Cationic polyfluorenes for intracellular delivery of proteins.
ABSTRACT: Two cationic polyfluorene derivatives, quaternary amine 1 and guanidine 2 sheathed systems, were prepared as potential carriers to mediate import of proteins into cells without requiring covalent attachment to the protein. Neither polymer showed significant cytotoxicities (IC(50) 100 ?M) when exposed to Clone 9 rat liver cells. Both polymers were shown to mediate import of a series of four proteins chosen because they have different pI values, sizes, and variable organic fluor attachments. Once inside the cells, the quaternary amine system 1 released more of its cargo into regions outside the lysosomes. In one exploratory experiment, pyrenebutyrate was shown to accelerate import of a protein system by polymer 1.
Project description:Hybridbio/-synthetic sensory conjugated polymer nanoparticles (CPNs) are developed for selective label-free detection of target ssDNA in serum. Carboxylic acid-functionalized anionic polyfluorene nanoparticles are rationally designed as signal amplifying unit to bioconjugate with amine functionalized single stranded oligonucleotides as a receptor. The covalent DNA coating can signi?cantly improve the photostability of the DNA-bioconjugated CPNs over a wide range of buffer conditions. Better ssDNA discrimination for the DNA-bioconjugated CPNs sensor is achieved owing to increased interchain interactions and more efficient exciton transport in nanoparticles. The distinguishable ?uorescent color for DNA-bioconjugated CPNs in the presence of target ssDNA allows naked-eye detection of ssDNA under UV irradiation.
Project description:Herein we report the topochemical modification of polymer surfaces with perfluorinated aromatic azides. The aryl azides, which have quaternary amine or aldehyde functional groups, were linked to the surface of the polymer by UV irradiation. The polymer substrates used in this study were cyclic olefin copolymer and poly(methyl methacrylate). These substrates were characterized before and after modification using reflection-absorption infrared spectroscopy, sessile water contact angle measurements, and X-ray photoelectron spectroscopy. Analysis of the surface confirmed the presence of aromatic groups with aldehyde or quaternary amine functionality. Enzyme immobilization and patterning onto polymer surfaces were studied using confocal microscopy. Enzymatic digests of protein were carried out on modified probes manufactured from thermoplastic substrates, and the resulting peptide analysis was completed using matrix-assisted laser desorption/ionization mass spectrometry. The use of functionalized perfluorinated aromatic azides allows the surface chemistry of thermoplastics to be tailored for specific lab-on-a-chip applications.
Project description:Cationic, amphiphilic polymers are currently being used as antimicrobial agents that disrupt biomembranes, although their mechanisms remain poorly understood. Herein, membrane association and disruption by amphiphilic polymers bearing primary, tertiary, or quaternary ammonium salt groups reveal the role of cationic group structure in the polymer-membrane interaction. The dissociation constants of polymers to liposomes of POPC were obtained by a fluorometric assay, exploiting the environmental sensitivity of dansyl moieties in the polymer end groups. Dye leakage from liposomes and solid-state NMR provided further insights into the polymer-induced membrane disruption. Interestingly, the polymers with primary amine groups induced reorganization of the bilayer structure to align lipid headgroups perpendicular to the membrane. The results showed that polymers bearing primary amines exceed the tertiary and quaternary ammonium counterparts in membrane binding and disrupting abilities. This is likely due to enhanced complexation of primary amines to the phosphate groups in the lipids, through a combination of hydrogen bonding and electrostatic interactions.
Project description:Charged polymer brushes grafted to surfaces are of great interest for antibacterial, biosensor, nanofluidic, and drug delivery applications. In this paper, chitosans with quaternary ammonium salts, CH-Q, were immobilized on silicon oxide and characterized by in situ quartz-crystal microbalance with dissipation, QCM-D, and in situ spectroscopic ellipsometry, SE. Both methods showed that the hydrated film exhibited a minimum thickness of ~40 nm near pH 5 that increased strongly (up to ~80 nm) at lower and higher pH. This symmetric swelling is surprising because CH-Q is a cationic polymer. The CH-Q grafted layer was stable for pH values from 3 to 8 and exhibited rapid, reversible swelling and contraction upon varying pH. The CH-Q layer also reduced S. aureus colonization by a factor of ~30× compared to bare silicon oxide and an amine terminated silane grafted to silicon oxide. This antibacterial characteristic of CH-Q is attributed to the quaternary ammonium salts and the flexible polymer brush.
Project description:Functionalized, dendrimer-stabilized gold nanoparticles (Au DSNPs) are of scientific and technological interest for biological applications. In this work, we show that acetamide-functionalized Au DSNPs can be formed by acetylation of amine-terminated poly(amidoamine) (PAMAM) dendrimers of generation 5 (G5.NH(2)) complexed with Au(III) ions (AuCl(4) (-)). In addition, hydroxyl-functionalized Au DSNPs can be formed by simply mixing the glycidol hydroxyl-terminated G5 dendrimers (G5.NGlyOH) with HAuCl(4). In both cases, no additional reducing agents were needed and the reactions were completed at room temperature. We also show that Alexa Fluor 594 dye-functionalized Au DSNPs can be formed by acetylation of Alexa Fluor 594-conjugated, amine-terminated G5 dendrimers complexed with HAuCl(4). All of these functionalized Au DSNPs are water-soluble and stable. Fluorescence spectroscopy studies reveal that the Alexa Fluor 594-functionalized Au DSNPs retain similar fluorescence intensity to the Alexa Fluor 594-functionalized dendrimers that lack Au nanoparticles. These preparations of Au DSNPs provide a straightforward approach to synthesizing functionalized metal nanoparticles for biomedical applications.
