Abstract
High cell density (HCD) fed-batch cultures are widely perceived as a requisite for high-productivity polyhydroxyalkanoate (PHA) cultivation processes. In this work, a reactive pulse feed strategy (based on real-time CO2 or dissolved oxygen (DO) measurements as feedback variables) was used to control an oxygen-limited fed-batch process for improved productivity of medium chain length (mcl-) PHAs synthesized by Pseudomonas putida LS46. Despite the onset of oxygen limitation half-way through the process (14 h post inoculation), 28.8 ± 3.9 g L−1 total biomass (with PHA content up to 61 ± 8% cell dry mass) was reliably achieved within 27 h using octanoic acid as the carbon source in a bench-scale (7 L) bioreactor operated under atmospheric conditions. This resulted in a final volumetric productivity of 0.66 ± 0.14 g L−1 h−1. Delivering carbon to the bioreactor as a continuous drip feed process (a proactive feeding strategy compared to pulse feeding) made little difference on the final volumetric productivity of 0.60 ± 0.04 g L−1 h−1. However, the drip feed strategy favored production of non-PHA residual biomass during the growth phase, while pulse feeding favored a higher rate of mcl-PHA synthesis and yield during the storage phase. Overall, it was shown that the inherent O2-limitation brought about by HCD cultures can be used as a simple and effective control strategy for mcl-PHA synthesis from fatty acids. Furthermore, the pulse feed strategy appears to be a relatively easy and reliable method for rapid optimization of fed-batch processes, particularly when using toxic substrates like octanoic acid.
Highlights
The detrimental effects from accumulation of plastic waste in natural environments call for change at both the regulatory and behavioral levels [1]
Despite constant aeration set at 6 LPM and a cascading stirrer reaching its maximal value, the dissolved oxygen (DO) could not be maintained at the intended set point of 40% beyond 11 h (Figure 1B)
The low residual levels of carbon and/or NH4 detected in the culture medium at this time may have slowed the growth rate and caused storage of carbon as PHA [34]. This is supported by the observation that Xr production continued until 14 h and declined to nearly negligible values due to DO limitation, which is consistent with previous work using octanoic acid [26]
Summary
The detrimental effects from accumulation of plastic waste in natural environments call for change at both the regulatory and behavioral levels [1]. There are two main classes of PHAs defined by the carbon chain-length of the monomer subunits: (1) short chain length (scl-) PHAs, which consist of C3 to C5 monomer subunits; and (2) medium chain-length (mcl-) PHAs, which consist of C6 to C18 monomer subunits [4]. Some of these PHA polymers have properties comparable to conventional petrochemical plastics, including polyethylene and polypropylene [5], and may be suitable alternatives
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