Interrupted CuAAC Ligation: An Efficient Approach to Fluorescence

5 days ago - We introduce a powerful variant of the copper(I)-mediated azide–alkyne cycloaddition (CuAAC) for the synthesis of miktoarm block copoly...
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Interrupted CuAAC Ligation: An Efficient Approach to Fluorescence Labeled Three-Armed Mikto Star Polymers Janin T. Offenloch,†,‡ Hatice Mutlu,*,†,‡ and Christopher Barner-Kowollik*,†,‡,§ †

Macromolecular Architectures, Institut für Technische Chemie und Polymerchemie, Karlsruhe Institute of Technology (KIT), Engesserstraße 18, 76128 Karlsruhe, Germany ‡ Soft Matter Synthesis Laboratory, Institut für Biologische Grenzflächen, Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, 76344 Karlsruhe, Germany § School of Chemistry, Physics and Mechanical Engineering, Queensland University of Technology (QUT), 2 George Street, QLD 4000, Brisbane, Australia S Supporting Information *

ABSTRACT: We introduce a powerful variant of the copper(I)-mediated azide−alkyne cycloaddition (CuAAC) for the synthesis of miktoarm block copolymers with a fluorescently labeled core. Via the simple addition of a functional azirine to a conventional CuAAC reaction of an azide and alkyne terminal polymer strand, a vinyl functionality is introduced at the focal point of the two polymers, which can subsequently be modified, via, e.g., radical thiol−ene addition. Concomitantly, a second functionality−a pyrene chromophore−stemming from the functional azirine is introduced enabling fluorescence detection and unimolecular micelle formation. Thus, by virtue of thiol terminal polymers an efficient two-step synthesis of midchain functional mikto-three armed star polymers is possible. Detailed NMR, SEC, DSC, UV−vis, and fluorescence studies underpin the successful formation of the functional miktoarm star with a fluorescence labeled core. Finally, yet importantly, dynamic light scattering (DLS) results demonstrate that the miktoarm polymers form self-fluorescent micelles in aqueous media and display robust micellar stability with unimolecular micellar characteristics.

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blocks.18 Nonetheless, the copper(I)-catalyzed azide−alkyne cycloaddition (CuAAC) is the most widely employed method for making covalent connections between building blocks containing various functional groups.19 The reaction enables the synthesis of 1,2,3-triazoles as linkage elements starting from azides and terminal alkynes catalyzed by Cu(I) species in a mild, efficient and modular manner. However, if a carbene or an electrophilic partner captures the triazolyl-Cu-intermediate, 5-functionalized 1,2,3-triazoles are obtained in the so-called “Cu-catalyzed interrupted click reaction” (Scheme 1a).20,21 Hence, arylated,22 iodo-substituted,23 aminated, and sulfenylated triazoles are accessible.24 For instance, enaminesubstituted 1,2,3-triazoles derivatives are formed as the product of a CuAAC reaction via a nitrene transfer process if a 2Hazirine is present (refer to Scheme S1 for the detailed mechanism of the reaction).25 Despite the facile and efficient utilization of Cu-catalyzed interrupted click reactions in organic synthesis, their powerful nature has never been exploited for the construction of complex macromolecular architectures. This comes as a surprise, as a

INTRODUCTION The properties of synthetic polymers are critically defined by their chemical composition. Thus, the fusion of polymers with disparate properties as separate segments into block copolymersfor example in terms of solubilityenables the formation of complex polymer architectures such as unimolecular micelles, vesicles, or elongated structures.1−3 The quest to design such advanced materials with predetermined properties has driven the development of efficient approaches that facilitate access to polymers with tailored properties. While conventional methods to complex mixed polymer entities include the sequential polymerization of disparate monomer units via reversible-deactivation radical or anionic polymerization in a macroinitiator approach,4−6 these polymerization methods and/or the necessary initiators limit the choice of monomers and monomer combinations.7 The alternative modular approach directly addresses this limitation by exploiting efficient chemical reactions to link the active chainends of existing polymer segments.8 Thiol−ene reactions,9,10 multicomponent reactions (i.e., Biginelli, Ugi or Passerini),11,12 strain-promoted azide−alkyne cycloaddition of cyclooctynes (SPAAC)13 in addition to thermally and photochemically activated cycloaddition including (hetero-) Diels−Alder reactions14−17 are classic examples for the ligation of polymer © XXXX American Chemical Society

Received: February 19, 2018 Revised: March 17, 2018

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DOI: 10.1021/acs.macromol.8b00383 Macromolecules XXXX, XXX, XXX−XXX

