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Streptomyces sp. strain M3 from marine algae produced poly-guluronate specific lyase. The alginate lyase gene was obtained by using PCR and elucidated to consist of 780bp and 259aa. The region from 1Met to 35Ala was expected to be a signal peptide by using SignalP 3.0 server. The amino acid sequence of M3 lyase showed 94% , 81%, 28% identity to alginate lyase of Streptomyces sp. ALG-5, Corynebacterium sp. ALY-1 and Sphingomonas sp. A1, respectively. Purified ALG-M3 lyase was activated by 1 mM of CaCl2, MnCl2, CoCl2 and MgCl2, while 1 mM ZnCl2 and HgCl2 inhibited lyase activity completely. ALG-M3 lyase showed temperature stability at 50-60℃, whereas ALG-5 lyase showed temperature stability at 30-40℃. We also carried out homology modeling and analyzed the energy level of M3 alginate lyase using Prosa 2003 software for improvement of enzyme activity or stability by site directed mutation. Acknowledgement : This work was financially supported by the 2008 Busan Techno Park program and the Ministry of Knowledge Economy (MKE) and Korea Industrial Technology Foundation (KOTEF) through the Human Resource Training Project for Strategic Technology.

Due to the finite and limited supply of fossil fuel resources, production of biofuels, such as bioethanol and biodiesel from biomass, has recently attracted considerable attention. Biofuel is generally defined as energy obtained from biomass, which can occur through biochemical, thermochmical and various other methods. Moreover, the development of marine biotechnology from macro- and microalgae have emerged recently bioenergy sources. Marine algae have high carbohydrate content and are easily degradable, making them a potential substrate for biofuel and chemical production. In this study, micro- and macroalgae was subjected to the acid and enzymatic hydrolysis method, with the goal of assessing whether or not micro- and macroalgae can be used as a potential resource for the production of sugars (glucose and galactose). [Acknowledgements: This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (KRF-2008-313-D00303).]

The soluble epoxide hydrolase from Zebrafish, Danio rerio (sZFEH) gene was cloned in pET21b(+) vector and expressed into Escherichia coli strain BL21(DE52). We investigated recombinant enzyme stability and the effect of temperature and pH on the EH activity. We also analyzed the energy level of the soluble enzyme using Prosa 2003 software. Some amino acids from the enzyme sequence were replaced using error-prone PCR and site directed mutagenesis. The mutated genes were successfully expressed in E. coli and the gene product exhibited an improved enatioselective hydrolysis activity for racemic styrene oxide mixure. Acknowledgment: This work was supported by the Marine and Extreme Genome Research Center Program, Ministry of Land, Transportation and Maritime Affairs and industrial Technology Foundation (KOTEF) through the Human Resources Training Project for Strategic Technology,, Republic of Korea Korea.

Mussel adhesive proteins (MAPs) have a potential in diverse biotechnological and medical applications. However, natural extraction is inefficient for large-scale use and mass-production of recombinant MAPs has limited or failed according to foot protein (fp) types in several expression systems. Previously, to improve the production yield of recombinant MAPs, we designed new types of MAP hybrids (fp-151 and fp-353) and tried to express in Escherichia coli. Due to their successful expression with superior properties such as high production yield, simple purification, and strong adhesive ability, they have been investigated in many practical uses. In the present work, we designed and constructed various MAP hybrids that are composed of fp-1 (6- or 12-repeated decapeptides), fp- 3, and fp-5 to investigate combination effect of each fp types and size effect of each MAP hybrid on adhesive ability, water resistance, and other properties in practical applications. We tried to express new MAP hybrids and optimized expression levels under various culture conditions.

