• Journal of Microbiology and Biotechnology
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• v.12 no.3
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• pp.476-482
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• 2002

Methanol dehydrogenase (MDH) and c-type cytochromes from marine methanol-oxidizing bacterium, Methylophaga sp. MP, were purified and characterized. The native MDH had a molecular mass of 148 kDa and its isoelectric point was 5.5. Two c-type cytochromes, $c_L\;and\;c_H$, were found, and their isoelectric points were 3.4 and 8.0, respectively. The purified MDH had higher thermal stability than that of the other soil methylotrophic bacteria. The electron flow rate from MDH to cytochrome $c_L$was higher than that from MDH to cytochrome $c_H$, indicating that the physiological primary electron acceptor for MDH is cytochrome $c_L$. The electron transfer from MDH to phenazine ethosulfate (PES, artificial electron acceptor) in the two dye (PES/DCPIP)-linked assay system was not inhibited by NaCl, whereas the electron flow from MDH to cytochrome $c_L$ in the cytochrome/DCPIP-linked assay system was suppressed significantly by NaCl. Metal chelating agents such as EDTA showed the same effects on the MDH activity.

• Journal of Microbiology and Biotechnology
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• v.10 no.5
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• pp.612-619
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• 2000

To investigate the biochemical properties of V. mimicus metalloprotease, whose gene was isolated previously from Vibrio mimicus ATCC33653, overexpression and purification were attempted. The 1.9 kb of open reading frame was amplified by PCR from pVMC193 plasmid which ligated the VMC gene with pUC19 and introduced into Escherichia coli BL21 (DE3) using the overexpression vector, pET22b (+). The overexpressed metalloprotease (VMC) was purified with Ni-NTA column chromatography and characterized with various protease inhibitors, pHs, temperatures, and substrates. The purified VMC showed the proteolytic activity against gelatin, soluble and insoluble collagens, and synthetic peptides. Unlike the observations made with all metalloproteases originated from other Vibrio sp., the VMC did not hydrolyze the casein. The proteolytic activity was critically decreased when the VMC was treated with metal chelating reagents, such as EDTA, 2,2-bipyridine, and 1, 10-phenanthroline. In particular, the 71 kDa VMC exhibited the hemagglutinating activity against human erythrocyte. As the purified VMC was treated with $CuCl_2$ and $NiCl_2$ for the chemical modification of metal binding, the proteolytic activity and hemagglutinating activity were profoundly influenced. The multialignment analysis made on the reported Vibrio metalloproteases showed the difference of amino acid sequence similarity between the two distinctive classes of Vibrio metalloproteases.

• Journal of Microbiology and Biotechnology
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• v.19 no.3
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• pp.257-264
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• 2009

A $\beta$-agarase gene, agaB34, was functionally cloned from the genomic DNA of a marine bacterium, Agarivorans albus YKW-34. The open reading frame of agaB34 consisted of 1,362 bp encoding 453 amino acids. The deduced amino acid sequence, consisting of a typical N-terminal signal peptide followed by a catalytic domain of glycoside hydrolase family 16 (GH-16) and a carbohydrate-binding module (CBM), showed 37-86% identity to those of agarases belonging to family GH-16. The recombinant enzyme (rAgaB34) with a molecular mass of 49 kDa was produced extracellularly using Escherichia coli $DH5{\alpha}$ as a host. The purified rAgaB34 was a $\beta$-agarase yielding neoagarotetraose (NA4) as the main product. It acted on neoagarohexaose to produce NA4 and neoagarobiose, but it could not further degrade NA4. The maximal activity of rAgaB34 was observed at $30^{\circ}C$ and pH 7.0. It was stable over pH 5.0-9.0 and at temperatures up to $50^{\circ}C$. Its specific activity and $k_{cat}/K_m$ value for agarose were 242 U/mg and $1.7{\times}10^6/sM$, respectively. The activity of rAgaB34 was not affected by metal ions commonly existing in seawater. It was resistant to chelating reagents (EDTA, EGTA), reducing reagents (DTT, $\beta$-mercaptoethanol), and denaturing reagents (SDS and urea). The E. coli cell harboring the pUC18-derived agarase expression vector was able to efficiently excrete agarase into the culture medium. Hence, this expression system might be used to express secretory proteins.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.50 no.5
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• pp.500-505
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• 2017

