Supporting Information

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1 Supporting Information A Combined Effect of the Picoloyl Protecting Group and Triflic Acid in Sialylation Samira Escopy, Scott A. Geringer and Cristina De Meo * Department of Chemistry Southern Illinois University Edwardsville Science Building West, Box 1652, Edwardsville, Illinois , cdemeo@siue.edu Contents: General Experimental Synthesis of Sialyl Donors 3a,b Synthesis of Diacetylated Sialyl Donors 8a and 8b Synthesis of Sialyl Donor 15 Synthesis of Disaccharides NMR Spectra S2 S2 S5 S6 S7 S11 S1

2 General Experimental The reactions were performed using commercial reagents (Aldrich, Acros, Carbosynth) and solvents purified according to standard procedures. Column chromatography was performed on silicagel 60 (Silicycle, mesh) or Sephadex LH-20 (GE Healthcare); reactions were monitored by TLC on TLC Silica Gel 60 F254S (Millipore). The compounds were detected by examination under UV light and by charring with 10% sulfuric acid in methanol. Solvents were removed under reduced pressure at <40 C. CH 2 Cl 2 and CH 3 CN were purified by MBraun solvent purification system (MB-SPS-800). Molecular sieves (3Å), used for reactions, were crushed and activated in vacuo at 390 C during 8 h in the first instance and then for 2-3 h at 390 C directly prior to application. ptical rotations were measured at Jasco P-1020 polarimeter. 1 H NMR spectra were recorded using a Bruker Ascend 400 MHz NMR Spectrometer and 13 C NMR spectra were recorded at 75 MHz. The 1 H NMR chemical shifts are referenced to the signal of the residual CHCl 3 (δ H = 7.27 ppm) for solutions in CDCl 3. The 13 C NMR chemical shifts are referenced to the central signal of CDCl 3 (δ C = ppm) for solutions in CDCl 3. HRMS determinations were made with the use of JEL MStation (JMS-700) Mass Spectrometer. Synthesis of Sialyl Donors 3a,b Methyl (phenyl 5-acetamido-3,5-dideoxy-8,9--isopropylidene-4--picoloyl-2-thio-Dglycero-α-D-galacto-non-2-ulopyranosid)onate (2a). To a solution of methyl (phenyl 5-acetamido-4,7,8,9-tetra--acetyl-3,5-dideoxy-2-thio-Dglycero-α-D-galacto-non-2-ulopyranosid)onate (1a 1, mg, 1.2 mmol) in MeH (66 ml), Ac Ac C 2 Me Ac SPh AcHN Ac 1a H AcHN H C 2 Me S1 SPh S2 C 2 Me H AcHN SPh Pico 2a freshly prepared MeNa (1.9 ml, 1M solution in MeH) was added and the solution stirred for 1 h, then neutralized with Dowex H +. the Dowex was then filtered off, rinsed with MeH, and concentrated in vacuo and dried. The residue was then dissolved in MeCN (11 ml), then 2,2- dimethoxypropane (1.1g, 6.3 mmol) and camphorsulfonic acid (14.6 mg, mmol) were added. The mixture was allowed to stir for 1 h under Ar at rt. The reaction was then neutralized with trimethylamine (TEA) and concentrated in vacuo. The residue was then purified by column chromatography on silica gel (ethanol/methylene chloride, 2% gradient), to afford methyl (1) Kirchner, E.; Thiem, F.; Dernick, R.; Heukeshoven, J.; Thiem, J. J. Carbohydr. Chem. 1988, 7, 453.

