New Steroidal Erythrityl Triesters from the Heat Processed Roots of Panax ginseng

Two new compounds stigmasta-3α-ol-3α-(2R,3S)-butane-1,2,3,4-tetraolyl-2,3-dioctadec-9/9-enoyl-4-octadec-9,12dienoate (1) and stigmasta-5-en-3β-ol-3β-(2R,3S)-butane-1,2,3,4-tetraolyl-2,3-dioctadec-9/9-enoyl-4-octadec-9,12dienoate (2) along with β-sitosterol-β-D-glucoside were isolated and identified from the heat processed roots of Panax ginseng. The structures of the new compounds were elucidated by 1D and 2D NMR (COSY, HSQC, and HMBC) spectroscopic techniques aided by FAB-MS, ESI FT/MS, and IR spectra.


Introduction
Ginseng (Panax ginseng C. A. Meyer, Araliaceae) is one of the most important oriental medicinal plants in Japan, Korea and China [1].Of the two kinds of ginseng, white ginseng is air dried, and red ginseng is produced by steaming raw ginseng at 98-100 ∘ C for 2-3 h.It has been reported that red ginseng is more effective in pharmacological activities than white ginseng [2][3][4][5].The differences in biological activities and chemical constituents of red and white ginsengs have been reported.Anticancer properties and other pharmacological activities of Panax ginseng [6][7][8] have been studied and ginsenosides are recognized as active anticancer compounds [6].Compared with Asian white ginseng, red ginseng has stronger anticancer activities [9,10].Recently, there was a report using a steaming process to treat American ginseng root [5].In the study, however, the treatment temperature was 100 ∘ C and, thus, chemical constituents did not change significantly.
Anticarcinogenic and antidiabetic effects of P. ginseng have been reported [9,11].Several other compounds and biological activities have been reported from the ginseng roots of P. ginseng [12][13][14].The most well-known chemical constituent of ginseng is ginsenosides, which are dammarane glycosides.Dammarane glycosides were reported from many parts of ginseng and heat processed P. ginseng roots [15,16].The chemical and morphological variations of Panax notoginseng and their relationship were recently described [17].

Materials and Methods
Optical rotation was measured with an instrument on an AA-10 model polarimeter (Instruments Ltd,.Seoul, Republic of Korea).IR spectra were recorded on an Infinity Gold FT-IR (Thermo Mattson, Waltham, MA, USA) spectrophotometer, which was available at Korea Institute of Science and Technology, Seoul, Republic of Korea.Both 1 H and 13 C-NMR spectra were obtained on a Bruker Avance 600 high-resolution spectrometer operating at 600 and 150 MHz, respectively.This NMR machine was available at Seoul National University (SNU), Seoul, Republic of Korea, and all NMR spectra were recorded at SNU (Instrument, Bruker, Germany).NMR spectra were obtained in deuterated chloroform using tetramethylsilane (TMS) as an internal standard, with chemical shifts expressed in ppm () and coupling constants () in Hz.FAB/MS data were recorded on a JMS-700 (Jeol, Mitaka, Japan) spectrometer instrument which was available at SNU, Seoul, Republic of Korea.All chemicals used were of analytical grade.Hexane, ethyl acetate, chloroform, methanol, ethanol, water, sulphuric acid and vanillin were purchased from Daejung Chemicals and Metals Co. Ltd., Republic of Korea.Precoated TLC plates (layer thickness 0.25 mm), silica gel for column chromatography (70-230 mesh American Society Testing Materials), and LiChroprep RP-18 (40-63 m) were from Merck (Darmstadt, Germany).Authentic standards of chemicals were purchased from Sigma-Aldrich, USA.Previously isolated authentic standards of -sitosterol--D-glucoside were available.
2.1.Plant Material.Fresh ginseng (P.ginseng) was cultivated of ground dried roots ginseng (6 years old) in Ganghwado, Republic of Korea.A voucher specimen (No.PG-R-11) has been deposited at the Department of Applied Life Science, Konkuk University.Korean red ginseng was prepared by using nonpeeled fresh ginseng, which was steamed at 98 ∘ C for 2 h using an autoclave.The steamed ginseng after drying and powdered 297.8 g was prepared for extraction.

Extraction of Korean Red Ginseng
Powder.The Korean red ginseng powder (297.8 g) was immersed in methanol (3 × 1 litre) for three days at room temperature and then the supernatant was concentrated under vacuum to yield 30.1 g of the extract, which was suspended in water and extracted with hexane, ethyl acetate, and n-butanol successively to produce 5 g, 8.9 g, and 14.2 g extract, respectively.