S. Hanamura et al. / Journal of Molecular Catalysis B: Enzymatic 128 (2016) 19–26
21
and 150.83 included two carbons. HRMS (ESI+): calculated for [M
(C42H46O21) + Na]+, 909.2429; found, 909.2432.
(s, 2H, H-2, H-6), 7.01 (d, J = 16.4 Hz, 1H, CH CH), 7.09 (d, J = 8.5 Hz,
2H, H-3ꢀ, H-5ꢀ), 7.49 (d, J = 8.5 Hz, 2H, H-2ꢀ, H-6ꢀ); 13C NMR: ␦ 20.76,
20.82, 21.29, 62.18, 68.44, 71.21, 72.22, 72.79, 98.94, 109.69, 112.80,
114.63, 121.89, 127.54, 127.76, 129.65, 134.55, 139.84, 150.56,
151.78, 157.67, 169.33, 169.48, 169.55, 170.35, 170.72; the signals
20.76, 20.82, 21.29, 121.89, 127.76 and 169.55 included two car-
bons. HRMS (ESI+): calculated for [M (C32H34O14) + Na]+, 655.1846;
found, 665.1855. This was employed for the next step without fur-
ther purification.
2.4. 5-Hydroxy-4ꢀ-methoxy-7-[(2-O-˛-l-rhamnopyranosyl-ˇ-d-
glucopyranosyl)oxy]flavone
(1f)
To a solution of 1e (301 mg, 0.339 mmol) in methanol (3.39 mL)
was added a solution of sodium methoxide in methanol (5.0 M,
13 L, 0.2 equiv.). The mixture was stirred for 1 h at room temper-
ature, then the reaction was quenched by adding acetic acid. The
resulting precipitates were collected by filtration, and washed with
water. After drying, 1f (177 mg, 88%) was obtained as yellow solid.
1H NMR (CD3OD): ␦ 1.33 (d, J = 6.1 Hz, 3H, rhamnose-H-6), 3.41
(dd, J = 9.4, 9.4 Hz, seemingly t, 2H, rhamnose-H-4, glucose-H-4),
3.53–3.56 (m, 1H, glucose-H-5), 3.59–3.73 (total 4H, glucose-
H-2, glucose-H-3, glucose-H-6a, rhamnose-H-3), 3.89–3.94 (total
6H, glucose-H-6b, rhamnose-H-2, rhamnose-H-5, −OMe), 5.20 (d,
J = 7.6 Hz, 1H, glucose-H-1), 5.28 (brs, 1H, rhamnose-H-1), 6.46 (d,
J = 2.2 Hz, 1H, H-8), 6.70 (s, 1H, H-3), 6.79 (d, J = 2.2 Hz, 1H, H-6), 7.08
(d, J = 8.8 Hz, 2H, H-3ꢀ, H-5ꢀ), 7.97 (d, J = 8.8 Hz, 2H, H-2ꢀ, H-6ꢀ). This
was employed for the next step without further purification.
2.7. 5-Acetoxy-3-(tetra-O-acetyl-ˇ-d-glucopyranosyl)oxy-4ꢀ-
hydroxystilbene
(2e)
To a solution of 2d (3.29 g, 5.13 mmol) in a mixture of 2-propanol
(10 mL) and THF (20 mL), which was pre-dried over anhydrous
Na2SO4 at room temperature overnight, was added an immobi-
lized form of Burkholderia cepacia lipase (Amano PS-IM, 250 mg).
The mixture was stirred for 12 h at 22 ◦C. The mixture was filtered
to remove insoluble materials with a pad of Celite. The precipi-
tates were washed with ethyl acetate. The combined filtrate and
washings were concentrated in vacuo to afford 2e (3.07 g, quant.)