3 (phenyl 5-acetamido-3,5-dideoxy-8.9--isopropylidene-2-thio-D-glycero-α-D-galacto-non-2- ulopyranosid)onate (S1, mg, 0.1 mmol, solid, white, 79% from 1a). EDC (569.5 mg, 2.7 mmol), DMAP (33.7 mg, 0.2 mmol), and Picolinic acid (339.6 mg, 2.7 mmol) were added to a solution of S1 (628.3 mg, 1.3 mmol) in CH 2 Cl 2 (18.8 ml). The mixture was stirred under argon for 2 h at rt. The resulting residue was then diluted with CH 2 Cl 2 and washed with brine (15 ml), sat. aq. sodium bicarbonate (15 ml), and brine (15 ml). The organic phase was dried over MgS 4, filtered, and concentrated in vacuo. The residue was purified by column chromatography on silica gel (acetone-methylene chloride, 10% gradient elution) to afford 2a (673.1 mg, 1.2 mmol, solid, white, 87%). Analytical data for 2a: R f = 0.55 (acetone/methylene chloride, 1/1, v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 8.74 (dd, J = 5.1 Hz, 1H, aromatic), 8.05 (d, J = 7.8 Hz, 1H, aromatic), 7.86 (td, J = 7.8, Hz, 1H, aromatic), (m, 2H, aromatic), 7.49 (ddd, J = 7.6, 4.7, 1.1 Hz, 1H, aromatic), (m, 3H, aromatic), 6.44 (d, J NH,5 = 8.0 Hz, 1H, NH), 5.37 (td, J 4,5 = 11.3, J 3eq,4 = 5.0 Hz, 1H, H-4), (m, 1H, 7-H), 4.30 (dd, J 7,8 = 13.8 Hz, J 8,9b = 6.4 Hz, 1H, H-8), (m, 2H H-5, 9a), 4.01 (dd, J 9a,9b = 8.5 Hz, 1H, H-9b), 3.55 (s, 3H, CH 3 ), 3.49 (d, J 6,7 = 7.5 Hz, 1H, H-7), 3.39 (dd, J 5,6 = 10.5 Hz, 1H, H-6), 3.00 (dd, J 3eq,3ax = 12.7 Hz, 1H, H-3eq), 2.24 (dd, J 3ax,4 = 11.7 Hz, 1H, H-3ax), 1.95 (s, 3H, NHCCH 3 ), 1.38 (s, 3H, CCH 3 ), 1.28 (s, 3H, CCH 3 ) ppm; 13 C NMR: δ, 172.8, 168.9, 165.6, 150.2, 146.8, 137.2, 136.8, 129.9, 129.0, 128.7, 127.5, 125.6, 108.7, 87.2, 74.5, 70.7, 70.3, 67.5, 52.3, 51.6, 37.6, 26.8, 25.4, 23.1 ppm; HR-FAB MS [M+H] + calcd for C 27 H 33 N 2 9 S , found Methyl (phenyl 5-acetamido-7,8,9-tri--acetyl-3,5-dideoxy-4--picoloyl-2-thio-D-glycero-α- D-galacto-non-2-ulopyranosid)onate (3a). Sialoside 2a (653 mg, 1.1 mmol) was dissolved in CH 2 Cl 2 (9.3 ml) and a solution of 10% TFA in CH 2 Cl 2 (2.3 ml) was added dropwise, followed by 9 drops of water. The mixture was allowed to stir under argon for 30 minutes, followed by neutralization with TEA. The residue was then concentrated in vacuo and carried on without further purification. The residue was then dissolved in pyridine (9 ml), and Ac 2 (4.3 ml) was added dropwise and allowed to stir for 16h at rt. The reaction was then quenched with MeH until no further heat was detected, concentrated in vacuo, and coevaporated with toluene. The resulting compound was purified by column chromatography on silica gel (acetone/hexane, 10% gradient) to afford 3a (498.4 mg, 0.7 mmol, solid, white, 67%). Analytical data for 3a: R f = 0.56 (acetone/toluene, 3/2, v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 8.80 (d, J = 4.7 Hz, 1H, aromatic), 8.06 (d, J = 7.8 Hz, 1H, aromatic), 7.83 (td, J = 7.7, 1.7 Hz, 1H, aromatic), (m, 3H, aromatic), (m, 3H, aromatic), 6.00 (d, J NH,5 = 9.7 Hz, 1H, NH), 5.41 (dd, J 8,9a = 1.9 Hz, 1H, H-8), (m, 1H, H-7), 5.26 (dd, J 4,5 = 10.9, J 3eq,4 = 4.1 Hz, 1H, H-4), 4.44 (dd, J = 12.4, 2.7 Hz, 1H, H-6), 4.25 (dd, J 8,9b = 6.1 Hz, 1H, H-9b), (m, 1H, H-5), 4.10 (dd, J 9a,9b = 10.7 Hz, 1H, H-9a), 3.54 (s, 3H, CH 3 ), 3.02 (dd, J 3eq,3ax = 12.7 Hz, 1H, H-3eq), (m, 7H, NHCCH 3, CCH 3, H- 3ax), 2.06, 2.05 (s, 6H, CCH 3 ) ppm; 13 C NMR: δ, 207.0, 170.7, 170.4, 170.2, 170.1, 167.8, 164.5, 150.3, 147.1, 137.3, 136.5, 129.9, 128.8, 128.6, 127.3, 125.6, 87.6, 74.59, 71.2, 70.1, 67.9, 62.1, 54.3, 52.7, 49.1, 38.3, 32.4, 30.9, 29.3, 23.1, 21.0, 20.8, 20.8 ppm; HR-FAB MS [M+Na] + calcd for C 30 H 34 N 2 12 SNa , found S3

