• BMB Reports
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• 제33권3호
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• pp.279-284
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• 2000

Using fluorescence probes, 2-(9-anthroyloxy) stearic acid (2- AS) and 12-(9-anthroyloxy) stearic acid (12-AS), we determined the differential effects of local anesthetics (tetracaine-HCI, bupivacaine-HCI, lidocaine-HCI, prilocaine-HCI and procaine-HCI) on the differential rotational rate between the surface (in carbon number 2 and its surroundings including the head group) and the hydrocarbon interior (in carbon number 12 and its surroundings) of the outer monolayer of the total phospholipid fraction liposome that is extracted from synaptosomal plasma membrane vesicles. The anisotropy (r) values for the hydrocarbon interior and the surface region of the liposome outer monolayer were$0.051{\pm}0.001$ and $0.096{\pm}0.001,$ respectively. This means that the rate of rotational mobility in the hydrocarbon interior is faster than that of the surface region. Local anesthetics in a dosedependent manner decreased the anisotropy of 12-AS in the hydrocarbon interior of the liposome outer monolayer, but increased the anisotropy of 2-AS in the surface region of the monolayer. These results indicate that local anesthetics have significant disordering effects on the hydrocarbon interior, but have significant ordering effects on the surface region of the liposome outer monolayer.

• 동의생리병리학회지
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• 제26권4호
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• pp.483-490
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• 2012

This study was designed to identify the effects of Polygoni cuspidati Radix(PCR) on the generation of superoxide anion radicals (${\cdot}O_2{^-}$), nitric oxide (NO), peroxynitrite ($ONOO^-$) in the renal epithelial cells of mouse(LLC-$PK_1$). The effects of PCR on the expression of inflammation-related proteins, IKK-${\alpha}$, phospho-$I{\kappa}B-{\alpha}$, NF-${\kappa}B$ (p50, p65), COX-2, iNOS, IL-$1{\beta}$, VCAM-1, were examined by western blotting. For this study, the fluorescent probes, namely dihydrorhodamine 123 (DHR 123), 2',7'-dichloro dihydrofluorescein diacetate (DCFDA), 4,5-diaminofluorescein (DAF-2) were used. Protein expression levels of IKK-${\alpha}$, phospho-$I{\kappa}B-{\alpha}$, NF-${\kappa}B$ (p50, p65), COX-2, iNOS, IL-$1{\beta}$, VCAM-1 were assayed by western blot. PCR reduced $H_2O_2$-induced cell death dose-dependently. It inhibited the generation of ${\cdot}O_2{^-}$, NO, $ONOO^-$ and $PGE^2$ in the $H_2O_2$-treated LLC-PK1 cells in vitro. PCR inhibited the espression of IKK-${\alpha}$, phospho-$I{\kappa}B-{\alpha}$, COX-2, iNOS, IL-$1{\beta}$ and VCAM-1 genes by means of decreasing the NF-${\kappa}B$ activation. These results suggest that PCR is an effective NO, ${\cdot}O_2{^-}$, $ONOO^-$ scavenger, and this substance recommended to be applied in treatment for the inflammatory process and inflammation-related disease.

• Preventive Nutrition and Food Science
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• 제9권2호
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• pp.150-155
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• 2004

The free radical scavenging activities and the protective effects of Rhus javanica extracts against oxidative damage induced by reactive oxygen species (ROS) were investigated. n-Hexane, ethyl acetate and water fractions were prepared from a methanol extract. DPPH radical, superoxide anion and hydroxyl radical scavenging activities were estimated. Intracellular ROS formation was quantified using fluorescent probes, 2', 7'-dichlorofluorescin diacetate (DCFH-DA) for hydroxyl radical and dihydroethidium (DHE) for superoxide anion. The oxidative DNA damage was investigated by the comet assay in HepG$_2$ cells exposed either to $H_2O$$_2$ or to menadione. The highest $IC_{50}$/ values for DPPH radical scavenging activity was found in the ethyl acetate fraction with a value of 5.38 $\mu\textrm{g}$/mL. Cells pretreated with $\geq$ 1 $\mu\textrm{g}$/mL of the ethyl acetate extract had significantly increased cell viability compared to control cells, which were not pretreated with the extract. Intracellular ROS formation and DNA damage in HepG$_2$ cells, which were pretreated with the various concentrations of Rhus javanica ethyl acetate extract and then incubated either with $H_2O$$_2$ or with menadione, reduced in a dose-dependent manner. These findings suggest that Rhus javanica might have biologically active components which have strong protective effects against ROS induced oxidative damages to the biomolecules, such as cell membranes and DNA.

