Sevda Senel

Mono-N-carboxymethyl chitosan (MCC) and N-trimethyl chitosan (TMC) nanoparticles for non-invasive vaccine delivery.

Mucosal application of a vaccine can effectively induce both systemic and mucosal immune responses. In general, mucosal applications of antigens result in poor immune responses. Therefore, adjuvant/delivery systems are required to enhance the immune response. Chitosan is a cationic biopolymer which exerts advantages as a vaccine carrier due to its immune stimulating activity and bioadhesive properties that enhance cellular uptake and permeation as well as antigen protection. Similar effects are also shown by chitosan derivatives. In this study, the nanoparticulate systems were prepared by using differently charged chitosan derivatives, N-trimethyl chitosan (TMC, polycationic), and mono-N-carboxymethyl chitosan (MCC, polyampholytic) for mucosal immunisation. The derivatives were synthesised and characterised in-house. The aqueous dispersions of the derivatives were also prepared for comparison. The cytotoxicity studies (MTT assay) on Chinese hamster ovary (CHO-K1) cell lines showed that cell viability was in the order of MCC, chitosan and TMC. Nanoparticles were prepared using ionic gelation method and loaded with tetanus toxoid (TT). Nanoparticles with high loading efficacy (>90% m/m), particle size within the range of 40-400nm, with a negative surface charge for MCC and positive surface charge for TMC and chitosan were obtained. The structural integrity of the TT in the formulations was confirmed by SDS-PAGE electrophoresis analysis. The effective uptake of the FITC-BSA loaded nanoparticles into the cells was demonstrated by cellular uptake studies using J774A.1 cells. Immune responses induced by the formulations loaded with tetanus toxoid were studied in vivo in Balb/c mice. Enhanced immune responses were obtained with intranasal (i.n.) application of nanoparticle formulations. Chitosan and TMC nanoparticles which have positively charged surfaces induced higher serum IgG titres when compared to those prepared with MCC which are negatively charged and smaller in size. Nanoparticle formulations developed in this study can be used as promising adjuvant/delivery systems for mucosal immunisation.

Effect of chitosan on a periodontal pathogen Porphyromonas gingivalis.

Local delivery systems of antimicrobial agents for treatment of the periodontal diseases received considerable attention during the past decade due to the disadvantages of the systemic administration. An ideal formulation should exhibit ease of delivery, a good retention at the application site, and a controlled release of the drug. The application of bioadhesive gels provides a long stay in the oral cavity, adequate drug penetration, high efficacy and acceptability. In dentistry and oral medicine, various applications of chitosan, which is a bioadhesive polymer have been proposed due to its favorable properties such as biocompatibility and biodegradability. The aim of this study was to determine the antimicrobial activity of chitosan formulations either in gel or film form against a periodontal pathogen, Porphyromonas gingivalis. The viscosity, bioadhesive properties and antimicrobial activity of chitosans at different molecular weight and deacetylation degree were evaluated in the absence or presence of chlorhexidine gluconate (Chx), incorporated into the formulations at 0.1 and 0.2% concentrations. The flow property of the gels were found to be suitable for topical application on the oral mucosa and to syringe into the periodontal pocket. Bioadhesion of the gels and films examined ex-vivo using fresh porcine buccal mucosa showed that both the film and gel formulations exert bioadhesive properties and was not affected by incorporation of Chx. Chitosan is shown to have an antimicrobial activity against P. gingivalis and this was higher with high molecular weight chitosan. The combination of chitosan with Chx showed a higher activity when compared to that of Chx alone, which would provide Chx application at lower concentrations thus avoiding its unwanted side effects. Chitosan films and gels seem to be promising delivery systems for local therapy of periodontal diseases with its bioadhesive property and antimicrobial activity.

The release of model macromolecules may be controlled by the hydrophobicity of palmitoyl glycol chitosan hydrogels

