# 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  
The fermentation conditions will vary depending on the specific microorganism involved, the composition of the fermentation medium, and the type of PHA being produced. This section focuses on _Cupravidus necator_, a bacterium that is used for the synthesis of PHBV from volatile fatty acids [132]. _Cupravidus necator_ (previously known as _Wautersiautropula_, _Ralstoniaeturopha_, and _Alcaligenes autropilus_) is a Gram-negative bacterium that belongs to the Burkholderiaceae family [143]. The wild-type C. _necator is_ capable of producing PHBV from an artificial mixture of VFA plus CO\({}_{2}\) and H\({}_{2}\), as well as VFAs from digestate [144]. Furthermore, an additional variant of C. _necator_ has been identified by [PERSON] et al. [122]., which is recombinant or modified and capable of utilizing an alternative \begin{table}
\begin{tabular}{c c c} \hline \hline
**Substrate** & **Bacteria Involved** & **Type of PHA** \\ \hline \multirow{8}{*}{Simple sugar} & _Alcaligenes latus_, & -P3 HB \\  & _Ralstonia eutropha_, & -Adding organic acids: P(3 HB-3 HV), \\  & _Haloferax mediterranei_, & P(3 HB-4 HB) \\  & _Azotobacter vinelandii_ & -P3 HHx, P3 HO, and other medium chain PHA \\ \cline{2-3}  & _Pseudomonas citronellosis_ & (mcl-PHA) \\ \cline{2-3}  & _Aeromonas hydrophilia_, & -Adding organic acids or alcohols: \\  & _Aeromonas scruiae_, & P(3 HB-co-3 HHx), P(3 HB-co-3 HO) and other \\  & _Rlodospirillum rubrum_ & scl-mcl copolymers. \\ \cline{2-3}  & _Bacillus megatherium_[140] & -PHB \\ \cline{2-3}  & _Bacillus thuringiensis_[141] & -PHBV \\ \hline \multirow{8}{*}{Tryacyylglycerols} & _Aeromonas hydrophilia_ & -P3 HB-3 HHx \\  & _Pseudomonas olevrorans_, & -P3 HHx, P3 HO and other mcl-PHA. \\ \cline{1-1}  & _Pseudomonas citronellosis_ & -P3 HHx, P3 HO and other mcl-PHA. \\ \cline{1-1} \cline{2-3}  & _Cupravidus necator_[142] & -P3 HB, P(3 HB-3 HV) \\ \hline \multirow{2}{*}{Hydrocarbons} & _Pseudomonas olevrorans_, & -P3 HHx, P3 HO, P3 HD and other mcl-PHA. \\ \cline{1-1}  & _Pseudomonas citronellosis_ & -P3 HHx, P3 HO, P3 HD and other mcl-PHA. \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Adapted from [139]. A summary of the substrates and the microorganisms that synthesize PHA. P3 HHx = poly(3-hydroxylhexanoate); P3 HO = poly(3-hydroxyoctaate); P3 HD = poly(3-hydroxydodecanoate); P3 HB = poly(3-hydroxybutyrate); P3 HV = poly(3-hydroxyvalerate). “Co” is used to indicate the copolymer. carbon source to produce PHBV. [PERSON] et al. [145] found that to facilitate the production of PHBV by using _C. necator_, it is essential to establish fermentation conditions that are conducive to this process. It is important to note that PHBV accumulation is a consequence of nutritional stress, whereby the accumulation of granules occurs when other nutrients are in short supply. For this accumulation to occur, a carbon source must remain available, and nutrients, such as nitrogen or phosphate, must be in short supply [146]. [PERSON] et al. [147] delineate the culture conditions employed in their investigation of PHBV production utilizing _C. necator_ as the producing microorganism. The strain employed was DSM 545, and the culture medium was a mixture of volatile fatty acids produced by anaerobic fermentation in a semi-continuous membrane reactor from food waste. The primary variables to be monitored and regulated were pH, C/N ratio, aeration, and nitrogen sources. The pH was maintained at 7, which is considered optimal for bacterial growth. The C/N ratio that was identified as optimal was 6, as this resulted in the greatest extent of bacterial growth and PHBV accumulation. However, a ratio of 12 was also tested, but this was ultimately excluded due to the lower biomass production, which consequently led to a reduction in the overall PHBV accumulation. The effect of aeration on the culture was also investigated, and a value of 1 vvm (volume of air per volume of medium per minute) was identified as the optimal condition.