Supporting information. Total Synthesis of (+)-ent-cyclizidine: Absolute Configurational Confirmation of Antibiotic M

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1 Supporting information Total Synthesis of (+)-ent-cyclizidine: Absolute Configurational Confirmation of Antibiotic M Stephen Hanessian, Udaykumar Soma, Stéphane Dorich and Benoît Deschênes-Simard Department of Chemistry, Université de Montréal, C. P. 6128, Succ. Centre-Ville, Montréal, P. Q., Canada H3C 3J7 Table of contents General Information S1 Experimental procedures S2 - S17 X-ray structure data S17- S20 References S21 1 H and 13 C NMR spectra S22 S62 General information. All non-aqueous reactions were run in flame-dried glassware under a positive pressure of argon with exclusion of moisture from reagents and glassware using standard techniques for manipulating air-sensitive compounds. Anhydrous solvents were obtained using standard drying techniques. Unless stated otherwise, commercial grade reagents were used without further purification. Reactions were monitored by analytical thin-layer chromatography (TLC) performed on pre-coated, glass-backed silica gel plates. Visualization of the developed chromatogram was performed by UV absorbance, aqueous cerium ammonium molybdate, iodine, or aqueous potassium permanganate. Flash chromatography was performed on mesh silica gel with the indicated solvent systems. Melting points are uncorrected. Infrared spectra were recorded on a FT-IR spectrometer and are reported in reciprocal centimeters (cm -1 ). Routine nuclear magnetic resonance spectra were recorded either on AV-300, AV-400, or AV-700 spectrometer. Chemical shifts for 1 H NMR spectra are recorded in parts per S1

2 million from tetramethylsilane with the solvent resonance as the internal standard (CHCl3, δ 7.27 ppm). Data are reported as follows: chemical shift, multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, qn = quintet, m = multiplet and br = broad), coupling constant in Hz, and integration. Chemical shifts for 13 C NMR spectra are recorded in parts per million from tetramethylsilane using the central peak of the solvent resonance as the internal standard (CDCl 3, δ ppm). All spectra were obtained with complete proton decoupling. ptical rotations were determined at 589 nm at ambient temperature. Data are reported as follows: [α]d, concentration (c in g/100 ml), and solvent. High-resolution mass spectra were performed by the Centre régional de spectroscopie de masse de l'université de Montréal using fast atom bombardment (FAB) or electrospray ionization (ESI) techniques. Low-resolution mass spectra were obtained using electrospray ionization (ESI). Experimental procedures: Propargyl alcohol 4: NBoc MgBr NBoc Me THF, rt, 92% H 3 4 To a stirred solution of the methyl ketone 3 1 (10.4 g, 42.7 mmol) in dry THF (100 ml) was added ethynyl magnesiumbromide (0.5 M sol. in THF, 213 ml, 107 mmol) at rt and stirred at the same temperature for 2 h. The reaction was quenched at 0 o C with an aqueous saturated solution of NH 4 Cl, and extracted with EtAc (2 times). The combined organic layers were then washed with water and brine sequentially, dried over anh. Na 2 S 4 and concentrated under reduced pressure. Flash column chromatography using ethyl acetate-hexane (12-15% EtAc/hexanes) afforded the propargylic alcohol 4 (10.5 g, 92%) as white solid. R f : 0.45 (10% EtAC in hexanes); [α] D o (c 1.39, CHCl 3 ); IR (neat): υ max 3273, 2999, 2920, 1645, 1368, 1061, 773, 677 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ 5.93 (s, 1H), 4.31 (s, 1H, br), (m, 2H), 2.47 (s, 1H), 1.67 (s, 3H), 1.51 (s, 12H), 1.44 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 155.8, 95.0, 86.5, 81.9, 71.7, 71.1, S2

3 65.5, 64.7, 27.9, 25.7, 23.9, 23.5; HRMS (ESIMS): calcd for C 14 H 23 N 4 Na: [M+Na] +, ; found: Allyl alcohol 5: NBoc NBoc LAH, THF 93% Me H Me H 4 5 To a solution of alkyne 4 (10.5 g, 39.0 mmol) in dry THF (100 ml) was added LAH (4.5 g, 117 mmol) in portions at 0 o C, then left to warm up to rt and stirred for 4 h. The reaction was quenched with an aqueous saturated solution of Na 2 S 4, and the mixture was extracted three times with EtAc. The combined organic washes were then washed with H 2 and brine, then dried over sodium sulfate and concentrated under vacuum. The crude product was purified by column chromatography (15% EtAc/hexanes) to afford the olefin 5 as a white solid (9.8 g, 93%). R f : 0.45 (10% EtAC in hexanes); [α] D o (c 1.34, CHCl 3 ); IR (neat): υ max 3375(br), 2980, 2936, 1698, 1662, 1393, 1367, 1251, 1171, 1066, 926, 849, 772 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ 5.98 (dd, J = 10.8, 17.1 Hz, 1H), 5.46 (s, 1H, br), 5.36 (d, J = 17.1 Hz), 5.13 (d, J = 10.8 Hz, 1H), 3.93 (s, 1H), 3.89 (t, J = 7.0 Hz, 1H), 3.78 (d, J = 8.5 Hz, 1H), 1.58 (s, 3H), 1.48 (s, 12H), 1.23 (s, 3H). 13 C NMR (CDCl 3, 100 MHz) δ 155.6, 142.3, 114.3, 94.4, 81.4, 75.1, 64.9, 64.6, 27.9, 25.8, 23.7, 21.2; HRMS (ESIMS): calcd for C 14 H 25 N 4 Na: [M+Na] +, ; found: Triol 6: NBoc AD-Mix-β NBoc H t-buh:h 2, CH 3 S 2 NH 2 Me H 80% Me H 5 6 H To a solution of the olefin 5 (9.74 g, mmol) in tert-butanol and water (1:1), (80 ml) at 0 o C was added AD-Mix-β (50.3 g, 1.4 g/mmol) followed by MeS 2 NH 2 (3.42 g, mmol) and stirred the resulting yellow slurry at rt for 24 h. The reaction mixture was then quenched with sodium sulfite (50 g) and the resulting greenish mixture was S3

