LCST behavior of poly(2-ethyl-2-oxazoline) containing diblock and triblock copolymers

European Polymer Journal 100 (2018) 57–66

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LCST behavior of poly(2-ethyl-2-oxazoline) containing diblock and triblock copolymers

T

Martin Sahn, Leanne M. Stafast, Michael Dirauf, Damiano Bandelli, Christine Weber, ⁎ Ulrich S. Schubert Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University Jena, Humboldtstr. 10, 07743 Jena, Germany Jena Center for Soft Matter (JCSM), Friedrich Schiller University Jena, Philosophenweg 7, 07743 Jena, Germany

A R T I C L E I N F O

A B S T R A C T

Keywords: Cationic ring-opening polymerization Block copolymer Poly(N-isopropylacrylamide) Poly(2-ethyl-2-oxazoline) Poly(ethoxytriethyleneglycol acrylate) Lower critical solution temperature

Mono- and bifunctional macro chain transfer agents were prepared via the living cationic ring opening polymerization of EtOx. A series of well-defined P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers, PEtOx-b-P (eTEGA) as well as PEtOx-b-PNiPAm diblock copolymers were obtained by subsequent reversible addition fragmentation chain transfer polymerization of eTEGA or NiPAm, respectively. While the molar mass of the PEtOx was kept constant, the DP of the PNiPAm or P(eTEGA) block was varied and covers a wide range. Turbidimetry, NMR spectroscopy and dynamic light scattering investigations of the aqueous polymer solutions revealed that the lower critical solution temperature behavior of the triblock copolymers is strongly affected by the hydrophobic end groups whereas the presence of hydrogen bond donating moieties does not play any role. For diblock copolymers with a sufficiently long PEtOx block, the PNiPAm blocks collapsed independently, resulting in multiple phase transitions and the formation of well-defined aggregated structures.

1. Introduction Thermoresponsive polymers are known as “smart materials” and are appealing for a wide range of applications due to their ability to alter the properties by an external trigger [1]. In particular, polymers exhibiting a lower critical solution temperature (LCST) are in focus of interest and well-studied [2]. Poly(N-isopropylacrylamide) (PNiPAm) is probably the most investigated polymer that reveals a “switch” from hydrophilic to hydrophobic properties and is considered to be the gold standard in that field [3]. Furthermore, its reversible coil-to globule phase transition temperature at 32 °C [4,5] is slightly below body temperature making it interesting for biomedical applications [6–10]. In this entropy driven process, hydrogen bonds between the polymer and water molecules are weakened at elevated temperatures leading to demixing, i.e. to the formation of a biphasic system comprising a higher and a lower concentrated polymer phase [11]. Tuning of the coil-toglobule transition temperature can be achieved by copolymerization of NiPAm with other monomers [12,13]. PNiPAm has been combined with permanently hydrophilic blocks such as, e.g., poly(ethylene oxide) (PEO) in different architectures to adjust the thermoresponsive behavior. In particular, aqueous solutions of PNiPAm-b-PEO-b-PNiPAm triblock copolymers exhibit a thermally induced reversible sol-gel transition due to the permanent solvation of

⁎

the middle block [14–16]. Furthermore, PEO-b-PNiPAm diblock copolymers undergo thermally induced self-assembly processes forming micelles or vesicles in aqueous solution upon elevated temperature [17,18]. Poly(ethoxytriethyleneglycol acrylate) (P(eTEGA)) represents a biocompatible [19,20] and thermoresponsive polymer with a reported molar mass dependent cloud point temperature (Tcp) of 34–39 °C [21–24] in aqueous solution that is similar to PNiPAm (Tcp = 32 °C). In contrast to PNiPAm, P(eTEGA) does not feature any hydrogen bond donor moieties. Also P(eTEGA) was combined in di-[25,26] and triblock [27] architectures with PEO and the resulting polymers showed the expected sol-gel transitions. However, the studies focused on P (eTEGA)-based building blocks comprising o-nitrobenzyl acrylate or acrylic acid in varying molar fractions [25–27]. To the best of our knowledge, a comprehensive direct comparison between PNiPAm and P (eTEGA) as thermo-responsive building blocks combined with a hydrophilic building block in di- and triblock architectures is missing to date. For this purpose, we selected a hydrophilic poly(2-oxazoline) (POx) block since it is known that PEO shows some drawbacks such as nonbiodegradability or possible accumulation in the body [28]. The hydrophilic poly(2-ethyl-oxazoline) (PEtOx) features appropriate properties, such as stealth effect and biocompatibility [28,29]. The cationic

Corresponding author at: Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University Jena, Humboldtstr. 10, 07743 Jena, Germany. E-mail address: [email protected] (U.S. Schubert).

https://doi.org/10.1016/j.eurpolymj.2018.01.014 Received 14 November 2017; Received in revised form 9 January 2018; Accepted 16 January 2018 0014-3057/ © 2018 Elsevier Ltd. All rights reserved.

