• Title/Summary/Keyword: Fluorescence anisotropy

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Effects of Parathyroid Hormone on the Fluidity of the Plasma Membrane Vesicles of Cultured Osteoblasts

  • Kang, Jung-Sook
    • Journal of Photoscience
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    • v.8 no.3_4
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    • pp.87-92
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    • 2001
  • Intramolecular excimer formation of 1,3-di(1-pyrenyl)propane (Py-3-Py) and fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) were used to investigate the effects of parathyroid hormone (PTH) on the bulk bilayer fluidity of the plasma membrane vesicles isolated from cultured osteoblasts (OB-PMV). In a dose-dependent manner, rat PTH-(1-34) [rPTH-(1-34)] increased the excimer to monomer fluorescence intensity ratio (I'/I) of Py-3-Py and decreased the anisotropy (r) of DPH in OB-PMV. This indicates that PTH increased both the lateral and rotational diffusion of the probes in OB-PMY. Selective quenching of DPH fluorescence by trinitrophenyl groups was utilized to examine the transbilayer fluidity asymmetry of OB-PMV. The anisotropy, limiting anisotropy, and order parameter of DPH in the inner monolayer were 0.024, 0.032, and 0.062 greater than calculated for the outer monolayer of OB-PMY. Selective quenching of DPH fluorescence by trinitrophenyl groups was also utilized to examine the transbilayer effects of PTH on the fluidity of OB-PMV. rPTH-(1-34) had a greater fluidizing effect on the outer monolayer as compared to the inner monolayer of OB-PMV. Thus, it has been proven that PTH exhibits a selective rather than nonselective fluidizing effect within transbilayer domains of OB-PMV.

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Fluorescence Anisotropy Study on the Effect of Phellodendri Cortex's Berberine on Regulation of the Function of DNA (황백(黃柏)의 berberine이 DNA의 기능조절에 미치는 영향에 관한 형광이방성 연구)

  • Lee, Seong Kyung;Han, Hyo Sang;Huh, Sung Ho
    • The Korea Journal of Herbology
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    • v.33 no.5
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    • pp.105-110
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    • 2018
  • Objectives : We tried to observe the fluorescence anisotropy and intensity of ethidium ion in the intercalating binding interaction between DNA and ethidium ions in the presence of berberine, and then tried to explain the effect of berberine on the intercalating interaction of ethidium ion with DNA. Methods : DNA(calf thymus DNA), berberine and ethidium bromide(EtBr) were purchased from Sigma-Aldrich Co. Proper amount of each compound was dissolved in 20 mM sodium phosphate buffer(pH 7.0) containing 100 mM of NaCl to prepare stock solutions. Collections of the fluorescence anisotropy and intensity data were performed on JASCO FP-8300 spectrofluorometer equipped with a polarizer and a Peltier temperature controller. The excitation of ethidium ion was done at 550 nm and the emission data were collected at 600 nm. For Stern-Volmer plot, the fluorescence data were collected at $18^{\circ}C$ and $30^{\circ}C$. Results : According to the results of this research, the weak competitive binding pattern between ethidium ion and berberine appeared in binding with DNA at low ratio of DNA to ethidium ion. But at high ratio of DNA to ethidium ion, this weak competition disappeared. Instead, berberine might bind to DNA by intercalating way. In other words, berberine could de-intercalate ethidium ion from DNA at low concentration of DNA relative to ethidium ion, but could not at high concentration of DNA relative to ethidium ion. In addition, the mechanism of fluorescence quenching of ethidium ion could also proceed differently, depending on the ratio of the amount of DNA to that of ethidium ion. Conclusions : The effect of berberine on the DNA-ethidium ion intercalating interaction could work differently, depending on the relative ratio of the amount of DNA to that of ethidium ion. This study also showed that fluorescence anisotropy analysis is very useful method to obtain detailed information for investigation of the complex binding interactions. In order to fully understand the mechanism of action of the pharmacological effect by berberine, studies on the effect of berberine on the action of proteins such as various enzymes closely related to berberine-induced medicinal effects should be continued.

Photophysical Model of 10-Hydroxybenzo[h]quinoline: Internal Conversion and Excited State Intramolecular Proton Transfer

  • Lee, Junghwa;Joo, Taiha
    • Bulletin of the Korean Chemical Society
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    • v.35 no.3
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    • pp.881-885
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    • 2014
  • Photophysics of 10-hydroxybenzo[h]quinoline (HBQ) has been in controversy, in particular, on the nature of the electronic states before and after the excited state intramolecular proton transfer (ESIPT), even though the dynamics and mechanism of the ESIPT have been well established. We report highly time resolved fluorescence spectra over the full emission frequency regions of the enol and keto isomers and the anisotropy in time domain to determine the accurate rates of the population decay, spectral relaxation and anisotropy decay of the keto isomer. We have shown that the ~300 fs component observed frequently in ESIPT dynamics arises from the $S_2{\rightarrow}S_1$ internal conversion in the reaction product keto isomer and that the ESIPT occurs from the enol isomer in $S_1$ state to the keto isomer in $S_2$ state.

