N-trimethyl Chitosan Chloride Research Articles

Chitosan, Chitosan/IgG-Loaded, and N-Trimethyl Chitosan Chloride Nanoparticles as Potential Adjuvant and Carrier-Delivery Systems.

This work proposes a feasible, reproducible, and low-cost modified method to manufacture chitosan, chitosan/IgG-protein-loaded, and trimethylated chitosan nanoparticles, using microfluidics combined with the microemulsion technique, which differs from the traditional batch process of chitosan-based nanoparticles. The synthesis process consists of generating microreactors of chitosan-based polymer in a poly-dimethylsiloxane ψ-shaped microfluidic device and then crosslinking with sodium tripolyphosphate outside the cell. Transmission electron microscopy demonstrates an improvement in size control and distribution of the solid-shape chitosan nanoparticles (~80 nm) compared to the batch synthesis. Regarding chitosan/IgG-protein-loaded nanoparticles, these presented a core-shell morphology having a diameter of close to 15 nm. Raman and X-ray photoelectron spectroscopies confirmed the ionic crosslinking between the amino groups of chitosan and the phosphate groups of sodium tripolyphosphate in the fabricated samples and the total encapsulation of IgG protein during the fabrication of chitosan/IgG-loaded nanoparticles. Then, an ionic crosslinking and nucleation-diffusion process of chitosan-sodium tripolyphosphate was carried out during the nanoparticle formation, with and without IgG protein loading. The use of N-trimethyl chloride chitosan nanoparticles in vitro on human-keratinocyte-derived cell line HaCaT did not show side effects independently of its concentration from 1 to 10 μg/mL. Therefore, the proposed materials could be used as potential carrier-delivery systems.

Synthesis of biodegradable antimicrobial pH-sensitive silver nanocomposites reliant on chitosan and carrageenan derivatives for 5-fluorouracil drug delivery toward HCT116 cancer cells

The current research relies on a one-pot green biosynthesis of silver nanoparticles (SNPs) with various ratios of silver (Ag) in the existence of N, N, N-trimethyl chitosan chloride (TMC) and carboxymethyl kappa-carrageenan (CMKC), to investigate the effectiveness of the synthesized silver nanocomposites (SNCs) as pH sensitive biodegradable carrier for orally intestinal delivery of 5-fluorouracil (5-FU) drug. FTIR, XRD, TEM and FE-SEM/EDX methods were utilized to demonstrate the structure of the prepared polyelectrolyte complex PEC (TMC/CMKC) and SNCs (TMC/CMKC/Ag). The results showed that the 5-FU encapsulation effectiveness inside all of the prepared SNCs samples was improved by increasing the concentration of Ag, reaching 92.16 ± 0.57 % with 3 % Ag. In vitro release behavior of 5-FU loaded SNC 3 % (TMC/CMKC/Ag 3 %), displayed slow and sustained release reaching 96.3 ± 0.81 % up to 24 h into pH 7.4 medium. The successful release of 5-FU from the loaded SNC 3 % was confirmed through occurrence of strong cytotoxicity, with an IC50 value of 31.15 μg/ml, and high % of apoptotic cells (30.66 %) within the treated HCT116 cells. Besides, SNC 3 % showed good biodegradability and antimicrobial properties against different bacterial strains. Overall, SNC 3 % can be suggested as an effective system for both controlled drug delivery and antibacterial action.

Mice immunized with trimethyl chitosan nanoparticles containing DENV-2 envelope domain III elicit neutralizing antibodies with undetectable antibody-dependent enhancement activity.

