Earticle

Flavonoids are natural compounds with diverse biological functions and served as antioxidants that protect against cardiovascular disease and certain forms of cancer in humans. Glycosylation of flavonoid has a remarkable influence on their pharmacokinetic properties. We generate E. coli BL21(DE3, Δpgi) by deletion of glucose phosohate isomerase (Pgi) gene in E. coli BL21(DE3). A whole-cell biotransformation system was designed by integrating of galU from E. coli K12, and expressing of calS8 (dehydrogenase) and calS9 (decrboxylase) from Micromonospora echinospora spp. calichensis and arGt-3 (3-Oglycosyltransferase) from Arabidopsis thaliana in E. coli BL21(DE3, Δpgi) to form an engineered host E. coli (US89Gt-3) which is expected to produce glycosylated flavonoids. To verify the system, quercetin was fed as a substrate in the engineered host and detected quercetin 3-O-gluronide, a glycosylated product of quercetin. The formation of product was verified by HPLC-LC/MS and MS/MS analyses. Reconstructed host can be applied for the production of various classes of glycosylated flavonoids.

From the cells of Bacillus lichemiformis which can degrade cellulose or other cellulosic materials, the putative cellulase gene, gene ID BLi01882, was cloned. For cell surface display of cellulase, the gene was inserted into the pET21a E. coli vector containing Ice Nucleation Protein (INP) gene as an anchor linker. The INP gene, which is expressed as an outer membrane protein, was obtained from Pseudomonas syringae. The ultimate purpose of artificial surface display is to perform saccharification and fermentation processes simultaneously at a step. Thus, expense and time for production of ethanol can be reduced. In this study, two different cloned vectors, with and without INP gene, were compared in respect of enzyme activity. Activities of the cellulases made, respectively, from two vectors were compared with the positive control using Congo red staining method. The clear zones of cloned products were smaller than the positive control, because it was thought that the cloned products have only one cellulase gene whereas the positive control has more than three kinds of cellulases. When two cloned products, with and without INP gene, were expressed, their enzyme activities did not make a difference on both Congo red staining and SDS-PAGE. It indicates that the fusion of INP gene did not affect either enhancement or inhibition of enzyme expression.

Deletion of large blocks of nonessential genes that are not needed for metabolic pathways of interest can reduce the production of unwanted byproducts, increase genome stability, and streamline metabolism without physiological compromise. Researchers have recently constructed a reducedgenome Escherichia coli strain MDS42 that lacks 14.3% of its chromosome. Here we describe the reengineering of the MDS42 genome to increase the production of the essential amino acid L-threonine. To this end, we overexpressed a feedback-resistant threonine operon (thrA*BC), deleted the genes that encode threonine dehydrogenase (tdh) and threonine transporters (tdcC and sstT), and introduced a mutant threonine exporter (rhtA23) in MDS42. The resulting strain, MDS-205, shows an ~83% increase in Lthreonine production when cells are grown by flask fermentation, compared to a wild-type E. coli strain MG1655 engineered with the same threoninespecific modifications described above. And transcriptional analysis revealed the effect of the deletion of non-essential genes on the central metabolism and threonine pathways in MDS-205. This result demonstrates that the elimination of genes unnecessary for cell growth can increase the productivity of an industrial strain, most likely by reducing the metabolic burden and improving the metabolic efficiency of cells.

Cyclosporine A (CyA), a cyclic polypeptide metabolite extracted from the fungus, Tolyplocadium inflatum, has been widely used as an immunosuppressive agent in organ transplantation both in human and veterinary medicine. To increase CyA productivity by cyclosporineproducing strain of T. niveum, a foreign bacterial gene, Vitreoscilla Hemoglobin gene (VHb), was transformed via protoplast regeneration, followed by its transcription confirmation using RT-PCR. One T. niveum transformant which stably contained and expressed VHb gene led to a 33.5% increase of CyA productivity comparing with the control strain. Although some protoplast-regenerated T. niveum transformants tend to lose CyA productivity, the optimized protoplast transformation should be an efficient alternative strategy to generate a commercially-valuable, yet metabolite low-producing, fungal species, such as CyA-producing T. niveum.

