On the Limits of Benzophenone As Cross-Linker for Surface-Attached Polymer Hydrogels

polymers

Article On the Limits of Benzophenone as Cross-Linker for Surface-Attached Polymer Hydrogels

Esther K. Riga †, Julia S. Saar †, Roman Erath, Michelle Hechenbichler and Karen Lienkamp * ID

Freiburg Center für Interactive Materials and Bioinspired Technologies (FIT) and Department of Microsystems Engineering (IMTEK), Albert-Ludwigs-Universität, Georges-Köhler-Allee 105, 79110 Freiburg, Germany; [email protected] (E.K.R.); [email protected] (J.S.S.) * Correspondence: [email protected]; Tel.: +49-761-203-95090 † These authors contributed equally to this work.

Received: 17 November 2017; Accepted: 4 December 2017; Published: 7 December 2017

Abstract: The synthesis of different photo-reactive poly(alkenyl norbornenes) and poly(oxonorbornenes) containing benzophenone (BP) via ring-opening metatheses polymerization (ROMP) is described. These polymers are UV irradiated to form well-deﬁned surface-attached polymer networks and hydrogels. The relative propensity of the polymers to cross-link is evaluated by studying their gel content and its dependency on BP content, irradiation wavelength (254 or 365 nm) and energy dose applied (up to 11 J·cm−2). Analysis of the UV spectra of the polymer networks demonstrates that the poly(oxonorbornenes) show the expected BP-induced crosslinking behavior at 365 nm, although high irradiation energy doses and BP content are needed. However, these polymers undergo chain scission at 254 nm. The poly(alkenyl norbornenes), on the other hand, do not cross-link at 365 nm, whereas moderate to good cross-linking is observed at 254 nm. UV spectra demonstrate that the cross-linking at 254 nm is due to BP cross-linking combined with a [2 + 2] cylcoaddition of the alkenyl double bonds. This indicates limitations of benzophenone as a universally applicable cross-linking for polymer networks and hydrogels.

Keywords: benzophenone cross-linking; coatings; hydrogels; photo-reactive polymers; ring-opening metathesis polymerization (ROMP); surface-attached polymer networks

1. Introduction Benzophenone (BP) is well-known for its photo-reactivity, which has been extensively reviewed [1,2]. In particular, BP forms covalent C–C bonds to molecules containing C–H bonds by UV-activated C, H–insertion reactions. More precisely, BP can absorb a photon upon UV irradiation at λ ~350 nm. This causes an electronic transition from the nonbonding n-orbital of oxygen to the antibonding π*-orbital of the carbonyl group (Figure1a). Slightly more energy (i.e., irradiation at λ ~250 nm) is needed to induce the transition of an electron from the bonding π-orbital to the antibonding π*-orbital of the carbonyl group (Figure1a). Either way, the singlet diradicaloid thus formed undergoes intersystem crossing (ISC, Figure1) into a triplet state, so that a diradical with one unpaired electron centered on oxygen and another one on the carbon of the former carbonyl group is generated. The oxygen can now interact with C–H σ-bonds of neighboring molecules (in our case polymer chains) by abstracting a hydrogen atom, thus forming an OH bond. This creates a radical on the carbon atom left behind. The new C radical can combine with the C radical on the former BP carbonyl C atom, so that a covalent C–C bond between the former BP group and the polymer chain is formed (Figure1b) [ 1,3]. The excitation of benzophenone is reversible, i.e. the diradical formed falls back into the ground state if no suitable reaction partner is found; thus its excitation can be repeated until favorable conditions (e.g., suitable reaction geometry and distance) for the reaction with a neighboring

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Polymers 2017, 9, 686 2 of 14 C–H-group are obtained [1]. As a result, efficient covalent modifications can be obtained using usingbenzophenone benzophenone [1]. Benzophenone [1]. Benzopheno wasne initiallywas initially used used in biochemistry in biochemistry (e.g., (e.g., as labeling as labeling reagent reagent for λ forphoto-affinity photo-affinity probes) probes) [4– 8[4–8],], since since the the longer long wavelengthser wavelengths needed needed for for BP BP activation activation ( ~360(λ ∼ 360 nm) nm) do donot not damage damage sensitive sensitive materials materials including including proteins proteins [9]. [9]. Prucker Prucker et al.et al. showed showed that that BP BP could could be be used used as asanchor anchor group group to functionalize to functionalize surfaces surfaces with thinwith polymer thin polymer layers [layers10]. Based [10]. onBased this principle,on this principle, polymer polymermonolayers monolayers made from made poly(vinyl from alcohol),poly(vinyl poly(styrene), alcohol), poly(acrylicpoly(styrene), acid) poly(acrylic and poly(methacrylic acid) and poly(methacrylicacid) have been attached acid) have to surfaces been attached [10,11]. to Next, surfac BP-containinges [10,11]. Next, polymers BP-containing that can self-crosslinkpolymers that upon can self-crosslinkUV irradiation upon were UV developed, irradiation so that were surface-attached developed, so polymerthat surface-attached networks and polymer hydrogels networks based on and BP hydrogelscould be obtained based on [12 BP–14 could]. Besides be obtained being a[12–14]. versatile Besides reagent being for covalent a versatile cross-linking, reagent for covalent benzophenone cross- linking,can also benzophenone be used but as acan photo-initiator also be used tobut induce as a photo-initiator cross-linking [to15 induce]. cross-linking [15].

Figure 1.1. (a(a)) Simpliﬁed Simplified Jablonski Jablonski diagram diagram of benzophenone of benzophenone illustrating illustrating its electronic its electronic states and states possible and possibletransitions transitions by internal by conversion internal (IC),conversion intersystem (IC), crossingintersystem (ISC), crossing ﬂuorescence (ISC (F)), fluorescence and phosphorescence (F) and phosphorescence(P) [16]; (b) Cartoon (P) illustration [16]; (b) ofCartoon a surface-attached illustration polymerof a surface-attached network with benzophenonepolymer network as surface with benzophenoneanchor group and as chain-chainsurface anchor cross-linker; group (cand,d) Structurechain-chain of the cross-linker; poly(alkenylnorbornenes) (c,d) StructureP(2-3-BP) of the poly(alkenylnorbornenes)and poly(oxonorbornenes) P(2-3-BP)P(4-5-BP) andused poly(oxonorbornenes) in this study. P(4-5-BP) used in this study.