Dengue is a disease that poses a significant global public health concern. Although a tetravalent live-attenuated dengue vaccine has been licensed, its efficacy is still debated due to evidence of vaccine breakthrough infection. To avoid this issue, dengue vaccines should stimulate a high degree of serotype-specific response. Thus, envelope domain III (EDIII), which contains serotype-specific neutralizing epitopes, is an attractive target for dengue vaccine development. In this study, we investigated how EDIII encapsidated in N, N, N-trimethyl chitosan chloride nanoparticles (TMC NPs) stimulates a serotype-specific response and whether this response exerts a potential in vitro breakthrough infection. The immune response to DENV-2 elicited by EDIII TMC NP-immunized mice was monitored. We demonstrated that immunization with EDIII TMC NPs resulted in a high level of anti-EDIII antibody production. These antibodies included IgG, IgG1, and IgG2a subtypes. Importantly, antibodies from the immunized mice exerted efficient neutralizing activity with undetectable antibody dependent enhancement (ADE) activity. We also found that EDIII TMC NPs activated functional EDIII-specific CD4+ and CD8+ T cell responses. In conclusion, EDIII TMC NPs stimulated humoral immunity with a strong neutralizing antibody response, as well as a cellular immune response against DENV-2.

Nanoparticle-mediated intravesical delivery of conditioned medium derived from mesenchymal stem cells for interstitial cystitis/bladder pain syndrome treatment

Interstitial cystitis/bladder pain syndrome (IC/BPS) is a critical disease characterized by debilitating pelvic pain associated with lower urinary tract symptoms. Effective therapeutic strategies remain challenging due to their elusive pathogenesis. Mesenchymal stem cell (MSC) therapy offers a promising therapeutic approach for IC/BPS treatment. Yet, the ethical arguments and tumorigenic risks from cell therapy by stem cells restrict the clinical application. Herein, we developed a nanoparticle that encapsulates the conditioned medium (CM) from stem cells and coated with cationic N-trimethyl chitosan chloride (TMC) on the surface to improve the penetration of epithelial barrier in intravesical therapy for IC/BPS. The TMC coating enhanced the accumulation of CM-loaded nanoparticles in the urothelium and subsequently improved the therapeutic efficacy on the mechanical threshold and micturition function through facilitating the effect of anti-inflammation and BDNF/TrkB signaling, which provides a great potential application on the transmucosal delivery platform for intravesical therapy of IC/BPS.

Healing Effect and Underlying Mechanism of Silver Nanoparticles Prepared Using Carboxymethyl Chitosan under Solar Irradiation

In our previous study, silver nanoparticles were prepared using AgNO3 and carboxymethyl chitosan (CMCS) which is commercially available with solar irradiation. In this study, the efficacy and safety of silver nanoparticles prepared by this method were evaluated for healing wounds in rats with diabetes. We also attempted to determine the underlying mechanism and influencing factors of the silver nanomaterial preparation method. Compared with growth factors, silver nanoparticles exhibit better healing effects for rats with diabetes. No remnant silver ions were detected in the major organs of these rats after the application of silver nanoparticles. Silver nanoparticles prepared using CMCS are less toxic than those prepared from the conventional method, promote the proliferation of skin fibroblasts, and are promising as a topical medication for infected wounds. An obvious breakage process of the particles is observed during the growth of nanocrystalline silver in CMCS. In this study, we also attempted to determine whether this method is suitable for synthesizing silver nanoparticles using N-trimethyl chitosan chloride and sodium alginate were used in this particular experiment. The characteristic UV-vis absorbance peak of silver nanoparticles was found only in the reaction mixture containing N-trimethyl chitosan chloride. Our study demonstrates that free radicals are the key factor in this silver nanoparticle preparation method.

Towards the development of Self-Nano-Emulsifying Drug Delivery Systems (SNEDDS) containing trimethyl chitosan for the oral delivery of amphotericin B: In vitro assessment and cytocompatibility studies

In the current study, a self-nanoemulsifying drug delivery system (SNEDDS) containing N-trimethyl chitosan chloride (TMC) was developed and evaluated for its potential to deliver amphotericin B (AmpB) via the gastrointestinal tract. SNEDDS comprising Captex 355, Kolliphor RH40 and Propylene Glycol were optimized and characterized for their formation spontaneity, droplet size and drug loading. Investigations on the effects of the application SNEDDS containing TMC to a model intestinal epithelium (Caco-2 monolayer) indicated the capability of the formulations to induce the transient opening of tight junctions. Caco-2 cell viability studies confirmed the safety of the SNEDDS, whereas in vitro transport studies of AmpB through Caco-2 cell monolayers showed the permeation enhancing ability of TMC. Our results suggest that chitosan derivative TMC combined with a SNEDDS may be used as permeation enhancer to facilitate oral delivery of AmpB.

