Supporting Information DIELS-ALDER CLICKABLE PLYMER BRUSHES: A VERSATILE CATALYST-FREE CJUGATI PLATFRM Yasemin ursel Yuksekdag, Tugce ihal Gevrek, Amitav Sanyal* Department of Chemistry, Bogazici University, Bebek, Istanbul 34342, Turkey *Corresponding author: Email address: amitav.sanyal@boun.edu.tr. Experimental Section Materials Furfuryl methacrylate (FuMA, 97%),,,,, -pentamethyldiethylenetriamine (PMDETA), Cu(I)Br, poly(ethylene glycol) methyl ether methacrylate (PEGMEMA, M n : 300 g/mol), 2,2 -bipyridine were purchased from Sigma-Aldrich. Cu(I)Cl (99%) was obtained from Acros rganics. The monomers FuMA and PEGMEMA were passed through aluminum oxide to remove inhibitors. BDIPY- 3 was synthesized according to literature. 1 rganic solvents were obtained from Merck and used as received. Qdot 605 streptavidin conjugate was purchased from Invitrogen molecular probes. ATRP initiator was synthesized and coated on silicon surfaces according to previously reported literature procedure. 2 Methods Prior to initiator immobilization, substrates were cleaned using a ovascan PSD Series UV/Digital zone System for 15 minutes. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy was performed on a Thermo Scientific icolet 380 FT-IR spectrophotometer equipped with Harrick Scientific GATR accessory and a Ge crystal. An initiator coated silicon wafer was used as a background during measurements on non-patterned regions of polymer brush coated substrates. Static water contact angles were determined using a
KSV s CAM 101 on the non-patterned part of a polymer brush coated substrate. Atomic force microscopy was performed on an Ambios-Quesant Q-Scope Universal SPM (Scanning Probe Microscope). To determine layer thicknesses, cross-sectional height profiles of patterned polymer brushes were recorded. X-ray photoelectron spectroscopy (XPS) was realized on the nonpatterned regions of a polymer brush by means of a K-Alpha instrument from Thermo Scientific. The X-ray source employed was a monochromatic Al Kα (1486.6 ev) source operated at 100 W and 1 10-9 mbar. Fluorescence microscopy was realized using LD-A-Plan 10x/0.30 objective in Zeiss Axio bserver inverted microscope (ZEISS Fluorescence Microscopy, Carl Zeiss Canada Ltd, Canada). Filter set 38 (Excitation BP 470/40, Emission BP 525/50) was used for imaging of BDIPY-maleimide functionalized polymer brushes and filter set 43 (Excitation BP 545/25, Emission BP 605/70) for imaging of Qdot 605 immobilized polymers brushes. btained fluorescence images were processed using Zeiss AxioVision software. Patterned polymer brushes were prepared by UV irradiation of ATRP initiator modified substrates using literature protocols. 3 Elemental analysis data were obtained from Thermo Electron S.p.A. FlashEA 1112 Elemental Analyzer (CHS separation column, PTFE; 2 m; 6x5 mm). Synthesis of polymer brushes via surface initiated ATRP Representative procedure for synthesis of polymer brushes with 90/10 PEGMEMA/FuMA (P1): 2,2-bipyridine (62.91 mg, 0.403 mmol) was weighted into a round bottom flask with a stir bar. FuMA (0.06 ml, 0.385 mmol), PEGMEMA (0.99 ml, 3.465 mmol), MeH (0.36 ml) and H 2 (0.4 ml) were added to this flask under a nitrogen atmosphere. This mixture was stirred and degassed using nitrogen flow for 15 minutes. Thereafter, Cu(I)Cl (14.11 mg, 0.143 mmol) was added to the flask and degassing was continued for 10 more minutes. The initiator coated Si/Si 2 wafer was placed in a vial and purged under 2. The mixture which containing the monomers and Cu(I) catalyst was added onto the silicon surface and the vial was placed in an oil bath at 60 ºC. After polymerization for a specified time, the surface was washed with MeH (3 2 ml) and H 2 (1 2 ml) and dried under a stream of nitrogen. Polymer brushes with 75:25 (P2), 60:40 (P3) PEGMEMA/FuMA and homopolymer brush of PEGMEMA (P0) were synthesized by changing monomer feed ratios.
80 70 Thickness (nm) 60 50 40 30 20 10 0 30 min 2h 3h 5h 24h 10% FuMA 24 42 53 71 69 25% FuMA 17 26 30 44 43 40% FuMA 15 17 19 24 25 Table S1. Thickness evolution of polymer brush films with polymerization time for 90/10 PEGMEMA/FuMA (blue), 75/25 PEGMEMA/FuMA (red), 60/40 PEGMEMA/FuMA (green) polymer brushes. Figure S2. XPS survey spectra of P1, P2 and P3 polymer brush surfaces.
