Fluorescent and paramagnetic chitosan nanoparticles that exhibit high magnetic resonance relaxivity: synthesis, characterization and in vitro studies.

We report water-in-oil (W/O) microemulsion synthesis of fluorescently bright and paramagnetically strong bimodal chitosan nanoparticles (BCNPs). The W/O microemulsion system provides a confined environment for producing monodispersed BCNPs. Average particle size as estimated by the Transmission Electron Microscopy was 28 nm. The water to surfactant molar ratio of 22 produced small size fairly monodispersed BCNPs. Fluorescein isothiocyanate (FITC, a fluorescent dye) and Gd-DOTA (a paramagnetic Gd ion chelating agent) were covalently attached to chitosan polymer backbone prior to BCNP synthesis. The purpose of the covalent attachment of fluorescent and paramagnetic labels to chitosan is to prevent leakage of these labels from the BCNPs. The BCNPs were cross-linked with tartaric acid using water-soluble carbodiimide coupling chemistry in order to maintain particulate integrity. Zeta potential value of +27.6 mV confirmed positive surface charge of cross-linked BCNPs. Fluorescence excitation and emission spectra of BCNPs were similar to that of bare FITC spectra, showing characteristic 520 nm emission at the 490 excitation. Paramagnetic gadolinium ion (Gd3+) concentration in the BCNPs was determined by inductively coupled plasma (ICP) emission spectroscopy. The longitudinal (T1) and transverse (T2) proton relaxation times were determined as a function of Gd3+ concentration in the BCNPs at 4.7 Tesla. Proton relaxivity (R1 value) of BCNPs was calculated to be 41.1 mM Gd(-1)s(-1). The reported R1 value of Gd-DOTA chelates is however 5.8 mM Gd(-1)s(-1). High proton relaxivity of BCNPs is attributed to hydrated chitosan environment around Gd chelates which additionally contributed to overall water exchange process. To demonstrate in vitro bioimaging capability, J774 macrophage cells were incubated with BCNPs. Confocal images clearly showed BCNP uptake by J774 cells. Internalization of BCNPs was confirmed by co-labeling J774 cells with a red-emitting membrane dye. BCNP green emission was mostly observed from middle of cells and within the red-emitting membrane boundary.

Nanobioimaging and sensing of infectious diseases.

New methods to identify trace amount of infectious pathogens rapidly, accurately and with high sensitivity are in constant demand to prevent epidemics and loss of lives. Early detection of these pathogens to prevent, treat and contain the spread of infections is crucial. Therefore, there is a need and urgency for sensitive, specific, accurate, easy-to-use diagnostic tests. Versatile biofunctionalized engineered nanomaterials are proving to be promising in meeting these needs in diagnosing the pathogens in food, blood and clinical samples. The unique optical and magnetic properties of the nanoscale materials have been put to use for the diagnostics. In this review, we focus on the developments of the fluorescent nanoparticles, metallic nanostructures and superparamagnetic nanoparticles for bioimaging and detection of infectious microorganisms. The various nanodiagnostic assays developed to image, detect and capture infectious virus and bacteria in solutions, food or biological samples in vitro and in vivo are presented and their relevance to developing countries is discussed.

Ultra-small water-dispersible fluorescent chitosan nanoparticles: synthesis, characterization and specific targeting.

A robust water-in-oil microemulsion method of making water-dispersible ultra-small (<30 nm) size fluorescent chitosan nanoparticles is reported for the first time and specific targeting of these FCNPs to human leukemia cells via aptamer recognition is demonstrated.

Silica-based multimodal/multifunctional nanoparticles for bioimaging and biosensing applications.

In the last decade, the field of nanoparticle (NP) technology has attracted immense interest in bioimaging and biosensing research. This technology has demonstrated its capability in obtaining sensitive data in a noninvasive manner, promising a breakthrough in early-stage cancer diagnosis, stem cell tracking, drug delivery, pathogen detection and gene delivery in the near future. However, successful and wide application of this technology relies greatly on robust NP engineering and synthesis methodologies. The NP development steps involve design, synthesis, surface modification and bioconjugation. Each of these steps is critical in determining the overall performance of NPs. It is desirable to obtain NPs that are highly sensitive, stable, imageable, biocompatible and targetable. It is also desirable to obtain multimodal/multifunctional NPs that will enable imaging/sensing of the target using multiple imaging/sensing modalities. In this review, we focus on silica NPs that have been developed for biosensing applications and silica-based multimodal/multifunctional NPs for bioimaging applications.

