Letter pubs.acs.org/macroletters
ROMPISA: Ring-Opening Metathesis Polymerization-Induced SelfAssembly Daniel B. Wright,†,‡,§,∥ Mollie A. Touve,†,∥,⊥ Lisa Adamiak,∥ and Nathan C. Gianneschi*,†,‡,§,∥,⊥,# †
Department of Chemistry, ‡Department of Materials Science and Engineering, and §Department of Biomedical Engineering, Northwestern University, 2145 Sheridan Road, Evanston, Illinois 60208-3113, United States of America ∥ Department of Chemistry and Biochemistry, ⊥Department of NanoEngineering, and #Materials Science and Engineering, University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, United States of America S Supporting Information *
ABSTRACT: Herein we report a polymerization-induced self-assembly (PISA) process with ring-opening metathesis polymerization (ROMP). We utilize a peptide-based norbornenyl monomer as a hydrophobic unit to provide a range of nanostructures at room temperature yet at high solids concentrations of 20 wt % in combination with an oligoethylene glycol based norbornenyl monomer. Evaluation of the polymerizations under mild conditions highlight that good control is maintained along with high monomer conversion of greater than 99%, indicating that the living polymerization is unaffected during the PISA process. The demonstration broadens the scope of the PISA process to a new living polymerization methodology toward the development of easily accessible and highly functionalized nanostructures in situ.
B
PISA process, ranging from spheres to vesicles, to exotic octopus-like structures.8,10,11,17,18 These polymerizations are often undertaken at elevated temperatures of greater than 50 °C and, in some cases, they must be quenched and cross-linked while at elevated temperature to retain the nanostructure when cooled, a process that can limit its utilization.18,19 Polymerization methods capable of PISA at room temperature are desirable to provide direct access to structures without requiring cross-linking, specifically because cross-linked systems can limit stimuli responsive behavior, for example, encapsulation and release in vesicle morphologies.10 Additionally, more hierarchical structures, such as supermicelles observed by Manners and co-workers, rely on a particle that can “initiate” and fuse, which in cross-linked systems would be entirely inhibited.27 Recent advances in photochemistry have paved the way for a unique answer to the requirement of conducting PISA at elevated temperatures with polymerizations instead directed by an external light stimulus.17,28−33 We sought to investigate an alternative route for conducting PISA at room temperature, which involves Ring-Opening Metathesis Polymerization (ROMP) and extends PISA to a broader range of polymerization methods. ROMP can be efficiently performed on complex monomers at room temperature with exceptional functional group tolerance.34−36 Until now, the formation of micellar nanoparticles via ROMP has been limited to standard
lock copolymer self-assembly offers a unique route for generating synthetic materials with highly complex morphologies.1−6 Despite advances, typical solution-based self-assembly is conducted at low polymer concentrations of less than 10 wt % and requires multiple steps, including switching solvents over extended time periods, limiting some applications.3,4,7 The formation of polymeric nanoscale morphologies in situ has been demonstrated, wherein both the polymer is synthesized and nanoparticles are formed simultaneously, a process termed polymerization-induced self-assembly (PISA).8−24 During the PISA process, a solvent-soluble polymeric macroinitiator is used to polymerize a second monomer. The second monomer must be solvent-soluble, with the newly formed polymer block being insoluble. Consequently, as the polymerization progresses, the second solventinsoluble block forces the polymers to spontaneously assemble into a variety of morphologies depending on the block lengths.8−25 This simple one-step nanoparticle formation is appealing for large-scale implementation and may also facilitate the formation of reproducible polymeric biomaterials in solution.10 PISA has been predominantly undertaken using radical polymerization processes, most notably, reversible addition− fragmentation termination (RAFT) polymerization, as this allows for a large range of functional monomers to be used.26 Extensive studies by Armes and co-workers and Sumerlin and co-workers have employed RAFT polymerization for the formation of a myriad of nanostructures in solution via the © 2017 American Chemical Society
Received: June 1, 2017 Accepted: July 25, 2017 Published: August 15, 2017 925
DOI: 10.1021/acsmacrolett.7b00408 ACS Macro Lett. 2017, 6, 925−929
