which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited.The development of green procedure for the synthesis of gold nanoparticles (AuNPs) has gained great interest in the field of nanotechnology. Biological synthetic routes are considered to be environmentally benign and cost-effective. In the present study, the feasibility of AuNPs' synthesis via intracellular protein extract (IPE) of Pycnoporus sanguineus was explored. The characteristics of generated particles of formation, crystalline nature, and morphology and dimension were analyzed by UV-vis spectroscopy, X-ray diffraction (XRD) and transmission electron microscopy (TEM), respectively. UV-vis spectra exhibited strong absorption peaks in 520 to 560 nm, indicating the formation of AuNPs. XRD analysis revealed that the formed AuNPs were purely crystalline in nature. TEM observation showed that AuNPs with various shapes including spherical, pseudo-spherical, triangular, truncated triangular, pentagonal, and hexagonal, ranging from several to several hundred nanometers, were synthesized under different conditions. The average size of AuNPs decreased from 61.47 to 29.30 nm as the IPE addition increased from 10 to 80 mL. When the initial gold ion concentration changed from 0.5 to 2.0 mM, the average size rose from 25.88 to 51.99 nm. As in the case of solution pH, the average size was 84.29 nm with solution pH of 2.0, which diminished to 6.07 nm with solution pH of 12.0. Fourier transform infrared (FTIR) analysis implied that the functional groups including hydroxyl, amine, and carboxyl were involved in the reduction of gold ions and stabilization of AuNPs. The catalysis results showed that 0.019 mg of AuNPs with average size of 6.07 nm could catalyze the complete degradation of 12.5 μmol of 4-nitroaniline within 6 min and the degradation rate increased drastically with the addition of AuNPs. All the results suggested that the IPE of P. sanguineus could be potentially applied for the eco-friendly synthesis of AuNPs.