• 미생물학회지
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• 제35권3호
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• pp.242-247
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• 1999

소양호에서 fluorescent in situ hybridization 방법을 이용하여 세균군집의 계절과 수심에 따른 변화를 조사하였다. 이 방법에 사용된 oligonucleotide probe는 EUB338, ALF1b, BET42a, GAM42a 와 CF probe 였다. 조사기간중 Proteobacteria $\alpha$-group 은 0.7~32.9% 이었으며 $\beta$-group은 1.0~25.8%, $\gamma$-group은 2.4~37.1%, Cytophaga-Flavobacterium group은 4.7~23.6%의 분포를 나타내었다. 계절별로 살펴보면 규조류가 우점하는 봄펄에는 $\gamma$-group 이, 유기물의 농도가 낮고 수온이 높은 여름철에는 $\alpha$-group 이 우점하였으며, 남조류가 우점하는 가을철에 총세균수에 대한 Eubacterial group 의 비율이 크게 감소하여 특정 group의 우점현상은 나타나지 않았다. 이처럼 소양호는 계절과 수심에 따라 군집구조가 변화하였으며, 특히 세균의 군집구조는 식물플랑크톤의 천이와 밀접한 관계를 보이는 것으로 나타났다.

• 대한한방내과학회지
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• 제24권1호
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• pp.75-83
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• 2003

Objectives : Peroxynitrite($ONOO^-$), formed from the reaction of $O2^-$ and NO, is a cytotoxic species that can oxidize several cellular components such as proteins, lipids and DNA. It has been involved in the aging process and age-related diseases such as Alzheimer's disease, rheumatoid arthritis, cancer and atherosclerosis. The aim of this study was to investigate scavenging activities of $ONOO^-$ and its precursors, NO and $O_{2^-}$ and its scavenging mechanism of Zingiberis Rhizoma (ZR). Methods : To investigate scavenging activities of $ONOO^-,\;NO,\;O_{2^-}$ and its scavenging mechanism, we used fluorescent probes like DCFDA, DAF-2 and DHR 123. The $ONOO^-$ scavenging activity on ZR was assayed by measuring oxidized dihydrorhodamine 123 (DHR 123) by fluorometry. The scavenging efficacy was expressed as IC50, showing the concentration of each sample that is required to cause 50% inhibition of DHR 123 oxidation. In a separate study, the protective effect of ZR on $ONOO^-$-induced nitration of bovine serum albumin was investigated through immuno-assay with a monoclonal anti-nitryrosine antibody, and a horseradish peroxidase-conjugated anti-mouse secondary antibody from sheep. Results : ZR markedly scavenged authentic $ONOO^-,\;O_{2^-}$ and NO. It also inhibited $ONOO^-$ induced by $O_{2^-}$ and NO which are derived from SIN-1. The data demonstrated that ZR led to decreased $ONOO^-$ mediated nitration of tyrosine through electron donation. It also inhibited the nitration of bovine serum albumin induced by $ONOO^-$ in a dose-dependent manner. Furtheremore, it blocked LPS-induced ROS and RNS generation. Conclusions : These results suggest that ZR can be developed as an effective $ONOO^-$ scavenger for the prevention of aging process and age-related diseases.

• 동의생리병리학회지
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• 제19권6호
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• pp.1615-1621
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• 2005

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are widely implicated in the aging process and age-related diseases. The present study was carried out to investigate scavenging activities of Junglans sinensis extract and its subfraction using fluorescent probes, DCF-DA, DAF-2 and DHR 123. Jungians sinensis was washed and crushed. The crushed Junglans sinensis was extracted 3times, each time with 3 volumes of methyl alcohol at $60^{\circ}C$ for 24 h. The extract was filtered and evaporated under a reduced pressure using a rotary evaporator to yield 16 g. Scavenging activities of $ONOO^-$ was measured by Kooy' method and ROS was measured by DCFDA assay. Junglans sinensis had the marked scavenging activites of $ONOO^-$, NO and $O_2^-$. Junglans sinensis scavenged $ONOO^-$ through electron donation and dose-dependently inhibited the nitration of bovine serum albumin by $ONOO^-$. Junglans sinensis also had ROS scavenging activity. Especially, ethylacetate fraction of Junglans sinensis showed the most effective scavenging activities for ROS and RNS. These results suggest that Junglans sinensis might be developed as an effective ROS and RNS scavenger Therefore, Junglans sinensis might be used as a preventive agent for the aging and relevant to aging of illness.

• 대한한방내과학회지
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• 제26권1호
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• pp.107-118
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• 2005

