## 3 Results and Discussion  
### Biodegradability Test  
Figure 2A shows methane production (expressed in mL of CH4 per kg\({}^{-1}\) of volatile solids fed) during the first phase of the biodegradability tests that corresponds to the start-up phase. Production rates were monitored every 2-3 days and results reveal a clear divergence in methane production among the different experimental conditions. On the one hand, the reactors containing the pellets (NB and OB) not only yielded the highest productivity, but also produced biogas with the largest CH4 content (Table 1). Although NB lagged behind OB during the first 21 days of operation, it surpassed OB in the final phase, reaching nearly 450 mL kg\({}^{-1}\) SV by day 26. The AD treatment showed moderate methane productivity, consistently lower than NB and OB for most of the duration of the experiment. As expected, methane production in the blank reactors was minimal, indicating negligible microbial activity in the absence of added substrates. In the second phase of the biodegradability tests (Figure 2B), biogas production was also monitored every 2-3 days, although during this phase, the blank did not produce gas. Compared to the initial phase (start-up), cumulative methane production reached a plateau sooner (particularly by day 12). This accelerated stabilization suggests that microorganisms have undergone an adaptation process, enhancing their efficiency in methane generation. Moreover, methane levels and methane content in the biogas across AD, NB, and OB are closer in magnitude, with less pronounced differences between them than observed in the first phase. This uniformity indicates that the adaptation has reduced variability between the different operational conditions, probably because microbial communities may have optimized their metabolic pathways across the three situations. Overall, these findings underscore the positive impact of NB and OB, especially during the inoculation and start-up phases (Figure 2A). The variability in methane production dur \begin{table}
\begin{tabular}{c c c c c} \hline  & **\%SV Cycle1** & **\%SV Cycle2** & **\%CH4 Cycle 1** & **\%CH4 Cycle 2** \\ \hline INITIAL & 1.84 & 2.54 & - & - \\ AD & 0.50 \(\pm\) 0.03 & 1.13 \(\pm\) 0.04 & 59 & 67 \\ NB & 0.58 \(\pm\) 0.13 & 1.21 \(\pm\) 0.18 & 66 & 67 \\ OB & 0.42 \(\pm\) 0.04 & 1.19 \(\pm\) 0.06 & 68 & 68 \\ \hline \end{tabular}
\end{table} 
Table 1: Percentage of Volatile Solids. ing the start-up phase observed in this experiment could reflect differences in the structural adaptation of granular sludge, with NB and OB pellets potentially enhancing the physical integrity or stability of microbial aggregates. However, the fading performance-enhancing role of NB and OB over time suggests that once microbial communities stabilize, other factors, such as substrate availability and metabolic uniformity, may dominate methane production, as previously reported in related research [33]. The structure of the microbial community did not show any significant differences between the experimental conditions (Figure 3). _Methanosaetaceae_, a well-known aceto-clastic methanogen, appears as a major family across all experimental conditions, being particularly dominant in the reactors filled with the OB pellets. This family relies on the  
Figure 2: Cumulative methane production over time in the biodegradability tests. (**A**) Cycle 1 and (**B**) Cycle 2. AD corresponds to conventional anaerobic digestion without fillers, NB to PLA/Carbon Black spherical fillers, OB to PLA/Graphene spherical fillers and BLANK is a digester without a substrate (water + inoculum) as control. availability of acetate in the environment and can also engage in direct interspecies extracellular electron transfer (DIET) in the presence of conductive solid surfaces [34], something that might explain the better performance of the reactors containing the conductive pellets (NB and OB), at least during the start-up phase.