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We confined the formation and characterization of heterogenous nano-catalysts and then used them to produce biodiesel from the novel non-edible seed oil of <i>Prunus aitchisonii</i>. <i>P. aitchisonii</i> seeds’ oil content was extracted at about 52.4 ± 3% with 0.77% FFA. Three different heterogenous nano-catalysts—calcined (CPC), KPC, and KOH-activated <i>P. aitchisonii</i> cake Titanium Dioxide (TiO₂)—were synthesized using calcination and precipitation methods. The mentioned catalysts were characterized through XRD, SEM, and EDX to inspect their crystallin dimension, shape, and arrangement. Titanium dioxide has morphological dimensions so that the average particle size ranges from 49–60 nm. The result shows that the crystal structure of TiO₂ is tetragonal (Anatase). The surface morphology of CPC illustrated that the roughness of the surface was increased after calcination, many macropores and hollow cavities appeared, and the external structure became very porous. These changes in morphology may increase the catalytic efficiency of CPC than non-calcined <i>Prunus aitchisonii</i> oil cake. The fuel belonging to PAOB stood according to the series suggested by ASTM criteria. All the characterization reports that <i>P. aitchisonii</i> is a novel and efficient potential source of biodiesel as a green energy source.