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Scheme 1. Schematic representation of the synthesis route towards P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers or PEtOx-b-P(eTEGA) and PEtOx-b-PNiPAm diblock copolymers, respectively.

filtration of potassium hydroxide it was dried over calcium hydride, distilled and stored under argon atmosphere. 4,4′-Bis(bromomethyl) biphenyl (BBMBP, ≥97% purity), N-isopropylacrylamide (NiPAm, ≥97% purity), acryloyl chloride (≥97% purity) and 4,4′-azobis(4-cyanovaleric acid) (ACVA, ≥98% purity) were purchased from SigmaAldrich. Triethylene glycol monoethylether (> 90.0% purity) was bought from TCI. 2-(Butylthiocarbonothioylthio)-propanoic acid (BTPA) was a kind gift from the BASF and used as received.

ring-opening polymerization (CROP) provides access to tailor-made POx. Its livingness enables the introduction of various functionalities into the polymer by using functional initiators [30] and end cappingagents [31], respectively. Sequential monomer addition [32–35] or crossover techniques [36,37] enable the preparation of block copolymers. In particular, block copolymers comprising PEtOx and PNiPAm building blocks have been obtained via a crossover of CROP and radical addition fragmentation transfer (RAFT) polymerization technique using PEtOx based macro chain transfer agents (MCTA) obtained by quenching the living CROP with a carboxylic acid functionalized CTA [37,38]. The utilization of methyl tosylate as CROP initiator resulted in a monofunctional MCTA providing access to diblock copolymers [37]. Bifunctional MCTA and the according symmetrical BAB triblock copolymers can be obtained using the bifunctional CROP initiator 4,4′-bis (bromomethyl)biphenyl (BBMBP). Both MCTA types should be efficient in controlling the polymerization of NiPAm as well as eTEGA, facilitating access to a range of block copolymers suitable to directly compare the influence of hydrogen bond donating moieties and the block copolymer architecture on the LCST behavior (Scheme 1). Featuring hydrogen bond accepting moieties, high molar mass PEtOx exhibits LCST behavior in water at temperatures above 70 °C [39]. The PEtOx building blocks were hence kept rather short, whereas the degree of polymerization (DP) of the PNiPAm and P(eTEGA) blocks, respectively, was varied over a broad range.

2.2. Instruments The polymerization of EtOx was performed in a Biotage Initiator Sixty microwave synthesizer. Size exclusion chromatography (SEC) was measured on an Agilent 1200 series equipped with a G1310A pump, a G1315D DA detector, a G1362A RI detector, and PSS GRAM 30 Å/1000 Å (10 µm particle size) columns in series at 40 °C using N,N-dimethylacetamide (DMAc) with 2.1 g L−1 LiCl as eluent at a flow rate of 1 mL min−1. The system was calibrated with PS standards (374–1,040,000 Da). 1 H NMR spectra were recorded on a Bruker Avance 300 MHz or a Bruker Avance 400 MHz using the residual solvent resonance as an internal standard. The chemical shifts are given in ppm relative to trimethylsilane. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectra were acquired with an Ultraflex III ToF/ToF instrument (Bruker Daltonics, Bremen, Germany). The instrument is equipped with a Nd-YAG laser. All spectra were measured in the positive reflector mode. The instrument was calibrated prior to each measurement with an external PMMA standard from PSS. Samples were spotted using the dried droplet technique applying 2,5-dihydroxybenzoic acid (DHB) as a matrix and NaI as a doping salt. For the electrospray ionization time-of-flight mass spectrometry (ESI-ToF-MS) measurements, samples were analyzed by using a microToF Q-II (Bruker Daltonics) mass spectrometer equipped with an automatic syringe pump from KD Scientific for sample injection. The mass spectrometer was running at 4.5 kV, at a desolvation temperature of 180 °C and was operated in the positive ion mode. Nitrogen was used as the nebulizer and drying gas. All fractions were injected with a constant flow rate (3 µL min−1) of sample solution. The instrument was calibrated in the m/z range from 50 to 3000 with a calibration standard

2. Experimental part 2.1. Materials Unless specified otherwise, all chemicals were used without further purification. 2-Ethyl-2-oxazoline (EtOx, Acros Organics) and methyl ptoluenesulfonate (MeOTs, Sigma Aldrich) were dried over barium oxide, distilled and stored under argon atmosphere. N,NDimethylformamide (DMF, Sigma Aldrich), acetonitrile (ACN, Sigma Aldrich) and dichloromethane were dried in a solvent purification system (Pure Solv EN, InnovativeTechnology) before use as polymerization or reaction solvent. Triethylamine (NEt3) was dried over potassium hydroxide overnight and refluxed for three hours. After 58

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Table 1 Characterization data of the synthesized P(eTEGA)-b-PEtOx20-b-P(eTEGA) triblock copolymers P1 to P6 synthesized from MCTA1. Sample

M/CTA

Conv. [%]a

DP eTEGAb

DP eTEGA (NMR)c

Composition eTEGA-EtOxeTEGAc

Mass% EtOxf

Mn (NMR) [g/mol]

Mn (SEC) [g/mol]d

Ðd

Tcp [°C]

MCTA1 P1 P2 P3 P4 P5 P6

– 20 40 60 80 100 120

– 51 41 41 58 57 67

– 10 16 25 47 57 80

– 10 18 28 36 46 62

0-20-0 5-20-5 9-20-9 14-20-14 18-20-18 23-20-23 32-20-32

76 40 29 22 18 15 12

2600 5000 6800 9100 11,000 13,300 17,000

3500 7300 8400 9900 11,400 12,900 17,200

1.14 1.14 1.14 1.17 1.16 1.20 1.25

– 18.5 22.4 29.0 31.2 32.2 34.6

e

a

Conversion determined from the 1H NMR spectra of the reaction solution. Overall degree of polymerization (DP) including both eTEGA blocks calculated from conversion and [M]/[CTA]. c Overall DP including both eTEGA blocks calculated from suitable peak integrals in the 1H NMR spectra of the purified polymers. d SEC: DMAc + 0.21% LiCl, RI detection, PS calibration. e 50% transmittance, 2nd heating run, heating rate 1 K min−1, concentration of 5 g L−1 in water. f End groups were considered separately from the EtOx fraction. b

