Guaiane dimers from Xylopia vielana

Phytochemistry 56 (2001) 335±340

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Guaiane dimers from Xylopia vielana Christine Kamperdick a, Nguyen Minh Phuong b, Tran Van Sung b, GuÈnter Adam a,* a Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle/Saale, Germany Institute of Chemistry, National Centre for Natural Science and Technology of Vietnam, Nghia Do, Tu Liem, Hanoi, Viet Nam

b

Received 27 March 2000; received in revised form 7 August 2000

Abstract From the leaves of Xylopia vielana (Annonaceae) the three dimeric guaianes vielanin A±C were isolated and structurally elucidated by mass and NMR spectroscopy as 1±3. The structure of 1 contains a bridged ring system formed probably via a Diels±Alder reaction of two di€erent guaiane monomers. Compounds 2 and 3 represent symmetric cyclobutanes formally generated from two equal guaiane moieties by [2+2] cycloaddition. # 2001 Elsevier Science Ltd. All rights reserved. Keywords: Xylopia vielana; Annonaceae; Vielanins A±C; Sesquiterpenoids; Guaiane dimers; Cyclobutanes

1. Introduction Xylopia vielana Pierre ex Fin. & Gagn. is a tree growing in Vietnam. In folk medicine the bark serves as an emmenagogue, whereas leaves are used for the treatment of rheumatism and pain. In South Vietnam, the plant is used against malaria (Do, 1991). The genus Xylopia is known to contain alkaloids of aporphine and tetrahydroberberine type and diterpenes (Hegnauer, 1964, 1989). In particular, some species contain dimeric guaianes (Martins et al., 1998) and dimeric diterpenes (Vilegas et al., 1991). X. vielana has not been phytochemically investigated until now. In continuation of our search for new biologically active compounds from Vietnamese medicinal plants (Porzel et al., 2000) we now report the isolation and structural elucidation of three guaiane dimers from the leaves of this plant. 2. Results and discussion The extract of the leaves (MeOH±H2O 95:5) of Xylopia vielana was partitioned between water and organic solvents of increasing polarity. Five compounds were isolated using normal phase column chromatography on silica gel and reversed phase chromatography. Besides the known ¯avonoids (ÿ)-epicatechin and * Corresponding author. Tel.: +49-345-5582-216; fax: +49-3455582-102. E-mail address: gadam@ipb- halle.de (G. Adam).

quercitrin, three new compounds belonging to the class of dimeric guaianes were obtained. The molecular mass of 506 for compound 1 was obtained from the negative ESIMS, which showed the [M±H]ÿ peak at m/z 505. The EIMS gave a very weak molecular ion (0.4%) at m/z 506.3064 leading to the molecular formula C32H42O5. The presence of an acetoxy group was revealed by loss of acetic acid producing the fragment at m/z 446 and by the corresponding signals in the NMR spectra (C 170.7 and 21.19, H 2.02). The remaining 30 carbon NMR signals belonged to two di€erent guaiane moieties. Other functional groups were three carbonyl groups ( 214.0, 206.9 and 202.9) and three fully substituted double bonds ( 168.6, 146.2, 140.8, 135.7, 132.5 and 129.4). The location of these functional groups within the two guaiane moieties and the assignment of the NMR signals were achieved by analysis of the CÿH long-range correlations from the GHMBC experiment. Some atoms like C-1, C-4, C-5, C-10 , C-40 , C-50 , H-3, H-6A, H-60 B, and H-150 showed surprisingly many correlation peaks. This was a ®rst hint to a bridged ring system and also explained the high chemical shifts of the quaternary carbons at  64.4 and 59.8. Careful analysis of all CÿH long-range correlations led to the depicted structure. Confusing for the structural elucidation was the presence of some 4JCH and 5JCH correlations through double bonds, which were of comparable intensitity with many 2JCH and 3 JCH correlations. This applied especially to the 4JCH correlations C-10 /H3-150 and C-80 /H3-120 and the 5JCH correlations C-50 /H3-120 , C-70 /H3-150 and C-100 /H3-150 .

