SEM and TEM studies were performed to study the surface morphology. Results of these studies are shown in Figures ​Figures44 and ​and5,5, respectively. These results confirmed that particles have

smooth surface and spherical shape. Figure 4 SEM images of nanoparticles. Figure 5 TEM images of nanoparticles. Table 1 Particle Inhibitors,research,lifescience,medical size, polydispersity, and entrapment efficiency of different batches of nanoparticles. Values are given as means ± SD (n = 3). One of the important goal of the present study was to achieve higher encapsulation of BSA in nanoparticles by employing minimal amounts of polymer (PLGA 85:15). Nanoparticles were prepared by employing two different ratios of protein: PLGA (1:5 and 1:10). BSA entrapment in nanoparticles was more than 65% in both cases (Table 1). This data clearly shows a significant entrapment of BSA in PLGA matrix. As the amount of PLGA was increased to prepare nanoparticles, entrapment of BSA in nanoparticles Inhibitors,research,lifescience,medical was enhanced as well. This could be attributed to enhanced hydrophobic interactions of BSA in HIP complex with PLGA polymer. Due to these hydrophobic interactions, partition of BSA (in HIP complex form) in the

polymeric matrix of PLGA was also significantly enhanced. The effect of HIP complexation and nanoparticle preparation on secondary structure Inhibitors,research,lifescience,medical of BSA was evaluated by CD spectra. Weak physical interactions such as electrostatic interactions, hydrogen bonds, Van-der-waals forces, and hydrophobic Inhibitors,research,lifescience,medical interactions stabilize secondary structure of the protein. During HIP complex formation, DS interacts extensively

with BSA which involves abovementioned forces. So, it is quite possible that DS has altered the native conformation of BSA. Similarly, Inhibitors,research,lifescience,medical during nanoparticle preparation, powder form of BSA-DS complex was sonicated in presence of organic solvents. These processes could possibly denature BSA. CD analysis was performed to understand the impact of these formulation factors on secondary structure of BSA. Freshly prepared BSA in 10mM Na2HPO4 solution was selected as control. Figure 6 depicts the CD spectra of standard BSA solution, BSA obtained from dissociation of HIP complex, and BSA released from both batches of nanoparticles. Results clearly show a significant overlap in peak shape throughout the EPZ5676 solubility dmso region studied. This data also confirms that the secondary structure of BSA was not perturbed due to HIP complexation or treatment others with organic solvent and sonication. Enhanced stability of BSA towards organic solvents and sonication may be explained by the following reasons. First, HIP complexation might have provided conformation stability and steric shielding to the BSA molecule. Moreover, with S/O/W emulsion method, the probability of protein denaturation has been significantly minimized compared to conventional method such as W/O/W emulsion method.