• Journal of Dairy Science and Biotechnology
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• v.34 no.2
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• pp.99-116
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• 2016

We herein performed animal safety assessment in accordance with Good Laboratory Practice (GLP) regulations with the aim of developing sialic acid from glycomacropeptide (hereafter referred to as "GMP") as an index ingredient and functional component in functional foods. GMP is a type of whey protein derived from milk and a safe food, with multiple functions, such as antiviral activity. A test substance was produced containing 7% (w/w) sialic acid and mostly-hydrolyzed whey protein (hereafter referred to as "7%-GNANA") by enzymatic treatment of substrate GMP. The maximum intake test dose level was selected based on 5,000 mg/kg/day dose set for male NOEL (no-observed-effect-level) and female NOAEL (no-observed-adverse-effect-level) determined by a dose-range finding (DRF) test (GLP Center of Catholic University of Daegu, Report No. 15-NREO-001) that was previously conducted with the same test substance. To evaluate the toxicity of a repeated oral dose of the test substance in connection with the previous DRF study, 1,250, 2,500, and 5,000 mg/kg of the substance were administered by a probe into the stomachs of 6-week-old SPF Sprague-Dawley male and female rats for 90 d. Each test group consisted of 10 male and 10 female rats. To determine the toxicity index, all parameters, such as observation of common signs; measurements of body weight and food consumption; ophthalmic examination; urinalysis, electrolyte, hematological, and serum biochemical examination; measurement of organ weights during autopsy; and visual and histopathological examinations were conducted according to GLP standards. After evaluating the results based on the test toxicity assessment criteria, it was determined that NOAEL of the test substance, 7%-GNANA, was 5,000 mg/kg/day, for both male and female rats. No animal death was noted in any of the test groups, including the control group, during the study period, and there was no significant difference associated with test substance, as compared with the control group, with respect to general symptoms, body weight changes, food consumption, ophthalmic examination, urinalysis, hematological and serum biochemical examination, and electrolyte and blood coagulation tests during the administration period (P<0.05). As assessed by the effects of the test substance on organ weights, food consumption, autopsy, and histopathological safety, change in kidney weight as an indicator of male NOAEL revealed up to 20% kidney weight increase in the high-dose group (5,000 mg/kg/day) compared with the change in the control group. However, it was concluded that this effect of the test substance was minor. In the case of female rats, reduction of food consumption, increase of kidney weight, and decrease of thymus weight were observed in the high-dose group. The kidney weight increased by 10.2% (left) and 8.9% (right) in the high-dose group, with a slight dose-dependency compared with that of the control group. It was observed that the thymus weight decreased by 25.3% in the high-dose group, but it was a minor test substance-associated effect. During the autopsy, botryoid tumor was detected on the ribs of one subject in the high-dose group, but we concluded that the tumor has been caused by a naturally occurring (non-test) substance. Histopathological examination revealed lesions on the kidney, liver, spleen, and other organs in the low-dose test group. Since these lesions were considered a separate phenomenon, or naturally occurring and associated with aging, it was checked whether any target organ showed clear symptoms caused by the test substance. In conclusion, different concentrations of the test substance were fed to rats and, consequently, it was verified that only a minor effect was associated with the test substance in the high-dose (5,000 mg/kg/day) group of both male and female rats, without any other significant effects associated with the test substance. Therefore, it was concluded that NOAEL of 7%-GNANA (product name: Helicobactrol) with male and female rats as test animals was 5,000 mg/kg/day, and it thus was determined that the substance is safe for the ultimate use as an ingredient of health functional foods.

• The Korean Journal of Pesticide Science
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• v.5 no.3
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• pp.1-11
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• 2001

