The purpose of this study was to develop a pH-triggered in situ gelling vehicle for ophthalmic delivery of puerarin. The effect of hydroxypropyl-beta-cyclodextrin (HP-beta-CD) on the aqueous solubility and in vitro corneal permeation of puerarin was also investigated. The puerarin solubility increased linearly and proportionally to the HP-beta-CD concentrations and 5% (w/v) HP-beta-CD enhanced its ocular permeability significantly. Carbopol 980NF was used as the gelling agent in combination with HPMC (Methocel E4M) which acted as a viscosity-enhancing agent. The optimum concentrations of Carbopol 980NF and HPMC E4M for the in situ gel-forming delivery systems were 0.1% (w/v) and 0.4% (w/v), respectively. When these two vehicles were combined, an in situ gel that had the appropriate gel strength and gelling capacity under physiological condition could be obtained. This combined solution could flow freely under non- physiological condition and showed the character of pseudoplastic fluid under both conditions. Both in vitro release studies and in vivo pharmacokinetics studies indicated that the combined polymer systems performed better in retaining puerarin than puerarin eye drops did. These results demonstrate that the Carbopol 980NF/HPMC E4M can be a viable alternative to conventional puerarin eye drops to enhance ocular bioavailability and patient compliance.