The nucleotides and their related compounds, including ATP (adenosine triphosphate), ADP (adenosine diphosphate), AMP (adenosine monophosphate), IMP (inosine monophosphate), HxR (inosine) and Hx (hypoxanthine), were nearly identical in oysters Crassostrea gigas from the two culture methods. The K-value was lower than the threshold value such as 11.2-12.1. Although oysters have low amount of IMP, it was detected in this experiment. DPPH radical scavenging activity did not vary significantly with sample amounts (100, 300, and $500{\mu}g/mL$). DPPH (1,1-diphenyl-2-picryl-hydrazyl) radical scavenging activity was 76.0-80.7% compare with the ascorbic acid standard. Superoxide anion scavenging activity reached 49.3% in the rack-and-bag culture sample at $500{\mu}g/mL$. However, the reducing power and $Fe^{2+}$ chelating activity were very low compared with their respective standard. The oyster culture methods did not affect oyster quality in terms of antioxidant activities.

• Nutrition Research and Practice
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• v.4 no.1
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• pp.16-22
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• 2010

Flavonoids are known to be effective scavengers of free radicals. In particular, proanthocyanidins are flavonoids that possess cardiovascular protection, antioxidative activities, and immunomodulatory activities. Here, we evaluated proanthocyanidin contents in the total polyphenolic compounds of pine needle extracts prepared by hot water, ethanol, hexane, hot water-hexane (HWH), and hot water-ethanol (HWE). Analysis of each extract indicated that the ethanol extract contained the highest proanthocyanidin concentration. The HWH and hexane extracts also contained relatively high concentrations of proanthocyanidin. On the other hand, proanthocyanidin content analyses out of the total polyphenolic compounds indicated that the HWH extract contained the highest content. These results suggest that HWH extraction is a suitable method to obtain an extract with a high level of pure proanthocyanidins and a relatively high yield. The HWH extract possessed superior activity in diverse antioxidative analyses such as 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferrous ion chelating (FIC), and ferric-ion reducing power (FRAP) assays. In addition, upon assessing the effects of the pine needle extracts on macrophages (Raw 264.7 cell), the HWH extract exhibited the highest activity. In this study, we discerned an efficient extraction method to achieve relatively pure proanthocyanidins from pine needles and evaluated the biological functions of the resulting extract, which could potentially be used for its efficacious components in functional food products.

• Journal of Microbiology and Biotechnology
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• v.23 no.11
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• pp.1536-1543
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• 2013

Proteases produced by Xenorhabdus are known to play a significant role in virulence leading to insect mortality. The present study was undertaken to purify and characterize protease from Xenorhabdus indica, an endosymbiont of nematode Steinernema thermophilum, and to decipher its role in insect mortality and its efficacy to control Helicoverpa armigera. A set of 10 strains of Xenorhabdus isolated from different regions of India were screened for protease activity on the basis of zone of clearing on gelatin agar plates. One potent strain of Xenorhabdus indica was selected for the production of protease, and the highest production (1,552 U/ml) was observed at 15-18 h of incubation at $28^{\circ}C$ in soya casein digest broth. The extracellular protease was purified from culture supernatant using ammonium sulfate precipitation and ion-exchange chromatography. The enzyme was further characterized by SDS-PAGE and zymography, which confirmed the purity of the protein and its molecular mass was found to be ~52 kDa. Further MALDI-TOF/TOF analysis and effect of metal chelating agent 1,10-phenanthrolin study revealed the nature of the purified protease as a secreted alkaline metalloprotease. The bioefficacy of the purified protease was also tested against cotton bollworm (Helicoverpa armigera) and resulted in $67.9{\pm}0.64%$ mortality within one week. This purified protease has the potential to be developed as a natural insecticidal agent against a broad range of agriculturally important insects.

• Nutritional Sciences
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• v.7 no.3
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• pp.144-150
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• 2004