New technologies will have a large impact on the discovery of new herbicide site of action. Genomics, combinatorial chemistry, and bioinformatics help take advantage of serendipity through tile sequencing of huge numbers of genes or the synthesis of large numbers of chemical compounds. There are approximately $10^{30}\;to\;10^{50}$ possible molecules in molecular space of which only a fraction have been synthesized. Combining this potential with having access to 50,000 plant genes in the future elevates tile probability of discovering flew herbicidal site of actions. If 0.1, 1.0 or 10% of total genes in a typical plant are valid for herbicide target, a plant with 50,000 genes would provide about 50, 500, and 5,000 targets, respectively. However, only 11 herbicide targets have been identified and commercialized. The successful design of novel herbicides depends on careful consideration of a number of factors including target enzyme selections and validations, inhibitor designs, and the metabolic fates. Biochemical information can be used to identify enzymes which produce lethal phenotypes. The identification of a lethal target site is an important step to this approach. An examination of the characteristics of known targets provides of crucial insight as to the definition of a lethal target. Recently, antisense RNA suppression of an enzyme translation has been used to determine the genes required for toxicity and offers a strategy for identifying lethal target sites. After the identification of a lethal target, detailed knowledge such as the enzyme kinetics and the protein structure may be used to design potent inhibitors. Various types of inhibitors may be designed for a given enzyme. Strategies for the selection of new enzyme targets giving the desired physiological response upon partial inhibition include identification of chemical leads, lethal mutants and the use of antisense technology. Enzyme inhibitors having agrochemical utility can be categorized into six major groups: ground-state analogues, group specific reagents, affinity labels, suicide substrates, reaction intermediate analogues, and extraneous site inhibitors. In this review, examples of each category, and their advantages and disadvantages, will be discussed. The target identification and construction of a potent inhibitor, in itself, may not lead to develop an effective herbicide. The desired in vivo activity, uptake and translocation, and metabolism of the inhibitor should be studied in detail to assess the full potential of the target. Strategies for delivery of the compound to the target enzyme and avoidance of premature detoxification may include a proherbicidal approach, especially when inhibitors are highly charged or when selective detoxification or activation can be exploited. Utilization of differences in detoxification or activation between weeds and crops may lead to enhance selectivity. Without a full appreciation of each of these facets of herbicide design, the chances for success with the target or enzyme-driven approach are reduced.

• The Korean Journal of Physiology
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• v.19 no.1
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• pp.15-23
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• 1985

Previous biochemical studies indicate that $(Na^++K^+)-ATPase$ is composed of two subunits, ${\alpha}$ and ${\beta}$, in a form of ${\alpha}_2{\beta}_2$ with a molecular weight of approximately 300,000 daltons. There is also suggestive evidence that the $Na^+$, $K^+$ pump in human erythrocytes occurs in a complex with some glycolytic enzymes. We assessed here in situ assembly size of the $(Na^++K^+)-ATPase$ of human erythrocytes by applying classical target theory to radiation inactivation data of the ouabain-sensitive sodium flux and ATP hydrolysis of intact cells and ghosts. Cells(in the presence of cryoprotective agent) and ghosts were irradiated at $-45^{\circ}C$ to $-50^{\circ}C$ with an increasing dose of a 1.5 MeV electron beam, and after thawing, the pump and/or enzyme activities were assayed. Each activity measured was decreased as a simple exponential function of radiation dose, from which a radiation sensitive volume (target size) was calculated. When intact cells were used, the target size of both $(Na^++K^+)-ATPase$ and $Na^+$, $K^+$ pump was found to be approximately 600,000 daltons. This target size of the ATPase was reduced to approximately 325,000 daltons if the cells were pretreated with strophanthidin. When ghosts were used, the target size of the ATPase was again approximately 325,000 daltons. Our target size measurement suggests that, in intact cells, the $(Na^++K^+)-ATPase/Na^+,K^+$ pump exists either as a dimer of $(\alpha\beta)_2$ which is a functional unit or as a monomer of $(\alpha\beta)_2$ but in tight complex with other enzyme or enzymes. The results also suggest that this dimeric or heterocomplex association is dissociated during ghost preparation and strophanthidin treatment.

• Journal of the Korean Society for Marine Environment & Energy
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• v.16 no.1
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• pp.9-16
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• 2013

An experiment was conducted to evaluate the biochemical adverse effect of increased carbon dioxide in seawater on marine polychaete, Perinereis aibuhitensis. We measured the available energy reserves, Ea (total carbohydrate, protein, and lipid content) and the energy consumption, Ec (electron transport activity) of Perinereis aibuhitensis exposed for 7-d to a range of $CO_2$ concentration such as 0.39 (control =390 ppmv), 3.03 (=3,030 ppmv), 10.3 (=10,300 ppmv), and 30.1 (=30,100 ppmv) $CO_2$ mM, respectively. The cellular energy allocation (CEA) methodology was used to assess the adverse effects of toxic stress on the energy budget of the test organisms. The results of a decrease in CEA effect of increased carbon dioxide in seawater from all individual in Ea and Ec. Increase of carbon dioxide reduced pH in seawater, significantly. The chemical changes in sea- water caused by increasing $pCO_2$ might cause stresses to test organisms and changes in the cellular energy allocations. Results of this study can be used to understand the possible influence of $CO_2$ concentration increased by the leakage from sub-sea bed storage sites as well as fossil fuel combustion on marine organisms.