(21 mL; 23.3 g; 257.53 mmol) in dry dichloromethane (100 mL) was added dropwise with continuous stirring (1 h). The mixture was stirred at room temperature for 18 h. The reaction was quenched by the addition of ice water. The organic layer was washed with brine (1 × 100 mL), deionized water (3 × 100 mL), collected and dried over anhydrous sodium sulfate. The crude product was concentrated (T < 30 °C) and purified by column chromatography using silica gel as stationary phase and mixture of ethyl acetate/hexane (50/50, v/v) as eluent. The desired fraction with Rf = 0.57 was collected. The volatiles were removed under reduced pressure (T < 30 °C) to isolate the final product as a pale yellow oil. Yield: 22.75 g (76%). The 1H NMR spectrum incl. peak assignment is shown in Fig. SI 1. 1 H NMR (300 MHz, CDCl3): δ = 6.39 (dd, J = 17.3, 1.0 Hz, 1H C]CeH), 6.11 (dd, J = 17.3, 10.4 Hz, 1H C]CeH), 5.80 (dd, J = 10.4, 1.2 Hz, 1H, C]CeH), 4.39–4.17 (m, 2H, CH2), 3.80–3.42 (m, 12H, OeCH2eCH2eO), 1.17 (t, J = 7.0 Hz, 3H, CH3) ppm.

(Tunemix solution) which is supplied from Agilent. All data were processed via Bruker Data Analysis software version 4.2. Cloud points were determined in a Crystal 16 from Avantium Technologies connected to a chiller (Julabo FP 40) at a wavelength of 500 nm. After sample preparation, all samples were stored in a fridge at 4 °C overnight to ensure the presence of homogeneous solutions. Prior to starting the measurement, all samples were kept at 1 °C for 10 min inside the instrument. Subsequently, the solutions were heated and cooled at a rate of 1 K min–1 in a temperature range between 1 °C and 100 °C. Three consecutive heating/cooling cycles were performed without interruption of the measurement using the heating program previously defined. If not specified otherwise the cloud point temperature Tcp was defined as the temperature where the transmittance decreased to 50% in the second heating run. Dynamic light scattering (DLS) measurements were performed on a Zetasizer Nano ZS (Malvern Instruments, Herrenberg, Germany) at a scattering angle of 173 °C. At each temperature, the samples were equilibrated for 240 s and then 3 × 30 runs were carried out with a delay time of 120 s. The number, intensity and the volume distribution of the hydrodynamic diameter were calculated applying the non-linear least square fitting mode.

2.3.4. General procedure for the RAFT polymerization to yield the P (eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers P1 to P6 For each polymerization, the [MCTA1]/[ACVA] ratio was kept as 1/ 0.25 and the initial monomer concentration was 2 mol L–1 in DMF. The [MCTA1]/[eTEGA] ratio was varied according to Table 1. In a representative example for P4 ([M]/[MCTA] = 80), eTEGA (2.0 g, 8.61 mmol), MCTA1 (289 mg, 0.11 mmol) and ACVA (7.54 mg, 0.03 mmol) were dissolved in DMF (4.30 mL) in a microwave vial. After taking a sample for the determination of the monomer conversion, the vial was closed with a suitable septum. The reaction mixture was subsequently flushed with a gentle flow of argon for 1 h. The reaction vessel was heated in an oil bath at 70 °C for 2 h and 15 min. The mixture was cooled to room temperature and a sample for determination of monomer conversion was taken. The solvent was evaporated under reduced pressure. The polymer was dissolved in THF and separated from residual monomer on a preparative size exclusion chromatography column (BioBeads SX-1). The solvent was removed under reduced pressure and the polymer was dried in a vacuum oven overnight. A representative 1H NMR spectrum incl. peak assignment is shown in Fig. 2. 1 H NMR (300 MHz, CDCl3) P1 to P6: δ = 7.57 (CeH (1)), 7.23 (CeH (2)), 4.61 (CH2 (3)), 4.18 (O]CeCH2 (4)), 3.84–3.30 (CH2 (5)), 2.40 (CH2 (6)), 2.05–1.37 (CH2 (7) + CH (8)), 1.21–1.12 (CH3 (9)), 1.12 (CH3 (10)) ppm.

