704 D. Y. W. LEE ET AL.
˚
chromatography was run on a silica gel 60 A (Merck
230–400 mesh). The HPLC was performed with waters
HPLC system (Waters 1525 binary HPLC pump and
Waters dual absorbance detector) at room temperature
and the wavelength for UV detection was 254 nm. YMC
HPLC column (YMC-Pack, ODS-A, 150 ꢀ 4.6 mm ID, S-
5 mm, 12 nm) was used. 1H, 13C NMR spectra were
recorded on Varian NMR-300 spectrometers using the
hydrogenated residue of the deuterated solvents
(CDCl3, DMSO-d6) and TMS as internal standards.
Radioactivity was measured by Multi-purpose Scintil-
lation Counter (Beckman) and counter efficiency is
50% with negligible background radiation. The total
radioactivity (mCi) is calculated by the formula
(s, 9 H), 3.46 (d, J ¼ 9:0 Hz, 1 H), 3.56 (dd, J ¼ 8:4,
9.0 Hz, 1 H), 3.65 (dd, J ¼ 8:7, 9.3 Hz, 1 H), 3.76
(m, 3 H), 5.15 (d, J ¼ 9:0 Hz), 6.88 (d, J ¼ 7:5 Hz, 2 H),
6.94 (d, J ¼ 9:3 Hz, 1 H), 7.43 (d, J ¼ 7:8 Hz, 2 H),
7.94 (s, 1 H), 8.14 (d, J ¼ 9:3 Hz), 8.90 (s, 1 H).
13C NMR (75 MHz, CDCl3): d –0.74, ꢁ0.11, 0.24, 0.94,
1.30, 60.00, 70.04, 73.40, 76.13, 78.60, 80.62,
111.13, 116.82, 117.76, 120.06, 124.44, 124.98,
127.64, 130.15, 151.62, 155.22, 155.28, 161.57,
175.85.
Compound 2 was unstable on silica gel to give 7-
hydroxy-3-(4-hydroxy-phenyl)-8-(3,4,5-tris-trimethyl-
silanyloxy-6-trimethyl-silanyloxy-methyl-tetrahydro-
pyran-2-yl)-chromen-4-one (3)
(mCi ¼ DPM ꢀ dilution factor=2:22 ꢀ 109)
and
the
specific activity is calculated as ðmCi=mmolÞ ¼
mCi ꢀ molecular weight (mg/mmol)/sample weight (mg).
White solid. M.p.168–1728C (dec). 1H NMR (CDCl3):
d –0.37 (s, 9 H), 0.15 (s, 9 H), 0.21 (s, 9 H), 0.22
(s, 9 H), 3.49 (d, J ¼ 9:3 Hz, 1 H), 3.58 (dd, J ¼ 8:7,
8.4 Hz, 1 H), 3.67 (dd, J ¼ 8:7, 9.3 Hz, 1 H), 3.79
(m, 3 H), 5.17 (d, J ¼ 8:7 Hz), 6.83 (d, J ¼ 7:5 Hz, 2 H),
6.98 (d, J ¼ 9:0 Hz, 1 H), 7.34 (d, J ¼ 7:8 Hz, 2 H),
7.94 (s, 1 H), 8.17 (d, J ¼ 9:3 Hz), 8.95 (s, 1 H).
13C NMR (75 MHz, CDCl3): d –0.74, ꢁ0.11, 0.94,
1.29, 60.05, 70.06, 73.40, 76.12, 78.56, 80.62,
111.16, 115.76, 117.01, 117.58, 123.50, 124.74,
127.64, 130.31, 151.88, 155.37, 156.24, 161.74,
176.47.
Puerarin (1)
Compound 5 (13.3 mg, 0.021 mmol) was dissolved in
0.6 ml THF, then the solution of 1.2 mg (0.032 mmol)
NaBH4 in 1.5 ml ethanol was added dropwise at 08C.
