Supporting Information. Copyright Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, 2008

Supporting Information Copyright Wiley-VC Verlag Gmb & Co. KGaA, 69451 Weinheim, 2008

The Guadinomines, Potent Inhibitors of Type-III Secretion Systems, Total Synthesis and Determination of the Absolute Configuration of Guadinomines B and C 2 Tomoyasu irose, Toshiaki Sunazuka,* Satoshi Tsuchiya, Toshiaki Tanaka, Yasuhiro Kojima, Ryuma Mori, Masato Iwatsuki, and Satoshi Ōmura* [a] Kitasato Institute for Life Science, Kitasato University 5-9-1 Shirokane, Minato-ku, Tokyo, 108-8641 (Japan)

Supplementary Figures 2 6 Guadinomine C 2 (4) 5 J value = 4.2 z for 1 -MR..E. (2.6%) This E and coupling constant could not indicate the relative configuration between C5 and C6. R 1 R 2..E.did not observe between - and -6. 5 6 C..E. (4.0%) R 1 = R 2 = 2 Figure S1. E observation in guadinomine C 2 (4). Me 5 1''' 6 0.7% of..e. L-Ala-L-Val-t-Bu 0.6% of..e. Figure S2. ESY correlation and E observations in 5,6-trans-piperazinone 2. ESY 6.6% of..e. Et Me Figure S. ESY correlation and E observation in 5,6-cis-piperazinone 28. 2

''' 2''' 1''' 5-C 6 5 2 -Me Guadinomine C 2 (4) Figure S4. Comparison of 100 Mz 1 C-MR (in 1% TFA/D 2 ) spectrum between 4, 8 and 1. The X- and Y-axes present position number of 4 and δ, respectivily. 1.% of..e. Bn Me Me (+)-259.0% of..e Figure S5. E observations in anti-diol derivative 8.

from 40 TBS Bu 2 Sn toluene, reflux then PMBCl TBAI, reflux 59% PMB 52 TBS (R)-(+) or (S)-( )-MTPA DCC, DMAP, C 2 Cl 2, r.t. PMB R TBS 5 : R = (R)-(+)-MTPA (94%) 54 : R = (S)-( )-MTPA (62%) +0.05 +0.07 Me 0.1 +0.02 +0.0 0.12 TBS (S) +0.09 0.07 +0.11 +0.08 +0.08 0.09 0.08!" = " S " R!" <0 MTPA C!" >0 Scheme S1. Conformation of the newly introduced hydroxy group by the modified Mosher s method for a derivative of 40. EtS PMB 42 TBS TiCl 4, C 2 Cl 2 20 C 82% EtS 55 TBS AgTf, DIPEA C 2 Cl 2, 0 C 80% 56 TBS 0.7% 2.8% 2.0% R 1.4% 56..E. Scheme S2. Derivatization from 42 to γ-lactone compound and E observations. 4

Experimental Section Contents Abbreviations 6 Experimental Procedures and Compounds Characterization 7 For synthesis of piperazinone model compounds 7 For synthesis of syn-4 10 For Scheme S1 11 For Scheme S2 14 (S,5S,6S,2 S, R,4 R)-guadinomine C 2 ( R)-4 16 Synthesis of (S,5S,6S,2 S, S,4 R)-guadinomine C 2 ( S)-4 20 Synthesis of (2S,S, 6R,7S,4 R)-guadinomine B 2 5 5

Abbreviations Ac = Acetyl, Bn = Benzyl, Bu = Butyl, Boc = tert-butyloxycarbonyl, CA = Cerium ammonium nitrate, Cbz = Benzyloxycarbonyl, DIBAL = Diisobutylaluminum hydride, DIPEA =, -Diisopropylethylamine, DMAP = 4-(Dimethylamino)pyridine, DMF =, -Dimethylformamide, DMS = Dimethyl sulfoxide, EDCI = 1-Ethyl--(-dimethylaminopropyl)carbodiimide, Bt = -ydroxybenzotriazole, Ms = Methanesulfonyl, MTPA = a-methoxy-a-(trifluoromethyl)phenylacetic acid, s = o-itrobenzenesulfonyl, p-s = p-itrobenzenesulfonyl, PMB = p-methoxybenzyl, PMP = p-methoxyphenyl, PPTS = Pyridinium p-touenesulfonate, PyBP = Benzotriazol-1-yloxytrispyrrolidinophosphonium hexafluorophosphite, TBAF = Tetrabutylammonium fluoride, TBDPS = tert-butyldiphenylsilyl, TBS = tert-butyldimethylsilyl, TEA = Triethylamine, Tf = Trifluoromethanesulfonyl, TFA = Trifluoroacetic acid, TF = Tetrahydrofuran. 6

Experimental Procedures and Compounds Characterization <For synthesis of piperazinone model compounds> (+)-tert-butyl-( -benzyloxycarbonyl-l-alanyl)-l-valinate Cbz 2 t-bu EDCI, Bt DIPEA, C 2 Cl 2 98% Cbz t-bu To a solution of tert-butyl-l-valinate (4.50 g, 21.5 mmol) in C 2 Cl 2 (215 ml) was added -benzyloxycarbonyl-l-alanine (5.80 g, 25.8 mmol), DIPEA (.7 ml, 21.5 mmol), Bt (5.80 g, 42.9 mmol), EDCI Cl (6.20 g, 2.2 mmol) at rt. After stirring for 70 min, the reaction mixture was added sat. 4 Cl aq. (15 ml). The organic layer was separated and water layer was extracted with CCl (50 ml x 2). The combined organic extracts were washed with 2 (100 ml), brine (100 ml) and dried over a 2 S 4, filtered, and evaporated under reduced pressure. Purification by flash chromatography on silica gel (hexane : AcEt = : 1) provided the title compound (8.00 g, 98%) as a colorless oil. 27 Rf = 0.4 (Silica gel, hexane : AcEt = 1 : 1); [α] D.57 (c 1.96, CCl ); IR (KBr) ν cm 1 52 (), 170 (C=, ester), 1664 (C=, amide), 157 ; 1 -MR (270 Mz, CDCl ) δ: 7.55-7.20 (complex m, 5, 2 --C-C 2 Ph), 6.6 (m, 1, 2 --C-C 2 Ph), 5.29 (m, 1, 2-), 5.12 (s, 2, 2 --C-C 2 Ph), 4.41 (dd, J = 8.6, 4.6 z, 1, 2-), 4.2-4.17 (br-m, 1, 1 -), 2.22-2.07 (br-m, 1, -), 1.47 (s, 9, 1-C(C ) ), 1.40 (d, J = 6.9 z,, - ), 0.90 (d, J = 6.6 z,, -C ), 0.87 (d, J = 6.9 z,, -C ); 1 C-MR (67.5 Mz, CDCl ) δ: 172.4, 170.6, 155.8, 16.2, 128.0 (2C), 127.5 (2C), 127.4, 81.5, 66.5, 57., 50.2, 1.0, 27.7 (C), 18.6, 17., 18.5; R-MS (FAB, BA matrix) m/z 79.2217 [M+] +, Calcd for C 20 1 2 5 79.22 [M+]. (+)-tert-butyl-l-alanyl-l-valinate; 19 Cbz t-bu 2, Pd/C Et 78% 2 19 t-bu To a solution of Cbz-peptide (8.00 g, 21.2 mmol) in Et (106 ml) was added 10 7

wt % Pd / C (2.26 g, 2.12 mmol) at RT. After stirring for 4.5 hr under 2, the reaction solution was filtrated with celite pad to remove the Pd catalyst, then the pad were washed with AcEt. The filtrate solution was evaporated to remove the solvent. Purification by flash chromatography on silica gel (CCl : Me = 100 : 1) provided the title compound 19 (4.00 g, 78%) as a colorless oil. Rf = 0.1 (Silica gel, CCl : Me = 8 : 1); [α] 29 D +12.6 (c 1.1, CCl ); IR (KBr) ν cm 1 60 (), 172, 1666 (C=); 1 -MR (270 Mz, CDCl ) δ: 7.72 (d, J = 7.9 z, 1, 2-), 4.41 (dd, J = 4.6, 9.2 z, 1, 2-),.59 (m, 1, 2 -), 2.18 (m, 1, -), 2.0-1.96 (br-m, 2, 2-2 ), 1.46 (s, 9, 1-C(C ) ), 1.6 (d, J = 6.9 z,, - ), 0.94 (d, J = 6.6 z,, -C ), 0.89 (d, J = 6.9 z,, -C ); 1 C-MR (67.5 Mz, CDCl ) δ: 175.1, 171.1, 81.8, 56.9, 50.8, 1.2, 28.7 (C), 21.6, 18.6, 17.; R-MS (FAB, BA matrix) m/z 245.1866 [M+] +, Calcd for C 12 25 2 245.1865 [M+]. tert-butyl-( -benzyloxycarbonyl-d-alanyl)-d-valinate Cbz 2 t-bu EDCI, Bt DIPEA, C 2 Cl 2 100% Cbz t-bu According to the procedure of the compound Cbz-L-peptide, the title compound (1.81 g, 100%) was obtained from tert-butyl-d-valinate (1.00 g, 4.77 mmol) and -benzyloxycarbonyl-d-alanine (1.28 g, 25.8 mmol). All analitycal data of Cbz-D-peptide are identical with Cbz-L-peptide, except for [α] D, R-MS. 2 [α] D +.55 (c 1.00, CCl ); R-MS (FAB, BA matrix) m/z 79.227 [M+] +, Calcd for C 20 1 2 5 79.22 [M+]. tert-butyl-d-alanyl-d-valinate Cbz t-bu 2, Pd/C Et 100% 2 t-bu According to the procedure of the compound 19, the title compound (1.20 g, 100%) was obtained from Cbz-D-peptide (1.80 g, 4.76 mmol). All analitycal data of D-peptide are identical with 19, except for [α] D, R-MS. 8

[α] D 2 5.45 (c 1.00, CCl ); R-MS (FAB, BA matrix) m/z 245.1868 [M+] +, Calcd for C 12 25 2 245.1865 [M+]. 9

<For synthesis of syn-4> (S)-(+)-5-(tert-Butyldimethylsiloxy)-2-(p-methoxybenzyloxy)-1-pentanol PMP (S) TBS DIBAL, C 2 Cl 2 78 C 9% PMB (S) >99% ee TBS Analytical PLC (normal phase) DAICEL CIRALCEL D 4.6 φ x 250 mm, hexane : 2-propanol = 90 : 10, 0.9 ml/min, temp. = 25 C, 226 nm Daicel CIRALCEL D 4.6φ X 250mm exane:2-propanol=90:10, 0.9 ml/min, t = 25 C, 226 nm PMB TBS (S) PMB TBS (R) 10

