H2O2-induced decomposition of layer-by-layer films consisting of phenylboronic acid-bearing poly(allylamine) and poly(vinyl alcohol).

Layer-by-layer (LbL) films were prepared by an alternate deposition of phenylboronic acid-bearing poly(allylamine hydrochloride) (PBA-PAH) and poly(vinyl alcohol) (PVA) on the surface of a quartz slide to develop thin films that can be decomposed in response to hydrogen peroxide (H2O2). The PBA-PAH/PVA films decomposed in the presence of H2O2; the degree and kinetics of decomposition depend on the concentration of H2O2 and on the pH of the solution. For example, the film decomposition completely occurred in 30 min in 1.0 mM H2O2 solution at pH 7.4, while it took 180 min in 0.1 mM H2O2 solution. The H2O2-induced decomposition of the film can be explained in terms of the oxidative scission of the carbon-boron bond in PBA residues in the PBA-PAH chains. A potential use of the PBA-PAH/PVA films in developing H2O2-sensitive delivery systems was suggested.

Poly(lactic acid) microparticles coated with insulin-containing layer-by-layer films and their pH-dependent insulin release.

Poly(lactic acid) (PLA) microparticles were coated with layer-by-layer (LbL) films containing insulin and the pH-dependent release of insulin was studied. The LbL films were prepared on the surface of PLA microparticles by the alternate deposition of insulin and poly(allylamine hydrochloride) (PAH) through the electrostatic attraction between insulin and PAH. The insulin loading on the PLA microparticles depended on the film thickness, which corresponded to the number of insulin layers, and on the pH of the solution used to deposit insulin. The insulin loading increased with the film thickness and when the film was prepared at pH 7.4. The LbL films decomposed upon exposure to acidic solutions because the electrostatic attraction between the insulin and the PAH in the films disappeared when the charge on insulin changed from negative to positive at an acidic pH, which resulted in the release of insulin. The temperature and salt concentration did not affect the pH stability of the LbL films. The pH threshold for insulin release was pH 5.0-6.0, which corresponds to isoelectric point of insulin, 5.4. The release of insulin from the microparticles was rapid, and was almost complete within a few minutes. The circular dichroism spectra showed that the released insulin retained its original secondary structure. Our insulin-loaded PLA microparticles may be useful for the controlled release of insulin.

Dendrimers in layer-by-layer assemblies: synthesis and applications.

We review the synthesis of dendrimer-containing layer-by-layer (LbL) assemblies and their applications, including biosensing, controlled drug release, and bio-imaging. Dendrimers can be built into LbL films and microcapsules by alternating deposition of dendrimers and counter polymers on the surface of flat substrates and colloidal microparticles through electrostatic bonding, hydrogen bonding, covalent bonding, and biological affinity. Dendrimer-containing LbL assemblies have been used to construct biosensors, in which electron transfer mediators and metal nanoparticles are often coupled with dendrimers. Enzymes have been successfully immobilized on the surface of electrochemical and optical transducers by forming enzyme/dendrimer LbL multilayers. In this way, high-performance enzyme sensors are fabricated. In addition, dendrimer LbL films and microcapsules are useful for constructing drug delivery systems because dendrimers bind drugs to form inclusion complexes or the dendrimer surface is covalently modified with drugs. Magnetic resonance imaging of cancer cells by iron oxide nanoparticles coated with dendrimer LbL film is also discussed.

Design and fabrication of flexible DNA polymer cocoons to encapsulate live cells.

The capability to encapsulate designated live cells into a biologically and mechanically tunable polymer layer is in high demand. Here, an approach to weave functional DNA polymer cocoons has been proposed as an encapsulation method. By developing in situ DNA-oriented polymerization (isDOP), we demonstrate a localized, programmable, and biocompatible encapsulation approach to graft DNA polymers onto live cells. Further guided by two mutually aided enzymatic reactions, the grafted DNA polymers are assembled into DNA polymer cocoons at the cell surface. Therefore, the coating of bacteria, yeast, and mammalian cells has been achieved. The capabilities of this approach may offer significant opportunities to engineer cell surfaces and enable the precise manipulation of the encapsulated cells, such as encoding, handling, and sorting, for many biomedical applications.

