Engineering Programmable Synthetic Vesicles with

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Engineering Programmable Synthetic Vesicles with Permeability Regulated by a Single Molecular Bridge Gong Cheng, and Juan Pérez-Mercader Chem. Mater., Just Accepted Manuscript • DOI: 10.1021/acs.chemmater.9b01635 • Publication Date (Web): 10 Jul 2019 Downloaded from pubs.acs.org on July 18, 2019

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Chemistry of Materials

Engineering Programmable Synthetic Vesicles with Permeability Regulated by a Single Molecular Bridge Gong Cheng, *, † and Juan Pérez-Mercader *, †, ‡ Department of Earth and Planetary Sciences and Origins of Life Initiative Harvard University, 20 Oxford Street, Cambridge, Massachusetts 02138, United States, ‡ Santa Fe Institute, Santa Fe, New Mexico 87501, United States †

ABSTRACT: Self-assembly of biomimetic vesicles has broad impacts in the development of functional materials and applications to a variety of fields. Although promising, facile construction of function programmable synthetic vesicles to meet special requirements is still in its infancy and highly desirable. We developed novel polymeric vesicles with photo/chemo programmable permeability through an aqueous based one-pot polymerization induced self-assembly (PISA) strategy. This is realized by using a newly designed macro chain transfer agent and installing a single azobenzene moiety to bridge the hydrophilic and hydrophobic blocks of the amphiphiles. These polymeric vesicles can realize either release of small molecules only by light-induced selective permeability or burst release of all contents from the lumen by chemoinduced degradation of the amphiphiles and disassembly of the vesicular structure. The proof-of-concept principles are demonstrated and supported by monitoring the structural evolution of these biomimetic vesicles. This work offers new inspiration for the design of “smart” systems in nanoreactors, drug delivery and molecular storage.

INTRODUCTION Cells and their internal organelles in extant living systems are compartmentalized structures spatially segregated by plasma membranes with controlled permeability for precise mass transport.1 These membranes are made with fatty acid amphiphilic molecules produced during metabolism. On the other hand, polymeric vesicles are ideal candidates for mimicking natural compartments, which have a lumen segregated from the external environment by an assembled membrane.2-4 Mimicking the biological compartmentalization for design and assembly of the polymeric vesicles with stimulus-responsive properties have recently attracted particular attention in fields of origin of life, artificial organelles, nanoreactors, and molecular delivery strategies.5-9 These “smart” artificial systems can sense changes in the environment and adjust their assembly structures to achieve interesting properties.10-14 In a living cell, the cell membrane is a sophisticated stimuli-responsive system.15 The cell membrane can either be triggered to be selectively permeable to small molecules but blocks the passage of certain macromolecules or be disassembled by the degradation of the lipid amphiphiles to lead to burst release of all cellular contents at the end of their lifetime.16-17 Therefore, developing new types of stimulus-responsive polymeric vesicles to mimic cell membrane behaviors can contribute to elucidate some basic biological activities and their function in a simplified and controlled way. In addition, the development of synthetic stimulus-responsive vesicles endowed with either selective release or burst release is well-suited to the design and implementation of artificial bioreactors and controlled drug release.18-19

