Metabolic Profiles Of Natural Co-Cultures Of Anaerobic Fungi And Methanogens In Vitro And The Production Of Lactate

In the rumen, anaerobic fungi are important degraders of lignocellulose materials and can produce a wide range of fibrolytic enzymes including celluases, hemicellulase and esterases. Co-culture with rumen methanogens can further enhance the degradation of lignocellulose and produce large amount of methane. However, most of the co-culture studies were focused on the mixture of pure methanogen and fungal strains. In our laboratory, natural co-cultures of anaerobic fungi with methanogens were isolated and evaluated for their ability to degrade agricultural residues, produce methane, and produce enzymes. The research aimed to investigate the fermentation profiles of the natural co-cultures under different carbon sources and the potential in production of lactate. This thesis was described in the following four sections.The first study was to study the effects of various carbon sources on fermentation profiles of natural co-cultures of anaerobic fungi and methanogens.Co-cultures of anaerobic fungi and methanogens were isolated from goat rumen (F1) and camel faeces (N3), respectively.10mL co-cultures were inoculated into90mL Orpin’s medium C containing penicillin (1600IU/mL) and streptomycin (2000IU/mL) and grown with different carbon sources, cassava (M), rice straw (D), rice straw treated with NaOH (A) and corncob (Y)(0.8g) at39℃without shaking. The present study aimed to investigate the fermentation profiles of the natural co-cultures under different carbon sources. Among all the treated groups, co-culture N3grown on cassava, had the highest production of total gas (233mL), methane (48.54mL), and acetate (51.42mM) productions (P<0.05). Total gas, methane, and acetate productions of treated with NaOH rice straw groups were significantly higher than that of untreated rice straw groups (P<0.05). Generally, co-culture N3had higher fibrolytic activities than Fl.There were positive correlations between total gas production and the corresponding methane production (Rz>0.98) except co-culture F1grown on cassava, which had the relative low level of acetate (37.30mM) and methane (24.86ml). Co-culture F1grown on cassava produced higher level of lactate(27.58mM) and ethanol(14.91mM), but all other cultures produced low level of lactate(<2mM) and ethanol(<6mM). In conclusion, with cassava as substrate, co-culture of rumen anaerobic fungi and methanogens has the potential for industry use in production of some useful industry products.The second study aimed to provide some evidences for the hypothesis showed in the first study. This study was designed to determine the fermentation characteristics of different co-cultures(F1, F2and F) grown on cassava and the metabolism characteristics of co-culture F1grown on different carbon sources(cassava, soluble starch, maltodextrin, maltose and glucose). With cassava as substrate, the metabolism characteristics of co-cultures F1and F2had no significant differences (P>0.05).The productions of total gas, methane, and acetate of F grown on cassava were higher than that of F1and F2grown on cassava significantly (P<0.05), but the lactate production of F grown on cassava(2.33mM) were lower than that of F1(26.28mM) and F2(25.20mM) grown on cassava significantly (P<0.05). With different carbon souces as substrates, F1grown on maltose had the highest total gas, methane, and acetate production (P<0.05),and F1grown on cassava had the lowest total gas, methane, and acetate production(P<0.05).F1grown on cassava had the highest lactate production(30.51mM), and followed in order by maltodextrin(28.00mM), soluble starch (16.94mM), maltose (4.87mM) and glucose (2.32mM).The third study aimed to explore the effects of different raw starchy substrates(cassava, sweet potato, potato, corn, rice and flour) on the fermentation characteristics and lactate production of co-culture F1. The results showed that different raw starchy substrates had impact on the fermentation characteristics and lactate production intensively. Fl grown on rice had the highest total gas production(310.42mL/g DM), and followed in order by corn, sweet potato.F1grown on sweet potato had the highest methane production, and followed in order by potato, rice,corn, and flour. With cassava as substrate had the lowest total gas and methane production (P<0.05). F1grown on rice had the highest lactate production(0.41g/g DM)(P<0.05), but the lactate production of rice group and cassava group(0.36g/g DM) had no significant difference (P>0.05). Further analysis of the results showed that there were no correlations between total gas production and the corresponding methane production.The lactate production increased with the amylopectin proportion of substrates for all groups except sweet potato group.The last study aimed to study the effects of different cassava concentration(0.8g、 1.0g、1.2g、1.6g、2.0g and2.4g) on fermentation characteristics and lactate production, and the major metabolism end-products productions changed with fermentation time when F1grown on cassava (0.8g).The lactate production changed with the cassava concentration. Generally, there was higher lactate production in the higher cassava concentration medium. With2.0g cassava as substrates had the highest lactate production (56.29mM). When the substrate concentration increased from1.0g, the methane and acetate production decreased gradually. when the cassava concentration increased in the medium, the metabolism fluxes of co-culture changed(from hydrogenosomal carbon metabolism to a cytosolic one.). After48h fermentation, the lactate production of F1grown on0.8g cassava reached to the highest.