(S,4S)-(+)-S-Ethyl-7-tert-butyldimethylsiloxy--hydroxy-4-(p-methoxybenzyloxy)- thioheptanate; 42 PMB (S) TBS (CCl) 2, DMS, TEA C 2 Cl 2, 78 C then 0 C C PMB TBS 41 EtS TMS TiCl (i-pr), C 2 Cl 2, 78 C 64% in 2 steps PMB EtS TBS (S) (S) 42 >99% de Analytical PLC (normal phase) DAICEL CIRALCEL D 4.6 φ x 250 mm, hexane : 2-propanol = 96 : 4, 0.8 ml/min, temp. = 25 C, 226 nm Daicel CIRALCEL D 4.6φ X 250mm exane:2-propanol=96:4, 0.8 ml/min, t = 25 C, 226 nm PMB EtS (S) (S) 42 TBS PMB EtS (R) (S) TBS Scheme S1 (2S)-( )-5-(tert-butyldimethylsiloxy)-1-(p-methoxyphenyloxy)-2-pentanol; 52 TBS Bu 2 Sn, toluene, reflux then PMBCl, TBAI, toluene 59% PMB TBS (S) from 40 52 To a solution of diol (25.2 mg, 108 µmol) in toluene (1.1 ml) was added Bu 2 Sn (28.1 mg, 11 µmol) at RT, and the mixture was heated to the condition of reflux. After stirring for 15 min, the reaction mixture was cooled to RT, and PMBCl (20.4 µl, 151 11

µmol) and TBAI (59.6 mg, 161 µmol) were added to the mixture. The resulting reaction mixture was stirred for 80 min under reflux condition, then the mixture was poured into 2 (7.0 ml) for quenching at RT. The layers were separated, and the aqueous layer was extracted EtAc (20 ml x ). The combined organic layers were washed with 2 (10 ml), dried over a 2 S 4, filtered and concentrated under reduced pressure. Flash chromatography on silica gel (hexsane : EtAc = 6 : 1) afforded the title compound 52 (22.4 mg, 59%) as a colorless oil. Rf = 0.4 (Silica gel, hexane : AcEt = 2 : 1); [α] D 25 55.9 (c 0.05, CCl ) IR (acl) ν cm 1 424 (-), 1250; 1 -MR (400 Mz, CDCl ) δ : 7.26 (dt, J = 2.0, 8.4 z, 2, 1--C 2 -Ph-C ), 6.88 (dt, J = 2.0, 8.4 z, 2, 1--C 2 -Ph-C ), 4.48 (s, 2, 1--C 2 -Ph-C ),.80 (m, 1, 2-),.80 (s,, 1--C 2 -Ph-C ),.64 (t, J = 5.9 z, 2, 5-2 ),.45 (dd, J = 4.0, 9.4 z, 1, 1-2 ). (dd, J = 7.6, 9.4 z, 1, 1-2 ), 1.64-1.58 (complex m, 2, 4-2 ), 1.58 (m, 1, - 2 ), 1.47 (m, 1, - 2 ), 0.09 (s, 9, 5-Si[C ] 2 C(C ) ), 0.04 (s, 6, 5-Si[C ] 2 C(C ) ); R-MS (FAB, BA matrix) m/z 55.2291 [M+] +, Calcd for C 19 5 4 Si 55.205 [M+]. (2S)-(+)-5-(tert-butyldimethylsiloxy)-1-(p-methoxyphenyloxy)-2-[(R)-a-methoxy- a-trifluoromethyl-phenylacetyl]-2-pentanol; 5 PMB TBS (R)-MTPA,DCC, DMAP C 2 Cl 2, r.t. 94% 52 5 CF Me (R) Ph TBS (S) To a solution of alcohol 52 (10.6 mg, 29.9 µmol) in C 2 Cl 2 (997 µl) was added (R)-MTPA (21.0 mg, 89.7 µmol), DCC (12. mg, 59.8 µmol), DMAP (.70 mg, 0.5 µmol) at RT under Ar. After stirring for 60 min, the reaction solution was concentrated. The residue was dissolved in Et 2 (20 ml), and insoluble salt were filtered off. The filtrate was washed with sat. 4 Cl aq. (5.0 ml), sat. ac aq. (5.0 ml) and 2 (5.0 ml), dried over a 2 S 4, filtered, and evaporated under reduced pressure. Purification by Prep. Silica gel TLC (hexane: AcEt = 2 : 1) provided the title compound 5 (16.0 mg, 94%) as a colorless oil. Rf = 0.74 (Silica gel, hexane : AcEt= 2 : 1); [α] D 25 +27.5 (c 0.80, CCl, ); IR (acl) ν cm 1 1747 (C=, ester); 1 -MR (400 Mz, CDCl ) δ : 7.56 (d, J = 7.1 z, 2, 2--C(CF )(C )Ph), 7.45-7.1 (complex m,, 2--C(CF )(C )Ph), 7.16 12

(dt, J = 1.8, 9.1 z, 2, 1--C 2 -Ph-C ), 6.84 (dt, J = 1.8, 9.1 z, 2, 1--C 2 -Ph-C ), 5.6 (dddd, J = 4.5, 5.5, 6.0, 7.5 z, 1, 2-), 4.40 (d, J = 11.6 z, 1, 1--C 2 Ph-C ), 4.4 (d, J = 11.6 z, 1, 1--C 2 Ph-C ),.80 (s,, 1--C 2 Ph-C ),.61 (apparent dt, J = 1.0, 6.0 z, 2, 5-2 ),.52 (d, J = 1.0 z,, 2--C(CF )(C )Ph),.52 (dd, J = 6.0, 11.0 z, 1, 1-2 ),.49 (dd, J = 4.5, 11.0 z, 1, 1-2 ), 1.77 (m, 1, - 2 ), 1.72 (m, 1, - 2 ), 1.59 (m, 1, 4-2 ), 1.54 (m, 1, 4-2 ), 0.90 (s, 9, 5-Si[C ] 2 C(C ) ), 0.04 (s, 6, 5-Si[C ] 2 C(C ) ) ; 1 C-MR (67.5 Mz, CDCl ) δ : 166.2, 159.2, 12.2, 129.5, 129.9 (2C), 129.2 (2C), 128.2 (2C), 127.6, 127.6, 11.7 (2C), 84.9, 75.4, 72.7, 70.7, 62.4, 55.4, 55.2, 28.4, 27., 25.9 (C), 18., 5.4 (2C); R-MS (FAB, BA matrix) m/z 59.2504 [M+a] +, Calcd for C 29 41 6 F Sia 59.2522 [M+a]. (2S)-( )-5-(tert-butyldimethylsiloxy)-1-(p-methoxyphenyloxy)-2-[(S)-a-methoxy- a-trifluoromethyl-phenylacetyl]-2-pentanol; 54 PMB TBS (S)-MTPA,DCC, DMAP C 2 Cl 2, r.t. 62% Me CF (S) Ph TBS (S) 52 54 According to the procedure of the (R)-MTPA ester 5, the title compound 54 (12.1 mg, 62%) was obtained from 52 (12.1 mg, 4.4 µmol). Rf = 0.74 (Silica gel, hexane : AcEt = 2 : 1); [α] D 25 6.4 (c 0.61, CCl ); IR (acl) ν cm 1 1747 (C=, ester); 1 -MR (400 Mz, CDCl ) δ : 7.59 (d, J = 7.1 z, 2, 2--C(CF )(C )Ph), 7.44-7.28 (complex m,, 2--C(CF )(C )Ph), 7.2 (dt, J = 2.0, 8.9 z, 2, 1--C 2 -Ph-C ), 6.86 (dt, J = 2.0, 8.9 z, 2, 1--C 2 -Ph-C ), 5.9 (1, dddd, J = 5.0, 5.1, 7.5, 11.0 z, 1, 2-), 4.51 (d, J = 11.5 z, 1, 1--C 2 Ph-C ), 4.44 (d, J = 11.5 z, 1, 1--C 2 Ph-C ),.81 (s,, 1--C 2 Ph-C ),.60 (dd, J = 5.1, 11.0 z, 1, 1-2 ),.57 (dd, J = 5.0, 11.0 z, 1, 1-2 ),.56 (d, J = 1.0 z,, 2--C(CF )(C )Ph),.54 (dt, J = 2.2, 6.0 z, 2, 5-2 ), 1.69 (m, 1, - 2 ), 1.6 (m, 1, - 2 ), 1.46 (m, 1, 4-2 ), 1.42 (m, 1, 4-2 ), 0.90 (s, 9, 5-Si[C ] 2 C(C ) ), 0.04 (s, 6, 5-Si[C ] 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 166., 159., 12.5, 129.8 (2C), 129.5 (2C), 128.7, 128.2, 127.4, 11.8 (2C), 84.9, 75.1, 72.7, 70.9, 62.4, 55.4, 55., 28.1, 27.2, 25.9 (C), 18., 5.4 (2C); R-MS (FAB, BA matrix) m/z 59.2504 [M+a] +, Calcd for 1

C 29 41 6 F Sia 59.2522 [M+a]. Scheme S2 (S,4S)-S-Ethyl-7-tert-butyldimethylsiloxy-,4-dihydroxy-thioheptanate; 55 PMB EtS (S) (S) TBS TiCl 4, C 2 Cl 2, 20 C EtS (S) (S) 82% 42 55 TBS To a solution of 42 (22.6 mg, 49.5 µmol) in C 2 Cl 2 (1.65 ml) was dropwised TiCl 4 solution (1.0 M in C 2 Cl 2, 52.0 µl, 52.0 µmol) at 20 C under Ar. After stirring for 5 min, the solution was quenched by addition of sat. aq. ac solution (1.0 ml) and stirred for 1 hr at RT. The mixture was diluted with CCl (5.0 ml) and sat. aq. Rochelle s salt solution (2.0 ml). The organic layer was then separated, and the aqueous layer was extracted with C 2 Cl 2 (7.0 ml x ). The combined organic extracts were washed with sat. aq. Rochelle s salt solution (5.0 ml ), dried over a 2 S 4, filtered, and evaporated under reduced pressure. Purification by flash chromatography on silica gel (exane : EtAc = 2 : 1) provided the title compound 55 (1.7 mg, 82%) as a colorless oil. Rf = 0. (Silica gel, hexane : AcEt = 2 : 1); 1 -MR (270 Mz, CDCl ), major isomer was indicated. δ :.96 (ddd, J = 4.6, 7.1, 7.8 z, 1, -),.74-.56 (complex m, 2, 7-2 ),.48 (m, 1, 4-), 2.91 (s, 2, 1-SC 2 C ), 2.8 (dd, J = 7.8, 15.5 z, 1, 2-2 ), 2.76 (dd, J = 4.6, 15.5 z, 1, 2-2 ), 1.77-1.42 (complex m, 4, 5-2, 6-2 ), 1.26 (t, J = 7. z,, 1-SC 2 C ), 0.90 (s, 9, 7-Si[C ] 2 C(C ) ), 0.07 (s, 6, 7-Si[C ] 2 C(C ) ). (4S,5S)-( )-5-[1 -(tert-butyldimethylsiloxy)-propane- -yl]-4-hydroxy-tetrahydro- furan-2-one; 56 EtS (S) (S) TBS 55 AgTf, DIPEA C 2 Cl 2, RT 80% 2 (S) (S) 5 56 1' TBS To a solution of alcohol 55 (1.7 mg, 40.7 µmol) in TF (1.6 ml) was added DIPEA (17. µl, 102 µmol) and AgTf (11.5 mg, 44.8 µmol) at RT under Ar. After stirring for 10 min, the reaction solution was quenched with sat. ac aq. (2.0 ml) The mixture 14

was diluted with CCl (7.0 ml), and the organic layer was then separated. The aqueous layer was extracted with C 2 Cl 2 (7.0 ml x ). The combined organic extracts were washed with sat. ac aq. (5.0 ml), dried over a 2 S 4, filtered, and evaporated under reduced pressure. Purification by flash chromatography on silica gel (exane : EtAc = 5 : 1) provided the title compound 56 (8.9 mg, 80%) as a colorless oil. Rf = 0.11 (Silica gel, hexane : AcEt = 2 : 1); [α] D 21 10.1 (c 0.50, CCl ); IR (acl) ν cm 1 48 (), 1770 (C=, lactone); 1 -MR (400 Mz, CDCl ) δ : 4.51 (apparent dd, J = 4.5, 5.0 z, 1, 4-), 4.7 (ddd, J = 4.5, 7.0, 8.0 z, 1, 5-),.76-.65 (complex m, 2, - 2 ),.08 (br-s, 1, 4-), 2.77 (dd, J = 5.0, 18.0 z, 1, - 2 ), 2.56 (apparent d, J = 5.0 z, 1, - 2 ), 2.05-1.91 (complex m, 2, - 2 ), 1.79 (m, 1, 2-2 ), 1.55 (m, 1, 2-2 ), 0.90 (s, 9, 1 -Si[C ] 2 C(C ) ), 0.09 (s, 6, 1 -Si[C ] 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 176.0, 84.8, 68.5, 62.7, 9.0, 27.5, 25.9 (C), 2.9, 18., 5.4 (2C); R-MS (FAB, BA matrix) m/z 297.1504 [M+a] +, Calcd for C 1 26 4 Sia 297.1498 [M+a]. 15

