Silibinin (SB), a flavonoid isolated from the milk thistle, Silybum marianum, has been shown to exhibit protective effects against skin damage. The objective of the present study was to investigate the effect of topical application of SB on levels of stromelysine 1 (STM1) gene expression, acetyl hexoseamines and collagen production during skin wound healing. Full-thickness skin wounds were topically treated with 10% and 20% SB extract in acetonitril:olive oil (AOO) (4:1) for 30 days, and expression level of STM1 transcript, n-acetyl glucoseamine (NAGLA), n-acetyl galactoseamine (NAGAA) and collagen contents were analyzed on the 10th, 20th and 30th days post wounding. SB in dose- and time-dependent manner accelerated wound closure time and increased levels of STM1 mRNA, hydroxyproline, NAGLA and NAGAA compared to the untreated and vehicle (AOO)-treated rats. The current study provides evidence that SB, by increasing STM1 gene expression and extracellular matrix constituents including glycosaminoglycans and collagen contents, promotes a faster wound healing process and can be used as a healing agent in future.