Spray-dried Chitosan Microparticles Research Articles

Design and evaluation of composite films for in situ synthesis and antibacterial activity of allicin vapour

Although allicin has potent antibiotic properties, its low stability, which is responsible for its persistent biological activity, has posed a significant challenge to its practical application in modern medicine. To harness the healing benefits of this phytochemical, known by humans for thousands of years, we propose a controlled in situ synthesis of allicin vapour near the site of infection. Considering the critical need for novel approaches to prevent pandemic scenarios caused by MDR bacteria, we suggest encapsulating and physically separating allicin precursors (substrate alliin and enzyme alliinase) in alginate-based films and spray-dried chitosan microparticles. The mechanical properties of the hydrogel films of various compositions were evaluated, as well as their ability to protect the encapsulated alliinase against thermal stress and control the overall rate of allicin release upon hydration. Furthermore, the non-contact antibacterial efficacy of free alliin/alliinase reaction mixture (aqueous solution) and three compartmentalised configurations, i.e. film-solution, film-particles, and double-film, were tested against selected bacterial strains, i.e. E. coli, S. epidermidis, and S. aureus. The results indicate that the formation of allicin vapour using the proposed compartmentalised systems addresses allicin’s stability issues and provides better control over the rate of allicin production. The observed antibacterial effect was comparable with directly formed allicin using higher initial amounts of both substances, which is given by diffusion limitations associated with encapsulation. These findings illustrate the potential of compartmentalised systems in developing nature-based wound dressings for infection prevention and promoting healing.

Spray-dried pH-sensitive chitosan microparticles loaded with Mycobacterium bovis BCG intended for supporting treatment of Helicobacter pylori infection.

Gram-negative spiral-shaped Helicobacter pylori (Hp) bacteria induce the development of different gastric disorders. The growing resistance of Hp to antibiotics prompts to search for new therapeutic formulations. A promising candidate is Mycobacterium bovis BCG (BCG) with immunomodulatory properties. Biodegradable mucoadhesive chitosan is a good carrier for delivering BCG mycobacteria to the gastric mucosal environment. This study aimed to show whether BCG bacilli are able to increase the phagocytic activity of Cavia porcellus-guinea pig macrophages derived from the bone marrow towards fluorescently labeled Escherichia coli. Furthermore, to encapsulate live BCG bacilli, in spray-dried chitosan microparticles (CHI-MPs), and assess the pH-dependent release of mycobacteria in pH conditions mimicking gastric (acidic) or gut (alkaline) milieu. Microparticles (MPs) were made of chitosan and coated with Pluronic F-127-(Plur) or N-Acetyl-D-Glucosamine-(GlcNAc) to increase the MPs resistance to low pH or to increase anti-Hp effect, respectively. Spray-drying method was used for microencapsulation of live BCG. The biosafety of tested CHI-MPs has been confirmed using cell models in vitro and the model of guinea pig in vivo. The CHI-MPs loaded with BCG released live mycobacteria at pH 3.0 (CHI-GlcNAc-MPs) or pH 8.0. (CHI-Plur-MPs). The CHI-MPs loaded with live BCG can be used for per os inoculation of Cavia porcellus to check the effectiveness of delivered mycobacteria in increasing anti-H. pylori host response.

SPRAY-DRIED CHITOSAN MICROSPHERES FOR SUSTAINED DELIVERY OF TRIFLUOPERAZINE HYDROCHLORIDE: FORMULATION AND IN VITRO EVALUATION

