A new acorane sesquiterpene from Lysionotus pauciflorus

Chinese Journal of Natural Medicines 2013, 11(2): 0185−0187

Chinese Journal of Natural Medicines

A new acorane sesquiterpene from Lysionotus pauciflorus WEN Qing-Fa, ZHONG Yu-Jiao, SU Xue-Hui, LI Cong-Ying, LIU Jing, LI Yan-Fang*, LIANG Bing Department of Pharmaceutics and Bioengineering, School of Chemical Engineering, Sichuan University, Chengdu 610065, China Available online 20 Mar. 2013

[ABSTRACT] AIM: To study the chemical constituents of the whole plant of Lysionotus pauciflorus. METHODS: Chromatographic separations on silica gel, Toyopearl HW-40F gel, and MCI gel were used to isolate the compounds. The structures were elucidated on the basis of spectral data. RESULTS: Three compounds were obtained and their structures were identified as 3, 10-dihydroxyacoronene (1), bis(2-butylhexyl)phthalate (2) and 3, 5-dimethoxy-4-hydroxy-trans-stilbene (3). CONCLUSION: Compound 1 is a new acorane sesquiterpene, this is the first report of acorane sesquiterpenes from the Lysionotus genus. [KEY WORDS] Lysionotus pauciflorus; Gesneriaceae; Sesquiterpene

[CLC Number] R284.1

1

[Document code] A

[Article ID] 1672-3651(2013)02-0185-03

Introduction

Lysionotus pauciflorus Maxim., one species of the Lysionotus genus (Gesneriaceae), is widespread in southern China, and has been used in traditional Chinese medicine for the treatment of lymph node tuberculosis, cough with tachypnoea and rheumatic pains [1]. Nevadensin, one of the main components of this plant, showed significant anti-tuberculosis, anti-inflammatory, and antitussive activities [2-5]. The previous phytochemical studies on this species led to the isolation of flavonoids, phenylethanols, β-sitosterol, and ursolic acid [6-11]. In the course of a search for anti-inflammatory active compounds, the constituents of an ethanolic extract of the whole plant of L. pauciflorus were investigated. The current paper describes the isolation and structure elucidation of a new acorane sesquiterpene (1).

2

Results and Discussion

Compound 1, a colorless oil, was assigned the molecular formula C15H24O3, as determined by positive HR-ESI-MS

[Received on] 02-Feb.-2012 [*Corresponding author] Li Yan-Fang: Associate Prof., Tel: 86-2885405221, E-mail: [email protected] These authors have no conflict of interest to declare. Published by Elsevier B.V. All rights reserved

([M + H]+ at m/z 253.179 4, Calcd. for 253.180 4). The 1H and 13C NMR spectroscopic data exhibited the presence of a trisubstituted olefin conjugated to a carbonyl group [δH 6.61 (1H, s), 1.72 (3H, t, J = 3.2 Hz); δC 134.1 (s), 150.7 (d), 200.3 (s)], two oxygenated methine [δH 4.74 (1H, s), δC 70.0 (d); δH 3.46 (1H, q, J = 6.8 Hz ), δC 77.8 (d)], the characteristic signals of an isopropyl group at δH 0.89 (3H, d, J = 7.2 Hz), 0.97 (3H, d, J = 6.8 Hz), and the spiro carbon at δC 53.0 (s) of acorane-type sesquterpene. Detailed 2D NMR analysis showed that compound 1 is an acorane type sesquiterpenoid[12]. Further analysis of the data of 1 (Table 1) showed that it was similar to the known compound 10-hydroxyacoronene from Illicium henryi [13]. The 10-hydroxy group was unambiguously confirmed by the HMBC experiment, which showed the correlations of H-10 (δH 4.74) with C-8 (δC 134.1) and C-9 (δC 150.7) (Fig. 1). The 3-hydroxy group was assigned by HMBC correlations from H-2 (δH 2.03, 1.58), H-4 (δH 2.19), and H-14 (δH 0.89) to C-3. The relative configuration of 1 was established by the ROESY spectrum, namely, the correlations between H-1 (δH 1.31) and H-4 (δH 2.19), and H-4 (δH 2.19) and H-10 (δH 4.74). These facts indicated that the relative configuration of 1 was as shown in Fig. 1. The skeleton and the relative configuration of compound 1 were similar to AC-1, but the CD spectrum data of compound 1 was different from that of AC-1 [12]. In the CD spectrum of 1, a strong negative Cotton