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ACS Macro Letters two-step processes, of first synthesizing polymeric amphiphiles and then allowing them to assemble in solution. Therefore, in this work, we explored a new approach to particle synthesis: Ring-Opening Metathesis Polymerization-Induced Self-Assembly (ROMPISA). For our initial demonstration of ROMPISA and given the need for rapid polymerization, as slow conversions can lead to limits on particle control,37 we chose to employ the highly efficient Grubbs’ modified third generation initiator (IMesH2)(C5H5N)2(Cl)2Ru=CHPh. Initially, a norbornenyl oligo(ethylene glycol) (OEG), monomer was used to generate a homopolymer via ROMP and for utilization as the macroinitiator stabilizer block, Figure 1. The OEG block length was
with the OEG monomer, the initiator, and solvent. A total of 30 min following the introduction of the initiator, an aliquot of solution was analyzed by 1H NMR to confirm the full consumption of the OEG monomer. After full consumption of the monomer, a solution of peptide monomer in the solvent mixture was added to give a final concentration of 20 wt % solids. The solution was then left to stir at room temperature for 4 h, after which ethyl vinyl ether was added to quench the reaction (see SI for experimental details). A sample of the reaction solution was analyzed by 1H NMR spectroscopy and size exclusion chromatography (SEC). The combination of both concluded successful chain extension, with >99% monomer conversion, to yield a single, low dispersity molecular weight distribution (Table 1). Furthermore, each sample showed an onset of opalescence, qualitatively indicating the presence of nanostructures. To analyze the postpolymerization solutions in more detail, the samples were diluted and analyzed by dynamic light scattering (DLS) and transmission electron microscopy (TEM), Figure 2.
Figure 1. Structure of the polymer peptide amphiphiles synthesized via ROMPISA. Blue highlights hydrophilic, red highlights hydrophobic.
critical as previous studies with a brush-like stabilizer block indicates that the repulsion of high density brush polymers in the coronal block offers larger surface curvature and prevents the production of morphologies other than spheres.38 Consequently, an OEG brush with a degree of polymerization of 4 was initially selected and where the initial polymer had a degree of polymerization of 20 (see Table 1, polymer 1). The core-forming monomer of choice was a protected peptide functionalized norbornene, with the sequence: GPLGLAGGERDG (Figure 1). The choice of solvent is vital to PISA. Therefore, we began by screening for an appropriate solvent. Mixtures of dimethylformamide (DMF) and methanol (MeOH), in DMF/MeOH ratios of 1:0, 1:2, 1:3, and 1:4, were chosen, with methanol acting as a poor solvent for the peptide. Reaction vessels under a dinitrogen atmosphere were charged
Figure 2. (a) Distributions of hydrodynamic radii from DLS for the PISA process with varying solvent composition with Polymer 2 (shown in Table 1). (b) Dry-state, stained TEM image of the nanostructures formed from polymer 2 with the solvent composition DMF/MeOH 1:2.
Both TEM and DLS data highlight two key results: (1) uniform nanostructures spontaneously formed in each solvent mixture during the polymerization, thus, confirming that PISA did indeed occur. In this regard, polymer 1 showed no structures by DLS and (2); as the methanol content was increased, the size of the structures increased. This latter observation can be understood that as the concentration of poor solvent (MeOH) is increased, the unfavorable interactions between the core block and solvent increased. These unfavorable interactions result in an increased aggregation of polymers and therefore larger structures are formed in solution.39,40 From these initial results, the most promising solvent mixture ratio appeared to be 1:2 DMF/MeOH, as from a packing argument the structures formed resemble most closely a spherical micelle given the block ratios of the polymer. Therefore, further ROMPISA experiments were undertaken with a 1:2 DMF/MeOH solvent ratio. To probe the structures formed by ROMPISA, the block lengths were changed in an effort to access different morphologies, for example, worms and vesicles. The chosen block lengths of the peptide block were 25, 50, 75, and 100 (polymers 3−6), and in each polymerization, identical reaction conditions were used, with a final concentration of 20 wt % solids. After the polymerization, each reaction was quenched with ethyl vinyl ether and their molecular characteristics determined by 1H NMR spectroscopy and SEC, which again resulted in a successful chain extension and low dispersity polymers.
