Ent-kaurene diterpenoids from Rabdosia yuennanensis

Phytochemistry, Vol. 30, No. 3, pp. 917-920, Printed in Great Britain.

1991 0

0031-9422/91 $3.00+0.00 1991 Pergamon Press plc

ENT-KAURENE DITERPENOIDS FROM RABDOSZA YUENNANENSZS Yu-QUN

CHEN,*

XIANG-LIN SHI and PEI-JU ZHENG~

Shanghai Institute of Organic Chemistry, Academia Sinica, 345 Lingling Lu, Shanghai 200032, China; tFudan University, 220 Handan Lu, Shanghai 200433, China (Received in revisedform 2 July 1990) Key Word Index-Rabdosia

yuennanensis; Labiatae; leaves; structure determination;

ent-kaurene diterpenoids.

Abstract-Three new diterpenoids rabyuennane A-C have been isolated from the leaves of Rabdosia yuennanensis and their structures established as ent-3/?,6a,7/?,15fl-tetraacetoxy-kaur-16-en-l l-one, ent-6a-hydroxy-3/?,7/_$15/?triacetoxy-kaur-16-en-11-one and ent-l/3,3/3,7/?,15/?-tetraacetoxy-kaur-16-en-lla-o1. The structure of rabyuennane A was confirmed by X-ray crystallographic analysis.

INTRODUCTION

Members of the genus Rabdosia (Labiatae) grow naturally in eastern Asia and various plants in Japan and China have been investigated chemically. Many diterpenoids of the ent-kaurane. B-seco-ent-kaurene and B/C seco-ent-kaurene types have been isolated and their biologically activities studied. We reported the structure elucidation of three new ent-kaurene diterpenoids, rabyuennane A-C from an ether extract of the leaves of Rabdosia yuennanensis (Hand-Ma@ Hara, collected in Dali, Yunnan provice, China. Three terpenoids were also isolated and shown to be identical with oleanolic, ursolic and crataegolic acids. RESULTSAND

DJSCU!SSION

The molecular formula of rabyuennane A (1) was determined as C,,H,,O, (m/z 518.2503) by HR mass spectroscopy. The major fragment ion peaks of the EI mass spectrum (m/z 458,398,338,278) resulting from [M -n-HOAc]+ (n= 1, 2, 3, 4) clearly indicated that 1 contained four acetoxy groups. This was consistentc with its NMR data (6 1.97,2.08,2.12,2.15 for acetyl methyls in ‘HNMR and 6169.0, 169.4, 170.1, 170.3 for ester carbonyls in 13CNMR). Because no hydroxy group was detected by IR, four 13C NMR doublet signals at 669.1, 75.6, 78.1, 79.2 revealed the carbons which had acetoxy groups were all methines. Due to the absence of the characteristic UV absorption at 230nm for an a#unsaturated exomethylene ketone, it was obvious that the exomethylene group [S 151.5 (s), 110.3 (t)] and the ketone (8208) were unconjugated. Thus, the nature of all nine oxygen atoms had now been elucidated. Considering the structure of many diterpenoids from the Rabdosia [ 11, 1 was presumed to be an ent-20-non-oxygenated kaurene diterpene ketone (three angular methyls 60.89, 1.04, 1.43 in ‘HNMR and three quatemary carbons 638.3, 43.4, 57.5 in 13C NMR) substituted with four acetoxy groups. Therefore, the signals of the six protons at low field in the

*Author to whom correspondence should be addressed.

‘HNMR should be attributed to two olefinic protons and the four protons attached to the carbon having an acetoxy group. The locations of the three acetoxy groups were examined by proton decoupling techniques. First, irradiation of the signal at 65.30 (lH, dd, J= 3.4,2.0 Hz) caused two signals at 64.93 (lH, d, J = 3.4 Hz) and 6 1.87 (lH, d, 2.0 Hz) to be decoupled. It revealed the environment of OAc rBB I I the partial structure -C--C-Cc--Cof II I I I AcO H the vicinal diacetoxy groups. According to the coupling constant (J = 2.0 Hz) of H-6 (6 5.30) and H-5 (6 1.87), H-6 must be j-oriented. Irradiation of the lowest field signal at 65.56 (lH, t, J=2 Hz) caused the signals at 65.12 and 4.98 (each lH, d, J = 2.0 Hz) to be decoupled, indicating that the partial structure AcO-CH-C=CH, was present in ring D. The remaining proton signal at 64.64 (lH, t, J = 1.7 Hz) was presumed to be located at C-3 according to the chemical shift and coupling constant. The chemical shift of C-5 (643.4 in “C NMR) showed a special upfield shift owing to the y-gauche effect [2] of the C-3 and C-7 disubstituents. Therefore, the two acetoxy groups at these two positions must be in the #I-orientation. For further proof of the structure of ring D [3], the following chemical transformations were performed. Mild hydrolysis [4] of 1 by K&O, in methanol at room temperature afforded a dihydroxy compound la as the sole product. Two signals appearing at 64.40 (t, J= 2.0 Hz, H-15) and 63.78 (d, J= 3.4 Hz, H-7) were observed in the ‘H NMR spectrum of la. It indicated that the two acetoxy groups at C-15 and C-7 were hydrolyzed simultaneously. Allylic oxidation [S] of la by freshly prepared MnO, in methylene chloride gave lb. The spectral data of lb [A.,., 230 nm, ~8700, in UV, 1730, 1635 cm-’ in IR and 66.16, 5.56 (each lH, s, H-17) in ‘HNMR] indicated the presence of a typical fivemembered ring ketone conjugated with an exomethylene group, a characteristic moiety in many Rabdane diterpenoids [S]. Catalytic hydrogenation [6] of la with Pd/C in

917

918

Y.-Q.