In spite of of its its overall overall versatility, versatility, there there are are some some limitations limitations to to the the use use of of BP BP in in polymer polymer chemistry. chemistry. For example, Rupp et al. observed chain scission wh whenen attempting to cross-linkcross-link polyethylene oxide (PEO) in ionic liquids using benzophenone as a photo-initiatorphoto-initiator [[17].17]. Teixeira et al.al. found that PEO would undergo chain scission at low concentrations of the added BP photo-initiator, while at higher BP concentrations cross-linking be becamecame dominant [18]. [18]. Ch Christensenristensen et et al. analyzed the UV-induced gelation of acrylate copolymers containing covale covalentlyntly attached benzophenone [19]. [19]. They found that when highly abstractable hydrogen atoms were presentpresent on the main polymer chain, BP induced chain cleavage rather than cross-linking [[19].19]. In ourour work work on surface-attachedon surface-attached polymer networkspolymer andnetworks hydrogels and made hydrogels from poly(norbornenes), made from wepoly(norbornenes), came across two we othercame across BP-containing two other polymer BP-containing families polymer that failed families to that show failed the to anticipated show the anticipatedcross-linking cross-linking behavior. Thebehavior. aim of The this aim work of this was work therefore was totherefore systematically to systematically study UV-activated study UV- activatedBP cross-linking BP cross-linking of these of polymers, these polymers, and to and identify to identify structural structural parameters parameters of the of polymers the polymers that thatprevented prevented cross-linking. cross-linking. For this, we synthesizedFor this, various we BP-containingsynthesized poly(alkenylnorbornenes) various BP-containing and poly(alkenylnorbornenes)poly(oxo-norbornenes) (Figure and1 c,d,poly(oxo-norbornenes) respectively) via ring-opening (Figure 1c,d, metathesis respectively) polymerization via ring-opening (ROMP). metathesis polymerization (ROMP). These consisted of alkenylnorbornene imide and oxonorbornene imide repeat units, and contained a quite large number of heteroatoms and double bonds. The imide sites were substituted either with BP moieties, propyl groups, or N-Boc protected amine/guanidin

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These consisted of alkenylnorbornene imide and oxonorbornene imide repeat units, and contained a quite large number of heteroatoms and double bonds. The imide sites were substituted either with BP moieties, propyl groups, or N-Boc protected amine/guanidin groups. By comparing the cross-linking behavior of these polymers when UV irradiated at λ = 254 or 365 nm, we could identify polymer components that are unfavorable for polymer cross-linking using benzophenone. Thus, we could derive design rules for the next generation of benzophenone cross-linkable poly(norbornenes), and potentially also for (surface-attached) networks and hydrogels made from other polymers.

2. Experimental

2.1. Materials All chemicals were obtained as reagent grade from Sigma-Aldrich (Taufkirchen, Germany), Carl Roth (Karlsruhe, Germany), Fluka (Taufkirchen, Germany), or Alfa Aeser (Karlsruhe, Germany), and used as received. High performance liquid chromatography (HPLC) grade solvents were purchased dry from Carl Roth (Karlsruhe, Germany), and used as received. Dichloromethane (DCM) was freshly distilled over P2O5 before use.

2.2. Instrumentation Gel permeation chromatography (GPC, in chloroform, calibrated with poly(methyl methacrylate) standards) was performed on a PSS SDV column (PSS, Mainz, Germany). NMR spectra were recorded on a Bruker 250 MHz spectrometer (Bruker, Madison, WI, USA) using acetone, MeOD and CDCl3 as solvent and tetramethylsilane (TMS) as internal reference. The thickness of the dry polymer layers on silicon wafers was measured with the auto-nulling imaging ellipsometer Nanoﬁlm EP3 (Nanoﬁlm Technologie GmbH, Göttingen, Germany), which was equipped with a 532 nm solid-state laser. For each sample, the average value from three different positions was taken. The irradiation of samples with UV light was conducted using a BIO-LINK Box (Vilber Lourmat GmbH, Eberhardzell, Germany) with different wavelengths (254 and 365 nm). UV spectra were taken using a Varian Cary 50 system (Varian, Darmstadt, Germany).

2.3. Synthesis

2.3.1. Synthesis of the Benzophenone-Containing Monomer M1 The benzophenone-containing monomer (M1) shown in Scheme1 was synthesized under nitrogen using standard Schlenk techniques. Oxonorbornene (1) was reacted with the brominated amine 2 following a literature procedure to yield N-(2-bromoethyl)-3,6-tetrahydrophthalimide (3) [20]. In the second step, 4-hydroxybenzophenone (19.8 g, 100 mmol) and K2CO3 (13.8 g, 100 mmol) were dissolved in dry acetone (250 mL), and the reaction mixture was stirred at 60 ◦C. Oxonorbornene imide 3 (2.72 g, 10.0 mmol) was slowly added, and the reaction mixture was reﬂuxed for 24 h. For work up, water was added and the solution was extracted with DCM. The combined organic phases were washed with NaOH solution (10%) and dried with MgSO4. The solvent was evaporated under reduced pressure. The thus obtained product was dissolved in DCM and hexane was added slowly, resulting in two solvent layers. The vial was put in the freezer allowing a slow mixing of the solvents, resulting in the crystallization of the pure product. After removing the solvent, the isolated yield was about 38% (1.48 g). 1H NMR (250 MHz, acetone, δ): 7.44–7.89 (m, 7H, benzophenone-CH), 7.04 (d, 2H, 2 3 JH–H = 8.85, benzophenone-CH), 6.58 (s, 2H, C=CH), 5.16 (s, 2H, C=CH–CH), 4.25 (t, 2H, JH–H = 5.84, 3 13 N–CH2–CH2), 3.88 (t, 2H, JH–H = 5.80, N–CH2–CH2), 2.96 (s, 2H, CH–C=O); C NMR (63 MHz, acetone, δ): 195.25, 177.05, 163.10, 139.32, 137.47, 133.09, 132.79, 131.36, 130.33, 129.22, 115.23, 81.85, + 65.23, 48.42, 38.42, 29.92; FTMS (ESI) m/z: (M + H) calcd for C23H20O5N, 390.13; found, 390.13. Polymers 2017, 9, 686 4 of 14 Polymers 2017, 9, 686 4 of 14

Scheme 1. Synthesis of the benzophenone-containing monomer M1. The The oxonorbornene oxonorbornene anhydride anhydride 1 is reacted with the brominated amine 2 to form the oxonorbornene imide 3. The The benzophenone benzophenone group group is introduced in the secondsecond reactionreaction stepstep toto yieldyield thethe monomermonomer M1M1..