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reduction reactions28 to the addition of nucleophiles29 and cycloadditions to dienes30 as well as 1,3-dipoles.31 In photochemistry,32−34 2H-azirine are powerful synthons with high quantum efficiency, even able to react in aqueous media in a bioorthogonal manner,35 and by participating in a rapid, catalyst-free visible light induced cycloadditions, when the 2Hazirine unit is tethered to pyrene.36 Concomitantly, there is a significant contemporary interest in miktoarm star polymer architectures, which stems from the wide property profile that can be established by the combination of virtually any type and number of polymer arms into a single unique architecture. Generally, miktoarm star polymers are synthetically challenging to access, since the polymer arms ideally vary in both molecular weight and chemical identity. Typically, multiple steps, protection/deprotection strategies, orthogonality, and combination of different polymerization methods are necessary for their successful synthesis, regardless of the specific type of miktoarm star polymer (e.g., ABC, A2B, AB2C2). In addition, self-assembly is a powerful avenue for controlling the complexity and hierarchy of these nanoscale materials, and promises to create a diverse range of distinctive properties. Thus, the provision of efficient synthetic routes that allow for the precision design of the building units is vital. Therefore, in the current contribution, we exploit the interrupted CuACC process for the construction of three-armed mikto star copolymers with a fluorescently labeled core via the simultaneously generated vinyl function as an orthogonally addressable reactive functional handle (Scheme 1b). By employing a pyrene-substituted azirine (Az-Py), the initial midreactive block copolymer formation can be witnessed by virtue of the optical properties of the block copolymers, i.e. the fluorescence of the pyrene moiety. Critically, the obtained block copolymer is further modified with thiol derivatives (either small molecule or thiol-terminated polymers) in a radical thiol−ene conjugation to form the noted miktoarm shaped architectures. By virtue of a detailed characterization via NMR, SEC, DLS, DSC, UV−vis, and fluorescence studies, we underpin the successful formation of the functional miktoarm star with a fluorescence labeled core. Furthermore, the solution properties of the multiarmed materials are established, demonstrating self-assembly behavior characteristic of unimolecular micelles, whose hydrophobic core (Py) may provide an ideal site to entrap molecular cargo.

Scheme 1. Copper(I)-Catalyzed Azide−Alkyne Cycloaddition Interrupted by 2H-Azirines: (a) Reaction Overview; (b) Two-Step Reaction Sequence to Miktoarmed Star Block Copolymersa

a

The initial step is the interrupted CuAAC reaction by Az-Py between PMMA-alkyne and PEG-N3. The subsequent step is the modification of the PMMA−PEG block copolymer with variable thiols via thiol− ene addition following a radical pathway.

careful literature survey indicates that 2H-azirines are interesting compounds due to their manifold features and reaction pathways.26 Their reactivity ranges from substitions,27

Figure 1. SEC (THF; A, RI detection; B, UV detection at 320 nm) traces of the block copolymer formation via interrupted CuAAC. B

DOI: 10.1021/acs.macromol.8b00383 Macromolecules XXXX, XXX, XXX−XXX

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Figure 2. (A) Comparative 1H NMR (400 MHz, CDCl3) spectra monitoring the block copolymer formation and synthesis of the ABC three-armed mikto star polymer μPMMA−PEG−PS. (B) COSY NMR spectra of PMMA−PEG and μPMMA−PEG−PS indicating the appearance of the resonances of the vinyl group after the interrupted CuAAC and their disappearance after thiol−ene addition of the PS block.

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RESULTS AND DISCUSSION In order to implement the Cu-catalyzed azide−alkyne cycloaddition interrupted by 2H-azirines and to quantify its coupling efficiency spectroscopically (specifically with 1H NMR to underpin the SEC measurements), polymer strands with low molecular weights were synthesized. Thus, we initially ligated a hydrophobic poly(methyl methacrylate) derivative (PMMAalkyne, Mn,SEC of 4000 g mol−1, and Đ of 1.1; refer to Figure 1A for SEC trace and Figure 2A for 1H NMR spectrum, respectively) and a readily available poly(ethylene glycol) methyl ether azide (PEG-N3, Mn,SEC = 2800 g mol−1, and Đ = 1.03; refer to Figure 1A for SEC trace and Figure 2A for 1H NMR spectrum, respectively). The well-defined PMMA-alkyne was synthesized via activators regenerated by electron transfer atom transfer radical polymerization (ARGET ATRP) (refer to Section A.2. in the Supporting Information for details).37 Subsequently, the interrupted CuAAC of PMMA-alkyne and PEG-N3 was exploited using 0.1 equiv of copper(I) iodide (CuI) and 1.4 equiv of N,N-diisopropylamine (DIPEA) as catalytic system in dichloromethane. In order to interrupt the click reaction, the pyrene-substituted azirine was mixed with the two polymer blocks (molar ratio: alkyne:N3:Az = 1.1:1.0:1.2.) in the presence of the catalyst at 0 °C. After 1 h, the mixture was thawed to ambient temperature and allowed to stand further for an additional 23 h (refer to Section A.3 in the Supporting Information for the detailed procedure). The small excess of PMMA-alkyne was readily removed via treatment with an azide-functionalized resin for 24 h at ambient temperature. The resulting PMMA−PEG amphiphilic block copolymer combining a spatially separated and orthogonally addressable reactive functional group (i.e., enamine) was isolated by precipitation in ice-cold hexane. The SEC trace of