Lectins at Interfaces-An Atomic Force Microscopy and Multi-Parameter-Surface Plasmon Resonance Study.

Lectins are a diverse class of carbohydrate binding proteins with pivotal roles in cell communication and signaling in many (patho)physiologic processes in the human body, making them promising targets in drug development, for instance, in cancer or infectious diseases. Other applications of lectins employ their ability to recognize specific glycan epitopes in biosensors and glycan microarrays. While a lot of research has focused on lectin interaction with specific carbohydrates, the interaction potential of lectins with different types of surfaces has not been addressed extensively. Here, we screen the interaction of two specific plant lectins, Concanavalin A and Ulex Europaeus Agglutinin-I with different nanoscopic thin films. As a control, the same experiments were performed with Bovine Serum Albumin, a widely used marker for non-specific protein adsorption. In order to test the preferred type of interaction during adsorption, hydrophobic, hydrophilic and charged polymer films were explored, such as polystyrene, cellulose, N,-N,-N-trimethylchitosan chloride and gold, and characterized in terms of wettability, surface free energy, zeta potential and morphology. Atomic force microscopy images of surfaces after protein adsorption correlated very well with the observed mass of adsorbed protein. Surface plasmon resonance spectroscopy studies revealed low adsorbed amounts and slow kinetics for all of the investigated proteins for hydrophilic surfaces, making those resistant to non-specific interactions. As a consequence, they may serve as favorable supports for biosensors, since the use of blocking agents is not necessary.

Functionalized Scaffold for in Situ Efficient Gene Transfection of Mesenchymal Stem Cells Spheroids toward Chondrogenesis.

Multicellular mesenchymal stem cell (MSC) spheroids possess enhanced chondrogenesis ability and limited fibrosis, exhibiting advantage toward hyaline-like cartilage regeneration. However, because of the limited cell surfaces in spheroid exposed to DNA/vector, it is difficult to realize efficient gene transfection, most of which highly rely on cell-substrate interaction. Here, we report a poly(l-glutamic acid)-based porous scaffold with tunable inner surfaces that can sequentially realize cell-scaffold attachment and detachment, as well as the followed in situ spheroid formation. The attachment and detachment of cells from scaffold is achieved by the capture and release of fibronectin (Fn) via reversible imine linkage between aromatic aldehyde groups of scaffold and amino groups of Fn. Together with N, N, N-trimethyl chitosan chloride condensing plasmid DNA encoding transforming growth factor-β1 (pDNA-TGF-β1), cell attachment realizes efficient surface-mediated gene transfection. Conversion of scaffold stiffness can affect the adhesion shape of cells. Stiffer scaffold reinforces the adhesion, leading to the amplification of peripheral focal adhesions and the promotion of cell spreading, as well as the promotion of gene transfection efficiency. After cellular detachment from the scaffold via lysine treatment, the subsequent spheroid formation with extensive cell-cell interaction up-regulates the corresponding protein expression with a prolonged term. With the induction effect of the expressed TGF-β1, significantly enhanced chondrogenesis of MSCs in spheroids is achieved at 10 d in vitro. Well-regenerated cartilage at 8 weeks in vivo indicates that the present gene transfection system is a platform that can be potentially applied toward cartilage tissue engineering.

Enhanced antitumor efficacy of arginine modified amphiphilic nanoparticles co-delivering doxorubicin and iSur-pDNA via the multiple synergistic effect

