• Bulletin of the Korean Chemical Society
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• v.22 no.4
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• pp.362-366
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• 2001

To show the effects of metallothionein (MT) on the activity of enzymes involved in the removal of reactive oxygen species, MT has been added to the assay systems of superoxide dismutase (SOD), catalase and peroxidase. We have used assay systems of SOD based on NADPH oxidation and nitrite formation from hydroxylammonium chloride as an assay of superoxide breakdown rate. The two assay systems showed different results at the high concentration of MT. MT showed the scavenging of superoxide in the SOD assay system in the presence and absence of SOD. MT added to the SOD assay system behaved as an activator of SOD, but apo-MT behaved as an inhibitor. When MT was added to the assay system in the presence of a fixed amount of SOD, the breakdown rate of superoxide increased. The effects of MT on the decomposition of hydrogen peroxide and the activity of catalase and peroxidase decomposing hydrogen peroxide were evaluated. MT decreased the activities of catalase and peroxidase. We have concluded that the function of MT as an antioxidant might effect the level of superoxide scavenging and not the level of hydrogen peroxide.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.350-355
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• 2004

Sulforhodamine B (SRB) assay is a rapid, sensitive, and inexpensive method for measuring cell proliferation and chemosensitivity. However, the lactate dehydrogenase (LDH) release assay is generally used to measure cytototoxicity of infectious microorganisms against host cells. In this study, we investigated the possibility of applying the SRB assay to determine cytotoxicity for infectious microorganisms, and compared the results with those obtained by the LDH release assay. We used Vibrio vulnificus as a model of infectious microorganisms. The SRB assay showed that V vulnificus strongly induced cytotoxic activity against human intestinal cells, Caco-2 and INT-407 cells. The degree of cytotoxicity closely correlated with infection time and number ratios of V. vulnificus to intestinal cells (MOI, multiplicity of infection). Furthermore, cytotoxicity values obtained by SRB assay correlated well with results obtained by the LDH release assay, and both assays gave a linear response with respect to MOI Heat-inactivation of V. vulnificus for 35 min at $60^{\circ}C$ did not induce cytotoxic activity, indicating that viability of V. vulnificus is crucial for cytotoxic activity against intestinal cells. Although both assays are suitable as cytotoxicity endpoints, the SRB assay is recommended for measuring cytotoxicity of infectious microorganisms against host cells because of its significantly lower cost and more stable endpoint than the LDH release assay.

• Advances in Traditional Medicine
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• v.1 no.1
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• pp.45-54
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• 2000

Cannabidiol derivatives (1, 2 and 3), and 5-fluorouracil (4, 5-FU) were tested for their growth inhibitory effects against human oral epitheloid carcinoma cell lines (KB) using two different 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and sulforhodamine B protein (SRB) assay. These compounds showed a potent inhibitory activity in vitro in the micromolar range against KB cell lines. In general, the antitumor activity of these compounds (1, 2, 3 and 4) was in a dose-dependent over the micromolar concentration ranges from $1\;{\mu}M\;to\;100\;{\mu}M$. The comparison of $IC_{50}$ values of these compounds in tumor cell lines shows that their susceptibility to these compounds decreases in the following order: CBD > 5-FU > CBDME > CBDDE by the MTT assay and SRB assay. Cannabidiol derivatives (1, 2 and 3), and 5-FU were tested for their cytotoxic effects on NIH 3T3 fibroblasts using two different MTT assay and SRB assay. These compounds exhibited potent cytotoxic activities in vitro in the micromolar range against NIH 3T3 fibroblasts. In general, the cytotoxic activities of these compounds (1, 2, 3 and 4) were in a dose-dependent over the micromolar concentration range $1\;{\mu}M\;to\;100\;{\mu}M$. The comparison of $CD_{50}$ values of these compounds on NIH 3T3 fibroblasts shows that their susceptibility to these compounds decreases in the following order; CBD > 5-FU > CBDDE > CBDME by MTT assay, CBD > 5-FU > CBDME > CBDDE by SRB assay. These results suggest that cannabidiol (1, CBD) retains the most growth-inhibitory activity against KB cell lines.