the resulting block copolymer obtained via RI detection (refer to Figure 1A) indicates a monomodal distribution with Đ of 1.1 and a shift of 2500 g mol−1 compared to the PMMA-alkyne precursor, thus confirming the successful ligation of the two discrete blocks. Importantly, the intrinsic fluorescence of the pyrene moiety enables imaging the formation of the block copolymer by SEC measurements via UV detection. Since three strong bands in the UV region characterize the absorption spectrum of Py, i.e., at close to 334, 320, and 307 nm, the SEC traces were recorded at the fixed wavelength of 320 nm. The SEC chromatogram of the PMMA−PEG block copolymer clearly indicates a polymer distribution, while the PMMA and PEG blocks generate no signal at 320 nm (refer to Figure 1B). As further evidence, the copolymer has an excitation wavelength at 244, 279, and 344 nm and emission wavelength at 384 and 405 nm (refer to Figure S10A and B). Furthermore, the 1H NMR spectrum of PMMA−PEG (refer to Figure 2A) displays new magnetic resonances in the range of 5.35 to 4.99 and 4.52 ppm, which are associated with triazole formation, while no resonances for the alkyne group at 4.64 and 2.46 ppm are present compared to the 1H NMR spectrum of PMMA-alkyne (refer to Figure 2A). Likewise, the NMR results indicate the complete disappearance of the characteristic resonances for the CH2 group next to the azide group of PEG-N3 at 3.43 ppm. In addition, in the 1H NMR spectrum of the block copolymer, resonances associated with the PMMA backbone between 3.62 and 3.57 ppm and for PEG at 3.66 and 3.62 ppm are visible, as are resonances for the aromatic protons of pyrene between 9.00 and 8.00 ppm. Importantly, the resonances associated with the vinylic protons of the newly formed double bond appear at 4.57 and 4.05 ppm, respectively. For further confirmation of the block copolymer formation, ATR-IR spectra for Az-Py, PMMA-alkyne, PEG-N3, and PMMA−PEG were recorded C

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tion.38 Thus, the thermal analysis via DSC of the PMMA−PEG block copolymer indicates no separate melting and recrystallization transitions for the PMMA or the PEG block in the second heating scan (refer to Figure S12). However, a Tg at close to 50 °C associated with the PMMA block is detected. The glass transition temperature of the PMMA block may decrease, caused by the presence of the soft PEG block. The PMMA−PEG block copolymer bearing a defined and inherently generated orthogonally reactive group in its fluorescence labeled core provides the basis to form an architecturally complex macromolecule, i.e., the miktoarm star polymer. Miktoarm polymers, are a unique class of macromolecules. Critically, their properties can be tailored by varying their polymer arms. The thiol−ene reaction is a very versatile reaction in which a double bond reacts with a thiol forming a thiol-ether bond. In some cases, the reaction even meets the criteria of a click reaction.18 Hence, miktoarm stars with a fluorescently labeled core can be prepared via a subsequent thiol−ene addition to the newly introduced enamine bond of PMMA−PEG. The first modification of the amphiphilic block copolymer includes the model thiol−ene addition with a small molecule thiol derivative (i.e., the commercially available 1dodecanethiol, Dode-SH). The ligation was executed in its radical form with 1.5 equiv of azobis(isobutyronitrile) (AIBN) as radical source in 1,4-dioxane at 60 °C for 29 h. To verify the architecture of PMMA−PEG-Dode, the polymer was isolated via precipitation in hexane and analyzed using 1H NMR spectroscopy (Figure S5). The complete disappearance of the characteristic enamine resonances of PMMA−PEG at 4.57 and 4.05 ppm confirms the successful thiol−ene reaction. The SEC curve of the parent copolymers disappeared entirely (Figure S3), and a new peak corresponding to the modified PMMA− PEG-Dode copolymer appeared at higher molecular weights with narrow Đ (Mn,SEC = 7700 g mol−1, Đ = 1.1). Once the thiol−ene ligation conditions were optimized, a polystyrene thiol derivative (PS-SH) was synthesized via reversible addition−fragmentation chain transfer (RAFT) polymerization and subsequent aminolysis with hexylamine (Mn,SEC of 2600 g mol−1, Đ of 1.1, refer to Figure 2A for 1H NMR spectrum, and Figure 4 for the SEC trace). PS-SH was subsequently employed in the miktoarm ABC star polymer synthesis. Figure S1 displays the SEC traces of the mixture before the modification and after the thiol−ene addition. By comparison of the traces, it can be concluded that the PS-SH species at lower retention times

Figure 3. Comparative ATR-IR spectra of Az-Py, PMMA-alkyne, PEG-N3, and PMMA−PEG, monitoring the successful block copolymer formation.

