Zu-Yan Wu‡
a,
Yang Wu‡a,
Guo-Dong Chen*b,
Dan Hua,
Xiao-Xia Lia,
Xiang Sunc,
Liang-Dong Guoc,
Yan Lid,
Xin-Sheng Yao*a and
Hao Gao*a
aInstitute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, Guangzhou 510632, PR China. E-mail: tghao@jnu.edu.cn; tyaoxs@jnu.edu.cn
bDepartment of Pharmaceutical Engineering, College of Natural Resources and Environment, South China Agricultural University, Guangzhou 510642, PR China. E-mail: chgdtong@163.com
cState Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100190, PR China
dState Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650204, PR China
First published on 17th October 2014
Four new sesquiterpenoids, sesquiterpenoids A–D (1–4), were isolated from solid cultures of the Xylariaceae fungus (no. 63-19-7-3). Their structures were determined through NMR analyses, CD calculation, the in situ dimolybdenum CD method, the modified Mosher's method, and X-ray data analysis. The cytotoxicities of all compounds against HL-60, SMMC-7721, A-549, MCF-7 and SW480 human cancer cell lines were assayed.
No. | 1 | 2 | 3 | 4 | ||||
---|---|---|---|---|---|---|---|---|
δH | δC | δH | δC | δH | δC | δH | δC | |
1 | 2.07, d (9.1), a | 40.8 | 2.08, d (9.5), a | 40.8 | 2.17, m, a | 30.2 | 2.05, m, a | 28.9 |
2.80, dt (9.1, 5.3), b | 2.80, dt (9.5, 5.5), b | 2.31, m, b | 2.15, m, b | |||||
2 | 2.45, br t (5.8) | 48.6 | 2.47, br t (5.7) | 48.4 | 7.09, m | 141.5 | 5.63, br s | 122.0 |
3 | 170.6 | 170.6 | 128.8 | 136.3 | ||||
4 | 5.73, q (1.0) | 121.5 | 5.74, q (1.4) | 121.5 | 1.80, m, a | 21.4 | 1.67, m, a | 23.1 |
2.31, m, b | 1.96, m, b | |||||||
5 | 204.4 | 204.2 | 1.48, m, a | 25.2 | 1.50, m, a | 25.4 | ||
1.95, m, b | 1.87, m, b | |||||||
6 | 2.71, br t (5.8) | 55.5 | 2.71, br t (5.7) | 55.8 | 45.4 | 45.7 | ||
7 | 57.1 | 57.1 | 146.4 | 146.8 | ||||
8 | 1.73, td (12.8, 4.0), a | 35.6 | 1.90, td (12.4, 4.5), a | 35.7 | 2.17, m, a | 30.0 | 2.17, m, a | 30.1 |
2.31, td (12.8, 4.0), b | 2.17, td (12.4, 4.5), b | 2.31, m, b | 2.33, m, b | |||||
9 | 1.34, m, a | 26.5 | 1.34, m, a | 26.5 | 1.47, m, a | 31.9 | 1.45, m, a | 32.0 |
1.48, m, b | 1.50, m, b | 1.86, m, b | 1.82, m, b | |||||
10 | 3.33, dd (10.3, 1.6) | 78.8 | 3.36, dd (10.4, 1.8) | 78.8 | 3.88, dd (11.9, 4.6) | 73.3 | 3.87, dd (11.9, 4.7) | 73.5 |
11 | 73.2 | 73.3 | 41.9 | 42.0 | ||||
12 | 1.16, s | 23.3 | 1.17, s | 23.2 | 0.76, s | 15.1 | 0.73, s | 15.0 |
13 | 1.21, s | 26.5 | 1.23, s | 26.7 | 1.02, s | 20.4 | 1.01, s | 20.4 |
14 | 0.96, s | 19.0 | 0.98, s | 19.0 | 4.44, br s, a | 112.0 | 4.55, br s, a | 111.9 |
4.92, br s, b | 4.92, br s, b | |||||||
15 | 2.00, d (1.0) | 23.6 | 2.01, d (1.4) | 23.6 | 171.7 | 3.92, br s | 67.2 |