Objectives : Peroxynitrite $(ONOO^-)$, fonned from the reaction of $O_2^-$ and NO, is a cytotoxic species that can oxidize several cellular components such as proteins, lipids and DNA. It has been implicated in the aging process and age-related disease such as Alzheimer's disease, rheumatoid arthritis, cancer and atherosclerosis. Due to the lack of endogenous enzymes to thwart $ONOO^-$ activation, developing a specific $ONOO^-$ scavenger is remarkably important. The aim of this study was to investigate scavenging activities of $ONOO^-$ and its precursors, NO and $O_2^-$ and its scavenging mechanism of Ojawhan. Methods : To investigate scavenging activities of $ONOO^-$, NO, $O_2^-$ and its scavenging mechanism using fluorescent probes, DCFDA, DAF-2 and DHR 123. The $ONOO^-$ scavenging activity on Ojawhan was assayed by measuring oxidized dihydrorhodamine 123 (DHR 123) by fluorometry. Oxidative stress was induced by strong oxidants t-butyl hydroperoxide (t-BHP). Endothelial cell (YPEN-1) was used for detection of intracellular oxidative stress. Results : Ojawhan markedly scavenged authentic $ONOO^-$, $O_2^-$ and NO. It also inhibited $ONOO^-$ induced by $O_2^-$ and NO which are derived from SIN-1. Furthennore, ${\underline{Ojawhan}}$ blocked lipopolysaccharide (LPS)-induced $ONOO^-$, $O_2^-$ and NO generation utilizing kidney homogenates of LPS-injected mouse and inhibited t-BHP-induced ROS and $ONOO^-$ in endothelial cell culture system. Conclusions : These results suggest that Ojawhan be developed as an effective $ONOO^-$ scavenger for the prevention of $ONOO^-$ involved diseases and age-related diseases.

• Archives of Pharmacal Research
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• 제25권2호
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• pp.143-150
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• 2002

Fluorescent probes bound to DNA typically display nanosecond decay times and reveal only nanosecond motions. We extend the time range of measurable DNA dynamics using $[Ru(pby)_2(dppz)]^{2+}$ (bpy=2.2'-bipyridine, dppz=dipyrido[3,2-a2',3'-c]phenazine) (RuBD) which displays a mean lifetime near 90 ns. To test the usefulness of RuBD as a probe for diffusive processes in calf thymus DNA, we compared the efficiencies of fluorescence resonance energy transfer (FRET) using three donors which display lifetimes near 5 ns for acridine orange (AO), 22 ns for ethidum bromide (EB) and 92 ns for RuBD, with nile blue (NB) as the acceptor. The F rster distances for AO-NB, EB-NB and RuBD-NB donor-acceptor pairs were 42.3, 52.3, and $30.6{\;}{\AA}$, respectively. All three donors showed dramatic decreases in fluorescence intensities and more rapid intensity decays with increasing NB concentrations. The intensity decays of AO and EB in the presence of varying concentrations of NB were satisfactorily described by the one-dimensional FRET model without diffusion (Blumen and Manz, 1979). In the case of the long-lifetime donor RuBD, the experimental phase and modulation somewhat deviated from the recovered values computed from this model. The recovered NB concentrations and FRET efficiencies from the model were slightly larger than the expected values, however, the recovered and expected values did not show a significant difference. Thus, it is suggested that the lifetime of RuBD is too short to measure diffusive processes in calf thymus DNA.

• Archives of Pharmacal Research
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• 제28권7호
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• pp.839-847
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• 2005

The aim of this study was to provide the basis to further examine the mode of action of ethanol. Fluorescent probes reported to have different membrane mobilities were used to evaluate the effect of dimyristoylphosphatidylethanol (DMPEt) on the lateral and rotational mobilities of liposome lipid bilayers. An experimental procedure, based on the selective quenching of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1,3-di(1-pyrenyl)propane (Py-3-Py) by trinitrophenyl groups, was used. DMPEt increased the bulk lateral and rotational mobilities, and had a greater fluidizing effect on the outer than the inner monolayer. These effects of DMPEt on liposomes may be responsible for some, but not all, of the general anesthetic actions of ethanol.

• 대한한방내과학회지
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• 제27권3호
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• pp.688-697
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• 2006

Objectives : Peroxynitrite ($ONOO^-$), $O_2^-$ and nitric oxide (NO) are cytotoxic species that can oxidize several cellular components such as proteins. lipids and DNA. It has been implicated in the aging process and age-related disease such as Alzheimer's disease, rheumatoid arthritis, cancer and atherosclerosis. The aim of this study was to investigate $ONOO^-$ scavenging activities. and that of its precursors. NO and $O_2^-$ of Trigonellae Semen. Methods : To investigate $ONOO^-$. NO. $O_2^-$ scavenging activities, fluorescent probes, namely 2'.7'-dichlorodihydrofluorescein diacetate (DCFDA). 4.5-diaminofluorescein (DAF-2) and dihydrorhodamine 123 (DHR 123) were used. Protein expression levels of iNOS, COX-2, NF-${\kappa}B$, and VCAM-1 were assayed by western blot. Results : Trigonellae Semen markedly scavenged authentic $ONOO^-$, $O_2^-$ and NO. It also inhibited $ONOO^-$ induced by $O_2^-$ and NO which are derived from SIN-1. Furthermore. Trigonellae Semen inhibited $ONOO^-$, $O_2^-$ and NO generation in LPS-treated ICR mouse kidney postmitochondria. Trigonellae Semen inhibited gene expression of iNOS, COX-2, VCAM-l and NF-${\kappa}B$ (p65) activation. Conclusions : These results suggest that Trigonellae Semen is an effective $ONOO^-$, $O_2^-$ and NO scavenger. and that this substance has a potential role as an inhibitor of the aging process, and in therapy against age-related diseases.