We studied the removal efficiency of five heavy metal ions (lead, cadmium, chromium, copper, and manganese) ions by three species of Korean seaweeds (Hizikia fusiformis, Laminaria and Undaria pinnatitida). Removal efficiencies of lead, cadmium, chromium and copper ions by these seaweeds were in the range of 80 to 96% as determined by using an Atomic Absorption Spectrophotometer. For pH variation from 2 to 8 at 21℃ and with agitation at 120 rpm for 120 min, the heavy metal adsorption was the lowest at pH 2 for all the five metal ions. This low level of adsorption at pH 2 was probably due to an unfavorable competition of heavy metal ions with hydrogen ions for their adsorption to seaweeds. At pHs between 3 to 8 the amount of adsorbed ions increased for lead, cadmium, and manganese ions. The amount of adsorbed lead ion increased rapidly for the first 10 min, and reached a near maximum value after 60 min. Our results indicate that pH is an important process variable when seaweeds are used as adsorbent.

The microbial production of 1,3-propanediol (1,3-PD) from rawglycerol, a byproduct of biodiesel (BD) process, has a great advantage from economical and environmental standpoints. In this study, the production of 1,3-PD was investigated with 5 types glycerol ; pure glycerol, raw glycerol and pretreated raw glycerol from waste vegetable oil-based BD process, and raw glycerol and pretreated raw glycerol from soybean oil-based BD process. Two strains of Klebsiella pneumoniae and 3 stains of Clostridium butyricum were utilized to investigate the effect of raw glycerol (with and without pretreatment) on 1,3-PD production. Klebsiella pneumoniae strains were generally resistant to all types of glycerol and there was no inhibitory effect of raw glycerol on 1,3-PD production. In contrast, the growth and 1,3-PD production of Clostridium butyricum strains was inhibited in the presence of raw glycerol. The production of 1,3-PD and the extent of growth inhibition were different between raw glycerol from waste vegetable oil-based BD and soybean oil-based BD process. When a simple acid treatment of raw glycerol was performed, the inhibition of Clostridium butyricum strains by raw glycerol was reduced up to the level obtained with pure glycerol.

The transestrification of fish oil was investigated with alcohol on the lipases. The immobilized Candida antarctica lipases and other lipases were induced in the reaction. The transesterification was carried out at various reaction conditions to determine the optimum reaction conditions. The optimum reaction conditions were 30% enzyme based on oil weight, 1:6 of oil/alcohol molar ratio, 40℃ reaction temperature, and 24 h of process time. The immobilized Candida antarctica lipases exhibited higher yield of fatty acid methyl esters (FAME) than other lipases. The FAME yield was 50% on the immobilized Candida antarctica lipases without additives. The additives such as hexane accelerated the yield of FAME and reduced process time.

Bio-ethanol is one of the alternatives for fossil fuel and its production is increasing in many countries. Stillage is the by-product from the distillation of ethanol following fermentation of carbohydrates. The amount of stillage can be up to 20 liters for each liter of ethanol produced and it contains high amount of organic matters, causing serious environmental problems. Therefore, a proper treatment of stillage is necessary. We are now examining whether stillage can be used as a feedstock for the production of bio-butanol, another alternative bio-fuel. As the first step, the effect of possible inhibitors contained in stillage, that is, lactic acid, acetic acid, and ethanol, on the butanol fermentation was investigated. Clostridium pasteurianum DSM 525 could produce 0.29~0.42 mole of butanol from 1 mole of glycerol in the presence of up to 9 g/L of lactic acid, 10 g/L of acetic acid, and 10 g/L of ethanol without serious inhibition of growth.

Biological denitrification among various denitrification methods is the most widely accepted process because of its economical and environmental advantages. Enzymes in microorganisms are known to be involved in biological denitrification process. However, the biological denitrification process has some problems. Most of all, the process needs other carbon sources for microbial activities. The carbon sources cause cost increase and 2nd pollution. For the reason, novel denitrification method required. In this research, Ochrobactrum anthropi SY509 was used as biocatalyst and the novel bioelectrode was made of silicone compound and graphite mixture. The graphite powder was used as conducting material. The graphite powder was mixed with biocatalyst for the electron transfer to the biocatalysts. And in the bioelectrode, the electrons were transferred directly to the immobilized biocatalysts from the electrode made of graphite powder. Using this electrode, a continuous nitrate removal system was made and high denitrification efficiency and durability was obtained. The details will be presented and discussed.