Lactate-induced decomposition of layer-by-layer films composed of phenylboronic acid-modified poly(allylamine) and poly(vinyl alcohol) under extracellular tumor conditions.

Multilayer films that decompose in the presence of lactate were prepared by depositing phenylboronic acid-modified poly(allylamine) (PBA-PAH) and poly(vinyl alcohol) (PVA) on a lactate oxidase (LOx) layer. The layers adhered through boronate ester bonds. The resulting LOx(PBA-AH/PVA)10 film was stable in pH 7.4 solution but decomposed following the addition of lactate. The carbon-boron bonds in PBA residues were cleaved by oxidative reaction with H2O2 produced by the enzymatic reaction of LOx. Approximately 90% of the film decomposed following exposure for 120 and 30min to 0.05 and 20mM lactate at pH 7.4, respectively. The multilayer film therefore decomposed under conditions comparable to the extracellular environment of tumors (20mM lactate at pH 6.5). Our results show that LOx/(PBA-PAH/PVA)10 multilayer film could be used for cancer drug delivery systems.

Effects of hydrogen peroxide on the electrochemical decomposition of layer-by-layer thin films composed of 2-iminobiotin-labeled poly(ethyleneimine) and avidin.

The effects of hydrogen peroxide on the electrochemical decomposition of layer-by-layer thin films composed of 2-iminobiotin-labeled poly(ethyleneimine) (ib-PEI) and avidin were studied. An ib-PEI/avidin thin film prepared on the surface of a platinum (Pt) film-coated quartz resonator was electrochemically decomposed in the presence of hydrogen peroxide (H(2)O(2)) in the solution. The resonant frequency of the thin-film-deposited quartz resonator was increased upon application of electric potential (0.4-0.6 V vs Ag/AgCl) to the Pt layer, suggesting that the mass on the quartz resonator was decreased as a result of decomposition of the ib-PEI/avidin film. It was found that decomposition of the film is highly accelerated in the presence of H(2)O(2) compared to the decomposition in the same buffer solution without H(2)O(2), due to a pH change originating from electrochemical oxidation of H(2)O(2) on the Pt surface. The rate of electrochemical decomposition of the ib-PEI/avidin film was highly dependent on the concentration of H(2)O(2,) buffer capacity, and pH of the solution.

Sugar-sensitive dendrimer films as a sacrificial layer for the preparation of freestanding multilayer films.

Multilayer thin films composed of poly(vinyl alcohol) (PVA) and phenylboronic acid-bearing poly(amidoamine) dendrimer (PBA-PAMAM) were used as a sacrificial layer for constructing freestanding polyelectrolyte films consisting of poly(styrenesulfonate) (PSS) and poly(allylamine hydrochloride) (PAH). Freestanding (PSS/PAH)15 films were successfully released from substrate by exposing composite (PVA/PBA-PAMAM)n/(PSS/PAH)15 films (n=5 and 10) to sorbitol solutions under mild conditions at pH7.0-9.0. The film release was accelerated in solutions of higher sorbitol concentrations at pH9.0 as well as in solutions with lower concentration of NaCl. The results were rationalized based on the scission of boronate ester bonds between PBA-PAMAM and PVA in the (PVA/PBA-PAMAM)n layer due to a competitive binding of sorbitol to PBA-PAMAM.

pH- and sugar-sensitive multilayer films composed of phenylboronic acid (PBA)-modified poly(allylamine hydrochloride) (PBA-PAH) and poly(vinyl alcohol) (PVA): A significant effect of PBA content on the film stability.

