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2869
28. Melvin, L. S.; Pfizer Inc. (Pfiz); p 79168-B.
(B) CH3CN. The gradient consisted of 10–35% B for 50 min at a flow rate of
0.5 mL/min. The column was washed with 100% B during 10 min and then
stabilized with the initial conditions during another 10 min.
29. Data for 6: 1H NMR (400.14 MHz, CDCl3), d (ppm): 7.63 (dd, J = 2.1, 8.4 Hz, 1H),
7.59 (d, J = 2.1, 1H), 7.49–7.29 (m, 5H, Bn), 6.92 (d, J = 8.4 Hz, 1H), 5.19 (s, 2H,
CH2Bn), 4.27 (s, 2H, CH2I), 3.96 (s, 3H, OCH3). 13C NMR (100.62 MHz, CDCl3), d
(ppm): 191.6 (C@O), 154.5 (C), 148.4 (C), 136.4 (C), 128.7 (2CH), 128.1 (CH),
127.5 (2CH), 126.4 (C), 124.2 (CH), 113.5 (CH), 110.5 (CH), 71.0 (CH2Bn), 56.1
(OCH3), 1.3 (CH2I); ESI-MS m/z: 383 [M+H]+.
35. Cyanidin-40-O-methyl-3-glucoside (9): 2,4-Diacetoxy-6-hydroxybenzaldehyde 2
(7.9 mg, 0.0332 mmol) and 2-(2,3,4,6-tetra-O-acetyl-b-
D-glucopyranosyloxy)-
30-benzyloxy-40-methoxyacetophenone
8
(20 mg, 0.0332 mmol) were
dissolved in dry EtOAc (1.5 mL) and anhydrous HCl (g) was bubbled through
the solution. The reaction was stirred from 0 °C to RT and a red color gradually
developed. The mixture was then stirred until all the starting materials were
consumed as monitored by TLC. No purification was undertaken at this stage.
The solvent was evaporated and the residue dissolved in MeOH. Debenzylation
was performed using 10 equiv of triethylsilane in 20% Pd/C at RT for 10 min.
The catalyst was removed by filtration. Complete deacetylation was performed
with KOH (3 equiv) in MeOH/H2O 1:1. The mixture was stirred at RT for 15 min
and then carefully acidified to pH 1 with HCl 1 M. After MeOH evaporation, the
aqueous solution was filtered to remove the majority of KCl precipitate and
further extracted with ethyl acetate to remove traces of toluene. The aqueous
fraction was eluted on silica gel C18-reversed phase to remove the acetic acid
and the desired compound was recovered in acidic MeOH. The final
purification was made by column chromatography using a TSK Toyopearl
HW-40 (S) gel (150 ꢁ 16 mm i.d.). The metabolite was eluted with 10%
aqueous methanol acidified with 2% HCl at a flow rate of 0.8 mL/min. The
fraction collection was made upon visual detection of the red band. After
removal of methanol on a rotary evaporator and freeze–drying, compound 9
was obtained as a red solid (3 mg, 18%). 1H NMR (600.13 MHz, CD3OD/TFA
98:2), d (ppm): 9.10 (s, 1H), 8.36 (dd, J = 2.3, 8.8 Hz, 1H), 8.02 (d, J = 2.3 Hz, 1H),
7.21 (d, J = 8.8 Hz, 1H), 6.93 (d, J = 1.6 Hz, 1H), 6.68 (d, J = 1.6 Hz, 1H), 5.31 (d,
J = 7.7 Hz, 1H), 4.04 (s, 3H, OCH3), 3.91 (dd, J = 2.2, 12.1 Hz, 1H), 3.72–3.65 (m,
2H), 3.56 (m, 1H), 3.54 (t, J = 9.1 Hz, 1H), 3.42 (t, J = 9.4 Hz, 1H). 13C NMR
(125.77 MHz, CD3OD/TFA 98:2), d (ppm): 169.6 (C), 162.7 (C), 156.7 (C), 156.6
(C),154.7 (C), 147.1 (C), 144.4 (C), 136.7 (CH), 127.9 (CH), 121.1 (C), 116.4 (CH),
112.7 (C), 111.5 (CH), 102.4 (CH), 102.1 (CH), 93.8 (CH), 77.5 (CH), 76.7 (CH),
73.4 (CH), 69.7 (CH), 61.0 (CH2), 55.5 (OCH3); LC-DAD/ESI-MS m/z: 463 [M]+.
31. Data for 8: 1H NMR (400.14 MHz, CDCl3), d (ppm): 7.56–7.29 (m, 7H), 6.90 (d,
J = 8.9 Hz, 1H), 5.22 (t, J = 9.5 Hz, 1H), 5.18 (s, 2H, CH2Bn), 5.12–5.04 (m, 2H),
4.89–4.75 (m, 2H), 4.67 (d, J = 7.9 Hz, 1H), 4.23 (dd, J = 4.6, 12.4 Hz, 1H), 4.15–
4.08 (m, 2H, CH2Oglc), 3.94 (s, 3H, OCH3), 2.07 (s, 3H, Ac), 2.02 (s, 3H, Ac), 2.00
(s, 6H, 2 ꢁ Ac); 13C NMR (100.62 MHz, CDCl3), d (ppm): 193.3 (C@O), 170.6
(C@O), 170.1 (C@O), 169.7 (C@O), 169.4 (C@O), 154.4 (C), 148.3 (C), 136.4 (C),
128.6 (2CH), 128.1 (CH), 127.7 (C), 127.5 (2CH), 123.2 (CH), 112.6 (CH), 110.6
(CH), 100.2 (CH), 72.6 (CH2Oglc), 71.9 (CH), 71.0 (CH2Bn), 70.9 (CH), 70.3 (CH),
68.3 (CH), 61.7 (CH2), 56.1 (OCH3), 20.6 (4 ꢁ CH3, Ac); ESI-MS m/z: 603 [M+H]+,
625 [M+Na]+.
32. LC-DAD/ESI-MS: LC-DAD/ESI/MS analyses were performed on
a Finnigan
Surveyor series liquid chromatograph equipped with Finnigan LCQ (Finnigan
Corp., S. J., Calif., USA) mass detector and an API source using an ESI interface.
The samples were analyzed on a reversed-phase column (150 ꢁ 4.6 mm, 5
lm,
C18) at 25 °C using the same eluents, gradients, and flow rates referred for
HPLC analysis. The capillary voltage was 4 V and the capillary temperature
275 °C. Spectra were recorded in positive and negative ion modes between m/z
120 and 1500. The mass spectrometer was programmed to do a series of three
scans: a full mass (MS), a zoom scan of the most intense ion in the first scan
(MS2), and a MS–MS of the most intense ion using relative collision energy of
30 and 60 (MS3).
34. HPLC–DAD: HPLC analyses were performed on a Merck–Hitachi L-7100 (Merck,
D., Germany) apparatus with a 150 ꢁ 4.6 mm i.d. reversed-phase ODS C18
column (Merck, Darmstadt) at 25 °C; detection was carried out using a L-
7450A diode array detector (DAD). The eluents were (A) H2O/HCOOH (9:1) and