concentrations and involves multiple steps.23 Recently, polymerization-induced self-assembly (PISA) techniques have become an efficient approach to construct high concentrated polymeric vesicles directly from a homogeneous precursor mixture in aqueous solution.24-30 It provides an autonomous, facile and highly efficient route for preparing functional vesicles31-33 and loading cargoes in one step.34-38 In the following, we report and demonstrate a strategy to construct novel photo/chemo-responsive polymeric vesicles using PISA. These vesicles can realize either release of small molecules only by light-induced selective permeability, or burst release of all of their contents from the lumen by chemo-induced hypoxic degradation of the amphiphiles and disassembly of the vesicular structure (Scheme 1). This was achieved by synthesizing a novel macromolecular reversible additionfragmentation chain transfer (macroRAFT) agent with a single molecular azobenzene (AZO) moiety to function as a switch. During PISA, the polymeric vesicles were formed, and, simultaneously, a layer of the responsive molecular gate was installed at the interface of the vesicle membrane. Notably, we only use a single AZO moiety to bridge the hydrophobic and hydrophilic blocks of the amphiphiles rather than numerous AZO moieties contained in a polymer as a block. As a consequence, the polymeric vesicles can mimic the cell membrane system in a very simplified way to enable different release behaviors and mechanism upon photo/chemo stimuli. The stimulus-response mechanism and morphology evolution of the polymeric nanovesicles have been studied, and selective photo/chemo-responsive release of small molecular and macromolecular cargoes have been tested and investigated.

Conventionally, stimulus-responsive polymeric vesicles are prepared based on postpolymerization self-assembly strategies of their molecular (i.e. amphiphiles) components.2022 However, this strategy is limited to very low polymer

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mixture was dialyzed in water to remove the solvent and salts. The product solution was lyophilized to obtain the final CTAAZO-PEG.

Scheme 1. One-pot synthesis of amphiphilic block polymers, vesicular assembly, and molecular cargo loading through polymerization induced self-assembly (PISA). The vesicle membrane is interfaced by the azobenzene (AZO) moieties. It enables both the size-dependent release by the photostimulus (1) and burst release by chemo-induced disassembly of the vesicular structure (2).

EXPERIMENTAL SECTION Materials. 4-Cyano-4-(phenylcarbonothioylthio)pentanoic acid N-succinimidyl ester (CTA-NHS), triethylamine (TEA), copper(I) bromide (CuBr), rhodamine 6g (Rh6G), 2Hydroxypropyl methacrylate (HPMA), lithium phenyl-2,4,6trimethyl benzoyl phosphinate (LP), fluorescein isothiocyanate-dextran (FITC-dextran), sodium dithionite (SD), N,N-dimethylformamide (anhydrous, DMF), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Alkyne terminated poly(ethylene glycol) (mPEG-alkyne, 5k) was purchased from Biochempeg Scientific Inc. (Watertown, MA, USA). All the chemical agents were used without further purification. Synthesis of the AZO interfaced poly(ethylene glycol) reversible addition-fragmentation chain transfer (CTA-AZOPEG). The CTA-AZO-PEG was synthesized by a two-step strategy (Figure S1). The heterobifunctional AZO moiety terminated with amine and azide terminated groups (N-(2Aminoethyl)-4-{-2-[4-(3-azido-propoxy)-phenyl]-vinyl}benzamide hydrochloride) was synthesized according to the previous report.39 First, an N-hydroxysuccinimide activated ester of 4-cyano-4-(phenylcarbonothioylthio) pentanoic acid (CTA-NHS) was conjugated to the terminal primary amine group on the heterobifunctional AZO moiety via carbodiimide coupling. Then, the alkyne-terminated poly(ethylene glycol) was covalently linked to form the CTA by click chemistry. Briefly, 100 mg of N-(2-Aminoethyl)-4-{-2-[4-(3-azidopropoxy)-phenyl]-vinyl}-benzamide hydrochloride and 141 mg of CTA-NHS were dissolved in 1 mL and 400 µL of DMF, respectively. The CTA-NHS solution was added in the other solution under vigorous agitation and followed by addition of 12 µL of triethylamine. The mixture was further stirred overnight. 1 g of mPEG-alkyne was dissolved in 0.5 mL DMSO and mixed with the above solution. Then, 35 mg of CuBr and additional 18 µL of triethylamine were added. The mixture was stirred under nitrogen atmosphere for 4 hours. Afterward, the