0031-9422/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved. PII: S0031-9422(00)00344-7

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C. Kamperdick et al. / Phytochemistry 56 (2001) 335±340

The resulting structure was con®rmed by the MS fragmentation, which was mainly determined by the Retro±Diels±Alder reaction giving monomer 1a at m/z 230 (19%) and monomer 1b at m/z 276 (6%). The base peak at m/z 234 resulted from loss of ketene from 1b, which is typical for unsaturated acetoxy compounds. According to the structure of 1, the dimer did not loose ketene but acetic acid, which is characteristic for saturated acetoxy compounds, thus proving the formation of a double bond by RDA. Important information about the con®guration was deduced from the NOESY experiment: the con®guration of the central bicyclo[2.2.1]heptane moiety with the endo-anellation of the ®ve-membered ring at C-10 and C-20 was revealed by a signi®cant NOE e€ect between the methyl protons H3-15 ( 1.60) and H-60 B ( 3.65), which was not possible for the exo-isomer. The NOE e€ects of H-2 ( 4.87) on the methyl groups H3-14 (

1.08) and H3-140 ( 0.90) and on H-20 ( 2.48) indicated the con®guration of the acetoxy group at C-2 as syn with respect to the
4 double bond and suggested that both methyl groups C-14 and C-140 were cis to the bridge of the norbornene moiety. The NOE e€ect between H3-120 ( 1.97) and H-60 B ( 3.65) assigned C-120 to be the methyl group in trans-position to the carbonyl group C-80 . In both seven-membered rings NOE e€ects across the ring were observed. They were only possible if both involved protons were in axial position and at the same side of the ring. Thus, the seven-membered ring on the left gave rise to NOE e€ects between H-60 A ( 2.93) and H-90 B ( 3.23). In the right-hand seven-membered ring the NOE interactions between H-7 ( 2.41) and H10 ( 2.88) showed that both were in axial position and cis so that the substituents at C-7 (isopropyl group) and at C-10 (methyl group) had also to be cis resulting in the depicted relative con®guration (Table 1). The new compound 1 was named vielanin A. Its biosynthetic precursors were supposed to be synthons of the instable guaiane-type monomers 1a and 1b, occurring also in the mass spectrum of 1. The HRMS of vielanin B (2) ([M]+ m/z 460.2596) led to the molecular formula C30H36O4. The 13C NMR showed only 15 carbon signals. Thus, 2 consisted of two equivalent C15H18O2 moieties, which were responsible for the base peak in the EIMS at m/z 230. The 1H NMR spectrum showed in addition to four methyl groups (3 singlets at  2.21, 2.04, 1.88 and one doublet at  1.08) only six other signals, which belonged to aliphatic protons. The carbon shifts revealed the presence of two carbonyl groups ( 203.7 and 205.1) and two double bonds ( 131.6, 143.8, 146.9 and 171.3). The high

C. Kamperdick et al. / Phytochemistry 56 (2001) 335±340

337

Table 1 NMR data (500/125 MHz) of vielanin A (1) in CDCl3 c

H

CÿH long-range correlations H-3 (m)b, H-6B (m), H-9A (w), H-9B (m), H-14 (s), H-15 (vw), H-20 (s) H-3 (m) H-15 (s), H-150 (w)

1

64.4

±

2 3

86.9 56.3

4.87 d (1.5) 3.10 s

4

135.7

±

5

132.5

±

6

25.7

7 8

57.0 214.0

9

48.2

10 11 12 13 14 15 10

28.5 27.8 21.13 20.1 17.1 14.3 59.8

A: 1.85 ddd (15.3/4.9/1.8) B: 2.33 dd (15.3/7.0) 2.41 m ± A: 2.16 dd (13.4/2.1) B: 2.71 dd (13.4/12.5) 2.88 dqd (12.2/6.6/2.5) 1.98 m 0.82 d (6.7) 0.94 d (6.7) 1.08 d (7.0) 1.60 s ±

20 30 40 50

51.8 206.9 140.8 168.6

60

30.7

70

129.4

A: 2.93 br d (14.0) B: 3.65 d (14.0) ±

80

202.9

±

90

47.3

100

34.6

110 120 130 140 150 CO-CH3 CO-CH3 a b

146.2 23.6 23.7 16.2 8.6 170.7 21.19

2.48 s ± ± ±

A: 2.57 dd (15.3/7.3) B: 3.23 dd (15.3/1.5) 2.46 m ± 1.97 s 2.05 s 0.90 d (7.3) 1.64 s ± 2.02 s