4 Methyl (ethyl 5-acetamido-3,5-dideoxy-8,9--isopropylidene-4--picoloyl-2-thio-D-glyceroα-D-galacto-non-2-ulopyranosid)onate (2b). To a solution of methyl (ethyl 5-acetamido-4,7,8,9-tetra--acetyl-3,5-dideoxy-2-thio-D-glyceroα-D-galacto-non-2-ulopyranosid)onate (1b 2, mg, 1.4 mmol) in MeH (71 ml), freshly prepared 1M solution of NaMe (2.1 ml) was added dropwise. The resulting mixture was stirred for 1 h under argon and then neutralized with Dowex H +. The Dowex was then filtered off, rinsed with MeH, and concentrated in vacuo and dried. The residue was then dissolved in MeCN (11 ml), then 2,2-dimethoxypropane (1.3 g, 7.4 mmol) and camphorsulfonic acid (17.2 mg, 0.1 mmol) were added. The mixture was allowed to stir for 1 h under Ar at rt. The reaction was then neutralized with TEA and concentrated in vacuo. The residue was then purified by column chromatography on silica gel (ethanol/methylene chloride, 2% gradient) to afford methyl (ethyl 5-acetamido-3,5-dideoxy-8.9--isopropylidene-2-thio-D-glycero-α-D-galacto-non-2- ulopyranosid)onate (S2, mg, 1.2 mmol, solid, white, 87%). EDC (536.5 mg, 2.6 mmol), DMAP (31.77 mg, 0.2 mmol), and Picolinic acid (320.0 mg, 2.6 mmol) were added to a solution of S2 (525.8 mg, 1.3 mmol) in CH 2 Cl 2. The mixture was stirred under argon for 2 h at rt. The resulting residue was then diluted with CH 2 Cl 2 and washed with brine (15 ml), sat. aq. sodium bicarbonate (20 ml), and brine (20 ml). The organic phase was dried over MgS 4, filtered, and concentrated in vacuo. The residue was purified by column chromatography on silica gel (acetone-methylene chloride, 10% gradient elution) to afford 2b (545.9 mg, 1.1 mmol, solid, white, 82%). Analytical Data for 2b: R f = 0.55 (acetone/methylene chloride, 1/1, v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 8.76 (d, 1H, aromatic), 8.07 (d, 1H, aromatic), 7.87 (m, 1H, aromatic), 7.50 (m, 1H, aromatic), 6.40 (d, 1H, J NH, 5 = 8.0 Hz, NH), 5.36 (td, 1H, J 4, 5 = 11.4 Hz, H-4), 4.36 (q, 1H, J 8, 9a = 6.2 Hz, 1H, 8-H), 4.20 (q, 1H, J 5, 6 = 10.5 Hz, H-5), 4.15 (dd, 1H, J 9a, 9b = 6.3 Hz, H-9a), 4.08 (dd, 1H, J 9b, 9a = 6.1 Hz, H-9b), 3.86 (s, 3H, CH 3 ), 3.56 (dd, 1H, J 7,8 = 7.0, H-7), 3.43 (dd, 1H, J 6,7 = 10.5, H-6), 2.94 (d, 1H, J 3eq, 4 = 5.0, H-3eq), 2.87 (m, 1H, SCH 2a CH 3 ), 2.74 (m, 1H, SCH 2b CH 3 ), 2.25 (t, 1H, J 3eq, 3ax = 11.7 Hz, H-3ax), 1.95 (s, 3H, NHCCH 3 ), 1.43, 1.40 (s x2, 6H, C(CH 3 ) 2 ), 1.25 (t, 3H, J = 7.5 Hz, SCH 2 CH 3 ) ppm; 13 C NMR: δ, , , , , , , , , , 83.59, 74.92, 70.65, 69.94, 67.08, 52.74, 51.76, 37.67, 26.93, 25.59, 23.60, 23.12, ppm; HR-FAB MS [M+H] + calcd for C 23 H 33 N 2 9 S , found (2) Tsvetkov, Y. E.; Nifantiev, N. E. Synlett 2005, 9, Methyl (ethyl 5-acetamido-7,8,9-tri--acetyl-3,5-dideoxy-4--picoloyl-2-thio-D-glycero-α- D-galacto-non-2-ulopyranosid)onate (3b). Sialoside 2b (526 mg, 1.0 mmol) was dissolved in CH 2 Cl 2 (8.2 ml), a solution of 10% TFA in CH 2 Cl 2 (2.0 ml) was added dropwise, followed by 8 drops of water. The mixture was allowed to stir under argon for 30 S4

