• Journal of Life Science
• /
• v.18 no.2
• /
• pp.264-272
• /
• 2008

The lactate dehydrogenase (EC 1.1.1.27, LDH) isozymes in tissues from Acanthogobius hasta were characterized by biochemical, immunochemical and kinetic methods. The activities of LDH in skeletal muscle and eye tissues were 65.30 and 53.25 units, but LDH activities in heart and liver tissues were very low. LDH/CS (EC 4.1.3.7, citrate synthase) in skeletal muscle was the highest as 22.29. Specific activities of LDH in brain, eye and skeletal muscle were 56.45, 38.04 and 11.0 units/mg, respectively. The LDH isozymes in tissues were separated by polyacrylamide gel electrophoresis after immunoprecipitation with antiserum against $A_4,\;B_4$ eye-specific $C_4$ and liver-specific $C_4$. LDH $AC_4$ isozymes were detected predominantly in skeletal muscle, brain and eye tissues, and $B_4$ isozyme was detected in heart. Anodal eye-specific $C_4$ and cathodal liver-specific $C_4$ were coexpressed in A. hasta. The eye-specific $C_4$ isozyme showed higher activity in eye tissue, but liver-specific $C_4$ isozyme showed lower activity in liver. As a result, one part of molecular structures in $A_4\;and\;C_4,\;A_4\;and\;B_4$, and eye-specific $C_4$ and liver-specific $C_4$ were similar, but in $B_4\;and\;C_4$ were different with each other. Therefore the subunit A may be conservative in evolution, and the evolution of subunit B seems to be faster than that of subunit A. The LDH $A_4$ isozyme of skeletal muscle was purified in the fraction from elution with NAD+ containing buffer of affinity chromatography and eye-specific $C_4$ isozyme was eluted right after $A_4$, so the structure of eye-specific $C_4$ isozyme is similar to $A_4$. And LDH activity remained 35.22-43.47% as a result of the inhibition by pyruvate, the Michaelis-Menten constant values for pyruvate was 0.080-0.098 mM, and Vmax were 153.85 units, 35.09 units in skeletal muscle and eye, respectively. Also the $B_4$ isozyme was the thermo-stablest and $C_4$ was stabler than $A_4$ isozyme. The optimum pH of LDH was 6.5. The results mentioned above indicate that isozymes in tissues showed the properties between LDH $A_4\;and\;B_4$ isozyme as A. hasta was adapted to hypoxic conditions. Also LDH seems to function more effectively under anaerobic condition because LDH in skeletal muscle and eye tissues have high affinity for pyruvate.

• Parasites, Hosts and Diseases
• /
• v.23 no.1
• /
• pp.156-164
• /
• 1985

To elucidate the effect of splenectomy on the development of experimental primary amoebic meningoencephalitis in mice, the death rate and survival time of mice infected intranasally with Naegleria fowleri trophozoites $5{\times}10^4$ cultivated in CGVS medium were compared according to the mouse age when splenectomy was done, and post-operation until experimental infection. Immunodigusion was undergone to detect the presence of serum antibod). due to N, fowleri infecttion in mice. Polyacrylamide gel electrophoresis was done to compare the protein fractions of mouse serum in each experimental groups. In experiment I, splenectomy was done 3 weeks and infection 4 weeks after birth, the death rate of control, sham operated and splenectomized group were 100%, 85% and 95%, and the mean survival time after infection 7.3 days, 7.5 days and 7.8 days, respectively. In experiment II, splenectomy was undergone 3 weeks and infection 6 weeks after birth, the death rate of of control, sham operated and splenectomized group were 95%, 95% and 95%, and the mean survival time after infection 12.1 days, 11.5 days and 11.5 days, respectively. In experiment III, splenectomy was done 5 weeks and infection 6 weeks after birth, the death rate of control, sham operated and splenectomized group were 95%, 90% and 95%, and the mean survival time after infection 8.1 days, 8.3 days and 8.5 days, respectively. By Ouchterlony immunodigusion, anti-JV. fowleri antibody in the serum of mouse with primary amoebic meningoencephalitis was detected against a N. fowleri antigen, which was prepared by ultrasonication of N, fowleri trophozoites, each reacting two lines of precipitation. The patterns of serum fractions by polyacrylamide gel electrophoresis were different between control and sham operated groups from splenectomized group in fraction II, III and V, the sera of which were collected after N. fowleri infection. This results may be summarized as that splenectomy has no effect on the development of primary amoebic meningoencephalitis in mice.