RAFT aqueous dispersion polymerization for one-pot synthesis and self-assembly of polyHPMA-AZO-PEG (PHAP) polymeric vesicles. One-pot step synthesis and self-assembly of PHAP vesicles were conducted in an aqueous solution using the PISA strategy.24, 35 Briefly, polymerization of the water-soluble monomer (2-hydroxypropyl methacrylate, HPMA) to the CTAAZO-PEG as the hydrophobic block (target degree of polymerization: 300) was carried out at room temperature (Figure 1A). 13.1 mg of CTA-AZO-PEG and 100 mg of HPMA were dissolved in 1 mL of H2O. Then, 800 µL of LP aqueous solution (4.2 mg mL-1) was added. The mixture was then purged with nitrogen for 30 minutes, and followed by stirring at room temperature under blue LED irradiation (405 nm) for one hour. After the reaction, the prepared polymeric vesicles were collected for NMR and TEM characterization. To load the molecular cargoes, 0.2 mg of Rh6G (small molecular cargo) or 1.0 mg of FITC-dextran (macromolecular cargo; Mw=70kDa) was added to the mixture of reactants and followed by the similar procedure to generate the polymeric vesicles with loaded cargoes. The polymeric vesicles with loaded molecular cargoes were isolated and purified by centrifugation and washing to remove unencapsulated molecular cargoes. UV-Vis spectrometry characterization of the synthesized PHAP block polymer and their photoisomerization. PHAP block polymers were collected by centrifugation of prepared polymeric vesicles, followed by wash and lyophilization. Then, the dry polymers were dissolved in methanol and the UV-Vis spectrum was recorded. For recording the UV-Vis spectrum of PHAP at cis status, the PHAP solution was exposed by a UV lamp (wavelength: 365; power: 100 mW) for half an hour and then measured directly. The dynamic profile of photoisomerization of the block polymer solution was conducted by measurement of the absorbance at 355 nm with decided intervals (30 seconds or 60 seconds) under UV light irradiation (UV) or ambient light (Vis). For cycling experiments, the polymer solutions were irradiated by UV light for 15 minutes, and immediately placed in the UV-Vis spectrometer to measure the absorbance at 355 nm, and then their recovery under ambient light was monitored in a similar way. This protocol was repeated 5 times. Photo-responsive and size-selective release of small molecular cargoes. To check the UV light-induced the release of small molecules from prepared polymeric vesicles, Rh6G dye molecules were caged in polymeric vesicles as described above. The purified vesicles with loaded Rh6G molecules were redispersed in aqueous solution with the final concentration of 10 mg mL-1. The vesicle dispersions in a constant temperature quartz cell with thermostated bath circulating water were either irradiated with a UV lamp (wavelength: 365; power: 100 mW) or covered by a foil paper at room temperature. Started from the beginning, 300 µL of dispersion was taken out at the decided intervals, and after immediate centrifugation, the supernatant was collected for monitoring the released Rh6G by fluorescence measurement (excitation wavelength: 490 nm, emission wavelength 550 nm). The concentration of released Rh6G molecules was determined by a calibration curve. As a comparison, the permeability of prepared polymeric vesicles for macromolecules was also determined. Similarly, the FITCdextran molecules were caged in polymeric vesicles as described above. The purified vesicles with loaded FITCdextran molecules were redispersed in aqueous solution with the final concentration of 10 mg mL-1. The vesicle dispersions


