Article Cite This: Macromolecules XXXX, XXX, XXX−XXX
pubs.acs.org/Macromolecules
Pushing the Limits of High Throughput PET-RAFT Polymerization Gervase Ng,† Jonathan Yeow,† Robert Chapman,‡ Naatasha Isahak,§ Ernst Wolvetang,§ Justin J. Cooper-White,§,∥ and Cyrille Boyer*,† †
Centre for Advanced Macromolecular Design and Australian Centre for NanoMedicine, School of Chemical Engineering, and Centre for Advanced Macromolecular Design and Australian Centre for NanoMedicine, School of Chemistry, The University of New South Wales, Sydney, NSW 2052, Australia § UQ-StemCARE Australian Institute for Bioengineering and Nanotechnology and ∥School of Chemical Engineering, The University of Queensland, Brisbane, QLD 4072, Australia Downloaded via KAOHSIUNG MEDICAL UNIV on September 20, 2018 at 22:48:54 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.
‡
S Supporting Information *
ABSTRACT: We investigate a high throughput approach to polymer synthesis by employing photoinduced electron/ energy transfer−reversible addition-fragmentation chain transfer (PET-RAFT) polymerization. Polymerization of a broad range of monomers, including acrylates, methacrylates, acrylamides, and styrenic monomers, was achieved directly in a multiwell plate by employing 5,10,15,20-tetraphenylporphine zinc (ZnTPP) as a photocatalyst under yellow LED light. Various parameters such as monomer concentration and degree of polymerization were investigated with respect to their effect on polymerization rate and the degree of control over the molecular weight and molecular weight distribution. Finally, the synthesis of well-defined multiblock copolymers (up to a hexablock copolymer) was shown to be achievable entirely within a multiwell plate without any intermediate purification. The versatility and ease of this oxygen tolerant polymerization in high throughput formats make it an excellent technique for the generation of polymer arrays.
■
reaction mixture.15−18 This is possible via the in situ removal of oxygen using chemical mediators such as enzymes19−22 or redox active metals to chemically convert oxygen into inactive species.23−32 Our group33,34 and others35−41 have exploited this oxygen tolerance to perform CLRP in a HT manner, allowing polymer libraries to be prepared on a benchtop and, importantly, while retaining excellent control over polymer molecular weight and architecture. Recently, our group in collaboration with Chapman’s group reported that the oxygen tolerance of the 5,10,15,20tetraphenyl-21H,23H-porphine zinc (ZnTPP) mediated photoinduced electron/energy transfer−reversible addition-fragmentation chain transfer (PET-RAFT) polymerization process42−48 can be exploited to synthesize a library of linear, 3arm, and 4-arm star polymers in HT using 96- or 384-well plates.49 By use of this HT PET-RAFT process, a mannose functional polymer library was rapidly synthesized and subsequently screened with a lectin binding assay generating a structure−activity relationship. We also reported the use of an online fluorescence-based ratiometric technique for monitoring polymerization, enabling monomer conversion to be determined without discrete sampling and thereby facilitating HT polymer synthesis at low volumes and
INTRODUCTION In recent years, high throughput (HT) approaches in the chemical sciences have become increasingly prevalent in both academia and industry.1−5 For example, in the pharmaceutical sciences, HT chemistry is commonly employed for performing rapid target identification and optimization of synthetic routes.6,7 More recently, these approaches have also been applied to the field of polymer chemistry for the screening of polymer structures. For example, Langer and co-workers have used UV-initiated free radical polymerization (FRP) to synthesize large polymer libraries in a microarray format directly on glass slides.8−10 Although these FRP strategies to HT synthesis allow the systematic study of polymer structure− property relationships, they are somewhat limited by their inability to control polymer composition and structure that ultimately influence the material properties. In comparison to conventional FRP, controlled/living radical polymerization (CLRP) techniques allow for exquisite control over the polymer molecular weight, dispersity, and architecture. Historically, one of the main limitations of CLRP in terms of increased throughput has been its strong susceptibility toward molecular oxygen.11 As a result, polymer libraries synthesized using CLRP typically require specialized equipment such as glove boxes to obtain a suitable level of control over the polymerization.12−14 As an alternative, several techniques have been proposed which allow for CLRP to be performed without prior physical deoxygenation of the © XXXX American Chemical Society
Received: July 26, 2018 Revised: September 10, 2018
A
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules
Scheme 1. (left) Proposed Mechanism for Oxygen Tolerant PET-RAFT Polymerization and (right) Chemical Structures of Catalyst, Monomers and RAFT Agents Employed in This Study
Table 1. PET-RAFT Polymerization of a Range of Monomer Families Conducted Directly in 96-Well Platesa monomer family acrylamide
acrylate
styrene methacrylate
no.
