Earticle

The principal histopathological feature of Alzheimer’s disease is the presence of β-amyloid (Aβ) aggregates in the brain, and researchers believe that various environmental factors play significant roles in the conformational change of Aβ peptides. Therefore, discovering a rapid and convenient analytical method of evaluating the environmental factors on Aβ aggregation would have a considerable impact. Herein we report our development of a novel microfluidic screening system enabling high-throughput analysis, low-consumption of reagents, and short analytical time. Microchannels were immobilized with Aβ monomers via N-hydroxysuccinimide ester activation, and then a fresh Aβ monomer solution mixed with different small molecules was continuously introduced into the microchannels. In this work, we investigated (1) the temporal evolution of Aβ aggregation within microchannels, (2) the high-throughput screening of the inhibitory effect of twelve small molecules against Aβ aggregation, and (3) the effect of different metal ions (Fe3+, Cu2+, Zn2+, and Al3+) on Aβ aggregation by using ThT-induced fluorescence microscopy and ex situ atomic force microscopy. The microfluidic system should contribute to a simultaneous analysis of multiple environmental factors in a parallel manner and to screen therapeutic small molecules prior to their in vivo evaluation.

paclitaxel extracted from plant cell cultures. However, the fractional precipitation process has been inherently problematic due to the lengthy precipitation time (~3 days) that is required. An improved fractional precipitation process could significantly reduce the precipitation time by increasing the purity of crude extract and the surface area available for precipitation. Glass beads (7 mm) were used to increase the surface area, and the optimal surface area per working volume (ie. volume of reaction solution) (S/V) for achieving the highest purity and yield of paclitaxel possible was found to be 0.428 mm-1. The content of paclitaxel dissolved in methanol that can be processed during fractional precipitation was evaluated, and it was established that up to 0.9% (w/v) pure paclitaxel content could be processed. This improved pre-purification process serves to minimize solvent usage and the size and complexity of the high performance liquid chromatography operation required for paclitaxel purification.

In this study a new type of biosorbent (PAAE) able to bind anionic metals were developed by surface modification of waste biomass of E. coli with pollyallylamine hydrochloride. The PAAE was used for removal of Pd (II) as chloro-complex from the aqueous solution was investigated. The parameters studied for performance evaluation of PAAE include the solution pH, contact time and initial metal concentration. Result of pH edge experiments showed that as increasing pH, the uptake of Pd (II) was increased at the below pH 5. The isotherm data were fitted with Langmuir isotherm model, showing that maximum uptake was determined to be 265.3 mg/g at pH 3 and 212.9 mg/g at pH 2, respectively. In order to clarify the biosorption kinetics, pseudo-first-order and pseudo-second-order model were applied to fit the kinetic experimental data. Pd (II) biosorption was rapidly happened in first 10 min, and almost achieved equilibrium within 60 min. For the desorption study, 5M HCl, 5M HNO3 and 0.1M thiourea in 1M HCl were selected as eluents. As a result, 0.1M thiourea in 1M HCl was powerful, showing the desorption efficiency of nearly 100%. Therefore, it may be concluded that PAAE can be useful cost-effective biosorbents for the removal and recovery of anionic precious metals as chlorocomplex form solutions containing hydrochloric acid produced in metal refining process.

Flavonoids are plant secondary metabolites with pharmacological and nutraceutical properties. To expand the applicability of Streptomyces venezuelae as a heterologous host to plant polyketide production, flavonoid biosynthetic genes were expressed in an engineered strain of S. venezuelae DHS2001 bearing a deletion of native pikromycin polyketide synthase gene. A plasmid expressing the 4-coumarate/cinnamate:coenzyme A ligase from Streptomyces coelicolor (ScCCL) and the chalcone synthase from Arabidopsis thaliana (atCHS) under the control of a single ermE*promoter was constructed and introduced into S. venezuelae DHS2001. The resulting strain produced racemic naringenin and pinocembrin from 4-coumaric acid and cinnamic acid. Placement of an additional ermE* promoter upstream of the codon-optimized atCHS (atCHSop) gene increased the yield of naringenin and pinocembrin. Expression of codonoptimized chalcone isomerase gene from Medicago sativa, together with ScCCL and atCHSop genes led to production of (2S)-flavanones, but the yield was reduced. This is the first report on the heterologous expression of plant phenylpropanoid biosynthetic pathways in Streptomyces genus.

