• Herbal Formula Science
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• v.15 no.1
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• pp.199-211
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• 2007

Inducible nitric oxide synthase (iNOS) are important inflammation enzyme and severe up-nitric oxide (NO) production by this enzyme has been intricated with pathogenesis of inflammation diseases as atopy dermatitis. The present study was designed in order to determine whether Sopungdojeok-san could inhibit atopy dermatitis through modulation of iNOS mRNA expression and NO production, We found that iNOS mRNA expression and NO production in RAW 264.7 macrophages stimulated with lipopolysaccharide dose-dependantly decreased by Sopungdojeok-san extract treatment (0.4 - 1.0 mg/ml). The distribution of iNOS positive reacted cell in atopy dermatitis elicited skin of mice were remarkably decreased by Sopungdojeok-san administration (2.5 ml/kg/day). The SOD ability of Sopungdojeok-san were dose-dependantly increased from 0.6 mg/ ml than butylated hydroxyanisole. These data likely indicate that Sopungdojeok-san may act as inflammatory regulator for atopy dermatitis may be possible to develop useful agent for chemopreventation of NO-intricate inflammatory diseases.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.5
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• pp.1092-1098
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• 2007

Inducible nitric oxide synthase (iNOS) are important inflammation enzyme and severe up-nitric oxide (NO) production by this enzyme has been intricate with pathogenesis of atopy dermatitis. The present study was designed in order to determine whether Cheonggi-san could inhibit atopy dermatitis through modulation of iNOS mRNA expression and NO production. We found that iNOS mRNA expression and NO production in RAW 264.7 macrophages stimulated with lipopolysaccharide dose-dependantly decreased by Cheonggi-san extract treatment (0.5 - 2.0 mg/ml). The distribution of iNOS positive reacted cell in NC/Nga mice with atopy dermatitis were decreased by Cheonggi-san extract treatment (2.5 ml/kg/day) and apoptosis were increased. These data likely indicate that Cheonggi-san may act as inflammatory regulator for atopy dermatitis and may be possible to develop useful agent for chemoprevention of NO intricate inflammatory diseases.

• THE JOURNAL OF KOREAN ORIENTAL ONCOLOGY
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• v.5 no.1
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• pp.137-150
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• 1999

Macrophage play a major role in host defence against infection and tumor development and this activity is regulated through the production of several mediators. In particular, the production of NO by macrophages mediates killing or growth inhibition of tumor cells, bacteria, fungi and parasites. However, over-expression of iNOS by various stimuli, resulting in over-production of NO, contributes to the pathogenesis of septic shock and some inflammator and auto-immune disease. Therefore, it would be valuable to develop potent and selective inhibitors of for potential therapeutic use. Thus the agent that supprees the expression of iNOS mRNA or enzyme protein will be usefull for the prevention of various diseases. We are intersted in identifying selective inhibitiors of iNOS which might be useful intreating inflammatory human diseases. In summary, we have demenstrated that scopoletin, isolated from Seman Armenicae and Radix Trichosanthis the production of NO induced by $IFN-\gammer$ plus LPS in RAW 264.7 macrophages, The mechanism for the inhibition of NO production was due to suppression of the expression of iNOS mRNA or enzyme protein.

• Bulletin of the Korean Chemical Society
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• v.25 no.1
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• pp.115-118
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• 2004

We have examined the feasibility of using the specific interaction between mistletoe lectin I (ML I) and ${\beta}$-Dgalactose instead of the anti-ML I antibody in developing a homogeneous type competitive binding assay for ML I. We also have examined the feasibility of adapting the biotin/avidin mediated homogeneous assay for this system. Alkaline phosphatase (AKP) was employed as a single substrate enzyme label. The dose-response curve shows a detection range of 1-25 ${\mu}$g/mL and a linear response with a correlation coefficient of 0.99. To demonstrate the analytical utility of this method, 10 ${\mu}$g/mL of ML I was spiked into distilled water. The results show that the mean recovery was 10.03 ${\mu}$g/mL with an SD of 0.18. The difference between the spiked value and the mean recovery was 0.03 ${\mu}$g/mL, with a relative error of 0.3 and 1.6 % of RSD.

