Purification And Identification Of Sterols, Antioxidant Activity Of Extracts In Ganoderma Atrum And Its Fingerprinting

The genus Ganoderma-also known as "Lingzhi" in Chinese, "Reishi" in Japanese, and "Youngzhi" in Korean-has been traditionally used as an ancient medicine for centuries. So far, more than 120 species of Ganoderma have been reported in the world, a majority of which was found in China.In addition to G. lucidum(red), other members of the genus Ganoderma that are commonly used are G. applanatum(brown), G. tsugae(red-brown), G. sinense(dark purple to black) and G. atrum(black). Today, the medicinal use of Ganoderma is widespread and growing, G.lucidum and its related species have been used for the treatment of a wide range of ailments and chronic diseases, such as migraine, hypertension, arthritis, bronchitis, asthma, anorexia, gastritis, haemorrhoids, diabetes, hypercholesterolaemia, nephritis, dysmenorrhoea, constipation, lupus erythematosis, hepatitis, and cardiovascular problems. In contrast to Ganoderma lucidum, from which many bioactive compounds have been isolated, Ganoderma atrum is one of the species of Ganoderma that many phytochemical components were not still identified. In this study, the purification, identification and antioxidant activities of bioactive components in Ganoderma atrum were investigated. Besides, a GC-MS method was developed to determine the sterol comoposition of Ganoderma for the classification of G. atrum and G. lucidum collected from different regions of China. These research results obtained in this dissertation are summarized as follows:1. Silica gel column chromatography and semi-preparation-RP-C₁₈ liquid chromatography were used to isolate and purify the components from the fruiting bodies of Ganoderma atrum. MS, ¹H-NMR and ¹³C-NMR were used to identify these components. The results showed four sterols had been isolated and purified and their structures were identified as ergosta-4,6,8(14),22-tetraen-3-one, ergosterol, ergosta-7,22-dien-3β-ol and ergosta-7-en-3β-ol.2. High speed counter-current chromatography(HSCCC) technique in semi-preparative scale was used to separate and purify one phytosterol component from the crude extracts of Ganoderma atrum. An ergostane-type sterol component was separated by HSCCC and the separation conditions were optimized. The structure of this compound was elucidated as ergosta-4,6,8(14),22-tetraen-3-one by spectrum methods including UV, EI-MS, ¹H-NMR and ¹³C-NMR.3. Using the assay system of DPPH, the antioxidant activities of Ganoderma atrum extracted by different solvents were studied and compared with BHT. The results showed that the antioxidative effect of the extracts is dose-dependent, and the scavenging effects on DPPH radical decreased in the order of extracts obtained from ethyl acetate>n-butanol>petroleun ether.4. A GC-MS method based on the profiling of sterols has been applied for the discrimination of Ganoderma samples from different species and regions. The sterol profiles dominated by six ergostane-type sterols were obtained and used for pattern recognition analyses such as hierarchical clustering analysis(HCA) and discriminant analysis(DA). The results demonstrated that G. lucidum samples could be differentiated from G. atrum and G. lucidum samples could be successfully classified in accordance with their cultivated regions.