4 stirred for 2 h and then extracted with EtAc (3-4 times) and washed with aqueous 2 N KH, water and brine. The organic phase was dried over Na 2 S 4, concentration under vacuum and purification by column chromatography (60-70% EtAC/hexanes) afforded the triol 6 (8.7 g, 80%) as a colorless syrup. R f : 0.3 (60% EtAC in hexanes); [α] D o (c 1.28, CHCl 3 ); IR (neat): υ max 3373 (br), 2979, 2937, 1659, 1395, 1377, 1367, 1250, 1171, 1064, 873, 770 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ 6.45 (s, 1H, br), 4.47 (d, J = 6.9 Hz, 1H), 4.11 (m, 1H), 4.0 (m, 1H), 3.88 (dd, J = 2.6, 9.2 Hz, 1H), 3.71 (dd, J = 2.6, 12.7 Hz, 1H), 3.38 (m, 1H), 1.60 (s, 3H), 1.52 (s, 3H), 1.51 (s, 9H), 1.15 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 156.1, 94.2, 81.9, 78.6, 73.5, 62.8, 64.4, 27.9, 25.6, 23.7, 13.7, 13.5; HRMS (ESIMS): calcd for C 14 H 27 N 6 Na: [M+Na] +, ; found: Silyl ether: NBoc H TBDPS-Cl, Et 3 N NBoc H H DMAP, CH 2 Cl TBDPS 2 Me H 82% Me H 6 To a solution of triol 6 (5.12 g, mmol) in CH 2 Cl 2 (45 ml), was added triethylamine (4.68 ml, mmol), TBDPSCl (4.72mL, mmol), and DMAP (205 mg, 1.68 mmol) at 0 o C. Stirred at rt for 3 h, then quenched with a saturated aqueous NH 4 Cl solution. The mixture was extracted three times with EtAc, and the combined organic layers were washed with water and brine. The organic layer was then dried over anhydrous Na 2 S 4 and concentrated under reduced pressure. Flash column chromatography of the crude product (6-10% EtAc/hexanes) gave the primary TBDPS ether (7.45 g, 82%) as a colorless viscous liquid. R f : 0.35 (15% EtAC in hexanes); [α] D o (c 2.42, CHCl 3 ); IR (neat): υ max 3363 (br), 2933, 2887, 2858, 1661, 1393, 1376, 1367, 1171, 1110, 1063, 824, 702, 504 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (m, 6H), 5.60 (s, 1H, br), 4.44 (d, J = 6.0 Hz, 1H), 4.02 (m, 1H), (m, 2H), 3.86, (m, 1H), 3.56 (m, 1H), 1.54 (s, 3H), 1.52 (s, 3H), 1,51 (s, 9H), 1.11 (s, 9H), 1.10 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 155.9, 135.2, 132.6, 129.5, 129.4, 127.4, 94.1, 81.4, 75.3, 76.0, 64.2, 62.6, 27.9, 26.5, 25.7, 19.1, 18.8; HRMS (ESIMS): calcd for C 30 H 45 N 6 NaSi: [M+Na] +, ; found: S4

5 BM ether (7): NBoc H Me H TBDPS BM-Cl, DIPEA ClCH 2 CH 2 Cl, TBAI, 50 o C, 78% NBoc Me H 7 BM TBDPS To a solution of TBDPS ether (7.3 g, mmol) in 1,2-dichloroethane (40 ml) was added DIPEA (5.97 ml, mmol), BMCl (60%, 3.42 ml, mmol) and TBAI (496 mg, 1.34 mmol) at rt, then heated at 50 o C for 12 h. The reaction mixture was cooled to rt and added water, extracted with EtAc, the organic phase was washed with aqueous saturated aqueous sodium thiosulfate solution, NaHC 3 solution, water, brine and dried (Na 2 S 4 ). The crude product was then purified by flash column chromatography (6-8% EtAc/hexanes) to afford 7 as a colorless liquid (6.9 g, 78%). R f : 0.45 (10% EtAc in hexanes); [α] D o (c 1.46, CHCl 3 ); IR (neat): υ max 3354 (br), 2933, 2858, 2888, 1661, 1427, 1391, 1376, 1366, 1172, 1112, 1037, 1026, 739, 701, 504 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (m, 10H), 5.33 (s, 1H, br), 5.04 (d, J = 6.8 Hz, 1H), 4.90 (d, J = 6.8 Hz, 1H), 4.79 (d, J = 11.8 Hz, 1H), 4.54 (d, J = 11.8 Hz, 1H), 4.44 (d, J = 6.5 Hz, 1H), 4.07 (d, J = 10.3 Hz, 1H), 3.93 (dd, J = 6.5, 11.0 Hz, 1H), 3.77 (m, 1H), 3.61 (m, 1H), 1.61 (s, 3H), 1.51 (s, 9H), 1.44 (s, 3H), 1.31 (s, 3H), 1.07 (s, 9H); 13 C NMR (CDCl 3, 100 MHz) δ 156.0, 135.3, 135.2, 135.1, 134.5, 129.6, 129.2, 128.1, 127.5, 95.4, 94.2, 83.6, 81.2, 69.9, 64.1, 63.6, 61.8, 28.0, 27.9, 26.6, 26.4, 24.4, 18.8, 18.7; HRMS (ESIMS): calcd for C 38 H 53 N 7 NaSi: [M+Na] +, ; found: Cyclic carbamate (8): NBoc BM N NaHMDS, THF TBDPS rt, 30 min, 81% Me Me H BM TBDPS 7 8 To a stirred solution of the tertiary alcohol 7 (6.14 g, 9.25 mmol) in dry THF (30 ml) was treated with NaHMDS (13.9 ml, 1 M solution in THF, 13.9 mmol) at 0 o C and stirred at rt for 30 min. The reaction was quenched with a aqueous saturated solution of NH 4 Cl, and washed three times with EtAc. The combined organic layer was then S5

