Cutaneous microdialysis. Methodology and validation.

This thesis describes the methodology and validation of cutaneous microdialysis for the study of skin penetration of various topically applied substances in experimental dermatological research. Microdialysis is a sampling technique which makes it possible to measure substances in the extracellular water space in human and animal skin in vivo. A microdialysis probe, i.e. a tubular semipermeable membrane connected to afferent and efferent tubings, is placed in the dermis and perfused. Substances from extracellular space may diffuse through the pores of the membrane and be collected in the dialysate for further analysis. Glucose, sodium fusidate, betamethasone 17,21-dipropionate and calcipotriol were chosen as model substances and were investigated by in vitro microdialysis. The perfusion rate, the length of the membrane, stirring rate and temperature influenced recovery of the substances. Lipophilic compounds tend to have low recoveries and differ in recovery and loss. Insertion of the microdialysis probe causes a trauma in the skin. Rat and human skin were studied in vivo. Increase in skin blood flow, erythema and skin thickness were demonstrated by laser Doppler perfusion imaging, Dermaspectrometer colorimetry, Minolta Chromameter colorimetry and ultrasound imaging of cross-sectional skin structure. In addition histamine was released in rat skin due to the needle insertion. An equilibration period of minimum 90 min in human skin and 30 min in rat skin after the insertion is necessary to allow the effects of trauma to diminish. To obtain measurable concentrations in the dialysate in rats treated topically with the lipophilic drug betamethasone 17-valerate, unrealistic high doses and penetration enhancement were required. The highly protein-bound drug fusidic acid was not measurable in the dialysate after topical application, probably due to very low concentrations of free diffusible drug. Measurable concentrations were only observed after high doses of oral administrations of fusidic acid. Calcipotriol could not be detected in the dialysate. The microdialysis technique is probably primarily useful for the study of hydrophilic substances and substances with low protein binding and low molecular weight. However, application of cutaneous microdialysis for the study of lipophilic substances need further methodologically development.