Improved fermentation strategies in a bioreactor for enhancing poly(3-hydroxybutyrate) (PHB) production by wild type Cupriavidus necator from fructose

Abstract

Poly(3-hydroxybutyrate) (PHB) belongs to the family of polyhydroxyalkanoates, biopolymers used for agricultural, industrial, or even medical applications. However, scaling up the production is still an issue due to the myriad of parameters involved in the fermentation processes. The present work seeks, firstly, to scale up poly(3-hydroxybutyrate) (PHB) production by wild type C. necator ATCC 17697 from shaken flasks to a stirred-tank bioreactor with the optimized media and fructose as carbon source. The second purpose is to improve the production of PHB by applying both the batch and fed-batch fermentation strategies in comparison with previous works of wild type C. necator with fructose. Furthermore, thinking of biomedical applications, physicochemical, and cytotoxicity analyses of the produced biopolymer, are presented. Fed-batch fermentation with an exponential feeding strategy enabled us to achieve the highest values of PHB concentration and productivity, 25.7 g/l and 0.43 g/(l h), respectively. The PHB productivity was 3.3 and 7.2 times higher than the one in batch strategy and shaken flask cultures, respectively. DSC, FTIR, 1 H, and 13 C NMR analysis led to determine that the biopolymer produced by C. necator ATCC 17697 has a molecular structure and characteristics in agreement with the commercial PHB. Additionally, the biopolymer does not induce cytotoxic effects on the NIH/3T3 cell culture. Due to the improved fermentation strategies, PHB concentration resulted in 40 % higher of the already reported one for wild type C. necator using other fed-batch modes and fructose as a carbon source. Thus the produced PHB could be attractive for biomedical applications, which generate a rising interest in polyhydroxyalkanoates during recent years.

PHA granule formation and degradation by Cupriavidus necator under different nutritional conditions

“…Quantitative estimation of PHA according to previous studies was performed [37,39]. The PHA content was calculated as the PHA percentage of the dry cell weight.…”

Evaluation of Fed-Batch Fermentation for Production of Polyhydroxybutyrate With a Banana Pulp Juice Substrate From an Agro Industrial By-Product

“…In this research, a roughly constant level of nitrogen was maintained; this may have prevented the C/N imbalance necessary for high PHB yields, or PHB synthesis may have been inhibited by the presence of nitrogen (Wang and Yu, 2007;Nygaard et al, 2021). The C/N ratio may explain the non-detectable levels of PHB in fermentations with fructose concentrations of 30 and 40 g/L.…”

“…A possible explanation for these results is the rapid consumption of fructose in the culture medium, which stimulates consumption of the polymer by the bacterial metabolism (Poomipuk et al, 2014). For fed-batch fermentations, PHB productivity was increased by replacing the single sugar supplement with several pulses of added sugar to maintain a relatively stable sugar concentration throughout the fermentation (Nygaard et al, 2021).…”

“…Polyhydroxybutyrate (PHB), the most common biodegradable homopolymer of polyhydroxyalcanoates (PHA) (Maity et al, 2020), has numerous applications in food, agriculture, medicine and other industries (Li et al, 2017;Sirohi et al, 2020). It naturally accumulates intracellulary in a large number of microorganisms (Oliveira et al, 2004;Mohapatra et al, 2020;Mostafa et al, 2020;Pati et al, 2020;Penkhrue et al, 2020;Nygaard et al, 2021). Some bacteria synthesize PHB as part of the primary metabolism; others, such as Cupriavidus necator, require a nutritional imbalance in the culture media to activate the synthesis metabolism (Grousseau et al, 2014;Getachew and Woldesenbet, 2016;Amini et al, 2020).…”

“…The major limitation for commercial use of biopolymers is the high cost of production (Li et al, 2017;Li and Wilkins, 2021). For this reason, several studies have focused on the use of low-cost raw materials from agricultural residues (Sirohi et al, 2020) and fermentation methods that favor competitive production (Shivakumar, 2012;Sallau et al, 2018;Sukruansuwan and Napathorn, 2018;Dalsasso et al, 2019), such as fed batch fermentation processes, which increase the productivity of PHB (Xia et al, 2017;Wilkins, 2020, 2021;Nygaard et al, 2021).…”