In this research, we successfully evaluated the dual biokinetic parameters between Nitrosomonas nitrosa and Nitrosomonas aestuarii using the new mathematical algorithm into which Monod model was modified. The process using industrial wastewater including 300 mg/L ammonium nitrogen was operated in the batch mode. From its molecular analysis, it was identified that there were Nitrosomonas europaea, N. nitrosa and N. aestuarii in the system through 16S rRNA DGGE/PCR. Furthermore, as a result of quantitative analysis with real-time quantitative PCR, it was investigated that the number of N. europaea could be nearly negligible since its population size was one hundred times smaller than the total one of other species in overall process of ammonia oxidation. Therefore, the dual kinetic algorithm, which has already been mentioned above, was applied to only two species, N. nitrosa and N. aestuarii. The three ordinary differential equations were numerically solved by 1st and 5th order Runge-Kutta Fehlberg methods. The solution calculated by the methods was compared with the experimental data in order to calculate the total sum of squares of residuals. Only in the final place, the error was numerically minimized by Levenberg-Marquardt method which is widely used as optimization tools. The values of μmax, Ks, Y and kd of N. nitrosa were 1.63 d-1, 7.3 mg NH4 +-N/L, 6.2 × 108 copy numbers/mg NH4 +-N and 0.2 d-1 , respectively. Also, those of N. aestuarii were 1.06 d-1, 19.5 mg NH4 +-N/L, 7.8 × 104 copy numbers/mg NH4 +-N and 0.036 d-1, respectively.

Cell organelles such lysosomes and peroxisomes, isolated from egg whites, are safe bioresources that probably be possible replaced for chemical based reagents to treat polluted compounds in the environments. In this study, phenol was efficiently degraded by various enzymes extracted from cell organelles, which isolated from egg whites. In optimal conditions for phenol (100 mg/L) degradation by cell organelle’s enzymes in dissolving buffer were found to be pH8.0 at 37℃, stirring at 200rpm. Phenol was reduced up to 20% for the 18 hours with various enzymes, which should be identification further. These results highlight the potential of cell organelle’s enzymes for removing aromatic compounds contaminated in the environments.

The bacterial ghost system showed a potential as a vaccine candidate against various fish pathogens. It provides versatile carrier functions for foreign antigens with natural intrinsic immune properties. The optimization of medium for E. coli K-12/pHCE-InaN-GAPDHGhost27 were determined as 10 g/L glucose, 5 g/L yeast extract, 50 mg/L thiamine with Riesenberg's semi-defined medium. The optimal batch fermentation parameters were determined as 10% inoculum size, 300 rpm agitation and 1.5 vvm aeration for the high cell density production. The optimal fed-batch process for high cell density fermentation of E. coli K-12/pHCE-InaN-GAPDH-Ghost27 was developed. The nutrient feeding strategy with Riesenberg's defined medium during fed-batch process was evaluated. The fermentation was conducted in four phases as follow: initial batch phase, fed-batch phase, temperature induction phase and heat shock phase. The maximum bacterial ghost vaccine was obtained about 40 g dcw/L. The protective efficacy of GAPDH ghost cell vaccination was determined by challenging immunized olive flounder with live S. iniae. Therefore, the ghost E. coli/ GAPDH antigen from S. iniae could be used as an effective vaccine in aquaculture.

Fractional precipitation is a simple, efficient method for pre-purifying paclitaxel from the plant cell cultures of Taxus chinensis. Crude paclitaxel was dissolved in methanol and the solution was diluted with distilled water to 61.5% (v/v) methanol. The water feeding and mixing method of fractional precipitation were evaluated in terms of the yield, purity and precipitation time of paclitaxel with a fixed methanol concentration in water (61.5%, v/v), paclitaxel content in the crude extract (0.5%, w/v) , and storage time ( 7 hr and 14 hr ). The maximum purity (~57.4%) and yield (~81.8%) of paclitaxel were obtained at pH 8 with ammonium phosphate buffer. The yield and purity of paclitaxel were increased by repeated intermittent addition of ammonium phosphate buffer during fractional precipitation. This evaluation of fractional precipitation process serves to minimize solvent usage and the size and complexity of the high performance liquid chromatography operation required for paclitaxel purification.