(refer to Figure 3). The IR data of the resulting midreactive block copolymer illustrates that the characteristic C−H stretching vibration of the terminal alkyne group of PMMAalkyne (3250 cm−1) and the band for azides at 2100 cm−1 have completely disappeared, while the distinctive polymer backbone adsorptions of PMMA- and PEG-blocks are still present. Respectively, at 1724 and 1148 cm−1, the C = O and C−O stretching vibrations of PMMA, and at 1100 cm−1, the C−O− C stretch of the ether polymer backbone are visible. Concurrently, the specific absorptions of the Az-Py, namely at 3042 and 1585 cm−1, have vanished and the C = C stretching vibration at 1640 cm−1 can be assigned to the newly formed enamine group tethered to the triazole linkage. The architectural changes can also be established by DSC analysis. For instance, the linear polymer PMMA-alkyne is completely amorphous, with a glass transition point at close to 102 °C (refer to Figure S12), whereas pure PEG-N3 is a highly crystalline polymer (i.e., PEG-N3 with Mn = 2000 g mol−1, Tm of 45−50 °C). In general, an environment with 60 wt % or more PMMA affects the chain organization of the PEG block within a PMMA-PEG copolymer and inhibits its crystalliza-

Figure 4. SEC (THF; A, RI detection; B, UV detection at 320 nm) recorded of the different species during the modification process of PMMA− PEG with PS-SH, resulting in the ABC miktoarm star polymer as μPMMA−PEG−PS. D

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Figure 5. DLS studies of μPMMA−PEG−PEG in water at different temperatures (c = 2 g mL−1) (A) and different concentrations (T = 25 °C) (B) reveal the stability of the micelles against temperature and concentration changes similar to unimolecular micelles.

anchored in the pores of the SEC column packing and elute abnormally. On the other hand, we assume that the shape of the SEC elugram may be associated with the SEC conditions. It is noteworthy that others have also reported SEC traces of miktoarmed star polymers with a certain degree of highmolecular-weight shoulders and low-molecular-weight tailing, although using somewhat different polymers and experimental parameters.44−51 PEG-based amphiphilic miktoarm star-shaped polymers have been widely investigated for their potential applications in the biomedical field.52 Moreover, fluorescent drug nanocarriers are highly desirable for both in vitro and in vivo applications as their fluorescence properties allow for tracking of the nanocarriers using a variety of microscopy imaging techniques. In fact, unimolecular micellesamphiphilic molecules that are able to act as a host for guest molecules53can serve as drug delivery vectors. In general, unimolecular micelles exhibit increased stability (typically unimolecular micelles are covalent in nature)54 against dilution and temperature compared to multimolecular systems that dissociate below the critical micelle concentration (CMC),55 therefore preventing the typically observed dynamic equilibrium between the individual amphiphilic molecules and the micelle. Here, however, due to the unique structure of the fluorescence labeled star polymers with arms of different polarity ranging from hydrophilic (i.e., PEG) to highly hydrophobic (i.e., PS), the synthesized miktoarm polymers are ideally suited for self-assembly in water. Thus, the miktoarm polymer μPMMA−PEG−PEG was dispersed in water, while a nanoprecipitation procedure was adopted for μPMMA−PEG−PS and the focal point modified block copolymer PMMA−PEG-Dode (for details refer to the DLS section in the Supporting Information). Dynamic light scattering (DLS) enables the investigation of the size of the self-assembled polymers and can be used to explore the impact of temperature and dilution on the particle size. For μPMMA− PEG−PS, the hydrophobic impact of the PMMA and PS arms disallowed the formation of stable micelles in water. For μPMMA−PEG−PEG and PMMA−PEG-Dode, particles with average hydrodynamic radii (DH) of 18.3 and 42.7 nm were identified at 25 °C, respectively. We speculate that PMMA− PEG-Dode is more hydrophobic, thus more amphiphilic molecules are required to assemble the hydrophilic shell around the PMMA arm and the dodecyl chain leading to larger micelles. For μPMMA−PEG−PEG, the two PEG arms are

vanishes, confirming the successful attachment of the PS arm to generate the μPMMA−PEG−PS miktoarm star polymer. The SEC evaluation of the miktoarm polymer results in a polymer with a Mn,SEC of 6600 g mol−1 and a Đ of 1.2 (refer to Figure 4A). The increment in the apparent weight-average molecular weight of the miktoarm polymer was insignificant in comparison to the block copolymer, consistent with the former having a star-shaped architecture and thus a smaller hydrodynamic volume than the latter. Note that the PS arm alone can form sandwich-like excimers through the association between intramolecular excited and unexcited phenyl groups along the backbone.39−43 Moreover, due to the interactions with the pyrene moiety in the core of the miktoarm polymer, the PS chain may even coil more effectively, leading to a lower hydrodynamic radius and therefore to the decreased apparent molecular weight value observed via SEC. The 1H NMR spectrum (refer to Figure 2A) of the precipitated polymer unambiguously confirms the successful formation of the ABC miktoarm polymer with a fluorescence labeled core. The magnetic resonances arising from the protons of the enamine unit at 4.57 and 4.05 ppm disappeared, while the new resonances for the thio ether bond collide with the resonances of the PMMA- and PEG-arms that lie in the range of 4.00 to 3.00 ppm. Additional analysis via a 2D NMR experiment (i.e., COSY) reveal cross signals for the enamine protons of PMMA−PEG as shown in the zoom-in (upper inset) of the COSY spectrum (refer to Figure 2B). Once the thiol−ene addition takes place, the cross signals are no longer detectable (dashed boxed area), further underpinning the successful formation of μPMMA−PEG−PS. We have additionally altered the synthetic route to the miktoarm star polymer by performing the thiol−ene reaction with a thiol derivative of PEG (PEG-SH, Mn,SEC = 2600 g mol−1,Đ = 1.1) to afford miktoarm AB2 stars. Figure S4A indicates that the SEC trace of the μPMMA−PEG−PEG species (Mn = 6300 g mol−1, Đ = 1.2) eluted at longer elution times than the monomodal SEC trace of its linear block copolymer analogue and the 1H NMR spectrum in Figure S6 shows the same characteristics as PMMA−PEG-Dode. As noted, this is consistent with the former one featuring a starshaped architecture and therefore a smaller hydrodynamic volume than the initial block copolymer.44 The SEC elugram of the miktoarm polymer additionally shows some low molecular weight tailing, which may be associated with the following two factors. On the one hand, the arms may be temporarily E