2.3. Synthesis 2.3.1. Synthesis of the bifunctional macro chain transfer agent (MCTA1) The synthesis of MCTA1 was performed as described in literature [38]. The 1H NMR spectrum incl. peak assignment is shown in Fig. SI 2. 1 H NMR (300 MHz, CDCl3): δ = 7.55 (CeH), 7.24 (CeH), 4.80 (CeH), 4.60 (CH2), 4.25 (CH2), 3.45 (CH2), 2.39 (CH2), 1.12 (CH3), 0.92 (CH3) ppm. DP = 20, DF = quant. (99.3% determined by 1H NMR). 2.3.2. General procedure for the synthesis of the monofunctional macro chain transfer agents (MCTA2 and MCTA3) MCTA2 and MCTA3 were synthesized as described in literature [37]. The 1H NMR spectra incl. peak assignments are shown in Fig. SI 3. 1 H NMR (300 MHz, CDCl3): δ = 4.83 (CH), 4.27 (CH2), 3.48 (CH2), 3.05 (CH3), 2.39 (CH2), 1.65 (CH2), 1.14 (CH3), 0.95 (CH3) ppm. MCTA2: DP = 20, DF = 96% MCTA3: DP = 50, DF = 99%. 2.3.3. Synthesis of triethylene glycol monoethylether acrylate (eTEGA) ETEGA was synthesized in a modified protocol known from literature [20,24]. Therefore, 35.7 mL (26.06 g; 257.5 mmol) of dry triethylamine was added to a solution of triethylene glycol monoethylether (25.0 mL; 22.95 g; 129.0 mmol) in 200 mL of dry dichloromethane. The mixture was cooled in an ice-bath and a solution of acryloyl chloride

2.3.5. General procedure for the RAFT polymerization to yield the PEtOx-bP(eTEGA) diblock copolymers P7 to P9 For each polymerization, the [MCTA2]/[ACVA] ratio was kept as 1/ 0.25 and the initial monomer concentration was 2 mol L–1 in DMF. The [MCTA2]/[eTEGA] ratio was varied according to Table 2. In a representative example for P7 ([M]/[MCTA2] = 13), eTEGA 59

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Table 2 Characterization data of the PEtOx20-b-P(eTEGA) diblock copolymers P7 to P9 synthesized from MCTA2. Sample

M/CTA

Conv. [%]a

DP eTEGAb

DP eTEGA (NMR)

MCTA2 P7 P8 P9

– 13 43 33

– 88 48 88

– 11 21 29

– 10 16 27

c

Composition EtOx-eTEGAc

Mass% EtOxf

Mn (NMR) [g/mol]c

Mn (SEC) [g/mol]d

Ðd

Tcp [°C]

20-0 20-10 20-16 20-27

91 44 33 23

2200 4600 6000 8500

3800 6600 8200 14,600

1.14 1.17 1.15 1.30

79.0 43.1 38.4 33.3

e

a

Conversion determined from the 1H NMR spectra of the reaction solution. Degree of polymerization (DP) calculated from conversion and [M]/[CTA]. c DP calculated from suitable peak integrals in the 1H NMR spectra of the purified polymers. d SEC: DMAc + 0.21% LiCl, RI detection, PS calibration. e onset, 2nd heating run, heating rate 1 K min−1, concentration of 5 g L−1 in water. f End groups were considered separately from the EtOx fraction. b

(325 mg, 1.4 mmol), MCTA2 (238 mg, 0.11 mmol) and ACVA (7.5 mg, 0.26 mmol) were dissolved in DMF (1.07 mL) in a microwave vial. After taking a sample for the determination of the monomer conversion, the vial was closed with a suitable septum. The reaction mixture was subsequently flushed with a gentle flow of argon for 1 h. The reaction vessel was heated in an oil bath at 70 °C for 1 h. The mixture was cooled to room temperature and a sample for determination of monomer conversion was taken. The solvent was evaporated under reduced pressure and the polymer was dissolved in THF and separated from residual monomer on a preparative size exclusion chromatography column (BioBeads SX-1). The solvent was removed under reduced pressure and the polymer was dried in a vacuum oven overnight. A representative 1H NMR spectrum incl. peak assignment is shown in Fig. SI 8. 1 H NMR (300 MHz, CDCl3) P7 to P9: δ = 4.21 (O]CeCH2), 3.84–3.32 (CH2), 3.05 (CH3) 2.38 (CH2), 2.11–1.50 (CH2 + CH), 1.24–1.15 (CH3) ppm.

polymers were dissolved in THF. The polymers were run over a preparative size exclusion chromatography column filled with BioBeads SX-1 to separate the residual MCTA2. The purified polymers were obtained by precipitation into cold diethyl ether, filtered and dried in a vacuum oven overnight. A representative 1H NMR spectrum incl. peak assignment is shown in Fig. 6. 1 H NMR (300 MHz, CDCl3) P10 to P17: δ = 6.30 (NH), 4.03 (CH), 3.47 (CH2), 3.04 (CH3), 2.38 (CH2), 2.10–1.55 (CH2 + CH), 1.16 (CH3) ppm. 3. Results and discussion 3.1. Synthesis of block copolymers comprising P(eTEGA) and PEtOx To explore if the block copolymer architecture affects the thermoresponsive properties, AB as well as BAB di- and triblock copolymers were synthesized keeping the DP of the PEtOx block (A block) constant at a constant value 20. For the bifunctional MCTA1, which was synthesized using BBMBP as bifunctional initiator for the CROP of EtOx, this corresponds to a DP of 10 per initiation moiety. As described previously, the CROP was quenched using the in situ deprotonated caboxylic acid functionalized CTA BTPA (Scheme 1) [38]. The according monofunctional MCTA2 was obtained using MeOTs as initiator for the CROP of EtOx at an initial [M]/[I] ratio of 20 and the same end-capping procedure [37]. Characterization by means of SEC, 1H NMR spectroscopy and ESI-MS as well as MALDI MS confirmed the narrow molar mass distribution and the covalent attachment of the desired trithiocarbonate at the ω-chain ends for both MCTAs (see supporting information). The MCTAs were utilized for the RAFT polymerization of eTEGA to obtain two series of P(eTEGA)-b-PEtOx20-b-P(eTEGA) triblock copolymers (P1-P6) and PEtOx-b-P(eTEGA) diblock copolymers (P7-P9),

