10
S. Hackbusch et al. / Carbohydrate Research 458-459 (2018) 1e12
127.85 (4C), 127.74, 127.70 (2C), 102.71 (C-1), 84.83 (C-3), 82.40 (C-
2), 77.99 (C-4), 75.81, 75.10, 75.00 (2C, C-5), 73.59, 71.25, 69.04 (C-
6). HRMS (DART-MS): m/z calculated for C41H46NO6 [MþNH4]þ
648.3320, found 648.3272.
give a pale yellow suspension, which was stirred for another 5 min.
The crude reaction mixture was then filtered and the retained
solids washed with saturated NaHCO3 (3 mL) and DCM (4 mL). The
organic layer was separated and the aqueous layer extracted with
DMC (3 ꢂ 4 mL). The combined organic layers were dried over
Na2SO4 to give a clear pale yellow solution, which was evaporated
with a rotavap to give a yellow sticky oil. The desired product was
isolated via column chromatography (95:5 to 8:2 hexane: EtOAc
gradient elution) to yield a colorless oil which solidified under high
vacuum (51.2 mg, 0.05 mmol, 27%). Rf 0.86 (8:2 hexane:EtOAc). 1H
4.1.2. Benzyl-6-O-acetyl-2,3,4-tri-O-benzyl-b-D-gluco-pyranoside 2
Based on literature procedures [48]. In a 3-neck flask with gas
adapter, placed ZnCl2 (10.7 g, 78.2 mmol) and heated under vacuum
to drive off residual water. After cooling under N2, added a mixture
of acetic acid:acetic anhydride (62 mL, 1:5) and stirred with cooling
on an icebath. To this, added benzyl 2,3,4,6-tetra-O-benzyl-
b-D
-
NMR (600 MHz, CDCl3): d 7.33e7.18 (m, 40H, Bn-CH), 4.94 (d,
glucopyranoside 1 (as a solution in acetic acid:acetic anhydride
(1:5, 62 mL)). Allowed the reaction mixture to come to RT slowly
over the course of 90 min. After another hour of stirring at RT, the
clear yellow reaction mixture was poured into approximately
300 mL of an ice-water mixture to precipitate the benzyl 6-O-
11.4 Hz, 2H, Bn-CH2), 4.91 (d, 10.8 Hz, Bn-CH2), 4.89 (d, 10.8 Hz, Bn-
CH2), 4.87 (d, 11.4 Hz, Bn-CH2), 4.84 (d, 10.8 Hz, Bn-CH2), 4.79 (d,
12.0 Hz, Bn-CH2), 4.78 (13.8 Hz, Bn-CH2), 4.78 (d, 10.8 Hz, Bn-CH2),
4.77 (d, 10.8 Hz, Bn-CH2), 4.70 (d, 10.8 Hz, Bn-CH2), 4.70 (d, 10.8 Hz,
Bn-CH2), 4.69 (d,10.8 Hz, Bn-CH2), 4.59 (d, 12.0 Hz, Bn-CH2), 4.58 (d,
10.8 Hz, Bn-CH2), 4.57 (d, 8.1 Hz, H-10), 4.55 (d, 12.3 Hz, Bn-CH2),
4.53 (dd, 1.8 Hz, 12.0 Hz, H-6proS), 4.49 (d, 8.4 Hz, H-1), 4.33 (dd,
5.1 Hz, 11.7 Hz, H-6proR), 3.95 (t, 8.4 Hz, H-50), 3.92 (t, 8.4 Hz, H-40),
3.66 (t, 8.7 Hz, H-30), 3.65 (t, 8.7 Hz, H-3), 3.60 (dd, 9.0 Hz, H-4), 3.58
(apt t, 8.4 Hz, H-20), 3.54 (ddd, 1.8 Hz, 4.8 Hz, 9.4 Hz, H-5), 3.51 (dd,
acetyl-2,3,4-tri-O-benzyl-b-D-glucopyranoside as a white solid
(6.2 g after drying). The crude compound was used without puri-
fication in the subsequent reaction. 1H NMR (600 MHz, CDCl3):