Bio-butanol has become another solution for the alternative to gasoline fuel. Its better characteristics of higher energy content, less hydroscopic and volatile which make it possible to use pre-existing transport infrastructure, than ethanol is regaining the reputation of conventional ABE (Acetone-Butanol-Ethanol) fermentation. However, the potent toxicity of butanol on the main ABE producers derived from Clostridium acetobutylicum ATCC 824, Clostridium beijerinckii NCIMB 8052, Clostridium saccharoperbutylacetonicum N1-4 etc., is still a major obstacle for economic production as a fuel. Extensive studies have been done only to reach the best titer around 20 g/L. For the production of n-butanol, we have constructed several recombinant C. beijerinckii strains by introducing E.coli-Clostiridum shuttle vecor. In this presentation, the detailed results will be presented.

Recently various kinds of macroalgae, such as Laminaia sp. and Gelidiaceae sp., are newly recognized as a biomass feedstock for the production of biofuels. We have studied on the potential of Laminaria japonica as a feedstock for the production of volatile fatty acids (VFAs) by anaerobic digestion. VFAs can be transformed into alcohols by chemical hydrogenation. We have tested the effects of various operating conditions including pH, retention time, and nutritional additives. The sludge taken from the acidogenesis reactor of an anaerobic digestion plant using livestock wastewater was used as the initial mixed inoculum. The optimization was carried out first by batch operations using flasks and then the result was confirmed in a semi-continuous bioreactor system. As a combined result of the optimization work and adaptation of the microbial flora to seaweed feedstock for 6 months, the VFA yields in the bioreactor was increased from 7.2-11.5% at the beginning to 20.2-21.6%. This result is equivalent to the mixed alcohol yield of 175-189 L/ton-VS (volatile solid), when 90% hydrogenation yield is assumed. The best yield achieved was 31.3%, which is equivalent to 273 L/ton-VS.

Microalgae are photosynthetic microorganisms that can harvest solar energy and convert CO2 and water to organic macromolecules such as carbohydrates, proteins, and lipids. Lipids (especially triacylglycerols) are the main storage compounds in many microalgae, and these could be effectively utilized as a source of biodiesel with dramatically low net carbon emissions than petroleum diesel. Cellular growth and lipid accumulation of microalgae are largely affected by various environmental and nutritional factors including seed condition, kinds and compositions of medium, CO2, pH, light intensity and temperature. Chlorella sp. KR-1 was isolated from thermal power plant and has been reported to have stable growth for high CO2 concentration up to 50%. In this study, before applying KR-1 strain for the production of biodiesel from the flue gas from 2 MW coal-power plant (KIER, Korea), the effects of several cultivation parameters such as inoculum preparation, kinds of medium (namely N-8, BG-11 and KR-1), and nitrate concentration were investigated for its cellular growth and lipid synthesis in flask cultures. The inoculum was cultivated under the same condition of main cultures and transferred at 10% (v/v) with and without centrifugation. Higher specific cell growth rates and cell-densities were observed with centrifugation (4000 rpm, 10 min) irrespective of media used. After 15 day incubation, cell-density with N8 medium was higher than those with KR-1 and BG-11 media when nitrate concentration was fixed at 5 mM. When nitrate concentration was varied in the range of 1 – 30 mM, optimal cell growth (1.0 g/L) was observed at 5 – 10 mM while highest intracellular fatty acids content (348 mg/g cell) was obtained at a low concentration of 1 mM. Maximal fatty acids production obtained under present study was 213 mg/L and major fatty acids were palmitic acid (27%, w/w), oleic acid (23%), and linoleic acid (27%). Other optimization experiments are under progress.