2.3.6. General procedure for the RAFT polymerization to yield the PEtOx-bPNiPAm diblock copolymers P10 to P17 For each polymerization, the [MCTA]/[ACVA] ratio was kept as 1/ 0.25 and the initial monomer concentration was 2 mol L–1 in DMF. The [MCTA]/[NiPAm] ratio was varied according to Table 3. In a representative example for P8, NiPAm (348 mg, 3.09 mmol), MCTA2 (172 mg, 0.08 mmol) and ACVA (5.4 mg, 0.02 mmol) were dissolved in DMF (1.43 mL) in a microwave vial. After taking a sample for the determination of the monomer conversion, the vial was closed with a suitable septum. The reaction mixture was subsequently flushed with a gentle flow of argon for 1 h. The reaction vessel was heated in an oil bath at 70 °C for 16 h. The mixture was cooled to room temperature and a sample for the determination of the monomer conversion was taken. The solvent was evaporated under reduced pressure and the

Table 3 Selected characterization data of the PEtOx-b-PNiPAm diblock copolymers P10 to P17 synthesized from MCTA2 (DP = 20) and MCTA3 (DP = 50). Sample

M/CTA

Conv. [%]a

DP NiPAmb

DP NiPAm (NMR)c

Composition EtOx-NiPAmc

Mass% EtOxf

Mn (NMR) [g/mol]c

Mn (SEC) [g/mol]d

Ðd

Tcp [°C]e

MCTA2 P10 P11 P12 P13 P14 MCTA3 P15 P16 P17

– 10 20 40 60 80 – 10 20 120

– 97 96 96 95 95 – 65 55 93

– 10 19 39 57 76 – 6.5 11 112

– 10 19 47 69 94 – 10 27 75

20-0 20-10 20-19 20-47 20-69 20-94 50-0 50-10 50-27 50-75

91 59 50 26 20 15 97 79 60 36

2200 3400 4400 7600 10,000 12,900 5100 6300 8300 13,700

3800 6700 9300 13,100 16,000 19,700 9900 12,000 13,000 20,500

1.14 1.09 1.10 1.07 1.08 1.09 1.09 1.08 1.10 1.13

79.0 54.3 52.1 45.2 40.6 39.0 – 85.9 72.0 49.0

a

Conversion determined from the 1H NMR spectra of the reaction solution. Degree of polymerization (DP) calculated from conversion and [M]/[CTA]. c DP calculated from suitable peak integrals in the 1H NMR spectra of the purified polymers. d SEC: DMAc + 0.21% LiCl, RI detection, PS calibration. e Onset, 2nd heating run, heating rate 1 K min−1, concentration of 5 g L−1 in water; partly multiple transmissions observed, see Fig. 7. f End groups were considered separately from the EtOx fraction. b

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a representative example for a P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymer. The spectrum of the PEtOx-b-P(eTEGA) diblock copolymer P7 is provided as example in the supporting information. As evident from the peak assignments, all signals deriving from the P(eTEGA) blocks overlap with signals assigned to the PEtOx block. However, the PEtOx α-end group signals remained isolated for both block copolymer types, and the absence of PEtOx homopolymer had been confirmed via SEC. The peaks derived from the biphenyl moiety (signal 1 in Fig. 2) and the methylene group next to the ester functionality of the P(eTEGA) block (signal 4 in Fig. 2) were hence utilized to estimate the DP of the P (eTEGA) blocks. The underlying four protons derived from the PEtOx block were taken into account during the calculation. The composition of the diblock copolymers P7 to P9 was determined in a similar fashion using the peaks of the methyl α-end group of the PEtOx block. The fact that end group signals had to be used to determine the copolymer composition via 1H NMR spectroscopy explains why the resulting values deviate from the values calculated from the initial [M]/[MCTA] and the monomer conversion.

Fig. 1. Overlay of the normalized SEC traces of the AB and BAB block copolymers comprising PEtOx and P(eTEGA) (DMAc/0.21% LiCl, RI detection, PS calibration). Bottom: P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers P1 to P6 obtained from MCTA1. Top: PEtOx-b-P(eTEGA) diblock copolymers P7 to P10 obtained from MCTA2.

respectively. For this purpose, ACVA was used as initiator at a molar ratio of [MCTA]/[ACVA] of 1/0.25. All polymerizations were conducted at 70 °C in DMF using an initial monomer concentration of 2 mol L–1. The ratio of [eTEGA] to [MCTA] was varied to obtain di- and triblock copolymers suitable to investigate the influence of the fraction of hydrophilic and thermoresponsive block on the LCST behavior of the aqueous solutions. Therefore, we aimed at a broad variation of the DP of the P(eTEGA) blocks (Tables 1 and 2). The polymerizations were stopped at moderate monomer conversions because a high molar mass shoulder occurred in the SEC traces at prolonged reaction times and, thus, higher conversions. This could be caused by chain coupling reactions during the RAFT polymerization or by a small amount of triethylene glycol bisacrylate that might have served as crosslinking agent during the polymerization. The resulting triblock copolymers were purified by preparative size exclusion chromatography (BioBeads SX-1) because the residual monomer could not be separated by precipitation. As expected for RAFT polymerizations, the SEC elugrams of both block copolymer series revealed monomodal molar mass distributions with narrow dispersity (Đ) values below 1.30 (Fig. 1). In addition, all block copolymer SEC traces are clearly shifted to lower elution volumes compared to those of the respective MCTA, demonstrating the absence of PEtOx homopolymers and the effectiveness of the MCTAs for the control of the molar mass during the RAFT polymerization process. However, the extracted molar mass values are based on PS calibration and, thus, represent relative values. The composition of the purified block copolymers was determined via 1H NMR spectroscopy. Fig. 2 depicts the 1H NMR spectrum of P3 as

