Proteins can be encapsulated in niosomes as they are known to protect proteins against the surrounding environment. Niosomes of Span 65/cholesterol/pluronic F -127 were prepared by thin film hydration method. Insulin and lysozyme were chosen as model proteins. Niosomes were characterised for morphology by Transmission Electron Microscopy (TEM) and vesicles size using Dynamic Light Scattering. The entrapment efficiency of protein in niosomes was determined by complete vesicle disruption using 50:50% isopropanol:buf fer, followed by analysis of the resulting solutions by HPLC method. Thermal behaviour of the niosomes was investigated using Differential Scanning Calorimetry (DSC). Protection of proteins against simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) were also assessed. The results showed that niosomes prepared with different molar ratios % of Span 65, cholesterol and pluronic F-127 successfully produced with insulin and lysozyme. For insulin containing niosomes, the ratio % of 64.7 (Span 65): 32.3 (cholesterol): 3.0 (pluronic F - 127) produced the highest protein encapsulation efficiency and the smallest vesicle size of 653.6 nm. For lysozyme containing niosomes, the maximum protein encapsulation was found in 72.75/24.25/3.00% molar ratio of Span 65/cholesterol/pluronic F -127 niosomes with vesicle size of 627.3 nm. The release study of proteins from the niosomal preparations in simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) revealed that insulin and lysozyme efflux from the niosomes is a biphasic process. The results indicate that Span 65 niosomes could be developed as controlled release dosage forms for delivery of peptides and proteins such as insulin and lysozyme.