< (S, 5S, 6S, 2 S, R, 4 R)-guadimonine C 2 ( R)-4> 1 -MR of atural and Synthetic ('R)-Guadinomine C 2 2 4''R 2'S 'R 6S 2'''S 5S 2''''S S atural ('R)-Synthetic 400Mz 1 -MR, in 1% TFA/D 2 16

1-MR (ppm) (400 Mz)* Positio atural 4 (in D 2 with 1% T FA) synthetic ( R)-4 (in D2 with 1% T FA) n 2 --- --- 4.09 (q, J = 7 z) 4.10 (q, J = 7.2 z) -C 1.5 (q, J = 7 z) 1.58 (q, J = 7.2 z) 5 4.50 (d, J = 4.2 z) 4.52 (d, J = 4.5 z) 5-C --- --- 6.99 (ddd, J = 4.2, 4.2, 11.2 z).99 (ddd, J = 2.5, 4.5, 11.2 z) 1 1.9 (ddd, J = 2.1, 5.6, 14 z) 1.77 (m) 1.92 (ddd, J = 2.1, 6.0, 14.0 z) 1.75 (ddd, J = 5.8, 10.0, 14.0 z) 2.61 (ddd, J = 2.1, 5.6, 9.8 z).61 (ddd, J = 2.1, 5.8, 10.0 z).48 (ddd, J = 2.1, 5.6, 8.4 z).48 (ddd, J =.0, 5.8, 10.0 z) 4 5 1.76 (m) 1.44 (m) 1.71 (m) 1.57 (m) 1.74 (m) 1.4 (m) 1.70 (m) 1.59 (m) 2 --- --- 4 4.27 (m) 4.25 (dddd, J = 5.8, 5.8, 6.0, 8.4 z) 5 4.2 (dd, J = 8.4, 8.4 z).79 (dd, J = 5.6, 8.4 z) 4.21 (dd, J = 8.4, 8.4 z).78 (dd, J = 5.8, 8.4 z) 1 --- --- 2 4.44 (q, J = 7 z) 4.44 (q, J = 7.2 z) 1.9 (d, J = 7 z) 1.8 (d, J = 7.2 z) 1 --- --- 2 4.07 (d, J = 7 z) 4.08 (d, J = 6.0 z) 2.12 (eighted, J = 7 z) 2.12 (dsep, J = 6.0, 6.8 z) -C -C 0.92 (d, J = 7 z) 0.92 (d, J = 7 z) * Reference is 4.76 ppm (D2). 0.92 (d, J = 6.8 z) 0.92 (d, J = 6.8 z) 17

Synthetic ( R)-Guadinomine C2 [( R)-4] TFA 100 Mz 1C-MR (in 1%TFA/D2) TFA We observed some shifts for value of signal (ppm) (1C-MR) between natural 4 and synthetic ( R)-4 as shown in Table on next pages. Therefore, the mixture of natural 4 and synthetic ( R)-4 (1:1) was measured for 1C-MR (in 1%TFA/D2) to be identical as shown in next chart. atural 4 + synthetic ( R)-4 (1 : 1) 100 Mz 1C-MR (in 1%TFA/D2) TFA TFA 18

1C-MR (100 Mz)* Positio n atural 4 (in D2 with 1% T FA) synthetic ( R)-4 (in D2 with 1% T FA) ppm ppm Δδ ppm** 2 172.1 170.8 1. 54.9 54.8 0.1 -C 17.0 16.5 0.5 5 59.8 59.7 0.1 5-C 168. 167.5 0.8 6 5.8 5.4 0.4 1 8.8 8.7 0.1 2 74.2 74.2 0 76.8 76.7 0.1 4 0.1 0.1 0 5 0.4 0.4 0 1 -C 158.9 158.4 0.5 2 158.9 159.0 0.1 4 54.0 54.0 0 5 5.2 5.1 0.1 1 176.6 177.1 0.5 2 52.2 52.1 0.1 19.2 19.2 0 1 179 177.7 1. 2 62.4 61.4 1.0 2.6 2. 0. -C 20.2 20.1 0.1 -C 21.2 20.9 0. * Reference is 0-200 ppm for all compound. ** Δd ppm = ppm(natural 4 synthetic ( R)-4). Although the 1 -MR spectrum of synthetic (R )-4 agreed with natural 4, the 1 C-MR spectra of synthetic vs. natural was different. This may be caused by contamination of phosphoric acid, which was used as additives of preparative PLC solvents for natural 2. Actuality, the data of MR and [α] D of the guadinomines are strongly influenced by this contamination. For these MR spectra, different batch of natural 2 were used. Therefore, the mixture of natural 4 and synthetic ( R)-4 (1:1) was measured for 1 C-MR (in 1%TFA/D 2 ) to be identical as shown in previous pages. 19

<Synthesis of (S, 5S, 6S, 2 S, S, 4 R)-guadimonine C 2 ( S)-4> syn-4 a g TBDPS C 2 Et 2 57 h n C 58 -L-Ala-L-Val-t-Bu Boc t-bu R R (R, R)-45 Co t-bu t-bu t-bu o 2 R 47 Boc Boc p, q -L-Ala-L-Val-t-Bu Boc Boc 60 R S2 invertion I 2, PPh r Boc Boc 61 R 59 R = Boc s u 2 -L-Ala-L-Val- 'S 4"R 2'S 6S 5S S ('S)-4 Reagents and conditions: a) (S)-16, n-bu 2 BTf, Et, C 2 Cl 2, 78 to 0 C, 80 min, 62% (2 steps); b) a, Et, 0 C, 20 min, 98% (as a mixture of trans/cis-epoxide; predominantly trans isomer; 5.8 : 1); c) a, 4 Cl, Et/ 2 (20/1), 60 C, 21 h, 58%; d) PPh, dehydrated MeC, 80 C, 4 h, 81% as a single trans-aziridine isomer; e) p-scl, 4-(dimethylamino)pyridine, Et, C 2 Cl 2, 0 C to RT, 4 h; f) a, DMF, RT, 1 h, 85% (2 steps) (>20:1 = a : b- ); g) PhS, DIPEA, MeC, RT, 6 h, 75%; h) (R)-2-bromopropionic acid R-10, PyBP, DIPEA, C 2 Cl 2, RT, 1 h, 95%; i) PPh, Et, MeC, 1 h; then 2, 60 C, 7 h; j) Boc 2, EtAc, 80 C, 1 h, 79% (2 steps); k) Li, TF/Me/ 2 (2/2/1), RT, 1 h; l) 2 -L-Ala-L-Val-t-Bu 19, PyBP, Bt, DIPEA, C 2 Cl 2, RT, 4 h, 68% (2 steps); m) TBAF, TF, RT, 4 h, 99%; n) S pyridine, DMS, Et, C 2 Cl 2, 0 C to RT, 1 h; o) (R, R)-salen-cobalt complex (R, R)-45, Me 2, DIPEA, C 2 Cl 2, 40 C, 46 h, 7% (2 steps), 90% de; p) C 2 4, 10% Pd/C, Me, RT, 2 h; q) 47, DIPEA, MeC, RT, 1 h, 76% (2 steps); r) I 2, PPh, imidazole, C 2 Cl 2, 0 C to RT, 2 h, 67%; s) PMPisocyanate, Ph, RT, 10 min; t) CA, MeC/ 2 (1/1), 0 C, 2.5 h; u) TFA/ 2 (/1), RT, 5 h 60% ( steps). 20

(2S,R,6S,7S)-Ethyl-9-(tert-butyldiphenylsilyloxy)-2,-epoxy- 6,7--isopropylidenenonanate; major (2R,R,6S,7S)-Ethyl-9-(tert-butyldiphenylsilyloxy)-2,-epoxy- 6,7--isopropylidenenonanate; minor TBDPS Bn Cl a, Et, 0 C 98% TBDPS TBDPS 6 7 trans 2 Et Et 5.8 : cis 1 According to the procedure of the preparation of trans-epoxide (pages 52), the mixture of the title compounds (trans/cis = 5.8/1) (1.46 g, 98%) was obtained from chlorohydrin (2.26 g,.26 mmol). 25 Rf = 0.54 (Silica gel, hexane : AcEt = 2 : 1); [α] D 8.50 (c 1.00, CCl ) as a mixture of trans and cis (5.8 : 1); IR (acl) ν cm 1 175 (C=, ester), 1429, 179, 1242, 1196, 1111, 1090, 104, 941, 82, 79, 704, 61, 505; 1 -MR (270 Mz, CDCl ), major isomer is indicated. δ : 7.70-7.60 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.47-7. (complex m, 6, 9--Si(Ph) 2 C(C ) ), 4.2-4.12 (complex m, 2, 1-C 2 C ),.8 (m, 1, 7-),.82 (dt, J = 1.0, 6.0 z, 2, 9-2 ),.64 (dt, J =.0, 8.2 z, 1, 6-),.2 (d, J = 2.0 z, 1, 2-),.20 (m, 1, -), 1.92-1.61 (complex m, 6, 4-2, 5-2, 8-2 ), 1.6 (s,, 6,7--i-Pr), 1.4 (s,, 6,7--i-Pr), 1.29 (t, J = 7. z,, 1-C 2 C ), 1.06 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ), major isomer is indicated. δ : 169.1, 15.5 (4C), 1.7, 1.6, 129.6 (2C), 127.6 (4C), 108.1, 80., 77.6, 61.4, 60.6, 58.2, 5.2, 5.6, 28.6, 28.5, 27.2 (2C), 26.8 (C), 19.1, 14.1; R-MS (FAB, BA + ai matrix) m/z 549.2671 [M+a] +, Calcd for C 0 42 6 Sia 549.2648 [M+a]. 21