3.2. Synthesis of PEtOx-b-PNiPAm diblock copolymers We previously reported the synthesis of a series of PNiPAm-b-PEtOxb-PNiPAm triblock copolymers with a constant DP of the PEtOx block of 20 [38]. This was now complemented by an according library of PEtOxb-PNiPAm diblock copolymers with varied DP of the PNiPAm block to facilitate a direct comparison of their solution behavior with the respective block copolymers containing P(eTEGA). Therefore, MCTA2 was utilized for the RAFT polymerization of NiPAM under same conditions as for the RAFT polymerization of eTEGA (P10 to P14, Table 3). In addition, PEtOx-b-PNiPAm diblock copolymers served as an example to assess the influence of the molar mass of the PEtOx block on the thermoresponsive behavior of the block copolymers. For this purpose, the monofunctional PEtOx based MCTA3 with a DP of 50 was utilized to gain access to block copolymers that featured also a significantly lower DP of the PNiPAM block (P15 to P17, Table 3). As evident from the SEC elugrams of the PEtOx-b-PNiPAm diblock copolymers, monomodal and narrow molar mass distributions with Đ < 1.15 were obtained even at high NiPAM conversions (Fig. 3). The SEC traces are shifted to lower elution volumes with increasing length of the PNiPAM block for both block copolymer series. In particular, the SEC trace of P14 is completely shifted compared to that of the corresponding MCTA2, showing the absence of residual MCTA. The copolymer composition was calculated from the 1H NMR spectra of the purified polymers by comparing the peak integrals of the four backbone protons of the PEtOx block (signal 4, Fig. 4) with the methine proton signals of the PNiPAm block (signal 3, Fig. 6). In contrast to the block copolymers containing P(eTEGA), the resulting values are more accurate as the method does not rely on the integration of end group signals.

Fig. 3. Overlay of the normalized SEC traces (DMAc/0.21% LiCl, RI detection, PS calibration) of the PEtOx-b-PNiPAm diblock copolymers. Bottom: MCTA2 and the diblock copolymers P10 to P14 featuring a PEtOx DP of 20. Top: MCTA3 and the diblock copolymers P15 to P17 featuring a PEtOx DP of 50.

Fig. 2. 1H NMR spectrum (300 MHz, CDCl3) of the P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymer P3 and assignment of the peaks to the schematic representation of the structure.

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the reported Tcp of a P(eTEGA) homopolymer solution (34–39 °C) [23], and solutions of polymers with longer P(eTEGA) blocks approach the Tcp of P(eTEGA) solutions. Apart from P1 and P2 featuring rather short P(eTEGA) blocks, which formed large aggregates at higher temperatures, the triblock copolymer solutions displayed sharp transitions with very little heating/cooling hysteresis (see Supporting Information). The direct comparison between both triblock series clearly showed that the evolution of the Tcp is independent from the presence (PNiPAM) or absence (P(eTEGA)) of hydrogen bond donor moieties (Fig. 6). In contrast, the unexpectedly low Tcp of the triblock copolymers most likely results from the effect of the hydrophobic end groups: In addition to the two trithiocarbonate end groups derived from the RAFT polymerization, the central biphenyl moiety derived from the CROP initiator adds to the hydrophobicity. Indeed, the Tcp showed a linear dependence on the mass fraction of hydrophobic end groups for both triblock copolymer series (Fig. 6). The fact that the y-axis intercepts are higher than the Tcp values of the two homopolymers, i.e. PNiPAM and P (eTEGA), can be explained by the contribution of the hydrophilic PEtOx to the overall hydrophilic hydrophobic balance of the triblock copolymers.

Fig. 4. 1H NMR spectrum (300 MHz, CDCl3) of the PEtOx-b-PNiPAm diblock copolymer P12 and assignment of the peaks to the schematic representation of the structure.

