# Valorization of Agri-Food Waste into PHA and Bioplastics: From Waste Selection to Transformation
## 3 Production of Bioplastics from Volatile Fatty Acids
### Accumulation of PHA Fermentation Conditions
Finally, two nitrogen sources, ammonium sulfate, and urea, were tested, and no significant differences in biomass production or PHBV accumulation were observed between them. The maximum PHA accumulation and PHA yield on biomass were 1.5 g/L and 56%, respectively. The accumulated PHA was found to be PHBV, with the percentage of hydroxybutyrate ranging from 91-96%. It is also noteworthy that PHA can be obtained through biological methods, although the carbon source can be synthetic (such as glucose) or derived from residual waste or by-products. For instance, as [PERSON] et al. [148] observed, in the case of lignocellulosic materials intended for PHA synthesis, pretreatments (physical, chemical, or biological) are necessary to eliminate lignin and render cellulose and hemicellulose accessible for the subsequent saccharification process. Furthermore, the generation of by-products, including furfural, vanillin, hydroxymethylfurral (HMF), and acetate, during pretreatment has been observed to exert an inhibitory effect on microbial fermentation, particularly on cell growth and PHA production. In the same vein, [PERSON] and [PERSON] [149] observed that higher biomass accumulation and PHB yields occurred when glucose was used as a carbon source as opposed to xylose and arabinose, and that yields were higher when pure glucose was used than mixed sugars. [PERSON] et al. [150] found that in the case of other wastes (fruit pulp waste), it has been demonstrated that the use of real waste as a substrate can result in the presence of impurities that are more difficult to remove from PHA. However, from an environmental, technological, and economic point of view, [PERSON] [151] stated that the use of agro-industrial waste in the production of PHA is positive, as it creates an alternative to the use of a by-product which in many cases has a high environmental impact or requires high economic investment for its treatment. It also promotes green technologies in the biopolymer industry, particularly PHB, by reducing production costs and adding value to a polluting raw material. In addition to _C. necator_, other microorganisms have been shown to synthesize PHAs For instance, _Bacillus megaterium_ has been found to possess a robust expression system and a lipopolysaccharide-deficient cell wall, making it an excellent microorganism for this purpose [152]. [PERSON] et al. [153] recently described the production of PHBV, with this bacterium using biogas digestate as the fermentation substrate. In this study, the Colombian bacterium strain _B. megaterium_ LVN01 was utilized for fermentation in bioreactors with a working volume of 3 L and agitation of 400 rpm, operated in batch mode. The pH was adjusted to 7.0 \(\pm\) 0.2, adjusted with HCl or NaOH 1M as needed. The temperature was 30.8 \({}^{\circ}\)C, and the fermentation time was 60 h. The optimal PHB yield was produced at 16 h, yielding 2.7 g PHB/g cell and a DCW (dry cell weight) of 0.133 g/L. However, the maximum DCW values were reached at 60 h, at 0.563 g/L, although the highest PHB production value was attained at 16 h, at 360 mg PHBV/L. The utilization of biogas digestate as a substrate resulted in an enhancement of accumulation efficiency during the initial phases (16 h), which can be ascribed to the stress conditions imposed by the complexity of the substrate [154]. In the context of such substrates, _B. megaterium_ experiences a prolonged acclimatization period prior to attaining optimal growth. This adaptation time must be taken into account when designing and optimizing fermentation processes, as it impacts the productivity and efficiency of the process [155]. In this study, _B. megaterium_ LVN01 has demonstrated its efficiency in producing PHBV from biogas digestate in a short fermentation period. Another PHA-producing microorganism is _Haloferax mediterranei_, an extremely halophilic marine archaeon. In the study by [PERSON] et al.