(2R,R,6S,7S)-Ethyl-2-azido-9-(tert-butyldiphenylsilyloxy)--hydroxy-6,7-isopropylidenenonanate; major (2S,S,6S,7S)-Ethyl--azido-9-(tert-butyldiphenylsilyloxy)-2-hydroxy-6,7-isopropylidenenonanate; minor TBDPS 7 6 2 Et TBDPS Et a, 4 Cl Et / 2, 60 C 58% TBDPS Et : 1 mixture of trans and cis (5.8 : 1) and two epimers at C-2 According to the procedure of the azidolysis (pages 5), the title compound (625 mg, 58%; as a mixture of : 1 = α- : β- ) was obtained from epoxide (1.00 g, 1.90 mmol). Rf = 0.50 (Silica gel, hexane : AcEt = 2 : 1); 1 -MR (270 Mz, CD 2 Cl 2 ), major isomer is indicated. δ : 7.72-7.65 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.48-7.4 (complex m, 6, 9--Si(Ph) 2 C(C ) ), 4.9-4.25 (complex m,, -, 1-C 2 C ),.9 (d, J = 9.0 z, 1, 2-),.92-.78 (complex m,, 7-, 8-2 ),.68 (m, 1, 6-), 1.84-1.52 (complex m, 6, 4-2, 5-2, 8-2 ), 1.41 (s,, 6,7--i-Pr), 1.8 (s,, 6,7--i-Pr), 1.5 (t, J = 6.8 z,, 1-C 2 C ), 1.07 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ), major isomer is indicated. δ : 169.4, 15.9 (4C), 14.2, 14.1, 10.1, 10.0, 128.1 (2C), 128.0 (2C), 108.7, 81.1, 78.0, 7.8, 71.9, 66.4, 61.0, 5.9, 0.0, 28., 27.7, 27.5, 27.2 (C), 19.6, 14.6; R-MS (FAB, BA + ai matrix) m/z 592.288 [M+a] +, Calcd for C 0 4 6 Sia 592.2819 [M+a]. 22

(2R,S,6S,7S)-( )-Ethyl-9-(tert-butyldiphenylsilyloxy)-2,-imino- 6,7--isopropylidenenonanate; trans (2S,S,6S,7S)-(+)-Ethyl-9-(tert-butyldiphenylsilyloxy)-2,-imino- 6,7--isopropylidenenonanate; cis TBDPS Et TBDPS Et : 1 PPh, MeC 80 C 81% TBDPS 7 6 trans 2 Et According to the procedure of the aziridine fromation (pages 54), the title compound trans aziridine (682 mg, 81%; as a single isomer) and cis aziridine (91 mg, 11%; as a single isomer) were obtained from azidoalcohol (912 mg, 1.60 mmol). trans aziridine: Rf = 0.29 (Silica gel, hexane : AcEt = 2 : 1); [α] 27 D 44.4 (c 2.00, CCl ); IR (acl) ν cm 1 286 (), 1728 (C=, ester), 1429, 171, 1244, 1211, 1111, 109, 104, 876, 82, 79, 704, 61; 1 -MR (400 Mz, CD 2 Cl 2 ) δ : 7.72-7.67 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.48-7.8 (complex m, 6, 9--Si(Ph) 2 C(C ) ), 4.19 (dq, J =.2, 6.8 z, 2, 1-C 2 C ),.82 (t, J = 7.0 z, 2, 9-2 ),.81 (ddd, J =.4, 5.5, 7.5 z, 1, 7-),.66 (dt, J =.4, 8.0 z, 1, 6-), 2.28 (d, J = 2.1 z, 1, 2-), 2.24 (dt, J = 2.1, 6.0 z, 1, -), 1.84 (m, 1, 8-2 ), 1.72 (m, 1, 8-2 ), 1.72-1.54 (complex m, 2, 5-2 ), 1.64-1.48 (complex m, 2, 4-2 ), 1.5 (s,, 6,7--i-Pr), 1.4 (s,, 6,7--i-Pr), 1.29 (t, J = 6.8 z,, 1-C 2 C ), 1.07 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 172.6, 15.5 (4C), 1.8, 1.7, 129.6 (2C), 127.6 (4C), 108.0, 79.9, 77.5, 61.4, 60.9, 9.0, 5.7, 5.2, 29.6, 29.0, 27.2 (2C), 26.8 (C), 19.1, 14.1; R-MS (FAB, BA matrix) m/z 526.2981 [M+] +, Calcd for C 0 44 5 Si 526.2989 [M+]. cis aziridine: Rf = 0.17 (Silica gel, hexane : AcEt = 2 : 1); [α] 28 D +6.90 (c 1.00, CCl ); IR (acl) ν cm 1 1728 (C=, ester), 179, 1198, 1111,109, 106, 82, 79, 702, 61; 1 -MR (270 Mz, CDCl ) δ : 7.70-7.45 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.4-7.0 (complex m, 6, 9--Si(Ph) 2 C(C ) ), 4.22 (dq, J =., 7. z, 2, 1-C 2 C ),.81 (t, J = 6.9 z, 2, 9-2 ),.79 (m, 1, 7-),.6 (m, 1, 6-), 2.75 (br-s, 1, 2-), 2.5 (br-s, 1, -), 1.88-1.61 (complex m, 6, 4-2, 5-2, 8-2 ), 1.6 (s,, 6,7--i-Pr), 1.4 (s,, 6,7--i-Pr), 1.28 (t, J = 7. z, 2

, 1-C 2 C ), 1.05 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 170.8, 15.6 (4C), 1.8, 1.7, 129.6 (2C), 127.6 (2C), 127.6 (2C), 108.1, 80.2, 77.6, 61., 60.8, 8.4, 5.8, 4.7, 0.7, 27. (2C), 26.8 (C), 24.6, 19.2, 14.; R-MS (FAB, BA matrix) m/z 526.012 [M+] +, Calcd for C 0 44 5 Si 526.2989 [M+]. (2S,S,6S,7S)-( )-Ethyl-2-azido-9-(tert-butyldiphenylsilyloxy)- -(p-nitrobenzensulfonylimino)-6,7--isopropylidenenonanate TBDPS Et 1) p-scl, DMAP, TEA C 2 Cl 2, 0 C then r.t. TBDPS Et p-s 2) a, DMF, r.t. 85% for 2 steps TBDPS 6 7 2 p-s Et regioselectivity; > 20 : 1 According to the procedure of the s fromation (pages 55) and the azidolysis (pages 55), the title compound azido (585 mg, 85% for 2 steps; as a single isomer) was obtained from aziridine (480 mg, 91 µmol). 2 Rf = 0.56 (Silica gel, hexane : AcEt = 2 : 1); [α] D 42.5 (c 1.00, CCl ); IR (acl) ν cm 1 286 (-), 2112 (-= + = ), 172 (C=, ester), 151 ( 2 ), 148 (S 2 ), 1165 (-S 2 ), 1111, 1090, 77, 702, 611; 1 -MR (270 Mz, CDCl ) δ : 8.6 (ddd, J = 2.0, 2.0, 8.9 z, 2, -S( 2 )-Ph- 2 ), 8.08 (ddd, J = 2.0, 2.0, 8.9 z, 2, -S( 2 )-Ph- 2 ), 7.69-7.62 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.47-7.4 (complex m, 6, 9--Si(Ph) 2 C(C ) ), 5.56 (d, J = 8. z, 1, -), 4.24 (q, J = 6.9 z, 2, 1-C 2 C ), 4.22 (d, J = 4.0 z, 1, 2-),.79 (t, J = 6. z, 2, 9-2 ),.78 (m, 1, 7-),.67 (ddd, J = 1.0, 4.0, 7.9 z, 1, -),.57 (dt, J = 2.0, 8.5 z, 1, 6-), 1.75-1.48 (complex m, 6, 4-2, 5-2, 8-2 ), 1.7 (s,, 6,7--i-Pr), 1. (s,, 6,7--i-Pr), 1.1 (t, J = 6.9 z,, 1-C 2 C ), 1.04 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 167.7, 150.0, 146.4, 15.5 (2C), 15.5 (2C), 1.7 (2C), 129.6 (2C), 128. (2C), 127.6 (2C), 127.6 (2C), 124. (2C), 108.2, 79.9, 64.8, 62.5, 60.5, 60.4, 55.1, 5., 27.9, 27.2, 27.1, 26.8 (C), 21.0, 19.1, 14.1; R-MS (FAB, BA + ai matrix) m/z 776.2761 [M+a] +, Calcd for C 6 47 5 9 SiSa 776.2761 [M+a]. 24

(2S,S,6S,7S)-( )-Ethyl--amino-2-azido-9-(tert-butyldiphenylsilyloxy)-6,7-- isopropylidenenonanate; 57 TBDPS p-s Et PhS, DIPEA MeC, r.t. 75% TBDPS 6 7 According to the procedure of the deprotection of s (pages 56), the title compound 57 (125 mg, 75%) was obtained from azido-s-amine (221 mg, 29 µmol). Rf = 0.12 (Silica gel, hexane : AcEt = 2 : 1); [α] D 26 40.2 (c 1.00, CCl ); IR (acl) ν cm 1 94 (-), 2108 (-= + = ), 179 (C=, ester), 1429, 179, 1244, 121, 111, 100, 870, 79, 704; 1 -MR (270 Mz, CD 2 Cl 2 ) δ : 7.69-7.62 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.45-7. (complex m, 6, 9--Si(Ph) 2 C(C ) ), 4.25 (q, J = 6.9 z, 2, 1-C 2 C ),.89 (d, J = 5.6 z, 1, 2-),.81 (m, 1, 7-),.82 (dt, J = 2.0, 7. z, 2, 9-2 ),.62 (m, 1, 6-),.11 (m, 1, -), 1.89-1.40 (complex m, 6, 4-2, 5-2, 6-2 ), 1.4 (s,, 6,7--i-Pr), 1.2 (s,, 6,7--i-Pr), 1.28 (t, J = 7. z,, 1-C 2 C ), 1.05 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 168.9, 15.5 (2C), 15.5 (2C), 1.8, 1.7, 129.6 (2C), 127.6 (4C), 108.1, 80.4, 77.5, 67.5, 61.9, 60.7, 5.1, 5.6, 29.8, 29.0, 27.2 (2C), 26.8 (C), 19.2, 14.2; R-MS (FAB, BA matrix) m/z 569.19 [M+] +, Calcd for C 0 45 4 5 Si 569.159 [M+]. 2 57 2 Et (2S,S,6S,7S,2 R)-( )-Ethyl-2-azido--(2 -bromopropanamido)-9-(tert-butyl- diphenylsilyloxy)-6,7--isopropylidenenonanate TBDPS 2 57 Et Br (R)-10 PyBP, DIPEA, C 2 Cl 2, r.t. 95% TBDPS 6 7 Et 2 Br 2' According to the procedure of the amido formation (pages 57), the title compound (52 mg, 95%) was obtained from 57 (01 mg, 528 µmol). Rf = 0.57 (Silica gel, hexane: AcEt = 2 : 1); [α] 27 D 41.1 (c 0.50, CCl ); IR (acl) ν cm 1 47 (-), 00 (-), 2112 (-= + = ), 1741 (C=, ester), 1662 (C=, amide), 171, 124, 1205, 111, 109, 1028, 82, 79, 704; 1 -MR (270 Mz, CD 2 Cl 2 ) δ : 7.70-7.62 (complex m, 4, 9--Si(Ph) 2 C(C ) ), 7.47-7.4 (complex m, 6, 9--Si(Ph) 2 C(C ) ), 6.59 (d, J = 8.9 z, -), 4.42 (m, 1, -), 4.8 (q, J = 6.9 z, 1, 2 -), 4.29 (q, J = 6.9 z, 2, 1-C 2 C ), 4.27 (d, J = 4.0 z, 1, 2-), 25

