# Urban Biorefinery Demonstration: Production of Polyhydroxyalkanoates from a Municipal Solid Waste
## 4 Discussion
### Accumulation of scl-PHA in Microorganisms  
The primary objective during the accumulation study was to maximize intracellular scl-PHA storage by optimizing the carbon feeding strategy. It can be seen how the scl-PHA concentration (expressed as wt.%) in the biomass exponentially grows with increasing VFA concentration (Figure 5) and once the COD feeding rate is higher than the COD consumption rate. Hence, it is observed in Table 4 that the low values of the scl-PHA storage yield show a slightly ascendant tendency as the COD feeding increases, which means that the total amount of VFAs employed during the accumulation process was not only employed for scl-PHA accumulation but also for MMC growth. [PERSON] et al. [68] studied the kinetics of stored wastewater substrates by a mixed microbial culture, they concluded that only a certain proportion of the COD present in industrial wastewater can be accumulated inside the cell. We detected that a COD feeding rate lower than 13.91 mg COD/L-min almost did not produce scl-PHA accumulated in the cells (0.05-0.12 g/L). When the COD feeding rate increased above 13.91 mg COD/L-min, a significant increase in accumulated scl-PHA in the biomass is achieved (0.18 g/L, accumulation n\({}^{\circ}\)6). Thus, we detected an increase in scl-PHA percentage storage in the biomass of 13.58 wt.% by increasing the COD feeding rate from (n\({}^{\circ}\)5) 13.80 up to (n\({}^{\circ}\)10) 24.22 mg COD/L-min. This low polymer accumulation was due to the DO cascade control. During the accumulation reaction the DO was higher than 20%, but in any case, rose up to 70%. With the selected DO cascade control, when the DO rises above 20% the aeration and/or agitation decreased keeping that DO condition. This control system turned out to be unfavorable for this kind of fermentation, causing large oscillations in aeration and agitation without providing a high DO concentration. The similar exponential tendency for the scl-PHA content in the biomass than the previous accumulation experiments 1 to 10 is observed in Figure 6. When the COD feeding rate applied is lower than the COD consumption rate in the enrichment fermenter (fermentation broth used for accumulation) the scl-PHA accumulated in the cells is low (0.66 g/L). Even though the obtained scl-PHA content in the biomass in accumulation batch n\({}^{\circ}\) 11 is higher than the obtained ones for accumulation batches from n\({}^{\circ}\) 1 to n\({}^{\circ}\) 5. Comparing Figures 5 and 6, the higher the COD consumption rate is in the enrichment fermenter, the higher is the scl-PHA storage yield. Moreover, the average scl-PHA accumulation between batches n\({}^{\circ}\) 11 and n\({}^{\circ}\) 29 (Table 5) was 20 wt.%, which is higher than the average one obtained between batches n\({}^{\circ}\) 1 and n\({}^{\circ}\) 10 (Table 4), 5.6 wt.%. Therefore, maintaining adequate aeration and agitation conditions that ensure a dissolved oxygen (DO) concentration above 70%, enhances scl-PHA accumulation by microorganisms. A precise understanding of aeration conditions in a bioprocess can significantly reduce productive costs by optimizing oxygen transfer efficiency and minimizing energy consumption. Proper aeration control ensures that microorganisms receive the necessary oxygen levels for metabolic activity, avoiding both oxygen limitations that could hinder MMC growth and excessive aeration that would lead to unnecessary energy consumption [69]. Additionally, well-regulated aeration enhances process stability, improves substrate utilization, and maximizes product yield [70]. In large-scale industrial applications, implementing advanced aeration strategies, such as dynamic oxygen supply control or real-time monitoring, can further contribute to sustainability by reducing operational costs and lowering the environmental footprint of the production process [69; 70].