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Chemistry of Materials were either irradiated with a UV lamp (100 mW) or covered by a foil paper at room temperature. Started from the beginning, 300 µL of dispersion was taken out at the decided intervals, and after immediate centrifugation, the supernatant was collected for determination of the released FITC-dextran molecules by fluorescence measurement (excitation wavelength: 465 nm, emission wavelength 515 nm). The concentration of released FITC-dextran molecules was determined by a calibration curve. The hypoxia-responsive release of molecular cargoes. To study the hypoxia-induced degradation and disassembly, the PHAP vesicles were incubated with SD solution, and 50 µL of suspension was taken out for TEM sample preparation at the determined time. After exposure to SD for 24 hours, the sample was collected for DLS analysis. For measurement of UV-Vis spectra, the suspension was lyophilized and dissolved in methanol. To test the hypoxia-responsive release of guest molecules from polymeric vesicles through vesicle disassembly under hypoxia circumstance, we first studied the influence of the SD concentration on the release of guest molecules. Polymeric vesicles with caged guest molecules (Rh6G) were incubated with sodium dithionite (SD) of different concentrations (0, 5, 10, 25, 50 mM) for 6 hours. The fluorescence spectra corresponding to the released Rh6G were recorded. Then, we studied the release profile of both the small molecular guest (Rh6G) and the macromolecule guest (FITCdextran). More specifically, Rh6G molecules and FITC-dextran molecules were caged in the polymeric vesicles as described above, respectively. The purified vesicles with loaded guest molecules were redispersed in either an aqueous solution or SD solution (25 mM, mimicking the hypoxia condition) with the final concentration of 10 mg mL-1. The vesicle dispersions were covered by a foil paper and stirred at room temperature. Started from the beginning, 300 µL of dispersion was taken out at the decided intervals, and after immediate centrifugation, the supernatant was collected for monitoring the released molecular cargoes by fluorescence measurement (Rh6G: excitation wavelength: 490 nm, emission wavelength 550 nm; FITC-dextran: excitation wavelength: 465 nm, emission wavelength 515 nm;). The concentration of released molecular cargoes was determined by the calibration curves. Characterization. Transmission Electron Microscopy (TEM) was used to characterize the morphology of the samples. The copper grids (carbon coated, TED PELLA) were immersed in the aqueous dispersion of samples (0.2% w/v), followed by Vanadium negative stain using NanoVan (Nanoprobes). Imaging was performed at 120/100 kV on a FEI Tecnai F20 microscope or a Hitachi HT7800 Microscope. DLS measurements of the aqueous dispersion of samples were conducted at 25 °C using a Malvern Instruments Zetasizer Nano. For NMR, polymers were dissolved in Methanol-D4 (SigmaAlrich) and analyzed by 1H NMR spectroscopy on a Varian INOVA 500 (I500B) NMR Spectrometer. Gel permeation chromatography (GPC) analysis was conducted on an Agilent 1260 system equipped with a refractive index detector, and DMF was used as the eluent at a flow rate of 1.0 mL min-1 at 50 °C. UV-Vis spectra were recorded at room temperature on an S2100UV+ UV/Vis spectrophotometer (UNICO). Fluorescence spectra were recorded on a Synergy H1 Hybrid Multi-Mode Reader (BioTek Instruments, Inc.).

RESULTS AND DISCUSSION A new macroRAFT agent (CTA-AZO-PEG) with a single molecular AZO moiety to bridge the RAFT and poly(ethylene glycol) was designed and synthesized as the hydrophobic block

(Figure S1 and S2). To construct the polymeric vesicle using a one-step PISA reaction in an aqueous solution, the RAFT aqueous dispersion polymerization of (2-hydroxypropyl methacrylate, HPMA) to the CTA-AZO-PEG was carried out at room temperature (Figure 1A). The target degree of polymerization (DP) was selected to be 300 for the formation of the polymeric vesicles.24, 35 The transparent solution gradually became turbid (Figure 1B), indicating the occurrence of the polymerization induced self-assembly. The polymerization was verified by 1H-NMR (Figure 1C) and FTIR (Figure S3). New peaks from the polyHPMA with high monomer conversion (>99%) are clearly observed, and the degree of polymerization is calculated to be ~312. Gel permeation chromatography (GPC) analysis (Figure S4) shows that the polydispersity of the final diblock copolymer is relatively low (Mw/Mn=1.54). Formation of polyHPMA-AZOPEG (designed as PHAP) polymeric vesicles was confirmed by TEM (Figures 1D and 1E). The vesicles are relatively uniform and have diameters around ~250 nm as seen upon further characterization using dynamic light scattering (DLS) (Figure 1F) which shows that the vesicles are monodispersed, agreeing well with the TEM results.

