Chitosan Derivative Nanoparticles Research Articles

Chitosan-derivative nanoparticles loaded with dihydromyricetin: Characterization, antibacterial and antioxidant activities

Bovine mastitis is a global problem and is mainly treated with antimicrobial agents that can significantly affect milk quality and threaten human health. The mammary gland can be damaged by the accumulation of reactive oxygen species (ROS), particularly during the perinatal period. Dihydromyricetin (DHM) has a wide range of pharmacological activities and is considered safe. However, its poor solubility, absorption, and bioavailability have limited its application. To address this issue, we designed and synthesized new chitosan-derivative nanoparticles (HACC-NPs) loaded with DHM, which have the potential to overcome its bioabsorption problems and enhance its antibacterial and antioxidant effects. We used the emulsification/crosslinking method to load DHM into chitosan-derivative nanoparticles (DHM/HACC-NPs); then, SEM, TEM and FTIR analyses were used to show a denser, more tightly packed matrix for the DHM/HACC-NPs, which were successfully prepared. The size (176.4 nm), polydispersity index (PDI, 0.21), and absolute value of the zeta potential (+26.3 mV) confirmed the stability of the DHM/HACC-NPs. More importantly, DHM/HACC-NPs had a stronger antibacterial effect on S. aureus (MIC = 0.52 mg/mL and MBC = 1.02 mg/mL) than on E. coli (MIC = 1.02 mg/mL and MBC = 2.05 mg/mL). Notably, DHM/HACC-NPs were concentration-dependent scavengers of DPPH and hydroxyl radicals. In summary, DHM/HACC-NPs exhibited superior biocompatibility and antimicrobial and antioxidant activities, offering new insights into the development of a robust, udder-stable drug platform as a promising strategy for treating dairy cow mastitis.

Loading Effect of Chitosan Derivative Nanoparticles on Different Antigens and Their Immunomodulatory Activity on Dendritic Cells.

Drug carrier nanoparticles (NPs) were prepared by the polyelectrolyte method, with chitosan sulfate, with different substituents and quaternary ammonium chitosan, including C236-HACC NPs, C36-HACC NPs, and C6-HACC NPs. To evaluate whether the NPs are suitable for loading different antigens, we chose bovine serum albumin (BSA), ovalbumin (OVA), and myoglobin (Mb) as model antigens to investigate the encapsulation effect of the NPs. The characteristics (size, potential, and encapsulation efficiency) of the NPs were measured. Moreover, the NPs with higher encapsulation efficiency were selected for the immunological activity research. The results showed that chitosan derivative NPs with different substitution sites had different loading effects on the three antigens, and the encapsulation rate of BSA and OVA was significantly better than that of Mb. Moreover, the NPs encapsulated with different antigens have different immune stimulating abilities to DCS cells, the immune effect of OVA-coated NPs was significantly better than that of BSA-coated NPs and blank NPs, especially C236-HACC-OVA NPs. Furthermore, we found that C236-HACC-OVA NPs could increase the phosphorylation level of intracellular proteins to activate cell pathways. Therefore, C236-HACC NPs are more suitable for the loading of antigens similar to the OVA structure.

Synthesis of Novel Chitosan-Grafted-Derivatives Nano Cationic Polymers as Antitumor Agents

Chitosan obtained from isolated chitin of shrimp shells by a modified chemical method with a high degree of deacetylation to improve its solubility. This was undergoing a graft copolymerization process to prepare acetyl chitosan (NACS), acetyl chitosan grafted polylactide (NACS-g-PLA), chitosan grafted polylactide (CS-PLA), and polylactide chitosan grafted poly(ethylene glycol) (CS- PLA-g-PEG) nanoparticles. They characterized by FT-IR and 1HNMR. Chitosan and its derivative nanoparticles morphology examined using SEM and average nanoparticle size counted by ImageJ program. After treatment with chitosan derivative nanoparticles (1 mg/mL) at various time intervals (24, 48 and 72 hour) for three different models of human breast cancer cell lines which are BT, MCF-7 and SKBR3 cell lines, the cell proliferation, cell viability percentage, and genotoxicity as a DNA fragmentation index (%DFI) were analysed by MTT assay, and flow cytometry techniques. The results displayed that CSNPs and its derivatives NACS-g-PLA, CS-g-PLA NPs, and CS- PLA-g-PEG, have strong antitumor activities by inducing in vitro treated BT, MCF-7, and SKBR3 cell lines as a highly significant effect, (p < 0.001) on cell proliferation growth as observed with untreated control cells in a different pattern. Furthermore, it can be said since genotoxicity results that prepared polymers NPs were considered as slight /or no effect on the nucleic material of the BT cell lines, as demonstrated as %DFI in compare with positive and negative control samples.