6 washed with brine and dried over anh. Na 2 S 4 and concentrated under reduced pressure. Flash column chromatography (15% EtAc/hexanes) afforded the cyclic carbamate 8 as a colorless oil (4.4 g, 81%). R f : 0.35 (25% EtAC in hexanes); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 2933, 2891, 2858, 1762, 1428, 1386, 1259, 1112, 1050, 1026, 741, 702, 504 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (m, 6H), (m, 5H), 4.92 (s, 2H), 4.71 (d, J = 11.9 Hz, 1H), 4.57 (d, J = 11.9 Hz, 1H), 4.51 (t, J = 6.1 Hz, 1H), (m, 2H), (m, 3H), 1.75 (s, 3H), 1.43 (s, 3H), 1.42 (s, 3H), 1.11 (s, 9H); 13 C NMR (100 MHz, CDCl 3 ) δ 155.2, 137.2, 135.2, 135.2, 132.3, 132.2, 129.7, 129.7, 128.1, 127.9, 127.6, 127.6, 127.4, 94.6, 93.9, 82.6, 80.3, 69.7, 63.8, 63.0, 62.8, 26.5, 26.4, 23.1, 18.8, 17.9; HRMS (ESIMS): calcd for C 34 H 43 N 5 NaSi: [M+Na] +, ; found: Hydroxy carbamate: N HN ptsa, MeH H Me rt, 2 h, 82%, Me BM TBDPS BM TBDPS 8 To a solution of bicyclic carbamate acetonide 8 (4.4 g, 7.47 mmol) in dry MeH (15 ml) at 0 o C was added ptsa (141 mg, 0.75 mmol), stirred at rt for 2 h, quenched the reaction mixture with NaHC 3 and extracted with EtAc (3 times) and washed the combined organic layers with water and brine, dried over anhydrous Na 2 S 4 and solvent was removed under reduced pressure to afford the primary alcohol as colorless liquid (3.36 g, 82%) after flash column chromatography using ethyl acetate- hexane (1:1) as eluant. R f : 0.35 (60% EtAc in hexanes); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 3306 (br), 2932, 2858, 1747, 1427, 1381, 1281, 1166, 1112, 1039, 740, 701 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (6H, m), (5H, m), 6.70 (1H, s, br), 4.93 (d, J = 6.8 Hz, 1H ), 4.87 (d, J = 6.8 Hz, 1H), 4.69 (d, J = 11.9 Hz, 1H) 4.55 (d, J = 11.9 Hz, 1H), 3.98 (s, 1H, br), (m, 3H), (m, 2H), 3.65 (1H, s, br), 1.42 (s, 3H), 1.09 (s, 9H); 13 C NMR (CDCl 3, 100 MHz) δ 158.8, 137.2, 135.3, 135.2, 132.4, 132.2, 129.7, 129.6, 128.1, 127.9, 127.6, 127.5, 127.3, 95.0, 84.2, 80.7, 69.7, 63.0, S6

7 61.6, 57.9, 26.3, 16.9, 13.4; HRMS (ESIMS): calcd for C 31 H 39 N 6 NaSi: [M+Na] +, ; found: Compound 9: H HN Me 1. DMP, DCM 2. Ph 3 P=CH 2 62%, 2 steps BM TBDPS BM TBDPS 9 To a stirred solution of primary alcohol (3.3 g, 6.01 mmol) in CH 2 Cl 2 (20 ml) was added Dess-Martin periodinane (3.82 g, 9.05 mmol) at 0 o C and the reaction mixture was stirred at rt for 1 h. The reaction mixture was quenched with an aqueous saturated solution of Na 2 S 2 3 /NaHC 3 (7:1) (30 ml). The mixture was stirred vigorously until the two layers were clear. The crude product was extracted with EtAc (50 ml x 3). The combined organic layers were washed with water, brine, dried over anhydrous Na 2 S 4 and concentrated under reduced pressure. The crude residue was then engaged in the subsequent olefination reaction without further purification. To a solution of the crude aldehyde in THF (20 ml) at 0 o C was added a freshly prepared Ph 3 P=CH 2 (generated in another flask by adding equivalent amount of K t Bu to Ph 3 PCH 3 Br in THF) in excess and the mixture was stirred for one hour at rt until there was total conversion of the starting material as indicated by a TLC. The reaction mixture was quenched with an aqueous saturated solution of NH 4 Cl and extracted with EtAc (3 times) washed with water, brine and dried over Na 2 S 4. The crude product was purified by flash chromatography on silica gel (30 % EtAc in hexanes) to give 9 as a colorless syrup (2.03 g, 62%). R f : 0.35 (30% EtAc in hexanes); [α] D o (c 1.16, CHCl 3 ); IR (neat): υ max 3262 (br), 2932, 2858, 1755, 1427, 1112, 1043, 1026, 741, 701 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (m, 6H), (m, 5H), 5.76 (m, 1H), 5.46 (s, 1H, br), 5.23 (d, J = 10.4 Hz, 1H), 5.16 (d, J = 17.1 Hz, 1H), 4.97 (d, J = 6.9 Hz, 1H), 4.90 (d, J = 6.9 Hz, 1H), 4.73 (d, J = 11.9 Hz, 1H), 4.58 (d, J = 11.9 Hz, 1H), 4.44 (d, J = 7.0 Hz, 1H), (m, 3H), 1.31 (s, 3H), 1.07 (s, 9H); 13 C NMR (CDCl 3, 100 MHz) δ 157.4, 137.3, 135.2, 135.2, 133.2, 132.5, 132.3, 129.6, 129.5, 128.1, HN Me S7

