This research presents a novel biological route for the biosynthesis of nickel oxide nanoparticles (NiO NPs) using marine macroalgae extract as a reducing and coating agent under optimized synthesis conditions. XRD and TEM analyses revealed that phytosynthesized NiO NPs are crystalline in nature with a spherical shape having a mean particle size of 32.64 nm. TGA results indicated the presence of marine-derived organic constituents on the surface of NiO NPs. It is found that biogenic NiO NPs with BET surface area of 45.59 m g is a highly efficient catalyst for benign one-pot preparation of pyridopyrimidine derivatives using aqueous reaction conditions. This environmentally friendly procedure takes considerable advantages of shorter reaction times, excellent product yields (up to 96%), magnetically viable nanocatalyst (7 runs), low catalyst loadings, and free toxic chemical reagents.