Multilayer thin films composed of phenylboronic acid (PBA)-modified poly(allylamine hydrochloride) (PAH), PBA-PAH, with different PBA contents were prepared to study the effect of PBA content on the stability of the films. An alternate deposition of PBA-PAH and poly(vinyl alcohol) (PVA) on the surface of a quartz slide afforded multilayer films through forming boronate ester bonds between PBA-PAH and PVA. The 10-layered (PBA-PAH/PVA)10 films constructed using PBA-PAHs containing 16% and 26% PBA residues were stable in aqueous solutions over the range of pH 4.0-10.0, whereas the multilayer films composed of PBA-PAHs with 5.9% and 8.3% PBA decomposed at pH 8.0 or lower. The pH-sensitive decomposition of the films was rationalized based on the destabilization of the boronate ester bonds in neutral and acidic solutions. In addition, the (PBA-PAH/PVA)10 films decomposed in glucose and fructose solutions as a result of competitive binding of sugars to PBA-PAH in the films. The sugar response of the films depended on the PBA content in PBA-PAH. The (PBA-PAH/PVA)10 films consisting of 16% and 26% PBA-substituted PBA-PAHs are sensitive to physiological relevant level of glucose at pH7.4 while stable in glucose-free solution, suggesting a potential use of the films in constructing glucose-induced delivery systems.

Multilayer assembly of calf thymus DNA and poly(4-vinylpyridine) derivative bearing [Os(bpy)(2)Cl](2+): redox behavior within DNA film.

Polyelectrolyte multilayer (PEM) films containing polycationic osmium (Os) bipyridyl (bpy) complex-attached poly(4-vinylpyridine) (PVP) derivative [Os(bpy)(2)Cl](2+)-PVP (Os-PVP) and polyanionic calf thymus DNA (CT-DNA) on the surface of gold (Au) electrodes were prepared using a layer-by-layer self-assembly method, and their redox properties were studied. Os complex shows different redox behavior with CT-DNA film in comparison with other PEM film which is composed of ordinary polymers. A cyclic voltammetric study suggested that the outermost polyanionic DNA layer does not hinder the redox reaction of Os complex within the Os-PVP/CT-DNA multilayer film, which may be helpful to understand the electron transfer mechanism with the DNA film. For all the Os-complex-containing PEM layers studied, a diffusion-free electron transfer from the Os complex moieties in these films to the electrode surface was observed. An electrocatalytic oxidation of ascorbic acid (AA) by this DNA-containing PEM film-covered electrode was also proposed.

Sugar-sensitive thin films composed of concanavalin A and sugar-bearing polymers.

Layered thin films composed of concanavalin A (Con A) and sugar-bearing polymers were prepared by a layer-by-layer deposition of Con A and the polymer on a solid surface. The sugar-induced disintegration was studied. Con A-polymer layered films could be successfully prepared using a maltose-bearing polymer (PV-MA), while melibiose- and glucose-bearing polymers (PV-MEA and PV-G) did not afford a layered film, due to a weak affinity of PV-MEA and PV-G to Con A. The Con A/PV-MA layered film was stable in pH 7 and 8 solutions, while in a pH 6 medium the film was slightly unstable. The Con A/PV-MA film was disintegrated upon the addition of sugars in solution owing to a preferential binding of the sugars to the binding site of Con A in the film. The disintegration rate was dependent on the type of sugar and its concentration. The Con A/PV-MA film was disintegrated rapidly upon the addition of methyl alpha-D-mannopyranoside, while the rate was slower upon the addition of the same concentration of D-mannose, D-glucose and methyl alpha-D-glucopyranoside. The present system may be useful for constructing sensitive devices that can release a drug or other functional molecules in response to sugars.

Loading and release of fluorescent dye from layer-by-layer film-coated magnetic particles in response to hydrogen peroxide.

Polymer-coated magnetic particles (MPs) were prepared to study the binding of fluorescence dye on the surface and its H2O2-induced release. For this goal, multilayer films were prepared by layer-by-layer deposition of shikimic acid-appended poly(allylamine hydrochloride) (SA-PAH) and poly(styrenesulfonate) (PSS) on the surface of MPs. 3-(Dansylamino)phenylboronic acid (DPBA) was loaded on the MPs through boronate ester bonding between SA-PAH and DPBA. DPBA was released from the MPs in response to H2O2 as a result of breakage of the boronate ester bond by an oxidative reaction with H2O2. DPBA release was dependent on the H2O2 concentration. For example, 65% and 93% of the DPBA was released from (SA-PAH/PSS)4SA-PAH film-coated MPs in 30min after the addition of 0.1 and 0.5mM H2O2, respectively. In addition, the multilayer film-coated MPs were further modified by using glucose oxidase (GOx) to develop glucose-induced release systems. GOx-modified MPs released DPBA in response to 0.1mM d-glucose as a result of H2O2 generation through a GOx-catalyzed oxidation reaction of d-glucose. The results suggest a potential use of the multilayer film-coated MPs in the development of H2O2- and/or glucose-sensitive drug delivery systems.