2.3.2. Synthesis of Model Polymer P(2-3-BP) 2.3.2. Synthesis of Model Polymer P(2-3-BP) To obtain P(2-3-BP), monomer M1 was copolymerized with the alkyl-functionalized norbornene To obtain P(2-3-BP), monomer M1 was copolymerized with the alkyl-functionalized norbornene monomers M2 and M3 (Scheme 2), which were synthesized following the method reported by Ilker monomers M2 and M3 (Scheme2), which were synthesized following the method reported by Ilker [ 21]. [21]. Grubbs’ third generation catalyst was synthesized as described by Love [22]. The amount of each Grubbs’ third generation catalyst was synthesized as described by Love [22]. The amount of each reagent used, and the molecular weight of the resulting polymers, as determined by GPC, can be reagent used, and the molecular weight of the resulting polymers, as determined by GPC, can be found in the supporting information (Table S1). A typical polymerization was performed under found in the supporting information (Table S1). A typical polymerization was performed under nitrogen atmosphere using standard Schlenk techniques. Monomers M2 and/or M3 were dissolved nitrogen atmosphere using standard Schlenk techniques. Monomers M2 and/or M3 were dissolved in dry DCM (5 mL) and stirred for 30 min. Grubbs’ third generation catalyst G3 was dissolved in dry in dry DCM (5 mL) and stirred for 30 min. Grubbs’ third generation catalyst G3 was dissolved in DCM (1 mg·mL−1) in a second flask and added to the monomer solution. The reaction mixture was dry DCM (1 mg·mL−1) in a second ﬂask and added to the monomer solution. The reaction mixture allowed to react for 2 h at 40 °C, and a solution of M1 in dry DCM (10 mL) was added dropwise was allowed to react for 2 h at 40 ◦C, and a solution of M1 in dry DCM (10 mL) was added dropwise simultaneously. Subsequently, an excess of ethylvinyl ether (1 mL) was added to terminate the simultaneously. Subsequently, an excess of ethylvinyl ether (1 mL) was added to terminate the reaction. After precipitation in ice-cold hexane (300 mL), the product was dried under reduced reaction. After precipitation in ice-cold hexane (300 mL), the product was dried under reduced 1 pressure. The isolated yields were between 18% and 67%. 1H NMR (250 MHz, CDCl3, δ): 6.85–7.85 pressure. The isolated yields were between 18% and 67%. H NMR (250 MHz, CDCl3, δ): 6.85–7.85 (m, 9H, benzophenone-CH), 4.85–5.70 (m, 2H, C=CH), 3.50–3.83 (m, 4H, CH2 and N–CH2), 3.00–3.40 (m, 9H, benzophenone-CH), 4.85–5.70 (m, 2H, C=CH), 3.50–3.83 (m, 4H, CH2 and N–CH2), 3.00–3.40 (m, 4H, C=C–CH and CH), 2.20–2.35 (m, 1H, M3–CH3–CH), 1.57–1.75 (m, 6H, M2–CH3), 1.41 (s, 9H, (m, 4H, C=C–CH and CH), 2.20–2.35 (m, 1H, M3–CH3–CH), 1.57–1.75 (m, 6H, M2–CH3), 1.41 (s, 9H, 13 boc-CH3), 0.79–0.90 (m, 6H, M3–CH3); 13C NMR (63 MHz, CDCl3, δ): 179.28, 177.63, 163.18, 161.45, boc-CH3), 0.79–0.90 (m, 6H, M3–CH3); C NMR (63 MHz, CDCl3, δ): 179.28, 177.63, 163.18, 161.45, 156.02, 142.51, 137.91, 137.67, 137.21, 120.59, 104.21, 97.26, 93.73, 84.31, 79.48, 79.29, 54.38, 48.90, 47.91, 156.02, 142.51, 137.91, 137.67, 137.21, 120.59, 104.21, 97.26, 93.73, 84.31, 79.48, 79.29, 54.38, 48.90, 47.91, 45.64, 38.53, 29.68, 28.34, 28.05, 26.26, 23.09, 21.57, 19.56. 45.64, 38.53, 29.68, 28.34, 28.05, 26.26, 23.09, 21.57, 19.56. For cleavage of the tert-butyl carbamate group (N-Boc protective group), the polymer made from For cleavage of the tert-butyl carbamate group (N-Boc protective group), the polymer made from 2.5% M1, 65% M2 and 32.5% M3 (60 mg, 0.85 mmol) was dissolved in trifluoroacetic acid (2 mL) and 2.5% M1, 65% M2 and 32.5% M3 (60 mg, 0.85 mmol) was dissolved in triﬂuoroacetic acid (2 mL) and allowed to react for 20 h at 45 °C. The crude product was precipitated in DCM (200 mL) and dried allowed to react for 20 h at 45 ◦C. The crude product was precipitated in DCM (200 mL) and dried under reduced pressure. The isolated yield was roughly 30% (20 mg). 1H NMR (250 MHz, MeOD-d4, under reduced pressure. The isolated yield was roughly 30% (20 mg). 1H NMR (250 MHz, MeOD-d4, δ): 6.86–7.84 (m, 9H, benzophenone-CH), 5.53–5.71 (m, 2H, C=CH), 3.67–3.90 (m, 4H, CH2 and N–CH2), δ): 6.86–7.84 (m, 9H, benzophenone-CH), 5.53–5.71 (m, 2H, C=CH), 3.67–3.90 (m, 4H, CH2 and N–CH2), 3.05–3.37 (m, 4H, C=C–CH and CH), 2.13–2.27 (m, 33% 1H, M3–CH3–CH), 1.61–1.80 3.05–3.37 (m, 4H, C=C–CH and CH), 2.13–2.27 (m, 33% 1H, M3–CH3–CH), 1.61–1.80 (m, 66% 6H, (m, 66% 6H, M2–CH3), 0.85–1.00 (m, 33% 6H, M3–CH3). M2–CH3), 0.85–1.00 (m, 33% 6H, M3–CH3).

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Scheme 2.2.Copolymerization Copolymerization of M1of withM1 (witha) norbornene (a) norbornene derivatives derivativesM2 and M3 M2, and and (b) oxonorborneneM3, and (b) imidesoxonorborneneM4 and M5imidesvia M4 ring-opening and M5 via metathesis ring-opening polymerization. metathesis po Thelymerization. monomers The were monomers initiated were with Grubbs’initiated thirdwith generationGrubbs’ third catalyst generationG3. The catalyst living chainG3. The end living was then chain quenched end was with then ethyl quenched vinyl etherwith toethyl yield vinyl the ether terpolymers to yieldP(2-3-BP) the terpolymersand P(4-5-BP) P(2-3-BP), respectively. and P(4-5-BP), respectively.

2.3.3. Synthesis of Model Polymer P(4-5-BP) To obtainobtainP(4-5-BP) P(4-5-BP),, monomer monomerM1 M1was was copolymerized copolymerized with with the the oxonorbornene oxonorbornene monomers monomersM4 andM4 M5and shownM5 shown in Scheme in Scheme2, which 2, which were synthesized were synthesized following following literature literature procedures procedures [ 23,24]. The[23,24]. amount The ofamount each reagentof each reagent used, and used, the and molecular the molecular weight weight of the resultingof the resulting polymer, polymer, as determined as determined by GPC, by canGPC, be can found be found in the in supportingthe supporting information information (Table (Table S4). S4). A A typical typical polymerization polymerization waswas performed under nitrogen atmosphere usingusing standardstandard SchlenkSchlenk techniques.techniques. MonomersMonomers M1M1,, M4M4 and/orand/or M5 were dissolved inin drydry DCMDCM (3(3 mL)mL) and and stirred stirred for for 30 30 min. min. Grubbs’ Grubbs’ third third generation generation catalyst catalyst was was dissolved dissolved in dryin dry DCM DCM (1 mg(1 mg·mL·mL−1)− in1) in a seconda second ﬂask flask and and added added to theto the monomer monomer solution. solution. The The reaction reaction mixture mixture was allowedwas allowed to react to react for 30 for min 30 min at room at room temperature. temperature. Subsequently, Subsequently, an excess an excess of ethylvinyl of ethylvinyl ether ether (0.6 mL)(0.6 wasmL) addedwas added to terminate to terminate the reaction. the reaction. After After precipitation precipitation in ice-cold in ice-cold hexane hexane (300 (300 mL), mL), the product the product was 1 1 driedwas dried under under reduced reduced pressure. pressure. The The isolated isolated yields yields were were about about 90%. 90%.H NMRH NMR (250 (250 MHz, MHz, CDCl CDCl3, δ3):, 6.89–7.85δ): 6.89–7.85 (m ,(m 9H,, 9H, benzophenone-CH), benzophenone-CH), 6.02–6.10 6.02–6.10 (m (m, 1H,, 1H, C=CH, C=CH,trans trans),), 5.70–5.83 5.70–5.83 ((mm,, 1H,1H, C=CH,C=CH, ciscis),), 4.93–5.15 (m,, 1H,1H, C=C–CH,C=C–CH, cis), 4.40–4.55 (m,, 1H, C=C–CH,C=C–CH, trans), 3.60–3.80 ((m,, 4H,4H, CHCH22 and N–CH 2),), 3.27–3.47 (m,, 2H, CH), 1.63–1.80 ((m,, 2H,2H, M5–CHM5–CH22), 1.48 ( s, 18H, boc-CH 3),), 0.81–0.90 0.81–0.90 ( (m,, 3H, 3H, M5–CH M5–CH3).).