In this presentation, we suggest two methods for the recovery of gold from gold-cyanide solutions. The biomass of Corynebacterium glutamicum generated from fermentation industry was used as a biosorbent. The capacity of the biosorbent for the binding of Au(I) ions from for cyanide solutions was examined through batch experiments, such as pH edge, isotherms and kinetics. For the pH edge experiment, the solution pH strongly influenced the sorption capacity. With the increase in the pH value, the uptake of Au(I) decreased, except at pH under 3. The Langmuir and Freundlich models were used to fit the experimental data. Between them, the Langmuir model described the experimental data very well with high correlation coefficient (R2=0.9946). According to Langmuir isotherm the maximum uptake of Au(I) at pH 2.5 were estimated to be 35.149 mg/g. The equilibrium of sorption process could be attained after 15 min. Gold ion (Au+) was able to be successfully eluted from the Auloaded biosorbent with D.I. water at pH 7, where the desorption efficiency was 91%. However, aqua regia was not a good eluent (41% of desorption efficient). From the results, it can be noted that the combined process of biosorption and desorption would be effective for the recovery of gold as the form of gold ion (Au+). We tried a different approach to recover zero-valent gold by incinerating the Auloaded biosorbents. Incineration resulted in the removal organic impurities and at the same time, reduction of the gold ions during oxidation of organics. As a result, the zero-valent gold content in ash was as high as 60 – 70%. It was also found that the purity of gold in ash could be enhanced by increasing the the uptake of gold by the biosorbent. Therefore, we may suggest that the combined method of biosorption and incineration can be useful for the recovery of zero-valent gold from goldcyanide solutions

White Spot Syndrome Virus (WSSV), an enveloped and rod-shaped virus, is one of the most virulent pathogen, causing high mortality of cultured shrimp1) and large economic loss in shrimp culturing industry. The virus particle was consists of at least five major proteins VP19, VP26, VP28, VP281 and VP466.2) This study was carried out Reverse Transcription-PCR (RT-PCR) for the identification of various WSSV infection stages. The RT-PCR is widely used for the quantification of steady-state mRNA levels and is a critical tool for molecular medicine and biotechnology. A diluted WSSV stock (1:200) was used for WSSV challenge test. At 11 days of post challenge, the shrimp in the positive control with WSSV showed 100% mortality. In this study, total RNA was obtained from frozen shrimp organs at various infection stages using RNA isolation kit (Takara, Japan). Isolated RNA was reverse-transcribed into complementary DNA (cDNA) using the reverse transcription reaction kit (Takara, Japan). The resulting cDNA was amplified using β-actin, VP19, and VP26 sets. From these results, the difference expression patterns were identified various WSSV infection stages.

Microalgae are widely used as live feeds, cosmetics, human food supplements, biomedicine and etc. Especially, the use of microalgae for a potential renewable liquid fuel source was proposed by many reports. In this study, characteristics of five microalgae: Tetraselmis suecica, Phaeodactylum tricornutum, Chaeotoceros calcitrans, Isochrysis galbana and Nannochloropsis oculata were examined on the ability to store intracellular neutral lipid and growth potential. The microalgae were grown in sterilized seawater with f/2 medium nutrients and photoautotrophically in batch cultures where inorganic CO2 from the atmosphere was the only source of carbon. Aeration was provided by air stone at regular pressure. The temperature was kept at 20℃. The strains were inoculated into the medium (18 L) in a 20-L column tank. Range of inoculation density was adjusted between 20~40ⅹ104cells/ml. A fluorescent spectrometric method was applied for lipid contents analysis. In the results, Specific growth rate of N. oculata was higher (0.325/day) and the lipid content of C. calcitrans estimated by fluorescence intensity (au /106 cells) was higher than other strains.

