Study On The Efficiency And Mechanism Of Flocculants Produced By The Fermentation Of Molasses And High Concentration Brewery Effluent To Harvest Oleaginous Yeasts

The rapid development of renewable energy can relieve the pressure of“more energy and lower carbon emissions”to some extent.Biodiesel is a typical“green renewable energy”.Microbial oil is a potential raw material for biodiesel production,and oleaginous yeasts are the best producers of microbial oil due to their high oil production rate,fast growth rate and easy cultivation.Soybean oil refining effluent?SORE?can be used to ferment Trichosporon fermentans to obtain microbial oil.However,microbial oil belongs to the intracellular product of yeasts,and only by harvesting yeasts efficiently can microbial oil be obtained.The flocculants produced by fungus M2-1 are non-toxic,do not cause secondary pollution to the environment,and can be used to harvest Trichosporon fermentans accumulating microbial oil in SORE,efficiently and greenly.However,the flocculants were obtained by fermenting M2-1 in flocculants producing medium?FPM?,which consumed a large amount of glucose and other nutrients and resulted in a high raw material cost of flocculants.In this study,M2-1 was fermented by the cheap raw material molasses to produce flocculants and the raw material cost of the flocculants was decreased.And then,the bioflocculants were used to harvest oleaginous yeasts fermented by SORE and molasses,respectively.M2-1 was also fermented by high concentration brewery effluent?HCBE?to produce flocculant and the raw material cost of the flocculants was further decreased.The produced flocculants were used to harvest oleaginous yeasts fermented by SORE and HCBE,respectively.The flocculation mechanism of oleaginous yeasts harvested by flocculants was expounded,and the effects of flocculants on the components of oleaginous yeasts and microbial oils were evaluated.This study laid a foundation for producing flocculants at low cost and harvesting oleaginous yeasts efficiently in engineering applications.The main research contents and achievements were as follows:?1?The production of flocculants by fermenting M2-1 in molasses medium for flocculants production?MMFP?and the harvest of Trichosporon fermentans were studied.Under the condition of optimal flocculants production with molasses,the yield of flocculants produced by M2-1 in MMFP?F-MMFP?reached 4.8 g/L,which was significantly higher than that of 3.9 g/L when M2-1 was fermented in FPM.The raw material cost of flocculants was decreased by 57%.The conditions of F-MMFP and the corresponding fermentation liquor producing flocculants?FLPF?to harvest Trichosporon fermentans in SORE were optimized,and the flocculation rate for the yeasts reached 97%and 99%,respectively.When Trichosporon fermentans was harvested by the FLPF,although calcium phosphate contributed to the flocculation rate and reduced the amount of flocculants,it prolongs the time needed to achieve the desired flocculation effect.In addition,the FLPF produced by M2-1 in MMFP was also used to harvest the Trichosporon fermentans which accumulated 1.21 g/L microbial oil in MMTF.Under the optimal conditions,the flocculation rate for the yeasts reached98%.?2?The production of flocculants by fermenting M2-1 in HCBE and the harvest of Trichosporon fermentans were studied.HCBE was diluted with domestic wastewater at a ratio of 1:1,and then an oleaginous fungus,Cunninghamella echinulata was fermented in the diluted HCBE?DHCBE?to consume about 90%of the ethanol and acetic acid.The resultant wastewater was used to ferment M2-1,and the gained FLPF showed 93.2%of flocculation rate for kaolin suspension.The raw material cost of flocculants was further reduced.The yield of flocculants produced by Cunninghamella echinulata and M2-1 in DHCBE?F-DHCBE?reached 4.2 g/L,and the above fermented Cunninghamella echinulata contributed to 0.75 g/L microbial oil.Under the condition of optimal flocculation,the flocculation rate of F-DHCBE and and the corresponding FLPF to harvest Trichosporon fermentans in SORE reached 93.9%and 94.4%,respectively.In addition,the FLPF produced by Cunninghamella echinulata and M2-1in DHCBE was also used to flocculate and harvest the Trichosporon fermentans which