2.3.4. Synthesis of Surface-Attached Polymer Networks Silicon substratessubstrates functionalized functionalized with with benzophenon benzophenon were were prepared prepared as follows: as follows: First, theFirst, cross-linking the cross- agentlinking triethoxy agent triethoxy benzophenone benzopheno silane (3EBP-silane)ne silane (3EBP-silane) was synthesized was sy asnthesized described as in thedescribed literature in [25the]. −1 Aliterature solution [25]. of 3EBP-silaneA solution of (40 3EBP-silane mmol·L (40in mmol·L toluene)−1 in was toluene) then spin was coatedthen sp onin coated a standard on a standard Si wafer −1 ◦ (800Si wafer rpm, (800 120 rpm, s and 120 500 s and rpm 500 s rpm). Subsequently, s−1). Subsequently, the wafer the wafer was was cured cured for for 30 min30 min at 100at 100C °C on on a preheateda preheated hot hot plate, plate, washed washed with with organic organic solvents solvents (first (first with with toluene toluene to remove to remove unbound unbound3EBP -silane,3EBP- thensilane, ethanol then ethanol for removal for removal of the polarof the leavingpolar leavin groupsg groups and easier and easier surface surfac drying).e drying). The surfaces The surfaces were thenwere driedthen dried under under a continuous a continuous flow offlow nitrogen of nitrogen gas. gas. To synthesize surface-attach surface-attacheded polymer polymer networks networks from from P(2-3-PB)P(2-3-PB) or orP(4-5-BP)P(4-5-BP), a ,solution a solution of the of − therespective respective polymer polymer in chloroform in chloroform (20 mg·mL (20 mg−·1mL) was1 )spin was cast spin on cast a 3-EBP on a 3-EBP treated treated silicon silicon wafer wafer(3000 − (3000rpm, 30 rpm, s and 30 s1000 and rpm·s 1000− rpm1). The·s 1films). The were films cross-linked were cross-linked at 254 or at at 254 365 or nm at 365with nm different with different energy energydoses, washed doses, washed with DCM with to DCM remove to remove unattached unattached polymer polymer chains, chains, and dried and dried in a nitrogen in a nitrogen stream. stream. For Fordeprotection, deprotection, the thesamples samples were were depo depositedsited in in4 molar 4 molar hydrochloric hydrochloric acid acid in in dioxane dioxane for for 12 12 h, h, washed with ethanol and dried in a nitrogen stream.stream.

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3. Results and Discussion The benzophenone-containing poly(alkenylnorbornenes) P(2-3-BP) (Figure1c) and poly(oxonorbornenes) P(4-5-BP) (Figure1d) used in this study were obtained by copolymerizing the benzophenone-containing monomer M1 (Scheme1) with the respective functional monomers M2 to M5 (Scheme2). To obtain M1, first oxonorbornene anhydride 1 was reacted with the brominated amine 2 to give N-(2-bromoethyl)-3,6-tetrahydrophthalimide (3, Scheme1). The latter was then reacted with hydroxybenzophenone and K2CO3 in dry acetone for 24 h under reflux, yielding monomer M1 (Scheme1). Details are given in the Experimental. To synthesize different variants of the target poly(alkenylnorbornene) P(2-3-BP), monomer M1 and the alkenylnorbornene derivatives M2 and M3 were mixed in various monomer feed ratios and polymerized via ring opening metathesis polymerization (ROMP) using the Grubbs’ third generation catalyst (G3) (Scheme2a). The catalyst-to-monomer ratio was adjusted to obtain copolymers with the desired molecular weight (target molecular weight 100,000 g·mol−1). The target content of BP repeat units was 2.5% to 5%. Since the polymerization rates of the monomers used varied significantly (M2 and M3 being much less reactive than the oxonorbornene imide M1)[26,27], the BP-containing monomer M1 was added dropwise over 120 min to the reaction mixture containing M2, M3 and the polymerization catalyst. That way, a more homogeneous distribution of the BP-monomer units along the polymer chain was obtained. The molar composition of the polymers thus obtained, and their molar mass distribution parameters (determined by gel permeation chromatography) are summarized in Table1. It was confirmed by 1H NMR spectroscopy via a comparison of the integrated peaks of the benzophenone group (6.85 to 7.85 ppm, 9H) and the N-Boc protective group (1.41 ppm, 9H) that the actual BP content of the polymers matched the target content. To obtain different variants of the polymers P(4-5-BP), monomer M1 was copolymerized with different amounts of the oxonorbornene derivatives M4 and M5 (Scheme2b). These three monomers had similar polymerizable groups and therefore similar polymerization rates, and were thus mixed and added at the beginning of the reaction. The target BP content was 5 to 15 mol %. As before, the comonomer feed ratio and catalyst-to-monomer ratio were adjusted to obtain the desired copolymer molecular weights and compositions (Table1). For some of the network formation studies presented below, the N-tert-butyloxycarbonyl (N-Boc) protective groups of the polymers were cleaved with trifluoroacetic acid before cross-linking. Unless otherwise specified, the reactions and measurements were performed on the protected polymers.

Table 1. Molar compositions, molecular weights (Mn and Mw) and polydispersity indices ÐM of the poly(alkenylnorbornenes) P(2-3-BP) and the poly(oxonorbornenes) P(4-5-BP). Mn, Mw and ÐM were determined by gel permeation chromatography in chloroform using PMMA standards.

Molar composition/mol % Molecular weight/kg·mol−1 Polymer ÐM M1 M2 M3 M4 M5 Mn,exp Mw,exp

P(490-BP10) 10 - - 90 - 83 147 1.76 P(485-BP15) 15 - - 85 - 82 156 1.91 P(295-BP5) 5 95 - - - 54 67 1.23 P(395-BP5) 5 - 95 - - 50 72 1.46 P(495-BP5) 5 - - 95 - 81 114 1.41 P(495-BP5) 5 - - 95 - 81 153 1.87 P(495-BP5) 5 - - 95 - 82 384 4.69 P(263-332-BP5) 5 63 32 - - 45 51 1.14 P(485-510-BP5) 5 - - 85 10 84 139 1.65 P(485-510-BP5) 5 - - 85 10 88 480 5.48 P(265-332.5-BP2.5) 2.5 65 32.5 - - 53 71 1.33 P(266.7-333.3) - 66.7 33.3 - - 24 59 2.50