Objective: Sustained release systems have the potential to enhance the therapeutic responses in the long-term management of psychiatric disorders. In the present study, cross-linked microspheres of the antipsychotic drug Trifluoperazine (TFP) were prepared using biodegradable polymer-chitosan and various in vitro evaluations were performed on the prepared microspheres. Methods: The spray drying technique was used to prepare TFP-loaded chitosan microspheres. Tripolyphosphate (TPP) was incorporated into the chitosan solutions as a cross-linking agent in varying concentrations. Different evaluations like production yield, encapsulation efficiency, drug-polymer compatibility, Scanning Electron Microscopy (SEM), X-ray diffraction studies (XRD), Differential Scanning Colorimetry (DSC), particle size, zeta potential analysis and in vitro drug release studies were performed on the developed formulations. Results: The formulated microparticles exhibited production yields ranging from 38.51 to 57.21% and had reasonably good encapsulation efficiencies (54.52-78.35%). The drug excipient compatibility was confirmed by Infrared Spectroscopy. All the microspheres showed positive zeta potential with a mean diameter ranging from 1.45-3.61µ. SEM images revealed the formation of spherical particles with indentations on the surface. XRD and DSC studies confirmed the presence of an amorphous form of the drug inside the microspheres. The in vitro release profile of TFP from cross-linked chitosan microspheres was influenced considerably by changing the concentration of polymer and crosslinking agent in the formulation. The drug release from (0.5%) chitosan microspheres reduced from 91% to 79%, when TPP concentration was increased from 10% w/w to 30% w/w. All the formulations clearly showed a burst release of the drug in the initial hours and a subsequent sustained release profile. Conclusion: The results of this study suggest that TPP crosslinked spray-dried chitosan microparticles could be a promising method for developing a long-acting drug delivery system intended to effectively treat schizophrenia.

Development of Methotrexate Complexes Endowed with New Biological Properties Envisioned for Musculoskeletal Regeneration in Rheumatoid Arthritis Environments.

Methotrexate (MTX) administration is the gold standard treatment for rheumatoid arthritis (RA), but its effects are limited to preventing the progression of the disease. Therefore, effective regenerative therapies for damaged tissues are still to be developed. In this regard, MTX complexes of general molecular formula M(MTX)·xH2O, where M = Sr, Zn, or Mg, were synthesized and physicochemically characterized by TGA, XRD, NMR, ATR–FTIR, and EDAX spectroscopies. Characterization results demonstrated the coordination between the different cations and MTX via two monodentate bonds with the carboxylate groups of MTX. Cation complexation provided MTX with new bioactive properties such as increasing the deposition of glycosaminoglycans (GAGs) and alternative anti-inflammatory capacities, without compromising the immunosuppressant properties of MTX on macrophages. Lastly, these new complexes were loaded into spray-dried chitosan microparticles as a proof of concept that they can be encapsulated and further delivered in situ in RA-affected joints, envisioning them as a suitable alternative to oral MTX therapy.

Direct solid-phase peptide synthesis on chitosan microparticles for targeting tumor cells

An EGFR-targeting peptide with Ala-Glu-Tyr-Leu-Arg sequence was assembled directly onto the surface of spray-dried chitosan microparticles using solid-phase peptide synthesis with Fmoc chemistry. Both targeted and scrambled peptides were cleaved from chitosan with enzymatic digestion, isolated using reversed-phase HPLC, then identified with LC/MS/MS and amino acid analysis. Particles with conjugated peptides and fluorescent-labeled were characterized for binding with A549 (cancer) and WI-26 VA4 (normal) lung cells using flow cytometry and confocal microscopy. The purity of peptide synthesis was in the range of 74–77%. The cell binding studies revealed that particles modified with the peptide bind to lung cancer cells 8.3 times higher than the normal lung cells. The binding potential of surface-modified targeted particles to the tumor cells was compared with scrambled and unmodified ones and was found to be significantly different. The binding was 7.3–7.5-fold higher than the scrambled and unmodified particles, respectively. Modification of chitosan particles with direct assembly of Ala-Glu-Tyr-Leu-Arg peptide on their surface enhanced their targeting potential to lung cancer cells and could be used as a potential carrier for delivery and therapy. This approach can further be utilized for assembly of other short peptide ligands on micro- or nanoparticulate systems for tumor targeting.