Stirred at 08C for 10 min, then 0.2 ml 10% HCl was
added. Stirred at this temperature for 10 min and then
another 5 min at room temperature. The solvent was
evaporated under a slow stream of nitrogen. The
residue was purified by preparative TLC (CHCl3:MeOH
= 9:1) to give 6.6 mg of white solid product (m.p. 187–
1898C; 75% yield). 1H NMR (300 MHz, DMSO-d6): d
3.19 (3H, m), 3.66 (1 H, d, J ¼ 10:5 Hz), 3.97 (1 H, brs),
4.51 (1 H, brs), 4.78 (2 H, m), 4.96 (2 H, m), 6.75 (2 H, d,
J ¼ 8:4 Hz), 6.94 (1 H, d, J ¼ 9:00 Hz), 7.34 (2 H, d,
J ¼ 8:4 Hz), 7.89 (1 H, d, J ¼ 9:00 Hz), 8.30 (1 H, s),
9.50 (1 H, s). 13C NMR (75 MHz, DMSO-d6): d 61.50,
70.60, 70.83, 73.48, 78.82, 81.90, 112.71, 115.07,
116.94, 122.59, 123.16, 126.37, 130.13, 152.76,
156.50, 157.22, 161.15, 175.03.
7-Hydroxy-8-(6-hydroxymethyl-3,4,5-tris-trimethylsi-
lanyloxy-tetrahydro-pyran-2-yl)-3-(4-hydroxy-phe-
nyl)-chromen-4-one (4)
To a solution of compound 2 (7.51 g, 9.7 mmol) in
MeOH (200 ml) at 08C, K2CO3 (1.33 g, 9.6 mmol) was
added. After being stirred at 08C for 4 h, the MeOH
solution was quickly passed through a short silica-gel
column. Solvent was evaporated in a rotary evaporator.
The residue was further purified by flash column
chromatography (ethyl acetate:hexane = 3:7 until low
polarity by-products elute out, then the ratio is
changed to 4:6) to afford compound 4 (2.61 g, 43%) as
a white solid. M.p.175–1788C (dec). 1H NMR (CDCl3): d
–0.36(s, 9 H), 0.21(s, 9 H), 0.23(s, 9 H), 0.88 (t,
J ¼ 9 Hz, 1 H), 3.73 (m, 6 H), 5.20 (d, J ¼ 9:3 Hz), 6.87
(d, J ¼ 8:7 Hz, 2 H), 6.97 (d, J ¼ 9:0 Hz, 1 H), 7.40 (d,
J ¼ 8:7 Hz, 2 H), 7.95 (s, 1 H), 8.18 (d, J ¼ 8:7 Hz, 1 H),
8.80 (broad, 1 H). 13C NMR (75 MHz, CDCl3): d –0.11,
0.89, 1.25, 61.57, 71.07, 73.58, 76.47, 78.63, 81.45,
110.96, 115.75, 116.78, 117.61, 122.93, 124.77,
127.73, 130.24, 152.02, 155.34, 156.56, 161.26,
176.55.
7-Hydroxy-3-(4-trimethylsilanyloxy-phenyl)-8-
(3,4,5-tris-trimethylsilanyloxy-6-trimethylsilanyloxy-
methyl-tetrahydro-pyran-2-yl)-chromen-4-one (2)
Puerarin (0.26 g, 0.6 mmol) and N-trimethylsilylaceta-
mide (0.69 g, 5.2 mmol) were mixed and stirred at
1758C for 15 min. Most of the by-product acetamine
was pumped off. The residue was dissolved in ethyl
acetate (7 ml). Hexane (14 ml) was added and the
precipitate (acetamine) was filtrated off. Solvent was
evaporated and the residue was purified by flash
column chromatography (very short column, ethyl
acetate) to afford compound 2 (0.42 g, 86%) as a white
solid. M.p. 161–1638C (dec). 1H NMR (CDCl3): d –0.39
(s, 9 H), 0.13 (s, 9 H), 0.19 (s, 9 H), 0.20 (s, 9 H), 0.27
Copyright # 2007 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2007; 50: 702–705
DOI: 10.1002.jlcr