CHENet

al.

O& Aco& AcO#

AR’

=

R1

=

OAc

1

RI

R3

2 3

R’ =

H, Rz =

R’ =

AC,

R’

=

R3 = =

lb

AC

R3 =

OAc IC

AC H

AcoBc Ace@ OH

3

methanol yielded compound lc. A newly formed methyl group at C-17 with 61.08 (3H, d, 5=7 Hz) was thus assigned to the /?-orientation. The proton of C-15 at 63.13 (lH, d, J = 12.6 Hz) was shown to be a-oriented. Thus, the C-15 acetoxy group in 1 was proved to be in the pconfiguration. It was therefore concluded that among the four acetoxy groups in A, except the one at C-6, the other three were all p-oriented and in the axial conformation. The ketone was assigned to C-11 based on the appearance of the signal of the proton at C-9 (62.76, lH, s). Compound 1 is the first naturally occurring compound with a C-7 and C-15 diacetoxy, ent-20-non-oxygenatedkaurene structure. Finally, the structure of A was unambiguously confirmed by X-ray analysis. The single crystal was orthorhombic, space group P2 212, with a=10.649 (l), b = 10.661 (l), c=24.974 (1) !& D,= 1.215g cmm3 for 2=4 (M, 518.6), (MoK,)=0.84 cm-‘. A colourless crystal 0.2 x 0.2 x 0.1 mm was chosen for determination of the cell parameters and for data collection. All reflection intensities were measured with w/20 scans, graphite-monochromatized MoK radiation in the range 0” < 28 < 50”. The corrections of LP factor and absorption based on psi scans were applied. The primary structural model was carried out by direct method (MULTAN82) and the remainder of atoms were positioned from the difference Fourier method. The positional parameters and anisotropit thermal parameters of all nonhydrogen atoms were refined by full-matrix least squares to the final R =0.071, R,=0.074 for 1170 observed reflections* with I >3o (I). There are no peaks greater than 0.326 eAm3 on the final difference Fourier map. Full crystal data are deposited at the Fudan Crystallographic Data centre. A perspective view of the molecule is shown in Fig. 1. The absolute configuration of 1 was proved by Cc [7] (&,ax,305nm + 1.2, MeOH; c 0.05). The FT-IR of rabyuennane B (2) showed strong absorption bands for a hydroxy group, four carbonyl groups (1740.8, 1733.0, 1718.6, 1706.0) and an exomethylene group. The ‘HNMR of 2 was similar to 1, especially the chemical shift and coupling constants of

Inflexinol

Fig. 1. Molecular structure of rabyuennane A (1) as determined by X-ray diffraction.

five protons (H-3, H-5, H-15, Ha-17, Hb-17) of 2 at lower field. Only the signals of.H-6 and H-7 were shifted upfield by 1.16 and 0.15 ppm, respectively. The signal of the C- 19 methyl was shifted downfield by 0.21 ppm. This suggested that the difference between 2 and 1 is a hydroxy group instead of an acetoxy group at C-6. The molecular formula would be C,,H,,O, (M, 476). The EI mass spectrum showed significant fragment ion peaks at m/z 416, 356, 296 relating to [M-n-HOAc]+ (n= 1, 2, 3). This result also supported the above suggestion. The IR spectrum of rabyuennane C (3) showed hydroxy and carbonyl groups. The molecular formula CzsH,o09 was established by FAB mass spectrometry [m/z 521 [M+ l]‘] and 13CNMR spectroscopy. It exhibited one secondary hydroxyl group (669.8, d) and four acetates of secondary hydroxyl groups by means of ‘HNMR, 61.94, 2.07, 2.09, 2.16 (3H, s, 4-MeCO) and “C NMR, S 169.0, 169.1, 169.7, 170.7 (ester carbonyl) and 675.0, 78.7, 78.9, 79.9 (all d, HC-OAc). 13CNMR also showed the existence of an exomethylene group [S 105.7 (t), 151.1 (s)] and three angular methyl groups. The

920

Y.-Q. CHEN et al.

int.): 417 [Mfl -HOAc]+ (I), 399 [417-H,O]+ (l), 356 [417 -1-HOAc]+ (lo), 341 [356-Me]+ (35), 296 [356-HOAc]+ (6). Rabyuennane C (3). Prisms. IR: 3450 (OH), 1730, 1720, 1700 (C=O), 1690 (C=CH,). FABMS m/z (ret. int.): 613 [M + 92+ l] + (35), 521 [M+l]+ (loo), 461 [521-HOAc]+ (80), 401 [461 -HOAc]+ (50), 281 [341-HOAc]+ (lOO), 263 [281-HH,O]+ (40). 13CNMR 675.0 (C-l), 35.3 (C-2), 79.9 (C-3), 38.5 (C-4), 42.46 (C-5), 33.0 (C-6), 78.7 (C-7)*, 46.1 (C-8), 50.3 (C-9), 38.7 (C-10). 69.8 (C-11), 36.6 (C-12), 39.3 (C-13), 37.8 (C-14), 78.9 (C-15)*, 151.1 (C-16), 105.7 (C-17), 26.6 (C-18), 23.6 (C-19), 12.4 (C-20) 170.7, 169.7, 169.1, 169.0 (for CO,-) *The signal may be interchanged. For ‘H NMR data see Table 1. Acknowledgement-We

of HRMS and FABMS.

thank Mr Chen lie-Fei for measurement

REFERENCES

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