Inhalable chitosan microparticles for simultaneous delivery of isoniazid and rifabutin in lung tuberculosis treatment

The direct delivery of antibiotics to the lung has been considered an effective approach to treat pulmonary tuberculosis, which represents approximately 80% of total cases. In this sense, this work aimed at producing inhalable chitosan microparticles simultaneously associating isoniazid and rifabutin, for an application in pulmonary tuberculosis therapy. Spray-dried chitosan microparticles were obtained with adequate flow properties for deep lung delivery (aerodynamic diameter of 4 µm) and high drug association efficiencies (93% for isoniazid and 99% for rifabutin). The highest concentration of microparticles that was tested (1 mg/mL) decreased the viability of macrophage-differentiated THP-1 cells to around 60% after 24 h exposure, although no deleterious effect was observed in human alveolar epithelial (A549) cells. The release of LDH was, however, increased in both cells. Chitosan microparticles further evidenced capacity to activate macrophage-like cells, inducing cytokine secretion well above basal levels. Moreover, the propensity of macrophages to internalize microparticles was demonstrated, with uptake levels over 90%. Chitosan microparticles also inhibited bacterial growth by 96%, demonstrating that the microencapsulation preserved drug antibacterial activity in vitro. Overall, the obtained data suggest the potential of chitosan microparticles for inhalable lung tuberculosis therapy.

Chemical cross-linking: A feasible approach to prolong doxylamine/pyridoxine release from spray-dried chitosan microspheres

Spray-dried chitosan microparticles have been widely exploited as vehicles for mucosal drug delivery. Despite their advantages as pharmaceutical formulations, one of the major challenges is achieving sustained drug release, which would diminish toxicity and dosage frequency. The aim of this study was to formulate mucoadhesive glutaraldehyde cross-linked chitosan microparticles loaded with doxylamine succinate and pyridoxine hydrochloride as potential nasal drug delivery systems with sustained release. Microparticle models were formulated via spray-drying technique, using glutaraldehyde in different concentrations (0.1–1.0 mg/mL) as a cross-linking agent for chitosan. The obtained particles were with spherical shape, smooth surface and median diameter of 4 μm. The drug entrapment efficiency was high (80.47%–94.25%), indicating a tendency to decrease at higher glutaraldehyde concentrations. FTIR data demonstrated that there were no chemical interactions between glutaraldehyde and the drugs. The in vitro studies showed that the cross-linking process substantially limited particles swelling. The cross-linked particles exhibited sustained drug release characteristics at pH 6.8 over a period of 5 h with an initial burst-effect in the first 30 min. Drug release followed Korsmeyer-Peppas kinetics. Although a decrease of the particles mucoadhesive properties was observed after modification, all cross-linked formulations demonstrated high in vitro adsorption of mucin. The proposed models of mucoadhesive microsphere with sustained drug release are a perspective ground for further development of a novel delivery system for nasal administration of doxylamine and pyridoxine.

Release and permeation profiles of spray-dried chitosan microparticles containing caffeic acid

Caffeic acid (CA), a phenolic compound found in plants with antioxidant and antimicrobial activity, induces collagen production and prevents premature aging of the skin. The objective of this study was to develop two types of chitosan microparticles (MP) containing CA and to relate the morphology with the release and permeation profiles. One type of MP was prepared from a hydroalcoholic solution (MPI) and the other from an aqueous solution (MPII). Their morphology and size was evaluated by high-resolution scanning electron microscopy. The release profile of CA was evaluated using the cellulose membrane from the two MPs in Franz diffusion cells and the permeation profile was evaluated using human abdominal skin samples; the epidermal membranes were prepared by the heat-separation technique. MPII was spherical with a smooth surface, suitable for the controlled release of substances, whereas MPI was porous with non-internalized residual material. This result was consistent with their release and permeation profiles because MPII exhibited a slower and more controlled release than MPI. Thus, the method of preparation of MP and their composition influence the release profile of CA. Therefore, the production conditions must be closely controlled.