Arginine and α-tocopherol succinate (α-TOS) double grafted N-trimethyl chitosan chloride (TMC) nanoparticles (TAS NPs) were designed and developed for effective co-delivery of doxorubicin (DOX) and Survivin shRNA-expressing pDNA (iSur-pDNA). With DOX loading into the hydrophobic core and iSur-pDNA combining to the hydrophilic shell, TAS/DOX/pDNA NPs demonstrated favorable structural stability and sustained release properties in vitro. With the special non-clathrin-dependent endocytosis, TAS/DOX/pDNA NPs presented higher cellular uptake and mainly distributed in ER and Golgi rather than lysosomes following internalization. The in vitro nuclear localization, gene silencing efficiency, cell apoptosis, and growth inhibition of tumor cells were significantly promoted by arginine modification. In the tumor-bearing mice model, TAS/DOX/pDNA NPs possessed the maximum antitumor efficiency as compared with single delivery of DOX or iSur-pDNA. Particularly, blank TAS NPs were selectively be toxic to tumor cells as evidenced by their capabilities to inhibit proliferation and induce apoptosis of tumor cells. The promising tumor treatment of TAS/DOX/pDNA NPs via a multiple synergistic manner arising from DOX and pDNA as well as the vectors would provide a potential strategy for a dual-delivery system to improve their therapeutic efficacies.

Development of MPC-DPA polymeric nanoparticle systems for inhalation drug delivery applications

Inhalation of nanoparticles for pulmonary drug delivery offers the potential to harness nanomedicine formulations of emerging therapeutics, such as curcumin, for treatment of lung cancer. Biocompatible nanoparticles composed of poly(2-methacryloyloxyethyl phosphorylcholine)-b-poly(2-(diisopropylamino)ethyl methacrylate) (MPC-DPA) have been shown to be suitable nanocarriers for drugs, whilst N-trimethyl chitosan chloride (TMC) coating of nanoparticles has been reported to further enhance their cellular delivery efficacy; the combination of the two has not been previously investigated. Development of effective systems requires the predictable, controllable, and reproducible ability to prepare nanosystems possessing particle sizes, and drug loading capacities, appropriate for successful airway travel, lung tissue penetration, and tumor suppression. Although a number of MPC-DPA based nanosystems have been described, a complete understanding of parameters controlling nanoparticle formation, size, and morphology has not been reported; in particular the effects of differing solvents phases remains unclear. In this current study a matrix of 31 solvent combinations were examined to provide novel data pertaining to the formation of MPC-DPA nanoparticles, and in doing so afforded the selection of systems with particle sizes appropriate for pulmonary delivery applications to be loaded with curcumin, and coated with TMC. This paper presents the first report of novel data detailing the successful preparation, characterisation, and optimisation of MPC-DPA nanoparticles of circa 150–180nm diameter, with low polydispersity, and a curcumin loading range of circa 2.5–115μM, tunable by preparation parameters, with and without TMC coating, and thus considered suitable candidates for inhalation drug delivery applications.

Development of chitosan–pullulan composite nanoparticles for nasal delivery of vaccines: optimisation and cellular studies

Nasal immunisation with nanoparticles has already shown promising results. In this study, nanoparticle composites carrying BSA for nasal vaccination prepared using electrostatic interaction process between polycation N-trimethyl chitosan chloride (TMC), chitosan glutamate (CG), chitosan chloride (CCl) and polyanion carboxymethyl pullulan (CMP). A mass ratio of 2:1 for TMC-CMP combination produced stable nanocarriers. For CCl-CMP and CG-CMP formulations needed a mass ratio of 3:1. Loading efficiency was >90% for all formulations. Nanoparticles’ size ranged from 207 to 603 nm. The surface charge of the complexes varied between +14 and +33 mV. SDS-PAGE integrity of the model antigen was also demonstrated. MTT studies showed that nanoparticle composites were less toxic to Calu-3 cells than the particles of cationic polymers alone. FITC-BSA loaded nanoparticles efficiently taken up by J774A.1 macrophages as confirmed by confocal microscopy highlighting the potential of these novel nanoparticulate carriers’ use for nasal vaccination.

Layer-by-Layer Assembly of Inactivated Poliovirus and N-Trimethyl Chitosan on pH-Sensitive Microneedles for Dermal Vaccination.