Butanol has several advantages to ethanol like higher energy density, low evaporation rate, and more hydrophobicity as a fuel source. These properties make butanol to use a real application without any modification of infrastructures that already installed (1, 2). Ceylon Moss is a red algae and fast growing biomass. It is able to be harvested by 5-6 times a year. Owing to low lignin content of macroalgae including Ceylon Moss it is easier saccharification step rather than that of lignocellulosic biomass used. Ceylon Moss Saccharified Liquid (CMSL) was used to produce butanol using solventogenic clostridia such as Clostridium acetobutyricum ATCC824, Cl. beijerlinkii NCIMB8052, Cl tetanomorphum ATCC49273 and Cl. aurantobutyricum NCIMB10659. CMSL has 18 g/L of galactose and 2 g/L of glucose as main carbohydrate. Clostridial species were cultivated on the modified Reinforced Clostridia Medium (RCM) with CMSL (50%) instead of glucose and starch. During ABE fermentation, n-butanol was produced 3-9 mM with small amount of ethanol (0.7-3 mM) and acetone (0.3-5 mM) when batch culture was conducted at 37℃ for 72 hrs. Our results indicated that solventogenic clostridia could utilize CMSL as carbon and energy sources, and produce butanol as main fermentation product.

Butyric acid has several potential applications in chemical industries and food technology. Currently, the most promising microorganism used for the bio-production of butyric acid is Clostridium tyrobutyricum. This microorganism is capable of producing butyric acid with high selectivity and can tolerate high concentrations of butyric acid. However, it is unable to use disaccharides such as sucrose as a carbon source. In this study, one strain of Clostridium diolis isolated from old fermented cow dung was screened and used in batch fermentation for the production of butyric acid using sucrose (50 g/l) as the sole carbon source. At pH 5.9, this strain was able to degrade all the sucrose in the medium to fructose and glucose and it produced 22 g/l of butyric acid. The yield also was promising. The drawbacks were relatively low tolerance to butyric acid as it was unable to consume the whole glucose and fructose in the medium and also slow production rate. At pH 6.8, the growth rate was faster, but the final conc. of butyric acid was only 13 g/l. In conclusion, this strain may be promising for the bio-production of butyric acid from sucrose.

Biobutanol has many characteristics that make it a better biofuel than bioethanol, now used in the formulation of gasohol. Despite the remarkable advantages of butanol as a fuel, biobutanol production has several limitations, including degeneration. Solventogenic clostridia lose their solvent formation capabilities during repeated sub-culturing of batch cultures or during a continuous fermentation. This phenomenon is known as 'degeneration'. The degeneration is caused by the loss of the pSOL1 plasmid that carries essential genes involved in solvent production. Therefore, a method is needed to detect and quantify the pSOL1 plasmid in Clostridium acetobutylicum ATCC 824. In this study, we designed two specific primer and probe sets for multiplex real-time qPCR detection of C.acetobutylicum ATCC 824 (the aceto set) and pSOL1 plasmid possessing C.acetobutylicum ATCC 824 (the DGS set). Specific primer and probe sets were designed on the basis of the 16S rDNA sequence and pSOL1 sequence. To confirm the effectiveness of the technique, the multiplex real-time qPCR results were compared with those obtained by plating. In conclusion, using multiplex real-time qPCR, degenerated Clostridium acetobutylicum was successfully detected and quantified.

Anaerobic digesters for the methane gas production from food wastes were comprised with methane fermentation, stabilized reactor. During one years, 25 Samples were collected from the reactors and the microbial communities were analyzed by T-RFLP using two different primer sets (63F-FAM/1389R and 27F-FAM/1492R) and three restriction enzymes (AluI, RsaI, and HaeI). Among these combinations, the combination of 63F-FAM/1389R primer set and restriction enzyme AluI showed the highest discrimination level of the microbial community and T-RFLP analyses with this combination could be used as a method for addressing microbial diversity of the anaerobic digesters.

n-butanol is a useful organic solvent used for various purposes in industry and thus could be released into the environment. The degradation of n-butanol by Gordonia sp. YK1 was investigated in this study. Gordonia sp. YK1 was cultivated in the M9 medium with adding different concentrations of n-butanol (2%, 4%, 6%, v/v) as the sole carbon source. Gordonia sp. YK1 showed different degradation and growth patterns that were strongly dependent on the initial concentration of the n-butanol. At a concentration of 0.6% (v/v), the degradation rate was fastest and butryic acid was produced by 0.25 g/L as a intermediate.