Table 1. Molecular Characterization of the Block Copolymers
a b
polymer
Mn,theoa (kDa)
Đb
mb
1 2 3 4 5 6 7 8 9 10 11 12
7.0 41.5 50.0 93.1 136.1 179.1 3.5 37.9 46.5 89.5 132.6 175.6
1.03 1.06 1.10 1.09 1.12 1.02 1.03 1.05 1.08 1.15 1.20 1.07
20 20 20 20 20 20 10 10 10 10 10 10
nb 20 25 50 75 100 20 25 50 75 100
Determined by monomer conversion from 1H NMR spectroscopy. From SEC-MALS based on poly(styrene) standards in DMF. 926
DOI: 10.1021/acsmacrolett.7b00408 ACS Macro Lett. 2017, 6, 925−929
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ACS Macro Letters From both TEM and DLS only spherical micelles were observed. As the block length of the peptide monomer increased, the size of the spherical micelle increased, Figure 3.
Figure 4. (a) Schematic of the OEG-b-peptide norbornene block copolymer particles prepared by ROMPISA. Dry-state TEM images of the nanostructures formed in situ: (b) polymer 9, (c) polymer 10, and (d) polymer 12. Figure 3. Dry-state TEM images of the nanostructures formed in situ: (a) polymer 4, (b) polymer 5, (c) polymer 6, and (d) schematic highlighting the change in size of the spherical micelles with increasing peptide block length.
The stabilizer block length is important because if it is too long, the surface curvature is too great and only spherical morphologies can form as observed in a previous poly(lauryl methacrylate-benzyl methacrylate) diblock copolymer PISA system described by Armes and co-workers.41 Therefore, a further study was undertaken where the polymerizations were repeated, but with an OEG norbornene stabilizer block length of 10. This smaller stabilizer block length was used in an effort to reduce the surface curvature and allow access to further morphologies.42 The targeted peptide block lengths were 25, 50, 75 and 100 (polymers 9, 10, 11, 12), and in each polymerization, identical reactions conditions were maintained, with a final concentration of 20 wt % solids. Interestingly, the DLS highlights the presence of nanostructures in solution which have similar sizes to the structures formed when the stabilizer block had a degree of polymerization of 25 (polymers 3, 4, 5, 6). Nevertheless, TEM confirms that with this shorter stabilizer block, new morphologies are found. Where increasing the core block length allows for a decrease in surface curvature and the formation of worms and vesicular structures are formed in situ. Specifically, pure worm and vesicle phases were observed, yet low dispersity polymers were still obtained, indicating that indeed the polymerization is unaffected, Figure 4 and SI for SEC chromatograms and kinetics. Moreover, the formation of higher order morphologies based upon both the core and coronal block length allows for a simple phase diagram to be produced for these ROMPISA particles, Figure 5. These results demonstrate for the first time that the PISA process can be utilized with a ROMP-based system and provides a very promising avenue for peptide-based nanoparticles of varying morphologies for therapeutics.43,44 In conclusion, we have expanded the PISA process with ROMP, using a DMF/MeOH solvent mixture. A library of OEG-b-Peptide norbornene block copolymers were synthesized which formed a range of nanostructures in situ. Importantly, these polymerizations occurred efficiently and still maintained a living character. For these brush block copolymers, the length of the stabilizer block had a significant impact on the morphology control.
Figure 5. Phase diagram constructed for OEG-b-peptide norbornene block copolymer particles prepared by ROMPISA in water at room temperature using the Grubbs 3rd generation catalyst. Ppt = precipitate.
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsmacrolett.7b00408. Synthesis of the polymers and further characterization of polymers and particles; kinetics of polymerization, TEM, and DLS of particles (PDF).
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AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected], Nathan.gianneschi@ northwestern.edu. ORCID
Nathan C. Gianneschi: 0000-0001-9945-5475 Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This research was conducted with Government support under and awarded by DoD through a MURI from the Air Force Office of Scientific Research (FA-9550-16-1-0150) and a National Defense Science and Engineering Graduate (NDSEG) Fellowship, 32 CFR 168a. We acknowledge the 927
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ACS Macro Letters
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use of the UCSD Cryo-Electron Microscopy Facility, which is supported by NIH grants to Dr. Timothy S. Baker and a gift from the Agouron Institute to UCSD. Dr. Matthew Thompson is kindly thanked for preparing the OEG norbornene monomer and Grubbs’ 3rd generation catalyst.
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