Effects of Barbiturates on Transbilayer Fluidity Domains of Phospholipid Model Membrane Monolayers (Barbiturates가 소의 대뇌피질 Synaptosomal Plasma Membrane Vesicles로 부터 추출 제제한 총지질 및 총인지질 인공세포막에 형성된 비대칭적 유동성에 미치는 비대칭적 영향)

  • Yun, Il;Kang, Jung-Sook
    • The Korean Journal of Pharmacology
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    • v.28 no.1
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    • pp.103-114
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    • 1992
  • Selective quenching of 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups was utilized to examine the transbilayer fluidity domains of the model membranes of total lipids (SPMVTL) and phospholipids (SPMVPL) extracted from synaptosomal plasma membrane vesicles. At $37^{\circ}C$, all anisotropy (r), limiting anisotropy $(r_{\infty})$, and order parameter (S) values of DPH in the SPMVTL were larger than those in SPMVPL. The anisotropy, limiting anisotropy, and order parameter of DPH in the inner monolayer were 0.025, 0.033, and 0.070, respectively, greater than calculated for the outer monolayer of SPMVTL. In SPMVPL, the anisotropy, limiting anisotropy, and order parameter of DPH in the inner monolayer were 0.014, 0.018, and 0.047, respectively, greater than calculated for the outer monolayer. Selective quenching of DPH by trinitrophenyl groups was also utilized to examine the effects of barbiturates on the transbilayer fluidity domains of SPMVTL and SPMVPL. Barbiturates did not affect the anisotropy of DPH in the transbilayer domains of SPMVTL. In contrast, barbiturates increased the fluorescence anisotropy, limiting anisotropy, and order parameter of DPH in the SPMVPL in a dose-dependent manner. Barbiturates showed a greater ordering effect on the outer monolayer as compared to the inner monolayer of SPMVPL. Hence, it has been demonstrated for the first time that the Sheetz-Singer hypothesis (1974) may be valid for phospholipid model membranes.

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Effect of Fatty Acid on the Membrane Fluidity of Liposomes (지방산 첨가가 리포좀 유동성에 미치는 영향에 관한 연구)

  • Lee, JinSun;Chi, Gyeong-Yup;Lim, JongChoo
    • Applied Chemistry for Engineering
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    • v.28 no.2
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    • pp.177-185
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    • 2017
  • In the present work, the interaction of fatty acid with vesicle membrane of phospholipids was investigated using 3 different kinds of fatty acids such as stearic acid (SA), oleic acid (OA) and linoleic acid (LA). Basically, the same trend has been found in 3 fatty acid systems. The addition of fatty acid produced a close packing of liposome due to the penetration of fatty acid molecules into liposome vesicles, which resulted in a decrease in size and an increase in zeta potential of liposome. However, excessive addition of fatty acid produced a transition from liposomes to aggregates of lipid particles having polymorphic structure. The membrane fluidity, characterized by measuring membrane deformability and fluorescence anisotropy ratio of liposomes, was in good agreement with measurement results of transmission electron microscopy (TEM) and particle size. The minimum size and closest packing of liposome with SA, OA and LA were found when the molar ratios of fatty acid to lecithin were 0.70, 0.50, and 0.25 respectively.

Membrane-Ordering Effects of Barbiturates on Pure Phospholipid Model Membranes

  • Knag, Jung-Sook;Chung, Young-Za;Cho, Goon-Jae;Byun, Won-Tan;Yun, Il
    • Archives of Pharmacal Research
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    • v.15 no.3
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    • pp.196-203
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    • 1992
  • Intramolecular excimer formation of 1, 3-di(1-pyrenyl)propane (Py-3-Py) and fluorescence polarization of 1, 6-diphenyl-1, 3, 5-hexatriene (DPH) were used to investigate the effects of barbiturates on the fluidity of model membranes of phosphatidycholine (SPMVPC), phosphatidylserine (SPMVPS), and phosphatidylinositol (SPMVPI) fractions of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex. In a dose-dependent manner, barbiturates decreased the excimer to monomer fluorescence intensity ratio (I'/I) of Py-3-Py and increased the anisotropy(r), rotational relaxation time (P), limiting anisotropy $(r_infty)$, and order parameter (S) of DPH in SPMVPC, SPMVPS and SPMVPI. This indicates that barbiturates decreased both the lateral and rotational diffusion of the probes in SPMVPC, SPMVPS and SPMVPI. The relative potencies of barbiturates in ordering the membranes were in the order: pentobarbital > hexobarbital > amobarbital > phenobarbital. This order correlates well with the anesthetic potencies of barbiturates and the potencies for enhancement of $\gamma$-aminobutyric acid-stimulated chloride uptake. Thus, it is strongly suggested that a close relationship might exist between the membrane ordering effects of barbiturates and the chloride fluxes across SPMV.

