Two new triterpenoid glycosides from the stems of Camellia oleifera Abel

Abstract Two new oleanane-type triterpenoid glycosides, 3-O-β-D-xylopyranosyl-(1→2)-α-L-arabinopyranosyl-(1→3)-[β-D-glucuronopyranosyl-(1→2)]-β-D-glucuronopyranosyl-22α-angeloyloxyolean-12-ene-15α,16α,28-triol(1) and 3-O-β-D-xylopyranosyl-(1→2)-α-L-arabinopyranosyl-(1→3)-[β-D-glucuronopyranosyl-(1→2)]-β-D-glucuronopyranosyl-21β-acetyl-22α-angeloyloxyolean-12-ene-16α,28-diol (2) were isolated from the stems of Camellia oleifera Abel. Their structures were elucidated by means of spectroscopic methods and chemical evidence. The cytotoxic activities of compounds 1–2 were evaluated against five human tumour cell lines (HCT-8, BGC-823, A5049, and A2780). Compounds 1–2 showed cytotoxic activity against five human cancer cell lines, with IC50 values ranging from 3.15 to 7.32 μM.


Results and discussion
The 95% etoH extract of the stems of C. oleifera was partitioned with CHCl 3 , etoAc and n-BuoH, successively. The n-BuoH-soluble portion was separated by a combination of silica gel, D101 macroporous resin chromatography and preparative HPLC (high-performance liquid chromatography) afforded two new compounds (1 and 2) (Figure 1). Their structures were elucidated by extensive NMR techniques including 1D NMR ( 1 H and 13 C NMR), 2D NMR (CoSY, NoeSY, HSQC and HMBC) and HReSIMS, as well as chemical evidence.

General experimental procedures
optical rotations were measured on an Autopol IV-T/V (Rudolph Research Analytical, New Jersey, uSA). uV spectra were recorded in MeoH on a Jasco V650 spectrophotometer (JASCo, Inc., easton, Maryland, uSA). The 1 H (600 MHz), 13 C (150 MHz) and 2D NMR spectra were recorded on a Bruker AVANCe → 600 instrument using TMS (Tetramethylsilane) as an internal reference (Bruker Company, Massachusetts, uSA). 1D and 2D NMR spectra were obtained at 500 and 125 MHz for 1 H and 13 C, respectively, on an INoVA 500 MHz spectrometer in pyridine-d 5 with solvent peaks as references. HReSIMS data were obtained on an Agilent 7890-7000A mass spectrometer (Agilent Technologies, Santa Clara, uSA). Preparative HPLC was conducted with an Angilent Technologies 1200 series instrument with a MWD detector using a YMC-pack oDS (octadecylsilyl)-A column (5 μm, 250 × 20 mm). Column chromatography (CC) was performed with silica gel (200-300 mesh, Qingdao Marine Chemical Ltd., Qingdao, China), Develosil oDS (50 μm, Nomura Chemical Co. Ltd., osaka, Japan), Sephadex LH-20 (Ge Healthcare Bio-Sciences AB, uppsala, Sweden). TLC (thin-layer chromatography) was carried out with glass precoated with silica gel GF 254 . Spots were visualised under uV light or by spraying with 10% sulphuric acid in etoH followed by heating.

Plant material
The stems of C. oleifera were collected from Guangfeng, Jiangxi, China, in May 2014 and identified by Prof. Guiping Yuan at Jiangxi Provincial Institute for Drug Control, China. A voucher specimen (No. 20140515) has been deposited in the Herbarium of Jiangxi Provincial Institute for Drug Control. The 95% etoH extract of the stems of C. oleifera was partitioned with CHCl 3 , etoAc and n-BuoH, successively.

Determination of absolute configurations of the sugar moieties in 1-2
The determination of the absolute configuration of the sugars in compounds 1-2 was conducted as described previously (Zhong et al. 2013).

Statistical analysis
The results of the cytotoxicity were expressed as means ± SD. Student's t test was used to determine statistical comparisons between the data-sets. p < 0.05 was considered to be significant.

Supplementary material
Supplementary material relating to this article is available online, alongside Figures S1-S16.

Funding
Financial support was provided by the Natural Science Foundation of Jiangxi, China [grant number 20142BAB215062].