Xylitol is a five-carbon sugar alcohol to be used as a sugar substitute because of its low caloric and anti-cariogenic properties. In this study, the effects of NAD(P)H regeneration on xylitol production were investigated by overexpression of glycolytic enzymes such as ALD6, ACS1 and GPD1 in recombinant Saccharomyces cerevisiae expressing Pichia stipitis xylose reductase (XR). The ALD6 and ACS1 genes encoding aldehyde dehydrogenase and acetyl-CoA synthetase, respectively were PCR-amplified from the genomic DNA of S. cerevisiae. The GPD1 gene coding for NADP+- dependent glyceraldehyde-3-phosphate dehydrogenase was originated from Kluyveromyces lactis. Each gene was introduced into the chromosome of S. cerevisiae BJ3505:δXR and expressed under the control of the GPD promoter. A glucose-limited fed-batch fermentation was performed in a 3.7 L-bioreactor with a complex medium containing 20 g/L glucose and 100 g/L xylose. Among three xylitol-producing systems, recombinant S. cerevisiae co-expressing XR and ACS1 produced a best xylitol concentration of 94.3 g/L at 1.62 g/L-hr its productivity, which were 1.2 and 1.3 times higher than those for the recombinant S. cerevisiae expressing XR only.

Information on metabolic fluxes can be obtained from a metabolic model and a set of measured fluxes, typically the uptake rates of substrates and secretion rates of. The analysis relies on the stoichiometry of cellular pathways, the metabolic demand for growth and optimization principles to estimate the intracellular carbon flux within the defined network. These stoichiometry-based metabolic models have been applied to a variety of organisms, including Escherichia coli. Experimental observations of in vitro concentrations of metabolites are used to metabolic flux analysis. The analysis of mechanism in metabolic system, regulation pattern, and regulation control were carried out by considering various factors in the glycolytic network. In order to compose the regulation map, effectors that mainly affect the enzyme reaction were selected, among the various effectors in the metabolic system of Escherichia coli. The data of enzymatic kinetics related to glycolysis metabolic network of Escherichia coli were taken from literature and databases. MATLAB program which developed in the previous research was used as a simulation tool. The key factors of glycolysis were studied by estimating the elasticity coefficients and parameter elasticities.

Metabolomics allows comprehensive phenotyping of environmentally modified plant systems. Thus, the metabolomics is an approach for measuring time-related biochemical responses and reactions. A timecourse metabolome analysis was conducted to elucidate the metabolic modulation in a response of plant cells to salt stress. Data mining results suggest that the methylation cycle for the supply of methyl groups, the phenylpropanoid pathway for lignin production, and glycinebetaine biosynthesis are synergetically induced as a short-term response against salt-stress treatment. Metabolomics is emerging as a powerful tool not only for research of system biology but also for substantial equivalence study of genetically modified (GM) crop. GM Chinese cabbage containing the bar gene and 24 non-GM Chinese cabbage varieties were analyzed to evaluate the undesirable changes of the GM crops using metabolic fingerprinting and profiling. The metabolome database was analyzed with PCA. The results clearly demonstrate that the time of sampling affects the metabolome and the metabolic fingerprints show a similar range of natural variability for the GM Chinese cabbage.

Genome of Streptomyces peucetius ATCC 27952 contains 23 cytochrome p450 genes. The ability of the cytochrome P450 heme monooxygenases to catalyze difficult oxidation reactions, often with high specificity and selectivity, makes them attractive for numerous biotechnological applications. However they are generally limited by low turnover rates and low stability, and their minimum requirements for catalysis include a cofactor as source of electrons (NAD(P)H), partner proteins for electron transfer, and dioxygen. Cytochrome P450 used in this study to investigate the hydroxylating property expressing in Escherichia coli belongs to the CYP105P2 family from S. peucetius. Owing to the inactivity of the isolated p450 enzymes,coexpression of three-component electron transfer chain in Escherichia coli was done from two plasmids with different selection markers and compatible origins of replication. The enzyme was supported in its activity with putidaredoxin reductase (CamA) and putidaredoxin (CamB) from Pseudomonas putida, supplying this enzyme with the reducing equivalents necessary in class I type electron transfer system. Hydroxylation of flavone ring was achieved as indicated by HPLC/GC-MS analysis.