To synthesize surface-attached polymer networks from the P(4-5-BP) polymers, they were dissolved in dichloromethane (DCM) and spin-coated onto silicon wafers that had been previously Polymers 2017, 9, 686 7 of 14 functionalized with benzophenone using the anchor group triethoxy benzophenone silane (3EBP)[25]. 3EBP contains a benzophenone moiety and a silane group, which links it covalently to the silicon surface. Upon UV irradiation, the BP moiety of the 3EBP anchor group formed a covalent bond to P(4-5-BP) and thus attached it to the surface, while the BP moieties that were part of P(4-5-BP) simultaneously formed cross-links between the polymer chains, so that overall a surface-attached polymer network was obtained (Figure1b). The degree of cross-linking of a network or hydrogel can be quantified by determining its gel content. The gel content of the here presented polymer films was determined by first measuring the overall dry film thickness d0 of the networks just after UV irradiation via ellipsometry (i.e., the thickness of the gel plus the sol). The networks were then extracted in DCM overnight to remove unbound polymer chains. After this treatment, the remaining dry layer thickness dextract (i.e., the gel thickness) was also measured via ellipsometry. From this, the gel content of each layer was calculated using Equation (1). d gel content = ( extract × 100) % (1) d0 When different polymers are cross-linked using comparable conditions (e.g., the same irradiation doses and wavelengths), the gel content of the networks or hydrogels formed can be considered as a quantitative measure for the relative propensity of each polymer to cross-link. With this in mind, the gel content of networks made from different members of the P(2-3-BP) and the P(4-5-BP) families was determined after UV irradiation at different wavelengths (λ = 254 or 365 nm, respectively) and different energy doses. In Figure2, the gel content of the P(4-5-BP) polymer networks thus obtained is shown. (For these polymers, the subscripts indicate the relative amount of each repeat unit, i.e., P(485-510-BP5) refers to a polymer made from 85% M4, 10% M5, and 5% M1, and P(495-BP5) to a polymer made from 95% M4, 0% M5, and 5% M1.) The thickness data onto which the gel content curves are based are listed in the Supporting Information (Tables S3–S5). Figure2a shows the gel content of P(4-BP) networks with 5–15 mol % of BP for various irradiation energy doses at λ = 365 nm. In this graph, the gel content first increases steeply to the optimum dose, and then levels off into a plateau at higher energy doses. This is the usual behavior described for BP cross-linked polymers in the literature [15], even though the energy doses needed to obtain a substantial amount of cross-linking in this polymer were quite high. Also in line with theory [12], the data shows that layers made from polymers with a higher BP content also had a higher gel content −2 when the same irradiation energy dose was used. For example, P(485-BP15) irradiated with 2 J·cm had a gel content of around 90%, while the gel content of P(490-BP10) was only 74%, and that of P(495-BP5) was only 24%. It is noteworthy, though, that surprisingly high BP contents and energy doses were needed to obtain significant amounts of cross-linking. When the P(4-5-BP) polymer networks were treated with acid to remove their tert-butyloxycarbonyl (Boc) protective groups (to obtain the corresponding polymer hydrogels), a layer thickness decrease −2 of 46% to 63% was observed. For example, the layer thickness of P(495-BP5) (energy dose 10 J·cm ) decreased from 229 to 113 nm after deprotection. This was mostly caused by the high mass loss occurring when the Boc groups were removed (42 mass %). At the same time, though, some polymer chains that are cross-linked predominantly via bonds to these Boc groups might also become detached, which would further increase the mass loss. To investigate if such chains formed was a significant fraction of the polymer networks obtained, the gel content of networks made from protected and the deprotected polymers P(495-BP5) (irradiated with different energy doses at λ = 365 nm) were determined and compared to the gel contents of the corresponding P(485-510-BP5) polymers. P(485-510-BP5) had the same benzophenone content as P(495-BP5), but 10 mol % repeat units containing propyl substituents, and thus more C–H bonds available for cross-linking that would not be cleaved by deprotection (Figure1d). When irradiated at 365 nm, P(485-510-BP5) consistently had higher gel contents than P(495-BP5) throughout the energy dose range investigated (Figure2b), even though it had a lower absolute number of C–H bonds compared to P(495-BP5). For example, P(485-510-BP5) had a gel Polymers 2017, 9, 686 8 of 14

−2 contentPolymers of 2017 around, 9, 686 81% after illumination with 8 J·cm , whereas that of P(495-BP5) was only 76% when8 of 14 irradiated with the same energy dose. This indicates that the propyl C–H groups per se were more easily availablemore easily for cross-linking available for than cross-linking the C–H bonds than the of theC–H two bonds CH2 ofCH the2N-Boc two CH fragments.2CH2N-Boc The fragments. same trend The wassame observed trend forwas the observed corresponding for the corresponding deprotected polymers deprotected of P(4 85polymers-510-BP5 of) and P(4P(485-59510-BP-BP55)) and(Figure P(4952b).-BP5) In(Figure Figure2 d,2b). the In relativeFigure 2d, thickness the relative loss duethickness to deprotection loss due to (in deprotection %) of the two (in polymers%) of the two is compared. polymers is Whencompared. all Boc groupsWhen areall cleaved,Boc groups the theoreticalare cleaved, mass the loss theoretical of deprotected mass P(4loss95 -BPof deprotected5) should be 42P(4 mass95-BP5) %,should corresponding be 42 mass to a %, thickness corresponding reduction to of a roughlythicknes thes reduction same percentage, of roughly while the same that of percentage,P(485-510-BP while5) shouldthat of be P(4 only85-5 slightly10-BP5) should lower (40be only mass slightly %). Figure lower2d (40 shows mass that %). the Figure mass 2d loss shows for the that deprotected the mass loss P(4for95-BP the5 )deprotectedpolymers was P(4 considerably95-BP5) polymers larger was than considerably for the P(4 85larger-510-BP than5) polymers,for the P(4 indicating85-510-BP5) thatpolymers, the formerindicating contained that athe larger former fraction contained of polymer a larger chains fracti thaton wereof polymer exclusively chains cross-linked that were viaexclusively bonds to cross- the Boclinked groups. viaIn bonds line withto the this Boc interpretation, groups. In line the with data this also interpretation, shows that for the both data polymers, also shows the that thickness for both decreasepolymers, becomes the thickness smaller with decrease increasing becomes irradiation smaller energy, with increasing i.e., in the stronglyirradiation irradiated energy, polymeri.e., in the networksstrongly (and irradiated the corresponding polymer networks deprotected (and the hydrogels), corresponding more cross-linksdeprotected to hydrogels), polymer parts more other cross- thanlinks the to Boc polymer group couldparts other be formed. than the Boc group could be formed.

FigureFigure 2. 2.Gel Gel content content of ofsurface-attached surface-attached polymerpolymer ne networkstworks and and hydrogels hydrogels made made from from different different P(4- P(4-5-BP)5-BP) polymerspolymers as as a a function function of of the the irradiation energy dose: ( a)) Polymers with varying amountamount of ofbenzophenone repeat repeat units units (5–15 (5–15 mol mol %, %, respectively), respectively), irradiated irradiated at at λλ == 365 365 nm nm (Table (Table S4); S4); (b) (b)polymers polymers with with 5% 5% benzophenone benzophenone repeat repeat units units and and varying varying amounts amounts of co-repeat of co-repeat units, units,black symbols black symbols= N-Boc = N protected-Boc protected polymers, polymers, grey symbols grey symbols = deprotected = deprotected polymers, polymers, irradiated irradiated at λ at= 365λ = nm 365 (Table nm (TableS5); S5);(c) Comparison (c) Comparison of cross-linking of cross-linking of ofthe the protected protected P(4P(485-58510-5-BP10-BP5) and5) and P(4P(495-BP955-BP) at5 λ) at= 365λ = 365nm nm(filled (ﬁlledsymbols, symbols, Table Table S5) S5) and and λ λ= =254 254 nm nm (open (open symbols, TableTable S3);S3);( d(d)) Thickness Thickness reduction reduction due due to to deprotectiondeprotection of theof theN-Boc N-Boc protected protected polymeric polymeric layers layers consisting consisting of P(4of P(485-58510-5-BP10-BP5)5and) andP(4 P(49595-BP-BP5)5)as as a a functionfunction of of the the irradiation irradiation energy energy dose dose (Table (Table S5). S5).P(495 P(4-BP955-BP) =5()M1 = /(M1M4//M4M5/M55:95:0), 5:95:0),P(4 90P(4-BP90-BP10) 10=) = (M1(M1/M4/M4/M5/M510:90:0), 10:90:0),P(4 P(48585-BP-BP1515))( (M1M1//M4/M5M5 15:85:0))15:85:0)) ( (P(4P(48585-5-51010-BP-BP5) 5()M1(M1/M4/M4/M5/M5 5:85:10)5:85:10) and and P(495- P(4BP95-BP5) (M15) (M1/M4//M4M5/ 5:95:0).M5 5:95:0).

