Krzysztof Szczepanowicz

Formation of biocompatible nanocapsules with emulsion core and pegylated shell by polyelectrolyte multilayer adsorption.

The aim of this work was to develop a novel method of preparation of loaded nanosize capsules based on liquid core encapsulation by biocompatible polyelectrolyte (PE) multilayer adsorption, with or without pegylated outermost layer. Using AOT (docusate sodium salt) as emulsifier, we obtained cores, stabilized by an AOT/PLL (poly-L-lysine hydrobromide) surface complex. These positively charged cores were encapsulated by layer-by-layer adsorption of polyelectrolytes, biocompatible polyanion PGA (poly-L-glutamic acid sodium salt), and biocompatible polycation PLL. We used the saturation method for formation of consecutive layers, and we determined the optimal conditions concerning concentration of surfactant and polyelectrolytes to form stable shells. The average size of the obtained capsules was 60 nm. Pegylated external layer were prepared using PGA-g-PEG (PGA grafted by PEG poly(ethylene glycol)). The capsules were stable for at least a period of 3 months. These nanocapsules were biocompatible when tested for cytotoxicity in a cellular coculture assay and demonstrated no or very low nonspecific binding to peripheral blood mononuclear cells when tested by flow cytometry. In order to study drug effects on leukemia cells, beta-carotene and vitamin A have been encapsulated as model drugs.

Novel approach to long sustained multilayer nanocapsules: influence of surfactant head groups and polyelectrolyte layer number on the release of hydrophobic compounds

Nanoemulsion-templated long sustained polyelectrolyte (PE) nanocapsules (average size < 200 nm) loaded with Oil Red O and two cyanine-type photosensitizers: IR-786 and IR-780 were successfully fabricated by using the layer-by-layer (LbL) technique. All nanoproducts were subjected to in vitro release characteristics and analysis of selected control parameters, i.e., type of surfactant head group, characteristic release time and surfactant–polyelectrolyte interactions. Their properties were characterized by means of dynamic light scattering (DLS) compared with scanning electron microscopy (SEM) and atomic force microscopy (AFM). In our studies for construction of oil-in-water nanoemulsion templates we selected three cationic surfactants with different nature of hydrophilic head groups, i.e., double-headed (or so-called dicephalic-type) N,N-bis[3,3′(trimethylammonio)propyl]dodecanamide dimethylsulfate (C12(TAPAMS)2), bulky saccharide-derived 2-(dodecyldimethylammonio)ethylglucoheptonamide bromide (D2GHA-12) and a classic dodecyltrimethylammonium bromide (DTABr) for comparison. The polyelectrolytes were the following: polyanion of poly(sodium 4-styrenesulfonate) (PSS) and polycation of poly(diallyldimethylammonium chloride) (PDADMAC). The in vitro release profile features, studied spectrophotometrically, were interpreted in the framework of diffusion-controlled processes and stability of the first interfacial PE–surfactant complex. Accordingly, the multicharge and bulky structure of the surfactant are found to be the most desirable factors for fabrication of long sustained and stable nanocapsules encapsulating a hydrophobic active substance.

Synthesis and antimicrobial activity of monodisperse copper nanoparticles.

Metallic monodisperse copper nanoparticles at a relatively high concentration (300 ppm CuNPs) have been synthesized by the reduction of copper salt with hydrazine in the aqueous SDS solution. The average particles size and the distribution size were characterized by Dynamic Light Scattering (DLS), Nanosight-Nanoparticle Tracking Analysis (NTA). The morphology and structure of nanoparticles were investigated using Scanning Electron Microscopy (SEM). The chemical composition of the copper nanoparticles was determined by X-ray Photoelectron Spectroscopy (XPS). Monodisperse copper nanoparticles with average diameter 50 nm were received. UV/vis absorption spectra confirmed the formation of the nanoparticles with the characteristic peak 550 nm. The antimicrobial studies showed that the copper nanoparticles had high activity against Gram-positive bacteria, standard and clinical strains, including methicillin-resistant Staphylococcus aureus, comparable to silver nanoparticles and some antibiotics. They also exhibited antifungal activity against Candida species.

