Study on Ethanol Fermentation Using D-Glucose Rich Fractions Obtained from Lignocelluloses by a Two-step Extraction with Sulfuric Acid and Issatchenkia orientalis MF 121

Abstract

As about 80% of the world s energy comes from non renewable fossil fuel, 1) if we go on using energy at the current rate, the world will shortly encounter the severe problem of its depletion. Most of the recent information has reported that the global oil will begin its inevitable decline and will reach its extreme depletion in the near future. It is also considered that the problem of global warming is mainly due to the overuse of fossil fuel. 2) From these problems, the production of ethanol from renewable resources such as lignocelluloses (hardwood, softwood and annual plant residues) has drawn attention. However, these lignocelluloses have very hard structures, various proportions of cellulose, hemicellulose and lignin and characteristic structures of hemicellulose. 3) Therefore, most studies on ethanol production have conducted experiments using a single biomass like wood or agricultural plants. Our previous study showed a unique method individually preparing oligosaccharides of hemicellulose and cellulose from wood and annual plant residues by using twostep acid extractions and comparatively purified D glucose could be prepared from complete saccharification of the cellulosic oligosaccharaides. 4) Although yeast can grow utilizing many monosaccha-rides, only a few monosaccharides contribute to ethanol fermentation. DGlucose is one of the most appropriate substrates for both growth and fermentation. The yeast used in this study was Issatchenkia orientalis MF 121, capable of fermenting ethanol from Dglucose, Dmannose and Dfructose in acidic medium of pH 2.0 with 5.0% (ww) salt. 5) In this study, performance of the twostep extraction using 43.5% (vv) sulfuric acid and 55% (vv) sulfuric acid was investigated in order to confirm the extraction of oligosaccharides of hemicelluloses and celluloses from various lignocelluloses. The treatment conditions were scaled up to the amount capable of ethanol fermentation. The saccharified solution of the second extraction containing mainly Dglucose was applied to a column of Diaion DSR01 to separate the sugar and most of the sulfuric acid used. The Dglucose rich fractions were tested on the adaptability for ethanol fermentation with the yeast. MATERIALS AND METHODS Materials. Six typical lignocelluloses were tested as follows. Japanese oak (Quercus serrata) and eucalyptus (Eucalyptus globules) were chosen as hardwood, Japanese cedar (Cryptomeria japonica) and cypress (Chamaecy-paris obtuse) as softwood, and sugar cane bagasse (SCB) and rice straw as annual plant residues. In the case of SCB, a sucrose free sample (SCBs) was prepared according to the method described previously. 4) Their powdery samples with approximately 500 µm chip size were used and examined as dried materialsmoisture content was about 8.0% (ww). The resin Diaion DSR01, used for separation of acid and sugar, was obtained from Mitsu-bishi Chemical Corporation, Tokyo, Japan. Abstract: Cellulosic oligosaccharides (720920 mg) were prepared by a two-step extraction using 43.5% sulfu-ric acid and then 55% sulfuric acid from lignocelluloses (2 g each) of Japanese oak and eucalyptus as hard-wood, Japanese cedar and cypress as softwood, and sugar cane bagasse and rice straw as annual plant residues. After the complete saccharification of the oligosaccharide and treatment with the appropriate amount of activated carbon, D-glucose with a comparatively high purity was prepared. As the D-glucose was in a strong sulfuric acid solution, the sugar solution was applied on a column packed with an ion exchanging resin (Diaion DSR01) in order to remove almost all of the acid. The fractions eluting the sugar were collected and concentrated with a rotary evaporator. Although the concentrated sugar solution showed an acidity of pH 23, acid tolerant Issatchenkia orientalis MF 121 could ferment ethanol successfully in YPD medium using the sugar as the sole source of carbon.