A glucose analog, 2-deoxy-D-glucose (2-DG), has been used to obtain derepressed mutants for various carbohydrate metabolisms. Pichia stipitis mutants for improved xylose metabolism were isolated by using ethyl methane sulfonate mutagenesis in the presence of xylose and 2-DG. The growths of mutants were faster than that of wild type strain. A mutant, KM-D1, completely consumed xylose in the presence of glucose and produced maximum ethanol (32 g/L) with productivity (0.44 g/L/h) after 72 h from 80 g/L glucose and xylose mixture.

이산화탄소는 지구온난화의 주요원인으로 알려져 있다. 여러 가지 이산화탄소 처리 방법 중, 액생 흡수제를 이용한 화학흡수법은 배가스 중의 이산화탄소를 처리하는데 가장 효과적인 공정으로 알려져 있다. 화학흡수법을 이용한 이산화탄소 처리 공정에서 monoethanolamin (MEA), diethanol amin (DEA), N-methyliethanolamine (MDEA), 2-amino-2-methyl-1-propanol (AMP) 등의 알카놀아민 (alkanolamone) 흡수제가 일반적으로 사용된다. 최근 암모니아 수용액이 상용 아민계 흡수제를 대체할 흡수제로 제안되고 있다. 암모니아 흡수제는 아민 흡수제에 비해 경제성을 가지고 , 이산화탄소 흡수능이 높으며, 탄화수소암모늄 (ammonium bicarbonate)의 분해 온도가 낮아 재생에 유리하며, 부식성이 적다는 장점을 가진 것으로 보고되고있다. 배가스 CO2처리에 암모니아수 사용이 제안되면서 흡수장치 설계에 필수적인 자료인 암모니아수의 CO2 흡수속도에 대한 자료가 필요하게 되었다. 아민계 흡수제에 대하여 방대한 양의 반응속도 자료가 보고된 것에 비하면 암모니아수 흡수제의 경우, 보고된 반응 속도 자료가 많지 않다. Andrew[7]는 세라믹 디스크가 설치된 흡수탑(disk absorption water)에서 CO2 분압 및 암모니아수 농도별 흡수실험을 수행하여 반응속도를 아레니우식(arrhenius equation) 형태로 보고한 바 있다. 여러 가지 흡수속도 측정 장치 중 교반셀 반응기 (stirred cell reactor)를 사용한 방법은 회분식으로 조업하기가 쉽고 기상 압력 변화로부터 비교적 간단한 방법으로 반응속도에 대한 자료를 얻을수 있기 때문에 기체흡수속도를 측정하는데 많이 상용되는 방법이다[8]. 본 연구에서는 교반셀반응기를 이용하여 6 wt%, 12 wt%, 18 wt% MEA첨가한 수용액에서 CO2흡수속도를 측정하였다. 일반적으로 흡수제 성능을 개선하기 위하여 비수용액 용매에 혼합한다. You 등[9]은 암모니아 흡수제가 가진휘발성을 감소시켜 암모니아가 기상으로 손실되는 단점을 개선하기 위하여 아민기와 수산화기를 가진첨가제를 사용하였다. 본 연구에서는 MEA (monoethanolamine)의 아민류 첨가제를 포함한 비수용액에서 CO2흡수 속도를 측정하여 아민류첨가제를 포함한 수용액에서와 비교하였다.