Glycolysis is the most important metabolic pathway for growth and maintenance of living organisms, and it is designed highly robust by nature. However, relevant modification of glycolytic activity is significant in metabolic engineering for the product accumulation and it is considered as one of main technical challenges for the strain improvement. gapA is a one of the key glycolytic genes and encodes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) which mediates oxidative phosphorylation of glyceraldehyde-3-phosphate into 1,3-diphosphoglycerate and has vital role in glycolysis. Wild-type GAPDH has strong activity and it is not easy to control its expression level by the conventional controllable promoters. Therefore, it is required to modify GAPDH for the sensitive regulation of glycolysis in E. coli. In this study, we developed a gapA mutant for the regulationsensitive glycolysis in E. coli by directed evolution approach using the gapAdeleted mutant obtained by Red recombination system and temperature sensitive promoter system. Directed evolution of GAPDH was conducted by error-prone PCR combined with growth rate based screening system. By several round of evolution processes, several mutants with the different levels of regulation sensitivities could be obtained. From the sequence analysis of the mutants, activity and sequence relationship was also examined.

Rhodobacter sphaeroides is non-sulfur purple photosynthetic bacteria exhibiting versatile growth capabilities in aerobic or anaerobic culture condition. In this study, seven organic acids were used in order to analyze the effect of cell growth according to various carbon sources. R. sphaeroides were cultured on sistrom’s minimal medium in the presence of acetate, citrate, lactate and 4 carbon-organic acids such as fumarate, malate, succinate and oxaloacetate during 60 hr, respectively. The result showed that growth rate under the culture condition with 4 carbon-organic acids was more increased than those with acetate, citrate and lactate. Moreover the growth rate of R. sphaeroides was drastically enhanced when used succinate as a carbon source. It means that R. sphaeroides is effectively able to use 4 carbon-organic acids as carbon source.

Opium poppy (Papaver somniferum) plant is known as a commercial source of morphinan alkaloids such as thebaine, codeine and morphinee. But callus induced from this plant are regarded to lose their ability to produce morphine and related alkaloids, which were derived from (R)-reticuline and produced in high concentration by the original intact plants, due to lack of enzymes transforming (R)-reticuline and codeine to next metabolites. Therefore, to elucidate the differences between secondary metabolism of plant cells and intact plants of Papaver somniferum and to determine the potential step for targeted metabolic engineering, we compared the expression profiles of gene transcripts and enzymes on the alkaloid biosynthetic pathway in P. somniferm using plant cells and intact plants parts such as leaves, stems, roots and capsules. Gene transcript and protein expression level of salutaridine oxidoreductase in cells was negligible compared to that in intact plants. Through integrated-omics study on secondary metabolic pathway of P. somniferum salutaridine oxidoreductase (SR) could be suggested as a potential target of metabolic engineering for cells whose secondary metabolism would be altered by genetic manipulation to produce morphinan alkaloids in vitro.

The third generation energy from seaweed as biomass can solve environmental and energy issues. In this study, the effective pretreatment and saccharification methods for common seaweeds such as Undaria pinnatifida, Laminaria japonica, Hizikia fusiforme, Sargassum fulvellum using physical, chemical and biological methods were evaluated for bioethanol production. The seaweeds were obtained from Gijang fisheries market in Busan and dried in sunlight or hot-air. The dried seaweeds after milling were pretreated and saccharified by the acid hydrolysis. The optimal acid hydrolysis method for the highest yields was determined as follows: 10% seaweed powder, 0.2% hydrochloric acid (HCl), heat treatment at 121℃ for 15min, agitation at 200 rpm for 30 min and neutralization by sodium hydroxide (NaOH). The physical properties of the saccharified products such as salinity, reducing sugar content and viscosity were evaluated. From these results, the optimal physical properties of seaweed saccharification were determined for bioethanol production.

The micelle process was developed for pre-purifying paclitaxel from plant cell cultures of Taxus chinensis, giving a high purity and yield. The approach in this work was to transfer paclitaxel in the crude extract to an aqueous surfactant solution as a micelle, allowing organic solvents to be used for removal of lipids and non-polar impurities. In this work, the effects of various surfactants such as CPC, CTMAC, LTMAC, SDS, AOT, Tween, PEG, and Triton were examined on the yield, purity, and phase separation time in micelle process. Among these surfactants, CTMAC (5%, w/v) gave the best result in terms of paclitaxel yield (~99%), purity (~21%), and phase separation time (30 min). The use of micelles in the pre-purification process allows for rapid and efficient separation of paclitaxel from interfering compounds and dramatically increases the yield and purity of crude paclitaxel for subsequent purification steps.

