The Journal of Organic Chemistry
Article
(C-6′), 54.9 (C-6T), 37.0 (C-6), 31.2 (CH2−triazole), 25.7, 25.6, 25.5,
25.4, 25.3, 25.2, 25.1 (CH3CO-). Anal. Calcd for C82H108N6O47S4: C,
47.86; H, 5.29; N, 4.08; S, 6.23. Found: C, 48.09; H, 5.59; N, 3.87; S,
6.02. HRMS (ESI): m/z [M + H]+ calcd for C82H109N6O47S4:
2057.5206, found: 2057.5236.
Hz, H-6′a), 3.63 (m, 2 H, H-5′, H-6′b), 3.56 (dd, 1 H, J3′,4′ = 3.3, J2′,3′
= 9.5 Hz, H-3′), 3.52 (t, 1 H, J1′,2′ = J2′,3′ = 9.6 Hz, H-2′), 3.50 (m, 1 H,
H-5), 3.42 (t, 1 H, J2,3 = J3,4 = 9.0 Hz, H-3), 3.39 (dd, 1 H, J1T,2T = 3.9,
J2T,3T = 9.9 Hz, H-2T), 3.32 (m, 2 H, H-2, H-4), 3.15 (t, 1 H, J3T,4T
=
J4T,5T = 9.6 Hz, H-4T), 2.89 (dd, 1 H, J5,6a = 2.1, J6a,6b = 14.3 Hz, H-
6a), 2.56 (dd, 1 H, J5,6b = 8.1, J6a,6b = 14.4 Hz, H-6b); 13C NMR (125
MHz, D2O) δ = 144.8 (C-4 triazole), 125.4 (C-5 triazole), 93.0 (C-
1G), 83.3 (C-1′), 82.7 (C-1), 79.4 (C-5), 79.2 (C-5′), 77.0 (C-3), 73.9
(C-3′), 72.6 (C-3T), 72.4, 72.3 (C-2, C-4), 70.9 (C-4G), 70.7 (C-2G),
70.4 (C-5G), 69.7 (C-2′), 68.8 (C-4′), 61.2 (C-6′), 51.0 (C-6T), 32.3
(C-6), 25.6 (CH2-triazole). Anal. Calcd for C42H68N6O27S4·H2O: C,
40.84; H, 5.71; N, 6.80; S, 10.38. Found: C, 40.90; H, 6.02; N, 7.12; S,
10.33. HRMS (ESI): m/z [M + H]+ calcd for C42H68N6NaO27S4:
1239.2913, found: 1239.2899.
Compound 21. Yield: 588 mg, 73%; [α]20D +22.6 (c = 0.3, CHCl3);
Rf = 0.31 (Hexane/EtOAc 1:8); 1H NMR (500 MHz, CDCl3
+
DMSO-d6) δ = 7.99, 7.89 (2 × H-triazole), 5.37−5.25 (m, 6 H, 2 × H-
3, 2 × H-4′, H-3C, H-3′C), 5.15 (m, 8 H, 2 × H-1, 2 × H-1′, 2 × H-3′,
H-1C, H-1′C), 5.00−4.85 (m, 9 H, 2 × H-2, 2 × H-4, 2 × H-2′, H-2C,
H-2′C, H-4′C), 4.81−4.50 (m, 4 H, H-6aC, H-6′aC, −6bC, H-6′bC),
4.30−3.85 (m, 12 H, 2 × H-5, 2 × H-5′, 2 × H-6′a, 2 × H-6′b, 2 ×
CH2-triazole), 3.70−3.50 (m, 3 H, H-4C, H-5C, H-5′C), 2.85 (m, 2 H,
2 × H-6a), 2.62 (m, 2 H, 2 × H-6b), 2.13, 2.01, 2.00, 1.99, 1.98, 1.97
(2 × ), 1.96, 1.95, 1.94, 1.93, 1.92, 1.91, 1.90 (13 s, 57H, 19 ×
CH3CO); 13C NMR (125 MHz, CDCl3 + DMSO-d6) δ = 170.4,
170.3, 170.2, 170.1, 170.0, 169.8, 169.7, 169.6 (-OCOCH3), 145.1,
144.9 (2 × C-4 triazole), 124.5, 124.0 (2 × C-5 triazole), 99.8 (C-
1′C), 81.0, 80.0 (2 × C-1′, 2 × C-1, C-1C), 77.6, 77.2, 76.9 (2 × C-5,
C-4C, C-5C), 74.2 (2 × C-5′), 73.6 (2 × C-3), 73.1, 72.7 (C-3C, C-
3′C, C-5′C), 71.5 (2 × C-3′), 71.3 (2 × C-2), 70.7 (2 × C-4), 70.5,
70.4, 70.3 (C-2C, C-2′C, C-4′C), 67.8 (2 × C-2′, 2 × C-4′), 61.5 (2 ×
C-6′), 50.7, 50.0 (C-6C, C-6′C), 32.7, 32.4 (2 × C-6), 26.5, 26.3 (2 ×
CH2-triazole), 21.0, 20.6, 20.5, 20.4, 20.3, 20.2 (CH3CO-). Anal. Calcd
for C160H210N12O90S9: C, 47.69; H, 5.25; N, 4.17; S, 7.16. Found: C,
47.42; H, 5.22; N, 4.45; S, 6.89. HRMS (ESI): m/z [M + 2Na]2+ calcd
for C160H210N12Na2O90S9: 2036.4748, found: 2036.4689.