• The Korea Journal of Herbology
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• v.29 no.6
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• pp.117-123
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• 2014

Objectives : Sinhyowoldo-san (SHWDS) is said to be a traditional medicine used for shigellosis, abdominal pain, diarrhea. But mechanism of SHWDS mediated-modulation of immune function is not sufficiently understood. To ascertain the molecular mechanisms of SHWDS 70% EtOH extract on pharmacological and biochemical actions in inflammation, we researched the effect of pro-inflammatory mediators in phorbol-12-myristate-13-acetate (PMA)+ A23187-activated human mast cell line (HMC-1). Methods : In the present research, cell viability was measured by MTS assay. pro-inflammatory cytokine production was measured by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to analyze the activation of mitogen-activated protein kinases (MAPKs), nuclear factor kappa-light-chain-enhancer of activated B cells ($NF-{\kappa}B$). The investigation focused on whether SHWDS inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8), MAPKs and $NF-{\kappa}B$ in PMA+A23187-activated HMC-1 cells. Results : SHWDS has no cytotoxicity at measured concentration (50, 100, and $250{\mu}g/ml$). SHWDS ($250{\mu}g/ml$) inhibits pro-inflammatory cytokine expression in PMA+ A23187-activated HMC-1 cells. Moreover, SHWDS inhibited cyclooxygenase (COX)-2 expression. In activated HMC-1 cells, SHWDS suppressed phosphorylation of extracellular signal-regulated kinase (ERK 1/2) and c-jun N-terminal Kinase (JNK 1/2). Then, SHWDS suppressed activation of nuclear factor $NF-{\kappa}B$ in nuclear, degradation of IkB ${\alpha}$ in cytoplasm. Conclusions : We propose that SHWDS has an anti-inflammatory therapeutic potential, which may result from inhibition of ERK 1/2, JNK 1/2 phosphorylation and $NF-{\kappa}B$ activation, thereby decreasing the expression of pro-inflammatory genes.

• The korean journal of orthodontics
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• v.28 no.1 s.66
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• pp.85-97
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• 1998

The purpose of this study was to observe the effects of sodium fluoride on the bony repair and regeneration processes after the rapid palatal expansion in the growing dogs. Eighteen dogs were divided into experimental and control groups. They were in the late mixed dentition. The rapid Palatal expansion was undertaken in all the animals($180^{\circ}$ turn/day) for ten days. The animals were sacrificed on 0, 15 and 45 days after the finish of expansion. One mg NaF/kg of body weight/day were given orally to the experimental group. Blood samples were drawn before and after expansion and the se겨m calcium, phosphate and alkaline phosphatase level were measured. The undecalcified bone section of midpalatal suture area was made, and observed under the light microscopy The results were as follows ; 1. The day after expansion, the infiltration of inflammatory cells were prominent and the new bone formation started at the edges of the two palatal plates bodering the midpalatal suture in both groups. Especially, the newly formed osteoid were very extensive and the osteoblasts lining the osteoid were very active in the experimental group. 2. At fifteen days after expansion, the active osteoblasts lining the osteold at the surface of trabecular bony spicules and active new bone formation were observed in the both groups. However, the cellular activity and new bone formation were more prominent In the experimental group. 3. At forty five days after expansion, the continuous osteoid and new bone formation and active osteoblasts were observed in the experimental group. But these phenomena were not observed in the control group. In the control group, the numerous osteoclasts were adjacent midpalatal suture and the bony remodeling process was begun. The serum alkaline phosphatase level was maintained highly in the experimental group, but decreased in the control. According to the above results, the author reached the conclusion that sodium fluoride has the stimulation effects on the osteoid production of the osteoblasts during the healing process after the rapid Palatal expansion more continuously.