Using a polyene cytochrome P450 hydroxylase (CYP)-specific genome screening strategy, Pseudonocardia autotrophica KCTC 9441 was previously determined to contain a biosynthetic gene cluster for a Nystatinlike novel polyene compound named NPP in the chromosome (1). The sequence information of an approximately 125.7 kb contiguous DNA region was completely characterized and revealed to contain a total 23 open reading frames (ORFs) involved in a nystatin-like NPP biosynthesis. It has been known that biosynthesis of secondary metabolites such as antibiotics producedby Streptomyces bspecies involves a complex regulatory network in response to nutritional and environmental factors. Among various regulatory genes, there is a family of genes called pathwayspecific regulatory genes, which control the expression of specific target antibiotic biosynthetic pathway gene cluster including a very large polyketide synthase (PKS) gene cluster. Therefore, analysis of the pathway-specific regulatory genes in the antibiotic biosynthetic gene clusters is essential for understanding the regulatory mechanisms as well as for designing strategies to construct strains with enhanced antibiotic productivity (2). Here, we describe the cloning and characterization of nppR genes, a group of NPP pathway-specific regulatory genes located at the flank region of the NPP biosynthetic gene cluster in Pseudonocardia autotrophica. The detailed characterization and their application will be further discussed.

The internal transcribed spacers (ITS) of the nuclear ribosomal 18S–5.8S–26S cistron have been the most popular target region in the nuclear genome. Taraxacum have been historically recognized as weedy plant and is familiar to us as medicinal herb. It comes into fashion as a new type of vegetable in recent year. In Korea, there is just fewness species type, ITS regions of wild type were amplified via the PCR method and then analysis sequencing to distinguish diversity of Taraxacum mongolicum that have been accessed in Gebank. In this study, there are difference nucleotide positions in the constitution of ITS among various type. The length of ITS regions among the two species ranged from 642 to 643bp. They had higher genetic similarities and closer relations, the species could be distinguished by their ITS sequences because they had difference genotypes on the ITS sequences. The ITS sequence of Taraxacum is relative conservation in different types. Compared in ITS sequence with Taraxacum mongolicum, result that high homology in ITS sequencing of them, the resemblance percent is above 90%.

Avermectins produced by Streptomyces avermitilis is a secondary metabolite with powerful anthelmintic and insecticidal activities. Avermectin is produced into eight derivatives. Among those derivatives avermectin B1a is known to be the most effective. S. avermitilis AJ-7, high avermectin B1a producing strain which was developed through random mutation with NTG, was used throughout this research. To increase avermectin B1a production, various materials related to B1a biosynthesis (biotin, L-isoleucine, S-adenosylmethionin) were added to the main culture medium at 3.0 μg/ml, 0.5 g/L, 0.1 mM respectively. And to optimize cultivation condition, research were performed on diverse agitation speeds and aeration condition. The average result obtained from this experiments, show that avermectin B1a was produced about 372.5 mg/L at 700 rpm, 0.8 vvm conditions. Results obtained from this research to increase avermectin B1a production with addition of various materials made contribution to lab scale medium optimization and batch culture using bioreactor.

Previously, Streptomyces interspecies DNA microarray analysis was successfully used to detect global changes in mRNA abundance associated with doxorubicin overproduction in S. peceutius, revealing wblA (SCO3579) as a pleiotropic down-regulator of antibiotic biosynthesis in Streptomyces species (Kang et al, 2007, J. Bacteriol.). Further analysis revealed another potential down-regulator in antibiotic biosynthesis, a tetR-family transcriptional regulator gene, SCO1712. Adventitious expression of SCO1712 reduced doxorubicin production in S. peucetius as well as two major S. coelicolor antibiotics, actinorhodin (ACT) and undecylprodigiosin (RED). The SCO1712 gene disruption from the S. coelicolor chromosome also resulted in a noticeable increase of both antibiotic productions. Moreover, transcripts encoded by major pathway-specific antibiotic activators in S. coelicolor (i.e., actII-ORF4 for ACT, redD/Z for RED, and cdaR for CDA) were stimulated in the SCO1712-disrupted S. coelicolor mutant, suggesting that SCO1712 acts broadly to down-regulate antibiotic biosynthesis in Streptomyces species. Interestingly, a previously-identified down-regulator wblA transcript was also stimulated in the SCO1712-disrupted S. coelicolor mutant, implying that the SCO1712 might down-regulate wblA expression as well as yet-unidentified antibiotic positive regulatory gene(s) which should bypass the wblA-dependent antibiotic down-regulatory system in S. coelicolor.

