Sequence-Defined Polymers via Orthogonal Allyl Acrylamide Building Blocks

Transcription

1 Sequence-Defined Polymers via Orthogonal Allyl Acrylamide Building Blocks Mintu Porel and Christopher A. Alabi* School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, United States Table of content Content Page number Materials and Methods... S2 Monomer synthesis S3-S4 Synthesis of polymer with fluorous support.. S4-S5 Figure S1: Phosphine catalyzed Michael addition kinetics.. S6-S7 Figure S2: Thiol-ene reaction kinetics S8 Figure S3: 1 H NMR spectra of fluorous Boc protected allyl amine. S9 Figure S4: Synthetic scheme of test oligomer.. S10 Figure S5: 1 H NMR spectra of test oligomer synthesis.... S11 Figure S6: 1 H NMR spectra: testing stoichiometry of the thiol-ene reaction... S12-S13 Figure S7: Testing Michael addition kinetics with fluorous substrate.. S14 Figure S8: Stepwise LCMS of test oligomer synthesis..... S15 Figure S9-S14: Stepwise 1 H NMR spectra of ISO1 synthesis.. S16-S21 Figure S15: Stepwise LCMS of ISO1 synthesis... S22 Figure S16: 1 H NMR spectra of ISO1 and ISO2 before and after cleavage. Figure S17: HPLC trace of purified ISO1 and ISO2... S23 S24 Figure S18: Tandem mass spectrum of ISO1... S25 Table S1: Table of calculated and observed m/z of the fragments of ISO1. S25 Figure S19: Tandem mass spectrum of ISO2... S26 Table S2: Table of calculated and observed m/z of the fragments of ISO2. S26 Figure S20: 1 H NMR spectra of 16-mer polymer. S27 Figure S21: Tandem mass spectrum of 16-mer polymer.. S28 Table S3: Table of calculated and observed m/z of the fragments of 16-mer... S29 Figure S22-S29: 1 H NMR spectra of monomers... Figure S30-S35: Assigned 1 H NMR spectra of test oligomers (A-F)... S30-S37 S38-S43 S1

2 Materials and Methods General chemicals were purchased from Sigma Aldrich. Precursors (amines and halides) for the monomer synthesis were purchased from Aldrich and Alfa Aesar. Fluorous BOC-ON (C 9 F 19 BOC-ON) and pre-packed fluorous silica cartridges were purchased from Boron Specialties. UV irradiation for the thiol-ene reaction was performed with a BlueWave 75 UV curing spot lamp (Dymax Corporation). 1 H NMR spectra were recorded on INOVA 400 spectrometers. NMR data was analyzed by MestReNova (version 8.1.1). 1 H NMR chemical shifts are reported in units of ppm relative to tetramethylsilane. NMR data are presented in the following order: chemical shift, peak multiplicity (s = singlet, d = doublet, t = triplet, m = multiplet, dd = doublet of doublet, dt = doublet of triplet), proton number, coupling constant. LCMS experiments were carried out on a Shimadzu HPLC LC20-AD and Thermo Scientific LCQ Fleet with a Sprite TARGA C18 column ( mm, 5 µm, Higgins Analytical, Inc.) monitoring at 215 and 260 nm with positive mode for mass detection. Solvents for LCMS were water with 0.1% acetic acid (solvent A) and acetonitrile with 0.1% acetic acid (solvent B). Compounds were eluted at a flow rate of 0.3 ml/min with 0% solvent B for 2 min, followed by a linear gradient of 0% to 10% solvent B over 2 min, followed by a linear gradient of 10% to 100% solvent B over 5 min, and finally 100% solvent B for 1 min before equilibrating the column back to 0% solvent B over 1 min. MALDI-TOF mass spectrometry was performed on a Waters MALDI micro MX MALDI-TOF mass spectrometer using positive ionization and a linear detector. MALDI samples were prepared by depositing the analyte dissolved in methanol and an alpha-cyano-4-hydroxycinnamic acid matrix onto a stainless steel sample plate. The plate was air dried before loading it into the instrument. HPLC purification was performed on a 1100 Series Agilent HPLC system equipped with a UV diode array detector and a 1100 Infinity analytical scale fraction collector using reverse phase C18 column (4.6 x 150 mm, 5 µm). The column compartment was kept at 25 C during fractionation. Solvents for HPLC were water with 0.1% trifluoroacetic acid (solvent A) and acetonitrile with 0.1% trifluoroacetic acid (solvent B). Compounds were eluted at a flow rate of 1 ml/min with 5% solvent B, followed by a linear gradient of 5% to 100% solvent B over 30 min, and finally 100% solvent B for 5 min before equilibrating the column back to 5% solvent B over 1 min. Polymers were collected based on their absorption at 254 nm. The fractionated polymer was transferred to a vial, dried and stored until further analysis. S2

