Titolo della tesi: Development of a continuous process for the production of Polyhydroxyalkanoates (PHA) from renewable resources and occupational health and safety assessment on PHA production pilot plant in Treviso.
Introduction
PHAs are a family of polyester completely biobased and fully biodegradable. These properties make PHAs more fascinating, and they have attracted a lot of attention in recent years. The current limitation in PHA production even on large scale is the production cost related to the use of pure microbial strains. Therefore, to reduce the PHA production cost, mixed microbial cultures (MMC) are employed through a continuous process in three stages (acidogenic fermentation, biomass selection, and PHA accumulation). Among these stages, the selection or enrichment stage is the most important since it gives a competitive advantage to those bacteria with the PHA storage ability. These MMC do not require sterile conditions like pure cultures, allowing them to grow on less expensive substrates, lowering production costs. In most cases, even on large scale, the selection process takes place in an SBR reactor, where a series of cycles and the feast and famine regime are used to boost the selection pressure. However, on large scale, to complete these series of cycles in the SBR in a short time and to separate liquid and solid, powerful pumps and oversized solid/liquid separation units are requested, which can increase the production cost. The same drawback also applies to the following PHA accumulation which is usually performed in batch reactors.
Aims
This Ph.D. thesis mainly deals with the development of an innovative continuous process for both biomass selection and PHA accumulation.
The process developed employed two continuous reactors for the biomass selection where the feast and famine regime were applied through continuous recirculation through the two reactors. Indeed, the first reactor was a tubular reactor (PF-like column) which was continuously fed with the influent substrate (with a feeding flow rate QE) and where the feast phase was carried out whereas the second reactor was a continuous stirred tank reactor (CSTR) which only received the effluent from the first reactor (at a flowrate QR) and in which the famine phase was performed. Furthermore, the two reactors were also connected through a specific recirculation back from famine to feast reactor (same recirculation flow rate QR). In this case, the selection occurred in the space rather than the time as in the SBR.
A synthetic mixture of acetic and propionic acids (OLR = 2.12 gCOD/Ld) was used as a feeding solution with the proportion of 65 % of acetic acid and 35 % propionic acid both on a COD basis and with 35 as C/N ratio. Several experiments were conducted on this novel configuration to evaluate the effectiveness of the process under the effect of changing recirculation factor (RC = RC1, RC2, RC4, RC8, defined as a ratio between the recirculation flow rate QR and the feeding flow rate QE), and of the OLR (2.12, 4.25 and 8.5 gCOD/Ld at RC1 and RC4 specifically). Then, different setups have been designed (setup 2 to setup 5) to also test the accumulation stage coupled continuously to the selection stage. As a result, the recirculation factor really influenced the biomass selection and different PHA content was obtained at the end of the selection stage (7±1, 18 ± 1, 34 ± 2, and 9.0 ± 0.10 % gPHA/gVSS) and at the end of accumulation (18 ± 6, 34 ± 2, 40± 1, 58±5, gPHA/gVSS) working at RC1, RC2, RC4, and RC8 respectively (at OLR 2.12 gCOD/Ld). On the other hand, the OLR also influenced the PHA production, and increasing the OLR had a negative impact on the PHA storage yield. The intermediate OLR (4.25 gCOD/Ld) was revealed to be more performant than the other in both selection and batch accumulation stages at RC1 and RC4. Furthermore, by investigating different setups for the continuous accumulation (setup 1 to setup 5), the stability of the process was enhanced, giving interesting results in terms of PHA production. Setup 5, is considered the most stable condition avoiding the biomass flake formation during the accumulation in the CSTR and allowing a better air dispersion in the feast tubular reactor. Globally, the PHA productivity of the entire system (selection + accumulation) in the different setups were 0.5,1,0.6, and 0.8 gCOD.L-1. d. -1 for setups 2, 3, 4, and 5 respectively. Overall, the PHA storage yield was 0.2, 0.3, 0.2, and 0.3 (mgCOD/mgCOD) respectively for setups 2, 3, 4, and 5 as well.
In summary, the research showed a very good potential for the continuous process to achieve both good biomass selection and PHA accumulation in order to produce PHA by using microbial mixed cultures.
This thesis also investigated the occupational risk and health assessment in PHA pilot scale production (case study Treviso PHA production pilot scale). This preliminary analysis aimed to highlight the possible deviations and hazards that could occur and cause concerns to the workers. To conduct this study, among the process hazard analysis tools, the HazOp (modified to BioHazOp) methodology has been applied to the first stage of the process since it can be considered the most hazardous stage (acidogenic fermenter). To guide comprehension, an initial P&ID has been designed to assist analysis by taking into account every single piece of equipment of the node under consideration. Finally, a list of potential deviations and hazards has been provided, along with several countermeasures and a final P&ID (which includes all of the pipes, process equipment, as well as its instrumentation and control systems to better regulate the process) to avoid, prevent, or limit the risk appearance.
The final section of this thesis included the results acquired at the world's first demonstrative PHA production plant using MMC and less expensive substrates (wastewater). This experiment aims to gather information for a larger industrial scale and to generate as much PHBV-rich biomass in direct accumulation without selection (with at least 40% gPHA/gVSS of intracellular PHA content) as possible in order to fulfill the PHA production target of 2500 tons/year.