H-2 (s), H-3 (vw), H-6A (vw), H-6B (w), H-15 (s) H-2 (s), H-3 (s), H-6B (m), H-7 (m), H-15 (s), H-20 (s) H-7 (w) H-6B (m), H-12 (s), H-13 (s) H-6A (vw), H-6B (m), H-7 (w), H-9A (w), H-9B (m), H-14 (w) H-14 (s) H-9A (m), H-14 (s), H-20 (s) H-6B (w), H-12 (s), H-13 (s) H-13 (s) H-12 (s) H-9A (vw) H-3 (w) H-20 (w), H-60 B (s), H-90 A (m), H-90 B (w), H-140 (s), H-150 (m) H-3 (s), H-15 (m) H-20 (s), H-150 (s) H-20 (w), H-60 A (w), H-60 B (s), H-150 (s) H-20 (s), H-60 A (w), H-60 B (s), H-100 (w), H-120 (w), H-130 (w), H-150 (s) ± H-60 A (w), H-60 B (s), H-90 A (w), H-120 (s), H-130 (s), H-150 (m) H-60 B (m), H-90 A (m), H-90 B (w), H-100 (w), H-120 (m), H-130 (w) H-100 (w), H-140 (s) H-3 (w), H-20 (s), H-90 A (m), H-90 B (w), H-140 (s), H-150 (w) H-60 A (w), H-60 B (s), H-120 (s), H-130 (s) H-130 (s) H-120 (s) H-90 B (w), H-100 (w) ± H-2 (m), CO-CH3 (s) ±

NOE e€ectsa

H-3, H-14, H-20 , H-140 H-2, H-15, H-20 , H-60 A, H-90 B, CO-CH3 (vw), H-140 (w)

H-7, H-13 (w) H-13, H-15 H-6A, H-10, H-12, H-13 H-10, H-14 H-11 (w), H-12 (w), H-14 (w) H-7, H-9A, H-14, H-20 H-9B, H-12, H-13 H-7 (w), H-9B (w), H-11, H-13 H-6A (w), H-6B, H-7, H-11, H-12, H-15 H-2, H-9A, H-9B (w), H-10, H-20 H-3, H-6B, H-13, H-60 A, H-60 B (w) H-2, H-3, H-10, H-14, H-140

H-3, H-15, H-90 B H-15 (w), H-120 , H-150

H-100 , H-140 (w) H-3, H-100 (w), H-60 A H-90 A, H-90 B, H-140 H-60 B, H-130 , H-150 H-120 , H-140 (w) H-2, H-20 , H-90 A, H-130 (w) H-60 B, H-120 H-3 (vw)

Obtained from the NOESY experiment. s=strong, m=medium, w=weak, v=very.

chemical shift of the ole®nic carbon at  171.3 was explained by conjugation to a carbonyl group and ring tension. Analysis of the CH long-range correlations from the GHMBC (Table 2) resulted in a guaiane moiety with two carbonyl groups in positions 2 and 8 and still free valencies at positions 3 and 4. Connecting the free valencies resulted in a dimer with a central cyclobutane ring. This was supported by the fact that C-3 ( 47.0) showed a 1JCÿH and simultaneously a long-range CÿH correlation (2JCH or 3JCH) to the protons at  2.21

(H-3/H-30 ). Because of the equivalence of each two cyclobutane carbons, it was not possible to distinguish between a 3-30 /4-40 or a 3-40 /4-30 connection on the basis of the CH long-range correlations. In the case of a 3-30 / 4-40 connection between the two guaiane moieties there would be a 3JHH coupling between H-3 and H-30 with an expected larger value, whereas in the other case (3-40 / 4-30 connection) a small 4JHH coupling would be observed. The coupling constant between H-3 and H-30 could not be observed directly because of their equiva-

338

C. Kamperdick et al. / Phytochemistry 56 (2001) 335±340

Table 2 NMR data (500/125 MHz) of vielanin B (2) in CDCl3 NOE e€ectsa

c

H

CH long-range correlations

1/10

146.9

±

2/20 3/30 4/40 5/50

205.1 47.0c 54.5 171.3

± 2.21 s ± ±

6/60

29.2

H-3/30 (w)b, H-6/60 A, H-6/60 B, H-9/90 A, H-9/90 B, H-14 H-3/30 H-3/30 , H-15/150 H-3/30 , H-6/60 A (w), H-6/60 B, H-15 H-3/30 , H-6/60 A, H-6/60 B, H-9/90 A (w), H-9/90 B (w), H-12/120 , H-13/130 , H-15/150 H-13/130 (w)