에너지의 대부분을 수입에 의존하는 우리나라에서는 국가안보와 지속적인경제성장을 유지하기 위해 에너지 수입 의존도를 최대한 감소시킬 수 있는 장기적인 노력과 대체에너지 개발이 필수적이다.더불어 최근 환경파괴와 지구온난화에 따른 기후변화가 심각하여 온실가스 배출에 대한 관심이증대하고 있다. 화석에너지는 발전과정에서 발생하는 온실가스가 문제가 되어 기존의 에너지원을 대체할 수 있는 다양한 에너지사업이 제기되고 있다. 이산화탄소나 메탄 등의 유해 온실가스를 감소시키기 위한 하나의 방안으로 제기된 신재생에너지의 필요성이 대두되고 있다. 특히 지속적인 유가의 상승으로 기존의 화석에너지 관련 법규의 규제로 신재생에너지로 전환하여 보급하는 것을 목표로 하여 투자와 지원을 강화하고 있는 추세이다. 이러한 온실가스 배출에 관한 문제를 해결하기 위해서 신재생에너지는 화석에너지와 비교시 경제성의 확보가 최우선 과제이며 각각의 신재생에너지에 관한 경제성 평가가 정확하게 이루어져야 한다. 경제성 평가를 바탕으로 기술성, 성장 가능성, 국내 환경조건에 맞는 적합성을 포함한 신재생에너지의 타당성이 검증되어야 한다. 때문에 기존의 석유 및 석탄에너지에서보다 의 발생량을 줄이고 경제성분석을 통해 다른 에너지원보다 저렴하게 이용할 수 있는 방안을 모색해보았다.

5-Hydroxymethylfurfural and levulinic acid are versatile biomass-derived platform compounds that can be used to synthesize a broad range of chemicals, and are currently derived from petroleum. This study focused on optimization of reaction conditions for formation of levulinic acid from marine algal biomass Gelidium amansii using acid catalyst and by using statistical approach. By this approach, optimal conditions for production of levulinic acid was found as follows: 160.0℃, 43.1 min and 3.0%. Under optimized conditions for levulinic acid formation, glucose and galactose concentrations were minimized. In addition, levulinic acid formed at a high reaction temperature, long reaction time, and high catalyst concentration. Based on the overall results, chemical intermediates (HMF and levulinic acid) produced from marine biomass will be used as platform compounds for use in synthesis of a broad range of chemicals currently obtained from petroleum. Our results may provide useful information in regard to development of more economical and efficient systems for production of sugars and chemicals from marine-biomass. [Acknowledgements: This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (KRF-2008-313-D00303).]

A large quantity of volatile organic compounds (VOCs) is released from various industrial plants and processes. These VOCs will become atmospheric pollutants and are facing increasingly stringent environmental regulations. Most VOC emitted to the atmosphere are likely to be harmful to human health as generating nausea, headaches and irritation, affecting the nervous and breathing systems, initiating cancers, etc. In this study, the removal of gaseous styrenee as experiment of TiO2/biofilter hybrid system was invesgated the industry. Removal performance of the photocatalyst biofilter hybrid system for the styrene was evaluated to optimize the operation conditions. Firstly to investigate the photocatalytic degradation of styrene, various nanocrystalline TiO2 thin films have been prepared by a sol-gel method. The optical fibers are employed as the light-diffusing guide with the immobilizing TiO2. A 500 W high pressure mercury UV lamp was used as the light source. The effects of the inlet concentration of styrene, flow rate, relative humidity and the thickness of TiO2 thin films on the degradation of the styrene were examined. In addition, the effects of some metal ions doped onto the TiO2 thin films were considered in order to get higher photocatalytic degradation. The hybrid system was operated at an EBRT (Empty Bed ResidenceTime) of 20~30 s under nitrogen deficient condition where nutrient solution was supplied only 1 L every 2 week. When ERBT was 30 s and average inlet concentration is 3.56~109.03 ppmv , average RE(Removal Efficiency) is 89.2 %. High elimination capacity of styrene could be achieved with the following operating conditions, 25~30℃, relative humidity 85~95 %, EBRT 20~30 s, mass loading rate 13.15~107.70 g/m3․h. During more than 3 months operation, maximal, hybrid system showed that the removal efficiencies were more than 80~99 % at inlet styrene concentration of 23.56~193 ppmv which corresponded to elimination capacity of 13.15~107.70 g/m3․h. Compared with other experiment, this study had equal removal efficiency in a hybrid system or reference UV reactor, and had more than 10~20 % efficiencies in a biofilter.