• Proceedings of the Korea Environmental Mutagen Society Conference
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• 2003.10a
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• pp.34-63
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• 2003

Occupational and environmental exposure to manganese continue to represent a realistic public health problem in both developed and developing countries. Increased utility of MMT as a replacement for lead in gasoline creates a new source of environmental exposure to manganese. It is, therefore, imperative that further attention be directed at molecular neurotoxicology of manganese. A Need for a more complete understanding of manganese functions both in health and disease, and for a better defined role of manganese in iron metabolism is well substantiated. The in-depth studies in this area should provide novel information on the potential public health risk associated with manganese exposure. It will also explore novel mechanism(s) of manganese-induced neurotoxicity from the angle of Mn-Fe interaction at both systemic and cellular levels. More importantly, the result of these studies will offer clues to the etiology of IPD and its associated abnormal iron and energy metabolism. To achieve these goals, however, a number of outstanding questions remain to be resolved. First, one must understand what species of manganese in the biological matrices plays critical role in the induction of neurotoxicity, Mn(II) or Mn(III)? In our own studies with aconitase, Cpx-I, and Cpx-II, manganese was added to the buffers as the divalent salt, i.e., $MnCl_2$. While it is quite reasonable to suggest that the effect on aconitase and/or Cpx-I activites was associated with the divalent species of manganese, the experimental design does not preclude the possibility that a manganese species of higher oxidation state, such as Mn(III), is required for the induction of these effects. The ionic radius of Mn(III) is 65 ppm, which is similar to the ionic size to Fe(III) (65 ppm at the high spin state) in aconitase (Nieboer and Fletcher, 1996; Sneed et al., 1953). Thus it is plausible that the higher oxidation state of manganese optimally fits into the geometric space of aconitase, serving as the active species in this enzymatic reaction. In the current literature, most of the studies on manganese toxicity have used Mn(II) as $MnCl_2$ rather than Mn(III). The obvious advantage of Mn(II) is its good water solubility, which allows effortless preparation in either in vivo or in vitro investigation, whereas almost all of the Mn(III) salt products on the comparison between two valent manganese species nearly infeasible. Thus a more intimate collaboration with physiochemists to develop a better way to study Mn(III) species in biological matrices is pressingly needed. Second, In spite of the special affinity of manganese for mitochondria and its similar chemical properties to iron, there is a sound reason to postulate that manganese may act as an iron surrogate in certain iron-requiring enzymes. It is, therefore, imperative to design the physiochemical studies to determine whether manganese can indeed exchange with iron in proteins, and to understand how manganese interacts with tertiary structure of proteins. The studies on binding properties (such as affinity constant, dissociation parameter, etc.) of manganese and iron to key enzymes associated with iron and energy regulation would add additional information to our knowledge of Mn-Fe neurotoxicity. Third, manganese exposure, either in vivo or in vitro, promotes cellular overload of iron. It is still unclear, however, how exactly manganese interacts with cellular iron regulatory processes and what is the mechanism underlying this cellular iron overload. As discussed above, the binding of IRP-I to TfR mRNA leads to the expression of TfR, thereby increasing cellular iron uptake. The sequence encoding TfR mRNA, in particular IRE fragments, has been well-documented in literature. It is therefore possible to use molecular technique to elaborate whether manganese cytotoxicity influences the mRNA expression of iron regulatory proteins and how manganese exposure alters the binding activity of IPRs to TfR mRNA. Finally, the current manganese investigation has largely focused on the issues ranging from disposition/toxicity study to the characterization of clinical symptoms. Much less has been done regarding the risk assessment of environmenta/occupational exposure. One of the unsolved, pressing puzzles is the lack of reliable biomarker(s) for manganese-induced neurologic lesions in long-term, low-level exposure situation. Lack of such a diagnostic means renders it impossible to assess the human health risk and long-term social impact associated with potentially elevated manganese in environment. The biochemical interaction between manganese and iron, particularly the ensuing subtle changes of certain relevant proteins, provides the opportunity to identify and develop such a specific biomarker for manganese-induced neuronal damage. By learning the molecular mechanism of cytotoxicity, one will be able to find a better way for prediction and treatment of manganese-initiated neurodegenerative diseases.