7/70 8/80

131.6 203.7

9/90

46.9c 0

10/10 11/110 12/120 13/130 14/140 15/150 a b c d

27.7 143.8 22.5 23.3 19.0 19.8

A: 3.27 d (17.3) B: 3.54 d (17.3) ± ± A: 2.68 dd (15.3/7.3) B: 3.02d m 3.01d m ± 1.88 s 2.04 s 1.08 d (7.0) 1.25 s

H-6/60 A (w), H-6/60 B, H-9/90 A, H-12/120 , H-13/130 H-6/60 A (w), H-6/60 B, H-9/90 A, H-9/90 B, H-12/120 , H-13/130 , H-14/140 H-10/100 (w), H-14/140

H-14/140 (w), H-15/150

H-6/60 B, H-9/90 B/H-10/100 , H-15/150 H-6/60 A, H-9/90 A (w), H-12/120 , H15/150

H-6/60 B(w), H-9/90 B, H-14/140 H-6/60 A, H-9/90 A, H-14/140 H-6/60 A, H-9/90 A, H-14/140

d

0A

0B

0

H-9/9 , H-9/9 , H-14/14 H-6/60 A (w), H-6/60 B, H-12/120 , H-13/130 H-13/130 H-12/120 H-9/90 A, H-9/90 B/10/100 H-3/30

d

H-6/60 A (w), H-6/60 B, H-13/130 , H-15/150 H-12/120 , H-14/140 H-9/90 A, H-9/90 B/H-10/100 , H-13/130 H-3/30 , H-6/60 A, H-6/60 B, H-9/90 10/100 , H-12/120

Obtained from the NOE di€erence spectra. w=weak. Assignment of 46.9 and 47.0 from APT spectrum, GHMBC correlations not resolved. Signals of H-9/90 B (3.02) and H-10/100 (3.01) overlapped.

lence. Thus, the coupling was deduced by analysis of the 13 C satellites (GuÈnther, 1992). Those were obtained from the one-dimensional GHSQC experiment showing exclusively the 13C satellites. The protons H-3/H-30 appeared as doublet of doublets with 1JCÿH of 146.9 Hz and 3JHH or 4JHH of 3.3 Hz. A 4JHH coupling for substituted cyclobutanes was expected to range from 0 to 2 Hz and thus was excluded. The 3JHH couplings of cyclobutanes range from 2 to 10 Hz and are normally unreliable for the determination of the relative con®guration, because the energy di€erence between the two puckered conformers is rather low, so that there exists an equilibrium between the conformers with an averaged HH coupling constant (Gaudemer, 1977). In the case of 2 however, the two anellated ®ve-membered rings caused a nearly planar rigid system. A cisoid-anellation of the two ®ve-membered rings with cis-con®guration of H-3 and H-30 and a dihedral angle of 0±15 would show a HH coupling of about 7 Hz, whereas the measured value of 3.3 Hz supported very well the transoid-anellation with trans-con®guration of H-3 and H-30 and a dihedral angle of ca 120 . Very interesting was the observation that in the GHMBC spectrum the correlation peaks between C-4/40 ( 54.5) and H3-15/150 ( 1.25) had more than the double intensity than all other correlation peaks originating from methyl groups. This was explained by the fact, that C-4 had one 2JCH and one 3JCH correlation to H3-

15 and H3-150 resulting in a strongly increased intensity of the correlation peak. This additionally supported the structure of 2, whereas an isomeric compound with 3-40 / 4-30 connection should have shown a double intensity of the correlation peak between C-3 and H3-15/150 . Analysis of the NOE di€erence spectra (Table 2) gave some further information: the di€erentiation between the two methyl groups at C-11/110 was deduced from the NOE e€ect between the signal at  1.88 (H3-12/120 ) and H-6/60 A/B ( 3.54 and 3.27). The NOE e€ects across the seven-membered ring between H-6/60 B and H-9/90 A showed that these protons were on the same side of the ring. Because of the conformational ¯exibility of the sevenmembered ring, the con®guration of the methyl group at C-10/100 could not be determined by NOE e€ects. Vielanin B (2) can be regarded as [2+2] cycloaddition product of two molecules of the instable guaiane monomer 2a which also appears as fragment in the mass spectrum of 2. Like vielanin B (2), also vielanin C (3) was a symmetric guaiane dimer. The positive ESIMS spectrum showed the [M+H]+ peak at m/z 525. The corresponding molecular ion with the mass 524 did not appear in the EIMS. The peak with the highest mass was that of the monomeric fragment at m/z 262, which easily lost water. The high resolution of this fragment at m/z 244.1104 (C15H16O3) led to the molecular formular C30H36O8 for the dimer containing 13 double bond equivalents.