Klebsiella pneumoniae is a typical microbial strain capable of producing 1,3-propanediol that is a valuable chemical that is used mainly for the synthesis of polymethylene terephalates by polymerization with terephthalates (1). The microbial production of 1,3-PD by K. pneumoniae involves the formation of various by-products which are synthesized through the oxidative pathway. 2,3-Butanediol, one of the main by-products, may serve as an obstacle for obtaining a high purity of 1,3-PD in downstream processes because of its similar boiling point (2). Hence, to eliminate the by-products synthesis, the oxidative branch of glycerol metabolism was inactivated by deleting from the chromosomal DNA the genes involved in the synthesis of by-products. The by-product formation except for acetate was eliminated in the resultant strains, giving the production yield of 0.57 mol/mol which was higher as compared to that of the wild type strain which gave the production yield of 0.47 mol/mol.

Many therapeutic proteins have been produced in mammalian cell culture. But the relatively high cost, complicated production technology, and potential contamination with animal viruses of the mammalian cell culture have been a major concern for its use in large-scale industrial production. Esherichia coli has been widely used as an alternative host for the mass production of therapeutic proteins. Especially, secretion of recombinant proteins to the periplasm of E. coli has several advantages over intracellular production, including enhanced biological activity, higher product stability and solubility, N-terminal authenticity of the expressed peptide and simplified downstream processing. We re-engineered the periplasm of E. coli by removing periplasmic protein genes that might interfere the expression of heterologous foreign proteins. Considering their abundance in the periplasm and their roles in cellular physiology, about 32 periplasmic genes were deleted from E. coli MG1655 using the rapid markerless deletion (NAR 36(14),: e84 (2008)). Alkaline phosphatase (AP) of E. coli, which forms an active enzyme only in the periplasm, was used as a marker to evaluate the secretion efficiency of the restructured periplasm. We could obtain an increased expression level of AP in the periplasm of the re-engineered E. coli strain. This periplasm restructured E. coli will be beneficial for the production of a wide range of human therapeutic proteins through increasing the protein productivity and simplifying the downstream processing.

Butanol has received considerable attention in relation to a possible alternative to gasoline fuel and can be produced by the biological process using Clostridia through acetone-butanol-ethanol (ABE) fermentation. Clostridium beijerinckii is one of the primary butanol producers. To improve the production of butanol in C. beijerinckii, we engineered the strain by random mutagenesis using NTG. Among the NTG-treated cells, we selected 3,000 colonies and transferred to 1 ml P2 medium in each well of 96-well deep plates and cultured. The concentration of butanol of each clone was measured by using gas chromatography. We further selected the mutant strains showing the increase in the butanol production of more than 10%, compared to the wild-type strain, and transferred to 10 ml P2 medium in each 15 ml conical tube and cultured. After two-step screening of 3,000 clones, N005, the selected mutant exhibiting the highest butanol production, showed about 30% increase in the level of butanol, compared to the wild-type strain. When batch cultivation of this mutant strain was carried out in 2 l fermenter containing 1 l P2 medium, the production level of butanol was approximately 14.6 g/l.

Escherichia coli has been explored as a host for butanol production because of its many advantages such as a fast growth and easy genetic manipulation. However, butanol toxicity is the major concern in the biobutanol production, using E. coli. In particular, E. coli growth is severely inhibited by butanol, almost completely stopped by 1% (vol/vol) butanol. Here we developed a new method to increase the butanol-tolerance of E. coli by artificial transcription factor (ATF) libraries which consist of zinc finger (ZF) DNA-binding proteins and an E. coli cyclic AMP receptor protein (CRP). Using these ATFs, we selected a butanol-tolerant E. coli which can tolerate up to 1.5% (vol/vol) butanol, with a concomitant increase in heat resistance. This result shows that E. coli can be engineered as a promising host for high-yield butanol production.

