Growth Characteristics, Demulsification Performance And Identification Of Extracted Products Of A Demulsifier Bacteria

A bio-demulsifier producing strain Alcaligenes sp. XJ-T-1 was isolated from oil-contaminated soil .The optimum conditions to culture XJ-T-1 in 5L fermentor were temperature 35℃, initial pH9.0, ventilation volume 3.0L/min and the rotational speed 400r/min.XJ-T-1 was a facultative alkaliphile which can grow at initial pH ranging from 6.0 to 11.0 and optimal pH range for its growth was between 9.0 and 11.0.XJ-T-1 produced extracellular bio-demulsifier only in alkaline environment while not produced in neutral and weak acidic conditions.By speculated of TEM images,surface morphology of XJ-T-1 changed by culturing in different initial pH,with the initial pH increased the fimbriae disappeared gradually.XJ-T-1 can tolerate 0~80g/L concentration of NaCl,the optimum growth concentration of NaCl was 20g/L,XJ-T-1 was a mild degree halophilic microorganisms.Biomass and demulsification rate of XJ-T-1 incresed slightly when XJ-T-1 cultured in fermentor in the optimal pH and optimal concentration of NaCl than cultured in flask.XJ-T-1 gradually decreased the pH of culture medium,the extracellular products may be acidic materials.A“dichloromethane elution +water extraction”method was established initially to extract crude cell associated products,but it can not extract all of the crude products that still need improve.Oil on the surface of XJ-T-1 can not break model emulsion,but other parts as residual bacteria, water extracted bacteria and crude cell associated products possessed effect of demulsifying.Effct of 500mg/L crude cell associated products on demulsification rate were lower than complete bacteria,residual bacteria and water extracted bacteria with same concentration.crude cell associated products may be mixed of lipopeptides and/or proteins and glycolipid,included four kinds of lipopeptides and/or proteins,one kind of glycolipid.A“improved resting cells”method was established to seperate crude extracellular products which did not contain any salt. crude extracellular products looked like yellow jelly and looked same as crude cell associated products.Improved resting cells could not make bacteria after stimulate dead or smash, effect of bacterias after stimulate to break emulsion were slightly lower than cells before stimulate,but still possess high demulsification rate. Crude extracellular products possess high demulsification raeffect more than crude cell associated products with the same concentration.Substance of Rf was 0.30 in thecrude extracellular products can not solve in methnol can be separated and purified,this substance possessed effect to break emulsion.crude extracellular products may be mixed of lipopeptides and/or proteins and glycolipid,included four kinds of lipopeptides and/or proteins,one kind of glycolipid.XJ-T-1 may produce 7 products ,3 of them may both crude cell associated products and crude extracellular products,and the other 4 products were extracellular products possessed 1 kind of peptide and/or protein and 1 kind of glycolipid,same as the cell associated products,but they were different products.On the base of same content of liquid paraffin as carbon source,compare with XJ-T-1 cultured by 4g/L NH4N03,five nitrogen source with same concentration such as triammonium citrate, leucine,valine,aspartic acid and glutamic acid can promote XJ-T-1 grow,but other two nitrogen souce alanine and isoleucine inhibited the cells growth. Biomass was little relevant to nitrogen content,same nitrogen content did not produce biomas similarly.Amino acids were used mainly as nitrogen source other than carbon source. Liquid paraffin was restrictive substrate of XJ-T-1.The optimum concentration of triammonium citrate cultured XJ-T-1 was 6g/L and optimum concentration of leucine was 2g/L,both of them can cuture biomass increase from around 4g/L to 6 g/L.