5 minutes, followed by neutralization with TEA. The residue was then concentrated in vacuo and carried on without further purification. The residue was then dissolved in pyridine (7 ml), and Ac 2 (3.5 ml) was added dropwise and allowed to stir for 16 h at rt. The reaction was then quenched with MeH until no further heat was detected, concentrated in vacuo and coevaporated with toluene. The resulting compound was purified by column chromatography on silica gel (acetone/hexane, 10% gradient) to afford 3b (531.7, 0.8 mmol, solid, white, 86% from 2b) Analytical Data for 3b: R f = 0.56 (acetone/toluene, 3/2 v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 8.81 (d, J = 4.7 Hz, 1H, aromatic), 8.08 (d, 1H, J = 7.8 Hz, aromatic), 7.86 (td, 1H, J = 1.7 and 7.8, Hz, aromatic), 7.52 (ddd, 1H, J = 7.6, 4.7, 1.1 Hz, aromatic), 5.79 (d, 1H, J NH,5 = 9.7 Hz, NH), 5.42 (bs, 2H, H-7,8), 5.30 (td, 1H, J 3ax,4 = 11.5 Hz, H-4), 4.37 (d, 1H, J = 11.7 Hz, H- 9a), 4.25 (dd, 1H, J 4,5 = 10.6 Hz, H-5), (m, 1H, H-9b), 4.04 (d, 1H, J 5,6 = 10.7 Hz, H- 6), 3.82 (s, 3H, CH 3 ), 2.93 (dd, 1H, J 3eq,3ax = 12.6 Hz, J 3eq,4 = 4.7 Hz, H-3eq), (m, 1H, SCH 2a CH 3 ), 2.59 (tt, 1H, J = 12.3, 8.3 Hz, SCH 2b CH 3 ), 2.19, 2.17, 2.06, 1.80 (4 s, 13H, NHCCH 3, 3 x CCH 3, H-3ax), 1.22 (t, 3H, J = 7.5 Hz, SCH 2 CH 3 ) ppm; 13 C NMR: δ, 170.7, 170.5, 170.3, 170.2, 168.4, 164.6, 150.2, 137.4, 127.4, 125.6, 83.4, 74.0, 71.2, 68.8, 67.5, 62.3, 52.9, 49.4, 38.2, 31.0, 23.3, 23.2, 21.2, 20.9, 20.8, 14.1 ppm; HR-FAB MS [M+H] + calcd for C 26 H 35 N 2 12 S , found Synthesis of Diacetylated Sialyl Donor 8a Methyl (phenyl 7,8,9-tri--acetyl-5-(N-acetyl)acetamido-3,5-dideoxy-4--picoloyl-2-thio-Dglycero-α-D-galacto-non-2-ulopyranosid)onate (8a) To a solution of 3a (267 mg, 0.4 mmol) in isopropenyl acetate (2.0 ml), toluenesulfonic acid (4.0 mg, 0.02 mmol) was added. The solution was allowed to reflux at 65 o C for 16 h under Ar atmosphere. The reaction was then neutralized with TEA, concentrated in vacuo and purified by column chromatography on silica gel (acetone/hexane, 10% gradient) to afford 8a (278 mg, 0.4 mmol, solid, white, 98%) Analytical data for 8a: R f = 0.5 (acetone/hexane, 1/1, v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 8.72 (dd, 1H, J = 5.9, 1.2 Hz, aromatic), 8.04 (d, 1H, J = 7.8 Hz, aromatic), 7.83 (td, 1H, J = 7.7, 1.7 Hz, aromatic), (m, 2H, aromatic), 7.48 (ddd, 1H, J = 7.6, 4.7, 1.2 Hz, aromatic), (m, 3H, aromatic), 5.83 (td, 1H, J 4,5 = 11.0 Hz, H-4), 5.23 (ddd, 1H, J 8,9a = 5.2 Hz, J 8,9b = 2.7 Hz, H-8), 5.16 (dd, 1H, J 7,8 = 7.7 Hz, H-7), 4.92 (dd, 1H, J 5,6 = 10.1, J 6,7 = 1.5 Hz, H-6), 4.35 (dd, 1H, J 9a,9b = 12.5 Hz, H-9a), (m, 2H, H-5, H-9b), 3.63 (s, 3H, CH 3 ), 3.18 (dd, 1H, J 3eq,3ax = 12.8, J 3eq,4 = 5.2 Hz, H-3eq), 2.34, 2.31, 2.17, 2.04, 2.03 (s, 16H, N(CCH 3 ) 2, CCH 3, H-3ax) ppm; 13 C NMR δ 174.6, 174.0, 170.6, 170.1, 169.9, 167.6, 163.9, 150.0, 147.3, 137.1, , 130.0, 128.8, 128.7, 127.2, 125.4, 87.4, 72.0, 69.8, 68.6, 67.5, 61.6, 57.4, 52.8, 39.2, 31.7, 29.3, 28.0, 25.9, 20.9, 20.9 ppm; HR-FAB MS [M+Na] + calcd for C 32 H 36 N 2 13 S , found Synthesis of Diacetylated Sialyl Donor 8b Methyl (ethyl 7,8,9-tri--acetyl-5-(N-acetyl)acetamido-3,5-dideoxy-4--picoloyl-2-thio-Dglycero-α-D-galacto-non-2-ulopyranosid)onate (8b) S5

