| Summary: | 博士 === 國立中央大學 === 化學工程與材料工程研究所 === 99 === In oriental culture, mushrooms, attracted attention because of their biochemical composition and medicinal characteristics, have been used as health foods and home remedies for thousands of years. The Basidiomycete fungus Agaricus blazei Murill, originally a native of Brazil, is one such species. The chemical components isolated from its fruiting bodies have been widely studied, especially its polysaccharides with antitumor activity. Besides, ergosterol and blazeispirol A are important bioactive components of A. blazei. However, yields and functionality of these bioactive metabolites produced by fermentation in submerged culture are highly dependent on the culture conditions, such as medium composition and environmental parameters including pH, temperature, agitation rate and aeration rate. Nevertheless, relatively few efforts have been focused on the influence of these operational conditions on the production of these metabolites. In previous studies, we reported the effects of medium composition, pH, temperature and agitation rate on the production of bioactive polysaccharides by A. blazei in batch cultures.
Therefore, the aim of this study was to propose a fermentation process of A. blazei with highly bioactive polysaccharides, ergosterol and blazeispirol A by investigating the effects of aeration rate and light intensity on their yields, the molecular weight distribution of polysaccharides, beta-(1,3)-glucan content in polysaccharides, beta-(1,3)-glucanase activity and TNF-alpha release capability on macrophage cells, to investigating the antitumor activity of ergosterol and blazeispirol A against the mouse hepatoma cell lines, Hepa-1c1c7, and human cervical epithelioid carcinoma cell line, HeLa, and to propose a novel approach to optimizing fermentation process of blazeispirol A using the conversion ratio of ergosterol into blazeispirol A.
First part of this research is to investigate the effects of aeration rate on the production of bioactive metabolites by A. blazei in batch cultures. The flask experiments were carried out by investigating three working volumes, 50, 100 and 150 mL. The results of flask experiments show that as the working volume increased, the value of surface of liquid per volume, beta-(1,3)-glucanase activity, and the yields of blazeispirol A (YP3/X, YP3/S) decreased, and the morphology of mycelia was changed from pellet to filamentous; however, the yields of ergosterol (YP2/X, YP2/S) enhanced. In addition, the optimal working volume for biomass, polysaccharides and beta-(1,3)-glucan formation was at 100 mL.
5 L stirred tank experiments were carried out by investigating five aeration rates, 0.1 to 0.5 vvm. The results of stirred tank experiments show that as the aeration rate increased, the oxygen transfer rate coefficient, kLa, enhanced, and the morphology of mycelia was changed from filamentous to pellet. Additionally, the optimal aeration rate for beta-(1,3)-glucan, high molecular weight fraction of polysaccharides and ergosterol formation was at 0.2 vvm, while that for biomass, polysaccharides and ?-(1,3)-glucanase activity formation was at 0.4 vvm, and the maximal bioactivity of polysaccharides was at the aeration rate of 0.2 vvm. However, the optimal aeration rate for blazeispirol A formation was at 0.3 vvm.
Second part of this research is to investigate the effects of light intensity on the production of bioactive metabolites by A. blazei in batch cultures. The flask experiments were carried out by investigating four light intensities, 0, 300, 500, 1000 lux. The results of flask experiments show that as the light intensity increased, the yields of biomass (YX/S) and polysaccharides (YP1/X, YP1/S), beta-(1,3)-glucan content, the average molecular weight of polysaccharides and beta-(1,3)-glucanase activity enhanced. Besides, the optimal light intensity for ergosterol and blazeispirol A formation was at 500 lux.
5 L stirred tank experiments were carried out by investigating four light intensities, 0, 500, 1000 and 1500 lux. The results of stirred tank experiments show that the optimal light intensity for ergosterol and blazeispirol A formation was at 500 lux, while that for biomass, polysaccharides, beta-(1,3)-glucan, high molecular weight fraction of polysaccharides and beta-(1,3)-glucanase activity formation was at 1000 lux, and the maximal bioactivity of polysaccharides was at the light intensity of 1000 lux.
Based on the result of the lower IC50 values of ergosterol and blazeispirol A against the proliferation of Hepa-1c1c7 and HeLa cells presented in this study, the contents of ergosterol and blazeispirol A in A. blazei play important roles in antitumor effects besides polysaccharides.
|
|---|