Dual pH-sensitive layer-by-layer films containing amphoteric poly(diallylamine-co-maleic acid).

Layer-by-layer (LbL) thin films were prepared using an amphoteric copolymer consisting of alternating diallylamine and maleic acid monomer units (PDAMA), and pH-induced decomposition of the films was studied. LbL deposition of poly(styrene sulfonate) (PSS) and PDAMA on a solid surface afforded thin films at pH 3.0, while thin films did not form at pH 8.0. In contrast, PDAMA-poly(N-ethyl-4-vinylpyridine) (PEVP) films were constructed by LbL deposition of PDAMA and PEVP at pH 8.0, though films could not be obtained at pH 3.0. These results were explained based on the amphoteric nature of PDAMA, which is net positively and negatively charged at pH 3.0 and 8.0, respectively. PSS-PDAMA films were stable at pH 3.0-4.0 in a 150 mM NaCl solution and decomposed upon immersion in solutions with pH 4.3 or higher as a result of the reversal of the net PDAMA charge from positive to negative. In contrast, PDAMA-PEVP films disintegrated at pH 4.0 or lower. Thus, PDAMA-containing LbL films were found to be dual pH-sensitive: the films can be decomposed at acidic or neutral pH depending on the type of counter polymer in the film. PSS-PDAMA and PDAMA-PEVP films were also sensitive to salt concentration, where a higher concentration of NaCl in the solution in which the films were immersed facilitated decomposition of the films. The effect of salt concentration was attributed to partial breakage of the ion pairing between PDAMA and the counter polymers as a result of competitive binding with Na(+) and Cl(-) ions.

Preparation of poly(methyl methacrylate) microcapsules by in situ polymerization on the surface of calcium carbonate particles.

Poly(methyl methacrylate) (PMMA) microcapsules were prepared by the in situ polymerization of methyl methacrylate (MMA) and N,N'-methylenebisacrylamide on the surface of calcium carbonate (CaCO(3)) particles, followed by the dissolution of the CaCO(3) core in ethylenediaminetetraacetic acid solution. The microcapsules were characterized using fluorescence microscopy, atomic force microscopy, scanning electron microscopy, and Fourier transform infrared spectroscopy. The average sizes of the CaCO(3) particles and PMMA capsules were 3.8±0.6 and 4.0±0.6 µm, respectively. A copolymer consisting of MMA and rhodamine B-bearing MMA was also used to prepare microcapsules for fluorescent microscopy observations. Fluorescein isothiocyanate-labeled bovine serum albumin was enclosed in the PMMA microcapsules and its release properties were studied.

Insulin-containing layer-by-layer films deposited on poly(lactic acid) microbeads for pH-controlled release of insulin.

Layer-by-layer (LbL) thin films containing insulin were deposited on the surface of biodegradable poly(lactic acid) (PLA) microbeads and the pH-triggered release of insulin was studied. The LbL films were successfully prepared by the alternate deposition of insulin and poly(vinyl sulfate) (PVS) or dextran sulfate (DS) at pH 4.0 through the electrostatic force of attraction between positively charged insulin and polyanions. The loading of insulin on the microbeads was dependent on the number of insulin layers and the type of polyanions used; higher insulin loading was observed for thicker films and when PVS was used as the polyanion. Insulin was released from the microbeads when they were exposed to neutral solution (pH 7.4) due to a loss of electrostatic attraction between the insulin and polyanions in the films, which in turn was caused by the charge reversal of insulin from positive to negative in the neutral medium. The pH threshold for insulin release was found to be pH 5.0-6.0. The released insulin retained its original secondary structure as evidenced by circular dichroism spectra. The insulin loaded on the microbeads was satisfactorily stable even in the presence of a digestive enzyme (pepsin) at pH 1.5. These results suggest a potential future use for insulin-loaded microbeads in the oral delivery of insulin.

pH- and sugar-sensitive layer-by-layer films and microcapsules for drug delivery.