To look more closely into the aspect of the surprisingly high-energy doses and cross-linker contents needed for substantial cross-linking of the P(4-5-BP) polymers, P(485-510-BP5) and P(495-BP5) were both irradiated with higher energy photons (λ = 254 nm) and different energy doses (Figure 2c).

Polymers 2017, 9, 686 9 of 14

To look more closely into the aspect of the surprisingly high-energy doses and cross-linker Polymerscontents 2017 needed, 9, 686 for substantial cross-linking of the P(4-5-BP) polymers, P(485-510-BP5) and P(4959-BP of 145) were both irradiated with higher energy photons (λ = 254 nm) and different energy doses (Figure2c). UnderUnder these these conditions, conditions, a a significant significant amount amount of of cros cross-linkings-linking was was achieved achieved even even with with an an energy energy dose dose −2 2 asas low low as as 0.5 J·cm J·cm (Figure(Figure 2c).2c). For For example, example, polymer P(4P(49595-BP-BP5)5 )had had a alayer layer thickness thickness of of 211 211 nm nm and and aa gel gel content of 75% when irradiatedirradiated withwith 0.50.5 JJ·cm·cm−−22. .However, However, when when this this polymer polymer was was irradiated irradiated withwith higher higher doses (up to 6 JJ·cm·cm−2),2), the the layer layer thickness thickness decreased decreased to to 20 20 nm nm and and the the gel gel content content to to only only 8%.8%. This This effect effect was was also observed for thethe copolymercopolymer P(4P(48585-510-BP-BP55) )(Figure(Figure 22c).c). This indicatesindicates thatthat higherhigher energy energy photons photons could, could, in in principle, principle, more more efficiently efficiently cross-link cross-link these these polymers. polymers. However, However, when when tootoo many many photons photons of of that that energy energy were were provided, provided, un unfavorablefavorable side side reactions reactions such such as as polymer polymer main main chainchain scissions, leadingleading to to lower lower gel gel content, content, become become dominant. dominant. To confirm To confirm this, UV this, absorption UV absorption spectra spectraof P(495 of-BP P(45) films95-BP5 that) films had that been had irradiated been irradiated at 254 nm at with254 nm different with energydifferent doses energy were doses measured were measured(Figure3a). (Figure 3a).

−2 FigureFigure 3. 3. UVUV absorption absorption spectra spectra of of P(4P(49595-BP-BP5) 5films) ﬁlms irradiated irradiated at ( ata) (λa) =λ 254= 254 nm nm with with 0 to 0 1.5 to J·cm 1.5 J·−cm2 and − (andb) λ( b=) 365λ = 365nm nmwith with 0 to 0 to50 50J·cm J·cm−2. The2. The insets insets show show the the reduction reduction of of the the adsorption adsorption band band of of the the

benzophenonebenzophenone moieties moieties as as a a func functiontion of of the the irradiation irradiation energy. energy.

TheseThese spectra spectra showed showed that that the the UV UV absorption absorption band band of of the the BP BP group (at ~280 nm) nm) decreased decreased continuouslycontinuously when when irradiated irradiated with with energy energy doses doses from from 0 0 to 0.5 JJ·cm·cm−−2 2atat λλ == 254 254 nm nm (Figure (Figure 3a),3a), i.e., thethe BP BP groups groups were were quantitatively quantitatively consumed consumed in in th thatat energy energy range. range. When When the the samples samples were were exposed exposed toto higher higher energy energy doses doses at at that that wavelength, wavelength, the the whole whole shape shape of of the the curve curve started started to to alter, alter, particularly particularly thethe peaks peaks at at 200 200 and and 235 235 nm nm decreased decreased significantly. significantly. This This confirms confirms that that other other reactions reactions besides besides BP BP activationactivation also also occurred occurred (Figure (Figure 2c).2c). It Itcan can be be sp speculatedeculated that that when when the the BP BPgroups groups were were used used up, up,the 254the nm 254 photons nm photons could could interact interact with withother otherfunctional functional groups, groups, which which could couldhave led have to cleavage led to cleavage of the 95 5 polymerof the polymer backbone. backbone. When the When P(4 the-BPP(4) films95-BP were5) films irradiated were irradiated with less withenergetic less energeticphotons ( photonsλ = 365 nm),(λ = 365the nm),UV absorption the UV absorption spectra spectradid not didshow not a show decre aase decrease in the inregion the region of 200 of to 200 ~260 to ~260nm, nm,even even for energyfor energy doses doses as high as high as 50 as J·cm 50 J−·2cm (Figure−2 (Figure 3b). Instead,3b). Instead, a steady a steady decrease decrease of the of UV the absorption UV absorption band ofband the ofBP the group BP groupat 280 atnm 280 with nm increasing with increasing energy energy dose was dose observed was observed (Figure (Figure 3b). Thus,3b). Thus,these lower these energylower energy photons photons had a hadlower a lower probability probability to trigger to trigger cross-linking cross-linking via via the the BP BP group, group, and and it it therefore therefore tooktook higher higher energy energy doses doses to to obtain obtain similarly similarly high high ge gell contents contents as as for for the the 254 254 nm nm photons. photons. On On the the other other hand,hand, the the 365 365 nm nm photons photons did did not not UV UV-activate-activate the the other other functional functional grou groupsps in in the the polymer, polymer, so so that that the the desireddesired polymer polymer networks networks were were obtained. AA similar similar analysis analysis was was performed performed with with the the P(2-3-BP)P(2-3-BP) polymerspolymers (Figure (Figure 1c).1 Polymerc). Polymer films filmswere alsowere obtained also obtained by spin-coating by spin-coating the therespective respective polymers polymers from from solution solution onto onto 3EBP3EBP-functionalized-functionalized substratessubstrates as described above forfor thethe P(4-5-BP)P(4-5-BP)samples. samples.Again, Again, the the polymer polymer films films were were cross-linked cross-linked at atwavelengths wavelengths of of 254 254 or or 365 365 nm nm with with defined defined energy energy doses.doses. TheyThey werewere extracted in dichloromethane overnightovernight to to remove remove unattached unattached polymer polymer chains, chains, an andd their their gel gel content content was was determined. determined. The The thickness thickness data obtained by ellipsometry for the different surface-attached P(2-3-BP) networks and hydrogels are listed in the supporting information (Tables S6 and S7) and summarized in Figure 4a. (As before, the subscripts of these polymers indicate the relative amount of each repeat unit in the polymer, i.e., P(265-332.5-BP2.5) is a polymer made from 65% M2, 32.5% M3, and 2.5% M1).