3 Monomers used in the current study: O N O N O N O N N 2a 2b 2c 2d O N H O N O N O N OH 2e 2f 2g 2h 1. Synthesis of Allyl-N-alkyl/aryl-amines Method A (monomer 2b, 2d and 2g): Br R NH 2 K 2 CO 3 HN R Primary amine derivatives were mixed with 1.2 equivalents of K 2 CO 3 in a round bottom flask and 0.2 equivalent of allyl bromide was added dropwise over a period of 30 min at room temperature and stirred overnight. The reaction mixture was then filtered through celite and washed with CH 2 Cl 2. The filtrate was then concentrated at reduced pressure. Excess primary amine was evaporated under high vacuum. The reaction mixture containing the secondary amine (desired product) and tertiary amine (side product) was used without purification for the subsequent reaction with acryloyl chloride. Method B (monomer 2f and 2h): Allyl amine was mixed with 1.2 equivalents of K 2 CO 3 in a round bottom flask and 0.2 equivalent of alkyl/aryl bromide was added dropwise over a period of 30 min at room temperature and S3

4 stirred overnight. The reaction mixture was then filtered through celite and washed with CH 2 Cl 2. The filtrate was then concentrated at reduced pressure. Allyl amine was evaporated under high vacuum. That reaction mixture containing the secondary amine (desired product) and tertiary amine (side product) was used without purification for the subsequent reaction with acryloyl chloride. Method C (monomer 2c): A solution of allylamine in isopropanol was stirred and treated portion wise with 0.3 equivalent 2-dimethylaminoethyl chloride hydrochloride, followed by 1.2 equivalent of K 2 CO 3. The mixture was allowed to stir at room temperature for one hour, then refluxed for two hours, cooled and diluted with 20 ml of a 6.25 M sodium hydroxide solution. The product was extracted with diethyl ether and dried over sodium sulfate. After evaporation of solvent, the residue was distilled to give the pure product (b.p. 50 C at 15 torr). 2. Synthesis of allyl-n-alkyl/aryl-acrylamide Allyl-N-alkyl/aryl-amines and one equivalent of triethylamine were dissolved in CH 2 Cl 2. The reaction mixture was cooled to 0 C, while being stirred. One equivalent of acryloyl chloride (diluted in 5 ml of CH 2 Cl 2 ) was added drop wise to the reaction mixture over a period of 1 h at 0 C. The reaction mixture was stirred at 0 C for 1 h and at room temperature for 1 h. The reaction mixture was washed twice with water and once with brine solution. The organic layer was then dried over anhydrous Na 2 SO 4, filtered, and concentrated at reduced pressure. The crude reaction mixture was purified by silica gel column chromatography. The product was eluted with 5% MeOH in CH 2 Cl 2. Purity was confirmed by 1 H NMR and LCMS. General method for Fluorous solid-phase extraction (FSPE) The fluorous organic mixture to be separated was loaded onto a 2 g pre-packed fluorous solidphase extraction (FSPE) cartridge. A fluorophobic wash (20% water in methanol) was used to elute all the non-fluorous molecules leaving the fluorous molecules retained on the fluorous silica gel. A fluorophilic wash (100% methanol) was then used to elute the fluorous molecules S4