Microalga Chlorella vulgaris C-169 was studied to optimize its growth and lipid synthesis in flask cultures. Important parameters investigated include kinds of medium (N8 and Bristol), nitrogen source (nitrate 1-20 mM, ammonium 1 – 30 mM), phosphate buffer (1-15 mM), Fe (0-0.3 mM), and pH (5.5-8.5). Cell density, pH and intracellular concentrations of protein, carbohydrate, chlorophyll, and fatty acids were monitored during cultivation. Overall nitrate was a better source than ammonium for cell growth and lipid synthesis. When phosphate buffer concentration increased, the final cell density increased although intracellular fatty acids content did not change. Optimal Fe concentration was 0.01 mM. Under tested pH range, optimal pH for cell growth was 6.5, but highest fatty acids accumulation was observed at 7.5. The maximal cell density and intracellular fatty acids content obtained under partially optimized conditions were 2.0 g/L and 320 mg/g cell, respectively. Major fatty acids were palmitic acid (15% of total fatty acids), stearic acid (8%), oleic acid (50%), linoleic acid (14%), and α- linolenic acid (13%). Other optimization experiments are under progress.

During past decades, considerable efforts have been made to utilize lignocellulose as biomass feedstock for the optimal production of bioethanol as an alternative source of fuel. Cellulase enzyme can be used for converting pre-treated lignocellulose material to glucose, which can be fermented to ethanol. The major concern in an enzymatic process is the instability of the enzyme under repetitive or prolonged use and inhibition by high substrate and product concentration. Immobilization is a very effective alternative in overcoming problems of instability and repetitive use of enzymes. Entrapment method of immobilization is advantageous over other methods as they do not involve chemical modification of the enzyme. In this work, the bioconversion of pretreated straw to fermentable sugar was investigated by using immobilized Cellulase enzyme in calcium alginate beads. The product released which is reducing sugar is determined by DNS method and then the enzyme activity for immobilized and free form were calculated.

In general, Industrial yeast strains used for baking or brewing lack selectable genetic marker and identification of hybrids by genetic complementation is impossible. Selection markers can be introduced by mutation into industrial strains. In the present study, respiratory deficient mutant and auxotrophic mutant for selection maker were induced by acriflavine treatment and mutagenesis with NTG, EMS, repectively. As a result, respiratory deficient petite mutants of industrial ethanol fermentative yeast, Saccharomyces cerevisiae CHY1011, CHFY0321 and Auxotroph of thermostable yeast, Kluyveromyces marxianus KCTC17631 have been isolated. The yeast strains can be used for protoplast fusion between S. cerevisiae CHY1011RD, CHFY0321RD and K. marxianus CHY1703 using tetrazolium overlay technique and selection medium.

Optimization of fermentation condition for production of high concentration (>13%) of ethanol from corn and triticale were conducted. Higher concentration of ethanol in the fermented mash results in lower distillation cost of ethanol. From 25 yeast strains, the best strain was selected for corn and triticale, respectively. The optimum conditions for high concentration of ethanol from corn were 33℃of fermentation temperature, 6 days of fermentation time, 1 × 108 cells/㎖ of initial cell number, 1.2 % of solid glucoamylase, 24 hr of seeding culture age, and 0.2 mM copper as medium additive. The optimum conditions for high concentration of ethanol from triticale were 36℃of fermentation temperature, 4 days of fermentation time, 1 × 107 cells/㎖ of initial cell number, 1.2 % of solid glucoamylase, 24 hr of seeding culture age, 12 mM urea and 0.2 mM copper as medium additive. Using the combination of the optimal conditions, almost of the fermentation for the corn and triticale were compleed in 3 days.