Polymers 2017, 9, 686 10 of 14 data obtained by ellipsometry for the different surface-attached P(2-3-BP) networks and hydrogels are listed in the supporting information (Tables S6 and S7) and summarized in Figure4a. (As before, the subscripts of these polymers indicate the relative amount of each repeat unit in the polymer, i.e., P(2Polymers65-3 32.52017-BP, 9, 6862.5) is a polymer made from 65% M2, 32.5% M3, and 2.5% M1). 11 of 14

Figure 4. (a(a) )Gel Gel content content of of polymer polymer networks networks co consistingnsisting of ofdifferent different versions versions of ofP(2-3-BP)P(2-3-BP) asas a afunction function of the of theirradiatio irradiationn energy, energy, irradiated irradiated at λ = at254λ nm= 254(open nm symbols, (open Table symbols, S7) and Table λ = S7) 365 andnm λ(filled= 365 symbols, nm (filled Table symbols, S6), P(3 Table95-BP S6),5) = P(3(M195/M2-BP/5M3) = 5:0:95), (M1/M2 P(2/M395-BP5:0:95),5) = (M1P(2/M295-BP/M35) 5:95:0),= (M1/ P(2M263/-3M332- 5:95:0),BP5) = (M1P(2/63M2-3/32M3-BP 5:63:32),5) = (M1 P(2/M265-3/32.5M3-BP5:63:32),2.5) = (M1P(2/M265/-3M332.5 2.5:65:32.5)-BP2.5) = (andM1/ P(2M266.7/M3-333.32.5:65:32.5)) = (M1/M2/ andM3 P(20:66.7:33.3).66.7-333.3 )(=b– (eM1) UV/M2 absorption/M3 0:66.7:33.3). spectra (ofb– P(2e) UV66.7-3 absorption33.3) films irradiated spectra of atP(2 (b66.7) 254-333.3 nm) filmsand ( irradiatedc) 365 nm atand (b )of 254 P(2 nm65-3 and32.5-BP (c)2.5 365) films nm andirradiated of P(2 65at-3 32.5(d) -BP2542.5 nm) films and irradiated (e) 365 nm. at ( dThe) 254 measurements nm and (e) 365 were nm. Theconducted measurements after irradiation were conducted with 0–9after J·cm− irradiation2. with 0–9 J·cm−2.

InWhen contrast the Boc to theprotectiveP(4-5-BP) groupspolymers, of the P(2-3-BP) no networks polymers formed were when removed different withP(2-3-BP) hydrochloricpolymers acid wereto obtain irradiated surface-attached at λ = 365 hydrogels, nm, regardless this treatm of theent energy near-quantitatively dose used (up todestroyed 9 J·cm− 2the) or network the amount (see ofTable BP S7 units of the in thesupporting polymer information). chain (Figure For4a andexample, Table the S6 inpolymer the Supporting network consisting Information). of P(3 On95-BP the5) −2 otherhad a hand,layer whenthickness the sameof 49 experimentsnm after irradiation were repeated with 9 with J·cm higher and extraction, energy photons which at decreasedλ = 254 nm, after gel deprotection to a thickness of only 5 nm. This suggested that in the P(2-3-BP) networks, most of the contents of up to 47% were reached (Figure4a). Networks made from polymer P(263-332-BP5) with cross-links formed via the C–H groups in the Boc groups. To confirm this, polymer P(265-332.5-BP2.5) 5% BP repeat units had a gel content that was similar to that of networks made from P(265-332.5-BP2.5) withwas first 2.5% deprotected BP repeat units,and then i.e., spin the coated number on of a BP3EBP units-functionalized did not signiﬁcantly wafer. After affect illumination the gel content. at λ = 245 nm with 9 J·cm−2, a surface-attached hydrogel with a gel content of 60% was formed, while the Interestingly, polymer P(266.7-333.3) with no benzophenone groups self-crosslinked to a signiﬁcant polymer coating made from the protected version of this polymer, treated under the same conditions, yielded only a gel content of 36%. This indicated that the Boc protective groups sterically shielded the other C–H bonds of the polymer, so that BP cross-linking mainly occurred with the C–H bonds in these protective group. In general, the ability of a polymer to effectively attach to a substrate by cross-linking depends on the exact circumstances of the network or hydrogel formation process. Körner et al. used a simple percolation model to describe the formation of polymer networks as a function of the reaction

