The Research Of Bio-ethanol Transformation By Using Marine Derived Sulfated Redseaweed Polysaccharide

At present, the feedstocks for bio-ethanol production studied are merely neutral polysaccharides chain, which has not only aroused bitter controversy over the food supply issue and the short supply with land, but also has resulted that the pretreatment of materials, the increase of saccharification yields and the utilization ratio of pentose are all in the technical bottleneck stage. Therefore, the derivation of bio-ethanol from other new resources has been the subject of interest worldwide.As a kind of characteristic polyanion polysaccharides, seaweed-based polysaccharide is a promising alternative source of bio-ethanol because of a variety of sources and low cost. However, the seaweed characteristic sugar chain converting into bio-ethanol has not been reported, the conversion of desulfurated K-carrageenan into fermentable reducing sugar will be another hot issue. The typical sulfated polysaccharide, K-carrageenan was chosen as the feedstock in the paper. And then the process of converting sulfated seaweed polysaccharides to bio-ethanol involved pretreatment to remove sulfate radical followed by saccharification and fermentation. The purpose of this study was to screen or compare the desulfuration way together with analyzing the structure of K-carrageenan and desulfurated K-carrageenan, and to optimize saccharification parameters, and then to justify the feasibility of sulfated seaweed polysaccharides applied to the production of bio-ethanol by analyzing compositions of saccharification liquids and optimizing fermentation condition. The results are determined as follows:Screening and comparing the desulfuration way:K-carrageenan was used to prepare desulfurated K-carrageenan through comparing improved methanolysis and organic solvolytic desulfation. The best desulfuration way was determined as improved methanolysis by testing DE vs TS as index. Based on the single factor and Orthogonal experiment, the parameters of methanolysis of K-carrageenan were0.5%of substrate concentration,1.0M HC1-methanol, temperature80"C, and20h of time and desulfuration rate reached93%, total sugar yielded90%, and the reducing sugar yielded18%, the desulfuration efficiency of desulfurated K-carrageenan was superior to that of organic solvolytic desulfation.Analyzing the structure of K-carrageenan and desulfurated K-carrageenan:The samples of K-carrageenan and desulfurated K-carrageenan were analyzed by IR to ascertain the structure of them. It displayed that there was no obvious change between the IR of K-carrageenan and desulfurated K-carrageenan, the relative content ratio of total sulfates was decreased from2.2to0.69, the relative content ratio of four-sulfate was decreased from1.34to0.09, the relative content ratio of3,6-Anhydro was decreased to zero. It proved that the desulfuration of κ-carrageenan had achieved a better effect and the3,6-Anhydro was damaged by improved methanolysis. Therefore, the desulfuration way not only realized the feasibility of sulfurated polysaccharide into netural sugar, but also laid a good foundation for further saccharification.Optimizing saccharification parameters of desulfurated κ-carrageenan: Desulfurated K-carrageenan was applied to saccharify by different enzymes including pectinase, cellobiase, hemicellulase, papain and carrageenase. The best saccharification enzyme was determined as pectinase and cellobiase by testing RS as index. The optimal pH of pectinase and cellobiase was4.0, and the time was72h. The desulfurated κ-carrageenan was saccharified by single enzyme and compound enzyme on the foundation of single factor experiment, and then the compound enzyme was determined as optimal saccharification enzyme. The best saccharification parameter of desulfurated κ-carrageenan was that the dosage radio of pectinase to cellobiase was1.1:0.1(v/w.v/w),96h reaction time and55.48℃temperature on the basis of response surface methodology experiment, and then the reducing sugar yielded81.29%. Simultaneously, desulfurated K-carrageenan and K-carrageenan were saccharified to obtain that the saccharification effect of desulfurated K-carrageenan was superior to that of κ-carrageenan. Hence, the saccharification of desulfurated K-carrageenan will lay a foundation for the further study on fermentation.Analyzing compositions of desulfurated K-carrageenan saccharification liquids: The saccharification liquids of desulfurated κ-carrageenan were isolated and purified by gel permeation chromatography (GPC) to separate different molecular weights. Then four components were obtained and the fourth component was the active peak. Simultaneously, the saccharification liquids of desulfurated K-carrageenan were detected by PMP-HPLC to obtain the composition. The result showed that the fourth component was composed of galactose with65.15%. The average molecular weight of active peak of the fourth component is determined to be180Da, which was proved by MS analysis. In brief, saccharification liquid of desulfurated K-carrageenan was majorly composed of fermentable galactose, which was released from saccharification of desulfurated K-carrageenan allowed for fermentation into ethanol.Optimizing fermentation condition of desulfurated K-carrageenan:The saccharification liquids of desulfurated K-carrageenan were inoculated with different strains. The best strain of fermentation was S. cerevisiae L-4by determining ethanol yields as index. Based on the single experiment, the best cultivation condition of fermentation was followed as that:2%galactose of seed liquors,2%substrate concentration, pH5.0,8%inoculation amount,42%temperature and72h reaction time. And then the parameters of the effect of inorganic salt on fermentation were determined as follows:3%。ammonium sulfate,5%。phosphate,4%o Mg2+. On the foundation of optimal parameters,1g κ-carrageenan was chosen as feedfock, the ethanol yielded0.7%(0.7g ethanol per100mL fermentation liquids) which was equal to0.273g ethanol/g K-carrageenan.The process of converting K-carrageenan into bio-ethanol is simple and feasible and the desulfuration products have a positive effect on the saccharification and fermentation. Compared to the traditional conversion way of bio-ethanol, the study shows the feasibility of converting seaweed polysaccharides with anionic type into bio-ethanol and it supplies a new kind of seaweed source for bio-ethnol.