monomer
RAFT agent
αb (%)
Mn,thc (g/mol)
Mn,GPCd (g/mol)
Mw/Mnd
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
DMA DAAm DEA NIPAM NAM HEAm MA BzA DEGA HEA Sty VBA MMA BzMA GMA HPMA MTEMA
BTPA BTPA BTPA BTPA BTPA BTPA BTPA BTPA BTPA BTPA BTPA BTPA CDTPA CDTPA CDTPA CDTPA CDTPA
98 92 97 98 97 95 96 77 92 91 20 25 71 74 75 68 60
19700 31400 24900 22400 27600 22100 16800 25200 34800 21400 4400 6800 14400 26300 21400 18100 19600
18900 22800 9000 24900 18700 33300 10100 18800 20500 24200 2600 7100 9500 17800 18300 22000 15100
1.16 1.09 1.08 1.12 1.08 1.11 1.07 1.14 1.25 1.11 1.18 1.21 1.29 1.34 1.38 1.26 1.37
a
Polymerizations were conducted for 4 h under yellow LED light (λmax = 560 nm, 9.7 mW/cm2) using a [M]:[RAFT]:[ZnTPP] = 200:1:0.02 and with [M] = 1 M. bMonomer conversion (α) was determined by 1H NMR spectroscopy using mesitylene as an internal standard. cTheoretical molecular weights (Mn,th) were calculated using the equation Mn,th = [M]/[RAFT] × α × MWmonomer + MWRAFT. dExperimental molecular weights (Mn,GPC) and polymer dispersities (Mw/Mn) were determined by GPC using PMMA standards for calibration. Monomers: N,N-dimethylacrylamide (DMA), diacetone acrylamide (DAAm), N,N-dimethylacrylamide (DEA), N-isopropylacrylamide (NIPAM), N-acryloylmorpholine (NAM), 2hydroxyethyl acrylamide (HEAm), methyl acrylate (MA), benzyl acrylate (BzA), di(ethylene glycol)ethyl ether acrylate (DEGA), 2-hydroxyethyl acrylate (HEA), styrene (Sty), 3-vinylbenzaldehyde (VBA), methyl methacrylate (MMA), benzyl methacrylate (BzMA), glycidyl methacrylate (GMA), 2-hydroxypropyl methacrylate (HPMA), and 2-(methylthio)ethyl methacrylate (MTEMA).
concentrations.33 However, in these studies, polymerizations were only conducted under a limited range of conditions with only detailed studies of acrylamide based monomers having been demonstrated. In this work, we expand the scope of HT PET-RAFT polymerization by exploring a broad range of synthetic conditions such as monomer families/functionalities, monomer concentrations, and target molecular weight as well as exploring the synthesis of multiblock copolymers (Scheme 1).
polymerizations were conducted in the presence of 2-(nbutyltrithiocarbonate)propionic acid (BTPA) as a RAFT agent and DMSO as solvent using a [M]:[RAFT]:[ZnTPP] = 200:1:0.02 at room temperature (∼20 °C). DMSO was selected as solvent to impart oxygen tolerance due to its ability to quench singlet oxygen generated by photosensitization.50 Notably, polymerization reactions were performed in 96-well plates without stirring and from visual observation appeared homogeneous after the irradiation period. Furthermore, the reaction mixture was homogenized by aspiration with a pipet prior to aliquot sampling for characterization by 1H NMR and GPC. Under yellow LED light irradiation (λmax = 560 nm, 9.7 mW/cm2), various acrylamide monomers were polymerized to high monomer conversion (>92%) within 4 h with good control over the molecular weight and molecular weight
■
RESULTS AND DISCUSSION PET-RAFT Polymerization of a Range of Monomer Families in 96-Well Plates. Initially, we investigated the oxygen tolerance of ZnTPP mediated PET-RAFT polymerization by polymerizing acrylamide monomers directly in 96well plates at a reaction volume of 200 μL. These B
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules
Figure 1. Molecular weight distributions of (A) acrylamide, (B) acrylate, (C) styrene, and (D) methacrylate families prepared using HT PETRAFT polymerization in 96-well plates. Polymerizations were conducted for 4 h under yellow LED light (560 nm, 9.7 mW/cm2) using a [M]: [RAFT]:[ZnTPP] = 200:1:0.02 and with [M]0 = 1 M.
Table 2. Polymerization Kinetics of ZnTPP Mediated PET-RAFT Polymerization of NAM and BzMA Conducted in 96-Well Plates in the Presence of 9,10-Dimethylanthracene (DMAn) and in DMF or Dioxane as Solventa no.
monomer
RAFT agent
DMAn (mg)
solvent
αb (%)
Mn,thc (g/mol)
Mn,GPCd (g/mol)
Mw/Mnd
1 2 3 4 5 6
NAM NAM NAM NAM BzMA BzMA
BTPA BTPA BTPA BTPA CDTPA CDTPA
1 1
DMF dioxane DMF dioxane DMF dioxane
66 35 7 4 53 42
18900 10100 2200 1400 18900 15000
16600 7500 N.R. N.R. 12400 11300
1.26 1.34 N.R. N.R. 1.49 1.40
1 1
a
Polymerizations were conducted for 4 h under yellow LED light (λmax = 565 nm, 9.5 mW/cm2) using [M]:[RAFT]:[ZnTPP] = 200:1:0.02 and an initial monomer concentration of 1.0 M. bMonomer conversion (α) was determined by 1H NMR spectroscopy using DMF as an internal standard. c Theoretical molecular weights (Mn,th) were calculated using the equation Mn,th = [M]/[RAFT] × α × MWmonomer + MWRAFT. dExperimental molecular weights (Mn,GPC) and polymer dispersities (Mw/Mn) were determined by GPC using PMMA standards for calibration. N.R. = not run.