as white amorphous solid. 1H NMR: ␦ 2.04, 2.05, 2.08 (each s,
total 12H, sugar-OAc), 2.31 (s, 3H, 5-OAc), 3.90 (ddd, J = 2.0, 5.4,
8.8 Hz, 1H, glucose-H-5), 4.18 (dd, J = 2.0, 12.2 Hz, 1H, glucose-H-
6a), 4.28 (dd, J = 5.4, 12.2 Hz, 1H, glucose-H-6b), 5.10 (d, J = 7.1 Hz,
1H, glucose-H-1), 5.16 (dd, J = 9.3, 9.3 Hz, seemingly t, 1H, glucose-
H-4), 5.26–5.33 (total 2H, glucose-H-2, glucose-H-3), 6.61 (s, 1H,
H-4), 6.81 (d, J = 16.4 Hz, 1H, CH CH), 6.82 (d, J = 8.6 Hz, 2H, H-3ꢀ, H-
5ꢀ), 6.93 (s, 2H, H-2, H-6), 6.98 (d, J = 16.4 Hz, 1H, CH CH), 7.34 (d,
J = 8.6 Hz, 2H, H-2ꢀ, H-6ꢀ); 13C NMR: ␦ 20.69, 20.73, 20.78, 21.26,
62.18, 68.41, 71.23, 72.11, 72.78, 98.88, 109.04, 112.52, 114.31,
115.84, 124.82, 128.24, 129.20, 130.28, 140.42, 151.64, 156.38,
157.58, 169.69, 169.72, 170.48, 170.94; the signals 20.73,115.84,
128.24 and 169.69 included two carbons. HRMS (ESI+): calculated
for [M (C30H32O13) + Na]+, 623.1741; found, 623.1748. This was
employed for the next step without further purification.
2.5. 5,7-Dihydroxy-4ꢀ-methoxyflavone (acacetin, 1a)
To a solution of 1f (101 mg, 0.170 mmol) in ethanol (1.70 mL)
was added conc. sulfuric acid (170 L). The mixture was heated
under reflux for 2 h, then the reaction was cooled at 0 ◦C. The
resulting precipitates were recovered by filtration and washed with
water to afford 1a (42 mg, 87%) as yellow solid. This was recrystal-
lized from methanol to give an analytical sample of 1a as yellow
needles, mp 256.0–257.8 ◦C [lit. [3] mp 254.6–257.5 ◦C]; 1H NMR
(DMSO-d6): ␦ 3.84 (s, 3H, −OMe), 6.18 (d, J = 2.2 Hz, 1H, H-6), 6.49
(d, J = 2.0 Hz, 1H, H-8), 6.86 (s, 1H, H-3), 7.10 (d, J = 9.0 Hz, 2H, H-3ꢀ,
H-5ꢀ), 8.02 (d, J = 9.0 Hz, 2H, H-2ꢀ, H-6ꢀ), 10.86 (s, 1H, OH-7), 12.91
(s, 1H, OH-5); 13C NMR (DMSO-d6): ␦ 55.58, 94.06, 98.91, 103.56,
103.78, 114.61, 122.84, 128.36, 157.36, 161.46, 162.33, 163.33,
164.24, 181.81; the signals 114.61 and 128.36 included two car-
bons. The identity of 1a was further confirmed after derivation to
the corresponding diacetate (1g) by an acetylation in a conven-
tional manner. This was recrystallized from methanol to give an
analytical sample of 1g as colorless needles, mp 200.1–201.0 ◦C
[lit. [2] mp 199 ◦C]; 1H NMR: ␦ 2.35, 2.44 (each s, 6H, 5-OAc, 7-
OAc), 3.89 (s, 3H, −OMe), 6.61 (s, 1H, H-3), 6.83 (d, J = 2.2 Hz, 1H,
H-6), 7.01 (d, J = 8.8 Hz, 2H, H-3ꢀ, H-5ꢀ), 7.34 (d, J = 2.2 Hz, 1H, H-8),
7.82 (d, J = 8.8 Hz, 2H, H-2ꢀ, H-6ꢀ). Its 1H NMR spectrum was in good
accordance to that reported previously [3].