C. Kamperdick et al. / Phytochemistry 56 (2001) 335±340

339

Table 3 NMR data of vielanin C (3) in CD3OD (500/125 MHz)

1/10 2/20 3/30 4/40 5/50 6/60 7/70 8/80 9/90 10/100 11/110 12/120 13/130 14/140 15/150 a b

c

H (500 MHz)

CH long-range correlations

147.8 208.8 48.8 54.5 163.9 114.5 166.1 104.2 41.5

± ± 2.10 s ± ± 6.00 s ± ± A: 1.82 dd (13.7 / 13.1) B: 2.21 dd (13.7 / 7.0) 3.00 m ± 1.49 s 1.56 s 1.23 d (6.7) 1.18 s

H-6/60 , H-9/90 B, H-10/100 , H-14/140 H-3/30 H-30 /3, H-15/150 H-3/30 , H-6/60 , H-15/150 H-3/30 , H-10/100 , H-15/150 ± H-6/60 , H-9/90 A (w)b, H-9/90 B, H-12/120 , H13/130 H-6/60 , H-9/90 A, H-9/90 B, H-14/140 (w) H-10/100 , H-14/140

29.6 86.6 27.5 23.5 19.2 20.4

H-9/90 A, H-9/90 B, H-14/140 H-6/60 , H-12/120 , H-13/130 H-13/130 H-12/120 H-9/90 A, H-10/100 H-3/30

NOE e€ectsa

H-15/150 , H-12/120 , H-13/130 H-6/60 , H-12/120 (w), H-14/140 H-10/100 H-9/90 B, H-14/140 H-6/60 , H-9/90 A (w), H-13/130 H-6/60 H-9/90 A, H-9/90 B, H-10/100 H-3/30 , H-6/60

Obtained from the NOE di€erence spectra. w=weak.

The 13C spectrum (Table 3) exhibited 15 signals, among them one carbonyl group ( 208.8) and two double bonds ( 166.1, 163.9, 147.8 and 114.5) for each monomeric moiety, thus leaving seven double bond equivalents for seven rings. The signals of the two guaiane moieties were assigned by analysis of the CÿH long-range correlations from the GHMBC spectrum (Table 3), locating the carbonyl groups at C-2/20 and the double bonds at positions 1(5)/10 (50 ) and 6/60 . The CÿH correlations of C-3 with H-3 (1JCH) and H-30 (2JCH or 3 JCH), the highly increased correlation peak between C4 ( 54.5) and H3-15/150 ( 1.18) and the similarity of the chemical shifts of the cyclobutane atoms compared with vielanin B (2) proposed an analogous structure and relative con®guration of the cyclobutane with vielanin C (3) containing two additional rings. Because of their chemical shift of  86.2 C-11/110 thus had to bear an oxygen atom. The shift of C-8 ( 104.2) was that of an acetalic or ketalic group. Connecting C-8/80 with C-11/ 110 via one oxygen atom accounted for the observed chemical shift and produced the two additional oxetane rings. Such an oxetane ring is known as structural element of a guaiane dimer from Xylopia aromatica (Martins et al., 1998). Comparing the so deduced structure with the molecular formula from the HRMS resulted in a discrepancy of two oxygen atoms. The chemical shifts of all atoms allowed no insertion of an additional oxygen into the molecule exept the formation of a peroxide resulting in the depicted structure of vielanin C (3). The peroxide group could account for the antimalarial activity of this plant as terpene peroxides like artemisinin are known to possess this activity (Thebtaranonth et al., 1995). H-9/90 A appeared as dd with a geminal coupling of 13.7 Hz with H-9/90 B and a vicinal coupling constant of

