Pure ZnO nanoparticles exhibit intrinsic antibacterial activity against a broad spectrum of bacteria, including Gram-positive and Gram-negative strains. This activity primarily stems from their ability to generate reactive oxygen species (ROS) and release zinc ions (Zn²⁺) [8] . Research indicates that the effectiveness of pure ZnO nanoparticles is often size-dependent; smaller nanoparticles typically show enhanced activity due to their larger surface area and increased number of active sites [6] . For example, facilely synthesized ZnO nanosheets demonstrate potent antibacterial activity against common pathogenic bacteria, suggesting their potential as effective antimicrobial agents [9] . Furthermore, ZnO nanoparticles show significant antibacterial potential against highly resistant strains like ESBL-producing Escherichia coli and Klebsiella pneumoniae [10] . Sonochemically grown ZnO nanorods also exhibit considerable antibacterial properties, highlighting ZnO’s intrinsic toxicity of ZnO to bacterial cells [8] .

The differential susceptibility between Gram-positive and Gram-negative bacteria to ZnO nanoparticles is influenced by their distinct cell wall structures. Gram-negative bacteria possess an outer membrane composed of lipopolysaccharides (LPS) that can hinder the penetration of nanoparticles and ions, whereas Gram-positive bacteria have a thick peptidoglycan layer that may offer different resistance mechanisms. These structural differences affect the efficacy of ROS penetration, Zn²⁺ influx, and physical disruption by ZnO nanostructures.

The shape and crystallographic facets of ZnO nanostructures critically modulate their antibacterial efficacy. Different morphologies—such as nanorods, nanosheets, nanospheres, nanowires, and quantum dots—possess varying surface energies, surface areas, and exposed reactive sites. These factors directly influence interactions with bacterial cells and the ability to generate ROS. Nanosheet structures, with their high surface area and exposed basal planes, consistently show excellent antibacterial performance. For instance, green-synthesized ZnO nanosheets from banana peel extract exhibit notable antibacterial activity, emphasizing morphology’s role in enhancing bioactivity [11] . Similarly, ZnO nanosheets synthesized by other methods demonstrate superior antibacterial efficacy, often attributed to their large surface-to-volume ratio and numerous active sites for ROS generation and direct physical interaction with bacterial membranes [9] . Comparative studies between nanosheet and nanosphere morphologies reveal that shape significantly governs both photocatalytic and antibacterial properties. Nanosheet-dominated structures typically offer more active sites, leading to enhanced ROS generation and consequently superior antibacterial performance compared to nanospheres [12] . This highlights that engineering ZnO morphology at the nanoscale is a crucial strategy for optimizing antimicrobial potential. The high aspect ratio of nanorods can also facilitate mechanical damage to bacterial membranes, contributing to their antibacterial action [8] . Additionally, exposed crystal facets (e.g., polar vs. non-polar surfaces) possess different surface energies and charge distributions, influencing the adsorption of bacterial components and catalytic reactions for ROS production.

Modifying ZnO with noble metals represents a key strategy to enhance its antibacterial efficacy. Noble metals like silver (Ag), platinum (Pt), and palladium (Pd) possess intrinsic antimicrobial properties and can act as cocatalysts, improving charge separation and photocatalytic activity in semiconductors [13] . Silver (Ag) modification involves Ag nanoparticles, which are powerful antimicrobials. Integrating them with ZnO typically yields a synergistic effect. Ag/ZnO composites often exhibit enhanced antibacterial activity due to combined Ag⁺ ion release and improved ROS generation from ZnO. Silver acts as an electron sink, facilitating electron-hole pair separation in ZnO, increasing charge carrier lifespan and promoting ROS production [14] . Platinum (Pt) and Palladium (Pd) modification similarly enhance ZnO’s photocatalytic activity by acting as electron traps, reducing electron-hole recombination and boosting ROS generation efficiency. These noble metals also catalyze reduction reactions. Pt and Pd coatings or doping can improve ZnO nanoparticle stability for long-term applications [15] . Their presence can also alter the surface charge of composite and hydrophobicity, promoting stronger interactions with bacterial cell membranes.