• Microbiology and Biotechnology Letters
• /
• v.44 no.3
• /
• pp.355-362
• /
• 2016

The phosphomannomutase/phosphoglucomutase gene (pmm/pgm) of Vibrio anguillarum (the causative agent of fish vibriosis) was cloned, and the open reading frame corresponded to a protein with 446 amino acids. The pmm/pgm gene showed a significant degree of sequence homology with the previously reported genes from V. mimicus, V. vulnificus, V. splendidus, and V. harveyi, with 92.3%, 91.4%, 89.9%, and 89.9% amino acid identity, respectively. By reverse transcriptase-polymerase chain reaction, we found that the pmm/pgm gene was upregulated under cold stress condition. The PMM/PGM protein is known to catalyze the interconversion between mannose-1-phosphate and mannose-6-phosphate or glucose-1-phosphate and glucose-6-phosphate, which are important intermediates for lipopolysaccharide (LPS) biosynthesis. To confirm the role of PMM/PGM in the LPS biosynthetic pathway, we constructed a knock out mutant by homologous recombination. The respective LPSs were isolated from the V. anguillarum wild-type and mutant strains, and changes were compared by subjecting them to sodium dodecyl sulfate polyacrylamide gel electrophoresis. Based on the different patterns of the LPSs, we expect the pmm/pgm gene to have an important role in LPS biosynthesis. The pmm/pgm-deficient mutant of V. anguillarum will contribute to further studies about the role of LPS in V. anguillarum pathogenesis.

• Applied Biological Chemistry
• /
• v.51 no.2
• /
• pp.142-147
• /
• 2008

Nuruk is the Korean traditional Koji that contains various microorganisms and has been used to make the traditional fermented foods including alcoholic beverages. Rhizopus oryzae KSD-815 was isolated from the alcohol-fermenting Nuruk used for manufacturing traditional alcohol. In this study, the authors reported the isolation and identification of four lipids from the Nuruk (Rhizopus oryzae KSD-815) that inoculated wheat with Rhizopus oryzae KSD-815. The dried and powdered Nuruk (Rhizopus oryzae KSD-815) were extracted three times at room temperature with 80% aqueous MeOH. The extracts were partitioned with EtOAc, n-BuOH, and water, successively. The EtOAc extract was suspended in 80% MeOH and partitioned repeatedly with n-hexane. From the n-hexane fraction, four lipids were isolated through the repeated silica gel and ODS column chromatographies. According to the results of physico-chemical data including NMR, GC and MS, the chemical structures of the compounds were determined as linolenic acid methyl ester (1), palmitic acid methyl ester (2), linoleic acid (3), palmitic acid (4). Cytotoxicity was evaluated in huamn breast cancer cells, MDA-MB-231 and human hepatocarcinoma, SK-HEP-1 cells using MTT assay. Exposure of compounds 1 and 3 led to a dose-dependent inhibition of cell viability in both cancer cell lines. In addition, treatment of RAW264.7 cells with compound 3 caused inhibition of lipopolysaccharide/interferon-${\gamma}$-induced nitric oxide production.