Synthesis of polymeric chitosan derivative Nanoparticles and their MTT and flow cytometry evaluation against breast carcinoma cell

Chitosan extracted from shrimp cortex was grafted with some different anhydride compounds like; acetic anhydride, propionic anhydride, succinic anhydride, and phthalic anhydride. The chitosan-grafted-anhydrides were copolymerized with L-lactide in ring opening polymerization. All grafted chitosan derivatives were obtained in very good yield, and they were characterized by FTIR and the resulted spectra confirmed the right structures of chitosan and its different synthesized derivatives, and then they were converted to nanoparticles in size by subjecting them to sonication method.In vitro cytotoxicity detection of different chitosan anhydride derivative nanoparticles and their grafted-polylactide were concerning three different types of human breast cancer cell lines as MTT assay, the results exhibit the highly significant cell growth inhibition of these tumor cells compared with a positive control; furthermore, the chitosan anhydride derivatives grafted polylactide demonstrated increasing in reducing cell viability in comparison with their non-grafted form. The DNA fragmentation index percentage was evaluated for some of the studied polymer nanoparticles, using acridine orange dye and the results were shown no or less effective against BT breast carcinoma cell lines DNA

Self-assembled nanoparticles for cellular delivery of peptide nucleic acid using amphiphilic N,N,N-trimethyl-O-alkyl chitosan derivatives.

Peptide nucleic acid (PNA) holds enormous potentials as antisense/antigenic drug due to its specific binding ability and biostability with DNA or RNA. However, the poor cellular delivery is the key obstacle in development of PNA therapy. To overcome this difficulty, we developed self-assembled nanoparticles (NPs) for delivery of PNA to living cells using amphiphilic CS derivatives. A series of N,N,N-trimethyl-O-alkyl chitosans (TMACs) with different lengths of alkyl chains were synthesized. The structures of these synthesized chemicals were characterized with FT-IR and 1H NMR. We found that the TMACs were all able to self-assemble in aqueous condition to form nano-size NPs. These nano-size NPs are spherical shape with a size range of around 100 nm and a zeta potential above +30 mV. PNA was easily encapsulated into chitosan derivative NPs by an ultrasonic method with entrapment efficiency up to 75%. The PNA-loaded TMAC NPs released the drug in a sustained manner in PBS (pH 7.4) at 37 °C. N,N,N-trimethyl-O-cetyl chitosan (TMCC) showed the best in vitro hemocompatibility and cell viability. These TMCC based NPs were able to dramatically increase the cellular uptake of PNA, specifically, 66-fold higher compared to without using these nanoparticles. The results suggest that the designed TMCC NPs might be a promising solution for improving cellular delivery of PNA.

Preparation on Chitosan Derivative Nanoparticles as Prolonged Releasing Drug Carrier

Ricinoleic acid grafted amphiphilic carboxymethy chitosan (CMC-g-RA) was synthetised as a carrier to load botanical drug rotenone (Rot). Then Rot/CMC-g-RA water dispersion in nanoscale was prepared, whose shape, zeta potential, loading efficiency and outdoor stability were characterized accordingly. The results indicated that the sizes, polydispersity index, zeta potential of Rot/CMC-g-RA particles were affected by concentrations of this water dispersion. When the ratio of carrier to drug ascended, the water dispersion had monodisperse nanoparticle sizes with negative charge on nanoparticle surface. And the water dispersion restrained Rot degradation in natural environment with higher loading efficiency.

