Encapsulation of bioactive compounds is of interest to the food industry. Zein is an excellent film former and has been used for oil encapsulation. Preliminary research showed that zein forms self-assembled nanoscale structures that might be used as carriers to encapsulate bioactive compounds and improve their bioavailability. OBJECTIVE: The objective of this study is to investigate the effect of pH and zein purity on the morphology of zein self-assembled structures. The effect of two surfactants was also investigated for their ability to alter zein morphology. METHODS: Two commercially available zein samples (low carotenoid content [LCC], high carotenoid content [HCC], 1mg/ml) were dissolved separately in 75% ethanol/water solution at different pH levels. pH of zein solutions was adjusted by HCl Solution (pH=2.0), chloroacetic acid solution (pH=3.6), and NaOH solution (pH=12.0). Zein solution as it is, pH~6.0, was used as control for comparison. Two surfactants, oleic acid (OA) and oleylamine (OL), were added separately to zein solutions prepared above. Both of them have similar lipophilic tails, however, carrying opposite charge on their polar heads. After zein-surfactant solutions were well mixed, they were dripped onto aluminum weighing pan, carefully spread, and dried in hood at ambient temperature. The surface micro and nano structures of zein deposits were characterized by focused ion beam/scanning electron microscopy (FIB/SEM). RESULTS: The pH level of the zein system significantly determines the morphology of zein structure. At pH=2, zein formed films regardless of the presence of surfactants, while zein formed particles at pH=12. In basic solutions, the surfactants had minor influence on the zein morphology resulting in distorted zein spheres. Exception is for the HCC zein sample with OA, uneven films formed instead. The effect of OA on zein morphology seems to be significant at all pH levels, however, the effect of OL is trivial. In HCl solution, zein for