• Korean Journal of Food Science and Technology
• /
• v.17 no.2
• /
• pp.121-127
• /
• 1985

In studying the structural work on ciguatoxin, parrot fish collected were identified as Scarus sordidus, S. frenatus, S. scaber and S. pectarlis, in which only S. sordidus contained toxic materials. Crude toxins obtained by silicic acid column chromatography, could be separated on a DEAE-cellulose column into two fractions, ST-1(less polar) and ST-2(polar) eluted with chloroform and chloroform-methanol(1:1). Furthermore ST-1 could be changed into ST-2 by repeated chromatography on DEAE-cellulose. Rf values of ST-1 and ST-2 were 0.60-0.75 and 0.30-0.54 on TLC coated with silica gel 60F-254 developed by chloroform-methanol-water-acetic acid (90:9.5:0.2:0.3) mixture. The peaks of ST-1 and ST-2 were not observed on each HPLC chromatogram at low sensitivity(2X), but by bioassay they were detected in the fraction of 24-27ml(less polar toxin, 120ng) and 22-27 ml (polar toxin, 150 ng). Less polar ciguatoxin from morey eel viscera also showed its peak in the same elution volume(25ml). Being subjected to chromatography on basic aluminum oxide (activity grade I) or to alkaline treatment, followed by basic aluminum oxide (activity grade I) chromatography ST-1 toxin was remarkably converted into the polar toxic component supposed to be polar ciguatoxin in both cases. In the latter case, approximately 74% of the residual toxicity was changed into the polar component, accompanied by about 50% loss of the initial toxicity. More than 26% of ST-2 toxicity was transformed into the less polar toxic component supposed to be less polar ciguatoxin on a deactivated aluminum oxide (activity grade V) column.

• Korean Journal of Environmental Biology
• /
• v.35 no.4
• /
• pp.694-705
• /
• 2017

The aim of this study was to isolate and identify marine bacterium with anti-methicillin-resistant Staphylococcus aureus (MRSA) activity, and to purify the anti-MRSA compound, as well as to determine its activity and synergistic effects. Among the marine bacteria isolated in this study, the YJ-1 isolate had the strongest anti-MRSA activity. The YJ-1 isolate was identified on the basis of its biochemical characteristics and an analysis of 16S rRNA gene sequences. The YJ-1 isolate showed over 99.2% homology with Pseudomonas stutzeri, and was designated as a Pseudomonas sp. YJ-1. The optimal culture conditions were $25^{\circ}C$ and initial pH 7.0. For the purification of the anti-MRSA compounds, the YJ-1 was cultured in Pa PES-II medium, and the culture filtrates were extracted by ethyl acetate, hexane, and 80% MeOH. The 80% MeOH fraction was separated by a $C_{18}$ ODS column, silica gel chromatography and a reverse phase HPLC, to yield three anti-MRSA agents, the MR1, MR2, and MR3 compounds. When the MR1 compound of $250{\mu}g\;mL^{-1}$ concentration was applied to the MRSA cells, over 95% of bacterial cells was killed within 48 hr. Compared with vancomycin and ampicillin, the MR1 compound showed significant anti-MRSA activity. In addition, the anti-MRSA activity was increased by dose and time dependent manners. Furthermore, the combination of an MR1 compound with vancomycin produced a more rapid decrease in the MRSA cells than did the MR1 compound alone. Taken together, our results suggest that the Pseudomonas sp. YJ-1 and its anti-MRSA compounds could be employed as a natural antibacterial agent in MRSA infections.

• The Korean Journal of Mycology
• /
• v.29 no.1
• /
• pp.1-8
• /
• 2001

To examine the structural properties of the proteoglycan (GMPG, Ganoderma lucidum mycelial proteoglycan) obtained from mycelia in Ganoderma lucidum IY009, we obtained the low and high molecular proteoglycan by ultrafiltration and sepharose CL-4B column chromatography. The physicochemical properties of these fractions were as follows. When the proteoglycan separated by ultrafiltration and sepharose CL-4B column chromatography, its was not fractionated completely. The molecular weight of high molecular proteoglycan by the gel column chromatography (CH) was 250 kD and 2,000 kD, and low molecular proteoglycan was 12kD. The total carbohydrate was consisted of 75.7% (UH) and 96.7% (CH), and the low fraction was 72.7% (UL) and 87.1% (CL), respectively. The sugar of high and low molecular proteoglycan composed of glucose, mannose, fructose, galactose, xylose, ribose and arabinose. Glucose contents of all fraction were ranged from $46.9%{\sim}82.4%$ of the total sugar and the ratio of ${\alpha}$\;and\;{\beta}-glucose$ was $0.84{\sim}1.14$, and its indicated the proteoglycan to be ${\beta}-glucan$. Amino acids pattern showed that the fractions contained a large amount of aspartie acid, glutamic acid, alanine and leucine. These fractions showed the characteristics of IR absorption for ${\beta}-glucan$ at $890\;cm^{-1}\;and\;^{13}C-NMR$ spectroscopy showed the presence of the ${\beta}-1,3-glucan$ and a ${\beta}-1,6-glucan$.