results reflect the relatively low propagation rate of styrenic monomers at room temperature compared to other monomer families. Similar results for the photoRAFT polymerization of styrene drivatives have previously been observed by Cai and co-workers.51 In contrast, polymerizations of methacrylates performed using 4-cyano-4-[(dodecylsulfanylthiocarbonyl)sulfanyl]pentanoic acid (CDTPA) as a RAFT agent were much faster, achieving conversions of 60−75% within 4 h despite the presence of oxygen (Table 1, no. 13−17). Molecular weight distributions were generally broader (Mw/ Mn ∼ 1.3) than obtained for acrylates and acrylamides (Figure 1D), which could be attributed to either a slow setup of the RAFT equilibrium due to the relatively stable methacrylyl radical52 or premature termination of growing chain radicals during propagation. Nonetheless, polymethacrylates with a range of functionalities such as epoxy (glycidyl methacrylate, GMA), hydroxy (hydroxypropyl methacrylate, HPMA), and thioether (2-(methylthio)ethyl methacrylate, MTEMA) moieties were successfully synthesized using this approach with a reasonable degree of polymerization control. Furthermore, for a range of monomers, similar degrees of polymerization control in multiwell plates were also achievable when the target degree of polymerization (DP) was reduced to 50 (Supporting Information, Table S1 and Figure S1).
distribution despite the lack of prior deoxygenation (Mw/Mn < 1.16, Table 1, no. 1−6, Figure 1A). More importantly, excellent control in polymerization was observed even when employing functional acrylamides such as ketone (DAAm) and hydroxy functional (HEAm) monomers as well as monomers which form thermoresponsive polymers (DEA and NIPAM). Encouraged by these initial results, HT PET-RAFT polymerization was next extended to a range of acrylate (Table 1, no. 7−10), styrenic (Table 1, no. 11 and 12), and methacrylate (Table 1, no. 13−17) monomers. In all cases, acrylate monomers such as the thermoresponsive di(ethylene glycol) ethyl ether acrylate (DEGA) and hydroxy functional 2hydroxyethyl acrylate (HEA) could be polymerized directly in a 96-well plate with good control over the molecular weight and molecular weight distributions (Mw/Mn < 1.14, Table 1, no. 7−10, and Figure 1B). In the case of some acrylamides and acrylates, a small high molecular weight shoulder was observed which we have attributed to bimolecular termination events occurring at high monomer conversion. For the polymerization of styrene (Sty) and 3-vinylbenzaldehyde (VBA), although reasonable dispersities were achieved (Mw/Mn < 1.21), the polymerization was relatively slow with less than 25% monomer conversion observed after 4 h of irradiation (Table 1, no. 11 and 12, and Figure 1C). These C
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules
Figure 2. (A) Polymerization kinetics of ZnTPP mediated PET-RAFT polymerization of NAM conducted in 96-well plates under yellow LED light (560 nm, 9.7 mW/cm2) using a [M]:[RAFT]:[ZnTPP] = 200:1:0.02 at an initial monomer concentration of 1.0, 0.5, or 0.1 M. Evolution of the GPC derived molecular weight and dispersity with monomer conversion at an initial monomer concentration of (B) 1.0 M, (C) 0.5 M, or (D) 0.1 M.
Table 3. Experimental and Characterization Data for the PET-RAFT Polymerization of NAM Conducted in 96-Well Plates at a Range of Initial Monomer Concentrationsa
Table 4. Experimental and Characterization Data for the PET-RAFT Polymerization of NAM Conducted in 96-Well Plates at a Range of Target Molecular Weightsa
no.
[M]:[BTPA]: [ZnTPP]
[M] (M)
αb (%)
Mn,thc (g/mol)
Mn,GPCd (g/mol)
Mw/Mnd
no.
[M]:[BTPA]: [ZnTPP]
αb (%)
Mn,thc (g/mol)
Mn,GPCd (g/mol)
Mw/Mnd
1 2 3 4 5 6 7 8
200:1:0.02 200:1:0.02 200:1:0.02 200:1:0.02 200:1:0.02 200:1:0.02 200:1:0.02 200:1:0.02
1.00 0.75 0.50 0.25 0.10 0.05 0.025 0.0125
99 99 99 99 94 91 80 43
28200 28200 28200 28200 26800 25900 22800 12400
22500 21800 22300 21800 22700 22900 20200 10900
1.09 1.10 1.10 1.10 1.15 1.19 1.28 1.33
1 2 3 4 5 6 7 8
25:1:0.02 50:1:0.02 100:1:0.02 200:1:0.02 500:1:0.02 1000:1:0.02 5000:1:0.02 10000:1:0.02
99 99 99 97 96 92 56 16
3700 7200 14200 27600 68000 130100 395500 226100
3300 6000 10500 18700 39100 60800 172900 187600
1.11 1.09 1.10 1.08 1.15 1.27 2.14 2.25
a
a
Polymerizations were conducted for 4 h under yellow LED light (560 nm, 9.7 mW/cm2) using [M]:[RAFT]:[ZnTPP] = 200:1:0.02. b Monomer conversion (α) was determined by 1H NMR spectroscopy using DMF as an internal standard. cTheoretical molecular weights (Mn,th) were calculated using the equation Mn,th = [M]/[RAFT] × α × MWmonomer + MWRAFT. dExperimental molecular weights and dispersities (Mw/Mn) determined by GPC using PMMA standards for calibration.
