In summary, we showed that BSA NPs with size smaller than 200 nm could be prepared in BmimCF3SO3 mediated system. It showed that zebrafish body is able to eliminate BSA NPs without any toxic side effects and encapsulation of Chl into NPs reduced the toxicity of free Chl. Furthermore, in vivo toxicity assessment of BSA NPs obtained in the IL system was conducted in the zebrafish animal model. On the other hand, cellular uptake of Chl loaded BSA NPs caused a significant MCF7 breast cancer cell death, whereas free Chl and unloaded BSA NPs did not have a significant effect on the cell viability. Monitoring the drug release by UV–Vis spectroscopy revealed a burst release at first 4 h, but two-thirds of drugs stayed with NPs upon diffusion method. Drug loading increased as increasing the ratio of Chl incubated with BSA NPs. The obtained BSA NPs have been used in drug loading and release studies with a hydrophobic anticancer drug chlorambucil (Chl). Bovine serum albumin (BSA) proteins dissolved in water were transformed into BSA NPs in a water/ Triton™X (TX-100), 1-butanol/1-butyl-3-methylimidazolium trifluoromethanesulfonate (BmimCF3SO3) microemulsion-like system by using a high-speed homogenizer and crosslinker glutaraldehyde. ILs are considered to be green and designer solvents with unique properties that can replace organic solvents in the synthesis of albumin NPs. Here, we prepared serum albumin NPs in an ionic liquid (IL) system. Different methods applied for the preparation of serum albumin NPs mostly used organic solvents. Serum albumin has been a preferred protein to generate biodegradable and non-toxic nanoparticles (NPs) for drug delivery applications. Synthesis of albumin nanoparticles in a water-miscible ionic liquid system, and their applications for chlorambucil delivery to cancer cells
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