexes self-assembled from 12 Pd(II) ions and 24 bipyridine ligands have been mostly studied [21,22]. An attractive aspect of this system is the capability of exohedral or endohedral cage functionalization via introducing side chains, which makes them highly attractive for areas like molecular recognition processes [23,24] and bioimaging [25]. What is more, being in possession of a rigid skeleton, both with a large cavity and many apertures that are permeable to small molecules, the Pd₁₂L₂₄ nanospheres can also be effectively employed for drug delivery [26,27,28,29] and as catalyst carriers [30].

Herein, in a continuation of our earlier work, we designed a soluble anisotropic COC ligand tethered with one bipyridine moiety. Through its coordination with naked Pd(II) ions, a well-defined virus-like cage hybrid (VCH) with 24 COCs attached symmetrically to the surface of Pd-based MOC were formed quantitively. They resemble the spike-like surface protein and capsid of a virus, respectively (Scheme 1).

2. Materials and Methods

All the reagents and solvents were purchased from commercial sources and used as received without further purification. All glassware were cleaned with aqua regia and rinsed several times with water prior to use.

2.1. Synthesis of COC Ligand

The COC ligand was synthesized through a substitution reaction according to the Scheme 2. Functional cage, compound A, was synthesized according to our previously reported method [16]. Compound A (0.22 g, 0.22 mmol, 1.0 eq.), Compound B (60.35 mg, 0.243 mmol, 1.1 eq.) and K₂CO₃ (39.47 mg, 0.286 mmol, 1.3 eq.) were added in tetrahydrofuran (THF, 150 mL) and the mixture was refluxed at 70 °C for 48 h. Then, the solvent was removed under reduced pressure. The residue was subjected to column chromatography with CH₂Cl₂:CH₃OH (100:1, v/v) as eluent to afford the COC ligand as a white solid (0.12 g, yield 45.2%). ¹H NMR (400 MHz, CDCl₃) δ 8.66 (d, J = 3.8 Hz, 4H), 8.03 (s, 1H), 7.75 (d, J = 4.8 Hz, 4H), 7.69 (s, 2H), 6.60 (s, 2H), 6.59 (s, 4H), 3.50 (t, J = 7.4 Hz, 8H), 1.57 (br, 8H), 1.27–1.23 (m, 40H), 0.83 (t, J = 2.66 Hz, 12H). ¹³C NMR (100 MHz, CDCl₃) δ 174.28, 174.16, 172.12, 167.49, 153.55, 152.77, 152.44, 150.51, 146.84, 141.12, 123.88, 121.79, 120.82, 116.14, 114.62, 47.66, 31.83, 29.43, 29.29, 27.61, 26.89, 22.68, 14.12. MS (MALDI TOF): Calculated for C₆₉H₈₅N₁₃O₇, [M+H]⁺ 1208.6695, found 1208.6726.

2.2. Synthesis of VCH

The COC ligand (24.2 mg, 0.02 mmol, 2.0 eq) in CDCl₃ (1.5 mL) and Pd(NO₃)₂ (2.3 mg, 0.01 mmol, 1.0 eq) in DMSO-d₆ (1.5 mL) were mixed and stirred at 50 °C for 12 h. After NMR analysis, the solution was evaporated to remove CDCl₃ and, then, freeze-dried for two days to remove DMSO-d₆, allowing the isolation of the product as a pale-yellow solid.

2.3. Characterization

¹H and ¹³C NMR spectra were recorded on a Bruker AVANCE III HD 400 at 400 and 100 MHz, respectively, using an internal deuterium lock for the residual protons in CDCl₃ and DMSO-d₆ at ambient probe temperature. Two-dimensional diffusion ordered spectroscopy (DOSY) and correlation spectroscopy (COSY) were recorded on Bruker AVANCE III HD 500 at 500 MHz. Chemical shift values (δ) are reported in ppm relative to the proton residual of the deuterated solvents, and coupling constants (J) are reported in Hertz (Hz).

The precise molecular weights of the synthesized compounds were recorded through mass spectra. Matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectra were performed on a Bruker Speed MALDI-TOF 7090 (Bruker, Germany). The sample was dissolved in CDCl₃ at a concentration of 4 mg/mL, and, then, mixed with DCTB solution (10 mg/mL, in CHCl₃) at a volume ratio of 1/5. Before testing the sample drop, we needed to make sure its solvent had volatilized.

Fourier transform infrared (FT-IR) spectra were recorded on a Bruker Tensor 27 FT-IR using ATR measurements for solids as neat samples. KBr was used as a diluting agent. The frequency range was 400 cm⁻¹ to 4000 cm⁻¹.

The morphologies and sizes of VCH were estimated by transmission electron microscopy (TEM) on Talos L120C G2 (Waltham, MA, USA) operated at an acceleration voltage of 120 kV. Carbon film supported by a standard copper grid was chosen as the sample carrier. Atomic force microscopy (AFM) images were obtained on MFP-3D (Oxford) by spin-casting a highly diluted THF solution of VCH (10 ng/mL) on freshly cleaved mica substrate. The sizes of the cage hybrids were recorded through dynamic light scattering (DLS) performed with a size and zeta potential analyzer (Malvern, UK).