.80 (dt, J = 1., 5.6 z, 2, 9-2 ),.77 (m, 1, 7-),.62 (dd, J =.0, 8. z, 1, 6-), 1.88 (d, J = 6.9 z,, - ), 1.91-1.44 (complex m, 6, 4-2, 5-2, 8-2 ), 1.6 (s,, 6,7--i-Pr), 1.4 (s,, 6,7--i-Pr), 1.0 (t, J = 6.9 z,, 1-C 2 C ), 1.05 (s, 9, 9--Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 169.5, 168.1, 15.5 (4C), 1.6 (2C), 129.6 (2C), 127.6 (4C), 108.2, 79.8 (2C), 64., 62., 60.6, 50.6, 44.5, 5.5, 28.6, 27.2 (2C), 26.8 (C), 26.7, 22.8, 19.2, 14.1; R-MS (FAB, BA matrix) m/z 70.25 [M+] +, Calcd for C 48 4 6 BrSi 70.2527 [M+]. (S,5S,6S, S,4 S)-( )-4-(tert-butoxycarbonyl)-6-[1 -(tert-butyldiphenylsilyloxy)-,4 --isopropylidene hexane-6 -yl]- 5-ethyloxycarbonyl--metyl-2-piperazinone TBDPS Et Br 1) PPh, TEA, MeC then 2, r.t. then 60 C TBDPS Et 2) Boc 2, AcEt, 80 C 79% in 2 steps TBDPS 4' ' Et 6 5 Boc According to the procedure of the piperizinone formation (pages 58) and the Boc protection (pages 58), the title compound (12 mg, 79% for 2 steps) was obtained from azido-amide (169 mg, 240 µmol). 29 Rf = 0.26 (Silica gel, hexane : AcEt = 1 : 1); [α] D 12.7 (c 0.50, CCl ); IR (acl) ν cm 1 1747 (C=, ester), 169 (C=, urethane), 1668 (C=, amide), 1471, 1429, 167, 117, 1169, 109, 100, 741, 702; 1 -MR (270 Mz, CDCl ), as two rotamers. δ : 7.69-7.62 (complex m, 4, 1 --Si(Ph) 2 C(C ) ), 7.47-7.2 (complex m, 6, 1 --Si(Ph) 2 C(C ) ), 6.27 (br-s, 1, 1-), 5.07 (br-s, /5, 5-), 4.79 (br-s, 2/5, 5-), 4.52-4.8 (br-m, 1, -), 4.19 (q, J = 7. z, 2, 1-C 2 C ),.82 (t, J = 5.9 z, 2, 1-2 ),.80 (m, 1, -),.68 (m, 1, 6-),.60 (dt, J = 2.0, 8. z, 1, 4 -), 1.88-1.52 (complex m, 6, 2-2, 5-2, 6-2 ), 1.60 (d, J = 2. z,, -C ), 1.50 (s, 9, 4-C-C(C ) ), 1.9, 1.6 (s, each,,4 --i-pr), 1.27 (t, J = 7. z,, 1-C 2 C ), 1.05 (s, 9, 1 --Si(Ph) 2 C(C ) ); 1 C-MR (67.5 Mz, CDCl ) δ : 170.9, 168.7, 154.0, 15.4 (4C), 1.6 (2C), 129.5 (2C), 127.5 (4C), 108., 81.2, 80.5, 77.5, 61.1, 60.5, 54.1, 5.6, 52.4, 5.4, 28.9, 28.7, 28.2 (C), 27.2, 27.0, 26.7 (C), 19.1, 18., 14.0; R-MS (FAB, BA matrix) m/z 697.884 [M+] +, Calcd for 26

C 8 57 2 8 Si 697.884 [M+]. (S,5S,6S, S,4 S)-( )-4-(tert-butoxycarbonyl)- 6-[1 -(tert-butyldiphenylsiloxy)-,4 --isopropylidenehexane-6 -yl]- 5-( 1 -tert-butyl-l-valinyl-l-alanyl)carbonyl--metyl-2-piperazinone TBDPS 2) PyBP, DIPEA C 2 Cl 2, r.t. 2 t-bu 19 68% for 2 steps Et Boc TBDPS 1''' 1) Li, TF/Me/ 2, r.t. ''' 4''' 2' t-bu 1' 1" 2" Boc TBDPS According to the procedure of the hydrolysis (pages 59) and the condensation (pages 58), the title compound (12 mg, 68% for 2 steps) was obtained from ethyl ester (12 mg, 189 µmol). Rf = 0.45 (Silica gel, hexane : AcEt = 1 : 2); [α] D 2 40.5 (c 2.00, CCl ); IR (acl) ν cm 1 6 5 286 (-), 1747 (C=, ester), 169 (C=, urethane), 1660 (C=, amide), 159, 1471, 171, 117, 122, 1157, 111, 849, 746, 704; 1 -MR (400 Mz, CDCl ) as two rotamers. δ : 7.67-7.64 (complex m, 4, 1 --Si(Ph) 2 C(C ) ), 7.44-7.4 (complex m, 6, 1 --Si(Ph) 2 C(C ) ), 7.0 (br-s, 1, 1-), 6.89 (br-d, J = 7.0 z, 1, 2 -), 6.58 (br-d, J = 8.8 z, 1, 2 -), 4.76 (br-s, 4/5, 5-), 4.52 (br-s, 1/5, 5-), 4.49-4.42 (br-m, 1, -), 4.46 (dq, J = 6.8, 7.0 z, 1, 2 -), 4.40 (dd, J = 4.5, 8.8 z, 1, 2 -),.81 (d, J = 5.2 z, 1, 1-2 ),.79 (d, J = 5.2 z, 1, 1-2 ),.78 (ddd, J =.0, 9.0, 11.2 z, 1, -),.67 (ddd, J = 4.5, 5.2, 8.0 z, 1, 6-),.57 (dt, J = 1.5, 9.0 z, 1, 4 -), 2.15 (dqq, J = 4.5, 6.8, 6.9 z, 1, -), 2.02-1.84 (complex m, 2, 6-2 ), 1.80 (m, 1, 2-2 ), 1.74-1.65 (complex m, 2, 2-2, 5-2 ), 1.50 (m, 1, 5-2 ), 1.51 (s, 9, 4-C-C(C ) ), 1.47 (d, J = 7.2 z,, -C ), 1.45 (s, 9, 1 -C(C ) ), 1.04 (s, 9, 1 --Si(Ph) 2 C(C ) ), 1.7 (s,,,4 --i-pr), 1.4 (s,,,4 --i-pr), 1. (d, J = 6.8 z,, - ), 0.89 (d, J = 6.9 z,, -(C ) 2 ), 0.87 (d, J = 6.9 z,, -(C ) 2 ); 1 C-MR (100 Mz, CDCl ) δ : 171. (C-1 ), 170.7 (C-1 ), 170.2 (C-2), 168.4 (1C, 5-C-), 154.5 (1C, 4-C-C(C ) ), 15.6 (2C, 1 --Si(Ph) 2 C(C ) ), 15.6 (2C, 1 --Si(Ph) 2 C(C ) ), 1.8 (1C, 1 --Si(Ph) 2 C(C ) ), 1.7, (1C, Boc 27

1 --Si(Ph) 2 C(C ) ), 10.0 (1C, 1 --Si(Ph) 2 C(C ) ), 129.6 (1C, 1 --Si(Ph) 2 C(C ) ), 127.6, (2C, 1 --Si(Ph) 2 C(C ) ), 127.6, (2C, 1 --Si(Ph) 2 C(C ) ), 108.5 (1C,,4 --i-pr), 82.1 (1C, 4-C-C(C ) ),82.0 (1C, 1 -C(C ) ), 80.6 (C-4 ), 77.8 (C- ), 60.7 (C-1 ), 57.5 (C-2 ), 5.7 (C-6), 52.5 (C-5), 52.0 (C-), 48.7 (C-2 ), 5.4 (C-2 ), 1. (C- ), 29.4 (C-5 ), 28.7 (C-6 ), 28.2 (C, 4-C-C(C ) ), 28.0 (C, 1 -C(C ) ), 27. (1C,,4 --i-pr), 27.1 (1C,,4 --i-pr), 26.8 (C, 1 --Si(Ph) 2 C(C ) ), 19.1 (1 --Si(Ph) 2 C(C ) ), 17.9 (C- ), 17.8 (1C, -C ), 18.9 (1C, -(C ) 2 ), 17.5 (1C, -(C ) 2 ); R-MS (FAB, BA + ai matrix) m/z 917.5048 [M+a] +, Calcd for C 48 74 4 10 Sia 917.5072 [M+a]. (S,5S,6S, S,4 S)-( )-4-(tert-Butoxycarbonyl)-5-( 1 -tert-butyl-l-valinyl- L-alanyl)carbonyl-6-[1 -hydroxy-,4 --isopropylidenehexane-6 -yl]- -metyl-2-piperazinone; ( )-26 ' t-bu 2' t-bu 1' 1" 2" TBAF, TF, r.t. TBDPS ''' 4''' " 6 Boc 99% 5 Boc 1''' According to the procedure of the deprotection of TBDPS (pages 60), the title compound (84 mg, 99%) was obtained from TBDPS ether (116 mg, 129 µmol). 27 Rf = 0.9 (Silica gel, CCl : Me = 10 : 1); [α] D 44.5 (c 2.00, CCl ); IR (acl) ν cm 1 7 (), 00 (), 172 (C=, ester), 1728 (C=, urethane), 1645 (C=, amide), 159, 1456, 171, 125, 1255, 1221, 1155, 1066, 918, 847, 7; 1 -MR (400 Mz, CDCl ) δ : 6.9 (br-s, 1, 2 -), 6.77-6.66 (br-m, 1, 1-), 6.76 (br-d, J = 8.6 z, 1, 1-), 4.74 (br-s, 1, 5-), 4.59-4.46 (br-m, 1, 2 -), 4.46-4.8 (br-m, 1, -), 4.9 (dd, J = 4.5, 8.6 z, 1, 2 -),.85-.75 (complex m, 2, 1 -),.74 (m, 1, -),.68 (m, 1, 6-),.67 (m, 1, 4 -),.0 (br-s, 1, 1 -), 2.15 (dqq, J = 4.5, 6.8, 7.0 z, 1, -), 1.96-1.82 (complex m, 2, 6-2 ), 1.86-1.78 (complex m, 2, 2-2 ), 1.81 (m, 1, 5-2 ), 1.51 (m, 1, 5-2 ), 1.50 (s, 9, 1 -C(C ) ), 1.46 (d, J = 7. z,, -C ), 1.45 (s, 9, 4-C-C(C ) ), 1.7 (s, 6,,4 --i-pr), 1.1 (d, J = 7.0 z,, - ), 0.91 (d, J = 7.0 z,, -(C ) 2 ), 0.88 (d, J = 6.8 z,, -(C ) 2 ); 1 C-MR (75.0 Mz, CDCl ) δ : 171.9 (C-1 ), 170.8 (C-1 ), 170. (C-2), 168.1 (1C, 5-C-), 154.7 (1C, 4-C-C(C ) ), 28

