7
52
H.M.P. Poumale et al.
internal standard. Mass spectra were obtained with a Jeol JMS-700 instrument.
Column chromatography was conducted on silica gel 60 (Kanto Chemical Co., Inc.,
Japan), Sephadex LH-20 (Pharmacia, Sweden) and ODS (Fuji Silysia, Japan). TLC
analysis was carried out by using precoated silica gel plates (Merck), and the spots
were detected by spraying with H SO /10% vanillin and then heating. Flash
2
4
chromatography was carried out on silica gel (230–400 mesh). R values were
f
measured on PolyGram SIL G/UV254 (Macherey-Nagel & Co.).
3
.2. Extraction and isolation
The twigs of D. harmsiana were collected in July 2008 at Kribi, South Cameroon.
A specimen has been deposited in the National Herbarium, Yaounde, Cameroon
Ref. No. 21642/SRF).
Powdered twigs (500 g) of D. harmsiana were successively extracted with
´
(
n-hexane and ethyl acetate in soxhlet apparatus. The solvent was removed under
reduced pressure to yield 12 and 9 g extract of n-hexane (extract A) and ethyl acetate
(extract B), respectively.
Extract A (12 g) was chromatographed over silica gel using hexane–acetone of
increasing polarity, which yielded two main fractions. Fraction I (2.2 g) was purified
by a small column chromatography using Sephadex LH-20 and CHCl : MeOH
3
(
7 : 2) as eluent, followed by preparative TLC to afford lupeol (1a, 85.1 mg)
Poumale, 2007) and lup-20(29)-en-3-nonanoate (1b, 28.0 mg). Fraction II (0.9 g)
(
was purified by a silica gel column chromatography eluted with hexane:ethyl acetate
(4 : 1) to yield lup-20(29)-en-3-undecanoate (1c, 24.3 mg).
Extract B (9.0 g) was passed through a Sephadex LH-20 column and subjected to
silica gel column chromatography and preparative TLC to afford lupeol (1a, 14.9
mg) and lup-20(29)-en-3-hexadecanoate (1d, 31.0 mg) (Razdan et al., 1996).
3
.2.1. Lup-20(29)-en-3-nonanoate (1b)
ꢁ
ꢁ
White powder; m.p.: 78–79 C; R ¼ 0.53 (CHCl ); [ꢃ] þ 26.7 (c 0.1, CHCl ); IR
f
3
D
3
(film) ꢀ : 2947, 2873, 1724, 1641, 1451, 1375, 1247, 1183, 1040, 1020, 975, 888
max
ꢀ
1 1
cm ; H NMR (400 MHz, CDCl ): ꢁ 4.62 (brd, 1H, J ¼ 1.1 Hz, Ha-29), 4.46 (brd,
3
1
9
H, J ¼ 1.1 Hz, Hb-29), 4.41 (dd, 1H, J ¼ 10.6, 4.1 Hz, H-3), 2.28 (ddd, 1H, J ¼ 11.8,
.7, 4.7 Hz, H-19), 2.18 (t, 2H, J ¼ 6.7 Hz, H-2’), 1.63 (s, 3H, H-30), 1.11-1.90
(overlapping multiplets, 39H, H-1, H-2, H-5, H-6, H-7, H-9, H-11, H-12, H-13, H-
0
0
0
0
0
0
0
1
2
5, H-16, H-18, H-21, H-22, H-3 , H-4 , H-5 , H-6 ; H-7 , H-8 , H-9 ), 1.12 (s, 3H, H-
3), 0.90 (s, 3H, H-24), 0.80 (s, 3H, H-27), 0.78 (s, 3H, H-26), 0.77 (s, 3H, H-25), 0.69
1
s, 3H, H-28); C NMR (100 MHz, CDCl ): ꢁ 174.0 (s, C-1 ), 151.3 (s, C-20), 109.7
3
0
(
(
(
(
(
3
t, C-29), 80.9 (d, C-3), 55.7 (d, C-5), 50.6 (d, C-9), 48.6 (d, C-18), 48.3 (d, C-19), 43.3
s, C-17), 43.1 (s, C-14), 41.2 (s, C-8), 40.3 (t, C-22), 38.7 (t, C-1) 38.7 (d, C-13), 38.6
0
t, C-15), 38.1 (s, C-4), 37.4 (s, C-10), 35.9 (t, C-16), 35.2 (t, C-7), 34.5 (t, C-2 ), 32.2
0
0
0
0
0
0
t, C-21), 30.1-27.7 (overlapping triplets, C-3 , C-4 , C-5 , C-6 , C-7 , C-8 ), 25.5 (t, C-
1
C-28), 16.9 (q, C-25), 16.4 (q, C-26), 16.3 (q, C-27), 14.8 (q, C-24), 14.4 (q, C-9 ); EI-
2), 25.4 (t, C-2), 21.2 (t, C-11), 24.1 (q, C-23), 18.5 (t, C-6), 19.6 (q, C-30), 18.2 (q,
0
þ
MS (70 eV): m/z (%) 566 (M , 9), 538(16), 530 (8), 513 (12), 460 (6), 425 ([M ꢀ
þ
C H CO] ,18), 409 (90), 407 (9), 367 (8), 365 (10), 307 (42), 289 (34), 229 (12), 203
8 17