RESUMEN
This study focused on defining the in vitro behavior of amphiphilic poly-N-vinylpyrrolidone (Amph-PVP) nanoparticles toward whole blood, blood plasma and blood cells in order to assess nanoparticle blood compatibility. In addition, possible effects on endothelium cell growth/viability were evaluated. The Amph-PVP nanoparticles were formed via self-assembling in aqueous media and composed of a hydrophobic alkyl core and a hydrophilic PVP outer shell. Their blood compatibility was evaluated by investigating their effect on red blood cells (RBCs) or erythrocytes, white blood cells (WBCs) or leukocytes, platelets (PLTs) and on complement system activation. Our results clearly demonstrate that the Amph-PVP nanoparticles are stable in presence of blood serum, have no significant effects on the function of RBCs, WBCs, PLTs and complement system activation. The Amph-PVP nanoparticles did not show considerable hemolytic or inflammatory effect, neither influence on platelet aggregation, coagulation process, or complement activation at the tested concentration range of 0.05-0.5â¯mg/ml. The Amph-PVP nanoparticles did not exhibit any significant effect on HMEC-1 microvascular skin endothelial cells' growth in in vitro experiments. The excellent blood compatibility of the Amph-PVP nanoparticles and the lack of effect on endothelium cell growth/viability represent a crucial feature dictating their further study as novel drug delivery systems.
Asunto(s)
Materiales Biocompatibles , Plaquetas/efectos de los fármacos , Eritrocitos/efectos de los fármacos , Leucocitos/efectos de los fármacos , Nanopartículas/toxicidad , Pirrolidinonas/toxicidad , Línea Celular , Células Endoteliales/efectos de los fármacos , Endotelio Vascular/efectos de los fármacos , Humanos , Técnicas In Vitro , Nanopartículas/química , Pirrolidinonas/química , Piel/irrigación sanguínea , Piel/citologíaRESUMEN
Novel amphiphilic poly-N-vinylpyrrolidone derivatives with different molecular weight of hydrophilic PVP fragment and one secondary di-n-alkyl terminal hydrophobic fragment of different length were synthesized to compare their inclination for formation of nano-scaled micelle-like aggregates in aqueous media with previously studied primary n-alkyl terminated poly-N-vinylpyrrolidones. The behavior of amphiphilies in water solutions was studied and critical aggregation concentration values for prepared polymer samples were determined by fluorescence spectroscopy and compared with those for primary n-alkyl derivatives. Polymeric micelle-like particles with or without encapsulated drug were prepared using dialysis or solvent evaporation techniques. Indomethacin was incorporated into hydrophobic inner core of these nanoparticles as a typical model drug. Dynamic light-scattering studies determined that the average size of particles formed was from 90 nm up to 600 nm with monodisperse size distribution and the nanoparticle size slightly increased with the amount of indomethacin encapsulated into inner core of the particles. In vitro release experiments carried out at different medium pH values using indomethacin-loaded nanoparticles exhibited slow and steady drug release into the medium.
Asunto(s)
Portadores de Fármacos/química , Indometacina/farmacocinética , Nanopartículas/química , Pirrolidinonas/química , Antiinflamatorios no Esteroideos/química , Antiinflamatorios no Esteroideos/farmacocinética , Composición de Medicamentos , Concentración de Iones de Hidrógeno , Interacciones Hidrofóbicas e Hidrofílicas , Indometacina/química , Cinética , Lípidos/química , Microscopía Electrónica de Transmisión , Nanopartículas/ultraestructura , Tamaño de la Partícula , Temperatura , Agua/químicaRESUMEN
Certain amphiphilic water-soluble polymers including amphiphilic derivatives of polyvinyl pyrrolidone (PVP) were found to be efficient steric protectors for liposomes in vivo. In this study, we have tried to develop synthetic pathways for preparing amphiphilic PVP and to investigate the influence of the hydrophilic/hydrophobic blocks on some properties of resulting polymers and polymer-coated liposomes. To prepare amphiphilic PVP with the end stearyl (S) or palmityl (P) residues, amino- and carboxy-terminated PVP derivatives were first synthesized by the free-radical polymerization of vinyl pyrrolidone in the presence of amino- or carboxy-mercaptans as chain transfer agents, and then modified by interaction of amino-PVP with stearoyl chloride or palmitoyl chloride, or by dicyclohexyl carbodiimide coupling of stearylamine with carboxy-PVP. ESR-spectra of the hydrophobic spin-probe, nitroxyl radical N-oxyl-2-hexyl-2-(10-methoxycarbonyl)decyl-4,4'-dimethyl oxazoline, in the presence of amphiphilic PVP demonstrated good accessibility of terminal P- and S-groups for the interaction with other hydrophobic ligands. Spontaneous micellization and low CMC values (in a low micromolar range) were found for amphiphilic PVP derivatives using the pyrene method. In general, S-PVP forms more stable micelles than P-PVP (at similar MW, CMC values for S-PVP are lower than for P-PVP). It was found that amphiphilic PVP incorporated into negatively charged liposomes effectively prevents polycation(poly-ethylpyridinium-4-vinylchloride)-induced liposome aggregation, completely abolishing it at ca. 10 mol% polymer content in liposomes. Additionally, the liposome-incorporated PVP prevents the fluorescence quenching of the membrane-incorporated hydrophobic fluorescent label [N-(4-fluoresceinthiocarbamoyl)dipalmitoyl-PE] by the free polycation. PVP-modified liposomes were loaded with a self-quenching concentration of carboxyfluorescein, and their destabilization in the presence of mouse serum was investigated following the release of free dye. Amphiphilic PVP with MW between 1,500 and 8,000 provides good steric protection for liposomes. The degree of this protection depends on both polymer concentration and molecular size of the PVP block.
Asunto(s)
Materiales Biocompatibles/síntesis química , Povidona/síntesis química , Animales , Materiales Biocompatibles/química , Portadores de Fármacos , Estabilidad de Medicamentos , Espectroscopía de Resonancia por Spin del Electrón , Colorantes Fluorescentes , Técnicas In Vitro , Liposomas , Ensayo de Materiales , Ratones , Micelas , Povidona/química , Electricidad Estática , Propiedades de Superficie , Tensoactivos/síntesis química , Tensoactivos/químicaRESUMEN
We have used resonance Raman spectroscopy and isotopic labeling techniques to unambiguously assign the dioxygen stretching frequency (vo-o) in the substrate-bound oxygenated complex of cytochrome P-450cam. The frequency found for Vo-o in the P-450cam system (1140 cm-1) is in remarkable agreement with recent studies of thiolate heme model compounds. The general features of the oxy-P-450cam Raman spectra are tabulated and comparisons are made with the oxy complexes of hemoglobin, myoglobin, and various model compounds. Most of the results are qualitatively explained by consideration of electron donation into the pi g (O2)/d pi (M) orbitals of the oxygenated complex (M = Fe or Co). It is also noted that the effect of the "extra" electron in the nitrogen base Co(II) oxy complexes, in some ways, parallels the effect of the lone pair electrons of thiolate in the oxy-P-450cam complex. This is evidenced by the enhanced resonance Raman activity of vo-o in both the Co(II) and P-450 systems as well as by the similarity of the vo-o frequencies.