108.4 (1C,,4 --i-pr), 82.1 (1C, 1 -C(C ) ), 81.9 (1C, 4-C-C(C ) ), 80.2 (C-4 ), 79.8 (C- ), 60.0 (C-1 ), 57.5 (C-2 ), 5. (C-6), 52.5 (C-5), 52.2 (C-), 48.5 (C-2 ), 5.0 (C-2 ), 1.2 (C- ), 0.9 (C-5 ), 28.6 (C-6 ), 28. (C, 1 -C(C ) ), 28.0 (C, 4-C-C(C ) ), 27.1 (2C,, 4 --i-pr), 18. (C- ), 18.0 (1C, -C ) 18.9 (1C, -(C ) 2 ), 17.6 (1C, -(C ) 2 ); R-MS (FAB, BA + ai matrix) m/z 679.907 [M+a] +, Calcd for C 2 56 4 10 a 679.894 [M+a]. (S,5S,6S,2 S,4 S,5 S)-( )-4-(tert-Butoxycarbonyl)-5-( 1 -tert-butyl-l-valinyl- L-alanyl)carbonyl-6-[2 -hydroxy-4,5 --isopropylidene-1 -nitroheptane-7 -yl]- -methyl-2-piperazinone; 59 Boc t-bu S Py, DMS, TEA C 2 Cl 2, 0 C then r.t. C Boc 58 t-bu t-bu RR Co t-bu t-bu t-bu (R,R)-salen-cobalt complex; (R, R)-45 Me 2, DIPEA, C 2 Cl 2, 40 C 7%, 90% de 2 1' 2' 4' 5' " 2" 1" t-bu 1"' 2"' 6 "' 5 Boc 59 According to the procedure of the oxidtion of primary alcohol (pages 62) and nitro aldol reaction (pages 62), the title compound 59 (15.2 mg, 7% for 2 steps; 90% de) was obtained from primary alcohol (20.5 mg, 1.2 µmol). 26 Rf = 0.2 (Silica gel, CCl : Me = 10 : 1); [α] D 119.9 (c 1.00, CCl ); IR (acl) ν cm 1 17 (), 1726 (C=, ester), 1666 (C=, amide), 1554 ( 2 ), 169, 125, 1255, 1219, 1159, 1097, 845; 1 -MR (400 Mz, CD D) δ : 8.15 (br-s, 1, 2 -), 8.10 (br-s, 1, 2 -), 4.86 (br-d, J =.2 z, 1, 5-), 4.65 (dd, J = 2.8, 11.8 z, 1, 1-2 ), 4.5-4.4 (complex m, 2, 2 -, 2 -), 4.9 (dd, J = 9.2, 11.8 z, 1, 1-2 ), 4.6 (q, J = 6.8 z, 1, -), 4.17 (dd, J =.0, 6.0 z, 1, 2 -),.88 (ddd, J = 2.0, 8.4, 10.0 z, 1, 4 -),.70 (ddd, J =.2, 8.0, 14.5 z, 1, 6-),.64 (ddd, J =.0, 5.8, 8.4 z, 1, 5 -), 2.1 (dsep, J = 6.0, 7.0 z, 1, -), 1.87-1.70 (complex m,, - 2, 7-2 ), 1.78-1.58 (complex m, 2, - 2, 6-2 ), 1.52 (d, J = 6.8 z,, -C ), 1.52 (s, 9, 4-C-C(C ) ), 1.51 (m, 1, 6-2 ), 1.46 (s, 9, 29

1 -C(C ) ), 1.9 (s,, 4,5 --i-pr), 1.7 (s,, 4,5 --i-pr), 1.6 (d, J = 6.8 z,, - ), 0.96 (d, J = 7.0 z, 6, -(C ) 2 ); 1 C-MR (75.0 Mz, CDCl ) δ : 172., 170.8, 170.7, 168.2, 155., 108.6, 82.1, 81.9, 81.2, 80.0, 77.8, 67.1, 57.6, 5.1, 52.9, 52.8, 48.4, 7.5, 1.1, 29.6, 28.5, 28.2 (C), 28.0 (C), 27.1 (2C), 18.9, 18.5, 18.1, 17.6; R-MS (FAB, BA + ai matrix) m/z 78.91 [M+a] +, Calcd for C 57 5 12 a 78.901 [M+a]. Analytical PLC (normal phase) DAICEL CIRALCEL D 4.6 φ x 250 mm, hexane : 2-propanol = 96 : 4, 0.8 ml/min, temp. = 25 C, 226 nm. (S,5S,6S,2 S,4 S,5 S)-( )-4-(tert-Butoxycarbonyl)-5-( 1 -tert-butyl-l-valinyl- L-alanyl)carbonyl-6-{1 -[,-di-(tert-butoxycarbonyl)guanidino]-2 -hydroxy- 4,5 --isopropylideneheptane-7 -yl}--methyl-2-piperazinone; 60 t-bu 1) C 2 4, Pd / C Me, r.t. t-bu 2 Boc 2 Boc 59 2) MeC, r.t., 47 Boc Boc 76% for 2 steps Boc 1' 2' Boc 4' 5' " 2" t-bu 1" 1"' 2"' 6 "' 5 Boc 60 According to the procedure of the reduction of nitro group (pages 6) and guanidine formation (pages 6), the title compound 60 (57.9 mg, 76% for 2 steps) was obtained from nitro aldol adduct (58.6 mg, 81.9 µmol). Rf = 0.7 (Silica gel, CCl : Me = 10 : 1); [α] 22 D 29.5 (c 1.00, CCl ); IR (acl) ν cm 1 19 (), 298 (), 1792 (C=, guanidyl), 1724 (C=, ester), 1647 (C=, amide), 169, 121, 1252, 1157, 1140, 1057, 771; 1 -MR (400 Mz, CD D) δ : 4.92-.98 (br-s, 1, 5-), 4.4 (br-q, J = 7.0 z, 1, 2 -), 4.7 (q, J = 7.0 z, 1, -), 4.18 (d, J = 6.0 z, 1, 2 -),.9 (m, 1, 2 -),.87 (dt, J = 2.6, 9.0 z, 1, 4 -),.71 (m, 1, 6-),.66 (dt, J = 2.6, 8.6 z, 1, 5 -),.62 (dd, J = 4.0, 14.0 z, 1, 1-2 ),.1 (dd, J = 7.0, 14.0 z, 1, 1-2 ), 2.1 (dsep, J = 4., 6.0, 7.0 z, 1, -), 1.86-1.70 (complex m, 2, 6-2, 7-2 ), 1.74 (ddd, J = 2.6, 9.0, 1.5 z, 1, - 2 ), 1.64 (ddd, J = 4.0, 9.0, 1.5 z, 1, - 2 ), 1.6-1.54 (complex m, 2, 6-2, 0

7-2 ), 1.5 (s, 9, 1 --C[C-C(C ) ][C-C(C ) ]), 1.52 (d, J = 7.0 z,, -C ), 1.51 (s, 9, 1 -C(C ) ), 1.47 (s, 18, 1 --C[C-C(C ) ][C-C(C ) ], 4-C-C(C ) ), 1.8 (s,, 4,5 --i-pr), 1.7 (s,, 4,5 --i-pr), 1.6 (d, J = 7.0 z,, - ), 0.96 (d, J = 7.0 z, 6, -(C ) 2 ); 1 C-MR (67.5 Mz, CD 2 Cl 2 ) δ : 172.2, 171.0, 170.4, 168.4, 16.2, 157. (2C), 15.1, 108.7, 8.4, 82.1, 81.8, 80.7, 79., 79.1, 78.5, 69.2, 58.0, 52.6, 52.4, 48.8, 47.6, 8.0, 1.4, 29.0, 28.7, 28.2 (C), 28.2 (C), 28.0 (C), 28.0 (C), 27.2 (2C), 19.0, 18.5, 18.1, 17.7; R-MS (FAB, BA matrix) m/z 928.5646 [M+] +, Calcd for C 44 78 7 14 928.5607 [M+]. (S,5S,6S,2 S, S,4 R)-( )-6-{1 -[, -Di-(tert-butoxycarbonyl)-2 -iminoimidazo- lidin-4 -yl]-2, --isopropylidenepentane-5 -yl}-4-(tert-butoxy-carbonyl)-5-( 1 -tert- butyl- L-valinyl-L-alanyl)carbonyl--methyl-2-piperazinone; 61 Boc Boc Boc 60 t-bu I 2, PPh, imidazole C 2 Cl 2, 0 C then r.t. 1" 2' ' 4" 67% Boc " Boc "' 2"' t-bu 1"' 1"" 2"" 6 "" 5 Boc 61 According to the procedure of the S 2 cyclization (pages 66), the title compound 61 (8.1 mg, 67%) was obtained from 60 (57.9 mg, 62.5 µmol). Rf = 0.2 (Silica gel, CCl : Me = 10 : 1); [α] D 21 51.7 (c 1.00, CCl ); IR (acl) ν cm -1 15 (), 1758 (C=, guanidyl), 174 (C=, ester), 170 (C=, urethane), 1668 (C=, urethane), 1606 (C=, amide), 15, 1456, 179, 169, 117, 1254, 1147, 1001, 849, 752; 1 -MR (400 Mz, CD D) δ : 4.84-4.78 (br-s, 1, 5-), 4.45 (m, 1, 2 -), 4.4 (dddd, J = 2.5,.0, 9.1, 10.0 z, 1, 4 -), 4.6 (q, J = 6.8 z, 1, -), 4.18 (d, J = 5.5 z, 1, 2 -),.78 (dd, J = 9.1, 12.2 z, 1, 5-2 ),.70 (m, 1, 2 -),.68 (m, 1, -),.68 (m, 1, 6-),.58 (dd, J = 2.5, 12.2 z, 1, 5-2 ), 2.1 (dsep, J = 5.5, 6.8 z, 1, -), 1.99 (ddd, J =.0, 9.0, 1.5 z, 1, 1-2 ), 1.89 (dd, J = 2., 10.0, 1.5 z, 1, 1-2 ), 1.76 (m, 1, 4-2 ), 1.75 (m, 1, 5-2 ), 1.62 (m, 1, 4-2 ), 1.56 (m, 1, 5-2 ), 1.55 (s, 9, 2 --C--C(C ) ), 1.52 (s, 9, -C--C(C ) ), 1.51 (d, J = 6.8 z,, -C ), 1.49 (s, 9, 4-C--C(C ) ), 1