Spray Dried Chitosan Microparticles for Intravesical Delivery of Celecoxib: Preparation and Characterization

Chitosan microparticles containing celecoxib (CB), were developed as chemoprevention of bladder cancer. Furthermore two inclusion complexes of CB with methyl-β-cyclodextrin (C1 and C2) were prepared to improve the solubility of the drug. C1 and C2 were obtained by freeze-drying and characterized in the solid state and in solution. Microparticles loaded with CB or C1 or C2 were prepared by spray drying and fully characterized. The yield and encapsulation efficiencies of microparticles depended by both the viscosity and the presence of the inclusion complex in the feed medium nebulised. Generally, the microparticles exhibited a spherical shape with mean diameter of approximately 2μm which was compatible with local intravesical administration using a catheter. The CB release studies from the microparticles allowed us to identify both immediate release systems (microparticles including the complexes) and prolonged release systems (microparticles including CB alone). The latter exhibited good adhesion to the bladder mucosa, as highlighted by a mucoadhesion study. Histological studies revealed a desquamation of the superficial cells when the bladder mucosa was treated with microparticles loaded with CB, while the morphology of the urothelium did not change when it was treated with microparticles loaded with the inclusion complex. A new CB intravesical formulation than can easily be administered with a catheter and is able to release the drug at the target site for several hours was realized. This new delivery system could be a good alternative to classic oral CB administration.

Spray-dried chitosan-magnesium aluminum silicate microparticles as matrix formers in controlled release tablets

The aim of this study was to prepare and characterize chitosan-magnesium aluminum silicate (chitosan-MAS) microparticles, which can be used as matrix formers in sustained release tablets. The chitosan-MAS composites were prepared using spray drying. Physicochemical and micromeritic properties of the spray-dried chitosan-MAS microparticles were investigated. Furthermore, physical properties and drug release kinetics of the respective chitosan-MAS tablets were characterized. The results showed that spray-dried chitosan-MAS microparticles had a spherical shape with a rough surface. An exfoliated nanocomposite was formed due to molecular interaction between chitosan and MAS and immediate drying at high temperatures. The spray-dried chitosan-MAS microparticles possessed better flowability than the spray-dried chitosan microparticles. The incorporation of MAS at higher ratios resulted in an increased hardness of spray-dried chitosan-MAS tablets when increasing the compression pressure. The swelling capacity of spray-dried chitosan-MAS tablets decreased with increasing MAS ratios in acidic and neutral media. Drug release behavior of the tablets was affected by matrix swelling and erosion processes. Additionally, the tablets prepared using the spray-dried 1:0.6 chitosan-MAS microparticles displayed the slowest drug release in acidic conditions. These findings suggest that the spray-dried chitosan-MAS microparticles show strong potential for use as a matrix forming agent in tablets intended for sustaining drug release.

Physicochemical aspects involved in methotrexate release kinetics from biodegradable spray-dried chitosan microparticles

Spray dried methotrexate (MTX) loaded chitosan microparticles were prepared using different drug/copolymer ratios (9%, 18%, 27% and 45% w/w). The physicochemical aspects were assessed in order to select particles that were able to induce a sustained drug release effect. Particles were successfully produced which exhibited desired physicochemical aspects such as spherical shape and high drug loading. XRD and FT-IR analysis demonstrated that drug is not bound to copolymer and is only homogeneously dispersed in an amorphous state into polymeric matrix. Even the particles with higher drug loading levels presented a sustained drug release profile, which were mathematically modeled using adjusted Higuchi model. The drug release occurred predominantly with drug dissolution and diffusion through swollen polymeric matrix, with the slowest release occurring with particles containing 9% of drug, demonstrating an interesting and promising drug delivery system for MTX.