• Journal of Microbiology and Biotechnology
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• v.6 no.4
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• pp.250-254
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• 1996

MR-387A [(2S, 3R)-2-hydroxy-3-amino-4-phenylbutanoyl-L-valyl-L-prolyl-(2, 4-trans)- L-4-hydroxy-proline] reversibly inhibits aminopeptidase N (BC 3.4.11.2) in a process that is remarkable for its unusual degree of time dependence. The time required to inactivate the enzyme by 50$%$ ($t_{1/2}$) for establishing steady-state levels of $EI^*$complex was approximately 5 minutes. This indicates that the inhibition is a slow-binding process. In dissociation experiments of $EI^*$ complex, enzymic activity was regained slowly in a quadratic equation, indicating that the inhibition of aminopeptidase N by MR-387A is tight-binding and reversible. Thus, the binding of MR-387A by aminopeptidase N is slow and tight, with $K_{i}$ (for initial collision complex, EI) and $K_i{^*}$ (for final tightened complex, $EI^*$) of $2.2\times10^{-8}$ M (from Lineweaver-Burk plot) and $4.4\times10^{-10}$ M (from rate constants), respectively. These data indicate that MR-387A and aminopeptidase N are bound approximately 200-fold more tightly in the final $EI^*$complex than in the initial collision EI complex.

• Korean Journal of Pharmacognosy
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• v.35 no.4 s.139
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• pp.309-314
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• 2004

By treating crude enzyme extract from Aspergillus kawachii, the liquiritigenin content in the licorice (Glycyrrhizae Radix) was significantly increased. The liquiritigenin content reached its maximum level (45.7 mg/g licorice extract) after 60 min of incubation with the crude enzyme extract at $37^{\circ}C$, while the inactivated crude enzyme treated control contained trace amount (about 0.11 mg/g) of liquiritigenin. The enzyme-treated licorice extract inhibited more than 50% DPPH radical at 100 ppm and this was about two times higher activity compared to the enzyme-untreated control.

• Korean Journal of Microbiology
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• v.24 no.3
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• pp.251-262
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• 1986

High-molecular-weight ${\beta}-glucosidase$ (EC 3.2.1.21) was purified from the culture filtrate of Trichoderma koningii through a four-step procedure including chromatography on Bio-Gel P-150, DEAE-Sephadex A-50 and SP-Sephadex C-50; and chromatofocusing on Polybuffer exchanger PBE 94. The molecular weight of the enzyme was determined to be about 101,000 by SDS-polyacrylamide gel electrophoreses, and the isoelectric point was estimated to be 4.96 by analytical isoelectric focusing. The temperature optimum for activity was about $55^{\circ}C$, and the pH optimumwas 3.5. The enzyme was considerably thermostable, for no loss of activity was observed when the enzyme was preincubated at $60^{\circ}C$ for 5h. Km values for cellobiose, gentiobiose, sophorose, salicin and $p-nitrophenyl-{\betha}-D-glucoside$ were 99.2, 14.7, 7.09, 3.15 and 0.70 mM, respectively, which indicates that the enzyme has much higher affinity towards $p-nitrophenyl-{\betha}-D-glucoside$ than towards the other substrates, especially cellobiose. Substrate inhibition by $p-nitrophenyl-{\betha}-D-glucoside$ and salicin was observed at the conecntrations exceeding 5mM. Gluconolactone was a powerful inhibitor against the action of the enzyme on $p-nitrophenyl-{\betha}-D-glucoside\;(K_i\;37.9\;{\mu}M)$, wherease glucose was much less effective ($K_i$ 1.95 mM). Inhibition was of the competitive type in each case. Transglucosylation activity was detected shen the readtion products formed from $p-nitrophenyl-{\betha}-D-glucoside$ by the enzyme were analysed using high-performance liquid chromatography.