1.47 (s, 9, 1 --C(C ) ), 1.6 (s, 6, 2, --i-pr), 1.5 (d, J = 6.2 z,, - ), 0.96 (d, J = 6.8 z, 6, -(C ) 2 ); 1 C-MR (100 Mz, CD D) δ : 174.4 (C-1 ), 172.9 (C-2), 172.1 (C-1 ), 169.7 (1C, 5-C--), 155.7 (C-2 ), 154.2 (1C, 4-C--C(C ) ), 152.5 (1C, -C--C(C ) ), 152.0 (1C, -C--C(C ) ), 110.0 (1C, 2, --i-pr), 84.6 (1C, 2 --C--C(C ) ), 82.7 (1C, 1 --C(C ) ), 82.7 (1C, -C--C(C ) ), 81.5 (1C, 4-C--C(C ) ), 82.0 (C- ), 78. (C-2 ), 59.8 (C-2 ), 55.7 (C-4 ), 54.2 (C-5 ), 55.0 (C-6), 5.5 (C-), 49.8 (C-2 ), 7. (C-1 ), 1.8 (C- ), 29.5 (C-4 ), 29.4 (C-5 ), 28.6 (C, 2 --C--C(C ) ), 28.5 (C, -C--C(C ) ), 28.4 (C, 4-C--C(C ) ), 28. (C, 1 --C(C ) ), 27.7, 27.5 (each 1C, 2, --i-pr), 19.7 (1C, -(C ) 2 ), 18.5 (1C, -(C ) 2 ), 18.5 (1C, -C ), 18.5 (C- ), Carbon at C-5 was not observed.; R-MS (FAB, BA matrix) m/z 910.5516 [M+] +, Calcd for C 44 76 7 1 910.5501 [M+]. (S,5S,6S,2 S, S,4 R)-( )-5-(L-Valinyl-L-alanyl)carbonyl- 6-[2, -dihydroxy-1 -(1 -carbamoyl-2 -iminoimidazolidin-4 -yl)pentane-5 -yl]- -methyl-2-piperazinone; ( S)-4 t-bu Boc Boc Boc 61 1) PMP isocyanate Benzene, r.t. PMP Boc Boc Boc t-bu 2) CA MeC / 2, 0 C 2 Boc Boc Boc t-bu "' 2"' 1"' 1"" 2"" ) TFA / 2, r.t. 2 1" 2' ' 4" 6 "" TFA 5 60% for steps " ('S)-4 According to the procedure of the cabamoyl formation and the deprotection of PMP, Boc and t-bu (pages 67), the title compound ( S)-4 (7. mg, 60% for steps) was 2

obtained from 61 (16.6 mg, 18.2 µmol). Rt = 6.76 min (Analitical PLC, Develosil C0 UG-5, 4.6 φ X 250 mm, 0.1% TFA / 15% Me / 2, 1.0 ml/min, 210 nm); [α] 22 D 4.5 (c 0.10, Me); IR (KBr) ν cm 1 419 (), 217 (), 170 (C=, carboxylic acid), 1672 (C=, amide), 1562, 1425, 1201, 118, 89, 800, 72; 1 -MR (400 Mz, D 2 with TFA) δ : 4.5 (d, J = 4.5 z, 1, 5-), 4.45 (q, J = 7.2 z, 1, 2 -), 4.25 (dddd, J = 5.8, 6.2, 6.8, 8.4 z, 1, 4 -), 4.21 (dd, J = 8.4, 8.4 z, 1, 5-2 ), 4.11 (q, J = 7.2 z, 1, -), 4.09 (d, J = 7.2 z, 1, 2 -),.97 (ddd, J = 2.1, 4.5, 11.2 z, 1, 6-),.79 (dd, J = 5.8, 8.4 z, 1, 5-2 ),.24 (ddd, J =.2, 6.8, 10.2 z, 1, 2 -),.45 (ddd, J =.2, 6.8, 8.6 z, -), 2.12 (1, dsep, J = 6.8, 6.8 z, 1, -), 1.89 (ddd, J = 6.2, 10.2, 14.2 z, 1, 1-2 ), 1.80 (ddd, J =.2, 6.8, 14.2 z, 1, 1-2 ), 1.6 (m, 1, 4-2 ), 1.58 (m, 1, 5-2 ), 1.55 (q, J = 7.2 z,, -C ), 1.54 (m, 1, 5-2 ), 1.54 (m, 1, 4-2 ), 1.8 (d, J = 7.2 z,, - ), 0.92 (d, J = 7.0 z, 6, -(C ) 2 ); 1 C-MR (100 Mz, D 2 with TFA), reference (0 and 200 ppm); δ : 177.9 (C-1 ), 177.1 (C-1 ), 170.9 (C-2), 167.6 (1C, 5-C-), 159.0 (C-2 ), 158.4 (1C, 1 -C- 2 ), 76.0 (C- ), 7.9 (C-2 ), 61.5 (C-2 ), 59.6 (C-5), 54.8 (C-), 5.1 (C-4 ), 54.0 (C-6), 5.4 (C-5 ), 52.1 (C-2 ), 9.5 (C-1 ), 2.4 (C- ), 1.2 (C-5 ), 0.4 (C-4 ), 21.1 (1C, -(C ) 2 ), 20.1 (1C, -(C ) 2 ), 19.1 (C- ), 16.6 (1C, -C ); R-MS (FAB, BA matrix) m/z 557.042 [M+] +, Calcd for C 2 41 8 8 557.047 [M+].

2 atural Guadinomine C2 (4) 2'''S 2 4''R 2'S 'R 2''''S 5S 6S S Synthetic ('R)-Guadinomine 2'''S 2 4''R 2'S 'S 2''''S 5S 6S S Synthetic ('S)-Guadinomine 400Mz 1-MR, in 1% TFA/D2 Synthetic ( S)-Guadinomine C2 [( S)-4] TFA 100 Mz 1C-MR (in 1%TFA/D2) TFA 4

<Synthesis of (2S, S, 6R, 7S, 4 R)-guadimonine B 2> -L-Ala-L-Val-t-Bu -L-Ala-L-Val-t-Bu TBDPS Boc Boc a c 2 Boc Boc d f 51 62 Boc Boc -L-Ala-L-Val-t-Bu 2 Boc g i Boc 6 -L-Ala-L-Val- 6R 4'R 7S S 2S 2 2 2 Reagents and conditions: a) TBAF, TF, RT, 1 h, 78%; b) S pyridine, DMS, Et, C 2 Cl 2, 0 C to RT, 6 h; c) (R, R)-salen-cobalt complex (R, R)-45, Me 2, DIPEA, C 2 Cl 2, 40 C, 12 h, 69% (2 steps), 90% de; d) C 2 4, 10% Pd/C, Me, RT, 2 h; e) 47, DIPEA, MeC, RT, 2 h, 68% (2 steps); r) I 2, PPh, imidazole, C 2 Cl 2, 0 C to RT, 2 h, 65%; s) PMPisocyanate, Ph, RT, 10 min; t) CA, MeC/ 2 (1/1), 0 C, 2 h; u) TFA/ 2 (/1), RT, 6 h 60% ( steps). (2S,S,6R,7S)-( )-( 1 -tert-butyl-l-valinyl-l-alanyl)-2,-bis(tert-butoxycarbonyl- amino)-9-hydroxy-6,7--isopropylidenenonanate TBDPS Boc Boc 51 t-bu TBAF, TF, r.t. 78% 9 7 6 2 Boc 2' t-bu 1' 1" 2" Boc According to the procedure of the deprotection of TBDPS (pages 60), the title compound (117 mg, 78%) was obtained from 51(200 mg, 212 µmol). Rf = 0.7 (Silica gel, CCl : Me = 10 : 1); m. p. 1-15 C (CCl ); [α] 27 D 28.5 (c 0.50, CCl ); IR (acl) ν cm 1 8 (-), 1722 (C=,ester), 1689 (C=, urethane), 1651 (C=, amide), 1525, 1456, 167, 1248, 1221, 1167, 1055, 1014, 870, 78; 1 -MR (270 Mz, CD 2 Cl 2 ) δ : 6.99 (br-d, J = 4. z, 1, 2 -), 6.5 (br-d, J = 8. z, 1, 2 -), 5.76 (br-d, J = 5.6 z, 1, -), 5.7-5.0 (br-m, 1, 2-), 4.5 (m, 1, 2 -), 4. (dd, J = 5.0, 8. z, 1, 2 -), 4.2 (m, 1, 7-), 4.20 (dd, J =.0, 7. z, 1, 2-), 4.0 (dd, J = 6., 12.9 z, 1, 6-),.86-.7 (br-m, 1, -), 5

.76 (d, J = 6. z, 1, 9-2 ),.72 (d, J = 6. z, 1, 9-2 ), 2.1 (m, 1, -), 1.77-1.22 (complex m, 6, 4-2, 5-2, 8-2 ), 1.45 (s, 9, -C-C(C ) ), 1.4 (s, 9, 2-C-C(C ) ), 1.41 (s, 9, 1 -C(C ) ), 1.6 (d, J = 7. z,, - ), 1.0 (s,, 6,7--i-Pr), 1.26 (s,, 6,7--i-Pr), 0.90 (d, J = 5. z,, -(C ) 2 ), 0.88 (d, J = 5. z, -(C ) 2 ); 1 C-MR (67.5 Mz, CD 2 Cl 2 ) δ : 171.7, 170.9, 170.5, 156.6, 155.8, 107.8, 81.8, 80.0, 79.6, 77.7, 77.7, 76.4, 60.5, 57.7, 57.4, 49.4, 2.4, 1., 29.0, 28., 28.2 (C), 28.1 (C), 27.8 (C), 26.8, 25.6, 18.9, 17.9, 17.6; R-MS (FAB, BA matrix) m/z 70.4528 [M+] +, Calcd for C 4 6 4 11 70.449 [M+]. 6