Project description:A series of nine poly(2-deoxy-2-methacrylamido glucopyranose)-b-poly(methacrylate amine) diblock copolycations The cationic block was varied in length and in the degree of methyl group substitution (secondary, tertiary, quaternary) on the pendant amine in an effort to optimize the structure and activity for plasmid DNA delivery. Upon a thorough kinetic study of polymerization for each polymer, the glycopolymers were prepared with well-controlled Mn and Ð. The binding and colloidal stability of the polymer-pDNA nanocomplexes at different N/P ratios and in biological media has been investigated using gel electrophoresis and light scattering techniques. The toxicity and transfection efficiency of the polyplexes has been evaluated with Hep G2 (human liver hepatocellular carcinoma) cells; several polymers displayed excellent delivery and toxicity profiles justifying their further development for in vivo gene therapy.
Project description:Polymer/inorganic thermoelectric composites have witnessed rapid progress in recent years, but most of the studies have focused on the traditional conducting polymers. The limited structures of traditional conducting polymers restrain the development of organic thermoelectric composites. Herein, we report the preparation and thermoelectric properties of a series of composites films based on SWCNTs and bipyridine-containing polyfluorene derivatives. The value of the power factor around 12 μW m-1 K-2 was achieved for the composite F8bpy/SWCNTs with a mass ratio of 50/50, and the maximum value of 62.3 μW m-1 K-2 was obtained when the mass ratio reached 10/90. Moreover, taking advantage of the bipyridine unit could chelate various kinds of metal ions to form polymer complexes. The enhanced power factor of 87.3 μW m-1 K-2 was obtained for composite F8bpy-Ni/SWCNTs with a mass ratio of 50/50. Finally, the thermoelectric properties of the bipyridine-containing polyfluorene derivative/SWCNT composites were conveniently tuned by chelating with different metal ions.
Project description:Amyloid oligomers have emerged as a key neurotoxin in Alzheimer's dementia. Amyloid aggregation inhibitors and modulators have therefore offered potential applications in therapeutics and diagnosis. However, crossing the blood-brain barrier (BBB) and finding the toxic aggregates among aggregates of different sizes and shapes remain a challenge. The ability of identifying early aggregates can provide a new approach to find inhibitors of the initial nucleation events correlating presenile dementia. In this study, we have prepared polyfluorene nanoparticles using chitosan as an additive, which enables it to cross BBB efficiently and employed as a highly efficient amyloid oligomer modulator. The polymer conjugate, polyfluorene-chitosan (PC), shows no toxicity in MTT assay and precludes self-aggregation of A?1-40 and human cerebrospinal fluid oligomers to final fibril formation. This modulation strategy is supported by thioflavin T assay, circular dichroism studies, atomic force microscope images, and Fourier transform infrared analysis. The polymer-protein interface exhibits the presence of co-aggregates and responded with a stable optical response. The simple synthesis to get desired sizes and shapes with necessary photophysical behavior, biocompatibility, and most prominently BBB permeability makes this polymer conjugate very unique and highly attractive for modulation of amyloid oligomers selectively as well as for developing next generation nanotheranostic materials toward presenile dementia.
Project description:This work presents three new experimental methods for studying molecular imprinting. The electric conductivity measurements of the pre-polymerization mixture of amine templates in an aprotic solvent provide evidence of ionic dissociation of the pre-polymerization complexes. The displacement measurement of the template propranolol from its molecularly imprinted polymer (MIP) using a quaternary ammonium ion in toluene, shows that this MIP behaves as an ion exchanger even in a non-polar solvent. The same experiment also shows that template binding to the MIP from toluene involves ionic interaction. The third experimental method introduced here serves to study the models of template binding on MIPs. To this end the binding isotherm of propranolol (PR) has been measured on a polymer mixture consisting of non-imprinted control polymer (NIP) and a stronger binding acidic polymer, respectively. All three methods are suitable for studying several other imprinting systems.
Project description:The intracellular, cytosolic, delivery of quantum dots is an important goal for cellular imaging. Recently, a hydrophobic anion, pyrenebutyrate had been proposed to serve as a delivery agent for cationic quantum dots as characterized by confocal microscopy. Using an extracellular quantum dot quencher, QSY-21, as an alternative to confocal microscopy, we demonstrate that quantum dots remain on the cell surface and do not cross the plasma membrane following pyrenebutyrate treatment, a result that is confirmed with transmission electron microscopy. Pyrenebutyrate leads to increased cellular binding of quantum dots rather than intracellular delivery. These results characterize the use of QSY-21 as a quantum dot quencher and highlight the importance of the use of complementary techniques when using confocal microscopy.