A non-covalently cross-linked palmitoyl glycol chitosan (GCP) hydrogel has been evaluated as an erodible controlled release system for the delivery of hydrophilic macromolecules. Samples of GCP with hydrophobicity decreasing in the order GCP12>GCP11>GCP21 were synthesised and characterised by 1H NMR. Hydrogels were prepared by freeze-drying an aqueous dispersion of the polymer in the presence or absence of either a model macromolecule fluorescein isothiocyanate-dextran (FITC-dextran, MW 4400), and/or amphiphilic derivatives Gelucire 50/13 or vitamin E d-alpha-tocopherol polyethylene glycol succinate. Gels were analysed for aqueous hydration, FITC-dextran release, and bioadhesion, and imaged by scanning electron microscopy. The gels were highly porous and could be hydrated to up to 95x their original weight without an appreciable volume change and most gels eventually eroded. Hydration and erosion were governed by the hydrophobicity of the gel and the presence of the amphiphilic additives. GCP gels could be loaded with up to 27.5% (w/w) of FITC-dextran by freeze-drying a dispersion of GCP in a solution of FITC-dextran. The controlled release of FITC-dextran was governed by the hydrophobicity of the gel following the trend GCP21>GCP11>GCP12. GCP gels were bioadhesive but less so than hydroxypropylmethylcellulose, Carbopol 974NF (7:3) tablets.

Effects of bile salts on transport rates and routes of FITC-labelled compounds across porcine buccal epithelium in vitro

Abstract In this study the penetration enhancing effect of bile salts on the transport of hydrophilic macromolecular compounds across porcine buccal mucosa was investigated in-vitro. Coadministration of 100 mM of the trihydroxy bile salts sodium glycocholate (GC) and sodium taurocholate (TC) and the dihydroxy bile salts sodium glycodeoxycholate (GDC) and sodium taurodeoxycholate (TDC) increased the in-vitro transport of fluorescein isothiocyanate (FITC) by a factor of a hundred or more, without a significant difference between the four bile salts. The concentration dependence of the enhancing effect of GDC was studied using FITC-labelled dextrans of increasing molecular weight as permeants (FD4, MW 4400; FD10, MW 9400; FD20, MW 19 600). The maximal enhancement was observed when GDC was coadministered in a concentration of 10 mM, resulting in an enhancement ratio of about 2000 for FD4. Using confocal laser scanning microscopy the effects of bile salts on the penetration pathways of hydrophilic compounds were investigated. The uniform distribution of FITC throughout the epithelium was changed by coadministration of 100 mM of bile salt to an increased amount of the fluorescent probe present in the intercellular domains. The intercellular distribution of both FD4 and FD10 was not changed by a low, but effective, concentration of GDC (2 mM, enhancement ratio of 72 for FD4). Increasing the concentration of GDC to 10 and 100 mM resulted in uptake of the fluorescent probe in the epithelial cells. From these results we conclude that the di- and trihydroxy bile salts studied increase the transport of hydrophilic compounds across buccal epithelium in vitro, below 10 mM by increasing the intercellular transport and at 10 mM and higher concentrations by opening up a transcellular route.

TMC–MCC (N-trimethyl chitosan–mono-N-carboxymethyl chitosan) nanocomplexes for mucosal delivery of vaccines

In this study, for the first time, TMC/MCC complex nanoparticles as a delivery system and as an adjuvant were developed and evaluated to obtain systemic and mucosal immune responses against nasally administered tetanus toxoid (TT). Nanoparticles were developed by complexation between the oppositely charged chitosan derivatives, N-trimethyl chitosan (TMC, polycationic) and mono-N-carboxymethyl chitosan (MCC, polyampholytic) without using any crosslinker for mucosal vaccination. The cellular viability was found to be higher with TMC/MCC complex compared to that of MCC and TMC alone. Size, zeta potential and morphology of the nanoparticles were investigated as a function of preparation method. Nanoparticles with high loading efficacy (95%) and positively charged surface were obtained with an average particle size of 283+/-2.5 nm. The structural integrity of the TT in the nanoparticles was confirmed by SDS-PAGE electrophoresis analysis. Cellular uptake studies indicated that FITC-BSA loaded nanoparticles were effectively taken up into the mouse Balb/c monocyte macrophages. Mice were nasally immunized with TT loaded TMC/MCC complex nanoparticles and compared to that of TMC and MCC nanoparticles. TMC/MCC complex nanoparticles were shown to induce both the mucosal and systemic immune response indicating that this newly developed system has potential for mucosal administration of vaccines.

Current status and the future of buccal drug delivery systems.

Background: The delivery of drugs through the buccal mucosa has received a great deal of attention over the last two decades, and yet there are not many buccal delivery products available on the market. Objective: This review outlines the advantages and disadvantages of buccal drug delivery, provides a historical perspective and discusses representative developmental and marketed drugs. Methods: The structure of the oral mucosa is briefly described to preface a description of the pathways for drug absorption and a critical discussion of permeation experiments. A brief historical perspective followed by a description of some of the currently marketed products provides a picture of where we are today. An indication is given of likely progress in this area and of the attributes of a successful business entity of the future. Conclusion: The authors provide an assessment of the future potential of buccal and sublingual drugs.