(2S,S,6R,7S,9S)-( )-( 1 -tert-butyl-l-valinyl-l-alanyl)-2,-bis(tert-butoxycarbonyl- amino)-9-hydroxy-6,7--isopropylidene-10-nitrononanate; 62 Boc Boc t-bu S Py, DMS, TEA C 2 Cl 2, 0 C then r.t. C Boc Boc t-bu t-bu RR Co t-bu t-bu t-bu (R,R)-salen-cobalt complex; (R, R)-45 Me 2, DIPEA, C 2 Cl 2, 40 C 69%, 90% de 2 10 7 6 2' t-bu 1' 1" 2" 2 Boc Boc 62 According to the procedure of the oxidtion of primary alcohol (pages 62) and nitro aldol reaction (pages 62), the title compound 62 (62.7 mg, 69% for 2 steps; 90% de) was obtained from primary alcohol (84. mg, 120 µmol). 24 Rf = 0.40 (Silica gel, CCl : Me = 10 : 1); [α] D 0.5 (c 0.50, CCl ); IR (acl) ν cm 1 44 (-, -), 1720 (C=, ester), 1691 (C=, urethane), 1651 (C=, amide), 1556 ( 2 ), 152, 1454, 167, 111, 1248, 1221, 1165, 104, 756; 1 -MR (400 Mz, CD 2 Cl 2 ) δ : 7.20 (br-d, J = 6.0 z, 1, 2 -), 6.70 (br-d, J = 8.6 z, 1, 2 -), 5.82 (br-d, J = 6.0 z, 1, 2-), 5.9 (br-d, J = 8.8 z, 1, -), 4.56 (dd, J = 2.5, 11.2 z, 1, 10-2 ), 4.5 (m, 1, 9-), 4.44 (dd, J =.6, 11.2 z, 1, 10-2 ), 4.9 (dq, J = 6.0, 7.2 z, 1, 2 -), 4.4 (dd, J = 4.6, 8.6 z, 1, 2 -), 4.1 (m, 1, 7-), 4.28-4.19 (br-m, 1, 2-), 4.10 (br-m, 1, 6-),.88-.76 (br-m, 1, -),.67 (br-d, J = 4.0 z, 1, 9-), 2.15 (dqq, J = 4.6, 6.8, 6.8 z, 1, -), 1.82-1.70 (br-m, 1, 4-2 ), 1.68-1.45 (complex m, 5, 4-2, 5-2, 8-2 ), 1.47 (s, 9, 1 -C(C ) ), 1.44 (s, 18, 2--C-C(C ), --C-C(C ) ), 1.41 (s,, 6,7--i-Pr), 1.1 (s,, 6,7--i-Pr), 1.7 (d, J = 7.0 z,, - ), 0.92 (d, J = 6.8 z,, -(C ) 2 ), 0.89 (d, J = 6.8 z,, -(C ) 2 ); 1 C-MR (100 Mz, CD 2 Cl 2 ) δ : 171.9 (C-1 ), 170.9 (C-1 ), 170.4 (C-1), 156.7 (1C, 2--C-C(C ) ), 155.9 (1C, --C-C(C ) ), 108.2 (1C, 6,7--i-Pr), 82.1 (1C, 1 -C(C ) ), 80. (1C, 2--C-C(C ) ), 79.8 (1C, --C-C(C ) ), 81. (C-10), 77.5 (C-6), 74.1 (C-7), 66.8 (C-9), 57.9 (C-2 ), 57.4 (C-2), 5.7 (C-), 49.6 (C-2 ), 4.2 (C-8), 1.5 (C- ), 29.0 (C-4), 28.4 (1C, 6,7--i-Pr), 28. (C, --C-C(C ) ), 28.2 (C, 2--C-C(C ) ), 28.0 (C, 1 -C(C ) ), 26.7 (C-5), 25.7 (1C, 6,7--i-Pr), 17.9 7

(C- ), 18.9 (1C, -(C ) 2 ), 17.7 (1C, -(C ) 2 ); R-MS (FAB, BA matrix) m/z 762.4514 [M+] +, Calcd for C 5 64 5 1 762.4501 [M+]. (2S,S,6R,7S,9S)-( )-( 1 -tert-butyl-l-valinyl-l-alanyl)-2,-bis(tert-butoxy- carbonylamino)-10-[,-di-(tert-butoxycarbonyl)guanidino]-9-hydroxy-6,7-- isopropylidenedecanate t-bu 1) C 2 4, Pd / C Me, r.t. t-bu 2 Boc Boc 62 2 Boc Boc 2) MeC, r.t., Boc 68% for 2 steps Boc 47 Boc 10 9 Boc 7 6 2 Boc ' 2' t-bu 1' 1" 2" " Boc According to the procedure of the reduction of nitro group (pages 6) and guanidine formation (pages 6), the title compound (15.8 mg, 68% for 2 steps) was obtained from nitro aldol adduct (18.1 mg, 2.8 µmol). 25 Rf = 0.42 (Silica gel, CCl : Me = 10 : 1); [α] D 18.4 (c 1.00, CCl ); IR (acl) ν cm 1 44 (-, -), 1724 (C=, guanidyl), 1687 (C=, ester), 1674 (C=, uretane), 1651 (C=, amide), 1614, 1525, 167, 15, 100, 1248, 116, 1140, 1055, 872, 77, 758; 1 -MR (400 Mz, CD 2 Cl 2 ) δ : 11.47 (br-s, 1, 10--C[C-C(C ) ][C-C(C ) ]), 8.64 (t, J = 5.0 z, 1, 10--C[C-C(C ) ][C-C(C ) ]), 7.12 (br-d, J = 6.0 z, 1, 2 -), 6.62 (br-d, J = 8.6 z, 1, 2 -), 5.82 (br-d, J = 6.0 z, 1, 2-), 5.6 (br-d, J = 8.0 z, 1, -), 4.8 (dq, J = 6.0, 7.0 z, 1, 2 -), 4.4 (dd, J = 4.8, 8.6 z, 1, 2 -), 4.29 (ddd, J =.2, 6.2, 9.4 z, 1, 7-), 4.26-4.19 (br-m, 1, 2-), 4.04 (m, 1, 6-),.90 (m, 1, 9-),.86-.77 (br-m, 1, -),.57 (ddd, J = 2., 5.8, 14.0 z, 1, 10-2 ),.8 (ddd, J = 5.4, 7.0, 14.0 z, 1, 10-2 ), 2.14 (dqq, J = 7.0, 7.0, 8.6 z, 1, -), 1.78 (m, 1, 4-2 ), 1.68-1.52 (complex m, 2, 4-2, 8-2 ), 1.52-1.40 (complex m,, 5-2, 8-2 ), 1.51 (s, 9, 1 -C(C ) ), 1.46 (s, 9, 2-C-C(C ) ), 1.45 (s, 9, -C-C(C ) ), 1.45 (s, 9, 10--C[C-C(C ) ][C-C(C ) ], 1.45 (s, 9, 8

10--C[C-C(C ) ][C-C(C ) ]), 1.1 (s,, 6,7--i-Pr), 1.41 (s,, 6,7--i-Pr), 1.7 (d, J = 7.0 z,, - ), 0.91 (d, J = 7.0 z,, -(C ) 2 ), 0.90 (d, J = 7.0 z,, -(C ) 2 ); 1 C-MR (100 Mz, CD 2 Cl 2 ) δ : 171.4 (C-1 ), 170.6 (C-1 ), 170.2 (C-1), 162.5 (1C, 10--C[C-C(C ) ][C-C(C ) ]), 157. (1C, 10--C[C-C(C ) ][C-C(C ) ]), 156.5 (1C, -C-C(C ) ), 155.6 (1C, 2-C-C(C ) ), 152.8 (1C, 10--C[C-C(C ) ][C-C(C ) ]), 107.6 (1C, 6,7--i-Pr), 8. (1C, 1 -C(C ) ), 81.7 (1C, 10--C[C-C(C ) ][C-C(C ) ]), 79.9 (1C, 2-C-C(C ) ), 79.4 (1C, -C-C(C ) ), 79.1 (1C, 10--C[C-C(C ) ][C-C(C ) ]), 77.6 (C-6), 74. (C-7), 69.1 (C-9), 57.5 (C-2 ), 57. (C-2), 5.6 (C-), 49. (C-2 ), 47.8 (C-10), 4.8 (C-8), 1.2 (C- ), 29.1 (C-), 28.1 (1C, 6,7--i-Pr), 28.0 (C, 10--C[C-C(C ) ][C-C(C ) ]), 27.9 (C, 10--C[C-C(C ) ][C-C(C ) ]), 27.8 (C, 2-C-C(C ) ), 27.6 (C, -C-C(C ) ), 27.6 (C, 1 -C(C ) ), 26.8 (C-5), 25.4 (1C, 6,7--i-Pr), 18.6 (1C, -(C ) 2 ), 17. (1C, -(C ) 2 ), 17.5 (C- ); R-MS (FAB, BA matrix) m/z 974.606 [M+] +, Calcd for C 46 84 7 15 974.6025 [M+]. (2S,S,6R,7S,4 R)-( )-( 1 -tert-butyl-l-valinyl-l-alanyl)-2,-bis(tert-butoxycarbonyl- amino)-8-[, -di-(tert-butoxycarbonyl)-2 -iminoimidazolidin-4 -yl]-6,7-- isopropylideneoctanate; 6 Boc t-bu Boc Boc Boc " 2" t-bu 1" 1"' I 2, PPh, imidazole 2"' C 2 Cl 2, 0 C then r.t. 1' 7 6 4' "' 2 Boc 65% Boc Boc ' Boc 6 According to the procedure of the S 2 cyclization (pages 66), the title compound 6 (10.0 mg, 65%) was obtained from guanidine alcohol (15.8 mg, 16.2 µmol). Rf = 0.1 (Silica gel, CCl : Me = 10 : 1); [α] D 24 26.4 (c 0.50, CCl ); IR (acl) ν cm 1 17 (-), 1759 (C=, ester), 1705 (C=, urethane), 1651 (C=, amide), 151, 167, 115, 1252,1149, 1001, 850, 768, 752; 1 -MR (270 Mz, CD 2 Cl 2 ) δ : 9

6.99 (br-d, J = 5.6 z, 1, 2 -), 6.57 (br-d, J = 8.6 z, 1, 2 -), 5.76 (br-s, 1, 2-), 5. (br-s, 1, -), 4.7 (m, 1, 4 -), 4.5 (dq, J = 5.6, 7. z, 1, 2 -), 4.2 (dd, J = 4.6, 8.6 z, 1, 2 -), 4.19 (dd, J = 2., 7.6 z, 1, 2-), 4.07 (m, 1, 7-), 4.04 (m, 1, 6-),.80 (m, 1, -),.7 (dd, J = 10., 10. z, 1, 5-2 ),.48 (dd, J = 2., 10. z, 1, 5-2 ), 2.1 (dqq, J = 4.6, 6.6, 8.6 z, 1, -), 1.94 (ddd, J =.0, 10.2, 10.2 z, 1, 4-2 ), 1.86-1.70 (complex m, 2, 4-2, 8-2 ), 1.69-1.40 (complex m,, 5-2, 8-2 ), 1.51 (s, 9, 1 --C(C ) ), 1.45 (s, 9, 2 --C--C(C ) ), 1.45 (s, 9, -C--C(C ) ), 1.45 (s, 9, 2-C--C(C ) ), 1.4(s, 9, -C--C(C ) ), 1.41 (s,, 6,7--i-Pr), 1.5 (d, J = 7. z,, - ), 1.27 (s, 2, 6,7--i-Pr), 1.26 (s, 1, 6,7--i-Pr), 0.90 (d, J = 6.6 z,, -(C ) 2 ), 0.88 (d, J = 6.9 z,, -(C ) 2 ); 1 C-MR (100 Mz, benzen-d6) δ : 172.2, 171.5, 171.1, 165.2, 159.0, 157., 156.5, 108.6, 8.1, 81.7, 80., 79.9, 78.4, 78.0, 74.5, 58.8, 58.5, 55.0, 54.7, 54.0, 50., 4.0, 2.0, 0.5, 29.7, 29.0 (C), 28.9 (C), 28.8 (C), 28.7 (C), 28.5 (C), 27.4, 26.2, 19.6, 18.5, 18.; R-MS (FAB, BA matrix) m/z 956.5911 [M+] +, Calcd for C 46 82 7 14 956.5920 [M+]. (2S,S,6R,7S,4 R)-( )-(L-Valinyl-L-alanyl)-2,-diamino-8-(1 -carbamoyl-2 - iminoimidazolidin-4 -yl)-6,7-dihydroxy-octanate; Guadinomine B 2 t-bu Boc Boc Boc Boc 6 1) PMP isocyanate Benzene, r.t. PMP Boc Boc Boc Boc t-bu 2) CA MeC / 2, 0 C 2 Boc Boc Boc Boc t-bu ) TFA / 2, r.t. 60% for steps " 2" 1" 1"' 2"' 2 1' 7 6 4' "' TFA 2 2 2 ' Guadinomine B 2 40