Synthesis of Polymeric Nanoparticles for biomedical delivery applications

Polymeric nanoparticles (PNPs) have attracted considerable interest over the last few years due to their unique properties and behaviors provided by their small size. Such materials could be used in a wide range of applications such as diagnostics and drug delivery. Advantages of PNPs include controlled release, protection of drug molecules and its specific targeting, with concomitant increasing of the therapeutic index. In this work, novel sucrose and cholic acid based PNPs were prepared from different polymers, namely polyethylene glycol (PEG), poly(D,L-lactic-co-glycolic acid) (PLGA) and PLGA-co-PEG copolymer. In these PNP carriers, cholic acid will act as a drug incorporation site and the carbohydrate as targeting moiety. The uptake of nanoparticles into cells usually involves endocytotic processes, which depend primarily on their size and surface characteristics. These properties can be tuned by the nanoparticle preparation method. Therefore, the nanoprecipitation and the emulsion-solvent evaporation method were applied to prepare the PNPs. The influence of various parameters, such as concentration of the starting solution, evaporation method and solvent properties on the nanoparticle size, size distribution and morphology were studied. The PNPs were characterized by using atomic force microscopy (AFM), scanning electron microscopy (SEM) and dynamic light scattering (DLS) to assess their size distribution and morphology. The PNPs obtained by nanoprecipitation ranged in size between 90 nm and 130 nm with a very low polydispersity index (PDI < 0.3). On the other hand, the PNPs produced by the emulsion-solvent evaporation method revealed particle sizes around 300 nm with a high PDI value. More detailed information was found in AFM and SEM images, which demonstrated that all these PNPs were regularly spherical. ζ-potential measurements were satisfactory and evidenced the importance of sucrose moiety on the polymeric system, which was responsible for the obtained negative surface charge, providing colloidal stability. The results of this study show that sucrose and cholic acid based polymeric conjugates can be successfully used to prepare PNPs with tunable physicochemical characteristics. In addition, it provides novel information about the materials used and the methods applied. It is hoped that this work will be useful for the development of novel carbohydrate based nanoparticles for biomedical applications, specifically for targeted drug delivery

Novel Unsaturated Sucrose Ethers and Their Application as Monomers

Novel unsaturated ethers were synthesised in good yields starting from sucrose,using a two-step mild and efficient procedure based on the Gassman method, whichconsists in forming a vinyl group by the elimination of ethanol from mixed acetals withtrimethylsilyl trifluoromethanesulfonate in the presence of alkyl amines. Mixed acetals arereadily obtained from the corresponding alcohols and ethyl vinyl ether, using an acidiccatalyst. Conventional etherification involving a primary halide was also examined. Themonomers thus obtained were successfully polymerised by a free radical mechanism,yielding unbranched linear and soluble polymers with pending sucrose moieties, and someof their physical properties were determined

Functional Group Coverage and Conversion Quantification in Nanostructured Silica by ¹H NMR

Silica nanostructured materials are important in many fields, including catalysis, imaging, and drug delivery, mainly due to the versatility of surface functionalization that can bestow a huge variety of chemical and physical properties. With most applications requiring precise control over this surface modification, characterization of surface composition and reactivity have become of extreme importance. We present a novel approach to track silica surface modification and quantify functional group coverage using only solution NMR. We test the method using different types of silica nanoparticles and surface modifications, to show that after dissolving the silica matrix, the ¹H NMR spectra can be resolved for every single component of the mixture. By using an internal standard, we are able to quantify the density of ligands and follow their sequential modification. Our work presents a fast, accurate, and straightforward method for surface characterization of silica nanostructures, using widely available NMR spectroscopy and small amounts of sample