The present review provides an overview on the recent progress in the development of pH- and sugar-sensitive layer-by-layer (LbL) thin films and microcapsules in relation to their potential applications in drug delivery. pH-sensitive LbL films and microcapsules have been studied for the development of peptide and protein drug delivery systems to the gastrointestinal tract, anti-cancer drugs to tumor cells, anti-inflammatory drugs to inflamed tissues, and the intracellular delivery of DNA, where pH is shifted from neutral to acidic. pH-induced decomposition or permeability changes of LbL films and microcapsules form the basis for the pH-sensitive release of drugs. Sugar-sensitive LbL films and microcapsules have been studied mainly for the development of an artificial pancreas that can release insulin in response to the presence of glucose. Therefore, glucose oxidase, lectin, and phenylboronic acid have been used for the construction of glucose-sensitive LbL films and microcapsules. LbL film-coated islet cells are also candidates for an artificial pancreas. An artificial pancreas would make a significant contribution to improving the quality of life of diabetic patients by replacing repeated subcutaneous insulin injections.

Avidin/PSS membrane microcapsules with biotin-binding activity.

Polyelectrolyte microcapsules with avidin-poly(styrene sulfonate) (PSS) membrane were prepared by a layer-by-layer deposition technique. The uptake and release of biotin-labeled fluorescein (b-FITC) as well as immobilization of biotin-labeled glucose oxidase (b-GOx) to the microcapsule were studied. The polyelectrolyte microcapsules were prepared by coating the surface of calcium carbonate (CaCO(3)) microparticles with an avidin/PSS multilayer membrane, followed by dissolution of CaCO(3) core in an ethylenediaminetetraacetic acid solution. Inner and outer poly(allylamine)/PSS films were required to isolate the microcapsules, whereas microcapsules could not be formed without the support. The uptake of b-FITC into the microcapsule was highly enhanced through a strong binding of b-FITC to avidin as compared with the uptake of biotin-free FITC. Release of b-FITC from the microcapsule was accelerated upon addition of biotin due to a competitive binding of the added biotin to the binding site of avidin. Similarly, the surface of microcapsule was modified with b-GOx with retaining its catalytic activity.

Electrode potential-dependent colorimetric response of fluorescein-modified layer-by-layer films in the presence of hydrogen peroxide.

Layer-by-layer (LbL) thin films composed of fluorescein-modified poly(allylamine) (F-PAH) and poly(styrenesulfonic acid) (PSS) were prepared on the surface of an indium-tin oxide (ITO) electrode and the electrode potential-dependent colorimetric response of the LbL films was studied in the presence of hydrogen peroxide (H(2)O(2)). The LbL films were prepared by an alternate deposition of F-PAH and PSS on the surface through an electrostatic force of attraction. The LbL films exhibited a UV-visible absorption band around 500 nm originating from fluorescein residues in the film and the intensity of the absorption band depended on the pH of the solution to which the LbL film is exposed. The absorbance of the film was higher at neutral pH than that in weakly acidic solutions. The intensity of the absorption band decreased when an electrode potential higher than 0.6 V was applied in the presence of H(2)O(2), while virtually no response was observed at lower electrode potential. The colorimetric response was suppressed in solutions with higher buffer capacity. The results were rationalized on the basis of the changes in local pH at the vicinity of the electrode surface, which in turn was induced by electrolysis of H(2)O(2) on the electrode surface. A possible application of the system for colorimetric sensing of H(2)O(2) was discussed.

Electrochemically controlled release of alpha,beta,gamma,delta-tetrakis(4-N-methylpyridyl)porphine from layer-by-layer thin films.