Escherichia coli B has been the major organism for production of various proteins and biomaterials including bioethanol in the field of biotechnology. Recently, The full genome information of two representative descendants of E. coli B strains, REL606 and BL21(DE3), has been determined by our co-workers. In this study, we have constructed 2-D subproteome maps of E. coli B REL606 by analyzing cytoplasmic, periplasmic, inner and outer membrane, and extracellular proteins based on the genome information using experimental and computational approaches. Thus this work represents one of the most comprehensive proteomic analyses of the subproteome of E. coli B. [This work was supported by the Korean Systems Biology Research Project (M10309020000–03B5002–00000) of the Ministry of Education, Science and Technology. Further supports by the LG Chem Chair Professorship, Microsoft, and IBM SUR program are appreciated.]

A shuttle vector, pTOPO-C, which replicated in Escherichia coli and Clostridium beijerinckii was constructed. pTOPO-C vector was derived from pCR2.1TOPO (Invitrogen) which contained pUC origin, ampicillin resistance gene and kanamycin resistance gene. The ampicillin resistance gene was removed from pCR2.1TOPO and the replication origin of the megaplasmid, pSOL1, from the Clostridium acetobutylicum and erythromycin resistance gene for the selection marker in C. beijerinckii was inserted. We also developed a method to transform plasmid into C. beijerinckii by electroporation. This shuttle vector and transformation protocol can provide the basis for metabolic engineering of C. beijerinckii.

Clostridium acetobutylicum is one of the major acetone-butanolethanol (ABE) producers. Cell density of C. acetobutylicum could relate to several factors including, nutrients, substrate inhibition, and limited heat dissipation, etc. To improve cell density, it was necessary; to design a balanced nutrient medium that contains all the necessary components for supporting cell growth, while avoiding inhibition. The maximum cell density at 600 nm (MaxOD600) was measured in order to determine the nutrient balance of each component. The cells were cultured at 37°C in the test tube containing varying concentration of component, anaerobically. From the results, the highest MaxOD600, 12.5, which was 1.8 times higher than that obtained at clostridial growth media (CGM), was obtained from the modified medium, designated C. acetobutylicum media 1 (CAM1). Batch-fermentations in CAM1 were carried out, and the cell concentrations increased gradually through cultivation up to OD600 values of 19.7, 23.7, and 18.6, which were 1.8, 2.6, and 1.4 times higher than those obtained in CGM, by ATCC 824, M5, and M5E1AB strains, respectively. [This work was supported by the Ministry of Knowledge Economy grant funded by the Korea government (10030795). Further supports by the GS-Caltex and the BioFuelChem].

Clostridium acetobutylicum is an attention-grabbing bacterium in that it produces several solvents, especially butan-1-ol. We reconstructed a genome-scale metabolic network of C. acetobutylicum ATCC 824 which comprises 502 reactions and 479 metabolites. Then, we carried out flux balance analysis using both linear and non-linear approach which is a modification of previous studies.1,2) Here, we present metabolic features and feasibilities of C. acetobutylicum, and hypothetical annotations derived during the reconstruction of the network. [This work was supported by the Korea–Australia Collaborative Research Project on the Development of Sucrose-Based Bioprocess Platform (N02071165) from the Korean Ministry of Knowledge Economy. Further support by LG Chem Chair Professorship and Microsoft are appreciated.]

We previously reported the novel β-glucosidase from Shinorhizobium meliloti that had distinct activity to the synthetic indican or plant extracts of Polygonum tinctorium. However, the Shinorhizobium β-glucosidase showed a relatively lower activity for indican and also mainly found in insoluble fraction when expressed solely without any tag or fusion partner, although it showed a quiet high activity toward MUG (4-Methylumbeliferyl-β-D-glucopyranoside) and X-gal in the screening procedures. These shortcomings of the enzyme were needed consideration to screen another enzyme with more high activity toward the target substrate indican or indican-containing plant extract. We, therefore, further attempted to screen the related enzyme from various sources of microbe minded in previous work. Resultantly, we found a potential enzyme with high activity to indican, from A. tumefaciens and then expressed functionally in E. coli. In this presentation, we characterized the enzyme in detail and further analyzed the potential of the enzyme in terms of expression level, solubility and specific activity for practical application of indigo production.