Figure 1. Preparation of stimulus-responsive PHAP polymeric vesicles. (A) Synthesis of PHAP block copolymer during polymerization induced self-assembly (PISA) of vesicles. (B) Photos of precursor solution and dispersion of PHAP polymeric vesicles prepared through PISA. (C) The 1H-NMR spectrum of synthesized PHAP block copolymers. The assignments for characteristic peaks are labeled in (A). (D and E) TEM images of PHAP polymeric vesicles. Scale bars: 200 nm. (F) Size distributions of as prepared PHAP polymeric vesicles measured by TEM (red bar chart) and DLS (blue curve), respectively. The photo-isomerization and unique light-responsiveness features of the AZO moieties containing block copolymers in the synthesized and self-assembled PHAP vesicles were studied by monitoring their UV-Vis spectra. The synthesized



vesicles were dissolved in methanol, and the UV-Vis spectra of the polymer solution under visible light and UV exposure were recorded (Figure 2A). The polymer solution shows a characteristic peak at 355 nm and a small peak at 450 nm under visible light. However, with UV exposure, the absorbance peak at 355 nm decreased due to the π to π* transition of AZO, and the absorbance at 450 nm increased slightly due to the n to π* transition.40 This indicates that AZO groups photoisomerize from the trans-to-cis form. The reversible cis-trans conversion only takes a few minutes (Figure 2B), and the responsive behavior did not show significant decay even after several cycles (Figure 2C). Interestingly, we did not see any apparent change in the PHAP polymeric vesicle solution after UV exposure (Figure 2D). As verified by TEM images (Figure 2E and 2F). and DLS (Figure 2G), after the actuation of light, the PHAP vesicles kept intact their vesicular structures and no apparent morphology or size change of the vesicles was observed.

Figure 2. (A) UV-Vis spectra of PHAP block polymer solution (from the PHAP polymeric vesicles) before and after UV light irradiation. (B) Dynamic profiles of absorbance at 355 nm of PHAP block polymer solution under UV light (UV) irradiation and visible light (Vis). (C) Repeated photoswitching of PHAP polymeric vesicles by alternating irradiation with UV and visible light. (D) Photos of PHAP polymeric vesicle dispersion before and after UV irradiation. (E) The TEM image of PHAP polymeric vesicles before UV irradiation. Scale bar: 200 nm. (F) The TEM image of PHAP polymeric vesicles after UV irradiation. Scale bar: 200 nm. (G) Dynamic light scattering (DLS) analysis of PHAP polymeric vesicle dispersion before and after UV irradiation. (H) Rh6G release profiles of PHAP polymeric vesicles under UV irradiation (UV+) and in the dark (UV-). (I) Fluorescence macromolecule release profiles of PHAP polymeric vesicles under UV irradiation (UV+) and in the dark (UV-). It has been recently discovered that the interface with AZO moieties undergoes a sudden perturbation upon light-induced isomerization, which would change the properties of the membrane.41-42 As a result, the membrane permeability can be modified by light irradiation. In virtue of the unique UV light responsiveness of the PHAP polymeric vesicles, we then