5 from the fluorous stationary phase. For a 50 mg (~ 0.08mmol) loading of crude mixture onto a 2 g pre-packed FSPE cartridge, two 10 ml fluorophobic washes were used to remove the nonfluorous compounds and one 10 ml fluorophilic wash was used to elute the desired fluorous compound. 3. Synthesis of fluorous Boc protected allyl amine C 9 F 19 O O O CN + H 2 N Et 3 N, THF RT, 2h C 9 F 19 O O N H Allyl amine (6.9 mg, 0.12 mmol) and triethylamine (20 mg, 0.2 mmol) were added to a solution of 94.7 mg (0.13 mmol) of 2-[2-(1H,1H,2H,2H-Perfluoro-9-methyldecyl) isopropoxycarbonyloxyimino]-2-phenylacetonitrile (fluorous BOC-ON) in 10 ml THF. The reaction mixture was stirred at room temperature for 2 h. Thereafter the reaction mixture was concentrated to ~0.3 ml under reduced pressure and purified by FSPE. Methanol was evaporated under reduced pressure to yield the fluorous Boc protected allyl amine as a white solid product. Purity of the product was confirmed by 1 H NMR. 1 H NMR (400 MHz, CDCl 3 ): δ 1.47 (s, 6H), 1.97 (m, 2H), 2.10 (m, 2H), 3.73 (s, 2H), 4.66 (m, 1H) and 5.11 (m, 2H), 5.82 (m, 1H). General method for Thiol-ene reaction 1,3-Propanedithiol (0.4 mmol) and 2,2-dimethoxy-2-phenylacetophenone (DMPA, 5 mol % of 1,3-propanedithiol) were added to a solution of corresponding fluorous-olefin (0.08 mmol) in methanol (300 µl). The reaction mixture was subjected to UV irradiation for 90 s at 20 mw/cm 2. The product (fluorous-thiol) was purified by FSPE. General method for Michael addition Allyl-N-alkyl/aryl-acrylamides monomer (0.16 mmol) and dimethyl phenyl phosphine (Me 2 PhP, 5 mol% of monomer) were added to corresponding fluorous-thiol (0.08 mmol) in methanol (300 µl) and stirred for 5 min at room temperature. The product was purified by FSPE. General method for fluorous Boc deprotection The fluorous Boc protected polymer was dissolved in a 50% TFA/CH 2 Cl 2 solution and stirred for 2.5 hours. The resulting mixture was purified by FSPE. In this case, the organic solution that elutes with the fluorophobic wash (20% water in methanol) is the desired product. The eluted product was dried under reduced pressure. Typical polymer concentration for the deprotection is 50 mm. S5

6 Kinetics experiment of Michael addition 1 H NMR spectra of a mixture of N-methyl allyl acrylamide (0.1 mmol) and Me 2 PhP (5 mol% of N-methyl allyl acrylamide) in CD 3 OH (600 µl) was recorded and taken as the 0 min time point. 1,3-propanedithiol (0.2 mmol) was added to this solution, mixed quickly and a 1 H NMR spectrum was recorded every minute. The addition, mixing and recording of the first 1 H NMR was complete within one minute. The progress of the Michael addition was monitored via disappearance of acryloyl olefin proton signal at 6.74 ppm. The percentage of consumed starting material was determined by integration of the 1 H NMR chemical shift at 6.74 ppm relative to that at 2.24 ppm (methyl proton signal from the Me 2 PhP catalyst in the reaction mixture as determined via integration of spectra B (i), see below), which remained constant throughout the reaction. The allyl olefin chemical shift at 5.2 ppm remained constant throughout the reaction indicating that there was no di-addition. S6

7 Figure S1 (A) 1 H NMR spectra (400 MHz, CD 3 OH) of (A) Michael addition reaction mixture mentioned above at different time intervals, (B) (i) Mixture of 2a and Me 2 PhP, (ii) Me 2 PhP (in CD3OD) and (iii) 2a (in CD 3 OD) and (C) kinetic plot of the reaction progress. S7