We have investigated the effect of calcium spirulan(Ca-SP) isolated from a blue-green alga Spirulina platensis, which is a sulfated polysaccharide chelating calcium and mainly composed of rhamnose and fructose, on invasion of both B16- BL6 melanoma cells, Colon 26 carcinoma and HT-1080 fibrosarcoma cells through reconstituted basement membrane (Matrigel). Ca-SP significantly inhibited the invasion of these tumor cells through Matrigel/fibronectin-coated filters in a concentration-dependent manner. Ca-SP also inhibited the haptotactic migration of tumor cells to laminin, but it had no inhibitory effect on tumor cell migration to fibronectin-coated filters. Ca-SP prevented the adhesion of B16-BL6 cells to Matrigel- and laminin-substrates but did not affect the adhesion to fibronectin. The pretreatment of tumor cells with Ca-SP inhibited the adhesion to laminin in a concentration-dependent fashion, while the pretreatment of laminin-substrates did not. Ca-SP had no effect on the production and activation of type IV collagenase in gelatin zymography. In contraset, Ca-SP significantly inhibited degradation of heparan sulfate by purified heparanase. The experimental lung metastasis was significantly reduced by co-injection of B16-BL6 cells with Ca-SP in a dose-dependent manner. Seven intermittent ⅰ.ⅴ. injection of 100$\mu\textrm{g}$ of Ca-SP caused a marked decrease of lung tumor colonization of B16-BL6 cells in a spontaneous lung metastasis model. These results suggest that Ca-SP, a novel sulfated polysaccharide, could reduce the lung colonization of B16-BL6 melanoma cells in experimental metastasis model, by inhibiting the tumor invasion of basement membrane Matrigel, probably through the prevention of the adhesion and migration of tumor cells to laminin-substrate and of the heparanase activity.

• Journal of Radiation Protection and Research
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• v.29 no.1
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• pp.1-8
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• 2004

Diphosil, a new version of the organic-inorganic composite resin developed by ANL has a structure of the chelating diphosphonic acid groups grafted to a silica support. To apply Diphosil for the treatment of liquid radioactive waste from nuclear power plants, the adsorption equilibrium and column experiments were carried out for the main radionuclides, $^{137}Cs\;and\;^{60}Co$, in the liquid radwaste stream. Through the adsorption equilibrium experiments, the removal efficiencies of $^{137}Cs\;and\;^{60}Co$, and the effects of non-radioactive ions on the removal efficiency have been measured in various conditions using radiotracers. The breakthrough curves for the tested tracers were obtained from the laboratory scale column tests using the simulated liquid radioactive waste. In addition, the removal capacity of Diphosil is compared with that of Amberlite IRN 77 resin, generally used in nuclear power plants.

• Journal of the Korean GEO-environmental Society
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• v.3 no.1
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• pp.37-45
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• 2002

EDTA-enhanced electrokinetic removal of copper and zinc from contaminated sandy soil was carried out. In desorption equilibrium tests, the required mass ratio of EDTA to metal was 10:1 to obtain over 90% of desorption from soil. The removal of heavy metals with chelating agent EDTA below pH 3 was limited because of EDTA precipitation. In electrokinetic experiments, the pH control at anode chamber was essential and 38% Cu and 56% Zn were removed under 30 mA for 1.5 days. Heavy metal removal was greatly improved by controlling anode and soil pH with circulation of anolyte with NaOH solution, in which >50% heavy metal was removed for 4 days and >70% for 9 days.

• Journal of the Korean Society of Food Science and Nutrition
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• v.27 no.3
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• pp.399-405
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• 1998

This study was carried out to investigate the effects of herb extracts on lipid oxidation and free radical reaction in iron sources reacted with active oxygen species. The catalytic effects of active oxygen on lipid oxidation in oil emulsion tended to show more active in the order of OH, H2O2 and KO2. Herb extracts tended to show a little catalytic effect and active oxygen scavenging ability of herb extracts didn't show. But herb extracts played role as a strong chelating agents to bind iron if Fe2+ ion exist in oil emulsion. The contents of Fe2+ ion and total iron in Salvia miltiorrhiza Bge. and Angelica gigas Nakai were higher than those of Prunus persica Stockes and pinus strobus. The content of asocrbic acid in Pinus strobus showed the highest (26.97ppm) among several herb extracts. Electron donating abilities of Pinus strobus and Salvia miltiorrhiza Bge. were 79.54% and 77.11%, respectively, which were higher contents than those of Prunus persical Stokes and Angelica gigas Nakai. The SOD-like activity of Prunus persca Stokes showed 0.16 optical density (O.D), which means the most strong antioxidant activity among other herb extracts. The nitrite scavening effects tended to be different depending on pH. Pinus strobus and Angelica gigas Nakai showed 99.8% and 98.6% nitrite scavening effects at pH 1.2. And the effects were decreased as pH was increased. Especially, they didn't show the nitrite scavenging effect in pH 6.0. In conculsin, the Prinus strobus extract among herb extracts were the most effective antioxidant by evaluating several functional tests.