2.8. 5-Acetoxy-3-(tetra-O-acetyl-ˇ-d-glucopyranosyl)oxy-4ꢀ-
[tert-butyl(dimethyl)silyl]oxystilbene
(2f)
To a solution of 2e (3.00 g, 5.00 mmol) and imidazole (680 mg,
10.0 mmol, 2.0 equiv.) in anhydrous N,N-dimethylformamide
(25 mL) tert-butyldimethylsilyl (TBS) chloride (1.50 g, 10.0 mmol,
2.0 equiv.) was added at 0 ◦C, then the mixture was warmed to
room temperature. After stirring for 4 h, the reaction was quenched
by adding cold water. The organic materials were extracted with
ethyl acetate twice. The combined extracts were washed with brine,
dried over Na2SO4, and concentrated in vacuo. The residue was
purified by silica gel column chromatography (30 g, hexane/ethyl
acetate = 2:1) to afford 2f (3.54 g, 99%) as slightly yellow amorphous
solid. 1H NMR: ␦ 0.21 (s, 6H, Me), 0.99 (s, 9H, tert-Bu), 2.04, 2.05,
2.06, 2.08 (each s, total 12H, sugar-OAc), 2.31 (s, 3H, 5-OAc), 3.90
(ddd, J = 2.2, 5.6, 9.1 Hz, 1H, glucose-H-5), 4.18 (dd, J = 2.2, 12.2 Hz,
1H, glucose-H-6a), 4.28 (dd, J = 5.6, 12.2 Hz, 1H, glucose-H-6b), 5.12
(d, J = 7.4 Hz, 1H, glucose-H-1), 5.16 (dd, J = 9.6, 9.6 Hz, seemingly
t, 1H, glucose-H-4), 5.26–5.32 (total 2H, glucose-H-2, glucose-H-
3), 6.62 (s, 1H, H-4), 6.83 (d, J = 8.5 Hz, 2H, H-3ꢀ, H-5ꢀ), 6.86 (d,
J = 16.4 Hz, 1H, CH CH), 6.94, 6.96 (each s, total 2H, H-2, H-6),
7.01 (d, J = 16.4 Hz, 1H, CH CH), 7.36 (d, J = 8.5 Hz, 2H, H-2ꢀ, H-6ꢀ);
13C NMR: ␦ −4.29, 18.34, 20.70, 20.72, 20.78, 21.15, 21.24, 25.77,
62.17, 68.42, 71.18, 72.17, 72.78, 98.93, 109.17, 112.44, 114.35,
120.52, 125.30, 127.99, 130.05, 130.30, 140.35, 151.72, 156.02,
157.62, 169.30, 169.44, 169.52, 170.30, 170.69; the signals −4.29,
120.52 and 127.99 included two carbons. the signal 25.77 included
2.6.
4ꢀ,5-Diacetoxy-3-(tetra-O-acetyl-ˇ-d-glucopyranosyl)oxystilbene
(2d)
To a solution of piceid (2c, 2.00 g, 5.13 mmol) in pyridine (10 mL)
were added Ac2O (10 mL) and 4-N,N-dimethylaminopyridine
(62.5 mg, 0.51 mmol, 0.1 equiv.) under argon atmosphere. The mix-
ture was stirred for 2 h at room temperature, then the reaction was
quenched by adding ice. Firstly separated gummy residue gradu-
ally solidified by grinding well with water in a mortar. The resulting
precipitates were filtered and washed with water and then was
dried in vacuo to afford 2d (3.29 g, quant.) as white solid. 1H NMR: ␦
2.04, 2.06, 2.08 (each s, total 12H, sugar-OAc), 2.31 (s, 6H, 5-OAc, 4ꢀ-
OAc), 3.91 (ddd, J = 2.0, 5.4, 8.8 Hz, 1H, glucose-H-5), 4.18 (dd, J = 2.0,
12.2 Hz, 1H, glucose-H-6a), 4.28 (dd, J = 5.4, 12.2 Hz, 1H, glucose-
H-6b), 5.12 (d, J = 7.2 Hz, 1H, glucose-H-1), 5.16 (dd, J = 9.3, 9.3 Hz,
seemingly t, 1H, glucose-H-4), 5.26–5.33 (total 2H, glucose-H-2,
glucose-H-3), 6.65 (s, 1H, H-4), 6.95 (d, J = 16.4 Hz, 1H, CH CH), 6.97