The partial relative configuration of 1 was established by the NOESY. The NOESY correlations between Hb-1 and Ha-8/Hb-8 indicated the relative configurations of C-2, C-6 and C-7 in 1 were 2S*, 6S*, and 7S*, respectively. The absolute configurations of C-2, C-6 and C-7 were determined by comparison of the experimental and the simulated circular dichroism (CD) spectra. Since the conformations of the flexible side chain in 1 had an insignificant effect on the CD spectrum, the simplified structures (2R,6R)-5 and (2S,6S)-5 were used for CD calculations (Fig. 3). The preliminary conformational analyses for a pair of enantiomers (2R,6R)-5 and (2S,6S)-5 were performed with Conflex version 7.0 via the MMFF94S force field. The corresponding minimum geometries were further fully optimized by DFT at the B3P86/6-31G(d) level as implemented in the Gaussian 09 program package. The obtained stable conformers were submitted to CD calculation by TDDFT [B3P86/6-311++G(2d,p)] method.5–8 The result showed that the experimental CD spectrum of 1 and the calculated CD spectrum of (2S,6S)-5 have similar negative Cotton effects (CEs) in the region of 320–350 nm (Fig. 3), which suggested that the absolute configurations of C-2, C-6 and C-7 in 1 were 2S, 6S, and 7S, respectively. In addition, the absolute configuration of the C-10 of 1 was determined by the in situ dimolybdenum CD method.9,10 The negative Cotton effect (∼310 nm), observed in the induced CD spectrum, permitted the assignment of 10R configuration according to the empirical rule proposed by Snatzke (Fig. 4). On the basis of the above deduction, the absolute configuration of 1 was assigned as 2S, 6S, 7S, 10R. Thus, the structure of 1 was established as (1S,5S,6S)-6-((R)-3,4-dihydroxy-4-methylpentyl)-4,6-dimethylbicyclo[3.1.1]hept-3-en-2-one, named as xylariterpenoid A.
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Fig. 3 Experimental CD spectrum of 1 in MeOH and calculated CD spectra of a pair of enantiomers of the corresponding simplified structures [(2R,6R)-5 and (2S,6S)-5]. |
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Fig. 4 CD spectra of 1 (red) and 2 (blue) in DMSO containing Mo2(OAc)4 with the inherent CD spectrum subtracted. |
Xylariterpenoid B (2) was isolated as a oil with the same molecular formula as 1. Except for the chemical shifts of protons around C-10 (Ha-8, Hb-8, Hb-9, H-10, and H3-12, H3-13, H3-14), the 1H and 13C NMR data (Table 1) of 2 were similar to those of 1, which disclosed that both the skeleton and the functional groups presented in 2 were same as those of 1, and suggested that 2 was a epimer of 1 different at C-10. The 1H, 1H–COSY and HMBC correlations (the details are available in the ESI†) observed fully supported that 2 had the same planar structure as that of 1. The NOESY correlation between Hb-1 and Ha-8 indicated that the relative configurations of C-2, C-6 and C-7 were also the same as 1, which confirmed that 2 was a epimer of 1 different at C-10. In the CD spectrum, 2 showed the same Cotton effects as 1 (Fig. 5), indicating that the absolute configurations of C-2, C-6 and C-7 were S. However, the positive Cotton effect at around 310 nm, observed in the in situ dimolybdenum CD spectrum, permitted the assignment of 10S configuration for 2 (Fig. 4). On the basis of the above data, the absolute configuration of 2 was assigned as 2S, 6S, 7S, 10S. Thus, the structure of 2 was determined as (1S,5S,6S)-6-((S)-3,4-dihydroxy-4-methylpentyl)-4,6-dimethylbicyclo[3.1.1]hept-3-en-2-one, named as xylariterpenoid B.