• Journal of Pharmacopuncture
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• v.3 no.1
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• pp.1-19
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• 2000

To study anti-cancer effect and molecular biological mechanism of bee venom for aqua-acupuncture, the effects of bee venom on cell viability and apoptosis were analyzed using MTT assay, tryphan blue assay, $[^3H]$thymidine release assay, flow cytometric analysis, and activity of caspase-3 protease activity assay. To explore whether anti-cancer effects of bee venom are associated with the transcriptional control of gene expression, quantitative RT-PCR analysis of apoptosis-related genes was performed. The obtained results are summarized as follows: 1. The MTT assay demonstrated that cell viability was decreased by bee venom in a dose-dependant manner. 2. Significant induction of apoptosis was identified using tryphan blue assay, $[^3H]$thymidine release assay, and flow cytomet1 ric analysis of sub $G_1$ fraction. 3. In analysis of caspase-3 protease activity, the activity had increased significantly, in a dose-dependant manner. 4. Quantitative RT-PCR analysis of the apoptosis-related genes showed that Bcl-2 and Bcl-$X_L$ were down-regulated whereas Bax was up-regulated by bee venom treatment.

• YAKHAK HOEJI
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• v.51 no.6
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• pp.500-507
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• 2007

To investigate the possibility of development as a whitening agent using bamboo extracts (Phyllostachys nigra var. henonis), we measured DPPH assay, NBT/XO assay, intracellular ROS scavenging assay, tyrosinase assay and MSH-induced melanin production in B-16 cells. Bamboo extracts had dose-dependently anti-oxidant activity in DPPH, NBT/XO and intracellular ROS assay. Bamboo extracts appear to inhibit xanthine oxidase directly. Bamboo extracts inhibited not only purified tyrosinase activity but also inhibited tyrosinase activity and melanin production in B16 melanoma cells stimulated by $1{\mu}M\;{\alpha}$-MSH. Anti-oxidant activity and cytotoxicity of ethyl acetate fraction was more potent than those of water fraction, whereas whitening effect of water fraction was stronger than ethyl acetate fraction. Therefore, these results suggest that water fraction of bamboo extracts may be useful for the development as whitening agents reducing cytotoxicity.

• Journal of Society of Preventive Korean Medicine
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• v.12 no.3
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• pp.91-98
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• 2008

80% MeOH extract of black rices fractionated with n-hexane, EtOAc, and n-BuOH. Copper-induced LDL oxidation inhibition assay and ACAT inhibition assay examined with fractions. n-Hexane and EtOAc fractions showed high inhibition activity. We divided n-hexane fraction into three sub-layers(H1 - H3) and EtOAc into eight sub-layers(E1 - E8). H1, H2, E6, and E7 are showed higher inhibition activity than standard in Lp-PLA2 inhibition assay and H1 and E6 are showed higher ACAT inhibition activity than standard.

• BMB Reports
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• v.28 no.1
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• pp.12-16
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• 1995

The branched-chain ${\alpha}$-keto acid dehydrogenase (BCKAD) complex is a rate limiting enzyme which catalyzes the oxidative decarboxylation of branched-chain ${\alpha}$-keto acids. Numerous studies have suggested that BCKAD is subject to covalent modification in vitro via phosphorylation and dephosphorylation, which are catalyzed by a specific kinase and phosphatase, respectively. The biggest difficulty in the assay of BCKAD activity is to arrest the interconversion between the active and inactive forms. BCKAD activity was determined from fresh rat heart and liver tissues using homogenizing and assay buffers containing inhibitors of phosphatase and kinase. The results suggest that a radiochemical assay using ${\alpha}$-keto[1-$^{14}C$]-isovalerate as a substrate for the enzyme can be applied as a reliable method to determine in vitro enzyme activity with arrested interconversion between the active and inactive forms of the BCKAD complex.

• Toxicological Research
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• v.16 no.2
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• pp.141-146
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• 2000

Phthalate esters are used extensively as a plasticizer in the manufacture of plastic products such as PVC bags and medical devices. Recently, phthalate esters have been shown to induce endocrine system mediated responses. However. only a Jew studies have been conducted for estrogenic activity of phthalate esters. In this study estrogenic activities of seven phthalate esters. butyl benzyl phthalate (BBP), di(2-ethylhexyl) phthalate (DEHP), di-n-butylphthalate (DBP), diethylphthalate (DEP), di-n-pentylphthalate (DPP), di-n-propylphthalate (DPrP) and dicyclohexylphthalate (DCHP), were examined in vitro using E-screen assay and competitive binding assay. From the E-screen assay, BBP. DEHP. DBP and DEP showed weak estrogenic activity at the concentration of 5 $\mu\textrm{M}$. The relative proliferative effect (RPE) and the relative proliferative potency (RPP) were 50~70% and 0.01%. respectively, when compared with 500 pM of 17$\beta$-estradiol (E2). In competitive binding assay with the rat uterine estrogen receptor (ER), BBP and DEP showed weak binding potency ［(l/$10^4$~1/$10^5$ of E2］ while DEHP and DBP scarcely bound to ER. These results suggest that some phthalate esters have weak estrogenic activities in vitro.

