## 4 Conclusions  
By adding a solution of Fe (II) and Fe (III) salts to another solution containing tea extract, the in situ green synthesis of zero-valent iron nanoparticles (nZVI) has been successfully performed. Operating in this manner, stable iron nanoparticles were obtained without unwanted flocculation phenomena. The influence of four working variables, specifically the initial concentrations of hydrogen peroxide, polyphenols, Fe(II), and Fe(III) in solution, on the efficiency of removing two commonly used dyes in medicine, chrysoidine G and methylene blue, was analyzed. Preliminary tests indicated a significant influence of the hydrogen peroxide parameter on the overall process. A marked synergistic effect on dye removal was observed when combining the treatment involving the formation of Fe\({}^{0}\) nanoparticles with the Fenton/Fenton-like process. For chrysoidine G, the degradation of the dye begins with the cleavage of the azo group's double bond (-N=N-), followed by hydroxylation and/or hydrogenation. In a subsequent stage, the hydroxylated or hydrogenated molecule undergoes successive bond cleavages, resulting in various reaction intermediates containing oxygenated benzene rings. These intermediate compounds can undergo further oxidation with ring opening, ideally leading to complete mineralization into CO\({}_{2}\) and H\({}_{2}\)O. The use of FT-IR spectroscopy has revealed that the removal of chrysoidine G by iron nanoparticles does not involve adsorption processes, where the dye remains retained on the surface of the nanoparticles. However, the adsorption of chrysoidine before the azo group's cleavage mentioned in the previous paragraph cannot be completely ruled out. For methylene blue, adsorption onto the active sites of nZVI remarkably contributed to the removal of dye. Statistical analysis of the experimental results has confirmed that, as suggested by preliminary tests, the initial concentration of hydrogen peroxide is the factor that most significantly influences the efficiency of chrysoidine removal. Through the use of the statistical design of experiments, optimization of the dye removal process has been achieved. Under suitable conditions, it is possible to eliminate 98.9% of the initially present chrysoidine in solution, and 99.1% of methylene blue. Moreover, these conditions result  
Figure 11: Schematic representation of the reactions involved in the Fenton process that is heterogeneously catalyzed by Fe\({}^{0}\) nanoparticles. in cost reduction by reducing the required amount of the two more expensive reagents, hydrogen peroxide and Fe(II) salts, by half. The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/app14156558/s1](https://www.mdpi.com/article/10.3390/app14156558/s1)]([https://www.mdpi.com/article/10.3390/app14156558/s1](https://www.mdpi.com/article/10.3390/app14156558/s1)), Table S1. Coded and real values of the operational variables in the experimental design; Table S2. Results of the analysis of variance (ANOVA); Figure S1. Preliminary runs. Influence of H\({}_{2}\)O\({}_{2}\) and polyphenol concentration (a) and Fe\({}^{2+}\) and Fe\({}^{3+}\) (b); Figure S2. Graphical analysis of the experimental design corresponding to the removal of methylene blue: (a) Pareto factor plot; (b) Main effects plot; (c) Interactions plot; (d) Response surface plot; Figure S3. FT-IR spectra of chrysodine-G and methylene blue; Figure S4. Schematic representation of radical generation in Fenton and/or Fenton-like processes. Conceptualization, E.M.C.-C.; methodology, E.M.C.-C.; validation, E.M.C.-C., M.F.A.-F. and C.F.-G.; formal analysis, M.F.A.-F., C.R.-R. and C.F.-G.; investigation, C.R.-R., M.C.-P. and A.G.-T.; writing--original draft preparation, M.F.A.-F., C.F.-G. and E.M.C.-C.; visualization, C.R.-R., A.G.-T. and M.C.-P.; supervision, E.M.C.-C. All authors have read and agreed to the published version of the manuscript. This research received no external funding.