1) This study was to compare effects of various solvent fractions of Rhodiola sachalinensis on several kinds of human cancer cell lines. Experimental studies were performed to measure anticancer activities of human cancer cell growth and human immune B and T cell growth. 2) Both human immune B and T cell growth was increased up to 17% adding R. sachalinensis extracts, compared to the control. 3) The cytotoxicity on human kidney cell (HEK293) was showed on lower 4.18% in adding 1.0 mg/mℓ of ethyl acetate fractions. 4) For human stomach adenocarcinoma cells, breast adenocacinoma cells, hepatocellular carcinoma, and lung carcinoma cells, Chloroform fraction extracts inhibited about 60% of cancer cells growth in 1.0 mg/mℓ of the extracts and its selectivity were about 6.5. 5) These results indicate that chloroform fractions of R. sachalinensis have low cytotoxicity on normal cells and high inhibition activity on cancer cell growth. The chloroform fractions could be developed take a of new functional foods material.

This study was performed to investigate the enhancement of antioxidant activities and skin-whitening effects of Ephedra sinica Stapf extracts by nano-encapsulation. Antioxidant activities and skinwhitening effects of E. sinica were investigated by using aqueous extracts associated with water extraction process at 60℃. Nanoencapsulated E. sinica extracts was showed the strongest free radical scavengering effect as 83.09% by adding 500 μg/mℓ of the sample, compared to the case of other conditions. Strong inhibition effect on melanin production test by Clone M-3 cells was also showed as 51.85% by adding nanoparticles. It also revealed high inhibitory potency on tyrosinase as 209.4% by adding 1 mg/mℓ of sample. From the preliminary observations, we considered that the nano-encapsulation process with edible materials could improve antioxidant and skinwhitening effects.

Chaga mushroom (Inonotus obliquus) has long been used as a folk medicine due to its numerous biological funtions such as antibacterial, anti-allergic, anti-inflammatory and anti-oxidative activities. In the present study, we found that the hot water extract of chaga mushroom activated adipogenesis of 3T3-L1 preadipocytes. The major constituent of the extract was glucose-rich polysaccharides with a molecular mass of 150 kDa. The extract enhanced differentiation of 3T3-L1 preadipocytes, increasing accumulation of lipid droplets in adipocytes in a dose-dependent manner. Immunoblot analysis revealed that the extract increased the expression of adipogenic makers such as PPARg, GLUT4, aP2 as well as accelerated triacylglycerol accumulation that is critical for acquisition of the adipocyte phenotype. In addition, even in the absence of adipogenic stimuli by insulin, the chaga extract strongly induced adipogenesis in 3T3-L1 preadipocytes. Taken together, these results indicate that the chaga extract can be developed as a PPARg agonist for the improvement of insulin resistance mediated with diabetes because ligands for this receptor have emerged as potent insulin sensitizers used in the treatment of type2 diabetes.

1) This study was conducted to enhance extraction yields and biologica activities by optomaizing.process parameters : extraction pressure and temperature by using response surface methodology. 2) Estimated conditions for maximizing extraction of biological active activity components including ginsenosides, total phenolic compounds were easily extracted at MPE (Medium Pressure Extraction, 121℃, 1.8atm, and 20min) process. It also found that the extracts under this condition could yield higher component than other extraction processes. 3) Ginsenosides were qualitatively monitored by TLC and HPLC. On the basis of HPLC analysis, ginsenosides of fresh ginseng was increased by MPE process, compared to other extraction processes. (water extraction, high pressure extraction and ethanol extraction) 4) 65-35% of α-glucosidase activity was inhibited in adding 1.0 mg/mℓ of MPE extracts. 5) It was found that an optimal extraction condition were estimated on 121℃, 1.8 atm for biological activities.

