polidocanol and dodecyloctaethyleneglycol-monoether

It has been proposed that intestinal sodium transport is mediated by two different active mechanisms: the ouabain-sensitive Na+/K(+)-ATPase and ouabain-insensitive Na(+)-ATPase. In order to determine the optimum conditions to solubilize the membrane-bound Na(+)-ATPase of enterocyte, basolateral plasma membranes were solubilized using different amounts of octyl glucoside (O.G), Tween 20, octaethylene glycol monododecyl ether (C12E8), and polyoxyethylene 9-lauryl ether (C12E9). Solubilized fractions were assayed for protein concentration and ATPase activity and characterized by electrophoresis analysis. Optimal solubilization of Na(+)-ATPase was obtained after mixing of 1 mg of basolateral plasma membrane with 1.5 mg of C12E9. Under these conditions, C12E9 solubilized over 60% membrane protein and Na(+)- and Na+/K(+)-ATPases activities were recovered over 80% in the soluble fraction without inactivation. In addition, when 25% glycerol and 2 mM ATP were added, the solubilized Na(+)-ATPase was stable after 3 days at 4 degrees C. The C12E9-solubilized Na(+)-ATPase presented the following kinetic characteristics: 1) is only stimulated by the Na+ salt, 2) K0.5 for Na+ = 4.62 +/- 0.06 mM, 3) is similarly stimulated by the Na+ salt of different anions, 4) optimal pH = 7.0, 5) inhibited by furosemide (IC50 = 0.52 +/- 0.10 nm). These kinetic properties of the solubilized Na(+)-ATPase were similar to those described to the native membrane-bound enzyme. This work reports for the first time, solubilization and characterization of a fully active and stable Na(+)-ATPase from basolateral plasma membranes of enterocyte using C12E9.

Topics: Animals; Cell Fractionation; Cell Membrane; Detergents; Electrophoresis, Polyacrylamide Gel; Epithelial Cells; Glucosides; Guinea Pigs; Intestinal Mucosa; Intestine, Small; Membrane Proteins; Ouabain; Polyethylene Glycols; Polysorbates; Sodium-Potassium-Exchanging ATPase; Solubility

The uptake, distribution, and clearance of three labeled nonionic surfactants, 14C-labeled dodecyl tetra(oxyethylene) ether [14C-C12-AE(4)], 14C-labeled dodecyl octa(oxyethylene) ether [14C-C12-AE(8)], and 14C-labeled dodecyl hexadeca(oxyethylene) ether [14C-C12-AE(16)] were investigated in carp (Cyprinus carpio) exposed to concentrations of 0.2-0.6 mg X liter-1, using whole-body autoradiography and the liquid scintillation counting method. 14C radioactivity was rapidly absorbed into the fish body and distributed in skin, nasal and oral cavities, gills, brain, hepatopancreas, kidney, gall bladder, and intestinal content at comparatively high concentrations. The calculated wet weight whole-body bioconcentration factor at steady state in the fish exposed to 14C-C12-AE(4), 14C-C12-AE(8), or 14C-C12-AE(16) for 72 hr was 310, 220, or 4.3, respectively. Clearance of 14C radioactivity from the fish body was rapid, with half-lives of 30-80 hr. The metabolites of 14C-C12-AE(4) were also examined in gills, blood, kidney, hepatopancreas, or gall bladder by thin-layer chromatography.

Topics: Animals; Autoradiography; Carps; Chromatography, Thin Layer; Cyprinidae; Polidocanol; Polyethylene Glycols; Software; Surface-Active Agents; Tissue Distribution