This research achieved the successful synthesis of zinc oxide (ZnO) NPs through an eco-friendly method, utilizing the leaf extract of Leonotis ocymifolia (L.O.). This innovative approach not only highlights the potential of green synthesis but also underscores the effectiveness of natural resources in nanoparticle production. The influence of annealing temperature on the properties and performance of the synthesized ZnO NPs was evaluated by varying the annealing temperatures as follows: unannealed (000), 350 °C (350), 550 °C (550), and 750 °C (750). The XRD analysis of L.O-mediated ZnO NPs confirmed the synthesis of highly crystalline wurtzite-structured ZnO NPs, with calculated average crystallite sizes that ranged between 13.8 and 20.4 nm. The UV–Vis spectra revealed a single strong absorption peak ranging from 354 to 375 nm, and the absorption peaks red-shifted with an increase in annealing temperature. The SEM micrographs showed that annealing temperature had an effect on the morphology, particle size, and distribution, with the average particle of 53.7–66.3 nm. The BET analysis revealed that the surface area of the prepared ZnO NPs was between 31.6 and 13.2 m2/g. In addition to its significant impact on the characteristics of the L.O-mediated, annealing temperature notably boosts the L.O-mediated capacity to photodegrade Methylene blue (MB) dye. Moreover, it exhibited significant antibacterial efficacy against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The photodegradation studies under UV irradiation and in 180 min revealed 750 (71.1%) had the highest degradation efficiency compared to 000, 350, and 550. The antibacterial tests showed that 000 had greater antibacterial efficacy than 350, 550, and 750. The results from this work suggest that annealing temperature had a significant effect on the structural, morphological, and optical properties and performance of L.O-mediated ZnO NPs.