A novel delivery system based on self-assembled liposome from multi-layered fibrous mucoadhesive membrane has been developed to improve the bioavailability of Carvedilol (Car). This system consisted of an electrospun layer (enable self-assembly of liposome once contacting with water), an adhesive layer (prolong the retention period in the mouth) and a backing layer. SEM, DSC and FTIR were applied to characterize the fiber. The TEM and fluorescence study demonstrated the formation of self-assembled liposome when electrospun fiber encountered water. The ratio of PC to Car and the molecular weight of PVP both had a significant impact on the drug encapsulation efficiency. In vitro and in vivo adhesive tests were conducted to evaluate the bioadhesive performance of the adhesive layer. The dialysis dissolution and permeation study through porcine buccal mucosa were carried out. The electrospun fiber showed excellent drug permeation amount compared to pure Car. The drug concentration–time curves, in rabbits, of fibrous mucoadhesive membrane and Car suspension were different, and possible reasons were analyzed. The pharmacokinetic study demonstrated 154% increase in the relative bioavailability compared to Car suspension. This drug delivery system offered a novel platform for potential buccal delivery of drugs with high first-pass effect.