The aim of this work was to coat pH-sensitive microneedle arrays with inactivated polio vaccine (IPV) particles and N-trimethyl chitosan chloride (TMC) via electrostatic interactions, and assess the immunogenicity of the vaccine after topical application of the coated microneedles in rats. The surface of 200 μm long microneedles was first chemically modified with pH-sensitive (pyridine) groups and then coated with negatively charged IPV and a positively charged polymer (TMC). To obtain a sufficient high antigen dose, 10 layers of IPV were alternately coated with TMC. The binding of IPV and TMC onto pH-sensitive microneedles was quantified and visualized by using fluorescently labeled TMC and IPV. The release of IPV and TMC from the microneedles was evaluated in ex vivo human skin by fluorescence and the immunogenicity of (unlabeled) IPV was assessed after topical application of the coated microneedles in rats. pH-sensitive microneedles were homogeneously coated with 10 layers of both IPV and TMC, resulting in 45 D antigen units IPV and 700 ng TMC per microneedle array. Fluorescence microscopy imaging revealed that both IPV and TMC were released into ex vivo human skin upon application of the coated microneedles. Finally, in vivo application of IPV-TMC-coated pH-sensitive microneedles in rats led to the induction of IPV specific antibody responses, illustrating that they are practically applicable. Topical administration of pH-sensitive microneedles coated with polyelectrolyte multinanolayers of antigens and oppositely charged polymers may be a useful approach for microneedle-based vaccination.

Positively Charged Chitosan and N-Trimethyl Chitosan Inhibit Aβ40 Fibrillogenesis.

Amyloid fibrils, formed by aggregation of improperly folded or intrinsically disordered proteins, are closely related with the pathology of a wide range of neurodegenerative diseases. Hence, there is a great deal of interest in developing molecules that can bind and inhibit amyloid formation. In this regard, we have investigated the effect of two positively charged polysaccharides, chitosan (CHT) and its quarternary derivative N-trimethyl chitosan chloride (TMC), on the aggregation of Aβ40 peptide. Our aggregation kinetics and atomic force microscopy (AFM) studies show that both CHT and TMC exhibit a concentration-dependent inhibiting activity on Aβ40 fibrillogenesis. Systematic pH-dependent studies demonstrate that the attractive electrostatic interactions between the positively charged moieties in CHT/TMC and the negatively charged residues in Aβ40 play a key role in this inhibiting activity. The stronger inhibiting activity of TMC than CHT further suggests the importance of charge density of the polymer chain in interacting with Aβ40 and blocking the fibril formation. The possible interactions between CHT/TMC and Aβ40 are also revealed at the atomic level by molecular docking simulation, showing that the Aβ40 monomer could be primarily stabilized by electrostatic interactions with charged amines of CHT and quaternary amines of TMC, respectively. Binding of CHT/TMC on the central hydrophobic core region of Aβ40 peptide may be responsible for blocking the propagation of the nucleus to form fibrillar structures. These results suggest that incorporation of sugar units such as d-glucosamine and N-trimethyl-d-glucosamine into polymer structural template may serve as a new strategy for designing novel antiamyloid molecules.

N-trimethyl chitosan chloride-coated PLGA nanoparticles overcoming multiple barriers to oral insulin absorption.

Although several strategies have been applied for oral insulin delivery to improve insulin bioavailability, little success has been achieved. To overcome multiple barriers to oral insulin absorption simultaneously, insulin-loaded N-trimethyl chitosan chloride (TMC)-coated polylactide-co-glycoside (PLGA) nanoparticles (Ins TMC-PLGA NPs) were formulated in our study. The Ins TMC-PLGA NPs were prepared using the double-emulsion solvent evaporation method and were characterized to determine their size (247.6 ± 7.2 nm), ζ-potential (45.2 ± 4.6 mV), insulin-loading capacity (7.8 ± 0.5%) and encapsulation efficiency (47.0 ± 2.9%). The stability and insulin release of the nanoparticles in enzyme-containing simulated gastrointestinal fluids suggested that the TMC-PLGA NPs could partially protect insulin from enzymatic degradation. Compared with unmodified PLGA NPs, the positively charged TMC-PLGA NPs could improve the mucus penetration of insulin in mucus-secreting HT29-MTX cells, the cellular uptake of insulin via clathrin- or adsorption-mediated endocytosis in Caco-2 cells and the permeation of insulin across a Caco-2 cell monolayer through tight junction opening. After oral administration in mice, the TMC-PLGA NPs moved more slowly through the gastrointestinal tract compared with unmodified PLGA NPs, indicating the mucoadhesive property of the nanoparticles after TMC coating. Additionally, in pharmacological studies in diabetic rats, orally administered Ins TMC-PLGA NPs produced a stronger hypoglycemic effect, with 2-fold higher relative pharmacological availability compared with unmodified NPs. In conclusion, oral insulin absorption is improved by TMC-PLGA NPs with the multiple absorption barriers overcome simultaneously. TMC-PLGA NPs may be a promising drug delivery system for oral administration of macromolecular therapeutics.