3.3.2. Diblock copolymers The effect of the hydrophobic end groups was significantly reduced for the corresponding diblock copolymer series, i.e. PEtOx-b-P(eTEGA) P7 to P9 and PEtOx-b-PNiPAm P10 to P14. Featuring the same PEtOx DP of 20, the diblock copolymers lack the biphenyl spacer and feature a methyl α-end group derived from the CROP initiator and one trithiocarbonate ω–end group. The PEtOx-b-PNiPAm and the PEtOx-b-P (eTEGA) diblock copolymers revealed the same trend regarding their solution behavior with increasing DP of the second thermoresponsive block (Fig. 7): An increase of the DP of the vinylic block led to a decrease of the Tcp of the aqueous solution of the respective diblock copolymer. In other words, the Tcp increased with increasing mass fraction of the hydrophilic PEtOx block (Fig. 8). For block copolymers with short vinylic blocks (P7 to P11), single phase transitions were observed via turbidimetry. Upon increase of the DP of the PNiPAm block (P12 to P14), multiple transitions became visible. A first decrease of the transmittance took place between 39 and 45 °C and can be associated to a collapse of the PNiPAm block. The PEtOx block remains still hydrated and acts as hydrophilic end group, explaining the elevated Tcp compared to a PNiPAm homopolymer solution. Further heating presumably led to a collapse of the PEtOx block, representing the second drop of the transmittance (between 45 and 55 °C). These values are significantly lowered compared to the collapse of the PEtOx MCTA2 at 79 °C (see supporting information) because the collapsed PNiPAm block acts as large hydrophobic “end group”. In direct comparison to PEtOx-b-P(eTEGA), solutions of the copolymers comprising PNiPAm revealed sharper transitions. Regardless if DP or the mass fraction were considered, turbidimetry did not hint at any individual collapse of the blocks for PEtOx-b-P(eTEGA). Hence, block copolymers comprising a longer PEtOx building block (DP = 50) focused on PNiPAm (see supporting information). In particular P17 featuring a PNiPAm DP of 75 exposed to a more complex two stage phase transition, whereas the aqueous solutions of the two block copolymers P15 and P16 with shorter PNiPAm blocks revealed a rapid decrease in transmittance above 70 °C.

3.3. LCST behavior of the aqueous block copolymer solutions Turbidimetry measurements [40] were performed to determine the cloud point temperature (Tcp) of the triblock copolymers in aqueous solution with a heating and cooling rate of 1 K min−1. Three heating/ cooling cycles were conducted in a temperature range between 1 and 100 °C. At lower temperatures, the polymers are completely dissolved while an increase of temperature leads to a phase separation and, thus, a decrease of the transmittance of the aqueous solutions. All phase transitions were fully reversible with no notable differences between the individual heating and cooling runs.

3.3.1. Triblock copolymers Recently, we reported single phase transitions of aqueous solutions of a series of PNiPAm-b-PEtOx-b-PNiPAm triblock copolymers [38]. The Tcp was lower compared to that of PNiPAm homopolymer solutions despite the presence of the central (hydrophilic) PEtOx block. The P (eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers P1 to P6 are structurally similar in terms of the central block and the DPs of the individual blocks. The fact that P(eTEGA) does not feature any hydrogen bond donating moieties enables the elucidation of a potential contribution of intramolecular hydrogen bonding between the blocks. Turbidimetry of aqueous solutions of P1 to P6 revealed a similar behavior (Fig. 5): The phase separation takes place at temperatures below

3.4. 1H NMR investigation of the thermo-responsive behavior in aqueous solution To evaluate if the blocks indeed collapsed independently from each other, NMR experiments in D2O were performed to monitor the solvation of the PEtOx and the P(eTEGA) and PNiPAm block, respectively, at varying temperatures below and above the Tcp. For this purpose, a representative example of each polymer type was selected and 1H NMR spectra were measured in steps of 5 K. To account for the different

Fig. 5. Transmittance curves of solutions of the P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers P1 to P6 at a concentration of 5 g L−1 in water (2nd heating run, heating rate 1 K min−1).

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Fig. 6. Comparison of the cloud point temperatures (Tcp) of the aqueous solutions of PNiPAm-b-PEtOx20-b-PNiPAm [38] and P(eTEGA)-b-PEtOx20-b-P(eTEGA) triblock copolymers (c = 5 g L−1, Tcp reported at 50% transmittance during the second heating run). DP refers to the DP of the P(eTEGA) and PNiPAm block, respectively.

The difference is less pronounced for the diblock copolymer PEtOx20-bPNiPAm47 P12, hinting towards the fact that the mobility of the rather short PEtOx block is strongly affected by the collapse of the PNiPAm block. In contrast, the longer hydrophilic PEtOx block of PEtOx50-bPNiPAm75 was almost unaffected by the collapse of the PNiPAm block. More than 80% of the PEtOx units remained solvated over the entire investigated temperature range up to 70 °C, and almost no difference could be detected at the Tcp determined by turbidimetry (onset value). However, the NMR experiments clearly indicated that the phase transition (or the re-organization of the block copolymer) takes place over a broader temperature range for both P12 and P17 as the PNiPAm blocks start to lose their mobility below the Tcp and are not fully immobilized even at 70 °C, i.e. more than 20 °C above the Tcp. The latter could be an effect of the covalently attached PEtOx block, which is still well solvated and, therefore, could enhance the mobility of parts of the PNiPAm block.

signal intensities assigned to the varying structural moieties of block copolymers, the spectra were first normalized according to the residual solvent peak. Subsequently, the integrals of each signal were related to the integral of the same signal at 298 K using Eq. (1) [41]. I represents the integral of a signal at the given temperature and I0 represents the integral of the same signal at 298 K, i.e. below the Tcp. The resulting value p hence describes the fraction of the respective structural moiety with reduced mobility compared to room temperature.

p= 1−(I0 /I)

(1)

Although the resolution of the individual peaks decreased in the 1H NMR spectra recorded above the Tcp of the solutions of the P(eTEGA)containing block copolymers P4 and P7, the signal integrals did not decrease significantly, regardless if assigned to the PEtOx or P(eTEGA) block (see supporting information). A similar behavior was reported for poly(methoxy triethylene glycol methacrylate) revealing mainly structural changes of the polymethacrylate backbone upon phase transition of the aqueous solution [42]. The fact that the according P(eTEGA) backbone signals partly overlap with signals assigned to the PEtOx block makes it difficult to draw clear conclusions about a potential individual collapse of the blocks in aqueous solutions of P4 and P7. Different observations were made for the PEtOx-b-PNiPAm diblock copolymers P12 and P17 (Fig. 9). Even without an integration of the signals in the 1H NMR spectra, it is visible that the signals of the PNiPAm block (A, D in Fig. 9) largely disappeared above the Tcp of the aqueous solution whereas the PEtOx signals (B, C) were fewer affected.

