Observation of molecular permeation through the stratum corneum by multiphoton fluorescence microscopy

• Main Authors: Chiu-Sheng Yang, 楊秋晟
• Other Authors: 董成淵
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/16242038191544421297

碩士 === 國立臺灣大學 === 應用物理所 === 101 === Currently there are three commonly applied methods for drug delivery including oral delivery, hypodermic injection and transdermal methodologies. Unfortunately, there are some disadvantages associated with oral delivery and hypodermic injection, therefore, in this research we focused on transdermal drug delivery system that has a handful of merits over conventional methods. The drug can diffuse into the deeper subcutaneous depth in the skin tissue by the process of permeation, reach the blood capillary, and finally enter the circulatory system for the therapeutic treatment. We utilized two-photon fluorescence microscopy to study the drug permeation on stratum corneum (SC) in order to calculate the diffusion coefficient and understand the detailed permeation process in skin specimens. The two fluorescent probes including SRB (Sulforhodamine B) and RBHE (rhodamine B hexyl ester) were served as the modeling of hydrophobic and hydrophilic drug agents, respectively, and were selected based on their similarity in molecular structure, and fluorescence emission range. These two fluorescent probes were used for the purpose of comparison and were applied in the following manner. We first mixed SRB with ethanol as well as with PBS as the control group combination. Secondly, we mixed RBHE, separately, with ethanol, PBS, and 3% v/v oleic acid, and used these combinations as the experimental group. Due to the fact that oleic acid can enhance the ability of drug permeation in skin, we compared the results with and without oleic acid. Our analytic results suggested that both SRB and RBHE reached the deeper subcutaneous layer via intercellular route. In addition we discovered that the ability of permeation of RBHE in intracellular area is better than that of SRB. The addition of oleic acid not only enhances the molecule’s permeative ability but also increase its heterogeneity and the revelation of the shape of the cell. Furthermore, there are several models illustrating the physical structure of stratum corenum. In our theoretical modeling we treated stratum corenum as a homogeneous membrane. By Fick’s law and the continuous equation we can obtain a differential diffusion equation. There is a unique diffusion coefficient associated with the diffusion equation for both the permeation of SRB and RBHE in the stratum corenum of skin. For both the permeation of SRB and RBHE we combined the Matlab with Labview, analyzed every pixel of images, fitted the diffusion equation, and obtained the diffusion coefficient from the same depth at seven different time points. Finally, we obtained the distribution diagram and recognized that the value of diffusion coefficient in intercellular domain is higher than that in the intracellular region for both SRB and RBHE. The difference of diffusion coefficients between intracellular and intercellular area were observed at different depths for particular type of cells in stratum corenum, and the results showed that the diffusion coefficients for SRB and RBHE decrease with the increasing depth both intracellular and intercellular regions.