Immunosuppressive terpenoids from Tripterygium wilfordii

Qian Shen a, ZhiYao b, Yoshihisa Takaishi c,Yan Wen Zhang a,b,c, Hong Quan Duan a,*

a School of Pharmaceutical Sciences, Basic Medical Research Center of Tianjin,

Tianjin Medical University, Tianjin 300070, Chinab Department of Immunology, Tianjin Medical University, Tianjin 300070, China

c Faculty of Pharmaceutical Sciences, University of Tokushima, Tokushima 770-8505, Japan

Received 18 October 2007

Abstract

Two new terpenes, triptobenzene P (1) and wilforone (2) were isolated from Tripterygium wilfordii, as well as 10 known

terpenes. Their structures were elucidated by spectroscopic methods. Compounds 2–4, 8, 10, and 11 showed significant

immunosuppressive activities.

# 2008 Hong Quan Duan. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

Keywords: Tripterygium wilfordii; Diterpene; Triterpene; Immunosuppressive activity

Tripterygium wilfordii Hook f. has been used as traditional Chinese medicine for treatment of cancer and as an

insecticide from ancient times [1,2]. In recent years, this plant has been used to treat rheumatoid arthritis, other

inflammatory and autoimmune diseases, skin disorders, and in male-fertility control in clinics [3–5]. In our previous

study, we have reported five immunosuppressive sesquiterpene alkaloids from T. wilfordii [6]. In this paper, two new

compounds, named triptobenzene P (1, 5.2 mg) and wilforone (2, 7.0 mg), and 10 known terpenes were isolated from

the ethyl-acetate extract of T. wilfordii. Their structures were elucidated by spectroscopic methods. Compounds 2–4, 8,

10, and 11 showed significant immunosuppressive activities.

Triptobenzene P (1), amorphous yellow powder, ½a�D25 þ 130:8 (c 5.2, CHCl3). Its HR ESI-MS showed [M]+ at m/z

315.2338 (calcd. 315.2324), corresponding to the molecular formula C21H30O2. Its IR spectrum [7] showed the

hydroxyl band at 3442 cm�1. The 1H NMR spectrum of 1 showed an isopropyl group [dH 1.21, 1.24 (d, each 3H,

J = 6.9 Hz), 3.30 (sept, 1H, J = 6.9 Hz)], one methyl [dH 0.98 (s, 3H)], one methoxyl [dH 3.73 (s, 3H)], an oxygenated

methylene [dH 3.96 (dd, 1H, J = 10.7, 6.6 Hz), 3.69 (dd, 1H, J = 10.7, 5.7 Hz)], two terminal double bond protons [dH

4.85,4.76 (brs, each 1H)], and two aromatic protons [dH 7.08 (s, 2H)]. The 13C NMR spectral data (Table 1) showed 21

carbon signals, including a benzene ring, a terminal double bond (dC 104.4, 150.9), three methyls, an oxygenated

methylene, and a methoxy group. From the above information, compound 1 was assumed to be an abietane-type of

diterpene, which has reported from the same plant. The 13C NMR spectral data of 1 was similar to those of triptohairic

acid (6) [8], except for the A ring. In the HMBC spectrum of 1, the proton signals of oxygenated methylene at dH 3.96

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Chinese Chemical Letters 19 (2008) 453–456

* Corresponding author.

E-mail address: duanhq@tijmu.edu.cn (H.Q. Duan).

1001-8417/$ – see front matter # 2008 Hong Quan Duan. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

doi:10.1016/j.cclet.2008.01.031

(H-18) correlated with the signals at dC 27.3 (C-2), 46.1 (C-3), and 150.9 (C-4), the signals of the terminal double bond

at dH 4.85 and 4.76 (H-19) with the signals at dC 46.1 (C-3), 48.5 (C-5), 150.9 (C-4). Thus, the partial structure of A—

ring was supposed as shown in Fig. 2. In addition, the signal at dH 3.96 (H-18) showed NOE correlation with the signal

at dH 4.85 (H-5). Thus, the hydroxyl methylene was assigned at 3a-position. Therefore, the structure of 1 was assigned

as shown (Fig. 1).