Diffusion Rates and Transport Pathways of Fluorescein Isothiocyanate (FITC)-Labeled Model Compounds Through Buccal Epithelium

The aim of this study was to characterize transport of FITC-labeled dextrans of different molecular weights as model compounds for peptides and proteins through buccal mucosa. The penetration of these dextrans through porcine buccal mucosa (a nonkeratinized epithelium, comparable to human buccal mucosa) was investigated by measuring transbuccal fluxes and by analyzing the distribution of the fluorescent probe in the epithelium, using confocal laser scanning microscopy for visualizing permeation pathways. The results revealed that passage of porcine buccal epithelium by hydrophilic compounds such as the FITC-dextrans is restricted to permeants with a molecular weight lower than 20 kDa. The permeabilities of buccal mucosa for the 4- and 10-kDa FITC-dextran (of the order of 10−8 cm/sec) were not significantly different from each other or from the much smaller compound FITC. The confocal images of the distribution pattern of FITC-dextrans showed that the paracellular route is the major pathway through buccal epithelium.

Enhancement of in vitro permeability of porcine buccal mucosa by bile salts: kinetic and histological studies

Abstract In this study the enhancing effects of di-and tri-hydroxy bile salts on buccal penetration were investigated using fluorescein isothiocyanate (FITC) as a model permeant. The permeability of the porcine buccal mucosa to this compound in the presence and absence of bile salts was determined in vitro. In conjunction with kinetic studies, histological studies were carried out to investigate the interaction between the bile salts and the buccal epithelium at the light and electron microscopic level. In the presence of the bile salts at 0.l M concentration, the permeability of buccal mucosa to FITC increased by a 100–200 fold compared to FITC alone. Calculation of enhancement ratios provided normalization of the permeability data for the enhancer treated tissue with respect to its own control. No significant difference was observed between the enhancing effects of di-hydroxy and tri-hydroxy bile salts. After the bile salt treatment significant morphological and ultrastructural changes occurred. At LM level, partial loss of superficial cells was observed as well as separation of the epithelium from the basal lamina. Upon a 4 hour bile salt treatment, two important changes were observed with FFEM: (1) the fracture plane preferentially ran across the cytoplasmic proteins and through the cytoplasmic space domains; (2) abnormally structured deposits were found in the cytoplasm. Similar to the kinetic studies, no differences were found between the histological effects of di- and tri-hydroxy bile salts.

Developing and advancing dry powder inhalation towards enhanced therapeutics.

Enhanced therapeutics are drug products derived from existing generic drugs that provide additional benefits to the patients and the healthcare system. Enhanced therapeutics are considered to be an important and relatively low risk source of innovation. Pulmonary drug delivery is the major delivery route to treat chronic respiratory diseases and has been proven as a potential delivery route for complex drugs that cannot be delivered orally. Development of dry powder inhalation systems targets the delivery of fine drug particles to the deep lung surface by a combination of drug formulation, primary packaging and a device, whereby each contributes to the overall performance. Various methodologies for the non-clinical and clinical performance testing of orally inhaled products have been proposed and applied with variable success. Regulatory pathways have been developed and applied since. Considerable efforts have been made during the past decade to understand and optimize pulmonary drug delivery including their efficient commercial manufacturing. Pulmonary drug delivery remains an area of future innovation in the effective treatment of pulmonary diseases as well as the systemic delivery of systemically active complex drugs.

Oral Mucosal Drug Delivery and Therapy

This volume provides a comprehensive overview of the current issues facing scientists working on delivering drugs locally and systemically via the membranes that line the mouth. The book describes the anatomical and physiological challenges of this route for drug delivery and how they impact the design of oral mucosal drug delivery systems. It also provides a detailed description of current oral mucosal drug delivery technologies that overcome these challenges alongside research, development and assessment methods. In 11 authoritative chapters, the book affords an in-depth evaluation of the major issues associated with this route of administration, namely the retention of the drug/product at the site of administration and increasing drug permeability through the oral mucosa. The book provides insights into the in vitro and in vivo methods available to assess drug permeability and retention, offers solutions on how to improve the permeation of the drugs through the oral mucosa, and explores approaches to prolong drug/product retention at the site of administration. It also indicates future directions in research and product development. Oral Mucosal Drug Delivery and Therapy is a key resource for those wishing to extend their knowledge of this field