Impact of Storage on In Vitro Permeation and Mucoadhesion Setup Experiments Using Swine Nasal Mucosa

Abstract

Intranasal topical administration offers a promising route for local and systemic drug delivery, with in vitro permeation and mucoadhesion studies often using porcine models. However, the impact of storage on mucosal integrity after the procedure remains unaddressed. This study aimed to standardize the preparation process and evaluated whether storage of porcine nasal mucosa impairs its integrity and permeability for experimental comparisons. Additionally, an optimized in vitro mucoadhesion experiment using texture analyzer equipment was investigated. Porcine nasal mucosa was subjected to different storage conditions ("fresh"; refrigerated at 4°C for 24 h and 48 h, and frozen at -20°C for two or three weeks) and assessed using optical and transmission electron microscopy. In vitro permeation assays were performed in a Franz-type vertical diffusion system with lidocaine hydrochloride (LDC). In vitro mucoadhesion assays were conducted using fresh nasal mucosa and a commercial nasal topical formulation using TA.XT. Plus texture analyzer. The variables involved (probe speed, contact time, and application force) in assessing mucoadhesive capacity (maximum mucoadhesive force F_max and work of mucoadhesion W_muc) were optimized using a Central Composite Design. Fresh tissues showed no alterations in histological arrangement or in the ultrastructure of adherence junctions. Stored tissues exhibited histological disorganization, reduced thickness, and loss of epithelial integrity. LDC permeability increased in storage tissues (p < 0.05). Contact force had a positive effect on F_max and W_muc (p < 0.0001), with a minimum required value of 0.48 N. Variations in contact time and probe speed did not affect the responses (p > 0.05). In conclusion, the preparation technique was adequate to maintain mucosa integrity for permeability studies. However, storing the mucosa at 4 or -20°C overestimated LDC permeation, which could mislead critical data for formulation development. Therefore, the use of fresh mucosa is recommended to ensure more reliable results. For in vitro mucoadhesion assays, a minimum contact force of 0.48N is required for optimal responses.

Graphical Abstract