Polymerizations were conducted for 4 h under yellow LED Light (560 nm, 9.7 mW/cm2) using a constant [RAFT]:[ZnTPP] = 1:0.02. b Monomer conversion (α) was determined by 1H NMR spectroscopy using DMF as an internal standard. cTheoretical molecular weights (Mn,th) were calculated using the equation Mn,th = [M]/[RAFT] × α × MWmonomer + MWRAFT. dExperimental molecular weights and dispersities (Mw/Mn) determined by GPC using PMMA standards for calibration.
Figure 3. Molecular weight distributions for the PET-RAFT polymerization of NAM conducted in 96-well plates at initial monomer concentrations of (A) 1.00−0.25 M and (B) 0.100−0.0125 M. Polymerizations were conducted for 4 h under yellow LED light (560 nm, 9.7 mW/cm2) using a [M]:[RAFT]:[ZnTPP] = 200:1:0.02. D
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules
Figure 4. (A) Molecular weight distributions for the PET-RAFT polymerization of NAM in a 96-well plate at a range of target DPs. Polymerizations were conducted under yellow LED light (560 nm, 9.7 mW/cm2) using a [RAFT]:[ZnTPP] = 1:0.02 and with [M]0 = 1 M. (B) Molecular weight distributions for PNAM synthesized at a range of target DPs but employing a fixed ZnTPP concentration of 0.1 mM.
sions (Table 2) in these solvents were relatively low compared to DMSO as has been previously observed.54 In addition, broader molecular weight distributions (Figure S2) were obtained along with some low molecular weight tailing, which can be ascribed to the differing solubility of oxygen in different solvents as well as the varying efficiencies of the singlet oxygen quenching process. Kinetics of PET-RAFT Polymerization in 96-Well Plates. Because of its relatively high propagation rate and excellent polymerization control, N-acryloylmorpholine (NAM) was selected as a model monomer for more detailed studies of the kinetics of HT PET-RAFT polymerizations conducted in 96-well plates. For these studies, polymerizations were performed in separate wells in the presence of BTPA as a RAFT agent and at an initial monomer concentration of [NAM]0 = 1.0, 0.5, or 0.1 M (Table S2). At all concentrations, the polymerization kinetics revealed little to no induction period as well as a linear evolution of ln([[M]0/[M]t) versus reaction time, suggesting a fairly constant concentration of propagating radicals throughout the polymerization (Figure 2A). Furthermore, the experimental molecular weights increase in reasonable agreement with the theoretical values, and the polymer dispersity generally decreases with increasing monomer conversion (Figure 2B−D). Finally, the molecular weight distributions obtained by GPC were narrow and unimodal throughout the polymerization indicating the livingness of the well plate polymerization despite the lack of prior deoxygenation (Figure S3). To determine the effect of oxygen on HT PET-RAFT polymerization, polymerization kinetics were performed in a nitrogen-filled glovebox (Table S3) and compared with results obtained on a benchtop under ambient conditions (Table S4). Because of instrument constraints, a different LED irradiation source (565 nm, 9.5
Table 5. Experimental Data and Polymer Characterization for the Synthesis of a PNAM Hexablock Copolymera block no.
time (min)
αb (%)
Mn,thc (g/mol)
Mn,GPCd (g/mol)
Mw/Mnd
1 2 3 4 5 6
120 120 180 180 240 240
96 98 98 97 96 88
3600 7400 11000 14500 18100 21500
3400 6700 10200 14100 18000 20900
1.10 1.11 1.11 1.11 1.15 1.19
a
Each chain extension was conducted under yellow LED light (560 nm, 9.7 mW/cm2) using a [M]:[RAFT] = 25:1. ZnTPP was added only for the polymerization of the first block at a [RAFT]:[ZnTPP] = 1:0.01. bMonomer conversion (α) was determined by 1H NMR spectroscopy using DMF as an internal standard. cTheoretical molecular weights (Mn,th) for a given block number (N) were calculated using the following equation: Mn,th(N) = [M]:[(macro)RAFT] × α × MWmonomer + MW(macro)RAFT(N−1) where the initial ratio [M]:[(macro)RAFT)] has been adjusted for the unreacted monomer from the polymerization of the previous block (calculations are given in Table S7). dExperimental molecular weights and dispersities (Mw/ Mn) as determined by GPC using PMMA standards for calibration.