8 Kinetics experiment of thiol-ene reaction The starting material N-allyl-N-methyl-3-(octylthio)-acrylamide (AMOA) was synthesized by mixing 1-octane thiol and N-methyl allyl acrylamide in presence of 5 mol% propyl amine as catalyst for 24 hours. After removal of propylamine, the reaction was deemed quantitative via 1 H NMR. 1,3-Propanedithiol (0.1 mmol) and DMPA (5 mol% of 1,3-propanedithiol) were added to a solution of AMOA (0.08 mmol) in methanol (300 µl). The reaction mixture was split equally into five vials and each was UV irradiated for 15, 30, 45, 60 and 90s respectively at 20 mw/cm 2. 1 H NMR spectra of the five reaction mixtures were recorded. 1 H NMR of AMOA was used as the 0 s time point. The reaction progress was monitored via integration of the 1 H NMR signals at 5.14 ppm (olefin proton) relative to that at 0.77 ppm. The latter represents the methyl proton of the octyl group, which remained constant throughout the reaction. Figure S2 (i) Partial 1 H NMR spectra (400 MHz, CDCl 3 ) of the thiol-ene reaction mixture mentioned above at different time intervals and (ii) kinetic plot of the reaction progress. S8

9 Figure S3 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) fluorous BOC-ON and (ii) BOC protected fluorous allyl amine; * represents the residual proton signal of CDCl 3. S9

10 Figure S4 Synthesis of the test oligomer using N-allyl-N-methylacrylamide and 1,3- propanedithiol S10

11 Figure S5 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) A, (ii) B, (iii) C, (iv) D, (v) E and (vi) F; * and represent the residual proton signals of CDCl 3 and MeOH respectively. Spectral assignments for compounds A-F are provided in Figure S30-S35. S11

12 (A) S12

13 Figure S6 (A) 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) A; product from the reaction mixture of A and 1,3-propanedithiol at a ratio of (ii) 1:5; (iii) 1:2; (iv) 1:1; (v) 1:0.5 and (vi) 0:1, i.e. only 1,3-propanedithiol. The reaction was performed in the presence of 5 mol% DMPA (w.r.t BOC protected fluorous allylamine), hν = 20 mw/cm 2, 90 sec; The blue and red dots represent olefin and thiol proton signals respectively. The reaction did not go to 100% conversion even after addition of 2 eq of dithiol, but was complete in presence of 5 eq dithiol. *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. (B) Expanded region of 1 H NMR spectra of a reaction mixture of A and 1,3-propanedithiol at a ratio of (i) 1:5; (ii) 1:0.5 and (iii) 0:1, i.e. only 1,3-propanedithiol. The proton signals of 1,3-propanedithiol undergo a slight chemical shift upon conjugation with the fluorous tag (see a, b, and c). Hence, the spectrum in B(ii) suggests that the thiol signal at 1.33 ppm is only from mono-addition product. S13

14 Figure S7 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) B; reaction mixture of B with 2 equivalent N-allyl-N-methylacrylamide in presence of 5 mol% of Me 2 PhP after reaction for (ii) 180 s and (iii) 300 s. Blue dots and red dots represent olefin proton signals and thiol proton signals respectively. The presence of the thiol proton signal after 180 s indicates that reaction was not complete, whereas disappearance of thiol signals after 300 s reactions confirms completion of the reaction. S14

15 Figure S8 LCMS of (i) B, calculated for (M+H) , observed ; (ii) C, calculated for (M+H) , observed ; (iii) D, calculated for (M+H) , observed ; (iv) E, calculated for (M+H) , observed ; (v) F, calculated for (M+H) , observed S15

16 Figure S9 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) A, (ii) G and (iii) H; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S16

17 Figure S10 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) I and (ii) J; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S17

18 Figure S11 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) K and (ii) L; * represents the residual proton signals of CDCl 3. S18

19 Figure S12 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) M and (ii) N; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S19

20 Figure S13 1 H NMR spectra (400 MHz, CDCl 3 ) of (i) O and (ii) P; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S20

21 Figure S14 1 H NMR spectra (400 MHz, CDCl 3 ) of (A) (i) Q, and (ii) R with assigned proton signals. The alkyl region could not be accurately assigned due to large number of overlapping proton signals. (B) (i) TFA: H 2 O = 1:1; (ii) TFA: H 2 O = 1:0.5 and (iii) TFA. The 1 H NMR spectra in B suggests that the proton signal of TFA is shifted up field upon hydration, indicating that the proton signal at 5.8 ppm of A(ii) is due to hydrated TFA and protonated amines. * and represent the residual proton signals of CDCl 3, and MeOH respectively. S21

