RESUMO
The interaction of adriamycin with cardiolipin in model membranes and in various membrane preparations derived from rat liver mitochondria was studied and the results are analyzed in the light of a possible specific interaction between adriamycin and cardiolipin. It was found that adriamycin binds to cardiolipin-containing model membranes with a fixed stoichiometry of two drug molecules per cardiolipin. Furthermore, the extent of drug complexation by mitochondria and mitoplasts (inner membrane plus matrix) is in reasonable agreement with their cardiolipin content. In contrast, adriamycin-binding curves of inner membrane ghosts and submitochondrial particles reveal considerable association to an additional site, presumably RNA. The evidence for the potential importance of RNA as a target comes from experiments on outer membranes and microsomes which both appear to bind substantial amounts of adriamycin. Removal of the major part of the RNA associated with these fractions by EDTA treatment is accompanied by a dramatic reduction of binding capacity. We propose that endogenous RNA present in mitochondria and mitoplasts is not accessible for adriamycin at low concentrations of the drug due to the presence of an intact lipid barrier. This potential site comes to expression in ghosts and submitochondrial particles, due to the absence of an intact lipid bilayer and due to the inside-out orientation of the limiting membrane, respectively. Electron microscopical studies show that adriamycin induces dramatic changes in mitochondrial morphology, similar to the uncoupler-induced effects described by Knoll and Brdiczka (Biochim. Biophys. Acta 733, 102-110 (1983). Adriamycin has an uncoupling effect on mitochondrial respiration and oxidative phosphorylation. The concentration dependence of this effect correlates with the adriamycin-binding curve for mitochondria which implies that only bound adriamycin actively inhibits respiration.
Assuntos
Cardiolipinas/metabolismo , Doxorrubicina/metabolismo , Membranas Intracelulares/metabolismo , Membranas Artificiais , Mitocôndrias Hepáticas/metabolismo , Animais , Masculino , Fosforilação Oxidativa , Consumo de Oxigênio , Ratos , Ratos Endogâmicos , Difração de Raios XRESUMO
99mTc(Sn)-MDP complexes have been prepared by reduction of 99mTcO4- by Sn(II) in the presence of MDP. These complexes were separated on an anion exchange column. The necessity of the addition of the ligand and the reducing agent to the eluent to avoid decomposition during chromatography is demonstrated. For the main component that is found at pH 5 the ionic charge was calculated according to the method of Wilson and Pinkerton [Anal. Chem. 57,246 (1985)] and the method of Russell and Bishoff [Int. J. Appl. Radiat. Isot. 35,859 (1985)]. With the first method a charge of -4.2 +/- 0.3 was obtained, with the second a charge of -5.1 +/- 0.6. An accurate method to determine the void volume of the ion exchange column is described.
Assuntos
Medronato de Tecnécio Tc 99m , Cromatografia por Troca Iônica , Concentração de Íons de Hidrogênio , Indicadores e Reagentes , Conformação Molecular , Medronato de Tecnécio Tc 99m/síntese químicaRESUMO
When the ligand and the reducing agent are added to the eluent used in the separation of 99mTc-diphosphonate complexes by column chromatography, a significant increase in recovery is obtained. The addition of the ligand and the reducing agent also influences the shape of the chromatogram.
Assuntos
Cromatografia/métodos , Difosfonatos/isolamento & purificação , Compostos de Tecnécio , Tecnécio/isolamento & purificação , Estabilidade de MedicamentosRESUMO
99mTc(Sn)-EHDP complexes have been produced by reduction of TcO4- with Sn(II) in the presence of EHDP at varying pH and EHDP concentration. The mixture was separated by means of gel-chromatography with an eluent of the same composition and pH as the reaction mixture. It appears that at neutral pH larger complexes are formed than under acidic or basic reaction conditions. Larger complexes are also formed at higher EHDP concentrations.
Assuntos
Osso e Ossos/diagnóstico por imagem , Ácido Etidrônico , Compostos Organometálicos , Compostos de Organotecnécio , Cromatografia em Gel/métodos , Ácido Etidrônico/isolamento & purificação , Humanos , Concentração de Íons de Hidrogênio , Compostos Organometálicos/isolamento & purificação , CintilografiaRESUMO
The binding of 99mTc(Sn)-MDP to human serum albumin and other blood proteins was investigated by gel chromatography and ultrafiltration. During gel chromatography dissociation of the 99mTc(Sn)-MDP-protein complex occurs: thus, it is not a suitable technique for the determination of protein binding. The values found with ultrafiltration have to be corrected for non-ultrafiltrable TcO2.nH2O. From the corrected values it can be concluded that binding of 99mTc(Sn)-MDP to blood proteins does not play a role in the biodistribution.
Assuntos
Proteínas Sanguíneas/metabolismo , Albumina Sérica/metabolismo , Medronato de Tecnécio Tc 99m/metabolismo , Cromatografia em Gel , Humanos , UltrafiltraçãoRESUMO
The influence of several variables on the adsorption of 99mTc(Sn)-diphosphonate complexes on tri-calciumphosphate was determined. The composition of the incubation medium influenced the percentage adsorption: with Hank's balanced salt solution (a medium frequently used for bone cell cultures) and Tris buffer lower percentage adsorption was obtained than with physiological saline as the incubation medium. The influence of addition to the incubation medium of some ions and neutral species, some of which occur in bone fluid, is very specific. Addition of Sn(II) or Mg(II) (a component of HBSS) reduces the amount of adsorption. Addition of Ca(II) and Al(III) had no effect. Addition of sodium-citrate and MDP to the medium and an increase of the pH of the medium decreased the percentage adsorption. The ligand that was used in the preparation of the complex mixture influences the percentage adsorption considerably. The Sn(II) concentration used during the preparation of the 99mTc(Sn)-MDP and 99mTc(Sn)-MHDP complexes showed no definite influence on the percentage adsorption. The pH and ligand concentration, used in the preparation, however, did effect the percentage adsorption. It was concluded that the 99mTc(Sn)-disphosphonate mixtures are part reversibly and part irreversibly bound to tri-calcium-phosphate.