Antimicrobial activity and local release characteristics of chlorhexidine diacetate loaded within the dental copolymer matrix, ethylene vinyl acetate.

In vitro results are presented for a novel oral drug-delivery system ultimately intended for treatment of oral infections in immunocompromised patients. Test samples of ethylene vinyl acetate copolymer (EVA) containing chlorhexidine diacetate (CDA) showed desirable antimicrobial properties and steady, slow release into aqueous and other media after an initial burst of drug release in the first day of liquid exposure. By washing away this initial burst, the proposed mouthguard device should be capable of sustained delivery of locally effective CDA concentrations far below systemically toxic levels. A prolonged room temperature shelf-life of at least 1 year, and effectivity against a wide range of oral bacteria and Candida species was demonstrated. Drug loaded films showed a top-to-bottom asymmetry in drug release, but good lateral homogeneity, and a linear relationship between initial CDA loading concentration (from 0.63 to 10 wt %) and days 3-14 release rates in a static aqueous environment. The EVA matrix containing CDA appears to possess many suitable properties for localized oral delivery of sustained antimicrobial activity.

Release of antimicrobial and antiviral drugs from methacrylate copolymer system: effect of copolymer molecular weight and drug loading on drug release.

OBJECTIVE: To study the release of antiviral drug acyclovir (ACY) and antibacterial drug chlorhexidine diacetate (CDA) from synthesized copolymers of ethyl methacrylate and hexyl methacrylate of different molecular weights. The effect of the copolymer molecular weight and the effect of drug loading into the copolymer on the release of the drugs have been studied. METHOD: Copolymers (I-IV) of ethyl methacrylate (EMA) and hexyl methacrylate (HMA) were synthesized by free radical solution polymerization with a yield of 76-82%. The copolymer composition was determined by proton NMR spectroscopy. The molecular weight of the copolymers was determined by gel permeation chromatography (GPC). Copolymers I and II were of higher molecular weight while copolymers III and IV were of lower molecular weight. The copolymers were impregnated with 2.5 wt.% of ACY and CDA individually and the release rate of these drugs in water at 37 degrees C was examined. Drug loading was studied with 2.5, 5.0 and 7.5 wt.% of ACY and CDA incorporated into a separate polymer. RESULTS: Measurements of the in vitro rate of drug release showed a sustained release of drug over extended period of time. ACY release rate increases with decrease in copolymer molecular weight from copolymers I to IV (p<0.001) while CDA showed a different release profile. CDA release rate was higher from higher molecular weight copolymers I and II than from lower molecular weight copolymers III and IV (p<0.001). ACY release rate increases steadily with increase in drug load (p=0.011) while CDA release rate had a leveling effect with increase in drug load. SIGNIFICANCE: Varying the copolymer molecular weight as well as the drug concentration alters the drug release rates and thus it is possible to control the drug release rates to a desired value.

Poly(ethylene-co-vinyl acetate) copolymer matrix for delivery of chlorhexidine and acyclovir drugs for use in the oral environment: effect of drug combination, copolymer composition and coating on the drug release rate.

OBJECTIVES: This study utilizes a bio-compatible ethylene vinyl acetate (EVA) copolymer to deliver drugs at therapeutic levels over extended periods of time. The release rate of an anti-fungal and an anti-microbial drug namely acyclovir (ACY) and chlorhexidine diacetate (CDA) from EVA was investigated individually and as a mixture. The effect of drug combination, the composition of the copolymer and the coating of the matrix with a different polymer on the rate of drug release are presented. METHOD: Polymer casting solutions were prepared by homogeneously dissolving EVA copolymer and the drugs in the ratio (40:1) in dichloromethane. The drugs ACY and CDA were used individually as well as in three different weight ratios maintaining the total drug concentration in the polymer at 2.5%. Different concentrations of vinyl acetate (VA) 28, 32 and 40% in the EVA matrix were used to study the release of either ACY or CDA alone while 40% VA was used for the release study of the individual drug as well as their mixtures. Thin square films of 3cmx3cm with a thickness of 0.7mm were cut from the dry sheet obtained by solvent evaporation. Coated films were prepared by dipping ACY and CDA drug-loaded EVA films (VA 40%) into EVA copolymer of VA 32% and then dried. All of the drug-loaded samples were extracted at 37 degrees C in 10ml distilled water that was replaced daily. The rate of individual drug release was measured by UV-spectrophotometer while the mixtures of drugs were measured by high performance liquid chromatography (HPLC). RESULTS: The release rate of ACY is higher than that of CDA both individually and in the ACY/CDA 50/50 mixture. In the other mixtures, the release of the drug is proportional to its concentration in the mixture. Total release of ACY is higher than CDA in most compositions. The effect of increasing the vinyl acetate content of the EVA matrix increased the drug release rate (p=0.02) while coating of films resulted in a decrease of the release rate of the drugs. SIGNIFICANCE: Measurements of the in vitro rate of drug release showed that there was a sustained release of drug at an almost constant concentration over extended period of time, thus providing a basis for oral treatment modality. We show that it is possible to alter the rate of drug release in the EVA matrix to a desired value by: (1) changing the composition of the EVA copolymer, (2) altering the mixtures of drugs and (3) coating the matrix with additional polymer. The use of mixtures of drugs that can enhance or decrease the rate of drug release may prove more effective in treating persistent oral infections in immunocompromised patients.