• The Korean Journal of Nuclear Medicine
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• v.33 no.3
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• pp.306-315
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• 1999

Purpose: We performed this study to evaluate the process of radiation induced apoptosis in A431 skin epithelial cancer cell line. Materials and Methods: Low to high dose radiation (0, 2, 5, 10, 25 Gy) was given to A431 cells by Cs-137 cell irradiator. Apoptosis was evaluated by cell morphology, dye exclusion test, and DNA laddering. Results: Cell viability decreased as the radiation dose increased. Number of apoptotic bodies increased as radiation dose increased. It increased most significantly at 12 hours after irradiation. Lactate dehydrogenase activity in culture medium increased according to radiation dose and time after irradiation. DNA ladders could be identified in irradiated cells, but, it had no correlation with radiation dose or time after irradiation. Conclusion: Radiation-induced apoptosis which was the main course of cell death in A431 cells could be analyzed quantitatively by counting apoptotic bodies under microscope. Apoptosis increased as radiation dose increased.

• Microbiology and Biotechnology Letters
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• v.39 no.4
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• pp.317-323
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• 2011

Strain A-3, an amylase-producing bacteria, was isolated from coastal seawater near Daecheon in the Republic of Korea. It was seen to possess a single polar flagella and grow well, on ASW-YP agar plates, at temperatures of between $20-37^{\circ}C$. However, it grew more slowly at the temperatures of $15^{\circ}C$ and $40^{\circ}C$. Similarly, it was observed to grow abundantly, in an Artificial Sea Water-Yeast extract-Peptone (ASW-YP) liquid medium, in a pH range of 6-9, but not grow at pHs of 4-5 and a pH of 10. Strain A-3 was noted as being close to Pseudoalteromonas phenolica O-$BC30^T$, Pseudoalteromonas luteoviolacea $NCIMB1893^T$, Pseudoalteromonas rubra $ATCC29570^T$, and Pseudoalteromonas byunsanensis $FR1199^T$, with 98.30%, 97.86%, 97.78%, and 97.25% similarities respectively, in its 16S rRNA sequence. A phylogenetic tree revealed that strain A-3 and P. phenolica O-$BC30^T$ belong to a clade. However, strain A-3 differed from P. phenolica O-$BC30^T$ in relation to a number of physiological characteristics. Strain A-3 exhibited no growth above 5% NaCl concentrations, no utilization of D-glucose, D-mannose, D-maltose, or D-melibose, and no lipase (C-14) activity. All of these properties strongly indicate that strain A-3 is distant from P. phenolica O-$BC30^T$ and thus led us to name it Pseudoalteromonas sp. A-3. Pseudoalteromonas sp. A-3 produces ${\alpha}$-amylase throughout growth. Maximal amylase activities of 144.48 U/mL and 149.20 U/mL were seen at pH 7.0 and $37^{\circ}C$, respectively. Pseudoalteromonas sp. A-3's high, stable production of ${\alpha}$-amylase in addition to its biochemical features, such as alkalitolerance, suggest that it is a good candidate for industrial applications.

• Journal of Applied Biological Chemistry
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• v.60 no.1
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• pp.79-86
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• 2017

Microbes in forest are very important due to not only to enhance soil fertility but also maintain a healthy ecosystem by supplying the energy available to living organisms by producing various kinds of enzymes related to degradation of lignocellulosic biomass. In order to isolate a lignocellulosic biomass degrading bacterial strain from the Jurassic park located in Gyeongnam National University of Science and Technology, We used the Luria-Bertani-Carboxymethyl cellulose (CMC) agar trypan blue method containing 0.4 % carboxymethyl cellulose and 0.01 % trypan blue. As a result, we isolated a bacterial strain showing both activity on the CMC and xylan. To identify the isolated strain, 16S rRNA sequencing and API kit analysis were used. The isolated strain turned out to belong to Bacillus species and then named Bacillus sp. GJY. In the CMC zymogram analysis, it showed that one active band of about 28kDa in size is present. Xylan zymogram analysis also showed to have one active band of about 25kDa in size. The optimal growth temperature of Bacillus sp. GJY was $37^{\circ}C$. The maximal activities of CMCase and xylanase were 12 hour after incubation. The optimal pH and temperature for CMCase were 5.0 and $40^{\circ}C$, respectively, whereas the optimal pH and temperature for xylanase was 4.0 and $40^{\circ}C$. Both activities for CMCase and xylanase showed to be thermally stable at 40and $50^{\circ}C$, while both activities rapidly decreased at over $60^{\circ}C$.