3.5. DLS investigations of PEtOx-b-PNiPAm diblock copolymers As the NMR experiments suggested an independent coil-to-globule transition of the PNiPAm block for P17, a thermo-induced self-assembly could be anticipated. As a consequence, the aqueous solution was investigated by means of DLS below the Tcp, at the onset of the transmittance drop at 50 °C and at the onset of the second drop in transmittance at 70 °C (Fig. 10). At 10 °C, a few polydisperse aggregates were observed whereas the major fraction of polymer chains remained

Fig. 7. Transmittance curves of the aqueous solutions of the PEtOx-b-P(eTEGA) diblock copolymers P7 to P9 and the PEtOx-b-PNiPAm diblock copolymers P10 to P14 featuring a PEtOx DP of 20 (c = 5 g L−1, 2nd heating run, heating rate 1 K min−1).

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Fig. 8. Comparison of cloud point temperatures Tcp of the aqueous polymer solutions of the PEtOx-b-PNiPAm and PEtOx-b-P(eTEGA) diblock copolymers (onset values, c = 5 g L–1). The asterisks indicate multiple transitions observed via turbidimetry. DP refers to the DP of the P(eTEGA) and PNiPAm block, respectively.

4. Conclusion

dissolved (Dh < 5 nm). Upon collapse of the PNiPAm block, the block copolymer assembled to particles with Dh ≈ 140 nm that formed very well-defined aggregates at 70 °C (PdI = 0.046) in aqueous solution upon the collapse of the PNiPAm block, which is in agreement with the results obtained from 1H NMR spectroscopy. As the conclusions drawn from NMR measurements of P12 were less clear, the sample was investigated more closely by means of DLS. Already below the Tcp, ill-defined aggregates of ≈30 nm were formed. These assembled to larger aggregates at the Tcp, featuring unimodal size distributions with Dh ≈ 300 nm. At the second drop in transmittance at 54 °C, a significant shrinkage of the structures was observed, which could relate to expelling of further water molecules from the collapsed macromolecules. In fact, this corresponds to the temperature at that the mobility of the PEtOx fraction of P12 was less decreased by further heating according to the NMR studies (compare signal C in Fig. 9, top). Further heating resulted in an increase of the detected size distributions, which could represent the beginning or the formation of larger concentrated phase droplets.

A series of well-defined P(eTEGA)-b-PEtOx-b-P(eTEGA) triblock copolymers, P(eTEGA)-b-PEtOx as well as PEtOx-b-PNiPAm diblock copolymers were synthesized by RAFT polymerization using a bifunctional and monofunctional PEtOx based macro chain transfer agent, respectively. The composition of the di- and triblock copolymers covers a wide range of different block ratios. The LCST behavior of the triblock copolymers in aqueous solution is strongly affected by the hydrophobic end groups, leading to lower Tcp values of polymers with small DP of the P(eTEGA) or PNiPAm block. Furthermore, there was no difference in the observed trends between di- or triblock copolymers when substituting the PNiPAm block with a P(eTEGA) block although PNiPAm provides a hydrogen bond donor moiety that can interact with the PEtOx block. 1H NMR experiments proved that the structure of the hydrophilic PEtOx block require a certain block length in order to keep the hydrophilic segment solvated above Tcp and that the coil to globule transition takes place over a broader temperature range. In agreement with this, the formation of well-defined aggregates was observed by DLS, hinting towards a thermo-induced self-assembly of selected diblock copolymers.

Fig. 9. 1H NMR spectra (D2O, c = 5 g mL−1) of PEtOx-b-PNiPAm diblock copolymers below and above the Tcp and the respective fractions p. The spectra are normalized according to the solvent signal. Selected signals are assigned to the polymer structure. Top: PEtOx20-b-PNiPAm diblock copolymer P12. Bottom: PEtOx50-b-PNiPAm diblock copolymer P17.

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Fig. 10. Comparison of the turbidity curves and the hydrodynamic diameters Dh (number distributions) obtained by DLS measurements of the PEtOx-b-PNiPAm diblock copolymers P17 and P12 in aqueous solution at varying temperatures below and above Tcp (c = 5 g L−1). The insets show the number (red) and intensity (black) size distributions at 10 °C, 50 °C and 70 °C for P17, respectively. The according data are supplied in the supporting information for P12.

Future research will focus on a more detailed investigation of the thermo-induced aggregate formation and its exploitation for the triggered release of guest molecules. To gain access to thermo-reversible gels made from the triblock copolymers, we will rely on the utilization of more hydrophilic bifunctional CROP initiators to enable a solvation of the central PEtOx block.

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