• Journal of Microbiology and Biotechnology
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• v.8 no.6
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• pp.560-567
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• 1998

We have found that Thermoactinomyces vulgaris KFB-Cl00 produces a chitinase. The optimum temperature and pH of the enzyme activity were $55^{\circ}C$ and 6.5. The enzyme was stable after heat treatment at $80^{\circ}C$ for 30 min and stable in acidic and basic conditions (PH 6.0~11.0). The thermostable endo-chitinase from Thermoactinomyces vulgaris KFB-C100 was cloned into the plasmid pBR322 by using E. coli DH5$\alpha$ as a host strain. The positive clone carrying a recombinant plasmid (PKCHI23) with a 4.1-kb fragment containing the chitinase gene was found. The recombinant plasmid was analyzed to determine the essential region for chitinase activity and obtained a 2.3-kb fragment, which was sub cloned into pTrc99A using the PstI and SalI sites to construct pTrc99A/pKCHI23-3. The resulting plasmid exerted high chitinase activity upon transformation of E. coli XL1-Blue cells. Chitinase was overproduced 14 times more in the clone cells than in the wild-type cells and the enzyme was purified to homogeneity. The purified enzyme showed the similar properties as the native chitinase from T. vulgaris in terms of molecular weight and substrate specificity. The catalytic action of the cloned enzyme was an endo type, producing chitobiose as a major reaction product.

• Journal of the Korean Society of Food Culture
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• v.14 no.5
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• pp.455-460
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• 1999

In order to establish the processing condition of salt-fermented liquefaction of sardine (Sardinops melanoslicta), effect of temperature, pH value, and concentration of salinity on crude enzyme activity of sardine viscera were investigated. The optimum temperature range of crude enzyme activity in sardine viscera was $45{\sim}50^{\circ}C$ and the optimum pH value of it was 9.8. According to the concentration of salinity increased the crude enzyme activity in sardine viscera decreased. The relationship between concentration of salinity (X) and the crude enzyme activity (Y) in sardine viscera is shown as follows; Y=-0.01363X+0.7676 (r=-0.88). For the purpose of processing conditions of rapid- and low salt-fermented liquefaction of sardine, changes of viable cell count, histamine content, and volatile basic nitrogen (VBN) in the chopped whole sardine with 8% NaCl during preheating process at $40^{\circ},\;45^{\circ}$ and $50^{\circ}C$ for 48 hrs were analyzed. During preheating, initial viable cell counts of chopped whole sardine were $10^{4-7}/g$, but they decreased $10^{1-5}/g$ after 48 hrs. Histamine contents during preheating process at $40^{\circ}\;and\;45^{\circ}C$ were gradually increased, whereas at $50^{\circ}C$ were almost the same level after 48 hrs. VBN contents were continuously increased during preheating, but preheating at $50^{\circ}C$ samples were lower level than that of $40^{\circ}\;and\;45^{\circ}C$ ones. For the purpose to accelerate the fermentation and liquefaction of chopped whole sardine, preheating at optimum temperature of crude enzyme activity for 48 hrs was useful processing method and the contents of viable cell count, histamine, and VBN were safety level for food sanitation.

• Biomolecules & Therapeutics
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• v.27 no.6
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• pp.577-583
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• 2019

Human cytochrome P450 2C9 is a highly polymorphic enzyme that is required for drug and xenobiotic metabolism. Here, we studied eleven P450 2C9 genetic variants-including three novel variants F69S, L310V, and Q324X-that were clinically identified in Korean patients. P450 2C9 variant enzymes were expressed in Escherichia coli and their bicistronic membrane fractions were prepared The CO-binding spectra were obtained for nine enzyme variants, indicating P450 holoenzymes, but not for the M02 (L90P) variant. The M11 (Q324X) variant could not be expressed due to an early nonsense mutation. LC-MS/MS analysis was performed to measure the catalytic activities of the P450 2C9 variants, using diclofenac as a substrate. Steady-state kinetic analysis revealed that the catalytic efficiency of all nine P450 2C9 variants was lower than that of the wild type P450 2C9 enzyme. The M05 (R150L) and M06 (P279T) variants showed high $k_{cat}$ values; however, their $K_m$ values were also high. As the M01 (F69S), M03 (R124Q), M04 (R125H), M08 (I359L), M09 (I359T), and M10 (A477T) variants exhibited higher $K_m$ and lower $k_{cat}$ values than that of the wild type enzyme, their catalytic efficiency decreased by approximately 50-fold compared to the wild type enzyme. Furthermore, the novel variant M07 (L310V) showed lower $k_{cat}$ and $K_m$ values than the wild type enzyme, which resulted in its decreased (80%) catalytic efficiency. The X-ray crystal structure of P450 2C9 revealed the presence of mutations in the residues surrounding the substrate-binding cavity. Functional characterization of these genetic variants can help understand the pharmacogenetic outcomes.