d
7.40e7.21 (m, 20H, Bn-CH), 5.02e4.92 (m, 3H, Bn-CH2), 4.84e4.49
(m, 6H, 5x CH2-Bn, H-1), 4.35 (dd, 1.2 Hz, 11.7 Hz, 1H, H-6a), 4.24
(dd, 4.3 Hz,11.7 Hz, H-6b), 3.67 (t, 8.3 Hz, H-3), 3.61e3.48 (m, 3H, H-
7.8 Hz, 9.0 Hz, H-2). 13C NMR (150 MHz, CDCl3):
d
168.34 (C¼O, C-
2, H-4, H-5), 2.05 (s, 3H, COCH3). 13C NMR (150 MHz, CDCl3):
d
170.6
60), 138.51, 138.44, 138.37, 138.25, 138.00, 137.84, 137.20, 137.05,
128.6e127.7 (40C, Bn-CH), 102.87 (C-10), 102.39 (H-1), 84.59 (C-3),
83.89 (C-30), 82.39 (C-2), 82.01 (C-20), 79.30 (C-40), 77.83 (C-4),
75.83 (2C), 75.31, 75.11, 75.05 (2C), 74.92 (C-50), 72.80 (C-5), 71.35,
71.06, 64.35 (C-6). HRMS (ESI-MS): m/z calculated for C61H61O12Na
[MþNaeC7H7]þ 1009.4133, found 1009.4240.
(C¼O), 138.3, 138.1, 137.6, 137.0, 128.4, 128.3, 128.2, 128.0, 127.9,
127.8, 127.7, 127.6, 102.2 (C-1), 84.5 (C-3), 82.0 (C-5), 77.2 (C-2), 75.6
(CH2), 74.8 (CH2), 74.7 (CH2), 72.7 (C-4), 71.0 (CH2), 63.0 (C-6), 20.8
(CH3).
4.1.3. Benzyl-2,3,4-tri-O-benzyl-b-D-glucopyranoside 3
Based on literature procedures [48]. In a 250 mL round-bottom
4.1.5. Gluco-pyranosyl-6,60 gluco-pyranuronate 5
flask, dissolved sodium metal (0.3 g) in dry methanol (70 mL). To
this, crude benzyl 6-O-acetyl-2,3,4-tri-O-benzyl-b-D-glucopyrano-
In a 50 mL three-neck round bottom flask, under nitrogen gas
atmosphere, added Pd/C (14 mg) in ethyl acetate (2.5 mL) and
rinsed the flask walls with methanol (1 mL). To this, 4 (83.9 mg,
side 2 was added and stirred at RT for 5 h, at which point TLC (2:1
hexane:ethyl acetate) indicated complete conversion of the starting
material. The clear yellow reaction mixture was poured onto
approximately 280 mL of ice-cold water to give a milky white
suspension, which was allowed to settle overnight. After cooling on
ice, the white solid product was filtered and washed with addi-
77 mmol) dissolved in methanol (2 mL) was added. Evaporated and
filled the flask with nitrogen gas thrice, then repeated with
hydrogen gas thrice, before attaching a double-walled balloon with
hydrogen gas and stirring the reaction mixture overnight. Checked
for completion by TLC (95:5 DCM:MeOH). After 24 h, removed the
hydrogen gas atmosphere and replaced with nitrogen. After filtra-
tion of the crude product mixture, the flask was rinsed with
methanol (1 mL). Washed the filter cake with an additional amount
of methanol (5 mL). The combined filtrates were evaporated using a
rotavap and the product was separated using column chromatog-
raphy (dryloading, silica, DCM:MeOH 95:5 / 7:3) to yield a clear
tional ice-cold water to give benzyl 2,3,4-tri-O-benzyl-b-D-gluco-