Functionalization with -Ethylmaleimide Furan containing polymer brushes were incubated in 1 ml of -ethylmaleimide solution in toluene (0.04 mm) at 60 o C for 16 hours and then washed with copious amounts of THF. In order to detach the maleimide-containing molecules via the retro Diels-Alder cycloreversion, - ethylmaleimide functionalized brush coated surfaces were heated in toluene (5 ml) at 110 ºC for 16 hours. Synthesis of BDIPY-maleimide Synthesis of furan protected BDIPY-maleimide. Synthesis of furan protected BDIPY-maleimide is represented in Figure S3. Furan protected maleimide bearing alkyne (2) was synthesized according to the literature procedure. 4 In the first step, BDIPY-azide (130 mg, 0.302 mmol) and furan protected maleimide bearing alkyne (2) (122.56 mg, 0.603 mmol) were dissolved in dry DMF (1.4 ml) and the mixture was stirred for 10 minutes under 2 gas. Degassed Cu(I)Br (8.56 mg, 0.06 mmol) and degassed,,,, - pentamethyldiethylenetriamine (PMDETA) (12.5 µl, 0.06 mmol) were added to the mixture and stirred for 15 minutes. This mixture was immersed in an oil bath at 40 ºC for 16 hours. Thereafter, the solvent was evaporated using a rotary evaporator. CH 2 Cl 2 (150 ml) was added to the remaining solid phase. The mixture was washed with H 2 (3 40 ml) until blue color of aqueous phase was disappeared. Anhydrous a 2 S 4 was added to the combined organic layers and concentrated after filtration and evaporation at 25ºC. The residue was purified by column chromatography with 50:50 ethyl acetate/hexane to obtain the pure furan protected BDIPYmaleimide (95.9 mg, yield: 50.3%). Deprotection of furan protected BDIPY-maleimide (5). Furan protected BDIPY-maleimide was put in a 10 ml toluene containing round bottom flask and was refluxed at 110 ºC. The solvent was evaporated and the residue was dissolved in ethyl acetate (3 ml). The residue was purified by column chromatography with 30:70 ethyl acetate/hexane. Thus, the pure BDIPY-maleimide product was obtained (m= 38.9 mg, yield: 87.2 %). 1 H MR (CDCl 3, δ, ppm): 7.5 (s, 1H), 6.6 (s, 2H), 6 (s, 2H), 4.75 (s, 2H), 4.25 (t, 2H), 2.9 (t, 2H), 2.5 (s, 6H), 2.39 (s, 6H), 1.85 (quintet J=7.8 Hz, 2H), 1.60 (m, 2H), 1.23-1.52 (broad, 12H) (Figure S4). Anal. Calcd. for 5 [C 30 H 39 BF 2 6 2 ]: C, 63.83; H, 6.96;, 14.89. Found: C, 62.36; H, 8.45;, 10.43.
Furan Maleic Anhydride 80 o C dry toluene, 12h Cycloadduct 1 1 72h H 2 MeH, 65 o C -propargylamine 2 2 3 3 +B- F F "CLICK REACTI" Cu(I)Br PMDETA dry DMF 40 o C 4 +B- F F retro Diels-Alder toluene 110 o C 5 +B- F F Figure S3. Synthesis of BDIPY-maleimide. Figure S4. 1 H MR spectrum of BDIPY-maleimide.
Conjugation of BDIPY-maleimide Furan containing polymer brushes were incubated in 1 ml solution of 8.86x10-3 mm dye in toluene at 60 o C for 16 hours. In order to realize deconjugation via the retro Diels-Alder, BDIPY-maleimide functionalized polymer brush surfaces were heated in toluene (5 ml) at 110 ºC for 16 hours. Modified surfaces were washed with copious amounts of toluene and THF and dried under a gentle stream of nitrogen. Immobilization of Biomolecules Polymer brushes were incubated in 1 ml solution of 4.4x10-3 mm biotin maleimide at 60 o C for 16 hours. Surfaces were rinsed with copious amounts of THF and water to remove all residual ligands. Commercial Quantum dots (Q-Dots) streptavidin conjugate solution (5 µl, 1µM) was diluted with distilled water (5 µl) and placed on the patterned biotinylated polymer brush for 30 minutes. Thereafter, surface was washed with copious amount of distilled water and dried under a gentle stream of nitrogen. Figure S5. (a) Conjugation/deconjugation of polymer brush with -ethylmaleimide through Diels-Alder cycloaddition. (b) Atomic percentages of 1s atom after conjugation with -ethyl maleimide at 40 ºC and 60 ºC and after heating up to 60 ºC. (c) XPS survey and high resolution XPS elemental scan of 1s peak of -ethylmaleimide functionalized polymer brush at 60 ºC (left) and after heating up to 60 ºC (right). (d) XPS survey and high resolution XPS elemental
scan of 1s peak of -ethylmaleimide functionalized polymer brush at 40 ºC (left) and after heating up to 60 ºC (right). Figure S6. High resolution XPS elemental scan of 1s peak and ATR-FTIR spectra of carbonyl regions of -ethylmaleimide functionalized poly(pegmema-ran-fuma) brushes REFERECES (1) Altin, H.; Kosif, I.; Sanyal, R. Macromolecules 2010, 43, 3801 3808. (2) Barbey, R.; Klok, H.-A. Langmuir 2010, 23, 18219-18230. (3) Paripovic, D.; Klok, H.-A. ACS Applied Mater. Interfaces 2011, 3, 910-917. (4) Le, D.; Montembault, V.; Soutif, J. C.; Rutnakornpituk, M.; Fontaine, L. Macromolecules 2010, 43, 5611-5617.