| Poria(Poria cocos (Schw.) Wolf), the Polyporaceae fungal is a traditional Chinese medicine (TCM) that is always applied both medicinal and edible. It was documented in the "Chinese Pharmacopoeia" over the years. In the "Chinese Pharmacopoeia", Poria was described to have efficacies such as excreting dampness and diuretic, comforting stomach and tonifying spleen, calming the mind and soothing the nerves, etc., it was also one of the commonly used TCM for the prevention and treatment of cancer. Modern pharmacological studies have shown that Poria has good biological activity of anti-tumor and regulating immune functions. Its main active components are polysaccharides and triterpenoids. Most of the literatures that imported its antitumor activity were about pachymarans, and there are relatively very limited research reports mentioned the antitumor activity of its triterpenoids so far.My thesis presented here is on the basis of the preliminary studies earried out in our laboratory. For the purpose of in-depth interpretation of the antitumor efficacy material basis of Poria, and to provide scientific evidences of improving the quality of the cultivated Poria, on the one hand, triterpenoids extract of Poria (the content of total triterpenes is above90%) was used for the systematical studies of the anti-tumor activity of Poria triterpenoids, on the other, the medicinal qualities of different strains of cultivated Poria were evaluated.1. Poria triterpenoids antitumor activity studySubjects with a variety of tumor cell lines, the anti-tumor activities of Poria triterpenoids were evaluated by using MTT assay. The results showed that Poria triterpenoids could induce noticeable proliferation inhibition effects in HepG2, A549, MDA-MB-231, Caski and SH-SY5Y tumor cell lines. The research results demonstrated that Poria triterpenoids have the anti proliferation activity for several tumor cell lines, in addtion, for HepG2tumor cell line, the anti proliferation activity tend to be very consistant.2. Poria triterpenoids antitumor mechanisms studyApoptosis morphology observation, apoptotic cells’DNA fragmentation analysis based on agarose gel electrophoresis and flow cytometry detection of cells apoptosis ratio were performed for understanding the antitumor mechanisms of Poria triterpenoids against human hepatoma cell line HepG2. Observations of the Poria triterpenoids treated HepG2cells under inverted microscope and optical fluorescence microscop showed that the shapes of Poria triterpenoids treated HepG2cells changed significantly, the number of cells decreased, there were visible compact dense stain, and some whitish colored nucleus. Agarose gel electrophoresis analysis of the apoptotic cells’ DNA fragmentation results demonstrated that after24,48h treatment with different concentrations of Poria triterpenoids HepG2cells, the typical DNA ladder of late stage apoptotic (180-200bp integer multiple of the oligonucleotides) could be identified. Flow cytometry analysis results illustrated that after40μg/ml,60μg/ml,80米g/ml and100μg/ml Poria triterpenoids36hours treatment, the HepG2cells apoptosis ratios were7.2%12.8%,39.3%and56%respectively, which proved that the Poria triterpenoids have apoptotic induction effects on HepG2cells. All evidences indicated that Poria triterpenoids could induce apoptosis of hepatoma cells efficiently.3. Poria triterpenoids isolation, identification and pharmacological evaluationModern chromatographic and spectroscopic techniques were applied to isolate and identify the chemical components in Poria triterpenoids. Eight terpenoids were isolated and identified, including poria acid, dehydroeburic acid, dehydropachymic acid,3β-hydroxy-lanostane-7,9(11),24-triene-21-acid, 15a-hydroxy-dehydrotumulosic acid,3β-hydroxy-16a-acetyloxy-lanosta-7,9(11),24-trien-21-oic acid, Trametenolic acid and polyporenic acid C. Their antitumor activities were evaluated by using MTT assay, and all the8compounds have shown moderate antitumor activities. The research result further illuminated the antitumor material basis of Poria triterpenoids.4. Quality analysis of different strains of cultivated PoriaThe TCM Poria is a fungus in the Polyporaceae family. As a result, in their cultivation process, the selection of strains is a key factor affecting the medicinal materials quality. In this study,27strains of Poria were analysed by using thin layer chromatography, content analysis and HPLC fingerprint comparison technologies. Comprehensive analysis indicated that the content of active components Poria acid, total triterpenes and polysaccharides in cultivated Porias by using strains from Macheng (Hubei), GZ (Guizhou) and ACCC50478were higher than all other samples. The qualities of the cultivated Porias from these strains were considered to be better than all the others. This study provides a reference for the TCM Poria standardized planting within the GAP project. |
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