pyranoside (4.5 g, 57% over two steps). The product was of sufficient
purity as determined by 1H NMR, when compared to material
recrystallized from ethanol. 1H NMR (600 MHz, CDCl3):
d 7.38e7.26
(m, 20H, Bn-CH), 4.95 (d, 10.8 Hz, Bn-CH2), 4.93 (d, 11.4 Hz, Bn-CH2),
4.92(d, 12.0 Hz, Bn-CH2), 4.86 (d, 10.8 Hz, Bn-CH2), 4.80 (d, 10.8 Hz,
Bn-CH2), 4.72 (d,10.8 Hz, Bn-CH2), 4.69 (d, 12.0 Hz, Bn-CH2), 4.63 (d,
10.8 Hz, Bn-CH2), 4.57 (d, 7.8 Hz, H-1), 3.87 (dd, 3.0 Hz, 12.0 Hz, H-
6a), 3.70 (dd, 7.2 Hz, 12.0 Hz, H-6b), 3.67 (t, 9.0 Hz, H-3), 3.57 (t,
9.3 Hz, H-4), 3.48 (dd, 8.1 Hz, 9.3 Hz, H-2), 3.36 (ddd, 3.0 Hz, 4.8 Hz,
colorless crystalline solid (13.7 mg, 38 mmol, 49%) after evaporation
and drying under vacuum overnight. Rf 0.45 (7:3 DCM:MeOH). For
1H and 13C NMR (600 MHz, 150 MHz, D2O) assignment, see tabular
representation below. HRMS (ESI-MS): m/z calculated for
9.6 Hz, H-5). 13C NMR (150 MHz, CDCl3):
d
138.59, 138.39, 138.05,
C
12H20O12Na [MþNa]þ 379.0847, found 379.0939.
137.36, 128.59 (4 C), 128.50 (2 C), 128.47 (2 C), 128.24 (2 C), 128.17
(2 C), 128.04 (4 C), 127.98 (2 C), 127.80, 127.76, 102.93 (C-1), 84.63
(C-3), 82.43 (C-2), 77.65 (C-4), 75.81 (CH2), 75.16 (CH2), 75.14 (C-5),
75.09 (CH2), 71.77 (CH2), 62.17 (C-6). HRMS (DART-MS): m/z
calculated for C34H40NO6 [MþNH4]þ 558.2851, found 558.2884.
4.2. Computational methods
4.2.1. Conformer search
The initial conformer search for compounds 5 and 6 was per-
formed using the Spartan14 software suite [61,62]. The input ge-
ometry was restrained to the preferred chair conformation, with
twist and boat conformations excluded. The conformer distribution
4.1.4. Benzyl-2,3,4-tri-O-benzyl-
(benzyl 2,3,4-tri-O-benzyl- -gluco-pyranuronate) 4
Adapted from the literature [47]. In a scintillation vial, combined
benzyl 2,3,4-tri-O-benzyl- -glucopyranoside (200 mg,
0.37 mmol), pyridine (60 L, 0.74 mmol, 2.0 equiv.), TEMPO (3 mg,
b-D
-gluco-pyranosyl-(6 / 60)-
b-D
was
computed
using
MAXCYCLES
¼
1000
and
b-
D
3
MAXCONFORMERS ¼ 20000 with 100% conformers kept at the HF/
6-31G*//AM1 level of theory (implicit solvent model used). The
lowest energy conformer fitting the above criteria was chosen as
the starting geometry for the molecular dynamics computations.
m
0.02 mmol, 5 mol%) and then dissolved in anhydrous acetonitrile
(1 mL). In a separate vial, dissolved TCCA (94 mg, 0.40 mmol, 1.1
equiv) in anhydrous acetonitrile (1 mL) and slowly added the clear
solution to the reaction mixture over the course of several minutes.
Stirred for 90 min, then quenched with saturated NaHCO3 (5 mL) to
4.2.2. Molecular dynamics
Molecular dynamics simulations of aa/bb-5 and aa/bb-6 were