Study On Enhancement Of Lipid Accumulation During Microalgae Cultivated In Sludge Liquor With Crude Glycerol Addition

Microalgae is considered as an ideal feedstock for biodiesel production due to fast growth rate,strong environmental adaptability and high intracellular lipid content.The microalgae is culturedin sludge liquor coupled with the addition of crude glycerol-a by-product of biodiesel,which could save water resources and inorganic nutrients.The cultivation mode integrating sewage treatment,harvesting of algal products and recycling of crude glycerol,is economic,efficient and sustainable.In order to increase the lipid production of Chlorella vulgaris cultured in sludge liquor,two-stage culture method was adopted.The biomass productivity of Chlorella vulgaris was increased by increasing inoculation density,adding crude glycerol(prepared in laboratory)and adding naphthoacetic acid(NAA).The lipid accumulation was improved by salt stress and iron stress.Finally,semi-continuous culture of Chlorella vulgaris was conducted in MPBR to optimize the culture conditions.The main research results are as follows:(1)When the inoculation density of Chlorella vulgaris was 1.1 g/L,the highest biomass pruductivity was 0.600 g/L/d,the highest lipid content was 25.7%and the highest lipid production was 0.59 g/L.The utilization rates of COD and NH₄⁺-N were 91.4%and 99.5%.(2)1.0?2.0 g/L crude glycerol was added to sludge liquor could promote the absorption of COD and NH₄⁺-N by Chlorella vulgaris.When the concentration of crude glycerol was 2.0 g/L,Chlorella vulgaris achieved the highest biomass of 2.81 g/L and the highest biomass pruductivity of 0.905 g/L/d.When the concentration of crude glycerol was 1.0 g/L,the highest lipid content was 22.3%and the highest lipid production was 0.62 g/L.The addition concentration of crude glycerol was determined to be 1.0?2.0 g/L.(3)When the concentration of NAA was 0.25 mg/L,Chlorella vulgaris achieved the highest biomass of 2.284 g/L and the highest biomass pruductivity of 0.642 g/L/d.The utilization rates of COD and NH₄⁺-N were 90.8%and 99.3%.The addition concentration of NAA was determined to be 0.25 mg/L.(4)The optimal NaCl concentration and stress time were 20 g/L and 48 h.Chlorella vulgaris achieved the highest lipid content of 45.4%,the highest lipid production of 1.29 g/L and the highest lipid productivity of 17.433 mg/L/h.The utilization rates of COD and NH₄⁺-N were 85.3%and 99.6%.During the stress culture,starch content decreased from 18.1%to3.67%,and lipid content increased from 21.6%to 45.4%,which indicated that salt stress could effectively block starch synthesis and force carbon transfer to lipid synthesis.The content of unsaturated fatty acid reached 78.93%and C18:1 fatty acid accounted for 34.92%.The length of carbon chain met the requirement of biodiesel production,which could be used as raw material for biodiesel production.(5)The optimal Fe³⁺concentration and stress time were 3 mg/L and 48 h.Chlorella vulgaris achieved the highest lipid content of 30.2%,the highest lipid production of 0.79 g/L and the highest lipid productivity of 8.533 mg/L/h.The utilization rates of COD and NH₄⁺-N were 92.4%and 98.2%.During the stress culture,starch content decreased from 24.6%to13.8%,and lipid content increased from 19.2%to 30.2%.The content of unsaturated fatty acid was 64.15%,and C18:1 and C16:1 fatty acid accounted for 35.17%.The length of carbon chain met the requirement of biodiesel production,which could be used as raw material for biodiesel production.(6)After two-stage culture of Chlorella vulgaris,the biomass of 20 g/L NaCl,3 mg/L Fe³⁺,20 g/L NaCl+3 mg/L Fe³⁺groups were 2.80 g/L,2.66 g/L and 2.75 g/L,all were lower than that of the control group(3.05 g/L).When the concentration of NaCl was 20 g/L,Chlorella vulgaris achieved the highest lipid content of 38.9%and the highest lipid production of 1.09 g/L.Stress culture led to increased cell size.The number and size of lipid droplets significantly increased.The number of starch granules decreased.The lipid droplets were fused.The results indicated that stress culture could enhance the lipid accumulation.The main component of lipid produced by Chlorella vulgaris was C16?C22 fatty acid.The highest sum of C18:1 and C16:1 fatty acid of 46.4%was achieved when Chlorella vulgaris was stressed under 20 g/L NaCl condition.The lipid accumulation could not be further enhanced by the double stress of salt and iron.However,both Na⁺and Fe³⁺could affect the fatty acid composition of Chlorella vulgaris.(7)In MPBR,Chlorella vulgaris was cultured semi-continuously with sludge liquor.The optimal HRT was 2 d.Chlorella vulgaris cultured for 48 h could achieve the highest lipid production of 0.966 g/L and the highest lipid pruductivity of 0.207 g/L/d.The removal rates of COD,NH₄⁺-N,TN and TP were 81.4?90.8%,95.7?96.0%,90.2?90.8%,and 73.7?82.2%.(8)After two-stage culture of Chlorella vulgaris in MPBR,the biomass productivity and the level of lipid accumulation were increased,and most nutrients were removed.In the stage of biomass accumulation,the highest biomass and biomass producticity were 3.84 g/L and1.165 g/L/d.In the stage of lipid accumulation,the highest lipid content,lipid production and lipid pruductivity were 40.2%,1.181 g/L and 11.737 mg/L/h,respectively.