UV-Vis measurements were performed on a UV-1800 (Kyoto, Japan) spectrophotometer with a 1 cm path length cuvette at 298 K. To investigate the thermal stability of VCH, we performed thermal gravimetric analysis (TGA) using a TG 209 F1 Libra thermos gravimetric analyzer in flowing air atmosphere.

3. Results and Discussion

The initial COC that we used was a conformationally rigid bis(tetraoxacalix[2]arene[2]-triazine) cage with D_3h symmetry and electron-deficient V-shaped clefts [16,31,32]. Of their three evenly distributed reactive sites, we modified two of them with alkyl chains to improve their solubility in organic solvents and the other one was coupled with a bipyridine moiety to construct Pd-based cuboctahedral MOC. The structure of the anisotropic COC ligand was well confirmed by NMR and MS (see Supplementary materials Figures S1 and S2). The external COCs did not interfere with the Pd-pyridine coordination bond, so the formation of Pd₁₂L₂₄ cage was not disrupted during the self-assembly process.

After the dissolution of COC ligand in CDCl₃ and Pd(NO₃)₂ in DMSO-d₆, respectively, the two solutions were mixed at 50 °C for 12 h, from which the quantitative self-assembly of a single product was detected by ¹H-NMR spectroscopy (see Supplementary Materials Figures S3 and S4). As shown in Figure 1a, only one set of ligand signals was observed, indicating a discrete and highly-symmetric structure of VCH. The characteristic broadened resonance signals also confirmed giant species formation, the motion of which was dynamically slow on the NMR time scale. Besides, some signals from protons adjacent to the pyridyl nitrogen atom exhibited distinct downfield shift (Δδ = 0.85–0.035 ppm for H_a, H_b, H_c, H_d), indicative of the coordination with metal ions.

The graph from DOSY showed a clear single bond after the assembly at 298 K, indicating the formation of a sole product. As shown in Figure 1b, the diffusion coefficient (D) of VCH (log D = −10.33) was apparently lower than that of COC ligand (log D = −9.77), which further proved that a giant species was formed from the coordination procession. According to the Stokes−Einstein equation ($D=k_{B}T/6\pi \eta r,$ where D is the Diffusion Coefficient, k_B is the Boltzmann constant, T is the temperature, η is the viscosity coefficient of solvents, r is the hydrodynamic radius.), the dynamic diameters of the COC ligand and VCH were calculated to be 1.06 and 3.84 nm. COSY proved the correlation between H_a and H_b, which were coupled to each other in the NMR spectrum (see Supplementary Materials Figure S5).

We obtained the FT-IR spectra of the structure of COC ligand and VCH (see Supplementary Materials Figure S6). There was little difference before and after the assembly. However, the characteristic peak at 1593 cm⁻¹, assigned to the vibration of C = N of the pyridine ring, was shifted to 1629 cm⁻¹, and the signal peak (1567 cm⁻¹), corresponding to the stretching vibration of C = C on the benzene ring, also moved to a higher wave number (1589 cm⁻¹), both of which changes were caused by the coordination of Pd(II) ions [33]. Tests showed that VCH could be easily dissolved in common solvents, like THF, DMSO, and DMF, which was very favourable to further experiments and practical application.

To detect the morphology and size of the obtained VCH, we performed a transmission electron microscopy (TEM) experiment by placing a drop of VCH solution in THF (about 0.2 mg/mL) on a copper grid. It was shown that the VCH were discretely and compactly spherical nanoobjects with a uniform diameter of about 4.0 nm (see Supplementary materials Figure S7). The result was consistent with the value calculated from the Stokes−Einstein equation. An AFM image was obtained by spin-casting a highly diluted THF solution of VCH (10 ng/mL) on freshly cleaved mica substrate. A clear AFM image was obtained showing discrete spherical-shaped particles with a uniform height of 4.9 nm above the surface, as shown in Figure 2. This also revealed the stability of the complex under AFM conditions. However, because of the unavoidable tip broadening effect [34], the measured widths of the dots were slightly larger than that from TEM measurement.

In dynamic light scattering (DLS) experiments, the hydrodynamic diameters of the COC ligand and VCH were measured to be 0.6 nm and 3.6 nm with a small polydispersity index (PDI) of 0.21 (see Supplementary materials Figure S8). This was consistent with the sizes estimated from DOSY measurement. Therefore, all the experimental results validated the formation of VCH.

We also performed TGA experiments to explore the thermal stability of VCH (see Supplementary materials Figure S9). The system remained thermally stable before 300 °C, only gradually losing solvent molecules. Alkyl chain decomposition occurred between 300 °C and 450 °C. As the temperature rose, the skeleton of the MOCs and COCs rapidly collapsed. The final stabilized mass residual was supposed to be PdO.

4. Conclusions

In summary, we demonstrated a novel virus-like cage hybrid with 24 COCs uniformly attached to one Pd-based MOC. The anisotropic COC ligand modified with one bipyridine moiety could coordinate with naked Pd(II) ions in a quantitative manner to form a supramolecular structure. The formed cage hybrid exhibited discrete, uniform and stable structures with good solubility. Climbing further up the ladder of constructing atomically-precise nanoobjects, the covalent attachment of COCs on the surface of MOC opens up new strategies to expand the complexity and functionality of PMCs. We envisage that the cooperation will package their relative properties and applications, thereby constituting promising multifunctional self-assembled nanoobjects in the future.