6 To a solution of 3b (220.8 mg, 0.3 mmol) in isopropenyl acetate (1.8 ml), toluene sulfonic acid (3.51 mg, 0.02 mmol) was added. The solution was allowed to reflux at 65 o C for 16 h under Ar. The reaction was then neutralized with TEA, concentrated in vacuo and purified by column chromatography on silica gel (acetone/hexane, 10% gradient) to afford 8b (214.4mg, 0.33 mmol, solid, white, 91%) Analytical Data for 8b: Rf = 0.5 (acetone/hexane, 1/1 v/v); [α]23d (c = 1, CHCl3); 1H NMR: δ, 8.72 (dd, J = 4.7 Hz, 1H, aromatic), 8.03 (d, J = 7.8 Hz, 1H, aromatic), 7.83 (td, J = 7.7, 1.7 Hz, 1H, aromatic), 5.83 (td, 1H, H-4), 5.37 (ddd, J 8, 9b = 4.8, J 8,9a = 2.7 Hz, 1H, H-8), 5.19 (dd, J 7,8 = 8.3 Hz, 1H, H-7), 4.89 (dd, J 6,7 = 1.8 Hz, 1H, H-6), 4.40 (t, 1H, H-5), 4.31 (dd, J 9a,9b = 12.5 Hz, 1H, H-9a), 4.18 (dd, 1H,H-9b), 3.89 (s, 3H, CH 3 ), 3.05 (dd, J H3eq,H3ax = 12.6, J 3eq,4 5.1 Hz, 1H, H-3eq), (m, 1H, SCH 2 CH 3 ), (m, 1H, SCH 2 CH 3 ), 2.38, 2.29, 2.16, 2.13, 2.03, 1.99 (6 s, 18H, N(CCH 3 ) 2, CCH 3 ), 2.13 (dd, J H3ax,4 = 11.3 Hz, 1H, H-3ax), 1.23 (t, J = 7.5 Hz, 3H, SCH 2 CH 3 ) ppm: 13 C NMR δ , , , , , , 83.34, 71.72, 68.61, 67.32, 67.19, 61.78, 57.14, 52.99, 39.22, 30.95, 28.01, 26.00, 23.38, 21.16, 21.03, 20.77, ppm: HR-FAB MS [M+Na] + calcd for C 28 H 36 N 2 13 SNa , found Methyl (phenyl 5-acetamido-7,8,9-tri--acetyl-3,5-dideoxy-4--benzoyl-2-thio-D-glycero-α- D-galacto-non-2-ulopyranosid)onate (15). Sialoside S1 (327 mg, 0.7 mmol) was H AcHN H C 2 Me SPh S1 Ac Ac Ac AcHN Bz C 2 Me dissolved in pyridine (4 ml) and benzoic anhydride was added (325 mg, 1.4 mmol) under Ar and the solution was stirred overnight upon completion. The reaction was then quenched with MeH until no further heat was detected, concentrated in vacuo, and coevaporated with toluene. The resulting compound was purified by column chromatography on silica gel (acetone/toluene, 10% gradient), and used directly for the removal of isopropilidene at C-8,9 by treatment with a solution of 10% TFA in CH 2 Cl 2 (0.3 ml) was added dropwise to a solution in CH 2 Cl 2 (3 ml), followed by 3 drops of water. The mixture was allowed to stir under argon for 10 minutes, followed by neutralization with TEA. The residue was then concentrated in vacuo and carried on without further purification. The residue was then dissolved in pyridine (4 ml), and Ac 2 (2 ml) was added dropwise and allowed to stir for 16h at rt. The reaction was then quenched with MeH until no further heat was detected, concentrated in vacuo, and coevaporated with toluene. The resulting compound was purified by column chromatography on silica gel (acetone/hexane, 10% gradient) to afford 15 (265.0 mg, 0.4 mmol, solid, white, 76%). Analytical data for 15: R f = 0.56 (acetone/toluene, 3/2, v/v); [α] 23 D (c = 1, CHCl 3 ); 1 H NMR: δ, 7.97 (dd, 2H, aromatic), (m, 3H, aromatic), (m, 5H, aromatic), 5.35 (dd, J 7.8 = 10.7 Hz, 1H, H-7), (m, 1H, H-8), 5.14 (d, J NH,5 = 9.7 Hz, 1H, NH), (m, J 4,5 = 10.8 Hz, 1H, H-4), 4.00 (dd, J 6,7 = 2.7 Hz, 1H, H-6), (m, J 8,9b = 7.7 Hz, 2H, H-5, H-9 b ), 4.42 (dd, J 9a,9b = 12.55, J 9a,8 = 2.8 Hz 1H, H-9a), 3.60 (s, 3H, CH 3 ), 3.02 (dd, J 3eq,3ax = 12.7 Hz, 1H, H-3eq), 2.17, 2.07, 2.06, 1.76 (4 s, 12H, NHCCH 3, CCH 3 ) ppm; 15 SPh S6

7 13 C NMR: δ, 170.0, 167.9, 166.5, 136.4, 133.5, 129.8, 129.1, 128.9, 128.6, 128.5, 87.6, 74.9, 70.5, 67.6, 52.7, 49.2, 38.4, 23.1, 21.0, 20.8, 20.8 ppm; HR-FAB MS [M+Na] + calcd for C 31 H 35 N 12 SNa , found Synthesis of Disaccharides General Sialylation procedure: Sialyl donors and acceptors were stirred overnight in methylene chloride, with molecular sieves 3Å, at -45 C under argon atmosphere. The following day, N- iodosuccinimide (NIS) and trifluoromethane sulfonic acid (TfH) were added and reaction monitored by TLC until disappearance of the acceptor or the donor. The solution was then diluted with CH 2 Cl 2, filtered through Celite and washed with a solution of 10% sodium thiosulfate and brine. The solution was then dried over magnesium sulfate, followed by filtration and concentration under vacuum. Isolation of the desired product was performed by size exclusion column (Sephadex LH-20) or in some cases, silica gel chromatography. 1,2:3,4-Di--isopropylidene-6--[methyl (5-acetamido-7,8,9-tri--acetyl-3,5-dideoxy-4-picoloyl-D-glycero-α-D-galacto-non-2-ulo-pyranosylonate]-α-D-galactopyranose (6). A solution of 3a (20.0 mg, 0.03 mmol), 1,2:3,4-di--isopropylidene-α-D-galactopyranose (4, 4.0 mg, 0.01 mmol), and molecular sieves 3Å (70 mg) in methylene chloride (0.6 ml) was stirred overnight at -45 C under argon atmosphere. The next morning, N-iodosuccinimide (14 mg, 0.06 mmol) was added followed by triflic acid (9.2 mg, 0.06 mmol, 6 µl) and stirred for 45 minutes until acceptor was consumed. Workup: dilute with methylene chloride, filter through Celite, wash with 10% sodium thiosulfate (5 ml) and brine (2 x 5 ml). The solution was then dried over magnesium sulfate, filtered, and concentrated in vacuo. Sephadex LH20 was performed on the residue to afford 6 (11.5 mg, 0.01 mmol, oil, colorless, 93%). Analytical data for 6: R f = 0.46 (acetone/toluene, 3/2 v/v); [α] 26 D (c = 1, CHCl 3 ); 1 H NMR: δ, 5.52 (d, 1H, J 1,2 =5.1 Hz, H- 1), 5.58 (bs, 1H, NH), (ddd, 1H, J 8,9a = 2.7 Hz, J 8,9b =5.1 Hz, J 7,8 = 7.9 Hz, H-8 ), 5.39 (dd, 1H, J 7,6 =1.3 Hz, H-7 ), (m, 1H, J 3eq,4 =4.6 Hz, J 4, 5 =12.7 Hz, H-4 ) 4.60 (dd, 1H, J 3,2 = 2.3 Hz, H-3), (m, 5H, H-6,H-9a',H-2, H-4, H-5 ), 4.18 (dd, 1H, J 9a,9b =12.6 Hz, H-9b ) (m, 2H, H-5, H-6 a ) (m, 1H, H-6 b ), 3.81 (s, 3H, C 2 CH 3 ), 2.83 (dd, 1H, J 3 eq,3 ax =12.8 Hz, J 3,4 =4.7 Hz, H-3 eq ), 2.10 (dd,1h, H-3 ax ), 2.15(x2), 2.05, 1.79 (4s, 12H, NHCCH 3, CCH 3 ), 1.54, 1.43, 1.33, 1.32 (4s, 12H, CCH 3 ) ppm; 13 C NMR for 5α: δ, 170.1, 169.8, 167.9, 128.6, 128.4, 126.1, 109.1, 108.4, 98.6, 96.3, 77.7, 71.7, 70.7, 70.6, 68.6, 67.4, 66.4, 63.0, 62.1, 52.7, 49.9, 37.8, 29.7, 29.2, 26.0, 24.9, 24.6, 23.3, 21.1, 20.8 ppm: HR- FAB MS [M+Na] + calcd for C 36 H 48 N 2 18 Na , found ,2:3,4-Di--isopropylidene-6--[methyl (7,8,9-tri--acetyl-5-(N-acetyl)acetamido-3,5- dideoxy-4--picoloyl-d-glycero-α-d-galacto-non-2-ulo-pyranosylonate]-α-dgalactopyranose (10). S7