• Applied Biological Chemistry
• /
• v.20 no.2
• /
• pp.221-227
• /
• 1977

Contents of proline and chloride in halophytes were $80-1700\;{\mu}g/gfw$ and 0.13-0.45 mM/gfw respectively. The content of proline was inversely proportional to that of chloride. Rhizomes of Phargmites communis Trin, a halophyte, were grown in non-saline medium and then taken to saline treatment for one or two weeks. Growth of P. communis was inhibited when salinized with 0.25M NaCl. Total nitrogen decreased and alcohol soluble nitrogen and proline increased when growth was retarded. The quantity of Fraction 1 protein decreased at 0.25M NaCl treatment. The accumulation of proline at high concentration in P. communis suggested that it might play a role in osmotic adjustment.

• Korean Journal of Food Science and Technology
• /
• v.35 no.1
• /
• pp.138-143
• /
• 2003

Total methanolic extract of Chrysanthemum coronarium L. var. spatiosum (Compositae) was revealed to have anti-hepatotoxic activity against galactosamine-induced toxicity on primary cultured rat hepatocytes. After successive partitioning with chloroform, n-butanol, and water, the chloroform fraction showed a significant inhibition activity of 51% at 50 ppm, compared with that of silybin, 45.9% at $100\;{\mu}M$. The chloroform fraction was subjected to silica gel column chromatography and yielded active CH-II, CH-V and CH-VI subfractions, and the anti-hepatotoxic activity of these subfractions were 47.6, 56.3, and 23.2%, respectively, at 50 ppm. Total glutathione contents of CH-II, CH-V, and CH-VI increased by 49.8, 43.9, and 47.5% of the control, respectively at 50 ppm, whereas that of silymarin was, 59.7% at $100\;{\mu}M$ after challenged with galactosamine. The ratio of (reduced glutathione) / (total glutathione) in CH-II, CH-V and CH-VI subfraction showed similar values of $0.86{\sim}0.87$ at 50 ppm, whereas that of silymarin was, 0.85 at $100\;{\mu}M$. The incorporation of $[^3H]-uridine$ uptake into RNA was not affected by these active subfractions.

• Korean Journal of Food Science and Technology
• /
• v.44 no.6
• /
• pp.763-771
• /
• 2012

In this study, total antioxidant properties of extracts from different parts of Lespedeza bicolor were determined using techniques of measuring 1,1-diphenyl-2-picryl hydrazyl/2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)-radical scavenging activity and total phenolic contents. The total antioxidant activities of leaf, stem and root extracts from various solvents (water, 50, 70, 100% ethanol, and hot-water) indicated that 50 and 70% ethanol extracts have high radical scavenging activities and phenolic contents. A systematic approach was used to determine the total antioxidant activity of different solvent fractions of the Lespedeza bicolor extracts, partitioning with chloroform, ethyl acetate, n-butanol, and water, and the ethyl acetate fraction was found to have the strongest antioxidant activity. Antioxidant assay-guided isolation was carried out to isolate potential antioxidant compounds. The ethyl acetate fraction of the leaf extract was subjected to silica gel, LH-20 and RP-18 column chromatography successively, and afforded compound 1, which was identified as eriodictyol by NMR and MS analysis, after which its antioxidant activity was determined.