Preparation and Characterization of Chitosan-Based Spray-Dried Microparticles for the Delivery of Clindamycin Phosphate to Periodontal Pockets

Biodegradable spray-dried chitosan microparticles loaded with clindamycin phosphate (CDP) were formulated to deliver drugs locally into the periodontal pocket. The effects of spray dryer conditions, drug/polymer ratio, and added amounts of glutaraldehyde (GA) solution on the characterization of microparticles were investigated by determining process yield, encapsulation efficiency, particle size and size distribution, surface morphology, drug release, release kinetics, thermal analysis, and antimicrobial efficacy of formulations. Burst release was obtained for all formulations due to the water solubility of the drug, but the increased amount of chitosan decreased the drug release rates. Microparticles with a more wrinkled surface were obtained by increasing the amount of the drug. Incorporation efficiencies higher than 80% were obtained for all preparation conditions. The addition of GA caused higher viscosity of the chitosan solution, leading to larger particles with more spherical and smooth surface characteristics. However, the increased GA amount did not significantly influence the drug release. The data obtained from in vitro release experiments were best fitted to the Weibull and Higuchi models. The amorphous nature of the drug-loaded microparticles was detected by differential scanning calorimetric (DSC) thermographs. A delayed drug release of more than one week could be obtained by loading the drug into the chitosan microparticles. Antimicrobial efficacy studies reflected a positive drug release profile. These results indicate that spray-dried clindamycin-loaded microparticles with sustained antimicrobial efficacy appear to be a promising periodontal therapy for drug delivery into the periodontal pocket.

Effect of cross-linking method on the activity of spray-dried chitosan microparticles with immobilized laccase

The effect of processing method on the properties of cross-linked chitosan microparticles and on the enzymatic activity of laccase immobilized in the particles has been investigated. Chitosan has been cross-linked by tri-polyphosphate (TPP) using two methods – the so called ex situ cross-linking whereby the solutions of chitosan, TPP and the enzyme have been pre-mixed and spray-dried by a standard two-fluid kinetic nozzle, and a novel in situ cross-linking method, whereby the solutions have been contacted at the tip of a three-fluid nozzle and cross-linking occurred within a drying droplet. The influence of the cross-linking method on the particle size and morphology, surface charge, and swelling ratio has been determined. The enzymatic activity of laccase toward the oxidation of a chromophore substrate (ABTS) has been systematically investigated and found to be superior in particles produced by the in situ cross-linking method.

Development of an ex vivo retention model simulating bioadhesion in the oral cavity using human saliva and physiologically relevant irrigation media

In recent years, there has been a particular interest in bioadhesive formulations for oromucosal drug delivery as this may promote prolonged local therapy and enhanced systemic effect. Saliva plays a vital role in oromucosal drug absorption by dissolving the drug and presenting it to the mucosal surface. However, the rheological, chemical, and interfacial properties of this complex biological fluid may strongly affect the adhesion of bioadhesive formulations. There is a need for well characterized in vitro models to assess the bioadhesive properties of oral dosage forms for administration in the oral cavity. Thus we aimed at developing an advanced ex vivo buccal retention model, with focus on choosing a physiologically relevant irrigation media closely resembling human saliva. Spray dried chitosan microparticles containing metformin hydrochloride as an example of a small hydrophilic drug, were employed as bioadhesive formulations. Chewing-stimulated human whole saliva was collected and characterized for use in retention studies in comparison with four artificial irrigation media; phosphate buffer, Saliva Orthana(®), porcine gastric mucin base media (PGM3), and xanthan gum based media (XG2). Retention of metformin, applied as spray dried microparticles on porcine buccal mucosa, greatly depended on the characteristics of the irrigation media. When rheology of the irrigation media was examined, changes in retention profiles could be interpreted, as irrigation media containing mucin and xanthan gum possessed a higher viscosity than phosphate buffer, which led to longer retention of the drug due to better hydration of the mucosa and the spray dried microparticles. Metformin retention profiles were comparable when human saliva, Saliva Orthana(®), or PGM3 were used as irrigation media. Moreover, PGM3 displayed physico-chemical properties closest to those of human saliva with regard to pH, protein content and surface tension. Saliva Orthana(®) and PGM3 are therefore considered as suitable irrigation media for further retention studies.