8 A solution of 8 (13.8 mg, mmol), 4 (2.6 mg, mmol), and 3Å molecular sieves (48 mg) in methylene chloride (0.40 ml) was stirred overnight at -45 C under argon atmosphere. The next morning, N-iodosuccinimide (9.0 mg, 0.04 mmol) was added followed by triflic acid (0.60 mg, mmol, 0.36 µl) and stirred for 4 hours until disappearance of the acceptor. Workup: dilute with methylene chloride, filter through Celite, wash with 10% sodium thiosulfate (5 ml) and brine (5 ml x2). Dried over magnesium sulfate, filter, and concentrated in vacuo to afford 10 (7.9 mg, mmol, oil, colorless, 94%). Analytical data for 10: Rf = 0.46 (acetone/hexane, 1/1 v/v); [α] 26 D (c = 1, CHCl3); 1H NMR δ; (d, 1H, aromatic) (dd, 1H, aromatic) (ddd, 1H, aromatic) (m, 1H, aromatic) (m, 1H, J 4,3ax = 5.4 Hz, J 3eq,4 = 5.3 Hz, H-4 ), 5.53 (d, 1H, J 1,2 = 5.0 Hz, H-1), (m, 1H, J = 2.5, 4.8, 7.8 Hz, H-8 ), 5.05 (dd, 1H, J 6,5 = 10.1 Hz, H-6 ), 5.20 (dd, 1H, J 7,6 = 1.8 Hz, H-7 ), 4.60 (dd, 1H, J = 8.0 Hz, H-3), (m, 5H, H-2, H-4, H-5, H-9 a,b ), (m, 2H, H-5, H-6 a ), 3.88 (s, 3H, C 2 CH 3 ), 3.71 (dd, 1H, J 6a,b = 8.3 Hz, J 6b,5 = 4.3 Hz, H-6 b ), 2.98 (dd, 1H, J 3 eq,3 ax =12.8 Hz, H-3 eq ), 2.14 (dd, 1H, H-3 ax ), 2.16, 2.15, 2.05 (3s, 9H, CCH 3 ), 1.53, 1.45, 1.35, 1.34 (4s, 12H, CCH 3 ) 2.40, 2.34 (2s, 6H, N(CCH 3 ) 2 ppm: 13 C NMR δ 174.7, 170.6, 170.0, 170.0, 167.6, 149.9, 137.3, 127.2, 125.3, 109.2, 108.5, 98.7, , 70.8, 70.6, 70.5, 69.8, 69.1, 68.5, 67.3, 66.7, 63.1, 61.7, 57.2, 52.8, 38.4, 30.9, 28.1, 26.1, 26.0, 24.9, 24.5, 21.1, 20.9, 20.8 ppm: HR-FAB MS [M+Na] + calcd for C 28 H 36 N 2 13 S , found Methyl 2,3,4-tri--benzyl-6--[methyl (7,8,9-tri--acetyl-5-acetamido-3,5-dideoxy-4-picoloyl-D-glycero-α-D-galacto-non-2-ulo-pyranosylonate]-α-D-galactopyranoside (13). A solution of 3a (13.0 mg, 0.02 mmol), methyl 2,3,4-tri--benzyl-α-D-galactopyranoside (11, 4.6 mg, 0.01 mmol), and 3Å molecular sieves (45 mg) in methylene chloride (0.4 ml) was stirred overnight at -45 C under argon atmosphere. The next morning, N-iodosuccinimide (9 mg, 0.04 mmol) was added followed by triflic acid (6 mg, 0.04 mmol, 3 µl) and stirred for 45 min. Workup: dilute with methylene chloride, filter through Celite, wash with 10% sodium thiosulfate (5 ml) and brine (5 ml x2). The solution was then dried over magnesium sulfate, filtered, and concentrated in vacuo. Sephadex LH20 was performed on the residue to afford 13 (7 mg, mmol, solid, white, 70%). Analytical Data for 13: R f = 0.46 (acetone/toluene, 3/2 v/v); [α] 26 D (c = 1, CHCl 3 ); 1 H NMR δ; (d, 1H, aromatic) (d, 1H, aromatic) (t, 1H, aromatic) (m, 1H, aromatic) (m, 20H, aromatic), (m, 3H, H-7, H-8, NH), 5.26 (m, 1H, H-4 ), (m, 7H, H-1, CH 2 (x3)), (m, 4H, J 9 b,8 = 4.9 Hz, J 9 a,9 b = 12.6 Hz, J 9 a,8 = 2.4 Hz, H-9 a,b, H-5, H-6 ), 4.03 (dd, 1H, J 1,2 = 5.5 Hz, H-2), (m, 4H, J 5,4 = 2.7 Hz, H-3, H-4, H-6 a,b ), 3.48 (ddd, 1H, J 5,6 = 8.6, H-5) 2.78 (dd, 1H, J 3 eq,3 ax =13.0 Hz, J 3eq,4 = 4.8 Hz, H-3 eq ), 2.09 (dd,1h, H-3 ax ), 3.68 (s, 3H, C 2 CH 3 ), 3.39 (s, 3H, CH 3 ), 2.15, 2.13, 2.00, 1.79 (4s, 12H, CCH 3 ) ppm: 13 C NMR for 5α: δ: S8

