Limited studies have exploited the silk fibroin potential as an oral drug delivery system. Herein, this study developed and characterized the polyethyleniminefunctionalized fibroin nanoparticles (PEI-FNP) and the FNP (as a reference), loaded with furosemide, a Biopharmaceutics Classification System (BCS) class IV drug, for oral application. Furosemide was incorporated into the systems by two different methods of co-condensation and adsorption. The optimal formulas possessed particles with sizes of 200 nm, positive charges of 60 mV, silk-II structured confirmed by infrared spectra, appropriate drug entrapment efficiencies of 65%, and insignificant hemolysis action at concentrations of up to 1 mg/mL. Additionally, the PEI-FNP significantly increased the drug solubility up to 1.5-folds. Kinetically, the drug adsorption processes followed the physical type and pseudo-second-order model. In the simulated gastrointestinal condition, the particles could protect the furosemide from the stomach acidic environment. Interestingly, the particles release profiles in the intestinal condition were heavily dependent on the formulation fabrication methods. Particles prepared by the co-condensation method demonstrated an initial burst release phase, followed by a sustained release phase. Conversely, the adsorption method yielded particles with an initial sustained release phase, followed by a burst release phase. Conclusively, the PEI-FNP showed much potentials in improving the oral bioavailability of furosemide. Proven by this case, PEI-FNP could be further explored to be a potential oral delivery system for BCS class-IV drugs.