Combining Chemical Permeation Enhancers for Synergistic Effects

Currently, macromolecular drugs such as proteins are mainly administered by means of injections due to their low intestinal epithelial permeability and poor stability in the gastrointestinal tract. This study investigated binary combinations of chemical drug absorption enhancers to determine if synergistic drug absorption enhancement effects exist. Aloe vera, Aloe ferox and Aloe marlothii leaf gel materials, as well as with N-trimethyl chitosan chloride (TMC), were combined in different ratios and their effects on the transepithelial electrical resistance (TEER), as well as the transport of FITC-dextran across Caco-2 cell monolayers, were measured. The isobole method was applied to determine the type of interaction that exists between the absorption enhancers combinations. The TEER results showed synergism existed for the combinations between A. vera and A. marlothii, A. marlothii and A. ferox as well as A. vera and TMC. Antagonism interactions also occurred and can probably be explained by chemical reactions between the chemical permeation enhancers, such as complex formation. In terms of FITC-dextran transport, synergism was found for combinations between A. vera and A. marlothii, A. marlothii and A. ferox, A. vera and TMC, A. ferox and TMC and A. marlothii and TMC, whereas antagonism was observed for A. vera and A. ferox. The combinations where synergism was obtained have the potential to be used as effective drug absorption enhancers at lower concentrations compared to the single components.

Preparation and characterization of bioadhesive microparticles comprised of low degree of quaternization trimethylated chitosan for nasal administration: effect of concentration and molecular weight.

Toward the development of microparticulate carriers for nasal administration, N-trimethylchitosan chloride (TMC) of low molecular weight (LMW) and high molecular weight (HMW) and low degree of quaternization (16% and 27%, respectively) was co-formulated into microparticles comprising of dipalmatoylphosphatidylcholine (DPPC) and poly(lactic-co-glycolic) acid (PLGA) via the spray-drying technique. The chitosan derivatives were characterized by means of nuclear magnetic resonance (NMR), differential scanning calorimetry (DSC), and Fourier transfrom infrared (FTIR) spectroscopy. The size and morphology of the produced microparticles were assessed by scanning electron microscopy (SEM), whereas their mucoadhesive properties were investigated by means of atomic force microscopy-force spectroscopy (AFM-FS). The results showed that microparticles exhibit mucoadhesion when TMC is present on their surface above a threshold of TMC (>0.3% w/w).

Oral absorption enhancement of salmon calcitonin by using both N-trimethyl chitosan chloride and oligoarginines-modified liposomes as the carriers

In this study both N-trimethyl chitosan chloride (TMC) and oligoarginine (Arg8) were utilized to modify liposomes as the multifunctional carriers (TMC-Arg8-Lips) for enhancing the oral absorption of salmon calcitonin. Two permeation enhancers with positive charges were sequentially adsorbed on the liposomal surface with negative charges by electrostatic interaction. Instead of salmon calcitonin, fluorescein isothiocyanate dextran (FD4) was loaded in TMC-Arg8-Lips for Caco-2 cell permeation test in vitro and penetration examination in rat intestinal tract in vivo. The results showed that the apparent permeability coefficient (Papp) of TMC-Arg8-Lips containing FD4 were 7.0-, 4.4-, 1.8- and 1.4-folds higher than FD4 solution, FD4-TMC solution, non-modified liposomes (Non-Lips) and TMC modified liposomes (TMC-Lips), respectively. A strong fluorescence was observed by confocal laser scanning microscope (CLSM) at rat intestinal wall isolated in different times after the FD4 loaded carriers were intragastrically administrated. Furthermore, the images revealed that TMC-Arg8-Lips could penetrate deeply inside the mucosal membrane. The pharmacodynamic study indicated that TMC-Arg8-Lips containing calcitonin were more efficient in enhancing the absorption and prolonging the reduction of blood calcemia in rats. The area above the plasma calcium concentration–time curve (AAC) of TMC-Arg8-Lips containing calcitonin was increased by more than 16.6- and 1.6-fold when compared to Non-Lips and TMC-Lips, respectively.

