Two New Compounds from Schisandra propinqua var. propinqua

Abstract Schisanpropinoic acid (1), a new bergamotane sesquiterpenoid, and schisanpropinin (2), a new tetrahydrofuran lignan with a rare epoxyethane unit, were identified from the stems and leaves of Schisandra propinqua var. propinqua. Their structures were determined based on comprehensive spectroscopic and mass spectrometric analysis. The absolute configuration of 1 was determined by X-ray analysis. Compounds 1 and 2 were tested for their cytotoxic activity against five human tumor cell lines. Graphical Abstract Electronic supplementary material The online version of this article (doi:10.1007/s13659-017-0129-7) contains supplementary material, which is available to authorized users.


Introduction
Schisandraceae family are a group of economically and medicinally valuable climbing woody vine plants, some of which are used as traditional Chinese medicines in Chinese folk for over 2000 years [1]. Since 1970s, researches on the plants of Schisandraceae family have been a hot topic [2]. In 2003, micrandilactone A, a highly oxygenated, skeleton rearranged, polycyclic nortritepenoid, was reported from S. micrantha by our group [3]. This new finding pushed the studies on the plants of Shisandraceae to a new climax. As a result, last ten years have seen the isolation and identification of large numbers of Schisandra nortriterpenoids (SNTs) from the Schisandraceae family [2,4]. S. propinqua (Wall.) Baill. var. propinqua is indigenous in Yunnan Province. Previous chemical investigation into this species has led to the identification of a series of new terpenoids and lignans, such as propindilactones E-O [5,6], lanopropic acid [7], methylisogomisin O [8], and propindilactones T and U [9]. Our further study on this species, which was collected from Weixi county, Yunnan Province, People's Republic of China to discover new compounds with interesting bioactivity led to the discovery of a new sesquiterpenoid and a new lignan (Fig. 1). This article discribes the isolation, structure elucidation and bioactivities of these two new compounds.
Even though the structure of 1 was deduced from extensive spectroscopic analysis, no available correlations in the ROESY spectrum were observed (Fig. 2). Thus, the structure of 1, especially the absolute configuration still needed to be further confirmed. Luckily, a single-crystal X-ray diffraction was performed successfully (Fig. 3). Through structural refinement, the Flack parameter with 0.0(4) and the Hooft parameter at 0.03 (17) for 1371 Bijvoet pairs [14,15], allowed an explicit assignment of the absolute configuration of 1 as 1S, 5S, 6S. Finally, the structure of 1 was determined, and named schisanpropinoic acid.
Compound 2, obtained as a yellowish solid (MeOH). Its IR absorptions at 3424, 1607, 1518 and 1462 cm -1 implied the presence of OH and phenyl groups. Its molecular formula was determined as C 20 (Table 1) displayed only ten carbon signals (six aromatic carbons, one oxygenated sp 3 quaternary carbon, one oxygenated sp 3 methine, one oxygenated methylene, and one methoxy group), suggesting the symmetrical structure of 2. The 1 H NMR signals (Table 1)   1,2,4-trisubstituted benzene. This deduction was also confirmed by the 1 H-1 H COSY (Fig. 4) correlation between H-5 and H-6, as well as the HMBC correlations (Fig. 4) from H-7 (d H 5.47, 1H, s) to C-1 (d C 129.9), C-2 (d C 113.2) and C-6 (d C 121.6). Taking the lignans from Schisandraceae family into consideration, the above-mentioned data indicated that 2 is identical to the characteristics of a 4H-furan-type lignan with an extra oxygen-containing ring. The HMBC correlations from H-2 and H-6 to C-4, and from methoxyl (d H 3.72, 3H, s) to C-3 (d C 148.6) suggested that the methoxy group was located at C-3 rather than C-4. The oxygen-containing ring was deduced to be formed between C-8 and C-8 0 by HMBC correlations from H-7 to C-8 (d C 89.4), from H-9 (d H 4.54 2H, s) to C-8, and from H-7 to C-9 (d C 77.0). Thus, the planar structure of 2 was determined. In the ROESY spectrum, both the correlation (Fig. 4) between H-9 and H-7 as well as the confusing correlations of H-2, H-6 and H-9 were observed. Then, a computer-aided 3D molecular model study of two possible relative structure of 2 was conducted. As a result, when H-9 and H-7 were at a same orientation, the internuclear distance between H-9 and H-7 was less than 3.0 Å . On the contrary, the internuclear distance was lager than 3.0 Å . At the same time, the confusing ROESY correlations of H-2, H-6 with H-9 could be explained by the free rotation of the C 1 -C 7 bond. Considering all these factors, the chemical structure of 2 with a relative configuration was determined (Fig. 1). 2 was named schisanpropinin and was a tetrahydrofuran lignan with a rare epoxyethane unit from Schisandraceae family [27].

General
1D and 2D NMR spectra were recorded on Bruker DRX 500 or 600 spectrometers using C 5 D 5 N as the internal standard. Chemical shifts (d) are expressed in ppm relative to the C 5 D 5 N signals. HRESIMS was performed on an API QSTAR Pulsar i spectrometer. UV spectra were obtained on a Shimadzu UV-2401PC spectrophotometer. IR spectra were obtained on a Bruker Tensor-27 FT-IR spectrometer using KBr pellets. Optical rotations were measured in MeOH with JASCO P-1020 polarimeters. Column chromatography (CC) was performed with silica gel (100-200 mesh; Qingdao Marine Chemical, Inc., Qingdao, People's Republic of China), MCI gel (CHP20P, 75-150 lm, Mitsubishi Chemical Corporation, Tokyo, Japan). Semi-

The Cytotoxicity Assay
The human tumor cell lines HL-60, SMMC-7721, A-549, MCF-7, and SW-480 were used in the cytotoxic assay. These cell lines were obtained from ATCC (Manassas, VA, USA). Cells were cultured in RMPI-1640 or DMEM medium (Biological Industries, Kibbutz Beit-Haemek, Israel) supplemented with 10% fetal bovine serum (Biological Industries) at 37°C in a humidified atmosphere with 5% CO 2 . The cytotoxicity assay was evaluated by the   (7) Table 1.