Polymers 2017, 9, 686 11 of 14 extent (19% gel content when irradiated with 9 J·cm−2) at 254 nm, but not when irradiated at the same dose at 365 nm (Figure4a). To understand this, the network formation of P(266.7-333.3) was also studied by recording UV spectra of films that had been irradiated at 254 nm (Figure4b) and 365 nm, respectively (Figure4c), and by comparing them to UV spectra of P(265-332.5-BP2.5) that had been likewise treated (Figure4d,e). The spectra of P(266.7-333.3) showed that the UV absorption bands at ~200 nm decreased from 0 to 9 J·cm−2 when the sample was irradiated at 254 nm (Figure4b) and that the whole shape of the curve started to alter. Since at the same time a network formation was observed (Figure4a, Table S7), this indicates a structural change of the polymer, which led to cross-linking. UV absorption at wavelengths around 200 nm are characteristic for an electron transition from the π-orbital to an antibonding π*-orbital of a C=C bond [28]. Since there are many double bonds in the P(2-3) polymer backbone, cross-linking can apparently occur also via those groups, presumably via a [2 + 2] cycloaddition involving the alkenyl group (which is not present in the corresponding poly(oxonorbornenes) that undergo chain cleavage when irradiated at 254 nm). When the same investigation was repeated with P(266.7-333.3)-films irradiated at λ = 365 nm, the UV absorption spectra did not show any decrease of the peak at 200 nm (Figure4c), which matches the finding that no network formation was observed (Figure4a). This verifies the assumption that cross-linking occurred also via a reaction involving the alkenyl double bonds, for which the activation energy is provided by photons at λ = 254 nm, while irradiation at 365 nm is insufficient. The same set of experiments was performed with P(265-332.5-BP2.5) films, which had a similar ratio of functional repeat units as P(266.7-333.3), but additionally a small amount of BP groups. When irradiated at 365 nm, the UV absorption spectra of P(265-332.5-BP2.5) only show a slight decrease of the peak at 200 nm, but a steady decrease of the UV absorption band of BP at ~280 nm with increasing energy dose (Figure4e). This indicates that the energy at λ = 365 nm was enough for the activation of the BP groups. However, thickness measurements showed that no network formed. When the P(265-332.5-BP2.5) films were irradiated at 254 nm, the UV absorption band of the BP group (~280 nm) vanished already at 0.5 J·cm−2 (Figure4d). Again, when the sample was irradiated with higher energy doses, the whole shape of the curve started to alter. Those results are consistent with the ones of the pure P(266.7-333.3)-films irradiated at 254 nm, only the gel content and thus the degree of cross-linking was higher for P(265-332.5-BP2.5). Thus, for the poly(alkenylnorbornenes) here investigated, network formation at λ = 254 nm occurs via a synergistic effect of BP-induced cross-linking and [2 + 2] cycloaddition. This finding also explains why P(395-BP5) cross-linked more efficiently than P(295-BP5) (Figure4a): the fully substituted double bond of P(295-BP5) is sterically more hindered than the only triply substituted one of P(395-BP5), and thus P(295-BP5) has a lower propensity to undergo a [2 + 2] cycloaddition. When the Boc protective groups of the P(2-3-BP) polymers were removed with hydrochloric acid to obtain surface-attached hydrogels, this treatment near-quantitatively destroyed the network (see Table S7 of the supporting information). For example, the polymer network consisting of P(395-BP5) had a layer thickness of 49 nm after irradiation with 9 J·cm−2 and extraction, which decreased after deprotection to a thickness of only 5 nm. This suggested that in the P(2-3-BP) networks, most of the cross-links formed via the C–H groups in the Boc groups. To confirm this, polymer P(265-332.5-BP2.5) was first deprotected and then spin coated on a 3EBP-functionalized wafer. After illumination at λ = 245 nm with 9 J·cm−2, a surface-attached hydrogel with a gel content of 60% was formed, while the polymer coating made from the protected version of this polymer, treated under the same conditions, yielded only a gel content of 36%. This indicated that the Boc protective groups sterically shielded the other C–H bonds of the polymer, so that BP cross-linking mainly occurred with the C–H bonds in these protective group. In general, the ability of a polymer to effectively attach to a substrate by cross-linking depends on the exact circumstances of the network or hydrogel formation process. Körner et al. used a simple percolation model to describe the formation of polymer networks as a function of the reaction conditions and the reaction time [29]. In this model, polymers first form gel clusters inside the polymer layer, and a network is obtained once these clusters percolate through the entire polymer film. For UV Polymers 2017, 9, 686 12 of 14 irradiation of polymer films from above (e.g., using poly(styrene) containing additional BP repeat units, cross-linked at λ = 365 nm), it was observed that there was a long induction period before any macroscopic network formation was detected, as the formation of the clusters started at the air-polymer interface, and slowly percolated down to the polymer-substrate interface. In this particular case, a significant amount of network formation occurred only at an energy dose of >10 J·cm−2 (corresponding to 100 min irradiation time at 2.2 mW·cm−2)[29]. Thus, it is possible that for polymers of the P(2-3-BP) family, such a gel cluster formation is also very slow, so that no network percolation is observed at 365 nm in the energy range investigated. When irradiated with 254 nm, however, these polymers formed surface-attached polymer networks, albeit with a low gel content, and with synergistic BP cross-linking and double-bond-induced cross-linking. The poly(oxonorbornene) family P(4-5-BP), on the other hand, degraded when irradiated at 254 nm at energy doses higher than 0.5 J·cm−2. Unlike P(2-3-BP), those polymers did not have the additional alkenyl substituent on the backbone. This double bond, however, seems to be a protection against main chain cleavage at λ = 254 nm, since it can absorb the high-energy photons via a π → π* transition. Additionally, this double bond assists network formation by its ability to self-crosslink at that wavelength. At 365 nm, on the other hand, the cross-linking behavior of the P(4-5-BP) copolymers was textbook-like, and with higher energy doses higher gel contents were reached. The finding that the P(4-5-BP) polymers undergo chain scission by photons having a wavelength of 254 nm is in line with the results of Christensen et al., who previously observed that highly abstractable hydrogens especially on the main chain tend to cause chain scission [19]. Another factor affecting the reactivity of a certain C–H bond in C–H insertion reactions, besides their electronic activation, is their steric availability. C–H abstraction can be sterically hindered by flexible, large side groups that can shield the C–H bonds near the polymer backbone [15]. For the polymers used in this study, the C–H bonds in the Boc group and the C–H bonds in the alkyl or alkenyl substituents were the sterically most easily accessible ones. In the norbornene polymers of the P(2-3-BP) family, the Boc groups effectively shielded the C–H bonds near the backbone, so that cross-linking via the BP units mainly occurred via the Boc group. As a result, these networks disintegrated when the Boc groups were cleaved off. Only when the Boc groups were removed before the network formation, the C–H groups nearer to the backbone were significantly involved in the cross-linking process. In case of the poly(oxanorbornenes) P(4-5-BP) family, on the other hand, BP could abstract hydrogen atoms from either the Boc groups or from the backbone. While the C–H bonds on the Boc group had less steric hindrance, the C–H bonds next to the oxygen and nitrogen atoms could be readily abstracted due to their lower bond energies, despite the higher steric hindrance.

4. Conclusions In this work, the formation of surface-attached polymer networks and hydrogels using several poly(oxonorbornenes) and poly(alkenylnorbornenes) with benzophenone-containing repeat units was investigated in detail. The polymers were spin-coated onto benzophenone-functionalized silicon wafers and irradiated at a wavelength of 254 or 365 nm, respectively, with different energy doses. The gel content of the resulting polymer networks and hydrogels was determined by ellipsometry, and the functional groups involved in the cross-linking process were analyzed using UV spectroscopy. The results show that the cross-linking of these polymers depended strongly on the radiation wavelength and the energy does used. This could be related to subtle changes of the repeat unit structures and to steric as well as electronic effects. Polymers belonging to the P(2-3-BP) family showed no network formation when irradiated at 365 nm. Irradiation with 254 nm, on the other hand, resulted in network formation due to synergistic effects between double bond cross-linking involving the alkenyl substituent on the ﬁve-membered ring of the polymer backbone on the one hand, and BP-induced cross-linking involving the Boc groups on the other hand. However, these networks were destroyed upon removal of the Boc group, as most network cross-links due to BP involved the Boc group. In case of the deprotected P(2-3-BP) polymers, Polymers 2017, 9, 686 13 of 14 on the other hand, hydrogels could also be formed via the other alkyl C–H bonds, which ceased to be sterically shielded after removal of the Boc groups. Thus, the presence of Boc groups leads to readily degradable polymer networks if such a feature is desired; if not, however, polymers should be deprotected prior to network formation by UV irradiation to reduce mass loss by deprotection. Apparently, the presence of the additional alkenyl substituent in the P(2-3-BP) polymers also protected them from main chain scission when irradiated at 254 nm. The poly(oxonorbornenes) of the P(4-5-BP) family had textbook-like cross-linking behavior when irradiated at λ = 365 nm, however they were degraded at λ = 254 nm when the irradiation energy dose was higher than 0.5 J·cm−2. It is likely that the large number of heteroatoms in these polymers, and thus the large number of C–H bonds with lower dissociation energy, led to side reactions ultimately causing chain scission (analogous to the situation in PEO). It appears that the oxonorbornene backbone is thus not suitable for these reaction conditions. This could be investigated in future studies by comparing poly(norbornenes) and structurally identical poly(oxonorbornenes). The design rules thus obtained (avoidance of heteroatoms, beneﬁcial effect of main-chain alkenyl substituents, avoidance of bulky, C–H rich protective groups) will help to pre-select interesting structures for the next generation of BP cross-linkable poly(norbornenes) intended for surface-attached polymer networks and hydrogels, or other kinds of UV-activated reactions.

Supplementary Materials: The following are available online at www.mdpi.com/2073-4360/9/12/686/s1. Acknowledgments: Funding of this work by the European Research Council (ERC Starting Grant REGENERATE) is gratefully acknowledged. Jürgen Rühe is gratefully acknowledged for very helpful discussions and sharing of many analytical instruments. Author Contributions: Esther K. Riga, Julia S. Saar and Karen Lienkamp conceived and designed the experiments; Esther K. Riga, Julia S. Saar and Michelle Hechenbichler performed the experiments; Esther K. Riga, Julia S. Saar and Karen Lienkamp analyzed the data; Roman Erath contributed reagents/materials/analysis tools; Esther K. Riga, Julia S. Saar and Karen Lienkamp wrote the paper. Conﬂicts of Interest: The authors declare no conﬂict of interest.

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