8 127.5, 127.4, 127.4, 127.3, 118.6, 94.8, 85.4, 80.9, 69.8, 63.0, 59.1, 26.4, 18.9, 18.6; HRMS (ESIMS): calcd for C 32 H 39 N 5 NaSi: [M+Na] +, ; found: Diene 10: NaHMDS, THF HN N Me Tf Me 78% BM TBDPS BM TBDPS 9 10 A solution of cyclic carbamate 9 (2.0 g, 3.66 mmol) in dry THF (6.0 ml) was treated with NaHMDS (5.5 ml, 1.0 M solution in THF, 5.5 mmol) at 0 o C, after 15 min., added a solution of freshly prepared 3-buten-1-ol-triflate ester (4.03 mmol) in THF (2.5 ml) in drop-wise. Stirred for 1-2 h until there was total dissaprearance of the starting material. Then, the reaction was quenched with a saturated aqueous solution of NH 4 Cl and extracted with EtAc (three times), washed the combined organic phases with water, brine, dried (Na 2 S 4 ) and concentrated in vacuo. The crude product was purified by flash chromatography on silica gel (20% EtAc/hexanes) to give 10 as a colorless liquid (1.71 g, 78%). R f : 0.4 (20% EtAC in hexanes); [α] D o (c 1.4, CHCl 3 ); IR (neat): υ max 3070, 2931, 2857, 2890, 1755, 1427, 1279, 1172, 1112, 1045, 1026, 741, 702 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 4H), (m, 11H), (m, 2H), 5.30 (d, J = 10.6 Hz, 1H), (m, 4H), 4.94 (d, J = 6.6 Hz, 1H), 4.75 (d, J = 11.9 Hz, 1H), 4.60 (d, J = 11.9 Hz, 1H), 4.16 (d, J = 9.0 Hz, 1H), 3.86 (d, J = 3.69 Hz, 1H), (m, 2H), 3.45 (m, 1H), 2.99 (m, 1H), (m, 2H), 1.30 (s, 3H), 1.08 (s, 9H); 13 C NMR (CDCl 3, 100 MHz) δ 156.2, 137.4, 135.3, 134.4, 132.8, 129.6, 128.0, 127.5, 127.4, 121.6, 116.7, 95.1, 82.3, 81.1, 69.7, 63.4, 63.2, 41.0, 31.3, 26.4, 18.8, 18.2; HRMS (ESIMS): calcd for C 36 H 45 N 5 NaSi: [M+Na] +, ; found: S8

9 Diene alcohol: N Me TBAF, THF 90% N Me BM TBDPS BM H 10 To a solution of diene 10 (1.7 g, 2.84 mmol) in THF (5 ml) was added TBAF (3.1 ml, 1.0 M in THF, 1.1 eq.) at 0 o C. The mixture was stirred ar rt until complete consumption of starting material. Aqueous saturated NH 4 Cl (20 ml) was then added and the aqueous phase was extracted three times with EtAc. The combined organic layers were washed with water, brine, dried over Na 2 S 4 and concentrated in vacuo. The crude product was purified by flash chromatography (50-60% EtAc/hexanes), to give the free alcohol as a colorsless liquid (922 mg, 90%). R f : 0.35 (50% EtAC in hexanes); [α] D -7.3 o (c 1.0, CHCl 3 ); IR (neat): υ max 3444 (br), 2935, 1731, 1418, 1382, 1167, 1026, 931, 738, 699 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 5H), (m, 2H), (m, 2H), (m, 2H), 5.00 (d, J = 6.9 Hz, 1H), 4.83 (d, J = 6.9 Hz, 1H), 4.76 (d, J = 11.7 Hz, 1H), 4.65 (d, J = 11.7 Hz, 1H), 4.19 (d, J = 9.1 Hz, 1H), 3.75 (m, 1H), (m, 2H), 3.48 (m, 2H), 3.11 (s, 1H, br), 3.02 (m, 1H), (m, 2H), 1.31 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 156.1, 136.5, 134.5, 132.2, 128.2, 127.7, 127.6, 122.0, 117.0, 95.8, 85.2, 82.0, 70.2, 63.3, 61.0, 41.0, 31.3, 18.5; HRMS (ESIMS): calcd for C 20 H 27 N 5 Na: [M+Na] +, ; found: ,4-Unsaturated piperidine 11: N BM Me H Grubbs II gen. catalyst CH 2 Cl 2, reflux, 2 h, 91% N BM Me H 11 To a stirred solution of diene (860 mg, 2.38 mmol) in CH 2 Cl 2 (230 ml, conc. 0.1 M) was added Grubbs II generation catalyst (100 mg, 0.12 mmol), after which the reaction was heated to 50 o C for two hours. The solvent was removed in vacuo, and the crude product was purified by flash chromatography on silica gel (60% EtAC in hexanes) to give 11 S9

