Biopolymer production through Purple Phototropic Bacteria (PPB) and Activated Sludge (AS) by different engineering processing

The issue of treatment and disposal of sludge produced by wastewater treatment plants (WWTPs), has become relevant worldwide due to the high amounts of residual sludge to be managed, the stringent requirements for agricultural use of sludge, and the cost and disposal constraints associated to landfilling. In Europe, where the availability of fossil fuels is limited, combined treatment of sewage sludge and biowaste, aimed at recovering material and energy, appears to be promising. The present work investigates the feasibility of applying the so-called bio-refinery approach to bio-sludge from WWTP and organic waste, through the analysis of a combined process for converting organic waste (agro-industrial wastes and sewage sludge) into biofuels (bio-H2) and biomaterials (biopolymers). The management of excess sludge from WWTPs is a critical issue due to the high costs and environmental implications. The conversion of a conventional WWTP into a biorefinery may contribute, while producing a high-quality effluent, to attaining the recovery of valuable elements, materials as well as energy. In particular, a suitable combination of treatment technologies may be directed to producing bio-H2 from mixtures of sewage sludge and agroindustrial biowaste in a first fermentative process stage, and bio-CH4 (under anaerobic conditions) or biopolymers (under aerobic/anaerobic conditions) in a subsequent treatment stage. Among the biopolymers, polyhydroxyalkanoates (PHAs), polyesters synthesized by numerous bacteria, appear to be the most attractive due to their thermoplastic properties similar to that of polypropylene (PP) (Carlozzi et al., 2018). So far, the application of mixed microbial cultures (MMCs) using complex residues for PHA production, through the application of transient carbon availability conditions (the so-called Feast and Famine (FF) strategy), has been widely investigated. However, PHA production with MMCs has been mainly restricted to the utilization of aerobic organisms. In recent years, different studies have proposed the utilization of phototrophic mixed cultures (PMCs). Phototrophic organisms can draw energy from sunlight and by not requiring oxygen to produce ATP, aeration is nonessential, and the high costs associated with system’s aeration can be eliminated. (Fradinho et al., 2013a). However, the research on the application of mixed microbial cultures (aerobic and anaerobic) for PHA production, have not yet achieved satisfactory results and needs further investigations applying waste complex feedstock in the process. The experimental campaign of this research project was developed in order to acquire the information necessary to derive the kinetics that describe the production processes of biopolymers, not only to evaluate the yields but also in qualitative terms, by different engineering processing. The specific targets of the research activities were the identification of the effect of different operating conditions on biopolymer production yields and kinetics and identification of the technical and environmental properties of the produced biopolymers. A first set of experiments, carried out at La Sapienza University of Rome with the participation of ACEA SpA, was designed to evaluate the optimal operating 3 conditions to enrich the PHA-accumulating biomass from the aerobic biological sludge collected at a full-scale WWTP located in Rome. To achieve these targets a synthetic substrate, mimicking the composition of a VFAs-rich effluent from the dark fermentation of mixtures of pre-treated sludge and cheese whey (Akhlaghi et al., 2017), was used as the substrate of an aerobic process for biopolymer accumulation. This study aims to explore the capabilities of the mixed microbial communities (MMCs) to produce PHAs from artificial substrate and complex substrates such as organic waste in a SBR reactor. Five different cycles of SBR were run and 5 different OLR were tested: SBR 1/ SBR 2 (OLR 33 mmolC/ld), SBR 3 (OLR 100 mmolC/ld) SBR 4 (OLR 300 mmolC/ld) and SBR 5 (OLR 150 mmolC/ld). Considering the elements compared, it can be concluded that the operating conditions adopted with SBR3, with an organic load set at 100 mmolC / l  d, were those that led to the best performance of the biomass selection and PHA production process, both in terms of productivity and polymer storage yields. PHA was reflected by a maximum accumulation percentage from 1 % up to 5 % (w/w) in 8 hours and a complete removal of C and N from the reactor. A second set of experiments was carried out at in collaboration with the Advanced Water Management Centre (AWMC) and the School of Chemical Engineering of the University of Queensland. This study was aimed to evaluate the PHA accumulation capacity by the use of another promising mixed culture: The Purple Phototrophic Bacteria (PPB). A continuous 2-L bench scale photo-anaerobic membrane reactor (PAnMBR), was used to test the ability to enrich PHA accumulating microorganisms from sewage sludge, using a synthetic substrate as in the previous experimental set of tests. The operative condition studied in this research had shown a potential capacity of PPB in the PHA accumulation process, reaching more than 15 % (W/W) content of PHA in the microbial cells and 61 % of COD removal and a complete N removal in each cycle. A third series of tests was carried out to investigate an alternative of the conventional biopolymer production setup, trying to minimize the cost of the oxygen supply and the use of expensive substrate. The Study of PHA accumulating capacity of PPBs growing under the natural light/dark cycle evaluating the effect of using molasses as sole carbon source were tested in an out-door 60 l flat plate photobioreactor. This study was carried in collaboration with the School of Chemical Engineering and Advanced Wastewater Management Centre (AWMC) of the University of Queensland (Australia). The operative condition studied in this research had shown a potential capacity of PPB in the PHA accumulation process from fermented molasses, reaching more than 13 % (W/W) content of PHA in the microbial cells and 61 % of COD removal. The results of the batch tests have confirmed the same accumulation capacity of the lab-selected culture so that strengthening the application of this experimental setup could help reaching more than 28 % of PHA content. The outcome of this study identified the PPB as one of the main interesting research challenges for industrial application of biopolymer production from waste.