To demonstrate the versatility of HT PET-RAFT polymerization, we explored polymerization in other solvents but with the addition of an exogenous singlet oxygen quencher, 9,10dimethylanthracene (DMAn).53 In the absence of DMAn, low monomer conversions (1000), polymerization rates become very slow, and the molecular weight distributions suggest uncontrolled free radical polymerization was occurring under these conditions. It should be noted that in these experiments the concentration of ZnTPP was varied with the varying concentration of RAFT agent to maintain a constant ratio of RAFT agent to photocatalyst ([RAFT]:[ZnTPP] = 1:0.02). To test whether higher concentrations of ZnTPP could be employed to improve the efficiency of deoxygenation and degree of polymerization control, we maintained the concentration of ZnTPP at 0.1 mM throughout while targeting different DPs. Notably, for comparable DPs, no significant improvements in monomer conversion or polymer dispersity were observed using this approach (Figure 4B and Table S5), suggesting that the presence of oxygen is not the main reason behind the loss of control at high target DPs. In addition, the relatively high concentration of ZnTPP at these high target DPs resulted in an increased generation of propagating radicals leading to significant bimolecular chain−chain coupling. This phenomenon can be associated with the mechanism of RAFT polymerization itself which has been known to limit the synthesis of high molecular weight polymers.56 Multiblock Synthesis Using PET-RAFT Polymerization in 96-Well Plates. Finally, we decided to investigate the livingness of this PET-RAFT system by synthesizing highorder multiblock copolymers of NAM entirely within a 96-well plate. We targeted a DP of 25 for each NAM block, and each subsequent chain extension was performed without prior purification. Furthermore, due to the decreasing monomer concentration with each block, the irradiation time was increased by 1 h every second block to ensure high monomer conversions (>95%) could still be obtained. After each chain extension, a fixed volume (80 μL) of the crude mixture was removed for NMR and GPC analysis followed by addition of fresh NAM as a stock solution in DMSO. After removal of a fixed volume (40 μL) for NMR analysis of the initial monomer concentration, the prepolymerization reaction volume was 200 μL. This process enabled successive chain extensions to be performed at a constant polymerization volume of 200 μL and was necessary to limit the reaction volume to the maximum well volume of 300 μL. A detailed description of the experimental setup is provided in the Supporting Information (Table S6). Apart from the sixth block, high monomer conversions of at least 96% were obtained for each chain extension (Table 5), and the molecular weight distributions shifted smoothly to higher molecular weight with little evidence of low molecular weight tailing (Figure 5A). Furthermore, narrow dispersities were obtained throughout the synthesis, yielding a dispersity of 1.19 for the final [PNAM25]6 hexablock. In addition, the experimental molecular weights derived from GPC were in good agreement with the theoretical values (Figure 5B), suggesting that the high chainend fidelity and livingness of the HT PET-RAFT system could be harnessed for successful multiblock synthesis in a multiwell plate. To determine whether additional ZnTPP was beneficial to the synthesis, we also performed an experiment whereby fresh ZnTPP was added to the chain extension mixture every second block. However, using this protocol, we observed no significant differences in the polymerization behavior in terms of monomer conversion and molecular weight distribution for each block (Figure S8 and Table S8). Together, these data indicate the high photocatalytic efficiency with which ZnTPP can photosensitize oxygen and mediate RAFT polymerization F
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules
libraries: review of state of the art. ACS Comb. Sci. 2011, 13 (6), 579− 633. (6) Macarron, R.; Banks, M. N.; Bojanic, D.; Burns, D. J.; Cirovic, D. A.; Garyantes, T.; Green, D. V.; Hertzberg, R. P.; Janzen, W. P.; Paslay, J. W.; Schopfer, U.; Sittampalam, G. S. Impact of highthroughput screening in biomedical research. Nat. Rev. Drug Discovery 2011, 10 (3), 188−95. (7) Bleicher, K. H.; Bohm, H. J.; Muller, K.; Alanine, A. I. Hit and lead generation: beyond high-throughput screening. Nat. Rev. Drug Discovery 2003, 2 (5), 369−78. (8) Anderson, D. G.; Levenberg, S.; Langer, R. Nanoliter-scale synthesis of arrayed biomaterials and application to human embryonic stem cells. Nat. Biotechnol. 