10 as a colorless liquid (720 mg, 91%). R f : 0.3 (60% EtAc in hexanes); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 3425 (br), 2935, 2890, 1751, 1453, 1429, 1303, 1166, 1025, 745 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 5H), 6.01 (dd, J = 10.5, 6.0 Hz, 1H), 5.60 (d, J = 10.5 Hz, 1H), 5.02 (d, J = 6.9 Hz, 1H), 4.88 (d, J = 6.9 Hz, 1H), 4.76 (d, J = 11.8 Hz, 1H), 4.67 (d, J = 11.8 Hz, 1H), 4.35 (m, 1H), 3.97 (dd, J = 13.3, 6.7 Hz, 1H), (m, 2H), (m, 2H), 2.33 (m, 1H), 2.12 (m, 1H), 1.31 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 155.2, 136.8, 128.2, 127.7, 127.6, 127.4, 122.8, 95.6, 84.0, 83.4, 70.2, 61.0, 56.5, 37.4, 23.3, 18.4; HRMS (ESIMS): calcd for C 18 H 23 N 5 Na: [M+Na] +, ; found: Amino diol: N BM 2N KH, EtH (1:1) Me reflux, 91% (BRSM) H BM NH H Me H 11 To a solution of carbamate 11 (375 mg, 1.13 mmol) in EtH (4 ml) was added aqueous solution of 2 N KH (4 ml), and the reaction mixture was refluxed for 12 h. The reaction mixture was then extracted three times with CH 2 Cl 2, washed the combined organic layers with H 2 and brine. The crude product was purified by flash chromatography on silica gel (10% MeH/ CH 2 Cl 2 ) to give the amino alcohol as a colorless liquid (250 mg, 91% BRSM). R f : 0.4 (10% MeH in CH 2 Cl 2 ); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 3318 (br), 3033, 2917, 1454, 1380, 1102, 1025, 738, 698 cm - 1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 5H), 5.99 (m, 1H), 5.76 (d, J = 10.6 Hz, 1H), 4.96 (d, J = 6.9 Hz, 1H), 4.89 (d, J = 6.9 Hz, 1H), 4.68 (dd, J = 17.4, 11.8 Hz, 2H), 4.12 (m, 1H), (m, 3H), 3.54 (m, 1H), 3.01 (m, 1H), 2.57 (m, 1H), 2.15 (m, 1H), 1.37 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 136.8, 128.1, 127.6, 127.5, 127.1, 125.1, 95.3, 83.0, 75.1, 70.1, 60.8, 59.1, 41.9, 24.4, 20.3; HRMS (ESIMS): calcd for C 17 H 26 N 4 : [M+H] +, ; found: S10

11 Fmoc-alcohol 12: BM NH H Me H Fmoc-Cl, THF: satd. Na 2 C 3 0 o C, 3 h, 89% NFmoc H Me H BM 12 To a solution of diol (250 mg, mmol) in THF (6 ml) and saturated aqueous Na 2 C 3 (2 ml) was added Fmoc-Cl (231 mg mmol) at 0 o C. The reaction mixture was stirred at the same temperature until there was total conversion of the starting material. Then, the reaction was quenched with a saturated solution of NH 4 Cl and extraction with EtAc (three times). The crude product was purified by flash chromatography on silica gel (40% EtAC in hexanes) to give 12 as a colorless syrup (383 mg, 89%). R f : 0.35 (40% EtAC in hexanes); [α] D o (c 1.05, CHCl 3 ); IR (neat): υ max 3348 (br), 2917, 1676, 1649, 1450, 1426, 1319, 1252, 1200, 1106, 1025, 988, 740, 698 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ 7.80 (d, J = 7.4 Hz, 2H), 7.61 (m, 2H), (m, 9H), 6.01 (m, 1H), 5.79 (d, J = 9.8 Hz, 1H), 5.03 (d, J = 6.8 Hz, 1H), 4.92 (d, J = 6.8 Hz, 1H), 4.82 (s, 1H, br), 4.72 (dd, J = 20.0, 11.7 Hz, 2H ), 4.50 (m, 2H), 4.30 (m, 1H), 4.10 (m, 1H), 3.98 (m, 1H), 3.82 (m, 2H), 3.64 (m, 2H), 3.41 (m, 1H), 2.05 (m, 2H), 1.28 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 157.4, 143.7, 143.4, 141.0, 136.8, 128.2, 128.0, 127.6, 127.5, 126.8, 124.6, 123.3, 119.7, 95.1, 83.3, 77.8, 70.2, 67.5, 61.9, 56.1, 53.1, 46.9, 39.3, 24.2, 20.4; HRMS (ESIMS): calcd for C 32 H 36 N 6 : [M+H] +, ; found: Epoxy-alcohol 13: NFmoc NFmoc H CF 3 CCH 3, xone H Me CH 3 CN: H 2, 0 o C, Me H 3h, 83% H BM BM To a solution of Fmoc-alkene 12 (340 mg, 0.64 mmol) in CH 3 CN (6.5 ml) and aqueous M Na 2 EDTA solution (2.5 ml) was added CF 3 CCH 3 (140 µl, 1.45 mmol), NaHC 3 (809 mg, 9.64 mmol), oxone (790 mg, 1.28 mmol) sequentially at 0 o C. The reaction mixture was stirred for three hours at same temperature until there was total S11

12 conversion of the starting material. Then, the reaction mixture was diluted with water and extracted with EtAc (three times). The crude product was purified by flash chromatography on silica gel (50% EtAc/hexanes) to give the epoxide 13 as a colorless liquid (288 mg, 83%). R f : 0.35 (50% EtAc in hexanes); [α] D o (c 1.31, CHCl 3 ); IR (neat): υ max 3438 (br), 2918, 1694, 1451, 1423, 1319, 1219, 1104, 1025, 906, 740, 698 cm -1 ; 1 H NMR (400 MHz, CDCl 3 ) δ (m, 3H), (m, 2H), (m, 8H), 4.96 (dd, J = 14.6, 6.8 Hz, 2H), (m, 2H), 4.68 (m, 1H, br), 4.58 (m, 1H), (m, 2H), 4.28 (t, J = 6.5 Hz, 1H), 3.94 (m, 1H), (m, 2H), 3.61 (m, 1H), (m, 2H), 3.21 (m, 1H), (m, 2H), 1.35 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 157.2, 143.7, 141.0, 136.4, 128.2, 129.8, 126.9, 126.7, 124.8, 124.6, 119.5, 96.6, 87.9, 75.3, 70.3, 66.8, 62.2, 57.4, 50.6, 48.3, 46.5, 36.2, 23.6, 21.5; HRMS (ESIMS): calcd for C 32 H 35 N 7 Na: [M+Na] +, ; found: Propargyl alcohol 15: NFmoc H Me H BM Dess-Martin periodinane DCM, rt, quant. 2. MgBr THF, -78 o C to 0 o C, 72% BM NFmoc H Me R 1 R 2 Dess-Martin periodinane DCM, rt, quant. 15: R 1 = H 16: R 2 = H 17: R 1, R 2 = Me NaBH 4, MeH, -78 o C, 80% To a solution of diol (140 mg, mmol) in CH 2 Cl 2 (3 ml) was added DMP (163 mg, 0.39 mmol) at rt. and stirred for one hour. The reaction was quenched with a saturated Na 2 S 2 3 :NaHC 3 (7:1) mixture, stirred the reaction mixture till the solution becomes clear (~15-20 min.), then extracted three times with EtAc, washed with water, brine, dried (Na 2 S 4 ) and concentrated in vacuo. The crude aldehyde was then passed through a short pad of silicagel and engaged in the next transformation. To a stirred solution of the crude aldehyde in dry THF (5 ml) was cooled to -78 o C, added 1-propynyl magnesiumbromide (2.05 ml, 1.02 mmol, 0.5 M sol. in THF) and stirred for two hours. S12