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process is a powerful addition to the synthetic toolbox of soft matter material science, chemical biology (i.e., fluorescence probing) or micelle formation for delivery systems.

more effective in prompting a hydrophilic shell around the PMMA arm independently, resulting in smaller micelles.56 To deliver polymeric unimolecular micelles effectively for biomedical applications, overcoming biological barriers in vivo such as serum instability and degradation susceptibility is prerequisite to fully achieving their therapeutic potential. In addition, upon intravenous injection, usually, the unimolecular micelles undergo a drastic dilution in the bloodstream and are exposed to a variety of serum proteins, which induce micelle dissociation and reduce circulation time. Moreover, the longterm stability for unimolecular micelles is commonly performed by storing them in three different environments at 4 ± 1 °C, ambient temperature, and 45 °C. Accordingly, we assessed the influence of temperature and dilution on the micelle integrity. By altering the temperature by 20 °C (refer to Figure 5A and S8), the average diameter of μPMMA−PEG−PEG changes in the range of 0.2 (1%) and 2.0 (11%) nm, respectively for 5 and 45 °C, thus showing that micelles of μPMMA−PEG−PEG retain their structural integrity. In contrast, the DLS analysis for PMMA−PEG-Dode reveals particles whose sizes deviates by 4% (1.8 nm) when the temperature is increased to 45 °C, and by 58% (24.7 nm) once the temperature is decreased to 5 °C (refer to Figure S8). This large difference is likely associated with hydrophobic interactions of Dode with the PMMA-block induced by the decrease in the temperature. Critically, dilution of the initial micelles of μPMMA−PEG−PEG by a factor of 50 has negligible impact on the morphological reorganization (refer to Figure 5B and S9), thus maintaining stable micelle formation with a standard deviation of 1.2 nm (7%). Interestingly, PMMA−PEG-Dode displays similar results (i.e., maximal deviations of 5%), indicating that dilution induces minimal changes in size (refer to Figure S9). Thus, the μPMMA−PEG−PEG based micelles maintain their structural integrity as a function of temperature and concentration, thus showing characteristics of unimolecular micelles while not being covalently connected. The herein introduced micellar constructs maintain their structural integrity against temperature changes and dilution, which is of high interest for an effective payload transport.

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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/acs.macromol.8b00383. Experimental section and additional characterization data (PDF)

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AUTHOR INFORMATION

Corresponding Authors

*(H.M.) E-mail: [email protected]. *(C.B.-K.) E-mail: [email protected], [email protected]. ORCID

Hatice Mutlu: 0000-0002-4683-0515 Christopher Barner-Kowollik: 0000-0002-6745-0570 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS C.B.-K. acknowledges continued support from the Karlsruhe Institute of Technology (KIT) in the context of the Helmholtz BioInterfaces in Technology and Medicine (BIFTM) and Science and Technology of Nanosystems (STN) programs as well as from the Queensland University of Technology (QUT) and the Australian Research Council (ARC) in the form of a Laureate Fellowship. J.T.O.’s Ph.D. studies are funded by the Fonds der Chemischen Industrie (FCI) as well as the Karlsruhe Institute of Technology (KIT). The authors thank Dr. P. Levkin for access to DLS equipment and Prof. M. Meier (both KIT) for access to ATR-IR and DSC instruments.

