Research On The Fermentation Process Of Amauroderma Rude And Its Polysaccharide Extraction

Xuezhi, also known as Amauroderma rude, a new basidiomycete in the Amauroderma Murrill(Ganodermataceae), mainly distributed in Tropical and Subtropical regions. Xuezhi polysaccharides is one of the main active ingredients in Amauroderma rude, the studies have shown that it has unique activities such as inhibition of tumor growth, enhance immunomodulation. In this study, the Amauroderma rude strain purchased by Huazhong Agricultural University was identified by the mushroom experiment and molecular identification, and observed its biological characteristics; using biomass and polysaccharide as target, optimized the liquid submerged fermentation conditions of Xuezhi; optimized the extraction process of intracellular polysaccharides. The main research results as follows:1. Xuezhi liquid strains as seed cultivated the fruit body of xuezhi, found its hyphae grew well in cultivation bags, and appeared the primordium, picked up a small part of fruit body, using the optical microscope to observe its spore morphology, discovered its spores was spherical, double wall, thickness of spore wall was uniformity, all consistented with literatures.2. The choice experiment of ITS sequence primers of Amauroderma rude, by comparing the five ITS primers(ITS1- ITS4, ITS1-ITS5, ITS1-ITS4-B, ITS1-F-ITS4, ITS1-F-ITS4-B), the results showed the specificity combination of the ITS1-F-ITS4-B primers and xuezhi DNA was the best, the stripes clearly visible.3. Xuezhi strain was identified by analyzing its 18 S rDNA and ITS rDNA sequences. When compared with the data in the NCBI database, it was conformed that the preserved strain was Amauroderma Murrill. Studied the biological characteristics of xuezhi, improved its cognition.4. Using biomass and polysaccharide as target, optimized the liquid submerged fermentation conditions of Xuezhi.The single factor optimization experiment results showed : the optimum culture conditions of the liquid fermentation was 5% inoculation, the optimal pH、temperature and rotating speed respectively was 6、28 and 130 r/min; ℃the optimal fermentation medium components: the optimum carbon source and nitrogen source was respectively glucose and peptone, their suitable ratio was 20/3、1 g/L MgSO4·7H2O were best. And based on the single factor experiments of the fermentation medium components, using the orthogonal experiment designs find the best fermentation condition: 30 g/L glucose,3 g/L peptone and 0.5 g/L MgSO4·7H2O.5. Studied polysaccharide extraction process of xuezhi mycelium by the single factor experiment including the material liquid ratio, temperature, time, pH, ultrasonic power, ultrasonic time, microwave power and microwave time, and using RSM find the optimum polysaccharide extraction condition, the analysis results of RSM displayed that the optimum polysaccharide extraction process was the material liquid ratio 1:50, pH=2, 100 ℃, 5 h by hot water extraction process of xuezhi, under this condition, the extraction of polysaccharide was 51.02 ±0.08 mg/g. Before the optimal extraction process of xuezhi, using ultrasound and microwave to process the powder of xuezhi mycelium, it can effectively improve the polysaccharides extraction.