Biocompatible long-sustained release oil-core polyelectrolyte nanocarriers: From controlling physical state and stability to biological impact

It has been generally expected that the most applicable drug delivery system (DDS) should be biodegradable, biocompatible and with incidental adverse effects. Among many micellar aggregates and their mediated polymeric systems, polyelectrolyte oil-core nanocarriers have been found to successfully encapsulate hydrophobic drugs in order to target cells and avoid drug degradation and toxicity as well as to improve drug efficacy, its stability, and better intracellular penetration. This paper reviews recent developments in the formation of polyelectrolyte oil-core nanocarriers by subsequent multilayer adsorption at micellar structures, their imaging, physical state and stability, drug encapsulation and applications, in vitro release profiles and in vitro biological evaluation (cellular uptake and internalization, biocompatibility). We summarize the recent results concerning polyelectrolyte/surfactant interactions at interfaces, fundamental to understand the mechanisms of formation of stable polyelectrolyte layered structures on liquid cores. The fabrication of emulsion droplets stabilized by synergetic surfactant/polyelectrolyte complexes, properties, and potential applications of each type of polyelectrolyte oil-core nanocarriers, including stealth nanocapsules with pegylated shell, are discussed and evaluated.

Encapsulation of liquid cores by layer-by-layer adsorption of polyelectrolytes.

The aim of this work was to develop the method of preparation of loaded, submicron nanocapsules based on the liquid core encapsulation by polyelectrolyte (PE) multilayer adsorption. The procedure of PE adsorption on the emulsion droplets requires a specific selection of surfactants, which have good properties as emulsifiers and provide a stable surface charge for sequential adsorption of PE without losing stability of emulsion. Using AOT as emulsifier this study obtained droplets, stabilized by AOT/PDADMAC surface complexes. These positively charged liquid cores were then modified by sequential adsorption of polyelectrolytes to obtain nanocapsules with the average size of 200 nm, with various combinations of polyelectrolytes (PDADMAC, CHIT, PAH, PSS, ALG). This study demonstrated the formation of consecutive layers of PE shells by measuring zeta potential of capsules after adsorption of each layer. It visualized the cores by dissolving fluorescent dye Coumarine6 in oil phase and multilayer shells by using FITC labelled polycation.

Polyelectrolyte multilayer capsules with quantum dots for biomedical applications.

The aim of this work was to encapsulate the CdTe quantum dots within the nanocapsules that were prepared by the layer-by-layer adsorption of polyelectrolytes. Two different polyelectrolyte pairs were used as components of the shell: synthetic polycation poly(allyamine hydrochloride) (PAH), together with anionic poly(sodium styrene sulfonate) (PSS), and biocompatible cationic poly-L-lysine hydrobromide in a pair with biocompatible anionic poly-D-glutamic acid sodium salt (PGA). The saturation method was used for formation of consecutive layers on the initial CdTe-polyelectrolyte complex. A growth of the polyelectrolyte shell was followed with the electrophoretic mobility and light scattering measurements, in order to determine the zeta potential and the size of capsules, respectively. The fluorescent spectra of the quantum dots, which are embedded within the capsules, were characterized with spectrofluorimeter. Later on, they were deposited on a negatively charged mica surface and studied by the means of atomic force microscopy (AFM). In order to estimate the cytotoxicity of capsules, their influence on the B-lymphoblastoid cell line proliferation and on unspecific binding to the P-blood mononuclear cells was examined using the flow cytometry.

Nanostructured multilayer polyelectrolyte films with silver nanoparticles as antibacterial coatings

Ultrathin polyelectrolyte films containing silver nanoparticles appear to be a promising material for antimicrobial coatings used in the medical area. The present work is focused on the formation of multilayer polyelectrolyte films using: polyethyleneimine (PEI) as polycation, Poly(sodium 4-styrenesulfonate) (PSS) as polyanions and negatively charged silver nanoparticles (AgNPs), which led to the polyelectrolyte-silver nanocomposite coatings. The film thickness and mass were measured by ellipsometry and quartz crystal microbalance with dissipation monitoring (QCM-D) and the structure and morphology of films were visualized using scanning electron microscopy (SEM). Systematic increase of the UV-Vis absorption confirmed formation of the consecutive layers of the film. The analysis of bacteria cell adhesion to films surface was done by the luminometry measurement. Three gram-negative bacterial strains with strong adhesive properties were used in this study: Escherichia coli, Aeromonas hydrophila, and Asaia lannenesis. It was found that nanocomposite films have antimicrobial properties, which makes them very interesting for a number of practical applications, e.g. for the prevention of microbial colonization on treated surfaces.