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REFERENCES

(1) Discher, D. E.; Eisenberg, A. Polymer Vesicles. Science 2002, 297, 967−973. (2) Riess, G. Micellization of Block Copolymers. Prog. Polym. Sci. 2003, 28, 1107−1170. (3) Shin, S.; Yoon, K. Y.; Choi, T. L. Simple Preparation of Various Nanostructures via in Situ Nanoparticlization of Polyacetylene Blocklike Copolymers by One-Shot Polymerization. Macromolecules 2015, 48, 1390−1397. (4) Davis, K. A.; Charleux, B.; Matyjaszewski, K. Preparation of Block Copolymers of Polystyrene and Poly (T-Butyl Acrylate) of Various Molecular Weights and Architectures by Atom Transfer Radical Polymerization. J. Polym. Sci., Part A: Polym. Chem. 2000, 38, 2274− 2283. (5) Epps, T. H., III; O’Reilly, R. K. Block Copolymers: Controlling Nanostructure to Generate Functional Materials − Synthesis, Characterization, and Engineering. Chem. Sci. 2016, 7, 1674−1689. (6) Lowe, A. B. Thiol-Ene “Click” Reactions and Recent Applications in Polymer and Materials Synthesis. Polym. Chem. 2010, 1, 17−36. (7) Quémener, D.; Davis, T. P.; Barner-Kowollik, C.; Stenzel, M. H. RAFT and Click Chemistry: A Versatile Approach to Well-Defined Block Copolymers. Chem. Commun. 2006, 48, 5051−5053. (8) Hillmyer, M. Block Copolymer Synthesis. Curr. Opin. Solid State Mater. Sci. 1999, 4, 559−564. (9) Boyer, C.; Granville, A.; Davis, T. P.; Bulmus, V. Modification of RAFT-Polymers via Thiol-Ene Reactions : A General Route to Functional Polymers and New Architectures. J. Polym. Sci., Part A: Polym. Chem. 2009, 47, 3773−3794.

CONCLUSIONS In summary, we provide an efficient and simple approach to fluorescent midchain reactive (i.e., enamine-functionalized) 1,2,3-triazole block copolymers by exploiting the efficient Cucatalyzed interrupted reaction of 2H-azirines with readily synthesized or commercially available homopolymer segments. Because of their defined midfunctionality, such structures are well-suited for the construction of miktoarm star polymers. Thus, the inherently generated double bond was reacted with thiol functionalized (macro)molecules in radical driven thiol− ene reactions. ABC- and AB2-miktoarm star architectures with chromophores in their core are thus readily accessible. Moreover, the miktoarm star polymers display the stability characteristics of unimolecular micelles making them prime candidates for delivery systems as they are stable against temperature and concentration changes. Furthermore, our micellar constructs are inherently fluorescent, as they contain a Py unit. We submit that the Cu-catalyzed interrupted reaction of 2H-azirines can not only be exploited as a conventional multicomponent reaction to efficiently ligate two polymers to quantitatively generate copolymers, yet also allows the introduction of different functionalities at the ligation points due to its multicomponent nature. Thus, the herein presented F