Wilforone (2), amorphous maize powder, ½a�D25 þ 23:3 (c 7.0, CHCl3). Its HR ESI-MS showed [M+Na]+ at m/z

463.3525 (calcd. 463.3552), corresponding to the molecular formula C30H48O2. Its IR spectrum [9] showed the

Q. Shen et al. / Chinese Chemical Letters 19 (2008) 453–456454

Table 113C NMR chemical shifts for compound 1 and 2

c 1 2

1 38.2 42.1

2 27.3 34.2

3 46.1 218.8

4 150.9 47.5

5 48.5 54.7

6 21.1 19.7

7 24.4 35.3

8 128.5 41.8

9 146.0 55.1

10 39.9 38.1

11 121.5 70.3

12 154.9 37.4

13 138.3 132.2

14 123.8 44.3

15 26.1 26.4

16 23.9 39.2

17 23.9 34.6

18 64.7 134.9

19 104.4 38.7

20 22.8 33.4

21 60.5 34.7

22 36.5

23 27.7

24 20.7

25 17.0

26 18.8

27 20.9

28 23.7

29 32.3

30 24.2

Fig. 1. The structures of compounds 1 and 2.

hydroxyl and ketone band at 3523, 1699 cm�1. The 1H NMR spectrum of 2 revealed eight methyls [dH 0.71, 0.88, 0.95,

1.00, 1.06, 1.08, 1.11, 1.19 (each 3H, s)], a oxygenated methine [dH 3.82 (dt, 1H, J = 10.9, 5.6 Hz)], and other five

proton signals [dH 1.92 (m, 1H), 2.27 (dd, 1H, J = 14.0, 2.1 Hz), 1.65 (m, 1H), 2.91 (dd, 1H, J = 4.1, 5.4 Hz)]. The 13C

NMR spectral data (Table 1) showed 30 carbons, including a double bond (dC 132.2, 134.9), an oxygenated methylene

(dC 70.3), a ketone (dC 218.8), and eight methyl carbon signals. The 13C NMR spectral data of 2 was similar to those of

hypodiol [10], except for the ketone in 2. By analysis of the HMBC spectral data (Fig. 2), the hydroxyl group was

located at C-11, the double bond was assigned at C-13, -18, and the ketone group was located at C-3. Further more, the

NOE correlation between H-11 (dH 3.82) with H3-25 and 26 (dH 1.08 and 0.88) indicated the hydroxyl group had a-

orention. Thus, the structure of 2 was determined as shown (Fig. 1).

The structures of the known compounds were identified by their spectral data in comparison with literature values

as follow: triptobenzene B (3) [11], wilforol E (4) [12], maytenoic acid (5) [13], triptohairic acid (6) [8], triptobenzene

H (7) [13], 3-oxo-olean-12-en-29-oic acid (8) [14], olean-12-ene-3b,29-diol (9) [15], 3-epikatonic acid (10) [16],

3b,29-dihydroxy-D:B-friedoolean-5-en (11) [17], wilforol D (12) [17].

In a screen for immunosuppressive agents from the extract of T. wilfordii, we examined the inhibitory effect of the

isolated compounds on lymphocyte transformation (Table 2). Compounds 2–4, 8, 10, and 11 revealed a significant

distinction compared to the concanavalin (ConA) control group ( p < 0.001, n = 6), and showed an inhibitory effect on

lymphocyte transformation by comparing with a reference compound (dexamethasone).

Q. Shen et al. / Chinese Chemical Letters 19 (2008) 453–456 455

Fig. 2. The HMBC and NOE correlations of 1 and 2.

Table 2

Inhibitiory effects of compound 1-12 on lymphocyte

Compounds Inhibition (%)

30 (mg/mL) 10 (mg/mL)

1 18.14 10.30

2 70.83 13.21

3 71.82 59.32

4 62.99 26.85

5 22.65 11.32

6 9.98 2.69

7 4.56 �4.56

8 71.63 14.09

9 14.02 10.81

10 70.06 9.02

11 60.52 55.95

12 7.69 0.10

Inhibition of dexamethasone 56.94% (50 mg/mL).

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