Development and evaluation of chitosan and chitosan derivative nanoparticles containing insulin for oral administration

Chitosan and chitosan derivative-based nanoparticles loaded with insulin were prepared by self-assembly, via electrostatic interactions between the negatively charged drug and the positively charged polymers. In the investigated chitosan derivatives, the amine groups were substituted to different extents (33, 52 or 99%) by 2-hydroxypropyl-3-trimethyl ammonium groups, rendering the polymers permanently positively charged, irrespective of the pH. This is an important property for this type of advanced drug delivery system, since the pH value changes throughout the gastrointestinal tract and electrostatic interactions are of crucial importance for the stability of the nanoparticles. Permanent positive charges are also in favor of mucoadhesion. In contrast, the electric charges of chitosan molecules depend on the pH of the surrounding medium. Since the solubility of the chitosan derivatives increased due to the introduction of quaternary ammonium groups, sodium tripolyphosphate (TPP) was added to the systems to create supplementary cross-links and stabilize the nanoparticles. The presence of TPP influenced both the dissolution of the polymer matrix as well as the resulting release kinetics. The underlying drug release mechanisms were found to be more complex than simple diffusion under constant conditions, likely involving also ionic interactions and matrix dissolution. The most promising formulation was based on a chitosan derivative with 33% substitution degree and characterized by a Z-average of 142 ± 10 nm, a zeta potential of 29 ± 1 mV, an encapsulation efficacy of 52 ± 3% and, most importantly, the release of insulin was sustained for more than 210 min.

Development of hybrid-type modified chitosan derivative nanoparticles for the intracellular delivery of midkine-siRNA in hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Most of the patients with HCC lose the surgical opportunity at the time of diagnosis. Some novel therapeutic modalities, like gene therapy, are promising for the treatment of HCC. However, the success of gene therapy depends on two aspects: efficient gene materials and gene delivery vectors. The present study was to develop new chitosan-based nanoparticles for a midkine-siRNA (anti-HCC gene drug) delivery. The novel gene delivery vector (MixNCH) was synthesized by hybrid-type modification of chitosan with 2-chloroethylamine hydrochloride and N, N-dimethyl-2-chloroethylamine hydrochloride. The chemical structure of MixNCH was characterized by FT-IR and 1HNMR. The cytotoxicity of MixNCH was determined by MTS assay. The gene condensation ability and size, zeta potential and morphology of MixNCH/MK-siRNA nanoparticles were measured. The in vitro transfection and gene knockdown efficiency of midkine by MixNCH/MK-siRNA nanoparticles was detected by qRT-PCR and Western blotting. Gene knockdown effect at the molecule level on the proliferation of HepG2 in vitro was determined by MTS assay. MixNCH was successfully acquired by aminoalkylation modification of chitosan. The MixNCH could condense MK-siRNA well above the weight ratio of 3. The average size of MixNCH/MK-siRNA nanoparticles was 100-200 nm, and the surface charge was about +5 mV. Morphologically, MixNCH/MK-siRNA nanoparticles were in regular spherical shape with no aggregation. Regarding to the in vitro transfection of nanoparticles, the MixNCH/MK-siRNA nanoparticles reduced MK mRNA level to 14.03%+/-4.03%, which were comparable to Biotrans (8.94%+/-3.77%). MixNCH/MK-siRNA effectively inhibited the proliferation of HepG2 in vitro. MixNCH/MK-siRNA nanoparticles could be effective for the treatment of hepatocellular carcinoma.

