## 3 Results and Discussion
catarrhalis_) bacteria using the 96-well microtiter method to determine the MIC, as indicated in Table 1. The results in Table 1 indicate that both the mushroom extract and _Pleurrotus ostreatus_-synthesized ZnO NPs exhibit antimicrobial activity against all tested microorganisms (Gram-positive and Gram-negative). However, when comparing mushroom extract with mushroom-synthesized ZnO NPs, the study showed that mushroom-synthesized ZnO NPs had enhanced antimicrobial activity for almost all the tested microorganisms. Therefore, this means that mushroom antibacterial activity is enhanced through nanoparticle synthesis. This activity of \begin{table}
\begin{tabular}{c c c c} \hline \hline
**Microorganisms** & **Mushroom ZnO NPs** & **Mushroom Extract** & *** Control Drug** \\ \hline _P. aeruginosa_ & 0.04 & 0.04 & 0.003 \\ _E. faecalis_ & 0.08 & 1.25 & 0.006 \\ _C. albicans_ & 0.31 & 0.63 & 0.003 \\ _K. pneumoniae_ & 0.16 & 0.63 & 0.008 \\ _B. cereus_ & 0.63 & 1.25 & 0.004 \\ _P. vulgaris_ & 0.31 & 0.16 & 0.001 \\ _S. aureus_ & 0.63 & 0.63 & 0.004 \\ _E. coli_ & 0.16 & 0.63 & 0.001 \\ _M. catarrhalis_ & 1.25 & 1.25 & 0.003 \\ _M. hominis_ & 0.16 & 0.63 & 0.004 \\ \hline \hline \end{tabular}  
* Control drug: gentamycin for all bacterial strains and amphotericin for _Candida albicans_.  
\end{table} 
Table 1: Antimicrobial activity of mushroom ZnO NPs (MIC in mg/mL) against selected pathogenic microorganisms. Figure 3: Zeta potential analysis of the biosynthesized ZnO NPs. mushroom-synthesized ZnO NPs may be attributed to the fact that both Gram-positive and Gram-negative microorganisms contain a cell wall that is negatively charged, hence causing interactions of the bacterial cell wall with positively charged ions released by nanoparticles [50]. Previous researchers have also noted that the ZnO NPs interact with the plasma membrane of bacteria through electrostatic attraction; hence, membrane permeability may be disrupted through metal ions (Zn\({}^{2+}\)), resulting in damage to bacterial DNA leading to cell death [51]. The results indicate that different microorganisms have varying susceptibility towards mushroom-synthesized ZnO NPs, probably due to differences in the polarity of the cell membranes of these bacteria [52]. To further confirm possible antibacterial activity of synthesized ZnO NPs, DNA cleavage assay was conducted in our study. Figure 4 shows that mushroom-synthesized ZnO NPs potentially damage bacterial DNA in a concentration-dependent manner. Lane C1 indicates pure bacterial DNA, whereas lane C2 shows DNA treated with H\({}_{2}\)O\({}_{2}\) (completely denatured DNA), and the remainder of the lanes indicate DNA treated with mushroom-synthesized ZnO NPs at increasing concentrations of 3.0, 6.25, 12.5, 25.0, 50.0, and 100 \(\upmu\)g/mL. It can be noted from Figure 4 that mushroom-synthesized ZnO NPs were able to digest/damage bacterial DNA; hence, higher concentrations of ZnO NPs, such as 50 and 100 \(\upmu\)g/mL, degraded the DNA structure. This observed DNA cleavage mechanism may be responsible for the antibacterial effects attained in other microorganisms. As expected, other researchers also found that DNA digestion can potentially be considered as an antimicrobial mechanism employed by nanoparticles [53]. These results suggest that ZnO NPs may compromise membrane permeability of bacteria, therefore causing Zn\({}^{2+}\) to enter the bacterial cytoplasm and causing damage to bacterial DNA, as reported by [PERSON] et al. [51]. The cytotoxicity of mushroom-synthesized ZnO NPs and mushroom extract was evaluated in vitro using HepG2 and Hek293 cell lines, which are the two frequently used cell lines for cytotoxicity testing. These cell lines were selected due to the vital role of liver cells in cellular metabolism and glucose homeostasis. In addition, the liver also detoxifies xenobiotics (including drugs) to water-soluble metabolites for excretion by the kidney [54]. The conversion of the yellow tetrazolium salt into purple  
Figure 4: The gel electrophoresis image of DNA cleavage of _P.