9 170.6, 170.1, 169.8, 167.8, 138.9, 138.4, 128.3, 128.1, 127.8, 127.6, 127.4, 127.3, 98.7, 78.9, 77.2, 76.3, 74.5, 73.5, 68.8, 68.3, 67.3, 63.3, 62.1, 55.3, 52.8, 49.9, 37.9, 30.9, 29.7, 23.3, 21.1, 20.9, 20.7 ppm: HR-FAB MS [M+Na] + calcd for C 52 H 60 N 2 18 Na , found Methyl 2,3,4-tri--benzoyl-6--[methyl (7,8,9-tri--acetyl-5-acetamido-3,5-dideoxy-4-picoloyl-D-glycero-α-D-galacto-non-2-ulo-pyranosylonate]-α-D-galactopyranoside (14). A solution of 3a (10.0 mg, 0.01 mmol), methyl 2,3,4-tri--benzoyl-α-D-galactopyranoside (12, 4.0 mg, 0.01 mmol), and 3Å molecular sieves (35 mg) in methylene chloride (0.3 ml) was stirred overnight at -45 C under argon atmosphere. The next morning, N-iodosuccinimide (7 mg, 0.01 mmol) was added followed by triflic acid (5 mg, 0.03 mmol, 3 µl) and stirred for 45 min. Workup: dilute with methylene chloride, filter through Celite, wash with 10% sodium thiosulfate (5 ml) and brine (5 ml x2). The solution was then dried over magnesium sulfate, filtered, and concentrated in vacuo. Sephadex LH20 was performed on the residue to afford 14 (4 mg, mmol, solid, white, 51%). Analytical Data for 14: R f = 0.46 (acetone/toluene, 3/2 v/v); [α] 26 D (c = 1, CHCl 3 ); 1 H NMR δ; (d, 1H, aromatic) (dd, 3H, aromatic) (d, 3H, aromatic) (t, 1H, aromatic) (d, 2H, aromatic) (t, 1H, aromatic) (m, 8H, aromatic) 5.99 (d, 1H, H-4), 5.68 (s, 1H, J 2,1 = 7.7 Hz, H-2), 5.65 (dd, 1H, J 3,4 = 3.4, 10.4 Hz, H-3), 5.45 (m, 1H, J 8,9a = 2.3, 5.9, 8.8 Hz, H-8 ), 5.33 (dd, 2H, J 7,6 = 2.0 Hz, J 7,8 = 9.1 Hz, H-7, NH) 5.18 (m, 1H, H-4 ), 4.78 (d, 1H, H-1) 4.39 (dd, 1H, J 9a b = 12.4 Hz, H-9 a ), (m, 2H, H-6, H-6 a ), 4.19 (t, 1H, H-5 ), 4.10 (dd, 1H, J 9b,8 = Hz, H-9 b ), 3.98 (ddd, 1H, J 6b,a = 6.1, J= 10.0 Hz, H-6 b ), 3.55 (m, 1H, H-5), 2.65 (dd, 1H, J 3eq,4 = 4.6 Hz, H-3 eq ), 2.10 (dd, 1H, J 3 eq,3 ax = 13.1 Hz, H-3 ax ), 3.60 (s, 3H, C 2 CH 3 ), 3.43 (s, 3H, CH 3 ) 2.23, 2.13, 2.05, 1.79 (4s, 12H, CCH 3 ) ppm: 13 C NMR for 5α: δ: 170.7, 170.5, 169.9, 165.4, 165.3, 165.2, 133.2, 133.1, 133, 129.8, 129.8, 129.7, 129.5, 129, 102.3, 98.9, 77.2, 71.9, 69.9, 68.1, 67.7, 67.3, 62.7, 57.3, 53.4, 52.6, 30.9, 29.7, 23.2, 21, 20.9, 20.8, 20.7 ppm: HR-FAB MS [M+Na] + calcd for C 52 H 54 N 2 21 Na , found ,2:3,4-Di--isopropylidene-6--[methyl (5-acetamido-7,8,9-tri--acetyl-3,5-dideoxy-4-benzoyl-D-glycero-α-D-galacto-non-2-ulo-pyranosylonate]-α-D-galactopyranose (16). A solution of 15 (12.0 mg, 0.02 mmol), 1,2:3,4-di--isopropylidene-α-D-galactopyranose (4, 2.4 mg, 0.01 mmol), and molecular sieves 3Å (42 mg) in methylene chloride (0.4 ml) was stirred overnight at -45 C under argon atmosphere. The next morning, N-iodosuccinimide (8.4 mg, 0.04 mmol) was added followed by triflic acid (5.5 mg, 0.04 mmol, 3 µl) and stirred for 15 minutes until acceptor was consumed. Workup: dilute with methylene chloride, filter through S9

