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The study investigated in situ biosynthesis of albumin capped 5-fluorouracil (5-FU) loaded gold nanoparticles (NPs) using bacterial extract for enhanced efficacy against MCF-7 and in silico prediction using a GastroPlus® software. The optimized formulations were characterized for morphology, size, zeta potential, drug loading (%DL) and entrapment (%EE), compatibility, in vitro drug release, in vitro hemolysis, cellular toxicity and apoptosis studies. The results exhibited highly dispersed albumin capped mono-metallic stable NPs. Spherical size, negative zeta potential and polydispersity index were in range of 38.25–249.62 nm, 18.18–29.87 mV and 0.11–0.283, respectively. F11, F7 and F3 showed a progressive increase in %DL and %EE with increased concentration of the cellular lysate (100% &gt; 50% &gt; 10%). The drug release was relatively extended over 48 h as compared to drug solution (96.64% release within 5 h). The hemolysis result ensured hemocompatibility (&lt;14%) at the explored concentration. The biogenic F11 was more cytotoxic (81.99% inhibition by F11 and 72.04% by pure 5-FU) to the MCF-7 cell lines as compared to others which may be attributed to the preferential accumulation by the tumor cell and capped albumin as the source of energy to the cancer cells. Finally, GastroPlus® predicted the key factors responsible for improved pharmacokinetics parameters and regional absorption from various segments of human intestine. Thus, the approach can be more efficacious and suitable to control breast cancer when administered transdermally or orally.

5-Fluorouracil Loaded Biogenic and Albumin Capped Gold Nanoparticles Using Bacterial Enzyme—In Vitro-In Silico Gastroplus® Simulation and Prediction