13 The reaction was quenched at 0 o C with a saturated aqueous solution of NH 4 Cl, and extracted with EtAc (2 times). The combined organic layers were then washed with brine and dried over Na 2 S 4 and concentrated under reduced pressure. Flash column chromatography using ethyl acetate-hexane (30-40% EtAc/hexanes) afforded a 3:2 diastereomeric mixture of propargylic alcohols (107 mg, 72%) as colorless syrup. The partially separated other diasteromer, 16 (50 mg, mmol) was then subjected to oxidation using Dess-Martin periodinane (54 mg, 0.12 mmol) as described before and the crude ketone thus obtained was dissolved in dry MeH (3 ml) and cooled to -78 o C, added NaBH 4 (13.6 mg, 0.36 mmol) and stirred for 2 h. The reaction was quenched at 0 o C with water, and extracted with EtAc (2 times). The combined organic layers were then washed with brine and dried over anh. Na 2 S 4 and concentrated under reduced pressure. Flash column chromatography using ethyl acetate-hexane (30-40% EtAc/hexanes) afforded the propargylic alcohol 15 (40 mg, 80%) as a colorless syrup. R f : 0.4 (40% EtAC in hexanes); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 3416 (br), 2958, 2917, 2849, 2217, 1737, 1703, 1451, 1373, 1250, 1106, 1044, 758, 742 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 3H), (m, 2H), (m, 8H), 5.00 (d, J = 6.6 Hz, 1H), 4.95 (d, J = 6.6 Hz, 1H), 4.77 (m, 1H), 4.69 (m, 1H), 4.61 (m, 1H), 4.60 (m, 1H), 4.39 (m, 1H), 4.31 (m, 1H), (m, 2H), 3.56 (m, 1H), 3.46 (m, 1H), (m, 2H), (m, 2H), 1.71 (s, 3H), 1.39 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 157.6, 143.6, 140.9, 128.2, 127.7, 127.6, 127.5, 126.8, 126.7, 124.6, 119.6, 97.0, 87.1, 81.6, 75.6, 70.6, 67.7, 62.6, 56.2, 50.7, 49.8, 46.7, 36.3, 29.3, 24.3, 23.0; HRMS (ESIMS): calcd for C 35 H 37 N 7 Na: [M+Na] +, ; found: Indolizidine alkyne 19: 1. Et NFmoc 3 N, MsCl, H Me H H CH 2 Cl 2, 0 o C to rt Me BM 2. Piperidine, CH H 3 CN N BM rt, 5 h, quant. (2 Steps) Me Me To a solution of propargylic alcohol 15 (34 mg, mmol) in CH 2 Cl 2 (2 ml) was added TEA (16.5 μl mmol) followed by MsCl (5 μl, mmol) at 0 o C. The S13

14 reaction mixture was stirred for one hour at rt until there was total conversion of the starting material. Then, the reaction was quenched with a saturated aqueous solution of NH 4 Cl and extracted with EtAc (2 times). The combined organic phases were washed with water, brine, dried (Na 2 S 4 ) and concentrated in vacuo. The crude mesylate obtained was dissolved in CH 3 CN (2 ml) and piperidine (0.4 ml) was added at rt, stirred the reaction mixture for 12 h. The volatiles in the reaction mixture were evaporated to dryness on a rotavapor and the crude mixture was purified by flash chromatography on silica gel (30% EtAc in hexanes) to give the bicyclic alkyne 19 as a colorless syrup (19.5 mg, quantitative). R f : 0.35 (30% EtAc in hexanes); [α] D o (c 1.0, CHCl 3 ); IR (neat): υ max 2919, 1454, 1361, 1170, 1105, 1041, 821, 737, 698 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 5H), 4.91 (d, J = 6.7 Hz, 1H), 4.83 (d, J = 6.7 Hz, 1H), 4.73 (d, J = 11.7 Hz, 1H), 4.62 (d, J = 11.7 Hz, 1H), 3.97 (d, J = 4.7 Hz, 1H), 3.33 (d, J = 4.1 Hz, 1H), 3.28 (m, 1H), 2.95 (m, 1H), 2.88 (m, 1H), 2.76 (m, 1H), 2.22 (s, 1H), 2.16 (m, 1H), 2.08 (m, 1H), 1.79 (d, J = 2.2 Hz, 3H), 1.37 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 138.0, 128.8, 128.4, 128.1, 94.5, 89.9, 81.5, 78.8, 70.1, 69.1, 60.6, 51.8, 51.5, 42.5, 30.1, 23.1, 17.6, 4.1; HRMS (ESIMS): calcd for C 20 H 26 N 4 : [M+H] +, ; found: E- Vinyl stannane: H Me H BM N 19 Me PdCl 2 (PPh 3 ) 2, Bu 3 SnH THF, rt, 30 min, 73% H Me H BM N Me SnBu 3 To a solution of alkyne 19 (18 mg, mmol) in THF (2 ml) was added PdCl 2 (PPh 3 ) 2 (3.6 mg, 0,0052 mmol), followed by slow addition of Bu 3 SnH (69 μl, 0.26 mmol) at rt over 20 min. After 30 min, there was total conversion of the starting material. Then, the volatiles in the reaction mixture were evaporated to dryness leaving the crude stannane on a rotavapor. The crude reaction mixture was purified by flash chromatography on silica gel (20% EtAc in hexanes) to give the E-vinyl stannane as a colorless syrup (24 mg, 73%). R f : 0.4 (20% EtAc in hexanes); [α] D o (c 0.5, CHCl 3 ); IR (neat): υ max 2956, 2923, 2853, 1462, 1376, 1048, 696 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ S14