Carboxypeptidase Y (CPY) is a useful enzyme involved in the amidation of therapeutic peptides and C-terminal sequencing of polypeptides. Endogenous CPY of Saccharomyces cerevisiae is initially expressed as a form of proCPY that contains the proregion and then is processed by vacuolar protease A for its conversion to the active form. CPY fused with the mating factor α (MFα) signal sequence was overexpressed under the control of the GAL1 promoter in recombinant S. cerevisiae. Batch fermentation of S. cerevisiae CENPK2-1D/p426αCPY resulted in 7.6 g/L dry cell mass and 14 mg/L CPY concentration. The mature form of CPY (61 kDa) and proCPY (67 kDa) were detected in the media. Pdi1p, an essential yeast protein catalyzing the oxidation, reduction, and isomerization of disulfide bond, was coexpressed at the same manner of CPY. Coexpression of Pdi1p enhanced the secretion of CPY. Finally, S. cerevisiae CENPK2-1D/p426αCPY+p424GalLPDI1 coexpressing CPY and Pdi1p produced 34 mg/L CPY, corresponding to a 2-fold increase compared with the control strain.

Nature utilizes solar energy for the synthesis of various organic chemicals, which remains as a target model for the development of artificial photosynthesis systems. Natural photosynthesis through the Z scheme of two photosystems, testifies an elegant example of how a fine nexus between photocatalysis and biocatalysis could be designed in a heterogeneous phase. Devising an artificial analogue in the heterogeneous phase has been a long-standing challenge in chemistry. The present work attempts the construction of such a photobioreactor device using a semiconductor photocatalyst through the integral coupling of photocatalysis and biocatalysis cycles. It unites these two catalytic functionalities in a novel photobiocatalytic system for the ultimate goal of utilizing solar energy in the visible region for photobiosynthesis of fine chemicals. Herein we report the new visible light photocatalyst W2Fe4Ta2O17 and its redox enzyme coupled application to a photobioreactor. A new potentially promising visiblelight driven photobioreactor synthesizes fine chemical by generating NADH in a non-enzymatic light-driven process and coupling it to the enzymatic dark reaction catalysis.

wblA (SCO3579) is a pleiotropic down-regulator of antibiotic biosynthesis in Streptomyces species, as well as modulates morphological differentiation (Kang et al. 2007). Although wblA is notable pleiotropic down-regulator in valuable secondary metabolite production, these regulatory networks are still largely unknown. In this study, total cosmid DNA library screening in doxorubicin (DXR)-producing S. peucetius resulted in the isolation of S. peucetius wblA gene (named wblA-spe). To examine biological significance of the wblA-spe on DXR production, the wblA-spe was disrupted from the chromosome of DXRoverproducing S. peucetius strain by PCR-targeting system, followed by quantitative DXR HPLC analysis. After the wblA-spe gene disruption, biosynthesis of DXR and DXR intermediate aklavinone were slightly enhanced. Furthermore, using Streptomyces interspecies DNA microarray analysis between the DXR high-producer S. peucetius strain and wblA-spe disruption mutant strain, several putative wblA-dependent genes were identified. To verify the biological significance of this comparative transcriptomics-guided targets, the five putative wblA-dependent target genes were individually cloned under the strong-and constitutive promoter(PermE*) of the Streptomyces integrative expression vector, followed by conjugation into S. coelicolor. Among the putative wblA-dependent genes tested, the overexpression of NADH dehydrogenase subunit, (SCO4571) conserved hypothetical protein (SCO4967), and putative oxidoreductase beta-subunit (SCO6569) in S. coelicolor enhanced biosynthesis of actinorhodin about 1.5-2-fold relative to that of S.coelicolor exconjugant harboring the empty vector alone. More detailed results will be discussed.