As the consumer’s concerns on health grew, there are more studies on commercial natural antioxidant substances than ever before. In this study, in order to increase the effects of anti-oxidation, sprouts grown under oxidative stress conditions were tested by extracting valid components according to temperature, time of extraction using certain concentration of ethanol and methanol. In the case of hot water extraction, 10 g of dried sprout was added to 100 ml of distilled water at an appropriate temperature for a certain amount of time to extract using heat. The anti-oxidizing effect of the extracted substance was compared by the analysis of the total content of phenol and flavonoid, and the result of an analysis of DPPH free radical scavenging. After comparing the change in amount of antioxidant substance content according to method of extraction using analysis of valid components, overall it showed that the content was the lowest when it was extracted with methanol, and the amount of content of antioxidant substance showed to be the highest when extracted with ethanol. However, there was little difference when compared with the hot water extraction case.

Inhibitory effects of 20 kinds of the Arctic seaweeds were investigated on growth of human cancer cells. The seaweeds were extracted by acetone/dichloromethane and methanol, respectively and then the combined extracts were used as a test sample. Growth inhibition effects of their crude extracts were evaluated in HT1080, HT-29 and AGS human cancer cells using MTT assay. The inhibitory effects of crude extract were increased as a dose dependent manner. Among them, Desmarestia aculeate and Palmaria palmate showed good inhibition effects on the growth of all human cancer cells tested. On the other hand, crude extracts of Laminaria digitata and Laminaria sp. exhibited inhibition effects on the growth of AGS and HT1080 cancer cells and chorda filum revealed the highest inhibition effects on the growth of HT1080 cancer cells.

1) High pressure extraction at 15Mpa 80℃ and 10Mpa 30℃ conditions included extraction yield up to 250% and 150% than conventional extraction(100℃). 2) Cytotoxicity of high pressure extracts on human normalcy kidney cell(HEK 293) was as low as 20.4% and 21.6%, in adding respectively the highest condition of 1.0 mg/ml. 3) Generally, All the from this process had showed higher 7% inhibition of A549, AGS cells through ultra high pressure process. 4) Acording to HPLC analysis, high pressure extraction of HPLC analysis was showed effects of high-pressure process efficiently brake cell wall and maintain metamorphose are expected immuno-modulatory activities. 5) In particular, the use of high-pressure techniques in the processing was examined new material of high biological activity in this research.

We have tested inhibition effects of crude extracts of the Arctic terrestrial plant Cerastium sp. on cell proliferation of HT 1080, HT-29, AGS and MCF-7 human cancer cells by the MTT method at 200, 100, 50, 10 and 1 μg/mL concentrations. The plant was extracted for two days with dichloromethane (CH2Cl2) and methanol (MeOH), respectively and then its crude extracts were combined. The inhibitory effects of crude extract increased as a dose dependent manner and showed potential cytotoxic effect in all human cancer cell lines tested, with IC50 0.18 mg/mL on MCF-7 cells, IC50 0.16 mg/mL on AGS cells, IC50 0.10 mg/mL on HT1080 cells, and IC50 0.09 mg/mL on HT-29 cells. The highest inhibition effects of the crude extracts were observed in HT-29 human colon cancer cell line. These results suggest that we can isolate active compounds from Cerastium sp. to show much more strong anticancer activity.

In general, the bamboo extracts was known to antimicrobial and pestcidal effects. This study was carried out to investigated to pestcidal effect of bamboo extracts (Phyllostachys bambuoides S. et Z, Phyllostachys nigra var. henoni Stapf, Phyllostachys pubescens Mazel, Sasa borealis Nakai) The bamboo extracts prepared from crushed dried bamboo in water, ethanol and methanol solution at 80℃ for 4hour. The pesticidal effect off bamboo extracts was tested by the leaf disk method using Tetranichus urticae. Pesticidal effects were investigated after 24 hour, 48 hour and 72 hour in room temperature. The extract of Phyllostachysnigra var. henonis was most effective when it was extraction with an ethanol. That of a stalk and a leaf extract was good for Fusarium oxysporum and Colletotrichum gloeosporioides and the Phyllostachysnigra var. henonis leaf's extract took effect on Tetranychus urticae pest control. The pesticidal effects of Tetranychus urticae was 45% with EtOH extract. Also, the thyme and rosemary extract was shown the low pesticidal effects. But, when bamboo extracts added herb components were sprayed on strawberry leaf. pesticidal effects increased up to 90%.