2004, 22 (7), 863−6. (9) Hook, A. L.; Chang, C. Y.; Yang, J.; Luckett, J.; Cockayne, A.; Atkinson, S.; Mei, Y.; Bayston, R.; Irvine, D. J.; Langer, R.; Anderson, D. G.; Williams, P.; Davies, M. C.; Alexander, M. R. Combinatorial discovery of polymers resistant to bacterial attachment. Nat. Biotechnol. 2012, 30 (9), 868−875. (10) Tweedie, C. A.; Anderson, D. G.; Langer, R.; Van Vliet, K. J. Combinatorial Material Mechanics: High-Throughput Polymer Synthesis and Nanomechanical Screening. Adv. Mater. 2005, 17 (21), 2599−2604. (11) Guerrero-Sanchez, C.; Keddie, D. J.; Saubern, S.; Chiefari, J. Automated parallel freeze-evacuate-thaw degassing method for oxygen-sensitive reactions: RAFT polymerization. ACS Comb. Sci. 2012, 14 (7), 389−94. (12) Cockram, A. A.; Bradley, R. D.; Lynch, S. A.; Fleming, P. C. D.; Williams, N. S. J.; Murray, M. W.; Emmett, S. N.; Armes, S. P. Optimization of the high-throughput synthesis of multiblock copolymer nanoparticles in aqueous media via polymerizationinduced self-assembly. React. Chem. Eng. 2018, DOI: 10.1039/ C8RE00066B. (13) Fijten, M. W. M.; Meier, M. A. R.; Hoogenboom, R.; Schubert, U. S. Automated parallel investigations/optimizations of the reversible addition-fragmentation chain transfer polymerization of methyl methacrylate. J. Polym. Sci., Part A: Polym. Chem. 2004, 42 (22), 5775−5783. (14) Zhang, H. Q.; Marin, V.; Fijten, M. W. M.; Schubert, U. S. High-throughput experimentation in atom transfer radical polymerization: A general approach toward a directed design and understanding of optimal catalytic systems. J. Polym. Sci., Part A: Polym. Chem. 2004, 42 (8), 1876−1885. (15) Yeow, J.; Chapman, R.; Gormley, A. J.; Boyer, C. Up in the air: oxygen tolerance in controlled/living radical polymerisation. Chem. Soc. Rev. 2018, 47 (12), 4357−4387. (16) Leibfarth, F. A.; Mattson, K. M.; Fors, B. P.; Collins, H. A.; Hawker, C. J. External regulation of controlled polymerizations. Angew. Chem., Int. Ed. 2013, 52 (1), 199−210. (17) Borská, K.; Moravčíková, D.; Mosnácě k, J. Photochemically Induced ATRP of (Meth)Acrylates in the Presence of Air: The Effect of Light Intensity, Ligand, and Oxygen Concentration. Macromol. Rapid Commun. 2017, 38 (13), 1600639. (18) Zhang, W.; Xue, W.; Ming, W.; Weng, Y.; Chen, G.; Haddleton, D. M. Regenerable-Catalyst-Aided, Opened to Air and SunlightDriven “CuAAC&ATRP” Concurrent Reaction for SequenceControlled Copolymer. Macromol. Rapid Commun. 2017, 38 (22), 1700511. (19) Liu, Z.; Lv, Y.; An, Z. Enzymatic Cascade Catalysis for the Synthesis of Multiblock and Ultrahigh-Molecular-Weight Polymers with Oxygen Tolerance. Angew. Chem., Int. Ed. 2017, 56 (44), 13852−13856. (20) Tan, J. B.; Liu, D. D.; Bai, Y. H.; Huang, C. D.; Li, X. L.; He, J.; Xu, Q.; Zhang, L. Enzyme-Assisted Photoinitiated PolymerizationInduced Self-Assembly: An Oxygen-Tolerant Method for Preparing Block Copolymer Nano-Objects in Open Vessels and Multiwell Plates. Macromolecules 2017, 50 (15), 5798−5806. (21) Liu, Z. F.; Lv, Y.; Zhu, A. Q.; An, Z. One-Enzyme Triple Catalysis: Employing the Promiscuity of Horseradish Peroxidase for
since it only needs to be present in the initial polymerization mixture.
■
CONCLUSION In this study, we have explored the scope and limits of ZnTPP mediated PET-RAFT polymerization when performed directly in low volume, 96-well plates. In particular, we show this HT process allows for excellent polymerization control with several monomer families (acrylamides, acrylates, methacrylates, and styrenes) as well as a range of monomer functionalities. Furthermore, we demonstrate that control over the polymerization can be achieved at monomer concentrations as low as 50 mM which may be useful for the synthesis of bioconjugates in which limited quantities of reagents are available. Significantly, despite some limitations in terms of absolute molecular weights that can be observed, the livingness of this process was sufficient to allow for the successful synthesis of a hexablock copolymer entirely within a 96-well plate and, notably, without the necessity for repeated deoxygenation steps and purification.
■
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.macromol.8b01600. Experimental details, NMR spectra, and all raw computational data (Figures S1−S9 and Tables S1− S8) (PDF)
■
AUTHOR INFORMATION
Corresponding Author
*E-mail
[email protected] (C.B.). ORCID
Justin J. Cooper-White: 0000-0002-1920-8229 Cyrille Boyer: 0000-0002-4564-4702 Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS C.B. acknowledges Australian Research Council (ARC) for the Future Fellowship (FT120100096) and Discovery Project (DP18010254). R.C. is grateful to the Australian Research Council (ARC) for funding through the Discovery Early Career Research Award (DE170100315).