15 (m, 5H), 5.43 (d, J = 9.2 Hz, 1H), 4.76 (m, 2H), 4.60 (m, 2H), 3.68 (d, J = 6.4 Hz, 1H), 3.35 (d, J = 3.9 Hz, 1H), 3.27 (s, 1H), 3.06 (m, 1H), 2.92 (s, 1H), 2.62 (m, 2H), 2.32 (s, 1H), 2.18 (m, 1H), 2.01 (m, 2H), 1.91 (s, 3H), 1.30 (s, 3H), (m, 21H), (m, 9H); 13 C NMR (CDCl 3, 100 MHz) δ 144.9, 139.6, 138.0, 128.8, 128.2, 128.1, 94.3, 89.9, 78.7, 69.9, 69.5, 65.4, 52.1, 51.6, 42.0, 29.6, 27.5, 20.1, 17.9, 14.1, 9.5; HRMS (ESIMS): calcd for C 32 H 54 N Sn: [M+H] +, ; found: E-Vinyl iodide 20: H Me H H Me H I 2 BM BM N CH 2 Cl 2, 0 o C to rt N Me quant. Me 20 SnBu 3 I To a solution of stannane (18 mg, mmol) in CH 2 Cl 2 (2 ml) was added Iodine (3.8 mg, 0.03 mmol) at 0 o C. The reaction mixture was stirred for 30 min at same temperature until there was total conversion of the starting material. Then, reaction was quenched with saturated aqueous solution of Na 2 S 2 3 and extracted with EtAc (2 times). The combined organic phases were washed with water, brine, dried (Na 2 S 4 ) and concentrated in vacuo. The crude product was purified by flash chromatography on silica gel (25% EtAc in hexanes) to give the vinyl iodide 20 as a colorless syrup (13.1 mg, quantative). R f : 0.3 (25% EtAC in hexanes); [α] D o (c 0.5, CHCl 3 ); IR (neat): υ max 2923, 2852, 1456, 1049 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (5H, m), 6.11 (dd, J = 12.5, 1.4 Hz, 1H), 4.79 (m, 2H), 4.60 (dd, J = 14.3, 6.2 Hz, 2H), 3.71 (d, J = 4.1 Hz, 1H), 3.33 (d, J = 4.1 Hz, 1H), 3.27 (m, 1H), 2.92 (s, 1H), 2.73 (s, 1H), 2.60 (m, 1H), 2.42 (d, J = 1.4 Hz, 3H), 2.31 (s, 1H), 2.13 (m, 1H), 2.04 (m, 2H), 1.28 (s, 3H); 13 C NMR (CDCl 3, 100 MHz) δ 140.4, 137.7, 129.0, 128.4, 128.3, 99.2, 94.7, 89.3, 78.6, 70.1, 69.4, 68.3, 51.8, 51.5, 42.1, 28.5, 26.2, 17.7; HRMS (ESIMS): calcd for C 20 H 27 IN 4 : [M+H] +, ; found: S15

16 BM cyclizidine 21: H Me H N BM Me B Pd[PPh 3 ] 4, Tl 2 C 3, rt, 3 h 81% 20 I 21 H Me H N BM Me To a solution of vinyl iodide 20 (13 mg, mmol) in THF: H 2 (4:1, 2 ml) at rt was added cyclopropyl vinyl boronpinacol ester (6.9 μl, mmol), followed by the addition of Pd[PPh 3 ] 4 (3.2 mg, 10mol%) and Tl 2 C 3 (25.8 mg, mmol). The reaction mixture was stirred for 3 h at rt until there was total conversion of the starting material. Then, the reaction mixture was diluted with EtAc and washed sequentially with water and brine, dried over Na 2 S 4, concentrated in vacuo. The crude product was purified by flash chromatography on silica gel (30% EtAc in hexanes) to give the BM-cyclizidine 21 as a white hygroscopic solid (9.2 mg, 81%). R f : 0.3 (30% EtAc in hexanes); [α] D o (c 0.35, MeH); IR (neat): υ max 2938, 2844, 1544, 1029 cm -1 ; 1 H NMR (CDCl 3, 400 MHz) δ (m, 5H), 6.21 (d, J = 15.6 Hz, 1H), 5.27 (d, J = 10.1 Hz, 1H), 5.18 (dd, J = 15.7, 8.9 Hz, 1H), 4.76 (dd, J = 10.1, 6.7 Hz, 2H), 4.60 (dd, J = 14.3, 6.2 Hz, 2H), 3.71 (d, J = 4.5 Hz, 1H), 3.35 (d, J = 4.0 Hz, 1H), 3.26 (m, 1H), 2.92 (dd, J = 9.9, 4.8 Hz, 1H), 2.58 (m, 1H), 2.31 (s, 1H), 2.07 (m, 1H), 1.99 (m, 2H), 1.73 (s, 3H), 1.47 (m, 1H), 1.39 (s, 3H), 0.77 (m, 2H), 0.43 (m, 2H); 13 C NMR (CDCl 3, 100 MHz) δ 137.2, 137.1, 133.7, 131.5, 129.0, 128.0, 127.7, 127.3, 93.7, 89.4, 77.9, 69.2, 69.0, 66.2, 51.2, 50.8, 41.3, 29.8, 25.5, 22.3, 17.0, 13.9, 6.9; HRMS (ESIMS): calcd for C 25 H 34 N 4 : [M+H] +, ; found: Cyclizidine: H Me H BM N Me LiDBB (0.5 M in THF) THF, -78 o C, 1 h 79% H Me H H N Me 21 cyclizidine S16

