## 4 Conclusions  
In the methanogenic phase of the TPAD, the removal of particulate matter was enhanced by the addition of nanoparticles, resulting in a 9% increase in TS removal and a 6% increase in VS removal. When combining the nanoparticles with ozone pre-treatment, these improvements were even more pronounced, with TS and VS removal reaching 27% and 24%, respectively. Additionally, the combined treatment improved total COD removal by 33% and soluble COD removal by 35%, thereby enhancing the overall efficiency of AD. Ozone pre-treatment also significantly improved VFA removal, with total acidity reduction reaching 95% in combined treatment tests, compared to 87% in control and nanoparticle-only tests.  
Regarding methane production, the addition of iron nanoparticles did not yield a statistically significant increase. However, ozone pre-treatment significantly improved methane production and increased the methane production yield by 22% (mL CH\({}_{4}\)/g VS\({}_{\text{added}}\)). The modified Gompertz and Cone models effectively described methane production, showing that ozone pre-treatment enhanced the production potential and maximum production rate while reducing the latency period from 10 days to 3 days.  
This study significantly contributes to the improvement of processes involved in TPAD by enhancing the isolated production of methane and hydrogen and optimizing the recovery of effluents rich in VFAs. Furthermore, it highlights the capability of producing highly valuable acids, such as butyric acid, which can be used as precursors of valuable chemicals, such as bioplastics. Importantly, the TPAD process can also be utilized for the treatment of other biodegradable wastes, including those with high pathogen content, such as livestock waste and sewage sludge. The implementation of a thermophilic phase ensures that the digestate obtained is free of pathogens, making it suitable for agricultural applications. In summary, these strategies enhance the production of different by-products, thereby aligning with a multiproduct biorefinery model and promoting a sustainable approach to waste management.  
Conceptualization, J.L.G.M. and E.D.D.; methodology, F.J.F.-M. and J.L.G.M.; software, J.M. and F.J.F.-M.; validation, J.L.G.M., J.M. and F.J.F.-M.; formal analysis, J.M. and F.J.F.-M.; investigation, M.E.I.L. and E.D.D.; resources, J.L.G.M., J.M. and F.J.F.-M.; data curation, M.E.I.L., F.J.F.-M. and E.D.D.; writing--original draft preparation, E.D.D. and J.L.G.M.; writing--review and editing, F.J.F.-M. and J.M.; visualization, F.J.F.-M. and J.L.G.M.; supervision, J.L.G.M., J.M. and F.J.F.-M.; project administration, J.L.G.M.; funding acquisition, J.L.G.M. All authors have read and agreed to the published version of the manuscript.  
This research was funded by the European Union within the framework of the ERDF Operational Program 2014-2020 and by the Ministry of Economic Transformation, Industry, Knowledge and Universities of the Junta de Andalucia. Project reference: FEDER-UCA18-107460, Verinsur, S.A., ZonoSistem. Ingenieria del ozono S.L. Co-financing of the Government of Spain and ERDF obtained for infrastructures and scientific-technical equipment, Call 2015 (UNCA15-CE-3476) and from Junta de Comunidades de Castilla-La Mancha Project BPLY/23/180225/000143. The authors thank the predoctoral contract of the Junta de Andalucia PREDOC-01870 (Encarnacion Diaz-Dominguez), the company Smallops for supplying the nanoparticles and the Plan Propio-UCA 2022-2023 for its assistance in co-financing publication expenses.  
Not applicable.  
The raw data supporting the conclusions of this article will be made available by the authors on request.  
The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.