Enzyme-mediated in situ preparation of biocompatible hydrogel composites from chitosan derivative and biphasic calcium phosphate nanoparticles for bone regeneration

Injectable chitosan-based hydrogels have been widely studied toward biomedical applications because of their potential performance in drug/cell delivery and tissue regeneration. In this study we introduce tetronic–grafted chitosan containing tyramine moieties which have been utilized for in situ enzyme-mediated hydrogel preparation. The hydrogel can be used to load nanoparticles (NPs) of biphasic calcium phosphate (BCP), mixture of hydroxyapatite (HAp) and tricalcium phosphate (TCP), forming injectable biocomposites. The grafted copolymers were well-characterized by 1H NMR. BCP nanoparticles were prepared by precipitation method under ultrasonic irradiation and then characterized by using x-ray powder diffraction (XRD) and scanning electron microscopy (SEM). The suspension of the copolymer and BCP nanoparticles rapidly formed hydrogel biocomposite within a few seconds of the presence of horseradish peroxidase (HRP) and hydrogen peroxide (H2O2). The compressive stress failure of the wet hydrogel was at 591 ± 20 KPa with the composite 10 wt% BCP loading. In vitro study using mesenchymal stem cells showed that the composites were biocompatible and cells are well-attached on the surfaces.

Downregulation of VEGF mRNA expression by triamcinolone acetonide acetate-loaded chitosan derivative nanoparticles in human retinal pigment epithelial cells

BackgroundThe purpose of this study was to investigate the downregulation of mRNA expression of vascular endothelial growth factor (VEGF) by triamcinolone acetonide acetate (TAA)-loaded chitosan nanoparticles in human retinal pigment epithelial cells.MethodsTAA-loaded deoxycholic acid-modified chitosan (TAA/DA-Chit) nanoparticles were prepared via a self-assembly mechanism, and their morphology and zeta potential were examined by transmission electron microscopy and zeta potential analysis, respectively. DA-Chit and TAA/DA-Chit nanoparticle toxicity was evaluated using a Cell Counting Kit-8 assay. The efficiency of cellular uptake was determined using fluorescein isothiocyanate-labeled DA-Chit nanoparticles, in place of TAA/DA-Chit nanoparticles, assessed by both inverted fluorescence microscopy and flow cytometry. Downregulation of VEGF mRNA expression by TAA/DA-Chit nanoparticles was further investigated by real-time reverse transcription polymerase chain reaction (RT-PCR) assay of the treated human retinal pigment epithelial cells.ResultsTAA/DA-Chit nanoparticles were prepared with a TAA-loading capacity in the range of 12%–82%, which increased the water solubility of TAA from 0.3 mg/mL to 2.1 mg/mL. These nanoparticles showed oblate shapes 100–550 nm in size in transmission electron microscopic images and had positive zeta potentials. The Cell Counting Kit-8 assay indicated that the DA-Chit and TAA/DA-Chit nanoparticles had no toxicity and low toxicity, respectively, to human retinal pigment epithelial cells. Fluorescein isothiocyanate-labeled DA-Chit nanoparticle uptake by human retinal pigment epithelial cells was confirmed by inverted fluorescence microscopy and flow cytometry. Real-time RT-PCR assay showed that the VEGF mRNA level decreased after incubation of human retinal pigment epithelial cells with TAA/DA-Chit nanoparticles.ConclusionTAA/DA-Chit nanoparticles had a downregulating effect on VEGF mRNA expression in human retinal pigment epithelial cells and low cytotoxicity, which might be beneficial characteristics for the development of future treatment for diabetic retinopathy.

Characterization of novel quaternary chitosan derivative nanoparticles loaded with protein

AbstractThe objective of this work was to characterize a novel quaternary chitosan derivative [O‐(2‐hydroxyl) propyl‐3‐trimethyl ammonium chitosan chloride (O‐HTCC)] nanoparticle system. O‐HTCC nanoparticles were prepared with a simple and mild ionic gelation method upon the addition of a sodium tripolyphosphate solution to a low‐molecular‐weight O‐HTCC solution. Highly cationic chitosan nanoparticles were prepared. Bovine serum albumin (BSA), a model protein drug, was incorporated into the nanoparticles. The physicochemical properties of the nanoparticles were determined with transmission electron microscopy (TEM), scanning electron microscopy (SEM), Fourier transform infrared analysis, differential scanning calorimetry, and X‐ray diffraction (XRD) patterns. The results showed that increasing the BSA concentration from 1.5 to 2.5 mg/mL promoted the BSA encapsulation efficiency from 57.3% to 87.5% and the loading capacity from 70.2% to 99.5%. Compared with the chitosan nanoparticles, the O‐HTCC nanoparticles had lower burst release. TEM revealed that the BSA‐loaded O‐HTCC nanoparticles were smaller than the O‐HTCC nanoparticles when the BSA concentration was 1.5 mg/mL; SEM showed that the size of the BSA‐loaded O‐HTCC nanoparticles was mostly affected by the BSA concentration, and the increase in size occurred with the concentration increasing. Thermograms and XRD of the BSA‐loaded nanoparticles suggested that polyelectrolyte–protein interactions increased with the BSA concentration increasing and greater chain realignment in the BSA‐loaded nanoparticles. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009