DOI: 10.1021/acs.macromol.8b00383 Macromolecules XXXX, XXX, XXX−XXX

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Macromolecules (10) Koo, P. S. K.; Stamenovic, M. M.; Prasath, R. A.; Inglis, A. J.; Du Prez, F. E.; Barner-Kowollik, C.; Van Camp, W.; Junkers, T. Limitations of Radical Thiol-Ene Reactions for Polymer − Polymer Conjugation. J. Polym. Sci., Part A: Polym. Chem. 2010, 48, 1699−1713. (11) Oelmann, S.; Solleder, S. C.; Meier, M. A. R. Controlling Molecular Weight and Polymer Architecture during the Passerini Three Component Step-Growth Polymerization. Polym. Chem. 2016, 7, 1857−1860. (12) Zhu, C.; Yang, B.; Zhao, Y.; Fu, C.; Tao, L.; Wei, Y. A New Insight into the Biginelli Reaction: The Dawn of Multicomponent Click Chemistry? Polym. Chem. 2013, 4, 5395−5400. (13) Glassner, M.; Maji, S.; de la Rosa, V. R.; Vanparijs, N.; Ryskulova, K.; De Geest, B. G.; Hoogenboom, R. Solvent-Free Mechanochemical Synthesis of a Bicyclononyne Tosylate : A Fast Route towards Bioorthogonal Clickable Poly(2-Oxazolines)s. Polym. Chem. 2015, 6, 8354−8359. (14) Delaittre, G.; Guimard, N. K.; Barner-Kowollik, C. Cycloadditions in Modern Polymer Chemistry. Acc. Chem. Res. 2015, 48, 1296−1307. (15) Pahnke, K.; Brandt, J.; Gryn’ova, G.; Lin, C. Y.; Altintas, O.; Schmidt, F. G.; Lederer, A.; Coote, M. L.; Barner-Kowollik, C. Polymer Chain Cleavage Entropy-Driven Selectivity for Chain Scission : Where Macromolecules Cleave Angewandte. Angew. Chem., Int. Ed. 2016, 55, 1514−1518. (16) Inglis, A. J.; Sinnwell, S.; Stenzel, M. H.; Barner-Kowollik, C. Ultrafast Click Conjugation of Macromolecular Building Blocks at Ambient Temperature. Angew. Chem., Int. Ed. 2009, 48, 2411−2414. (17) Gruendling, T.; Oehlenschlaeger, K. K.; Frick, E.; Glassner, M.; Schmid, C.; Barner-Kowollik, C. Rapid UV Light-Triggered Macromolecular Click Conjugations via the Use of O-Quinodimethanes. Macromol. Rapid Commun. 2011, 32, 807−812. (18) Barner-Kowollik, C.; Du Prez, F. E.; Espeel, P.; Hawker, C. J.; Junkers, T.; Schlaad, H.; Van Camp, W. “Clicking” Polymers or Just Efficient Linking: What Is the Difference? Angew. Chem., Int. Ed. 2011, 50, 60−62. (19) Blasco, E.; Sims, M. B.; Goldmann, A. S.; Sumerlin, B. S.; Barner-Kowollik, C. 50th Anniversary Perspective : Polymer Functionalization. Macromolecules 2017, 50, 5215−5252. (20) Wei, F.; Wang, W.; Ma, Y.; Tung, C.; Xu, Z. Regioselective Synthesis of Multisubstituted 1,2,3-Triazoles: Moving beyond the Copper-Catalyzed Azide-Alkyne Cycloaddition. Chem. Commun. 2016, 52, 14188−14199. (21) Ackermann, L.; Potukuchi, H. K. Regioselective Syntheses of Fully-Substituted 1,2,3-Triazoles : The CuAAC/C−H Bond Functionalization Nexus. Org. Biomol. Chem. 2010, 8, 4503−4513. (22) Ackermann, L.; Potukuchi, H. K.; Landsberg, D. CopperCatalyzed “Click” Reaction/Direct Arylation Sequence : Modular Syntheses of 1,2,3-Triazoles. Org. Lett. 2008, 10, 3081−3084. (23) Schwartz, E.; Breitenkamp, K.; Fokin, V. V. Synthesis and Postpolymerization Functionalization of. Macromolecules 2011, 44, 4735−4741. (24) Wang, W.; Peng, X.; Wei, F.; Tung, C.; Xu, Z. Copper (I)Catalyzed Interrupted Click Reaction: Synthesis of Diverse 5-HeteroFunctionalized Triazoles. Angew. Chem., Int. Ed. 2016, 55, 649−653. (25) Zhou, W.; Zhang, M.; Li, H.; Chen, W. One-Pot ThreeComponent Synthesis of Enamine-Functionalized 1,2,3-Triazoles via Cu-Catalytic Azide − Alkyne Click (CuAAC) and Cu - Catalyzed Vinyl Nitrene Transfer Sequence. Org. Lett. 2017, 19, 10−13. (26) Khlebnikov, A. F.; Novikov, M. S. Recent Advances in 2H -Azirine Chemistry. Tetrahedron 2013, 69, 3363−3401. (27) Pinho e Melo, T. M. V. D.; Gonsalves, A. M. d’A. R.; Lopes, C. S. J. Synthesis and Reactivity of 2-Halo-2H-Azirines towards Nucleophiles. Tetrahedron Lett. 2000, 41, 7217−7220. (28) Palacios, F.; Ochoa de Retana, A. M.; Gil, J. I.; Ezpeleta, J. M. Simple Asymmetric Synthesis of 2H-Azirines Derived from Phosphine Oxides. J. Org. Chem. 2000, 65, 3213−3217. (29) Pinho e Melo, T. M. V. D.; Lopes, S. J.; Beja, A. M.; Paixao, A. Reactivity of 2-Halo-2H-Azirines. 1. Reactions with Nucleophiles. J. Org. Chem. 2002, 67, 66−71.