Compound 22. Yield: 224 mg, 92%; [α]20 −41.3 (c = 0.4, H2O);
D
Rf = 0.15 (BuOH/EtOH/H2O, 2.5:1:1, double development); 1H
NMR (500 MHz, D2O) δ = 8.02, 7.81 (2s, 2 H, 2 × H-triazole), 4.80
(m, 4 H, 2 × H-1, H-6′aC, H-6aC), 4.66 (d, 2 H, J1′,2′ = 9.6 Hz, 2 × H-
1′), 4.60 (dd, 1 H, J5C,6bC = 7.8, J6aC,6bC = 14.7 Hz, H-6bC), 4.50 (d, 1
H, J1′C,2′C = 7.9 Hz, H-1′C), 4.30 (dd, 1 H, J5′C,6b′C = 9.5, J6a′C,6b′C
=
14.4 Hz, H-6b′C), 4.05 (d, 1 H, J1C,2C = 10.1 Hz, H-1C), 3.88 (m, 6 H,
2 × H-4′, 2 × CH2-triazole), 3.78 (m, 1 H, H-5′C), 3.70 (m, 2 H, 2 ×
H-6′a), 3.64 (m, 4 H, 2 × H-5′, 2 × H-6′b), 3.60 (m, 2 H, 2 × H-3′),
3.56−3.46 (m, 7 H, H-3C, H-4C, H-3′C, 2 × H-2′, 2 × H-5), 3.41 (m,
4 H, 2 × H-3, H-2C, H-2′C), 3.34 (m, 4 H, 2 × H-2, 2 × H-4), 3.20
(m, 2 H, H-4′C, H-5C), 2.95 (m, 2 H, 2 × H-6a), 2.65 (m, 2 H, 2 ×
H-6b); 13C NMR (125 MHz, D2O) δ = 145.0, 143.9 (2 × C-4
triazole), 125.4, 125.1 (2 × C-5 triazole), 102.1 (C-1′C), 83.3 (2 × C-
1′), 83.2 (2× C-1), 82.7 (C-1C), 79.2, 79.1 (2 × C-5, 2 × C-5′), 78.6
(C-3C), 77.0 (2 × C-3, C-5C), 75.6, 75.3 (C-4C, C-3′C), 74.0 (2 × C-
3′), 72.8, 72.5, 72.4, 72.3, 72.0 (C-2C, C-2′C, C-5′C, 2 × C-2, 2× C-
4), 70.7 (C-4′C), 69.7 (2 × C-2′), 68.8 (2 × C-4′), 61.2 (2 × C-6′),
50.9 (C-6C, C-6′C), 33.1, 32.6 (2 × C-6), 26.0, 25.8 (2 × CH2-
triazole). Anal. Calcd for C84H134N12O52S9.H2O: C, 41.17; H, 5.59; N,
6.86; S, 11.78. Found: C, 40.90; H, 5.32; N, 7.02; S, 11.63. HRMS
(ESI): m/z [M + 2Na]2+ calcd for C84H134N12Na2O52S9: 1238.2740,
found: 1238.2738.
General Procedure for the O-Deacetylation. Compounds 9,
17, 19, and 21 (0.10 mmol) were suspended in a mixture of MeOH/
Et3N/H2O 4:1:5 (10 mL) and stirred at room temperature. The solid
was progressively dissolving, and after 4−6 h TLC (EtOAc or EtOAc/
MeOH, 9:1) showed complete consumption of the starting material.