Rice straw is a lignocellulosic biomass that is a renewable organic substance and alternative source of energy. For the first time, rice straw was pretreated in a novel manner using hypochlorite-hydrogen peroxide (Ox-B) solution. The optimum pretreatment condition was analyzed by response surface methodology and the pretreated rice straw was hydrolyzed using two kinds of enzymes: Cellulase and Spezyme. Following hydrolysis, Saccharomyces cerevisiae and Pichia stipitis were inoculated for ethanol production. The optimum condition was 60 min pretreatment using 5% Ox-B solution (consisting of 10 ml hypochlorite and 50 ml hydrogen peroxide). The Ox-B solution treatment was an essential step for efficient hemicelluloase hydrolysis. Under these conditions, 406.8 mg glucose and 224.0 mg xylose were obtained from 1g rice straw. The structural change of rice straw after pretreatment and enzyme hydrolysis was examined by scannin gel ectron microscopy. With the 5% initial sugar concentration, the final ethanol concentration was about 1.67%, which is 87.3% of stoichiometric and fermentation efficiency yield.

Recently, we have studied on the liquefaction of waste newspaper using ethylene glycol (1). From the results, cellulose, lignin and hemicellulose were removed by approximately 10%, 84% and 70% based on initial content at 10 minutes, respectively. This means that most of lignin and hemicellulose can be removed while keeping cellulose intact, and indicates that ethylene glycol can be used as a pretreatment reagent under proper reaction conditions. Pretreatment by ethylene glycol does not need high pressure equipment because of its high boiling point (197.6℃), whereas conventional pretreatment by aqueous liquor does. The major parameters to affect on pretreatment performance are temperature, acid concentration, and reaction time. To compare the pretreatment performance of a reaction with other’s, a unified parameter that combines temperature, acid concentration, and reaction time, called as severity parameter, can be used. Since the pH of nonaqueous solvent like ethylene glycol cannot be measured, we modified Chum et al.’s equation (2) by replacing hydrogen ion concentration with sulfuric acid concentration (% weight). The result indicated that severity factor could be well correlated with the removal of cellulose, hemicellulose and lignin, and enzymatic digestibility, respectively.

Nowadays, microalgal fuel technology is in the limelight again as a new biomass feedstock to produce biofuel because novel biological technologies are being developed and climate and energy crisis is more and more serious. Biological CO2 fixation process using microalgae can be one of technological solutions because it is possible to recover CO2 directly from exhaust gas of industrial facilities without additional CO2 separation process. In addition, microalgal biomass is available for producing biofuel such as biodiesel and bioalcohol. In this study, Isochrysis galbana and Chlorella minutissima, which were proved to have relatively higher CO2 fixation capacity and lipid contents than other microalgal strains in last study, were co-cultured under various conditions for enhancing both CO2 fixation capacity and lipid contents. In addition, Isochrysis galbana was cultured in pilotscale photobioreactor, which was based on various hydrodynamic tests performed for designing high efficient photobioreactor, for mass production of microalgae.

Because of high contents of cellulose and hemicellulose, barley straw seems to be a potential lignocellulosic biomass for production of bioethanol. However, lignocellulose such as barley straw is difficult to hydrolyze using only cellulolytic enzymes due to its recalcitrant and heterogeneous structure. Barley straw is considered to account for the largest portion of available biomass and most of barley straw is wasted following harvesting in Korea. Ethanosolv applied organosolv pretreatment is more convenient method than other pretreatment. To provide milder ethanosolv condition and enhance the enzymatic saccharification, metal salts as a catalyst were employed in ethanosolv. Barley straw was soaked in aqueous ethanol solution (50 wt% ethanol, 1 wt% H2SO4) with 0.2~25mmol/L metal salt (FeSO4, Fe2(SO4)3, FeCl3, AlCl3, Al2(SO4)3, MgSO4). After soaking, the impregnated barley straw is heated at 170oC for 1hr.