17 A solution of BM-cyclizidine 21 (9 mg, mmol) in THF (2 ml) was treated with freshly prepared LiDBB (0.5 M, 1.3 ml) at -78 o C. The reaction mixture was stirred for 1 h, then quenched with H 2. The aqueous layer was extracted three times with CH 2 Cl 2 and the combined organic layers were dried over Na 2 S 4 and concentrated under reduced pressure. The crude product was purified by flash chromatography on silica gel (60% EtAc/hexanes) to give cyclizidine, as colorless crystals. (5 mg, 79%). R f : 0.4 (EtAc); [α] D o (c 0.5, MeH); IR (neat): υ max 3390 (br), 2921, 2850, 1462, 1069, 955,822 cm -1 ; 1 H NMR (CDCl 3, 700 MHz) δ 6.22 (d, J = 15.6 Hz, 1H), (m, 2H), 3.63 (d, J = 4.2 Hz, 1H), 3.34 (d, J = 4.0 Hz, 1H), 3.27 (d, J = 4.0 Hz, 1H), 2.84 (m, 1H), 2.61 (m, 1H), 2.35 (s, 1H) 2.11 (m, 1H), (m, 2H), 1.81 (d, J = 1.0 Hz, 3H), 1.49 (m, 1H), 1.38 (s, 3H), (m, 2H), (m, 2H); 13 C NMR (CDCl 3, 100 MHz) δ 137.8, 133.9, 131.3, 86.1, 78.0, 69.0, 68.0, 51.5, 51.1, 41.8, 25.8, 16.9, 14.2, 13.1, 7.3; HRMS (ESIMS): calcd for C 17 H 26 N 3 : [M+H] +, ; found: We have also recorded the optical rotation of the natural cyclizidine, sample provided by Professor Leeper in the same concentration, as measured for the synthetic cyclizidine. Natural cyclizidine (recorded by us) [α] D o (c 0.5, MeH); Lit. value 2 : [α] D o (c 2.0, MeH). Explanation for the absolute configuration of an optically active compound by X- ray analysis: The determination of absolute configuration by crystallographic methods is based on the anomalous dispersion of X-rays by one or more atoms in the structure. 3 This effect is enhanced by X-ray wavelengths that are not at or below the absorption edge of atoms. Furthermore, the higher the absorption for an atom, the more the anomalous scattering of X-rays is important. These phenomena will make differences in path lengths and intensities of reflections in Friedel 4 pairs in the presence of crystal structures from optically pure substances. The accurate measurement of small differences (Bijvoet 5 anomalies) in Friedel pairs leads to the determination of a Flack 6,7 factor, elaborated in 1983, that tells us the absolute stereochemistry. With modern systems and techniques using copper radiation souce, it is possible to assign the absolute configuration of even S17

18 light atom-containing structures, but the molecule should contain at least an oxygen atom. 8 The relative stereochemistry of natural cyclizidine, reported from the isolation in 1982, was determined by molebdenum radiation which is not suitable for absolute configuration determination of light atom-containing structures because of low absorption of the wavelengths for light atoms. 2 Modern copper radiation systems and techniques enable us to measure with better precision, the small differences in Friedel pairs involved in light atom-containing structures and thus determine the absolute configuration of natural cyclizidine. S18

19 rtep Diagrams: (a) Synthetic, (+)-cyclizidine: RTEP view of synthetic, (+)-cyclizidine. Ellipsoids drawn at 30% probability level. Hydrogen atoms are represented by sphere of arbitrary size. S19

20 (b) Natural, (-)-cyclizidine: RTEP view of natural, (-)-cyclizidine. Ellipsoids drawn at 30% probability level. Hydrogen atoms are represented by sphere of arbitrary size. S20

21 References: 1. For the preparation of methyl ketone 3, see: Ageno, G.; Banfi, L.; Cascio, G.; Guanti, G.; Manghisi, E.; Riva, R.; Rocca, V. Tetrahedron, 1995, 51, Freer, A. A.; Gardner, D.; Greatbanks, D.; Poyser, P.; Sim, G. A. J. Chem. Soc. Chem. Commun. 1982, Glusker, J. P.; Lewis, M.; Rossi, M. Crystal Structure Analysis for Chemists and Biologists VCH Publishers, Inc: New York, USA 1994; Chapter 14, p Friedel, G. Comptes Rendus, Acad. Sci. Paris 1913, 157, Bijvoet, J. M.; Peerdeman, A. F.; van Bommel, A. J. Nature, 1951, 168, Flack, H. D. Acta. Cryst. A 1983, 39, Flack, H. D.; Schwarzenbach, D. Acta. Cryst. A 1988, 44, For examples see: (a) Fun, H.-K.; Quah, C. K.; Chantrapromma, S. Acta. Cryst. E 2010, 66, (b) Fun, H.-K.; Yeap, C. S.; Chantrapromma, S. Acta. Cryst. E 2010, 66, (c) Fun, H.-K.; Joycharat, N.; Voravuthikunchai. S. P.; Chantrapromma, S. Acta. Cryst. E 2010, 66, S21

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