The release of alpha,beta,gamma,delta-tetrakis(4-N-methylpyridyl)porphyrin (TMPyP) from layer-by-layer assembled thin films composed of poly(allylamine hydrochloride) (PAH) and poly(acrylic acid) (PAA) was electrochemically controlled. The release rate of TMPyP was enhanced when a positive electrode potential (+1.2 to +1.5 V) was applied to the glassy carbon electrode on which TMPyP-loaded PAH/PAA film was coated, while the effect of lower electrode potentials (0 to +1.0 V) was negligibly small. The effect of applied potential on the release rate was more significant at pH 7.4 than in the solutions of pH 8.0 and 9.0. The electrochemically enhanced release of TMPyP was rationalized based on local pH change in the vicinity of the electrode surface, which was induced by electrolysis of H(2)O.

Layer-by-layer assembled thin films composed of carboxyl-terminated poly(amidoamine) dendrimer as a pH-sensitive nano-device.

Dendrimer-containing multilayer thin films have successfully been prepared by a layer-by-layer deposition of carboxyl-terminated poly(amidoamine) dendrimer (PAMAM-COOH) and poly(methacrylic acid) (PMA) on a solid surface at pH 4.0, while the multilayer film did not form at pH 7.0. The PMA/PAMAM-COOH multilayer films prepared at pH 4.0 are decomposed at neutral pH due to electrostatic repulsion between negatively-charged carboxylate residues. The results suggest that the primary force for the successful deposition of PAMAM-COOH and PMA at pH 4.0 is hydrogen bonding between COOH residues on the surface of the dendrimer and PMA. The multilayer films are decomposed also at strongly acidic pH, suggesting an electrostatic force of attraction between the protonated tertiary amino groups in PAMAM-COOH and a small fraction of COO(-) residues in PMA contributes in part to the multilayer formation at pH 4.0. The PMA/PAMAM-COOH thin films can accommodate model dyes, Rose Bengal and 5,10,15,20-tetraphenyl-21H,23H-porphinetetrasulfonate, and the release can be controlled by changing pH.

Redox reactions of ferricyanide ions in layer-by-layer deposited polysaccharide films: a significant effect of the type of polycation in the films.

Redox reactions of ferricyanide ions, [Fe(CN)6]3-, in polysaccharide thin films that were prepared by layer-by-layer (LbL) deposition on the surface of a gold electrode were studied electrochemically by cyclic voltammetry. LbL films composed of alginic acid (AGA) and carboxymethylcellulose (CMC) were successfully prepared using poly(ethyleneimine) (PEI) and poly(diallyldimethylammonium chloride) (PDDA) as the cationic counterparts in the electrostatic LbL deposition. The deposition behavior of the PEI-based films significantly depended on the pH of the solutions from which the LbL films were deposited, while the effects of pH were negligibly small for the PDDA-based films due to the pH-independent positive charges on the PDDA chains. The cyclic voltammograms (CVs) of [Fe(CN)6]3- ions on the LbL film-coated electrodes revealed that all the LbL films tested are permeable to [Fe(CN)6]3- ions and that the redox reactions of [Fe(CN)6]3- ions proceed smoothly in the LbL polysaccharide films. It was found that [Fe(CN)6]3- ions are concentrated in the films from the bulk solution, depending on the pH of the medium and on the type of polycations in the film. The PEI-based films concentrated [Fe(CN)6]3- ions more effectively in an acidic solution than in neutral and basic media, while the pH effect was not observed for the PDDA-based films. In addition, we found that the [Fe(CN)6]3- ions are confined in the LbL films due to a strong binding of the ions to the positively charged sites arising from the protonated amino groups in the films. The confined [Fe(CN)6]3- ions exhibited redox reactions in the films, with the redox potentials being shifted to the positive or negative direction in the PEI- or PDDA-based film, respectively, as compared to the redox potential of diffusing [Fe(CN)6]3- ions. Thus, significant effects of the type of polycation in the LbL films on the redox reactions of [Fe(CN)6]3- ions were observed.