22 Figure S15 LCMS of (i) H, calculated for (M+H) , observed ; (ii) J, calculated for (M+H) , observed ; (iii) L, calculated for (M+H) , observed , (M+2H) ; (iv) N, calculated for (M+H) , observed , (M+2H) ; (v) P, calculated for (M+H) , observed , (M+2H) S22

23 Figure S16 1 H NMR spectra (400 MHz, CDCl 3 ) of (A) before cleavage of (i) ISO2 and (ii) ISO1 and (B) after cleavage of (i) ISO2 and (ii) ISO1 with assigned proton signals. The alkyl region could not be assigned due to large number of overlapping proton signals; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S23

24 Figure S17 HPLC trace of purified (i) ISO1 (retention time = 17.8 min) and (ii) ISO2 (retention time = 17.1 min); * represents the product signal and represents residual loading solvent signals (these peaks show up in a blank run). S24

25 M 1 M2 M 3 M 4 M 5 N 5 N 4 N 3 N 2 N 1 + c ESI Full ms N Instrument scan range = 190 to 2000 M 1 =166.08, was out of range N M N N 4 M 3 M M 5 N m/z Figure S18 Tandem mass spectrum of ISO1 Table S1 Calculated and observed m/z of the fragments of ISO1 Fragment Calculated Observed Fragment Calculated Observed (m/z) (m/z) (m/z) (m/z) M n/a * N M N M N M N M N *m/z was out of the instrument scan range S25

26 N 2 N 3 M 1 M 2 M 3 M 4 M 5 N 5 N 4 N 3 N 2 N 1 T: ITMS + c ESI Full ms @cid50.00 [ ] M M M 3 M m/z N 4 N m/z m/z Instrument scan range = 190 to 2000 M 1 = and N 1 = , were out of range Figure S19 Tandem mass spectrum of ISO2 Table S2 Calculated and observed m/z of the fragments of ISO2 Fragment Calculated Observed Fragment Calculated Observed (m/z) (m/z) (m/z) (m/z) M n/a * N n/a * M N M N M N M N *m/z was out of range of the instrument scan range S26

27 Figure S20 1 H NMR spectra (400 MHz, CDCl 3 ) of 16-mer polymer (i) before and (ii) after cleavage from fluorous tag with assigned proton signals. The alkyl region could not be assigned due to large number of overlapped proton signals; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and MeOH respectively. S27

28 Figure S21 Tandem mass spectrum of 16-mer polymer S28

29 Table S3 Calculated and observed m/z of the fragments of 16-mer polymer Fragment Calculated Observed Fragment Calculated Observed (m/z) (m/z) (m/z) (m/z) M n/a * N n/a * M N ** M N M N M N M N M N *** M *** N n/a * *m/z was out of range of the instrument scan range ** (M+H-H 2 O) + was observed *** (M+2H) 2+ was observed S29

30 Figure S22 1 H NMR spectra (400 MHz, CDCl 3 ) of 2a; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and tetramethylsilane respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 2.95 (m, 1.7H), δ 2.98 (m, 1.3H), 3.93 (m, 1H), 4.01 (m, 1H), 5.14 (m, 2H), 5.62 (m, 1H), 5.74 (m, 1H) and 6.42 (m, 2H), inset: LCMS calculated for (M+H) , observed S30

31 Figure S23 1 H NMR spectra (400 MHz, CDCl 3 ) of 2b; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and tetramethylsilane respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 0.86 (m, 3H), 1.25 (m, 2H), 1.48 (m, 2H), 3.24 (t, 1H, J=8 Hz), 3.32 (t, 1H, J=8 Hz), 3.89 (m, 1H), 3.97 (m, 1H), 5.11 (m, 2H), 5.60 (m,1h), 5.75 (m,1h), 6.38 (m,2h); inset: LCMS calculated for (M+H) , observed S31