Eudragit® L100/N-Trimethylchitosan Chloride Microspheres for Oral Insulin Delivery

Effective oral delivery of protein and peptide drugs remains an active topic in scientific research. In this study, matrix type microspheres were prepared with Eudragit® L100 containing N-trimethylchitosan chloride to improve the permeation of insulin across the intestinal epithelium via the paracellular pathway. Insulin loaded microspheres were initially formulated in accordance with a factorial design (23) and manufactured by means of a single water-in-oil emulsification/evaporation method. Based on external and internal morphology two microsphere formulations were selected from the initial formulations for further investigation in terms of particle size, dissolution behaviour and in vitro insulin transport across excised rat intestinal tissue. The initial eight microsphere formulations exhibited drug loading capacities ranging from 27.9–52.4% with different shapes and internal structures. The two selected microsphere formulations had average particle sizes of 157.3 ± 31.74 µm and 135.7 ± 41.05 µm, respectively, and mean dissolution time values for insulin release of 34.47 and 42.63 min, respectively. In vitro transport of insulin across excised rat intestinal tissue from the two selected microsphere formulations was 10.67–fold and 9.68–fold higher than the control group (insulin alone). The microsphere delivery system prepared from Eudragit® L100 containing N-trimethylchitosan chloride is therefore a promising candidate for effective oral insulin delivery.

A once-a-day dosage form for the delivery of insulin through the nasal route: in vitro assessment and in vivo evaluation.

An in situ thermogelling, mucoadhesive formulation based on N-trimethyl chitosan chloride has been evaluated for its potential to affect the transmucosal delivery of insulin via the nasal route. In vitro studies at a physiologically relevant temperature (ca. 35 °C) have shown that the formulation releases most of its insulin load (ca. 70%) in a non-Fickian manner during the timescale over which the sol-to-gel transition (ca. 8 min) takes place, and also that, once gelation is complete, the release of the remainder of the therapeutic content follows first order kinetics over at least sixty minutes. Investigations on the effects of the application of the same formulation to a modelled nasal mucosa (Calu-3 cell monolayer) have indicated the capability of the formulation to induce the transient opening of tight junctions. Cytotoxic investigations have shown that the formulation exhibits negligible detrimental effects to the integrity of these monolayers. The in vivo potential of the nasal formulation to act as a once-a-day dosage form for the intranasal delivery of insulin has been demonstrated in a diabetic-rat model.

Synthesis and Hydroxyl Radicals Scavenging Activity of 2-Pyridine-acetyl-N-trimethyl Chitosan Chloride

A novel chitosan derivative with double quaternary ammonium salt-2-pyridine-acetyl-N-trimethyl chitosan chloride (PATMCS) was synthesized and the antioxidant activity of PATMCS against hydroxyl radicals was assessed. The results indicated that PATMCS had potent hydroxyl scavenging activity. The <TEX>$IC_{50}$</TEX> of PATMCS was 0.13 mg/mL. PATMCS showed 100% scavenging effect at a dose of 0.8 mg/mL which markedly better than that of N-trimethyl chitosan chloride (TMCS). It was confirmed that quaternary chitosan derivatives showed potent antioxidant activity. PATMCS has double quaternary ammonium salt structure in the molecules. Therefore, the antioxidant activity of PATMCS was better than TMCS. The above results are theoretically fundamental for further development and making use of chitosan resources to prepare new antioxidants.