Biocompatible Polymeric Nanoparticles as Promising Candidates for Drug Delivery

The use of polymeric nanoparticles (NPs) in pharmacology provides many benefits because this approach can increase the efficacy and selectivity of active compounds. However, development of new nanocarriers requires better understanding of the interactions between NPs and the immune system, allowing for the optimization of NP properties for effective drug delivery. Therefore, in the present study, we focused on the investigation of the interactions between biocompatible polymeric NPs and a murine macrophage cell line (RAW 264.7) and a human monocytic leukemia cell line (THP-1). NPs based on a liquid core with polyelectrolyte shells were prepared by sequential adsorption of polyelectrolytes (LbL) using AOT (docusate sodium salt) as the emulsifier and the biocompatible polyelectrolytes polyanion PGA (poly-l-glutamic acid sodium salt) and polycation PLL (poly l-lysine). The average size of the obtained NPs was 80 nm. Pegylated external layers were prepared using PGA-g-PEG (PGA grafted by PEG poly(ethylene glycol)). The influence of the physicochemical properties of the NPs (charge, size, surface modification) on viability, phagocytosis potential, and endocytosis was studied. Internalization of NPs was determined by flow cytometry and confocal microscopy. Moreover, we evaluated whether addition of PEG chains downregulates particle uptake by phagocytic cells. The presented results confirm that the obtained PEG-grafted NPs are promising candidates for drug delivery.

Linseed oil based nanocapsules as delivery system for hydrophobic quantum dots.

In the present work, the CdSe/ZnS hydrophobic quantum dots were embedded within the polyelectrolyte nanocapsules. The core of the capsules, which consists of a mixture of the linseed oil with chloroform, was prepared using the spontaneous emulsification technique. The obtained emulsions were stabilized with lecithin and encapsulated using the layer-by-layer (LbL) adsorption of polyelectrolytes. The pair of biocompatible polyelectrolytes was used: the cationic poly-l-lysine hydrobromide (PLL) together with the anionic poly-d-glutamic acid sodium salt. The saturation LbL method, which is based on the stepwise formation of consecutive layers on the initial emulsion without the intermediate rinsing step, was applied to form the capsule shells. Their growth was evidenced by the capsule size and electrophoretic mobility measurements. The emulsion and the capsules were deposited on a mica surface and the deposit topology was examined by the means of atomic force microscopy (AFM). The presence of quantum dots within the oil cores was confirmed by recording the fluorescent spectra of the samples containing CdSe/ZnS. In order to evaluate cytotoxicity of the capsules, their influence on the viability of mouse embryonic fibroblasts was examined using the MTT test, followed by optical-microscope observation of morphology of the cells after hematoxylin-eosin staining.

In Vitro Interaction of Polyelectrolyte Nanocapsules with Model Cells

The nanocapsules based on a liquid core with polyelectrolyte shells prepared by the technique of sequential adsorption of polyelectrolytes (LbL) were investigated to verify capsules bioacceptance. Using AOT (docusate sodium salt) as emulsifier, we obtained liquid cores, stabilized by the interfacial complex AOT/PLL (poly-l-lysine hydrobromide). These liquid cores were encapsulated by sequential adsorption of polyelectrolytes using biocompatible polyanion PGA (poly-l-glutamic acid sodium salt) and biocompatible polycation PLL. The average size of the formed capsules was 60-80 nm. The influence of a number of polyelectrolytes layer in the shell (thickness of polyelectrolytes shell), surface charge, and capsule doses on cell viability was studied in a cellular coculture assay. In order to improve nanocapsules biocompatibility, the PEG-ylated external layers were prepared using PGA-g-PEG (PGA grafted by PEG poly(ethylene glycol)). For the most toxic nanocapsules (with only one polycation layer) about 90% of cells could survive when the concentration of nanocapsules was below 0.2 × 10(6) per one cell. That suggests that they use as a delivery vehicles is quite safe for living cells. Analysis of internalization of AOT(PLL/PGA)4-g-PEG in HEK 293 cells indicates that tested nanocapsules can easily penetrate cells membrane.