is a new species of the genus Antrodia (family polyporaceae, Aphyllophorales) that is parasitic on the inner cavity of the endemic species Cinnamomum kanehirai HAY. This endangered species has been used in Taiwan to treat food, alcohol, and drug intoxication, diarrhea, abdominal pain, hypertension, skin itching and liver cancer as a Chinese folk medicine.
1)The hot water extract of A. cinnamomea has hepatoprotective effect, 2,3) but the responsible component remains unclear. Here we describe the separation and purification of a neutral polysaccharide named ACN2a from a hot water extract of the mycelium of A. cinnamomea. We examined the hepatoprotective activity of the extract in mice with Propionibacterium acnes (P. acnes)-lipopolysaccharide (LPS) induced hepatotoxicity and elucidated the structure of this polysaccharide (ACN2a) by chemical and spectroscopic means.
ExperimentalMaterials Antrodia cinnamomea mycelia (Lot No. #C082002-1) were provided by Simpson Biotech Co. Ltd. (Taiwan). Pullulans (Shodex Standard P-82) were purchased from Showa Denko Co. Ltd. (Tokyo, Japan) and used as standard molecular weight markers.General Optical rotation was determined in H 2 O using a JASCO DIP-360 automatic polarimeter and UV absorption was measured using a Shimadzu UV-2200 UV-VIS recording spectrometer. Infrared (IR) spectra were recorded in a KBr disk or liquid film using a JASCO FT/IR-230 IR spectrometer and NMR spectra were recorded on a Varian Unity Plus 500 ( 1 H, 500 MHz; Analysis by GC-MS proceeded on a Shimadzu GC-17A gas chromatograph equipped with a JEOL mass spectrometer. Samples were separated by thin-layer chromatography (TLC) on pre-coated silica-gel 60 F 254 and cellulose F plates (Merck Darmstadt Germany, 0.25, 0.1 mm, respectively), and spots were detected by spraying with 10% H 2 SO 4 or aniline hydrogen phthalate (AHP) and then heating at 100°C. Carbohydrate concentrations were determined using a phenol-H 2 SO 4 method.
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Structural Analysis of a Neutral Polysaccharide from Antrodia cinnamomeaExtraction and Fractionation of Polysaccharides Freeze-dried mycelia of A. cinnamomea (1.5 kg) were extracted three times with 4 l of CHCl 3 at room temperature for 1 d, then combined extracts filtered and evaporated in vacuo to yield a CHCl 3 extract. The residue was dried and dipped into H 2 O (20 volumes) at room temperature for 1 h and extracted for 2 h 3 times at 100°C. The combined hot water extracts were concentrated to 800 and 3200 ml of EtOH was added. The mixture was stirred and left overnight in a refrigerator. The precipitates were filtered, washed with cold EtOH, dried and suspended in 10% trichloroacetic acid (TCA). The TCA-soluble fraction obtained by centrifugation (2500ϫg for 10 min) was dialyzed against distilled water for 3 d. The nondialyzable portion was lyophilized in vacuo to yield 14.25 g of a brownish residue (AC).Ion-Exchange Column Chromatography of AC AC (100 mg) dissolved in H 2 O was eluted through a column (Whatman International Ltd., Maidstone, England. 2.0ϫ20 cm) containing DE-52 ...