accumulated 1.52 g/L microbial oil in HCBE.Under the optimal conditions,the flocculation rate for the yeasts reached 95.8%.?3?The mechanism of flocculants flocculating oleaginous yeast was analyzed.F-MMFP and F-DHCBE mainly contain sugars and their derivatives,a small amount of proteins,nucleic acids,and a large amount of ash.The monosaccharides in F-MMFP mainly include galacturonic acid and xylose,while the monosaccharides in F-DHCBE mainly include mannose and glucose,etc.The differences in the components of the two flocculants made them have different flocculation time for oleaginous yeasts.In addition to the common constituent elements of organic molecules such as carbon,hydrogen,oxygen,nitrogen and sulfur,the two flocculants also contain phosphorus with mass concentration of more than 20%and potassium and sodium with total mass concentration of more than 20%,which makes the ash content in the flocculants high.The main functional groups of the two flocculants include-OH,-COOH and-PO₄^3-groups,among which the-PO₄^3-group is connected with biological macromolecules such as sugars and proteins in the form of secondary bonds such as ionic bonds,strongly adsorbs Ca²⁺,and enhances the aggregation capacity and flocculation activity of the flocculants.The mechanism of F-MMFP and F-DHCBE to flocculate Trichosporon fermentans is mainly based on the bivalent positive dissociation theory.In the range of pH4-10,the flocculants are negatively charged,but the main functional group in the flocculants is a good adsorption site of Ca²⁺,which can form a bridge through Ca²⁺to absorb negatively charged Trichosporon fermentans and result in flocculation.At the same time,the three-dimensional network structure of the flocculants can produce the net catching and sweeping effect and promote flocculating the Trichosporon fermentans.The results of adsorption kinetics showed that the adsorption rate of F-DHCBE was faster,the adsorption process was more in line with the second order kinetic reaction,and chemisorption was the main speed limiting step.The adsorption thermodynamic results showed that the adsorption of the flocculants on oleaginous yeasts conforms to the Langmuir,Freundlich and Dubinin-Radushkevich adsorption isothermal equations,and chemisorption and physical adsorption play roles in the adsorption process,together.?4?The effects of the flocculants on components of oleaginous yeasts and microbial oil were evaluated.The results of thermogravimetric analysis showed that compared with MMTF and HCBE,the Trichosporon fermentans cultured from SORE had high lipid content and less carbohydrate and protein.The F-MMFP and F-DHCBE had no effect on the components of the Trichosporon fermentans,but the more ash in the flocculants made the carbon surplus increase slightly after thermogravimetry.The flocculants also had little effect on the oil yield and oil content.The oil yield and oil content of the Trichosporon fermentans cultivated in SORE increased slightly after flocculation,because the flocculants flocculated a small amount of waste oil in the wastewater while harvesting the oleaginous yeasts.The flocculants did not affect the quality of microbial oils.The F-DHCBE was used to harvest Trichosporon fermentans cultivated in SORE,and in the fatty acid of the gained microbial oils,the content of unsaturated fatty acids,linoleic acid and oleic acid accounted for 73.1%of the total oil fatty acid.The fatty acid composition was similar to that of vegetable oils,and the microbial oils can be used as a potential raw material of biodiesel.The carbon chain of alkane substances in the pyrolysis oil obtained from the Trichosporon fermentans harvested by the F-DHCBE became longer,and the types and proportion of them increased,while the proportion of oxygen-containing compounds decreased.Meanwhile,the solid pyrolysis products also increased slightly,which improved the quality of the pyrolysis oil and the added value of the solid pyrolysis products.The technology of producing flocculants from molasses and HCBE was established in this study,which provides technical support for the low cost recovery of Trichosporon fermentans in engineering applications,and is conducive to promoting the resource utilization of waste and wastewater in food industry.