1α-Methoxy-3-oxo-8α-hydroxy-10αH-eremophila-7(11)-en-12,8β-olide from Ligularia fischeri

Abstract

One unreported eremophilane sesquiterpenoid, 1α-methoxy-3-oxo-8α-hydroxy-10αH- eremophila-7(11)-en-12,8β-olide (1), was isolated from Ligularia fischeri. The structure of 1 was identified by detailed 1D and 2D NMR and HRMS analyses.

1. Introduction

Ligularia fischeri Turcz. is a perennial herb belonging to the Asteraceae family. The traditional Chinese medicine “Shanziwan” is derived from the roots of L. fischeri and is used to treat bronchitis and asthma [1]. The L. fischeri plantlets are also utilized for the amelioration of indigestion, pulmonary abscess and toxemia [2]. During our investigation of natural products from L. fischeri [3], a previously undescribed eremophilane sesquiterpenoid, 1α-methoxy-3-oxo-8α-hydroxy-10αH-eremophila-7(11)-en-12,8β-olide (1), was obtained from the title plant. The structure of 1 was identified by HRMS and NMR analyses.

2. Results and Discussion

One novel eremophilane derivative 1α-methoxy-3-oxo-8α-hydroxy-10αH-eremophila- 7(11)-en-12,8β-olide (1) was purified from the whole plant of L. fischeri (Figure 1).

Compound 1, obtained as yellow powder, possessed the molecular formula C₁₆H₂₂O₅ based on the molecular ion peak at m/z 295.1542 [M + H]⁺ (calcd for C₁₆H₂₃O₅, 295.1545) in HRESIMS. The IR spectrum showed the absorbance at 3355 cm⁻¹, 1758 cm⁻¹ and 1712 cm⁻¹ for a hydroxyl group, an α, β-unsaturated-γ-lactone and a ketone carbonyl, respectively [4]. The ¹H NMR spectrum showed signals of four methyl groups at δ_H 3.33 (3H, s), 1.78 (3H, s), 1.01 (3H, d, J = 6.7 Hz), 0.53 (3H, s) and an oxymethine at δ_H 3.22 (1H, m). The ¹³C NMR and DEPT spectra exhibited signals corresponding to 16 carbons. Except for one methoxyl group (δ_C 55.5), the remaining 15 carbons were classified into three methyls (δ_C 11.6, 6.6, 6.3), three methylenes (δ_C 46.0, 36.3, 35.7), three methines (δ_C 77.9, 54.5, 46.8), and six quaternary carbon (δ_C 209.1, 172.8, 158.7, 123.1, 103.6, 41.5). The signal at δ_C 103.6 was attributed to the hemiketal carbon at C-8 of 1 [5,6]. These spectroscopic features implied that 1 might be an eremophilenolide derivative, since they were isolated frequently from the genus Ligularia [7]. The NMR data of 1 was similar to the known compound fischerin A, except for the presence of one hydroxyl group in 1 instead of the methoxyl group at C-8 in fischerin A [3], which was supported by ¹H–¹H COSY correlations of H-2/H-1/H-10/H-9 and HMBC correlations of H₃-OMe to C-1, H₃-15 to C-3 and C-5, H₃-14 to C-4, C-6 and C-10, H₃-13 to C-7 and C-12, H-1 to C-9, and H-6 to C-8 (Figure 2). Consequently, the planar structure of 1 was determined.

The NOESY correlations established the relative configuration of 1 (Figure 2). The eremophilane sesquiterpenoids from Asteraceae generally exhibited the β-orientation for CH₃-14 and CH₃-15 [8]. The NOESY correlation of H₃-14/H₃-15 and H₃-14/H-1 revealed the β-orientation of H-1, while the presence of NOESY correlation of H-4/H-10 supported the α-orientation of H-10. Additionally, the OH-8 was assigned as α-oriented by comparing the proton signals of CH₃-14 and CH₃-15 based on the Naya rule [9]. The structure of 1 was therefore determined and named 1α-methoxy-3-oxo-8α-hydroxy-10αH-eremophila-7(11)-en-12,8β-olide.

3. Materials and Methods

3.1. General Experimental Procedures

The experimental or plant materials were as reported previously [3]. The IR spectrum (KBr) was acquired on a Bruker TENSOR 37 instrument (Bruker, Ettlingen, Germany). The NMR spectra were recorded in CD₃OD at room temperature by a Bruker AV 400 spectrometer (Bruker, Fallanden, Switzerland). HR-ESIMS spectrum was measured by an LTQ-Orbitrap mass spectrometer (Thermo Fisher Scientific, Bremen, Germany).

3.2. Extraction and Isolation

The whole plants of L. fischeri (9.8 kg) were extracted with methanol three times under reflux. The combined filtrate was evaporated in vacuo to yield a MeOH extract (975 g), which was further suspended in H₂O and partitioned with EtOAc. The EtOAc-soluble part (265 g) was chromatographed on silica gel column eluting with a petroleum ether/acetone (100:0–100:30, v/v) gradient to give eight fractions (F₁–F₈). Fraction F₆ was chromatographed on the MPLC (75–90% MeOH in H₂O) to afford eight subfractions F_6-1–F_6-8. The further purification of F_6-1 by semipreparative HPLC (54% MeOH in H₂O) gave compound 1 (t_R = 31.3 min, 10.9 mg).

1α-methoxy-3-oxo-8α-hydroxy-10αH-eremophila-7(11)-en-12,8β-olide (1): yellow powder, ${\left[\mathrm{\alpha }\right]}_D^{19}$-115.6 (c 0.06, CH₂Cl₂); IR (KBr) ν_max 3355, 2971, 1758, 1712, 1438, 1385, 1339, 1253, 1211, 1113, 1084, 955, 736, 697; ¹H-NMR (400 MHz, CD₃OD) and ¹³C-NMR (100 MHz, CD₃OD) data, see

Table 1. ¹H-NMR (400 MHz, CD₃OD) and ¹³C-NMR (100 MHz, CD₃OD) spectroscopic data for compound 1.

Position	δH (J = Hz)	δC	Position	δH	δC
1	3.22 m	77.9	9α	1.48 t (13.1)	35.7
2α	2.90 dd (13.3, 4.8)	46.0	9β	2.72 m
2β	2.39 m		10	2.28 m	46.8
3		209.1	11		123.1
4	2.68 m	54.5	12		172.8
5		41.5	13	1.78 s	6.6
6α	2.32 m	36.3	14	0.53 s	11.6
6β	2.70 m		15	1.01 d (6.7)	6.3
7		158.7	1-OMe	3.33 s	55.5
8		103.6

; HRESIMS m/z 295.1542 [M + H]⁺ (calcd for C₁₆H₂₃O₅ 295.1545).

4. Conclusions

An unreported member of the eremophilane class was isolated from L. fischeri based on a search across SciFinder. Its structure was identified as 1α-methoxy-3-oxo-8α-hydroxy-10αH-eremophila-7(11)-en-12,8β-olide through IR, HRESIMS and NMR analysis, which added a member to the Ligularia eremophilane family.