WEN Qing-Fa, et al. /Chinese Journal of Natural Medicines 2013, 11(2): 185−187

effect at 245 nm (Δε = −23.06), which was based on the π→π* transition of a conjugated cyclohexanone [14], and a small positive Cotton effect at 329 nm (Δε = + 0.85), which was based on the n→π* transition of an α, β-unsaturated ketone [15] are visible. Based on the above data, the absolute configuration of 1 was concluded to be 1R, 3S, 4R, 5R, 10R. Thus, on the basis of above evidence, compound 1 was determined to be 3, 10-dihydroxyacoronene. Table 1 1H (400 MHz) and 13C NMR (100 MHz) data for compound 1 in CD3OD (J in Hz) No.

δH

δC

HMBC

1

1.31 (1H, m)

52.9 d

10,12

2

2.03 (1H, m), 1.58 (1H, dd)

37.3 t

1, 3, 11

3

3.46 (1H, q, 6.8)

77.8 d

1, 14

4

2.19 (1H, q)

44.4 d

1, 3, 10, 14

5

53.0 s 2.42 (1H, d, 16.8), 2.55 (1H, d 17.2)

6

46.2 t

7

200.3 s

8

134.1 s

1, 4, 7, 8, 10

9

6.61 s

150.7 d

7, 10, 15

10

4.74 br s

70.0 d

4, 5, 8, 9

11

1.98 (1H, m)

27.7 d

4, 13

12

1.03 (1H, d, 6.4)

23.6 q

1, 11, 13

13

0.97 (3H, d, 6.8)

21.0 q

1, 12, 13

14

0.89 (3H, d, 7.2)

15.0 q

3, 4, 5

15

1.72 (3H, t, 3.2)

13.6 q

7, 8, 9

Fig. 1 Structure and key NOESY correlations of 1

The two known compounds, bis (2-butylhexyl) phthalate (2) and 3, 5-dimethoxy-4-hydroxy-trans-stilbene (3), were also isolated and identified by comparing their spectral data with literature values [16-17].

3 3.1

Experimental

General Optical rotation was measured on a WZZ-3 polarimeter (Shanghai Shenguang Co., Shanghai, China) using a 0.8 mL cell. The CD spectrum was determined on a ChirascanTM circular dichroism spectroscopy instrument (Applied Photophysics Ltd, UK). UV spectra were obtained on a UV-2100 UV spectrophotometer (Rayleigh Co. Beijing, China). The high resolution mass spectrometric analysis was performed by quadrupole time of flight (Q-TOF) premier spectrometer

coupled with an ESI source (Micromass, Simonsway, Manchester, UK). The 1H and 13C NMR spectra were recorded on a Bruker AM-400 spectrometer with TMS as the internal standard. The chemical shifts were reported in ppm using CD3COCD3, CD3OD as solvents. Silica gel (200−300 mesh) for column chromatography and GF254 (10−40 μm) for TLC were supplied by the Qingdao Marine Chemical Inc., China. Spots on TLC were monitored under UV lamp and by heating silica gel plates sprayed with 10% H2SO4 in EtOH. Toyopearl HW-40F gel (Tosoh, Japan) and MCI gel CHP-20P (Mitsubishi Kasei Industry Co., Ltd., Japan) were used for column chromatography. 3.2 Plant material The whole plant of L. pauciflorus were purchased in May 2010 from a local herbal medicine store in Chengdu, Sichuan Province, China, and identified by Dr. LI Yan-Fang (Sichuan University). A voucher specimen (D201002) was deposited at the Department of Pharmaceutics & Bioengineering, Sichuan University, Chengdu, China. 3.3 Extraction and isolation The air dried whole plants (5.0 kg) of L. pauciflorus were powdered and extracted with 95% ethanol (25 L × 72 h) for four times at room temperature. The combined extracts were concentrated under reduced pressure to afford a dark brown residue (387 g). This residue was suspended in distilled water (2 L) and partitioned successively between petroleum ether (PE, 60−90 °C, 5 × 1.2 L), ethyl acetate (5 × 1.2 L), and n-BuOH (3 × 1.2 L). Each fraction was concentrated under reduced pressure to give PE crude extract (72 g), ethyl acetate crude extract (44 g), and n-BuOH crude extract (60 g). The EtOAc crude extract (44 g) was subjected to silica gel CC (160 g, 50 mm × 470 mm) using PE-acetone with increasing polarity (100 : 1-1 : 5) collecting 151 fractions of 250 ml each which were combined on the basis of TLC analysis to afford 15 fractions (Fr. A to Fr. O). The Fr. E (720 mg) was chromatographed over MCI gel (18 mm × 320 mm, acetone-H2O, 1 : 1 to 1 : 0) and six fractions (Frs. E1-E6) collected. The Fr. E4 (28 mg) eluted with 70% acetone was rechromatographed over Toyopearl HW-40F gel CC (18 × 650 mm, CH2Cl2-MeOH, 1 : 1) to give 3 (8 mg). The Fr. E6 (60 mg) eluted with 80% acetone followed by PTLC afforded 1 (19 mg). The Fr. H (750 mg) was further fractionated on a MCI gel column (18 mm × 320 mm, acetone-H2O, 1 : 1 to 1 : 0) to afford six fractions (Frs. H1-H6). The Fr. H2 (138 mg) was further separated by Toyopearl HW-40F column (11 mm × 530 mm, CH2Cl2MeOH, 1 : 1) and followed by PTLC yielded 1 (60 mg).