32 Figure S24 1 H NMR spectra (400 MHz, CDCl 3 ) of 2c; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and tetramethylsilane respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 2.22 (s, 6H), 2.43 (m, 2H), 3.39 (t, 0.8H, J= 8 Hz), 3.48 (t, 1.2H, J= 8 Hz), 3.98 (m, 1.2H), 4.04 (m, 0.8H), 5.15 (m, 2H), 5.64 (m, 1H), 5.76 (m, 1H), 6.43 (m, 2H); inset: LCMS calculated for (M+H) , observed , is for (M+H- NHMe 2 ) +. S32

33 Figure S25 1 H NMR spectra (400 MHz, CDCl 3 ) of 2d; *, and represent the residual proton signals of CDCl 3, CH 2 Cl 2 and tetramethylsilane respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 0.89 (m, 3H), 0.91 (m, 3H), 1.96 (m, 1H), 3.13 (d, 1H, J= 8 Hz), 3.24 (d, 1H, J= 8 Hz), 3.97 (m, 1H), 4.04 (m, 1H), 5.16 (m, 2H), 5.66 (m, 1H), 5.79 (m, 1H), 6.46 (m, 2H); inset: LCMS calculated for (M+H) , observed S33

34 Figure S26 1 H NMR spectra (400 MHz, CDCl 3 ) of 2e; * and represent the residual proton signals of CDCl 3 and tetramethylsilane respectively. 1 H NMR (400 MHz, CDCl 3 ): δ 3.89 (m, 2H), 5.10 (m, 2H), 5.58 (dd, 1H, J= 0.8, 0.4 Hz), 5.79 (m, 1H), 6.20 (m, 2H), 6.66 (s, 1H); inset: LCMS calculated for (M+H) , observed S34

35 Figure S27 1 H NMR spectra (400 MHz, CDCl 3 ) of 2f; * and represent the residual proton signals of CDCl 3 and CH 2 Cl 2 respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 3.44 (t, 0.6H, J=4 Hz), 3.52 (t, 1.4H, J= 4 Hz), 3.69 (t, 0.7H, J= 4 Hz), 3.73 (t, 1.3H, J= 4 Hz), 3.82 (s, 1H), 4.01 (m, 2H), 5.15 (m, 2H), 5.66 (m, 1H), 5.77 (m, 1H), 6.40 (m, 2H); inset: LCMS calculated for (M+H) , observed S35

36 Figure S28 1 H NMR spectra (400 MHz, CDCl 3 ) of 2g; * represents the residual proton signals of CDCl 3. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 0.88 (t, 3H, J=7.6 Hz), 1.58 (m, 2H), 3.24 (t, 1H, 7.6 Hz), 3.33 (t, 1H, 7.6 Hz), 3.94 (m, 1H), 4.03 (m, 1H),5.15 (m, 2H), 5.64 (m, 1H), 5.77 (m, 1H), 6.44 (m, 2H); inset: LCMS calculated for (M+H) , observed S36

37 Figure S29 1 H NMR spectra (400 MHz, CDCl 3 ) of 2h; * and represent the residual proton signals of CDCl 3 and CH 2 Cl 2 respectively. The product is a mixture of two rotational isomers. 1 H NMR (400 MHz, CDCl 3 ): δ 2.90 (m, 2H), 3.61 (m, 2H), 3.85 (m, 1.1H), 4.04 (m, 0.9H), 5.16 (m, 2H), 5.68 (m, 1H), 5.80 (m, 1H), 6.42 (m, 2H), 7.26 (m, 5H); inset: LCMS calculated for (M+H) , observed S37

38 a c c e g c, c b d f g A f g,g d e a b Figure S30 1 H NMR spectra (400 MHz, CDCl 3 ) of A with assigned proton signals S38

39 Figure S31 1 H NMR spectra (400 MHz, CDCl 3 ) of B with assigned proton signals S39

40 Figure S32 1 H NMR spectra (400 MHz, CDCl 3 ) of C with assigned proton signals S40

41 Figure S33 1 H NMR spectra (400 MHz, CDCl 3 ) of D with assigned proton signals S41

42 Figure S34 1 H NMR spectra (400 MHz, CDCl 3 ) of E with assigned proton signals S42

43 Figure S35 1 H NMR spectra (400 MHz, CDCl 3 ) of F with assigned proton signals S43