• The Korean Journal of Food And Nutrition
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• v.30 no.3
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• pp.454-463
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• 2017

Ganoderma lucidum has been traditionally used as a medicine for treatment of bronchitis, arthritis, and high blood pressure, and it has been reported to display many biological activities including anticancer and immune activities. Since mushroom mycelium is known to have excellent biological activities together with mushroom fruiting body, studies on biological activities of mushroom mycelium have been actively conducted. Thus, the present study compared the biological activities before and after the cultivation of Ganoderma lucidum mycelium on Atractylodes rhizoma. When the radical scavenging activity was assessed by the DPPH assay, ARGL (ethanol extract of Atractylodes rhizoma mycelium fermented with Ganoderma lucidum) showed radical scavenging activity of 5.58~82.56% at concentrations of $10{\sim}500{\mu}g/assay$, while AR (ethanol extract of Atractylodes rhizoma) showed radical scavenging activity of 5.27~72.08% at the same concentrations. When measured by using the ABTS assay, ARGL showed higher radical scavenging activity than AR, which was consistent with the result obtained by the DPPH assay. In the MTT assay, the cytotoxicity of ARGL against all cell lines was higher than that of AR. In particular, the cytotoxicities of AR and ARGL against Hep3B at a concentration of $400{\mu}g/assay$ were 71.81% and 86.40%, respectively. In addition, the result obtained by the SRB assay was consistent with the result obtained by the MTT assay. According to the results mentioned above, there is a high probability that medicinal herb cultures using mycelium can be used as sources of functional foods since the cytotoxicities against cancer cells and antioxidant activities increased when the mycelium was fermented with Atractylodes rhizoma.

• Journal of Oriental Neuropsychiatry
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• v.11 no.1
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• pp.1-18
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• 2000

To study the effects of Ramulus et Uncus Uncariae (REUU) on oxygen free radical-mediated damage by hydrogen peroxide $(H_{2}O_{2})$ on cultured spinal sensory neurons, in vitro assays such as MTT assay, NR assay, neurofilament enzymeimmuno assay (EIA), sulforhodamine B (SRB) assay, assay for lactate dehydrogenase (LDH) activity and assay for lipid peroxidation were used in cultured spinal dorsal root ganglion neurons derived from mice, Spinal dorsal root ganglion neurons were cultured in media containing various concentrations of $H_{2}O_{2}$ for 5 hours, after which the neurotoxic effect of $H_{2}O_{2}$ was measured by in vitro assay. The protective effect of the herb extract, Ramulus et Uncus Uncariae (REUU) against H2O2-induced neurotoxicity was also examined. The results are as follows. 1. In NR assay and MTT assay, $H_{2}O_{2}$ significantly decreased the cell viability of cultured mouse spinal dorsal root ganglion neurons according to exposure concentration in these cultures. An additional time course study was done on these cultures. 2. Cultured spinal dorsal root ganglion neurons which were exposed to various concentrations of $H_{2}O_{2}$ showed a quantitative decrease of neuronal cells by EIA and of total protein by sulforhodamine B (SRB) assay, while they showed an increase of both lipid peroxidation and LDH activity. 3. The effect of Ramulus et Uncus Uncariae (REUU) on $H_{2}O_{2}$ induced neurotoxicity showed a quantitative increase in both neurofilament and total protein, but showed a decrease of lipid peroxidation and LDH activity. These results suggest that $H_{2}O_{2}$ has a neurotoxic effect on cultured spinal dorsal root ganglion neurons from mice and that the herb extract, Ramulus et Uncus Uncariae (REUU), was very effective in protecting $H_{2}O_{2}$ induced neurotoxicity by decreasing lipid peroxidation and LDH activity.