■
REFERENCES
(1) Zhang, H.; Hoogenboom, R.; Meier, M. A. R.; Schubert, U. S. Combinatorial and high-throughput approaches in polymer science. Meas. Sci. Technol. 2005, 16 (1), 203. (2) Xiang, X. D.; Sun, X.; Briceno, G.; Lou, Y.; Wang, K. A.; Chang, H.; Wallace-Freedman, W. G.; Chen, S. W.; Schultz, P. G. A combinatorial approach to materials discovery. Science 1995, 268 (5218), 1738−40. (3) Danielson, E.; Golden, J. H.; McFarland, E. W.; Reaves, C. M.; Weinberg, W. H.; Wu, X. D. A combinatorial approach to the discovery and optimization of luminescent materials. Nature 1997, 389 (6654), 944−948. (4) Senkan, S. M. High-throughput screening of solid-state catalyst libraries. Nature 1998, 394 (6691), 350−353. (5) Potyrailo, R.; Rajan, K.; Stoewe, K.; Takeuchi, I.; Chisholm, B.; Lam, H. Combinatorial and high-throughput screening of materials G
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
Article
Macromolecules Synthesis and Functionalization of Well-Defined Polymers. ACS Macro Lett. 2018, 7 (1), 1−6. (22) Enciso, A. E.; Fu, L.; Russell, A. J.; Matyjaszewski, K. A Breathing Atom-Transfer Radical Polymerization: Fully OxygenTolerant Polymerization Inspired by Aerobic Respiration of Cells. Angew. Chem., Int. Ed. 2018, 57 (4), 933−936. (23) Matyjaszewski, K.; Coca, S.; Gaynor, S. G.; Wei, M. L.; Woodworth, B. E. Controlled radical polymerization in the presence of oxygen. Macromolecules 1998, 31 (17), 5967−5969. (24) Gody, G.; Barbey, R.; Danial, M.; Perrier, S. Ultrafast RAFT polymerization: multiblock copolymers within minutes. Polym. Chem. 2015, 6 (9), 1502−1511. (25) Chapman, R.; Gormley, A. J.; Herpoldt, K. L.; Stevens, M. M. Highly Controlled Open Vessel RAFT Polymerizations by Enzyme Degassing. Macromolecules 2014, 47 (24), 8541−8547. (26) Xu, J.; Jung, K.; Atme, A.; Shanmugam, S.; Boyer, C. A robust and versatile photoinduced living polymerization of conjugated and unconjugated monomers and its oxygen tolerance. J. Am. Chem. Soc. 2014, 136 (14), 5508−19. (27) Schneiderman, D. K.; Ting, J. M.; Purchel, A. A.; Miranda, R.; Tirrell, M. V.; Reineke, T. M.; Rowan, S. J. Open-to-Air RAFT Polymerization in Complex Solvents: From Whisky to Fermentation Broth. ACS Macro Lett. 2018, 7 (4), 406−411. (28) Wilson, O. R.; Magenau, A. J. D. Oxygen Tolerant and Room Temperature RAFT through Alkylborane Initiation. ACS Macro Lett. 2018, 7 (3), 370−375. (29) Oytun, F.; Kahveci, M. U.; Yagci, Y. Sugar overcomes oxygen inhibition in photoinitiated free radical polymerization. J. Polym. Sci., Part A: Polym. Chem. 2013, 51 (8), 1685−1689. (30) Liarou, E.; Whitfield, R.; Anastasaki, A.; Engelis, N. G.; Jones, G. R.; Velonia, K.; Haddleton, D. M. Copper-Mediated Polymerization without External Deoxygenation or Oxygen Scavengers. Angew. Chem. 2018, 130 (29), 9136−9140. (31) Yang, Q. Z.; Lalevee, J.; Poly, J. Development of a Robust Photocatalyzed ATRP Mechanism Exhibiting Good Tolerance to Oxygen and Inhibitors. Macromolecules 2016, 49 (20), 7653−7666. (32) Wu, C.; Shanmugam, S.; Xu, J.; Zhu, J.; Boyer, C. Chlorophyll a crude extract: efficient photo-degradable photocatalyst for PET-RAFT polymerization. Chem. Commun. 2017, 53 (93), 12560−12563. (33) Yeow, J.; Joshi, S.; Chapman, R.; Boyer, C. A Self-Reporting Photocatalyst for Online Fluorescence Monitoring of High Throughput RAFT Polymerization. Angew. Chem., Int. Ed. 2018, 57 (32), 10102−10106. (34) Yeow, J.; Chapman, R.; Xu, J. T.; Boyer, C. Oxygen tolerant photopolymerization for ultralow volumes. Polym. Chem. 2017, 8 (34), 5012−5022. (35) Cosson, S.; Danial, M.; Saint-Amans, J. R.; Cooper-White, J. J. Accelerated Combinatorial High Throughput Star Polymer Synthesis via a Rapid One-Pot Sequential Aqueous RAFT (rosa-RAFT) Polymerization Scheme. Macromol. Rapid Commun. 2017, 38 (8), 1600780. (36) Chapman, R.; Gormley, A. J.; Stenzel, M. H.; Stevens, M. M. Combinatorial Low-Volume Synthesis of Well-Defined Polymers by Enzyme Degassing. Angew. Chem., Int. Ed. 2016, 55 (14), 4500−3. (37) Tan, J.; Xu, Q.; Li, X.; He, J.; Yuxuan, Z.; Xiaocong, D.; Liangliang, Y.; Ruiming, Z.; Li, Z.; et al. Enzyme-PISA: An Efficient Method for Preparing Well-Defined Polymer Nano-Objects under Mild Conditions. Macromol. Rapid Commun. 2018, 39 (9), 1700871. (38) Siegwart, D. J.; Leiendecker, M.; Langer, R.; Anderson, D. G. Automated ARGET ATRP Accelerates Catalyst Optimization for the Synthesis of Thiol-Functionalized Polymers. Macromolecules 2012, 45 (3), 1254−1261. (39) Wu, H. B.; Yang, L.; Tao, L. Polymer synthesis by mimicking nature’s strategy: the combination of ultra-fast RAFT and the Biginelli reaction. Polym. Chem. 2017, 8 (37), 5679−5687. (40) Lv, C.; He, C.; Pan, X. Oxygen-Initiated and Regulated Controlled Radical Polymerization under Ambient Conditions. Angew. Chem., Int. Ed. 2018, 57 (30), 9430−9433.