4

Identification

3, 10-Dihydroxyacoronene (1): Colorless powder; [α]20D −82.1 ° (c 0.0525, CH3OH); CD (MeOH, c 0.194) 245, and MeOH 329 nm (Δε −23.06 , +0.85); UV λmax (nm):(log ε ): 240

(1.40); 1H and

13

C NMR: see Table 1. HR-ESI-MS m/z

WEN Qing-Fa, et al. /Chinese Journal of Natural Medicines 2013, 11(2): 185−187

253.179 4 [M + Na]+ (Calcd. for C15H25O3: 253.180 4). Bis(2-butylhexyl)phthalate (2): Colorless powder; ESI-MS m/z 579.2 [2M + Na]+ (C32H44O8Na); 1H NMR (CD3COCD3, 400 MHz) δ: 7.65 (2H, dd, C3, C6-H), 7.75 (2H, dd, C4, C5-H), 4.27 (4H, t, C8, C8′-H), 1.72 (4H, m, C9, C9′-H), 1.43 (4H, m, C10, C10′-H), 0.95 (6H, t, C11, C11′-H); 13C NMR (CD3COCD3, 100 MHz) δ: 132.5 (C-1, 2), 128.7 (C-3, 6), 131.1 (C-4, 5), 167.1 (C-7, 7′), 65.0 (C-8, 8′), 30.4 (C-9, 9′), 18.9 (C-10, 10′), 13.1 (C-11, 11′). 3, 5-Dimethoxy-4-hydroxy-trans-stilbene (3): pale yellow solid; HR-ESI-MS (m/z): 257.1174 [M + H]+ (Calcd. for C16H17O3: 257.117 8); 1H NMR (CD3COCD3, 400 MHz) δ: 6.37 (1H, t, J = 2 Hz, C6′-H), 6.73 (2H, d, J = 2.0 Hz, C4′, 8′-H), 6.86 (2H, d, J = 8.4 Hz, C2, C6-H), 6.98 (1H, d, J = 16.0 Hz, C2′-H), 7.45 (2H, d, J = 8.4 Hz, C3, C5-H), 7.18 (1H, d, J = 16.4Hz, C1′-H).

[5]

[6]

[7] [8]

[9]

[10]

[11]

Acknowledgments NMR measurements afforded by the Centre of Testing & Analysis (Sichuan University) and CD spectral measurements provided by the Kunming Institute of Botany (Chinese Academy of Sciences) are gratefully acknowledged.

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石吊兰全草中的一个新菖蒲烷类倍半萜 文庆发，钟玉蛟，苏雪会，李聪颖，刘

静，李延芳*，梁

冰

四川大学化学工程学院制药与生物工程系，成都 610065 【摘 要】 目的：研究石吊兰全草的化学成分。方法：采用硅胶、MCI 等多种层析柱分离手段。运用 NMR 和 MS 等波谱 技术鉴定化合物的结构。结果：从石吊兰全草中分离并鉴定了 3 个化合物：3, 10-dihydroxyacoronene (1)、邻苯二甲酸二丁酯 (2) 和 3', 5'-二甲氧基 4 羟基顺式二苯乙烯(3)。结论：化合物 1 为新的菖蒲烷类倍半萜，为首次从该属植物中分离得到的菖蒲烷类倍半 萜类化合物。 【关键词】 石吊兰；苦苣苔科；倍半萜