RESUMO
Poor absorption of protein antigens across mucosal membranes necessitates the use of drug delivery systems. Pre-formed dextran microspheres [sephadex] possess a good potential for delivery of protein and peptide drugs to mucosal membranes. However they have rarely been used for mucosal immunization. Regards the advantages of mucosal over parenteral immunization and also the penetration enhancement potential of dextran microspheres, we examined their efficacy as mucosal antigen delivery system. Dextran microspheres loaded with tetanus toxoid [TT] were prepared by suspending microspheres in TT solution followed by freeze-drying. Size and morphological features of microspheres were studied by optical and electron microscopes. Encapsulation efficiency of TT was determined by micro-BCA protein assay. In vitro release study was performed in a model, simulating nasal cavity. Immunoreactivity of encapsulated antigen was assayed by ELISA. Gamma-scintigraphy [GS] was used for determination of mucoadhesion and clearance rate of microspheres, by spraying of technetiumlabelled microspheres to the nose of human volunteers. Local irritating potential of microspheres was studied by nasal administration to human nose. Microspheres with a mean diameter of about 150 micro m and a TT encapsulation efficiency of 20.3% [n=3] were obtained. Antigenicity of encapsulated TT was determined as 90.5+/-1.8% [n=3] that of the original TT. Release studies showed that almost 100% of TT was released in 1 h. GS studies showed that after 4 h, 36.9+/-3.4% [n=4] of microspheres were cleared from nasopharynx region of volunteers. Considering the suitable release characteristics, desirable preservation of the antigenic activity of TT, appropriate mucoadhesive properties and also safety of TT loaded Sephadex microspheres, these microspheres could be used as a proper mucosal antigen delivery system. However further studies are needed regarding the immune response induced by these systems in an animal model
RESUMO
Surfactants are widely used as detergents, emulsifiers, solubilizers and in recent years as penetration enhancers. In the present study, the hemolytic effects of four non-ionic surfactants from the polysorbate Tween® series including Tween® 20, 40, 60 and 80 were investigated on human red blood cells as an indication of their membrane disrupting potentials. The results showed that all four surfactants were hemolytic to different extents such that the maximum hemolysis occurred at 1.6, 40, 80 and 8G mM by Tween® 20, 40, 60 and 80 respectively. This indicates that Tween® 20 is much more damaging to biological membranes than the other three surfactants. The effect of temperature on hemolysis induced by surfactants was also examined, and it was found that an increase in temperature from 25 to 37degreeC resulted in a significant increase in hemolysis. When 2 mM Tween® 80 was added to the solution of Tween® the hemolysis reduced from 75% to about 1% which shows a mitigating effect for Tween® 80 on the hemolysis caused by Tween®20