Effects Of Ionic Liquids On Ethanol Fermentation Of Pichia Stipitis

In this thesis,the inhibitory effect of residual ionic liquids（[C₈mim][BF₄] and [C₄mim][PF₆]）on subsequent ethanol fermentation was investigated.The effects of ionic liquid concentration on the growth and ethanol fermentation of Pichia stipitis were investigated.The effects of ionic liquid chemical structure on the cytotoxicity were also investigated.The mechanism of action of ionic liquid on yeast cell structure was investigated by using yeast cell wall as the starting point.Using a variety of methods and means to remove the ionic liquid in the water phase and the removal efficiency was compared.The main results were as follows:The solubility of ionic liquids [C₈mim][BF₄] and [C₄mim][PF₆] in aqueous solution was 16.61 and 18.70 g/L respectively,determined by using UV spectrophotometry.Different concentrations of ionic liquids（0.0025-0.0125 mol/L [C₈mim][BF₄]and 0.01-0.05 mol/L [C₄mim][PF₆]）were added in the xylose medium for ethanol fermentation of P.stipitis.The results showed that the higher the concentration of ionic liquid,the stronger the inhibitory effect on the growth and ethanol fermentation of P.stipitis and [C₈mim][BF₄] was stronger than [C₄mim][PF₆].Based on [C₈mim][BF₄] and [C₄mim][PF₆],the added ionic liquid composition structure was changed.The ionic liquids of different cationic alkyl side chains,the ionic liquids of different anionic species and several intermediates of ionic liquids were added to the culture medium respectively to investigate their effects on yeast cytotoxicity and ethanol fermentation.The results showed that the toxicity and inhibition of imidazole ionic liquid increased with the increase of the length of the cationic alkyl side chain,and the anion species had little effect on the toxicity of the ionic liquid.When the N-methylimidazole,BF₄-,PF₆-were added separately,they had little effect on the growth and fermentation of yeast.When the cationic structure of ionic liquid was [C₈mim],the growth and ethanol fermentation of yeast were significantly inhibited,ionic liquids might cause irreversible damage to yeast cells.The cytotoxicity of imidazolium ionic liquids was mainly determined by the length of the cation and its alkyl side chain.The changes of the morphology and structure of the yeast cells were observed by the scanning electron microscope.It was presumed that the ionic liquid could affect the normal growth and metabolism of yeast cells by dissolving cell wall.Four kinds of wall breaking methods were used,includingliquid nitrogen grinding and ultrasonic disruption,repeated freezing and thawing and ultrasonic disruption,salt breakage and glass bead vortex.Among them,the glass beads vortex method was the best wall breaking method with 81.0 % maximum breaking rate.The change of ionic liquid concentration and the change of cell wall dry weight before and after ionic liquid treatment showed that [C₈mim][BF₄] and [C₄mim][PF₆] could act on yeast cell wall in different ways.[C₈mim][BF₄] dissolved part of the cell wall while [C₄mim][PF₆] went into the cell interior by changing the permeability of the cell wall.The polysaccharides in the cell wall of P.stipitis were determined by ion chromatography,which was composed of three kinds of monosaccharides: galactose,glucose and mannose.According to the change trend of yeast cell wall monosaccharide content in the fermentationand combined with the above conclusions,it was speculated that [C₈mim][BF₄] was highly toxic and dissolves part of the cell wall polysaccharide,and [C₄mim][PF₆] was less cytotoxic,and yeast cells synthesize more cell wall polysaccharides under the stress.Respectively,using advanced oxidation method,organic solvent extraction method,activated carbon adsorption method to remove ionic liquid in water.The results showed that the removal rates of [C₈mim][BF₄] and [C₄mim][PF₆] were 33.91 % and 71.86 %,respectively,by heat activated ammonium persulfate.Dichloromethane was used as extractant to extract ionic liquid solution for 2 hours with the organic-aqueous phase ratio of 3:1,the removal rates of [C₈mim][BF₄] and [C₄mim][PF₆] were 88.07 % and 73.75 %,respectively.Powdered activated carbon was used to adsorb ionic liquids at 25 ℃ and 200 r/min for 24 hours,and the removal rates of [C₈mim][BF₄] and [C₄mim][PF₆] were 96.81 % and 96.76 %,respectively.