PTA (purified terephthalic acid) wastewater contains various aromatic compounds such as phthalate isomers, p-toluate, benzoate, trimellitic acid, etc. Most of these aromatic compounds can be finally converted into methane biogas under anaerobic condition. In this study, we investigated the anaerobic degradation mechanisms of the aromatic compounds in the PTA wastewater, especially focusing on p-toluate and trimellitic acid of which detailed mechanisms have not yet fully elucidated. We also conducted microbial community analysis using the 16S rDNA gene-based molecular techniques such as denaturing gradient gel electrophoresis and gene cloning in order to identify the microorganisms responsible for their degradation. In the investigation of the anaerobic degradation characteristics of the PTA wastewater, it was revealed that p-toluate and trimellitic acid could be degraded only by a syntrophic culture system. In the community analysis, on the other hand, it was identified that Desulfotomaculum and Pelotomaculum related bacterial populations were responsible for the initial degradation of p-toluate, trimellitic acid and phthalate isomers while Methanothirx, Methanosaeta, and Methanospirillium related methanogens were involved in the final methane production. In addition, the population abundance and distribution of the identified bacteria and archaea within the sludge could be further confirmed using scanning electron microscopy.

Over the past 40 years, there was significant increase in research towards the field of molecular electronic to overcome the limitations of silicon based devices. Among several types of molecular devices, molecular memory using biomaterial is a major research theme nowadays. The naturally occurring biomaterials like metalloproteins are nano-sizes and possess inherently charge retention property, which make them to use as a high density molecular device platform. Pseudomonas aeruginosa azurin, the protein of interest in this research, has copper as a key element in the electron transfer mechanisms. This can be used as an electron acceptor in the development of molecular electronic device by mimicking biological mechanism. To achieve their information storage properties on solid state with reproducibility and reliability, the robust fabrication of molecular layers with fine orientation are one of the key processes. We have modified azurin with different number of cysteine residues at its amino acid chain based on site-directed mutagenesis. The resulting recombinant protein, azurin, retained its original redox property in the same manner as native azurin. Recombinant azurin was immobilized on Au substrate by strong affinity between thiol of cysteine and gold. The orientations of recombinant azurin with cystein residues immobilized on the Au substrate were analyzed by fluorescence microscope, scanning tunneling microscope, and surface plasmon resonance. Our data revealed that binding activity of recombinant azurin with two cystein residue on the Au substrate significantly increased in comparison to single residue auzrin. Immobilization of highly oriented recombinant azurin based on cysteine-modification could be useful for the nanoscale fabrication of bioelectronic device. Acknowledgments: This research was supported by the Nano/Bio Science & Technology Program of the Ministry of Education, Science and Technology (MEST), by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (2006-05374), and by Seoul Research and Business Development Program (10816).

In this study, we optimized the important process parameters of crystallization to obtain a high purity and yield of glycopeptide antibiotics vancomycin in a purification step. The optimal ratio of acetone/water, pH, conductivity, and initial vancomycin concentration were 3.5/1 (v/v), 2.5, 20 (ms/cm), and 0.1 (g/mL), respectively. The morphology of the vancomycin crystal was analyzed during crystallization using a video microscopy system. The vancomycin was efficiently purified by cryatallization, increasing in purity from 88% to over 97%, with a yield of 95%.

In this study, we intended to increase mass production of Bacillus subtilis through medium optimization by statistical experimental method. For the seed culture of B. subtilis, tryptic soy broth and 15 hours were selected as the basic medium and inoculum age respectively. Final dry cell weight and protease contents of B. subtilis by basic production medium were 1.8 g/L and 0.04 U/mL, respectively. First, maltose, yeast extract and potassium phosphate dibasic were selected as carbon and nutrient sources for mass production of B. subtilis by using one factor at a time method. And then Plackett-Burman experimental design was used to determine the key factors which are critical to produce the biological products and to culture the cell by fermentation. According to the result of Plackett-Burman experimental design, key factors were selected yeast extract and Na2CO3. to obtain the optimum concentrations of two selected variables the response surface methodology was performed . The optimized medium composition found consisted of 20 g/L maltose, 18.5 g/L yeast extract, 2 g/L K2HPO4, 0.36 g/L Na2CO3, 0.4g/L MgSO4 and 0.01g/L CaCl2. Dry cell weight (22.3 g/L) by optimum production medium were increased 12 times, as compared to those determined with basic production medium. Fermentation conditions were examined for the mass production of B. subtilis. The effect of temperature, agitation speed, pH and aeration rate on the mass production of B. subtilis were also observed in a batch fermenter which was carried out in a 2.5 L bioreactor with a working volume of 1.5 L containing optimized production medium. As a result, Dry cell weight of batch culture was 83.8 g/L at 37℃, 300 rpm, pH 7.0 and 3 vvm.