The solution was concentrated, and the residue was dissolved in water
(1 mL) and passed through a column filled with Dowex MR-3C mixed
bed ion-exchange resin. The eluate was concentrated and further
purified by filtration through an Octadecyl C18 minicolumn.
Evaporation of the solvent afforded the free product, which showed
a single spot by TLC (n-BuOH/EtOH/H2O, 2.5:1:1) whose Rf are
indicated in each case.
Deacetylation of Compound 9. Compound 23. Thiodisacchar-
ide 9 (100 mg, 0.14 mmol) was dissolved in 0.5 M NaOMe in MeOH
(0.4 mL). The mixture was stirred for 2 h at room temperature. The
solution was neutralized with acidic ion-exchange resin Dowex 50X
[H+], filtered, and concentrated. The residue obtained was purified by
reverse-phase chromatography to afford compound 23 (52 mg, 89%).
[α]20D −19.6 (c = 0.4, H2O); Rf = 0.55 (BuOH/EtOH/H2O, 2.5:1:1);
1H NMR (500 MHz, D2O) δ = 4.80 (d, 1 H, J1,2 = 10.0 Hz, H-1), 4.69
(d, 1 H, J1′,2′ = 9.7 Hz, H-1′), 3.90 (dd, 1 H, J4′,5′ < 1, J3′,4′ = 3.3 Hz, H-
4′), 3.70 (dd, 1 H, J5′,6′a = 8.8, J6′a,6′b = 12.6 Hz, H-6′a), 3.65 (dd, 1 H,
Compound 18. Yield: 54 mg, 85%; [α]20D −29.1 (c = 0.4, H2O); Rf
= 0.27 (BuOH/EtOH/H2O, 2.5:1:1); 1H NMR (500 MHz, D2O) δ =
7.96 (s, 1 H, H-triazole), 4.80 (d, 1 H, J1,2 = 10.1 Hz, H-1), 4.75 (dd, 1
H, J5G,6aG = 2.3, J6aG,6bG = 14.6 Hz, H-6aG), 4.69 (d, 1 H, J1′,2′ = 9.6 Hz,
H-1′), 4.66 (d, 1 H, J1G,2G = 3.8 Hz, H-1G), 4.53 (dd, 1 H, J5G,6bG
=
8.2, J6aG,6bG = 14.6 Hz, H-6bG), 3.91 (dd, 1 H, J4′,5′ < 1, J3′,4′ = 3.3 Hz,
H-4′), 3.88 (d, 2 H, J = 5.6 Hz, CH2-triazole), 3.83 (ddd, J5G,6aG = 2.3,
J5G,6bG = 8.2, J4G,5G = 9.6 Hz, H-5G), 3.70 (dd, 1 H, J5′,6′a = 8.9, J6′a,6′b
=
12.4 Hz, H-6′a), 3.65 (m, 2 H, H-5′, H-6′b), 3.60 (dd, 1H, J3′,4′ = 3.3,
J5′,6′b = 2.1, J6′a,6′b = 12.6 Hz, H-6′b), 3.63 (ddd, 1 H, J4′,5′ < 1, J5′,6′b
=
J
2′,3′ = 9.3 Hz, H-3′), 3.58 (t, 1 H, J2G,3G = J3G,4G = 9.7 Hz, H-3G), 3.53
2.1, J5′,6′a = 8.8 Hz, H-5′), 3.60 (dd, 1H, J3′,4′ = 3.3, J4′,5′ = 9.5 Hz, H-
3′), 3.57 (ddd, 1 H, J5,6a = 2.5, J5,6b = 8.7, J4,5 = 9.5 Hz, H-5), 3.53 (t, 1
H, J1′,2′ = J2′,3′ = 9.6 Hz, H-2′), 3.42 (t, 1 H, J2,3 = J3,4 = 9.0 Hz, H-3),
3.35 (dd, 2 H, J = 2.6 Hz, CH2S), 3.34 (t, 1 H, J1,2 = J2,3 = 10.0 Hz, H-
2), 3.33 (t, 1 H, J3,4 = J4,5 = 9.4 Hz, H-4), 3.16 (dd, 1 H, J5,6a = 2.5, J6a,6b
= 14.4 Hz, H-6a), 2.76 (dd, 1 H, J5,6b = 8.4, J6a,6b = 14.4 Hz, H-6b),
2.62 (t, 1 H, JCCH,CH2CCH = 2.6 Hz, CCH); 13C NMR (125
MHz, D2O) δ = 83.0 (C-1′), 82.3 (C-1), 80.8 (CCH), 79.2 (C-5′),
79.0 (C-5), 77.0 (C-3), 73.9 (C-3′), 72.5, 72.4 (C-2, C-4), 72.2 (C
CH), 69.6 (C-2′), 68.8 (C-4′), 61.1 (C-6′), 32.6 (C-6), 19.3 (CH2S).