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proceeded to study the UV light-induced release of guest molecules from prepared PHAP polymeric vesicles. Fluorescence dye Rh6G and fluorescence FITC-dextran (Mw=70k), where identified as model small molecules and macromolecules respectively, and were entrapped in the PHAP polymeric vesicles. The cargo molecules could be easily encapsulated in the polymeric vesicles without significant change of their morphology or size during polymerization induced self-assembly (Figure S5). A dispersion of PHAP polymeric vesicles was exposed under light irradiation or wrapped by foil paper. The concentration of released dye molecules was monitored periodically by measurement of their fluorescence intensities and calculated based on the calibration curves (Figure S6). It should be noted that light irradiation has limited influence on the fluorescence intensity of the Rh6G dye or fluorescence dextran (Figure S7). As shown in Figure 2H, in comparison to that without light actuation, the encapsulated Rh6G dye molecules were released into the bulk solution rapidly when under UV exposure. This indicates that the PHAP polymer vesicles become more permeable for Rh6G dye with UV light actuation, thereby releasing the dye molecules. Interestingly, the release of Rh6G is dependent on the continuous light inflow and its intensity (Figure S8A). Therefore, the on-demand release of Rh6G could be realized by switching on and off the UV light source (Figure S8B). However, for those PHAP versicles with entrapped macromolecules of FITC-dextran, no matter whether the light is applied or not, the fluorescence intensity of the solution has no significant increase (Figure 2I). These results reveal that the permeability of the PHAP polymer vesicles for macromolecules is limited. These results, therefore, demonstrate that the PHAP polymeric vesicle has light-responsive and size-selective permeability for small molecules. Besides its photo-isomerization, another unique feature of AZO is its chemo-sensitivity: AZO moieties are chemoselectively cleavable under inadequate oxygen supply or low oxygen (hypoxia).43-44 Notably, for the PHAP polymeric vesicles in our design, the AZO modules are installed at the interface of the amphiphiles assembled membrane. We hypothesized that the cleavage of AZO moieties would result in the degradation of the PHAP polymeric vesicles (Figure 3A), and therefore release the cargo molecules confined in the lumen without size limitation. To test our hypothesis, we incubated the PHAP polymeric vesicles with sodium dithionite solution (SD, mimicking the condition of hypoxia), and monitored their morphology evolution by TEM. As shown in Figure 3B, we can clearly see some vesicles started to aggregate after incubation in the SD solution, possibly due to loss of some hydrophilic block (PEG) to maintain the vesicular structure. After 6 hours, many vesicles were broken, and their morphology also changed. In the end, no vesicular structure was observed, and only nanometer-size objects and aggregates were observed (Figure 3C). DLS analysis shows that the resulting samples have a much broader size distribution than the PHAP polymeric vesicles (Figure 3D), indicating the degradation of the vesicles. We further confirmed the cleavage of AZO by UV-Vis spectra (Figure 3E). The characteristic absorbance peak of AZO at 355 nm disappeared. Furthermore, GPC analysis of the polymer from the cleaved PHAP vesicles (Figure S9) shows that a new shoulder peak was presented at the molecular weight, which can be ascribed to the cleavage of the AZO bridge of the block polymers. These results support and demonstrate our hypothesis that the redox-induced cleavage of AZO could lead to the disassembly of polymeric vesicles.


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Chemistry of Materials We then further tested the chemo-responsive release of confined (entrapped) molecules from our PHAP polymeric vesicles through vesicle disassembly under hypoxia conditions. Similarly, PHAP polymeric vesicles with caged small molecules (Rh6G) were incubated with sodium dithionite (SD) solution of different concentrations (Figure 3F). Clearly higher concentrations of the SD solution lead to a faster release of the encapsulated dye molecules. Figure 3G depicts the timedependent release of the Rh6G to the bulk solution in a 25 mM SD (SD+) or without SD (SD-). In the absence of SD, the dye release was limited. However, in the presence of SD, the dye molecules were rapidly released into the bulk solution, and dye release reached a saturation value after 12 hours. These results indicate that the cleavage of the AZO moiety led to the disassembly of the PHAP polymeric vesicles and, as a result, burst dye release. More importantly, for those PHAP vesicles with caged FITC-dextran macromolecules, similar release profiles were observed (Figure 3H). In the presence of SD, the PHAP polymeric vesicles also show burst release of FITCdextran macromolecules. These results could be ascribed to the disassembly of the vesicular structure due to the cleavage of the hydrophobic block and the hydrophilic block of the amphiphiles.