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Effects of Barbiturates on the Fluidity of Phosphatidylethanolamine Model Membranes (Barbiturates가 소의 신선한 대뇌피질 Synaptosomal Plasma Membrane Vesicles로 부터 추출하여 제제한 Phosphatidylethanolamine 인공세포막의 유동성에 미치는 영향)

  • Yun, Il;Kim, Hyung-Il;Hwang, Tae-Ho;Kim, Jong-Ryol;Kim, In-Se;Chung, Yong-Za;Shin, Yong-Hee;Jung, Hyun-Ok;Kang, Jung-Sook
    • The Korean Journal of Pharmacology
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    • v.26 no.2
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    • pp.209-217
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    • 1990
  • Intramolecular excimer formation with 1,3-di(1-pyrenyl)propane (Py-3-Py) and fluorescence polarization with 1,6-diphenyl-1,3,5-hexatriene (DPH) were used to evaluate the effects of barbiturates on the bulk fluidity of the model membranes of phosphatidylethanolamine fraction of synaptosomal plasma membrane vesicles (SPMVPE) isolated from bovine cerebral cortex. In the SPMVPE, barbiturates decreased the excimer to monomer fluorescence intensity ratio (I'/I) of Py-3-Py and increased the fluorescence polarization (P), anisotropy (r), limiting anisotropy $(r_{8})$, order parameter (S) and rotational relaxation time $({\bar{P}})$ of DPH in a dose-dependent manner. The relative potencies of barbiturates to order the SPMVPE were in the order: pentobarbital > hexobarbital > amobarbital > phenobarbital. Hence, it is concluded that barbiturates have ordering effects on the SPMVPE. And the membrane-ordering potencies of barbiturates appear to be correlated with the potencies for enhancement of GABA-stimulated chloride influx and with the anesthetic effects of barbiturates.

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Effect of Ganglioside $G_{M3}$ on the Erythrocyte Glucose Transporter (GLUT1): Conformational Changes Measured by Steady-State and Time-Resolved Fluorescence Spectroscopy

  • Yoon, Hae-Jung;Lee, Min-Yung;Jhon, GiI-Ja
    • BMB Reports
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    • v.30 no.4
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    • pp.240-245
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    • 1997
  • Interactions between ganglioside $G_{M3}$ and glucose transporter, GLUT1 were studied by measuring the effect of $G_{M3}$ on steady-state and time-resolved fluorescence of purified GLUT1 in synthetic lipids and on the 3-O-methylglucose uptake by human erythrocytes. The intrinsic tryptophan fluorescence showed a GLUT 1 emission maximum of 335 nm, and increased in the presence of $G_{M3}$ by 12% without shifting the emission maximum, The fluorescence lifetimes of intrinsic tryptophan on GLUT1 consisted of a long component of 7.8 ns and a short component of 2,3 ns and $G_{M3}$ increased both lifetime components. Lifetime components were quenched by acrylamide and KI. Acrylarnide-mduced quenching of long-lifetime components was partly recovered by $G_{M3}$ However. KI-induccd quenching of short- and long-lifetime components was not rescued by $G_{M3}$. The anisotropy of 1.6-diphenyl-1.3.5-hexatriene (DPH)-probed dimyristoylphosphatidylcholine (DMPC) model membrane was also increased with $G_{M3}$ incorporation, The transport rate of 3-O-methylglucose increased by 20% with $G_{M3}$ incorporation on the erythrocytes, Therefore, $G_{M3}$ altered the environment of lipid membrane and induced the conformational change of GLUT1.

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Differential Effects of Local Anesthetics on Rate of Rotational Mobility between Hydrocarbon Interior and Surface Region of Model Membrane Outer Monolayer

  • Chung, In-Kyo;Cha, Seong-Kweon;Chung, Yong-Za;Kim, Bong-Sun;Choi, Chang-Hwa;Cho, Goon-Jae;Jang, Hye-Ock;Yun, Il
    • The Korean Journal of Physiology and Pharmacology
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    • v.4 no.1
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    • pp.41-46
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    • 2000
  • Using fluorescence polarization of 12-(9-anthroyloxy)stearic acid (12-AS) and 2-(9-anthroyloxy)stearic acid (2-AS), we evaluated the differential effects of local anesthetics on 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 lipid fraction liposome extracted from synaptosomal plasma membrane vesicles. The anisotropy (r) values for the hydrocarbon interior and the surface region of the liposome outer monolayer were $0.078{\pm}0.001$ and $0.114{\pm}0.001,$ respectively. This means that the rate of rotational mobility in the hydrocarbon interior is faster than that of the surface region. In a dose-dependent manner, the local anesthetics 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.

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