Background: The use of nanotechnology in various biomedical and pharmaceutical fields is expanding rapidly. Objectives: The aim of this study was to synthesize zinc nanoparticles using Hibiscus sabdariffa plant extract and investigate their antimicrobial properties. Methods: In this experimental study, the aqueous extract of sour tea was mixed with a 0.1 M zinc sulfate solution at a 1:1 ratio and allowed to react for 15 minutes at room temperature to produce nanoparticles. The synthesis of zinc oxide nanoparticles was confirmed using spectrophotometric methods, measuring the average diameter of nanoparticles, X-ray diffraction, and scanning electron microscopy (SEM). The minimum inhibitory concentration (MIC) of the zinc oxide nanoparticles was then measured against standard strains. Results: The results showed that the largest inhibition zone diameter was observed at a concentration of 1024 μg/mL against Listeria monocytogenes, while the smallest inhibition zone was observed against Aeromonas hydrophila. At a concentration of 512 μg/mL, an inhibition zone of 10 mm was developed against two strains: Vibrio cholerae and Escherichia coli. Conclusions: Based on the results of this study, zinc oxide nanoparticles, along with the sour tea aqueous extract and zinc sulfate, exhibited antibacterial properties. However, the most pronounced antimicrobial effects were observed with the zinc oxide nanoparticles.