Hybrid Mesoporous Nanoparticles for pH-Actuated Controlled Release

Among a variety of inorganic-based nanomaterials, mesoporous silica nanoparticles (MSNs) have several attractive features for application as a delivery system, due to their high surface areas, large pore volumes, uniform and tunable pore sizes, high mechanical stability, and a great diversity of surface functionalization options. We developed novel hybrid MSNs composed of a mesoporous silica nanostructure core and a pH-responsive polymer shell. The polymer shell was prepared by RAFT polymerization of 2-(diisopropylamino)ethyl methacrylate (pKa ~6.5), using a hybrid grafting approach. The hybrid nanoparticles have diameters of ca. 100 nm at pH &lt; 6.5 and ca. 60 nm at pH &gt; 6.5. An excellent control of cargo release is achieved by the combined effect of electrostatic interaction of the cargo with the charged silica and the extended cationic polymer chains at low pH, and the reduction of electrostatic attraction with a simultaneous collapse of the polymer chains to a globular conformation at higher pH. The system presents a very low (almost null) release rate at acidic pH values and a large release rate at basic pH, resulting from the squeezing-out effect of the coil-to-globule transition in the polymer shell

Silica nanoparticles with thermally activated delayed fluorescence for live cell imaging

Thermally activated delayed fluorescence (TADF) has revolutionized the field of organic light emitting diodes owing to the possibility of harvesting non-emissive triplet states and converting them in emissive singlet states. This mechanism generates a long-lived delayed fluorescence component which can also be used in sensing oxygen concentration, measuring local temperature, or on imaging. Despite this strong potential, only recently TADF has emerged as a powerful tool to develop metal-free long-lived luminescent probes for imaging and sensing. The application of TADF molecules in aqueous and/or biological media requires specific structural features that allow complexation with biomolecules or enable emission in the aggregated state, in order to retain the delayed fluorescence that is characteristic of these compounds. Herein we demonstrate a facile method that maintains the optical properties of solvated dyes by dispersing TADF molecules in nanoparticles. TADF dye-doped silica nanoparticles are prepared using a modified fluorescein fluorophore. However, the strategy can be used with many other TADF dyes. The covalent grafting of the TADF emitter into the inorganic matrix effectively preserves and transfers the optical properties of the free dye into the luminescent nanomaterials. Importantly, the silica matrix is efficient in shielding the dye from solvent polarity effects and increases delayed fluorescence lifetime. The prepared nanoparticles are effectively internalized by human cells, even at low incubation concentrations, localizing primarily in the cytosol, enabling fluorescence microscopy imaging at low dye concentrations

TADF Dye-Loaded Nanoparticles for Fluorescence Live-Cell Imaging

Thermally activated delayed fluorescence (TADF) molecules offer nowadays a powerful tool in the development of novel organic light emitting diodes due to their capability of harvesting energy from non-emissive triplet states without using heavy-metal complexes. TADF emitters have very small energy difference between the singlet and triplet excited states, which makes thermally activated reverse intersystem crossing from the triplet states back to the singlet manifold viable. This mechanism generates a long-lived delayed fluorescence component which can be explored in the sensing of oxygen concentration, local temperature, or used in time-gated optical cell-imaging, to suppress interference from autofluorescence and scattering. Despite this strong potential, until recently the application of TADF outside lighting devices has been hindered due to the low biocompatibility, low aqueous solubility and poor performance in polar media shown by the vast majority of TADF emitters. To achieve TADF luminescence in biological media, careful selection or design of emitters is required. Unfortunately, most TADF molecules are not emissive in polar media, thus complexation with biomolecules or the formation of emissive aggregate states is required, in order to retain the delayed fluorescence that is characteristic of these compounds. Herein, we demonstrate a facile method with great generalization potential that maintains the photophysical properties of solvated dyes by combining luminescent molecules with polymeric nanoparticles. Using an established swelling procedure, two known TADF emitters are loaded onto polystyrene nanoparticles to prepare TADF emitting nanomaterials able to be used in live-cell imaging. The obtained particles were characterized by optical spectroscopy and exhibited the desired TADF emission in aqueous media, due to the polymeric matrix shielding the dye from solvent polarity effects. The prepared nanoparticles were incubated with live human cancer cells and showed very low cytotoxicity and good cellular uptake, thus making fluorescence microscopy imaging possible at low dye concentrations