Anal. Calcd for C15H24O9S2·H2O: C, 41.85; H, 6.09; S, 14.90. Found:
C, 42.00; H, 6.22; S,15.23. HRMS (ESI): m/z [M + H]+ calcd for
C15H24NaO9S2: 435.0759, found: 435.0756.
(t, 1 H, J1′,2′ = J2′,3′ = 9.6 Hz, H-2′), 3.49 (m, 1 H, H-5), 3.46 (dd, 1 H,
J1G,2G = 3.8, J2G,3G = 9.8 Hz, H-2G), 3.41 (t, 1 H, J2,3 = J3,4 = 9.0 Hz, H-
3), 3.33 (t, 1 H, J1,2 = J2,3 = 9.7 Hz, H-2), 3.32 (t, 1 H, J3,4 = J4,5 = 9.3
Hz, H-4), 3.17 (t, 1 H, J3G,4G = J4G,5G = 9.6 Hz, H-4G), 2.88 (dd, 1 H,
J5,6a = 2.2, J6a,6b = 14.2 Hz, H-6a), 2.58 (dd, 1 H, J5,6b = 8.1, J6a,6b = 14.2
Hz, H-6b); 13C NMR (125 MHz, D2O) δ = 145.0 (C-4 triazole),
125.3 (C-5 triazole), 99.1 (C-1G), 83.2 (C-1′), 82.6 (C-1), 79.2 (C-
5′), 79.1 (C-5), 77.0 (C-3), 74.0 (C-3G), 73.0 (C-3′), 72.5, 72.3 (C-2,
C-4), 71.1 (C-2G), 70.9 (C-4G), 69.9 (C-5G), 69.7 (C-2′), 68.8 (C-
4′), 61.2 (C-6′), 54.8 (-OCH3), 50.9 (C-6G), 32.6 (C-6), 25.6 (CH2-
triazole). Anal. Calcd for C22H37N3O14S2·H2O: C, 40.67; H, 6.05; N,
6.47; S, 9.87. Found: C, 40.90; H, 6.32; N, 6.62; S, 9.73. HRMS (ESI):
m/z [M + H]+ calcd for C22H38N3O14S2: 632.1790, found: 632.1799.
Compound 20. Yield: 97 mg, 80%; [α]20D −49.6 (c = 0.4, H2O); Rf
= 0.10 (BuOH/EtOH/H2O, 2.5:1:1); 1H NMR (500 MHz, D2O) δ =
7.92 (s, 1 H, H-triazole), 4.80 (d, 1 H, J1,2 = 10.0 Hz, H-1), 4.73 (dd, 1
H, J5T,6aT = 2.4, J6aT,6bT = 14.7 Hz, H-6aT), 4.68 (d, 1 H, J1′,2′ = 9.6 Hz,
H-1′), 4.49 (dd, 1 H, J5T,6bT = 8.4, J6aT,6bT = 14.7 Hz, H-6bT), 4.47 (d,
1 H, J1T,2T = 4.0 Hz, H-1T), 3.94 (ddd, 1 H, J5T,6aT = 2.3, J5T,6bT = 8.4,
J4T,5T = 9.8 Hz, H-5T), 3.88 (m, 3 H, H-4′, CH2-triazole), 3.69 (t, 1 H,
J2T,3T = J3T,4T = 9.6 Hz, H-3T), 3.67 (dd, 1 H, J5′,6′a = 8.9, J6′a,6′b = 12.3
Enzymatic Assays. Stability of the Glycoclusters toward β-
Galactosidase. Escherichia coli β-galactosidase (grade VIII, EC
3.2.1.23, 117 U/mg) was purchased from Sigma-Aldrich. A solution
of the glycoclusters (1 mM) in sodium phosphate buffer (100 mM, pH
7.3, MgCl2 1.2 mM, 2-mercaptoethanol 100 mM) was incubated with
the β-galactosidase from E. coli (0.6 U/mL) at 37 °C. Aliquots of the
solution were taken after 8, 16, and 24 h, evaporated, and examined by
TLC and NMR. o-Nitrophenyl-β-D-galactopyranoside was used for the
control experiment.
J
dx.doi.org/10.1021/jo500883v | J. Org. Chem. XXXX, XXX, XXX−XXX