Selection of biomass which is inexpensive and long term-supplied became an important issue due to environmental and ethical reason. Marine biomass is getting more attention as one of the candidates for substrates for biofuels since algae are rich in carbohydrates and minerals and can be cultivated without limitation of water and land. Unlike use of starch-based substrates, corn and sugarcane, use of algae as substrates for biofuel production can avoid the ethical issue such as shortage of food supply. In addition, it is also expected that the hydrolysates of algae can be better fermentation substrates due to its lower content of lignin compared to lignocellulosic materials. In the case of red algae, they are high in galactan and glucan, which can be converted into galactose and glucose upon hydrolysis, respectively. Fermentation of galactose is a problematic since butanol is produced at low yield when galactose is used as a sole carbon source. In order to enhance the conversion of galactose into butanol, we performed a chemical mutagenesis using clostridium beijerinckii and selected for both galactose-hyperutilizing and butanol-hyperproducting phenotypes. Two mutant strains were isolated and characterization of mutant strains is currently undertaken.

Nanotechnologies are developing rapidly in many industries. Especially titanium dioxide has been widely used in many ways as an additive, including as a white pigment, foodcolorant, and sunscreens and cosmetic cream. However, with useful properties, concerns about their potential toxicity and environmental impact are arising . The purpose of this study is to test of many used commercial nanoparticle, and to prove that antibacterial effect. Antimicrobial activity with commercial product of TiO2 nanomaterials were tested using Gram-negative Escherichia coli and Gram-positive Bacillus subtilis, and yeast, Saccharomyces cerevisiaeyeast strain. In percentage survival rate, S. cerevisiae showed highest survival rate than other two species. TiO2 nanoparticles were tested effect of wavelength and maintenance effect. Then to compare antimicrobial activity, TiO2 nanoparticles have antibacterial effect only UVC range.

In the previous study, we reported that indigo was produced by recombinant E. coli harboring a flavin-containing monooxygenase gene. This recombinant E. coli produced 5.6g/L/day of indigo for 5day on the tryptophan medium (0.2% tryptophan, 1% NaCl, 0.5% yeast extract) through fed-batch cultivation. In this study, investigated the expression of fmo gene in the E. coli DH5α, Bacillus subtilis and Corynebacterium glutamicum to improve the stability of indigo production. The fmo gene(1686bp) was amplified and subcloned into the vectors such as PHY300PLK, pEKEx2, pHCEIIB, pGEX4T-1 and PBSK. The recombinant plasmids produced indigo such as 182 mg/L (B. subtilis/pHY300PLK), 377mg/L (C. glutamicum/pEKEx2), 672.3 mg/L (E. coli DH5α/ pBSK SKII), 400.2 mg/L (E. coli DH5α/pHCE II B) and 638.5 mg/L (E. coli DH5α/pGEX4T-1), respectively.

Dextran is a generic term for a bacterial exopolysaccharide synthesized from sucrose and composed of chains of D- glucose units connected by α -1,6-linkages by using dextransucrases. Dextran can be branched out by α -1,2, α -1,3, or α -1,4-linkages, to varying degrees, depending on the strain of Leuconostoc sp. Recently, dextrans have been investigated for delivery of drugs, proteins/enzymes, and imaging agents. L. mesenteroides ATCC 13146 produces two dextransucrases which synthesize two branched dextrans: soluble (S) type and less soluble (L) type. Soluble dextran (precipitated by 45% ethanol) contains 50% of α -D-(1-6) glucose linkages and 50% of α -(1-3) branch linkages. Less soluble dextran (precipitated by 39% alcohol) is made up of 87% of α -D-(1-6) glucose linkages and 13% of α -D-(1-4) branch linkages. In this study dextrans of L. mesenteroides ATCC 13146 was produced by batch fermentation and separated into two different kinds. The less soluble dextrans were precipitated by an ethanol concentration of 39% (v/v) and soluble dextrans were precipitated by an ethanol concentration of 45% (v/v). The molecular weight of less soluble dextran was between MW 500,000 and 2,000,000. Soluble dextran was between 70,000 and 150,000. The molecular weight average of total dextran (50% alcohol, v/v) was between 150,000 to 500,000. The enzymatic hydrolyzates of dextrans by Penicillium or Lipomyces dextranase showed branched oligodextran series of which consisted of glucose, isomaltose, isomaltotriose, and isomaltooligosaccharides greater than DP4. References1. Seymour, F. R., and R. D. Knapp. Structural analysis of dextrans from strains of Leuconostoc and related genera that contain 3-O- α -D-glucosylated α -D-glucopyranosyl residues at the branch points, or in consecutive, linear positions (1980). Carbohydr. Res. 81, 105-129.2. Seymour, F.R., R.D. Knapp, S.H. Bishop, and A.A. Jeanes. High-temperature enhancement of 13C-NMR chemicals shift of unusual dextrans and correlation with methylation structural analysis (1979). Carbohydr Res. 68, 123-140. 3. Mehvar, R. Dextrans for targeted and sustained delivery of therapeutic and imaging agents (2000). J. Control. Release 69, 1–25.

