This study continues long-standing efforts to develop protein delivery systems based on cyclodextrin-conjugated polyester in our laboratory. The crude products of ethylenediamino bridged bis(beta-cyclodextrin)-conjugated poly(DL-lactic-co-glycolic acid) were used in this study to make full use of unreacted reactant. With bovine serum albumin (BSA) as a model protein, the encapsulation effects (the encapsulation efficiency and particle size) of nanoparticles were similar to those of using pure conjugated products. Besides, a water-in-oil-in-water emulsification technique was conveniently modified. By adding polyvinyl alcohol (PVA) in the internal aqueous phase, a more stabilized emulsion was formed. Consequently, less PVA (similar to 0.05%) was needed in the outer aqueous phase and less PVA (0.14 g/g nanoparticles) remained in the nanoparticles. This modification resulted in improved encapsulation efficiency (similar to 89-94%) of BSA and an enlarged particle size (340-390 nm). Furthermore, the burst release of BSA at the 1st day was less pronounced (7-12% of the encapsulated amount) than that of nanoparticles with no PVA added in the internal aqueous phase. Degradation studies using transmission electron microscope and gel permeation chromatography suggested that the mechanism for protein release was mainly through nanoparticles erosion. (C) 2007 Wiley Periodicals, Inc.