Effects of solubilizing surfactants and loading of antiviral, antimicrobial, and antifungal drugs on their release rates from ethylene vinyl acetate copolymer.

OBJECTIVES: This study investigates the effects of surfactants and drug loading on the drug release rate from ethylene vinyl acetate (EVA) copolymer. The release rate of nystatin from EVA was studied with addition of non-ionic surfactants Tween 60 and Cremophor RH 40. In addition, the effect of increasing drug load on the release rates of nystatin, chlorhexidine diacetate and acyclovir is also presented. METHOD: Polymer casting solutions were prepared by stirring EVA copolymer and nystatin (2.5wt.%) in dichloromethane. Nystatin and surfactants were added in ratios of (1:1), (1:2) and (1:3). Drug loading was studied with 2.5, 5.0, 7.5, and 10.0wt.% proportions of nystatin, chlorhexidine diacetate and acyclovir incorporated into a separate polymer. Three drug loaded polymer square films (3cmx3cmx0.08cm) were cut from dry films to follow the kinetics of drug release at 37 degrees C. Ten milliliters of either distilled water or PBS was used as the extracting medium that was replaced daily. PBS was used for nystatin release with addition of surfactants and water was used for the study on drug loading and surfactant release. The rate of drug release was measured by UV-spectrophotometer. The amount of surfactant released was determined by HPLC. RESULTS: The release of nystatin was low in PBS and its release rate increased with the addition of surfactants. Also, increasing surfactant concentrations resulted in increased drug release rates. The release rates of chlorhexidine diacetate (p<0.0001), acyclovir (p<0.0003) and nystatin (p<0.0017) linearly increased with increasing drug loads. The amount of surfactants released was above the CMC. SIGNIFICANCE: This study demonstrates that the three therapeutic agents show a sustained rate of drug release from EVA copolymer over extended periods of time. Nystatin release in PBS is low owing to its poor solubility. Its release rate is enhanced by addition of surfactants and increasing the drug load as well.

Influence of additives on the properties of Bis-GMA/Bis-GMA analog comonomers and corresponding copolymers.

OBJECTIVES: The purpose of this study was to evaluate the effect of two additives: propionaldehyde (aldehyde) and 2,3-butanedione (diketone) on the properties of Bis-GMA diluted with TEGDMA and the synthesized Bis-GMA analogs, propoxylated Bis-GMA (CH(3)Bis-GMA) and propoxylated fluorinated Bis-GMA (CF(3)Bis-GMA). METHODS: Nine experimental comonomers were prepared combining Bis-GMA and TEGDMA, CH(3)Bis-GMA, CF(3)Bis-GMA, with aldehyde (32mol%) and diketone (32mol%). Photopolymerization was effected by using Camphorquinone (0.2wt%) and N,N-dimethyl-p-toluidine (0.2wt%). Experimental comonomer viscosity (Brookfield viscometer), polymerization shrinkage (gravimetrically), degree of conversion (FT-IR) and contact angles (contact angle goniometer) were determined. Comonomer and copolymer T(g)s (DSC and Fox equation) were also evaluated. Data were analyzed by one-way ANOVA and Tukey test (alpha=0.05). RESULTS: Bis-GMA/CH(3)Bis-GMA and Bis-GMA/CF(3)Bis-GMA with additives exhibited lower viscosities (p<0.01). Inclusion of additives into the comonomer systems did not produce significant increase in polymerization shrinkage (p>0.05). A significant increase in degree of conversion was shown for Bis-GMA/TEGDMA and Bis-GMA/CH(3)Bis-GMA with additives (p<0.01). Additives reduced contact angle and comonomer T(g) values, whereas the corresponding copolymers with additives showed an increase in T(g). SIGNIFICANCE: Use of novel comonomer systems with the addition of aldehyde and diketone functional groups would improve dental resin composite properties.