13.1 Hz with H-10/100 , thus showing the trans-diaxial conformation of H-9/90 A and H-10/100 . Thus, the methyl group at C-10/100 was equatorial. As in vielanin B (2), the relative con®guration could not be determined by the observed NOE e€ects (Table 3). 3. Experimental 3.1. General experimental procedures EIMS: AMD 402, 70 eV. ESIMS: Finnigan TSQ 700. NMR: Varian Gemini 300, Unity 500. CC: siliga gel 60, 40±63 mm (Merck), Lichroprep RP-18, 25±40 mm (Merck). Prep. TLC: precoated plates, silica gel 60, F254, thickness 0.5 mm (Merck). 3.2. Plant material Leaves of X. vielana (Lour.) Tan. Pierre ex Fin. & Gagn. were collected in August 1997 in Vinh phuc, North Vietnam, and identi®ed by Ngo Van Trai, Institute of Materia Medica, Hanoi. A voucher specimen is deposited at the Institute of Ecology, National Centre for Natural Science and Technology, Hanoi, Vietnam. 3.3. Extraction and isolation The plant material was dried (1.54 kg), ground and extracted 4 with MeOH±H2O (95:5). The organic solvent was evaporated under red. pres. and the aq. residue extracted with n-hexane, EtOAc and n-BuOH, successively (each 3) giving 27.4 g n-hexane extract, 16.9 g EtOAc extract and 72.9 g n-BuOH extract. The EtOAc extract was separated on silica gel (238 g, 63±200 mm)

340

C. Kamperdick et al. / Phytochemistry 56 (2001) 335±340

with solvents of increasing polarity: CHCl3, followed by 10%, 20% and 30% MeOH in CHCl3 and ®nally CHCl3±MeOH±H2O (60:35:3). 16 Frs were collected. Fr. 11 (1.2 g, eluted with CHCl3±MeOH 7:3) yielded after fractionation on silica gel with CHCl3±MeOH (85:15) 45 mg (ÿ)-epicatechin. Fr. 6 (1.7 g, eluted with CHCl3±MeOH 8:2) was separated on siliga gel (160 g, 63±200 mm) with n-hexane±acetone (8:2) giving 149 fractions. Frs 24±27 (139 mg) were further puri®ed on silica gel using n-hexane±acetone (8:2) followed by crystallization to give 20 mg pure vielanin A (1) and 33 mg mother liquid. Frs 87±101 (62 mg) were chromatographed on silica gel with n-hexane±acetone (7:3) to yield 3 mg of vielanin B (2). Vielanin C (3, 8 mg) was obtained after reversed-phase chromatography of Frs 142±149 (24 mg) on RP-18 with MeOH±H20 (6.4). Frs 13±15 of the ®rst column (2.48 g) yielded after chromatography on silica gel and on RP-18 38 mg of quercitrin. 3.3.1. Vielanin A (1)  Crystals from MeOH, m.p. 190±191 C. []27 D ÿ8 EtOH (CHCl3, c 0.5). UV lmax nm (log ): 261 (4.44). IR 3 (cmÿ1): 2964, 2937, 2875, 1741, 1724, 1662, 1635, CHCl max 1597, 1458, 1440, 1381, 1369, 1333, 1243, 1184, 1073, 1062, 1033, 1007, 991. EIMS m/z (rel. int.): 506.3064 [M]+, (0.4), (C32H42O5 requires 506.3032), 446.2825 [M±CH3COOH]+ (11), (C30H38O3 requires 446.2821), 276.1724 [monomer 1b]+ (6) (C17H24O3 requires 276.1725), 234.1618, [monomer 1b Ð C2H2O]+ (100), (C15H22O2 requires 234.1620), 230.1322 [monomer 1a]+ (19), (C15H18O2 requires 230.1307) 216.1492 [monomer 1b Ð CH3COOH]+ (81), (C15H20O requires 216.1514), 174.1412 (15), (C13H18 requires 174.1409), 135.0806 (22), (C9H11O requires 135.0810), 108. 0531 (12) (C7H8O requires 108.0575). 3.3.2. Vielanin B (2) m.p. 210±212 C (EtOAc). EIMS m/z (rel. int.): 460.2596 [M]+ (47) (C30H36O4 requires 460.2614), 244 (25), 231 [Monomer+H]+ (62), 230.1352 [monomer]+ (100), (C15H18O2 requires 230.1307), 229 (54), 215 [monomer Ð CH3]+ (30), 204 [monomer Ð CH3 ÿOH]+ (25), 187.1123 [monomer Ð CH3ÿCO]+ (33), (C13H15O requires 187.1123), 173 (17), 159 (20), 145 (13), 105 (13), 91 (14), 69 (17), 55 (13). 3.3.3. Vielanin C (3)  m.p. 108±110 C (acetone). []26 D ÿ120 (CHCl3, c 0.5). EtOH UV lmax nm (log ): 292 (4.22), 204 (4.19). EIMS m/z (rel. int.): 262 (8) [monomer]+, 244.1104 (59) [monomer Ð H2O]+ (C14H13O3 requires 244.1099), 229.0859 (100) [monomer Ð H2OÿCH3]+ (C14H13O3