Beyond noble metals, doping ZnO with metal ions or forming composites with other metal oxides and carbon-based materials offers another powerful approach to tailor and enhance antibacterial properties. These modifications can alter electronic band structure, increase defect sites, enhance surface area, or provide additional antimicrobial mechanisms. Metal doping, such as with transition metals like cobalt (Co) or neodymium (Nd), shows significant promise. For example, Co-doped ZnO quantum dots exhibit high-performance antibacterial activity primarily driven by enhanced superoxide anion (O₂∙⁻) generation [8] . Similarly, Nd-doped ZnO nanoparticles demonstrate improved antibacterial effects against ESBL-producing strains, indicating rare-earth element doping can boost intrinsic antimicrobial of ZnO potential against resistant bacteria [10] . Doping introduces lattice defects and alters the local electronic environment, creating active sites for ROS generation or enhancing metal ion release. Metal oxide composites, such as Cu₂O/ZnO and TiO₂/ZnO, combine ZnO with other metal oxides to create heterojunctions that improve charge separation and expand light absorption range, enhancing photocatalytic and antibacterial efficiency. CuO/ZnO composites leverage copper oxide’s inherent antimicrobial properties, creating synergistic effects through the p-n heterojunction between CuO and ZnO, which facilitates charge transfer and enhances ROS generation and the release of both Zn²⁺ and Cu²⁺ ions [16] . TiO₂/ZnO composites utilize titanium dioxide’s photocatalytic properties, showing improved photoefficiency and broader-spectrum antibacterial activity compared to individual components [17] . Integrating ZnO nanoparticles with graphene oxide (GO) or reduced graphene oxide (rGO) is highly effective, as GO and rGO offer excellent conductivity, high surface area, and inherent antibacterial activity. ZnO/GO or ZnO/rGO composites benefit from enhanced charge separation, where GO/rGO acts as an electron acceptor, promoting efficient electron-hole pair separation in ZnO and increasing ROS production [18] . They also provide increased surface area for more active sites and bacterial cell interaction, along with direct antibacterial activity from sharp GO nanosheet edges that physically damage bacterial membranes and induce oxidative stress. The composite leverages these multiple mechanisms for superior antibacterial efficacy, making it a candidate for advanced biomedical and environmental applications.

The antibacterial activity of nano ZnO is multifaceted, primarily involving two main pathways: reactive oxygen species (ROS) generation and zinc ion (Zn²⁺) release, often complemented by direct physical damage.

Under light irradiation (especially UV, but also visible light due to defects), ZnO acts as a photocatalyst. Photon absorption excites electrons from the valence band to the conduction band, creating electron-hole pairs (e⁻/h⁺). These charge carriers react with adsorbed oxygen and water to form ROS such as superoxide radicals (O₂∙⁻), hydroxyl radicals (∙OH), and hydrogen peroxide (H₂O₂) [8] . In dark conditions, surface defects and oxygen vacancies in ZnO can also mediate ROS generation, though typically at a lower rate.

Zn²⁺ release occurs gradually, especially in slightly acidic environments or upon contact with bacterial exudates. Released Zn²⁺ ions disrupt cell membrane integrity by interacting with thiol groups in proteins, leading to enzyme inactivation, and cause leakage of intracellular components. Additionally, direct physical interaction, particularly with high-aspect-ratio morphologies like nanosheets or nanorods, can mechanically damage bacterial cell walls [8] [9] .

The relative contribution of each mechanism depends on factors such as nanoparticle size, morphology, concentration, surface properties, environmental conditions, and bacterial type.

While ZnO nanoparticles exhibit strong antibacterial properties, their potential cytotoxicity to mammalian cells remains a critical concern for clinical and environmental applications. Studies have shown that ZnO nanoparticles can induce oxidative stress, inflammatory responses, and cell death in human cells, depending on concentration, exposure time, particle size, and surface functionalization [19] . Factors such as high Zn²⁺ release and ROS generation—which contribute to antibacterial activity—may also cause damage to host tissues. Therefore, balancing antibacterial efficacy with biocompatibility is essential [20] . Surface modification, doping, and composite formation can modulate toxicity profiles by controlling ion release kinetics and reducing nonspecific interactions with mammalian cells [21] . Further research into safe-by-design approaches and long-term toxicological assessments is necessary to ensure the safe application of ZnO-based nanomaterials [22] .