(41) Richards, S. -J.; Jones, A.; Tomás, R. M. F.; Gibson, M. I. Photochemical “In-Air” Combinatorial Discovery of Antimicrobial Co-polymers. Chem. - Eur. J. 2018, DOI: 10.1002/chem.201802594. (42) Allegrezza, M. L.; DeMartini, Z. M.; Kloster, A. J.; Digby, Z. A.; Konkolewicz, D. Visible and sunlight driven RAFT photopolymerization accelerated by amines: kinetics and mechanism. Polym. Chem. 2016, 7 (43), 6626−6636. (43) Judzewitsch, P. R.; Nguyen, T. K.; Shanmugam, S.; Wong, E. H. H.; Boyer, C. Towards Sequence-Controlled Antimicrobial Polymers: Effect of Polymer Block Order on Antimicrobial Activity. Angew. Chem., Int. Ed. 2018, 57 (17), 4559−4564. (44) Mao, T.; Liu, G.; Wu, H.; Wei, Y.; Gou, Y.; Wang, J.; Tao, L. High Throughput Preparation of UV-Protective Polymers from Essential Oil Extracts via the Biginelli Reaction. J. Am. Chem. Soc. 2018, 140 (22), 6865−6872. (45) Fu, Q.; Ruan, Q. S.; McKenzie, T. G.; Reyhani, A.; Tang, J. W.; Qiao, G. G. Development of a Robust PET-RAFT Polymerization Using Graphitic Carbon Nitride (g-C3N4). Macromolecules 2017, 50 (19), 7509−7516. (46) Cabannes-Boue, B.; Yang, Q. Z.; Lalevee, J.; Morlet-Savary, F.; Poly, J. Investigation into the mechanism of photo-mediated RAFT polymerization involving the reversible photolysis of the chaintransfer agent. Polym. Chem. 2017, 8 (11), 1760−1770. (47) Watanabe, A.; Niu, J.; Lunn, D. J.; Lawrence, J.; Knight, A. S.; Zhang, M. W.; Hawker, C. J. PET-RAFT as a facile strategy for preparing functional lipid-polymer conjugates. J. Polym. Sci., Part A: Polym. Chem. 2018, 56 (12), 1259−1268. (48) Chu, Y.; Huang, Z.; Liang, K.; Guo, J.; Boyer, C.; Xu, J. A photocatalyst immobilized on fibrous and porous monolithic cellulose for heterogeneous catalysis of controlled radical polymerization. Polym. Chem. 2018, 9 (13), 1666−1673. (49) Gormley, A. J.; Yeow, J.; Ng, G.; Conway, Ó .; Boyer, C.; Chapman, R. An Oxygen-Tolerant PET-RAFT Polymerization for Screening Structure−Activity Relationships. Angew. Chem., Int. Ed. 2018, 57 (6), 1557−1562. (50) Corrigan, N.; Rosli, D.; Jones, J. W. J.; Xu, J. T.; Boyer, C. Oxygen Tolerance in Living Radical Polymerization: Investigation of Mechanism and Implementation in Continuous Flow Polymerization. Macromolecules 2016, 49 (18), 6779−6789. (51) Zhang, H.; Deng, J.; Lu, L.; Cai, Y. Ambient-Temperature RAFT Polymerization of Styrene and Its Functional Derivatives under Mild Long-Wave UV−vis Radiation. Macromolecules 2007, 40 (26), 9252−9261. (52) Xu, S. H.; Ng, G.; Xu, J. T.; Kuchel, R. P.; Yeow, J.; Boyer, C. 2(Methylthio)ethyl Methacrylate: A Versatile Monomer for Stimuli Responsiveness and Polymerization-Induced Self-Assembly in the Presence of Air. ACS Macro Lett. 2017, 6 (11), 1237−1244. (53) Ng, G.; Yeow, J.; Xu, J. T.; Boyer, C. Application of oxygen tolerant PET-RAFT to polymerization-induced self-assembly. Polym. Chem. 2017, 8 (18), 2841−2851. (54) Shanmugam, S.; Xu, J.; Boyer, C. Exploiting Metalloporphyrins for Selective Living Radical Polymerization Tunable over Visible Wavelengths. J. Am. Chem. Soc. 2015, 137 (28), 9174−9185. (55) Tucker, B. S.; Coughlin, M. L.; Figg, C. A.; Sumerlin, B. S. Grafting-From Proteins Using Metal-Free PET−RAFT Polymerizations under Mild Visible-Light Irradiation. ACS Macro Lett. 2017, 6 (4), 452−457. (56) Braunecker, W. A.; Matyjaszewski, K. Controlled/living radical polymerization: Features, developments, and perspectives. Prog. Polym. Sci. 2007, 32 (1), 93−146.
H
DOI: 10.1021/acs.macromol.8b01600 Macromolecules XXXX, XXX, XXX−XXX
