Research On Transdermal Absorption And Local Toxicity Of Triptolide Peptide Derivative

Triptolide(TP),an active compound isolated from Tripterygiumwilfordii Hook F(TWHF),displays multiple bioactivities such as anti-inflammatory and immunosuppressive activities.However,TP displays diverse side effects on various organs,which limits its clinical application.Therefore,improving the dosage form of TP is of great significance to reduce its toxicity.But TP can cause significant skin irritation,which hinders the development of its transdermal formulations.It is previously reported that the significant toxicity of TP is caused by the presence of its C14-OH.Thus,this research aims to using hepta-arginine(R7)to modify the C14-OH of TP to reduce its skin toxicity while examining its possibility of its transdermal administration,and the conjugate product is named TP-S-S-CR7.In vitro and in vivo experiments were conducted to test dermal toxicity of TP-S-S-CR7.It was demonstrated that,at concentrations of 1 and 0.1 ?M,TP-S-S-CR7 displayed less toxicity on the immortal human keratinocyte(HaCaT)cell line(P<0.01).The results of in vivo skin irritation test showed that severe dermal toxicity appeared on both stratum corneum and dermis after smearing TP on the belly skin of guinea pigs for 7 days successively.However,there was no obvious histopathological changes in the TP-S-S-CR7 treated group.The results of in vivo experiment showed that after transdermal administration of TP and TP-S-S-CR7 for 6 h,the concentration of TP in TP administration group went to stable,at the same time,that in TP-S-S-CR7 administration group increased with a steady rate.TP could not be detected in rat plasma the TP administered group within 72 h.In the TP-S-S-CR7 administration group,a relatively low concentration of TP could be detected,which further proved the penetration enhancing ablility of TP-S-S-CR7.In this study,a bio-stable fluorescein rhodamine(Rh b)derivative and R7 covalently linked compound RHB-S-S-CR7 was used to observe the fluorescence in different layers of the skin at different times after transdermal administration to speculate the percutaneous route of R7.After 2 h of transdermal administration of RHB-S-S-CR7,the fluorescence could be observed mainly in hair follicle,and then after 8 h,fluorescence could be seen in the deeper layer of skin.Based on such result,it could be speculated that the transdermal route of R7 is via hair follicle.In order to further improve the transdermal penetration ability of TP,peptides were designed and synthesized instead of R7.A series of liner and cyclic peptides were designed and synthesized based on TD-34(ACSSKKSKHCG,N-1 and N-2 amino acids were substituted by Lysine,Tryptophan or Arginine),and their transdermal enhancing ability was investigated by in vitro penetration experiment using TP as model drug.Result showed that amidated peptides have better transdermal enhancing activity(P<0.05).All together,in this research,TP-S-S-CR7 was synthesized,which could reduce dermal toxicity of TP,improved the transdermal accumulative penetration amount of TP at the same time.Cyclicpeptide DLCC-2 and the C-terminal amidated linear peptide DL-2,which has the advantage of simple structure,and good transdermal penetration enhancing activity.Such results have laid a solid foundation for further TP toxicity reduction and transdermal enhancing investigation.