requires 229.0865), 216 (20) [244 Ð CO]+, 204 (4) [monomer (CH3)2CO]+, 201 (19) [229 Ð CO]+, 189 (29) [204 Ð CH3]+, 161 (23), 159 (24), 148 (19), 145 (22), 115 (24), 91 (31), 59 (41), 58 (52). Positive ESIMS m/z (rel. int.): 547 (44) [M+Na]+, 525 (100) [M+H]+. 3.3.4. (ÿ)-Epicatechin  m.p. 233±235 C. []27 D ÿ47 (MeOH, c 1.0). Negative ESIMS m/z (rel. int.): 298.0 (100) [MÿH]ÿ. The compound was identi®ed by comparison of its 13C NMR data with reference values (Agrawal, 1989). 3.3.5. Quercitrin  Amorphous solid, []30 D ÿ110 (MeOH, c 1.0). Negative ESI MS m/z (rel. int.): 447.2 (91) [MÿH]ÿ, 119.1 (100). The compound was identi®ed by comparison of its 13C NMR data with reference values (Agrawal, 1989). Acknowledgements We thank the VW-Stiftung, Hannover, and the Bundesministerium fuÈr Bildung, Wirtschaft, Forschung und Technologie, Bonn, for ®nancial support. References Agrawal, P.K., 1989. Studies in Organic Chemistry 39: Carbon-13 NMR of Flavonoids. Elsevier, Amsterdam, p. 336, 446 Do, T.L., 1991. Nhung cay thuoc va vi thuoc Viet Nam (Glossary of Vietnamese Medicinal Plants). Nha xuat ban Khoa hoc va Ky thuat (Science and Technics Publication), Hanoi, Vietnam, p. 583 Gaudemer, A., 1977. Determination of con®gurations by spectrometric methods. In: Kagan, H.B. (Ed.), Stereochemistry: Fundamentals and Methods. Vol. 1. Thieme, Stuttgart, pp. 83±89. GuÈnther, H., 1992. NMR-Spektroskopie: Grundlagen, Konzepte und Anwendungen der Protonen- und Kohlensto€-13-KernresonanzSpektroskopie in der Chemie. Thieme, Stuttgart, pp. 200±204. Hegnauer, R., 1964. Chemotaxonomie der P¯anzen III. BirkhaÈuser, Basel, p. 117. Hegnauer, R., 1989. Chemotaxonomie der P¯anzen VIII. BirkhaÈuser, Basel, p. 44. Martins, D., Osshiro, E., Roque, N.F., Marks, V., Gottlieb, H.E., 1998. A sesquiterpene dimer from Xylopia aromatica. Phytochemistry 48, 677±680. Porzel, A., Lien, T.P., Schmidt, J., Drosihn, S., Wagner, C., Merzweiler, K., Sung, T.V., Adam, G., 2000. Fissistigmatins A±D: novel type natural products with ¯avonoid-sesquiterpene hybrid structure from Fissistigma bracteolatum. Tetrahedron 56, 865±872. Thebtaranonth, C., Thebtaranonth, Y., Wanauppathamkul, S., Yuthavong, Y., 1995. Antimalarial sesquiterpenes from tubers of Cyperus rotundus: structure of 10,12-Peroxycalamenene, a sesquiterpene endoperoxide. Phytochemistry 40, 125±128. Vilegas, W., D'Arc Felicio, J., Roque, N.F., Gottlieb, H.E., 1991. Diterpenic adducts from Xylopia species. Phytochemistry 30, 1869± 1872.