Novel chitosan derivative nanoparticles enhance the immunogenicity of a DNA vaccine encoding hepatitis B virus core antigen in mice

Chitosan has been shown to possess useful properties such as non-toxicity, high biocompatibility and non-antigenicity that offer advantages for vaccine delivery systems. In this study, we prepared novel chitosan derivative nanoparticles as DNA vaccine carriers and the potential and mechanism of the DNA-nanoparticle complexes in inducing augmented immune responses were explored. The pVAX(HBc)DNA-nanoparticle complexes as vaccine delivery systems were studied in several aspects: the protection against DNase I degradation was measured by an in vitro inhibition assay; the sustained expression of the plasmid in vivo was determined by RT-PCR; the elevated uptake efficiency by phagocytes was observed with confocal microscopy; the biocompatibility was evaluated by cytotoxicity and histology assay; the complexes were administrated to C57BL/6 mice and the humoral and cellular immune responses were evaluated by ELISA, IFN-gamma production and cytolytic T lymphocyte (CTL)-specific lysis assay. The remaining relative activity of DNase I after inhibition varied from 32.3% to 77.6%. The complexes were observed with higher uptake efficiency by phagocytes than naked DNA. Three types of nanoparticles did not induce significant cytotoxicity at concentrations<or=400 microg/ml. No specific histological alteration related to the injection of the complexes was observed. The formulations of DNA-nanoparticle complexes significantly enhanced the immunogenicity in several parameters: elevated antibody production, higher level of IFN-gamma secretion, and augmented specific cell lysis. This study demonstrated the potential of the novel chitosan derivative nanoparticles for safe and effective DNA vaccine delivery.

Antibacterial and mechanical properties of bone cement impregnated with chitosan nanoparticles

Although total joint replacement has become commonplace in recent years, bacterial infection remains a significant complication following this procedure. One approach to reduce the incidence of joint replacement infection is to add antimicrobial agents to the bone cement used to fix the implant. In this in vitro study, we investigated the use of chitosan nanoparticles (CS NP) and quaternary ammonium chitosan derivative nanoparticles (QCS NP) as bactericidal agents in poly(methyl methacrylate) (PMMA) bone cement with and without gentamicin. The antibacterial activity was tested against Staphylococcus aureus ( S. aureus) and Staphylococcus epidermidis ( S. epidermidis). A 10 3-fold reduction in the number of viable bacterial cells upon contact with the surface was achievable using QCS NP at a nanoparticle/bone cement weight ratio of 15%. The inhibition of S. aureus and S. epidermidis growth on the surface of the CS NP and QCS NP-loaded bone cements was clearly shown using the LIVE/DEAD Baclight bacterial viability kits and fluorescence microscopy. The CS NP and QCS NP also provided a significant additional bactericidal effect to gentamicin-loaded bone cement. The antibacterial effectiveness remained high even after the modified bone cements had been immersed for 3 weeks in an aqueous medium. No cytotoxic effect of the CS NP- and QCS NP-loaded cements was shown in a mouse fibroblast MTT cytotoxicity assay. Mechanical tests indicated that the addition of the CS and QCS in nanoparticulate form allowed the retention of a significant degree of the bone cement's strength. These results indicate a new promising strategy for combating joint implant infection.