and in absence of SD (SD-). (H) Fluorescence macromolecule release profiles of PHAP polymeric vesicles in presence of SD (SD+) and in absence of SD (SD-). The above results have shown the construction of novel stimulus-responsive nanovesicles through a one-pot and onestep polymerization induced self-assembly strategy. Besides the convenient and relatively mild synthetic approach, the unique photo/chemo responsiveness properties of the PHAP nanovesicles are achieved by the installation of one single molecular module at the interface between hydrophilic and hydrophobic blocks. This concept differentiates our PHAP nanovesicles from other reported stimulus-responsive vesicles. In natural vesicular structures, living cells and their organelles show selective permeability and can be completely disassembled by the degradation of the lipid amphiphiles. Interestingly, these stimulus-responsive PHAP nanovesicles show some similar features. On the one hand, the PHAP nanovesicles can undergo photo stimulus without destroying their vesicular structure and lead to the reversible and selective release of small molecular cargoes. These properties make these polymeric vesicles into excellent candidates for caging and protecting bioactive macromolecules, as for the application to bioreactors. On the other hand, the above PHAP nanovesicles can be disassembled when exposed to chemo stimulus (hypoxia conditions), resulting in the bursting release of all lumen contents. Notably, hypoxia, which is caused by inadequate oxygen supply, is present on a variety of diseases, including cancer, cardiopathy, ischemia, and vascular diseases.45 From the above, we infer that these PHAP vesicles also have the potential for biomedical applications as nanocarriers to load fluorescence dyes and therapeutic drugs.

CONCLUSIONS

Figure 3. (A) Illustration and mechanism of chemoresponsive disassembly of PHAP polymeric vesicles due to the cleavage of AZO moieties. (B) TEM images of PHAP polymeric vesicles incubated with sodium dithionite (SD) at the indicated time points. Scale bars: 500 nm. (C) Photos of PHAP polymeric vesicle dispersion before and after incubation in SD solution. (D) Dynamic light scattering (DLS) analysis of PHAP polymeric vesicle dispersion before and after incubation with SD. (E) UV-Vis spectra of polymer solution (from vesicles) before and after treatment with SD solution. (F) Fluorescence spectra of Rh6G released from PHAP polymeric vesicles in the presence of SD at different concentrations. (G) Rh6G release profiles of PHAP polymeric vesicles in presence of SD (SD+)

In summary, we have demonstrated the one-pot synthesis and self-assembly of new stimulus-responsive polymeric vesicles endowed with photo/chemo-responsive permeability by using a polymerization induced self-assembly approach. We used a novel molecular design that installs photo/chemoresponsive moieties on the membrane interface of the polymeric vesicles and enables controllable, UV-induced, sizeselective permeability or hypoxia-induced burst disassembly of the vesicular structure. Under the irradiation of UV light, the PHAP polymeric nanovesicles can release small molecular cargoes while retaining the macromolecular cargoes; however, under hypoxic conditions, the PHAP polymeric vesicles disassembled and result in sustained release of encapsulated guest molecules. Considering the complicated procedures and enormous difficulties in the construction of stimulusresponsive polymeric vesicles, our molecular design, and facile one-pot approach provide new insights and opportunities to develop next-generation self-assembling and functionalized supramolecular objects. It is anticipated that our photo/chemo-responsive PHAP polymeric vesicles have extensive specialized applications as bioreactors, active biosensors, and drug delivery vehicles in various basic and applied research and industrial fields.

ASSOCIATED CONTENT Supporting Information. Additinoal NMR, FTIR, GPC, TEM, fluoresence spectra and DLS data of the polymers and vesicles. This material is available free of charge via the Internet at http://pubs.acs.org

AUTHOR INFORMATION



Corresponding Author * E-mail: [email protected]; * E-mail: [email protected].

Author Contributions The manuscript was written through the contributions of both authors, which give approval to the final version of the manuscript.

Notes The authors declare no competing financial interest.

ACKNOWLEDGMENTS This work was funded by Repsol, S.A., Spain. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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