Elevation of the intraocular pressure (IOP) is recognized as a risk factor for glaucoma development. Latanoprost (LAT) is a prostaglandin analog used to reduce the (IOP). Thymoquinone (TQ) is a major bioactive ingredient of Nigella sativa. The aim of this study was to develop novel liposomal drug carriers for ocular delivery of LAT, TQ and a mixture of them to investigate their IOP lowering efficacy upon subconjunctival injection in glaucoma-induced rabbit's eye. The aim of the present work extends also to study the effect of the different liposome formulations on the aqueous humor oxidative stress. Liposome samples were prepared using thin film hydration method. The physiochemical properties of the prepared drugs were characterized. The IOP was recorded for 70 rabbits using Schiotz-tonometer. Malondialdehyde (MDA), reduced glutathione (GSH), catalase (CAT) activities and total antioxidant activity of the aqueous humor were estimated. Fourier transform infrared and differential scanning calorimetric studies confirmed the interaction between the drug and the vesicles, which resulted in high drug encapsulation efficiency ≥88%. The size of the prepared liposomes was less than 10μm which make them suitable in ophthalmic applications. The sustained effect was achieved by liposome samples of Lip (LAT) and Lip (LAT+TQ) which were able to reduce the IOP significantly up to 84h. Morever, the treatment of glaucomatous rabbits with liposome formulations containing TQ in their preparation [Lip (TQ) and Lip (LAT+TQ)] greatly improved the ocular tissue-induced histopathological lesions. None of the prepared liposome formulations succeeded to improve the glaucoma-induced oxidative stress damage.