Spray-Dried Chitosan Microparticles for Cellular Delivery of an Antigenic Protein: Physico-chemical Properties and Cellular Uptake by Dendritic Cells and Macrophages

Spray-dried chitosan microparticles for cellular delivery of antigen to dendritic cells (DC) and macrophages (Mϕ) were investigated. Chitosan microparticles were prepared by spray drying. For comparison, poly(lactic-co-glycolic acid) (PLGA) and poly(α-butyl cyanoacrylate) (BCA) micro-/nanoparticles were generated. Bovine serum albumin (BSA) was used as a model antigen. The particles were characterized in terms of size, morphology, surface charge, surface composition, protein content, entrapment efficiency, in vitro release, and protein integrity. Additionally, they were subject to cell viability and cellular uptake study with DC and Mϕ. Size of chitosan, PLGA, and BCA micro-/nanoparticles ranged between 3.11-7.18, 0.94-6.26, and 0.30-6.34μm, respectively. Particle morphology and in vitro protein release varied, depending on polymer type, particle composition and preparation process parameters. Chitosan microparticles were cationic, while PLGA microparticles were neutral. BCA micro-/nanoparticles were either anionic or cationic, according to polymerization pH. Protein content and entrapment efficiency of chitosan and PLGA microparticles were relatively consistent. Only integrity and conformational structure of protein encapsulated in chitosan microparticles were completely retained. Chitosan and PLGA microparticles were non-toxic to DC and Mϕ, but the former were internalized more efficiently. Spray-dried chitosan microparticles delivered the antigen efficiently to DC and Mϕ.

Retained integrity of protein encapsulated in spray-dried chitosan microparticles

Chitosan microparticles for delivery of proteins were prepared by spray-drying technique. The effects of formulation (molecular weight and concentration of chitosan) and process variables (inlet drying air temperature and spray rate) on size and morphology of microparticles were characterized. Size of microparticles was mainly controlled by formulation variables, while particle morphology was influenced by both formulation and process variables investigated in this study. Bovine serum albumin (BSA), as a model protein, was loaded into microparticles at different levels. BSA-loaded chitosan microparticles were characterized in terms of physicochemical properties and integrity of encapsulated protein, which was studied by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and circular dichroism. Size of microparticles ranged between 3.760–8.681 µm, of which BSA-loaded microparticles were larger in size than their corresponding blank microparticles. All microparticles showed dented or distorted surface, especially when BSA was incorporated, with positive surface charge exposed. Burst release of protein was observed. The effect was more pronounced as BSA loading level was increased. Integrity of entrapped protein could be retained when BSA was incorporated at high loading level. In conclusion, chitosan microparticles for delivery of protein could be efficiently prepared by spray-drying technique. The encapsulated protein was capable of retaining its integrity after the preparation process.

Spray-Dried Chitosan Microspheres as a pDNA Carrier

ABSTRACT Spray-dried chitosan microparticles with controlled size were prepared and crosslinked with the biocompatible reactant d,l-glyceraldehyde. Glutaraldehyde, the conventional cross-linking agent for proteins, was used as a control. The crosslinking degree was characterized through thermal analysis, X-ray diffraction, and infrared spectroscopy. In addition, the crosslinking effects were analyzed in terms of particle size, swelling capability, and surface charge. By properly choosing d,l-glyceraldehyde concentration and crosslink time, a high degree of control was achieved over the physical properties of particles. Yields around 78% as well as images obtained through fluorescence microscopy demonstrated the capability of the crosslinked chitosan microspheres to pDNA complexation.