nystatin-a1 and fluorexon

The changes induced by biologically active substances in the permeability to K+ and calcein of liposomes composed of egg phosphatidylcholine and cholesterol were measured simultaneously in order to rapidly screen the sizes of pores formed in a membrane, using different sized markers. The substances examined in the present study were classified into three types based on differences in the rates at which K+ and calcein were released. The first type released only K+, and included gramicidin A. The second type predominantly released K+, preceding the release of calcein, and included amphotericin B and nystatin. The third type, including antimicrobial peptides, such as gramicidin S, alamethicin, and melittin, and several membrane-active drugs, like celecoxib (non-steroidal anti-inflammatory drug), 1-dodecylazacycloheptan-2-one (named azone; skin permeation enhancer), and chlorpromazine (tranquilizer), caused the release of K+ and calcein simultaneously. Thus, the sizes of pores formed in a liposomal membrane increased in the following order: types one, two, and three. We determined the size more precisely by conducting an osmotic protection experiment, measuring the release of calcein in the presence of osmotic protectants of different sizes. The radii of pores formed by the second type, amphotericin B and nystatin, were 0.36 - 0.46 nm, while the radii of pores formed by the third type were much larger, 0.63 - 0.67 nm or more. The permeability changes induced by substances of the third type are discussed in connection with a transient pore formed in a lipid packing mismatch taking place during the phase transition of dipalmitoylphosphatidylcholine liposomes.

Topics: Alamethicin; Amphotericin B; Azepines; Celecoxib; Chlorpromazine; Fluoresceins; Gramicidin; Liposomes; Melitten; Membranes, Artificial; Nystatin; Permeability; Potassium; Pyrazoles; Sulfonamides

The stability and spectral properties of nystatin-encapsulating liposomes, composed of various combinations of dipalmitoyl phosphatidylcholine (DPPC), cholesterol (CH) and distearoyl-N-(monomethoxy poly(ethylene glycol)succinyl) phosphatidylethanolamine (DSPE-PEG), were studied in order to elucidate the molecular state and localization of nystatin encapsulated in liposomes. Localization of nystatin at the surface region of the liposomal membrane was investigated by PEG/dextran two-phase partition and measurement of the fluorescence quenching of nystatin by p-xylene-bis-pyridinium bromide (DPX). In DPPC/DSPE-PEG liposomes and DPPC/CH/DSPE-PEG liposomes, containing 151 and 160 mcg nystatin per mg lipid, respectively, nystatin appeared to be present at the surface region of the liposomal membranes. Self-quenching of nystatin fluorescence was observed in DPPC/CH and DPPC/CH/DSPE-PEG liposomes even at low encapsulated amounts, suggesting the localization of nystatin in CH-incorporating membranes. In CH-free liposomes, nystatin molecules were at first delocalized in the membranes and then self-associated at a higher level of encapsulation. Absorption and circular dichroism (CD) spectra were also measured to examine the monomeric and aggregated states of nystatin in liposomes. High encapsulation efficacy was observed in DPPC and DPPC/DSPE-PEG liposomes, but the highest stability and retention of nystatin in liposomes were observed in DPPC/CH/DSPE-PEG liposomes, evaluated in terms of the nystatin and calcein release from nystatin-encapsulating liposomes in vitro. From the results, possible encapsulation mechanisms of nystatin in liposomes narrowed down to the following three points; interaction with lipid membrane, adsorption on the liposomal surface and complex formation with DSPE-PEG.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Algorithms; Cholesterol; Circular Dichroism; Drug Carriers; Drug Compounding; Fluoresceins; Fluorescence; Indicators and Reagents; Liposomes; Nystatin; Particle Size; Pharmaceutical Vehicles; Polyethylene Glycols; Surface Properties

Protocols to reconstitute channels into planar bilayers via fusion methods have now been developed. The greater the intravesicular pressures generated, the greater is the fusion. These pressures can be calculated exactly for any experimental configuration. For some of the configurations, adding nystatin channels to the vesicle membrane will greatly aid fusion. The configurations of the 1990 Method (Figs. 4 and 5) are optimal for fusing vesicles that are reconstituted with ion-selective channels to planar membranes. Greater binding, and ultimately greater fusion, is achieved by ejecting vesicles directly at the membrane rather than by simply adding material to the cis compartment.

Topics: Amphotericin B; Ergosterol; Fluoresceins; Fluorescent Dyes; Formamides; Indicators and Reagents; Ion Channels; Kinetics; Lipid Bilayers; Membrane Fusion; Methods; Models, Biological; Nystatin; Phospholipids; Thermodynamics