Inonotus obliquus is a white rot fungus that belongs to the family Hymenochaetaceae. Extracts from this fungus have exhibited biological activities, including antitumor, antifungal and antiviral. Especially, the PBP(protein-bound polysaccharide) extracted from Inonotus obliquus have been used as an immune-boosting agent, diabetes and cancer therapies. In this study, the effect of particle size on extraction of PBP form both dried fruit body and mycelium of Inonotus obliquus was determined. Two kinds of milling systems were used. One is a blender(PB) which pulverizes Inonotus obliquus (particle sizes are higher than 100 μm), and the other is a ultra-fine pulverization (UFP) which is good enough to make the fine particles(particle sizes are less than 100 μm). The various extraction methods were also applied to extract the PBP like hot water extraction, microwave extraction and cascade extraction system which is consisted with the 1st step(hot water extraction) and the 2nd step(microwave extraction after hot water extraction). The total content of PBP by using cascade extraction system was increased 3.6(fruit body) and 4.8(mycelium) times as compared to that of PB powder, respectively(fruit body; 31.25 mg/g, mycelium; 63.76 mg/g). Especially, the content of PBP by using microwave extraction using UFP powder was higher then total content of PBP by using combined 2 step extraction method. As a result, UFP might be useful to increase the extraction yield of both dried fruit body and mycelium of Inonotus obliquus.

In this study, the optimum growth conditions of algae in a photobioreactor were investigated. An alga strain was isolated from a lake and identified as Chlorella vulgaris. The isolated strain was cultivated in photobioreactor using BBM (Bold Basal Medium) and 1% glucose for 2 weeks at 28℃ and adjusting pH 7.8. When the effect of lighting hour for the growth of the alga strain, about 12 hr of lighting produced the highest biomass productivity in the presence of urea (1%). In the presence of glucose (1%), a higher growth rate was observed compare to the CO2 as a sole carbon source. At optimal growth conditions, the specific growth rate (μmax) of the isolated strain was 0.045 (in normal condition: μmax = 0.017). In the case of dry cell weight in optimum condition was 3.6 g/L (in normal condition: 1.3 g/L). These results indicate that the isolated alga strain could be used for the biomass production in a photobioreactor.

In this study we attempted to optimize conditions of purifying glutaminase from Lactobacillus reuteri KCTC3594 and to investigate the properties of the enzyme. When the cells of L. reuteri were extracted with high pressure of 15,000 psi three times, the crude extract containing supernatant showed the maximum glutaminase activity, 15.515 U/ml. Glutaminase was purified approximately 20.67-fold with a yield from the cell-free extract of L. reuteri KCTC3594 by protamine sulfate treatment and chromatography methods including anion exchange and gel filtration [1]. The sizes of a major unit and a subunit of the enzyme were presumed 45 kDa and 60 kDa, respectively, by SDS-PAGE. The enzyme activity was optimal at 40 ℃ and pH 7.5, respectively. It was shown that the glutaminase was salt-tolerant as the activity was maintained 50 % at 15 % (w/v) salt. On the other hand, the enzyme was significantly inhibited up to 80% by DON (10 mM) and iodoacetate (50 mM). The glutaminase of L. reuteri can be partially purified by previous methods and showed most of properties were similar with those of other bacteria [2]. In addition, it has high salt-tolerance. The purified glutaminase can be used in food and pharmaceutical industries.