(30) Dubinina, G. G.; Yoshida, W. Y.; Chain, W. J. On the Preparation of Azepinones. Tetrahedron Lett. 2010, 51, 5325−5327. (31) Brown, D.; Brown, G. A.; Andrews, M.; Large, J. M.; Urban, D.; Butts, C. P.; Hales, J.; Gallagher, T. The Azomethine Ylide Strategy for -Lactam Synthesis. Azapenams and 1-Azacephams. J. Chem. Soc., Perkin Trans. 2002, 1, 2014−2021. (32) Padwa, A. Azirine Photochemistry. Acc. Chem. Res. 1976, 9, 371−378. (33) Padwa, A.; Smolanoff, J. Photocycloaddittion of Arylazirenes. J. Am. Chem. Soc. 1971, 93, 548−550. (34) Albrecht, E.; Mattay, J.; Steenken, S. [3 + 2] Cycloadditions and Protonation by Alcohols of Photochemically Generated Nitrile Ylides from 2H -Azirines. Formation and Reactivities of Azaallenium Cations. J. Am. Chem. Soc. 1997, 119, 11605−11610. (35) Lim, R. K. V.; Lin, Q. Azirine Ligation: Fast and Selective Protein Conjugation via Photoinduced Azirine − Alkene Cycloaddition W. Chem. Commun. 2010, 46, 7993−7995. (36) Mueller, J. O.; Schmidt, F. G.; Blinco, J. P.; Barner-Kowollik, C. Visible-Light-Induced Click Chemistry. Angew. Chem., Int. Ed. 2015, 54, 10284−10288. (37) Kwak, Y.; Matyjaszewski, K. ARGET ATRP of Methyl Methacrylate in the Presence of Nitrogen-Based Ligands as Reducing Agents. Polym. Int. 2009, 58, 242−247. (38) Chen, N.; Yan, L. T.; Xie, X. M. Interplay between Crystallization and Phase Separation in PS-B-PMMA/PEO Blends: The Effect of Confinement. Macromolecules 2013, 46, 3544−3553. (39) Yanari, S. S.; Bovey, F. A.; Lumry, R. Fluorescence of Styrene Homopolymers and Copolymers. Nature 1963, 200, 242−244. (40) Vala, M. T.; Haebig, J.; Rice, S. a. Experimental Study of Luminescence and Excitation Trapping in Vinyl Polymers, Paracyclophanes, and Related Compounds. J. Chem. Phys. 1965, 43, 886− 897. (41) Nishiyama, N. Nanomedicine: Nanocarriers Shape up for Long Life. Nat. Nanotechnol. 2007, 2, 203−204. (42) Kloepffer, W. T. Energy Transfer in Films of Polymers with Aromatic Side-Groups. Ann. N. Y. Acad. Sci. 1981, 366, 373−386. (43) Hirayama, F. Intramolecular Excimer Formation. I. Diphenyl and Triphenyl Alkanes. J. Chem. Phys. 1965, 42, 3163−3171. (44) Cui, S.; Pan, X.; Gebru, H.; Wang, X.; Liu, J.; Liu, J.; Li, Z.; Guo, K. Amphiphilic Star-Shaped Poly(sarcosine)-Block-Poly(ε-Caprolactone) Diblock Copolymers: One-Pot Synthesis, Characterization, and Solution Properties. J. Mater. Chem. B 2017, 5, 679−690. (45) Wang, F.; Bronich, T. K.; Kabanov, A. V.; Rauh, R. D.; Roovers, J. Synthesis and Evaluation of a Star Amphiphilic Block Copolymer from Poly(e-Caprolactone) and Poly(ethylene Glycol) as a Potential Drug Delivery Carrier. Bioconjugate Chem. 2005, 16, 397−405. (46) Ozlem, S.; Iskin, B.; Yilmaz, G.; Kukut, M.; Hacaloglu, J.; Yagci, Y. Synthesis and Pyrolysis of ABC Type Miktoarm Star Copolymers with Polystyrene, Poly(lactic Acid) and Poly(ethylene Glycol) Arms. Eur. Polym. J. 2012, 48, 1755−1767. (47) Iskin, B.; Yilmaz, G.; Yagci, Y. Synthesis of ABC Type Miktoarm Star Copolymers by Triple Click Chemistry. Polym. Chem. 2011, 2, 2865−2871. (48) Hong, L.; He, J.; Chen, Y.; Kakuchi, T. Synthesis of ABB′ and ABC Star Copolymers: Via a Combination of NMRP and ROP Reactions. Polym. Chem. 2016, 7, 3599−3607. (49) Gao, H.; Matyjaszewski, K. Synthesis of Functional Polymers with Controlled Architecture by CRP of Monomers in the Presence of Cross-Linkers: From Stars to Gels. Prog. Polym. Sci. 2009, 34, 317− 350. (50) Bahadori, F.; Dag, A.; Durmaz, H.; Cakir, N.; Onyuksel, H.; Tunca, U.; Topcu, G.; Hizal, G. Synthesis and Characterization of Biodegradable Amphiphilic Star and Y-Shaped Block Copolymers as Potential Carriers for Vinorelbine. Polymers 2014, 6, 214−242. (51) Altintas, O.; Hizal, G.; Tunca, U. ABC-Type Hetero-Arm Star Terpolymers Through “Click” Chemistry. J. Polym. Sci., Part A: Polym. Chem. 2006, 44, 5699−5707. G

DOI: 10.1021/acs.macromol.8b00383 Macromolecules XXXX, XXX, XXX−XXX

Article

Macromolecules (52) Choi, Y. K.; Bae, Y. H.; Kim, S. W. Star-Shaped Poly(etherEster) Block Copolymers: Synthesis, Characterization, and Their Physical Properties. Macromolecules 1998, 31, 8766−8774. (53) Newkome, G. R.; Yao, Z.; Baker, G. R.; Gupta, V. K. Micelles. Part 1. Cascade Molecules: A New Approach to Micelles. A [27]Arborol. J. Org. Chem. 1985, 50, 2003−2004. (54) Chen, G.; Wang, L.; Cordie, T.; Vokoun, C.; Eliceiri, K. W.; Gong, S. Multi-Functional Self-Fluorescent Unimolecular Micelles for Tumor-Targeted Drug Delivery and Bioimaging. Biomaterials 2015, 47, 41−50. (55) Wang, Y.; Li, L.; Li, J.; Yang, B.; Wang, C.; Fang, W.; Ji, F.; Wen, Y.; Yao, F. Stable and pH-Responsive Polyamidoamine Based Unimolecular Micelles Capped with a Zwitterionic Polymer Shell for Anticancer Drug Delivery. RSC Adv. 2016, 6, 17728−17739. (56) Hashidzume, A.; Mizusaki, M.; Yoda, K.; Morishima, Y. Interaction of Unimolecular Micelles of Hydrophobically-Modified Polyelectrolytes with Nonionic/ionic Mixed Surfactant Micelles. Langmuir 1999, 15, 4276−4282.

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DOI: 10.1021/acs.macromol.8b00383 Macromolecules XXXX, XXX, XXX−XXX