10 Celite, wash with 10% sodium thiosulfate (5 ml) and brine (2 x 5 ml). The solution was then dried over magnesium sulfate, filtered, and concentrated in vacuo. Sephadex LH20 was performed on the residue to afford 16 (4.6 mg, mmol, oil, colorless 62%). Analytical data for 16: R f = 0.46 (acetone/toluene, 3/2 v/v); [α] 26 D (c = 1, CHCl 3 ); Selected 1 H NMR: δ, 5.45 (m, J 3eq,4 = 5.1 Hz, 1H, H-4 β ), 5.05 (m, J 3eq,4 = 4.9 Hz, 1H, H-4 α ), 4.29 (m, 1H, H-5 β ), 4.26 (m, 1H, H-5 α ), 2.81 (dd, J 3 eq, 3 ax =12.8 Hz, H-3 eq.α ), 2.09 (dd,1h, H-3 ax.α ), 2.68 (dd, J 3 eq, 3 ax=13.2 Hz, H-3 eq. β ), 2.00 (dd,1h, H-3 ax.β ) ppm; 13 C NMR for 16: δ, 170.7, (x2), 170.4, 170.2, 170.1, 169.8, 167.1, 166.7, 166.5, 133.4, 129.3, 128.5, 109.6, 108.8, 108.5, 98.7, 96.3, 72.9, 70.0, 69.6, 68.9, 67.4, 66.3, 63.0, 62.7, 62.1, 61.7, 52.8, 52.7, 49.3, 37.9, 37.4, 30.9, 29.7, 26.0, 25.9, 24.9, 24.7, 23.2, 23.1,21.0, 20.8 ppm: HR-FAB MS [M+Na] + calcd for C 37 H 49 N 18 Na , found S10

11 H AcHN Pico C 2 Me 2a SPh CDCl 3 400MHz CDCl 3 75MHz S11

12 H AcHN Pico C 2 Me 2a SPh S12

13 Ac Ac Ac AcHN Pico C 2 Me 3a SPh CDCl 3 400MHz CDCl 3 75MHz S13

14 Ac Ac Ac AcHN Pico C 2 Me 3a SPh S14

15 H AcHN Pico C 2 Me SEt 2b CDCl 3 400MHz CDCl 3 75MHz S15

16 H AcHN Pico C 2 Me SEt 2b S16

17 Ac Ac Ac AcHN Pico C 2 Me 3b SEt CDCl 3 400MHz S17

18 CDCl 3 75MHz Ac Ac Ac AcHN Pico C 2 Me SEt 3b S18

19 Ac Ac Ac Ac 2 N Pico C 2 Me 8a SPh CDCl 3 400MHz CDCl 3 75MHz S19

20 Ac Ac Ac Ac 2 N Pico C 2 Me 8a SPh S20

21 CDCl 3 400MHz CDCl 3 75MHz S21

22 Ac Ac Ac Ac 2 N Pico C 2 Me 8b SEt CDCl 3 400MHz S22

23 CDCl 3 400MHz S23

24 CDCl 3 75MHz CDCl 3 400MHz S24

25 Ac Ac Ac 2 N Ac C 2 Me Pico 10 α:β3.8:1 CDCl 3 400MHz CDCl 3 75MHz S25

26 Ac Ac C 2 Me Ac Ac 2 N Pico 10 α:β3.8:1 S26

27 CDCl 3 400MHz CDCl 3 75MHz S27

28 S28

29 CDCl 3 400MHz CDCl 3 75MHz S29

30 S30

31 Ac Ac AcHN Ac Bz C 2 Me 15 SPh CDCl 3 400MHz CDCl 3 75MHz S31

32 Ac Ac AcHN Ac Bz C 2 Me 15 SPh S32

33 Ac Ac Ac AcHN Bz 16 C 2 Me CDCl 3 400MHz CDCl 3 75MHz S33

34 Ac Ac Ac AcHN Bz 16 C 2 Me S34