Investigation of ?-caryophyllene as terpene penetration enhancer: Role of stratum corneum retention

Terpenes are usually used as penetration enhancers (PE) for transdermal drug delivery (TDD) of various mole-cules. However, TDD of hydrophilic macromolecules is becoming an urgent challenge due to their potent ac-tivities. The aim of this study was to investigate the potential application of beta-caryophyllene (beta-CP), a sequiterpene, as PE for TDD of hydrophilic macromolecules for the first time. Commonly used PEs, namely azone and 1,8-cineole (1,8-CN), were applied as controls. Transepidermal water loss (TEWL) analysis revealed that the reduction of skin barrier function caused by beta-CP was reversible. Transdermal experiments showed that when skin was treated with beta-CP or azone, there was a significant permeation-enhancing effect on fluorescein iso-thiocyanate (FITC) and FITC-dextran with different molecular weight (MW) of 4k or 10k. CLSM analysis confirmed that beta-CP and azone can facilitate the penetration of FD-4k through epidermis and dermis. However, the cytotoxicity of azone against epidermal keratinocytes was significantly higher than beta-CP and 1,8-CN. Additionally, application of beta-CP and 1,8-CN didn't increase erythema index (EI) but the EI values of azone group increased significantly and irreversibly, indicating the high biocompatibility of the natural terpenes. beta-CP had better permeation-enhancing effect and higher stratum corneum (SC) retention than 1,8-CN due to its increased carbon chain length and lipophilicity, as further demonstrated by molecular dynamics (MD) simulation studies. Skin electrical resistance (SER) and attenuated total reflection fourier transform infrared spectroscopy (ATR-FTIR) studies revealed a significant interfering effect of beta-CP on SC lipids. Taken together, beta-CP exhibited significant penetration enhancement of hydrophilic macromolecules due to its SC retention and SC lipid fluid-ization ability.