Xylariterpenoid C (3) was assigned as C15H22O3 (five degrees of unsaturation) on the basis of its HRESIMS at m/z 273.1465 [M + Na]+ (calcd for C15H22O3Na, 273.1467). The IR spectrum showed bands corresponding to hydroxyl (3439 cm−1). The 1H NMR spectrum (Table 1) exhibited two methyls, five sp3 methylenes, one oxygenated sp3 methine and three olefinic protons. The 13C NMR and DEPT spectra of 3 (Table 1) displayed 15 carbon signals consisting of two methyls, five sp3 methylenes, one oxygenated sp3 methine, two sp3 quaternary carbons, four olefinic carbons, and one carbonyl carbon. The proton resonances were assigned to relevant carbon atoms through the HSQC experiment. Interpretation of the 1H, 1H-COSY spectrum of 3 identified three isolated proton spin systems, which were C-1–C-2, C-4–C-5, and C-8–C-9–C-10 (Fig. 2). Combined with the analysis of 1H, 1H-COSY spectrum, the HMBC correlations from H3-13 to C-6/C-10/C-11/C-12, from H3-12 to C-6/C-10/C-11/C-13, from H2-14 to C-6/C-8, from H-2 to C-4/C-6/C-15, from Hb-4 to C-3/C-6/C-15, from Hb-5 to C-1 and from Ha-5 to C-6/C-7 completed the planar structure of 3 (Fig. 1).
The relative configuration of 3 was established by the NOESY data analysis. The orientation of H-10 was the same as those of H2-5 and H3-13, which was determined by the NOESY correlations between H-10 and H3-13/Hb-5. Thus, the relative configuration of 3 was designated as 6R*, 10S*. The absolute configuration of 3 was determined by the modified Mosher method.11 The Δδ values of the (S)- and (R)-MTPA esters (3a and 3b) indicated the S configuration for C-10 (Fig. 6). On the basis of the above deduction, the absolute configuration 6R, 10S was proposed for 3. Therefore, the structure of 3 was elucidated as (6R,8S)-8-hydroxy-7,7-dimethyl-11-methylenespiro[5.5]undec-2-ene-3-carboxylic acid, named as xylariterpenoid C.
Xylariterpenoid D (4) was isolated as a crystal. Its HRESIMS analysis gave a quasi-molecular ion peak at m/z 259.1667 [M + Na]+ (calcd for C15H24O2Na, 259.1674), 14 mass units less than 3. The 1D NMR data of 4 revealed its structural features were similar to those of 3, except that the resonance for C-15 [δC 171.7 (C-15)] was replaced by an oxygenated methylene group [δC/H 67.2 (C-15)/3.92 (br s, H2-15)]. At the same time, the 1H, 1H-COSY and HMBC correlations (the details are available in the ESI†) observed fully supported the locations of functional groups. Thus, the planar structure of 4 was established as shown in Fig. 1.
The relative configuration of 4 was established on the basis of the NOESY data. The NOESY correlation between H-10 and Hb-5 indicated that 4 had the same relative configuration with 3. In the meanwhile, the relative configuration of 4 was confirmed by single-crystal X-ray crystallographic analysis (Fig. 7). Furthermore, the values of the Flack parameter (−0.1(3)) and the Hooft y parameter 0.04(16) allowed the assignment of the absolute configuration of 4 as 6R, 10S. Therefore, the structure of 4 was determined as (2S,6R)-9-(hydroxymethyl)-1,1-dimethyl-5-methylenespiro[5.5]undec-8-en-2-ol, named as xylariterpenoid D.§
Footnotes |
† Electronic supplementary information (ESI) available: The general experimental procedure, fungus material, extraction and isolation, spectroscopic data of 1–4, quantum chemical CD calculation of 5, single-crystal X-ray data of 4, the in situ dimolybdenum CD method, preparation of (S)- and (R)-MTPA esters of 3 (3a and 3b), cytotoxicity assay, and 1D and 2D NMR spectra of compounds 1–4. CCDC 1023174. For ESI and crystallographic data in CIF or other electronic format see DOI: 10.1039/c4ra10365c |
‡ These authors have contributed equally to this work. |
§ Crystal data of xylariterpenoid D (4): data were collected using a Sapphire CCD with a graphite monochromated Cu Kα radiation, λ = 1.54184 Å at 173.00 (10) K. Crystal data: C15H24O2, M = 236.34, orthorhombic, space group P212121; unit cell dimensions were determined to be a = 6.5259(3) Å, b = 13.1402(5)Å, c = 15.6863(7) Å, α = 90.00°, β = 90.00°, γ = 90.00°, V = 1345.13(10) Å3, Z = 4, Dx = 1.167 g cm−3, F (000) = 520, μ (Cu Kα) = 0.586 mm−1. 10![]() |
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