For this research, a comparative evaluation was performed in which the addition of 3D-printed biocarriers fabricated with 13X and Halloysite (13Χ-Η) and commercial Kaldnes K1 biocarriers were evaluated as opposed to the non addition of biocarriers in a semi-pilot scale MBBR-MBR unit. For the evaluation of the MBBR-MBR efficiency, various physicochemical parameters were measured while static light scattering and optical microscopy observations were additionally used. Biofilm extracted from the biocarriers was also evaluated. It was observed that in the MBBR-MBR K1 unit, the membrane filtration improved while in the MBBR-MBR 13X-H unit the membrane fouling rate was the same as in the control MBBR-MBR unit. This is due to the production of large amount of SMP which resulted from the large amount of biofilm created in the 13X-H biocarriers. An optimal biodegradation of the organic load was concluded for all three MBBR-MBR units. The nitrification and denitrification processes were improved at the MBBR-MBR units using 13X-H and K1. The dry mass produced on the 13X-H biocarriers was three orders of magnitude larger than that produced on the K1. Finally, it was observed that mostly EPS was produced in the biofilm of K1 biocarriers while in the biofilm of 13X-H mostly SMP.

Biogas is mainly produced by anaerobic digestion (AD) using organic matter as a substrate. In Mediterranean countries, a promising substrate is lignocellulose biomass of perennial grass Miscanthus x giganteus, due to its potentially high biogas yields, which could be comparable to maize silage. During AD, bacteria convert biomass into more minor compounds, which are further converted to methane by methanogenic archaea. The selection of appropriate microbes for the degradation of the substrate is crucial, and the enhancement of this step lies in the immobilization of microbes on biocarriers. Here, a microbial consortium, de-novo isolated and conditioned to degrade the Mischantus biomass, was immobilized onto several natural biocarriers: natural zeolitized tuff (NZ), ZeoSand®, perlite, and corncob. There was no statistically signifi-cant difference in the numbers of immobilized bacteria between the materials. Therefore, all proved to be suitable for the immobilization of the consortium. In the consortium, five bacterial species with different shares in the consortium were identified: Enterobacter cloacae, Klebsiella pneumoniae, Enterobacter asburiae, Leclercia adecarboxylata, and Exiguobacterium indicum. After im-mobilization on each carrier, the share of each species changed when compared to starting conditions, and the most dominant species was E. cloacae (71-90%), while the share for other species ranged from 2-23%. The share of E. indicum was around 14 % at the start. However, it diminished to less than 1% because it was overgrown during the competition with other bacterial species, not due to an inability to immobilize. For future research, single species could be separately immobilized onto carriers, depending on their degradation capacity, thus creating the targeted and optimized biocarriers.

In the present work, a comparative evaluation of an MBBR unit performance was carried out for the following cases: when adding 3D-printed biocarriers fabricated with 13X and bentonite, when using K1 commercial biocarriers and when not adding biocarriers at all. For the evaluation of the MBBR efficiency, various physicochemical parameters were measured, while static light scattering and optical microscopy observations were additionally used. Finally, biofilm extracted from the biocarriers was evaluated. The findings suggest that there is an optimal biodegradation of the organic load in all MBBR units. The nitrification and denitrification process were improved at the 3D MBBR as compared to the control MBBR and MBBR K1. The dry mass of the biofilm in the 3D-printed biocarriers was two orders of magnitude larger than the one in the K1 biocarriers. What is more, in the K1 biocarriers the mass of the biofilm varied in relation to time, due to the fact that it could not be kept inside the holes, something that was not observed to happen with the 3D-printed biocarriers. Finally, it was observed, mostly in the 3D MBBR and less in the K1 MBBR, that the growth of nitrifying bacteria and heterotrophs inside the units increased the biomass production in the form of SMP, which in turn favored the adhesion of biomass on the surface of biocarriers.

The addition of biocarriers can improve the biological processes appearing in a bioreactor since their surface allows the immobilization, attachment, protection, and growth of microorganisms. In addition, the development of a biofilm layer allows the colonization of microorganisms in the biocarriers. The structure, composition, and roughness of the biocarriers' surface are crucial factors that affect the development of the biofilm. In the current work, the aluminosilicate zeolites 13X and ZSM-5 were examined as the main building components of the biocarrier scaffolds, using bentonite, montmorillonite, and halloysite nanotubes as inorganic binders in various combinations. 3D printing was utilized to form pastes into monoliths that underwent heat treatment. The 3d-printed biocarriers were subjected to a mechanical analysis, including density, compression, and nanoindentation tests. Furthermore, the 3d-printed biocarriers were morphologically and structurally characterized using nitrogen adsorption at 77 K (LN2), scanning electron microscopy (SEM), and X-ray diffraction (XRD). The stress-strain response of the materials being studied was obtained through nanoindentation tests combined with finite element analysis (FEA). These tests were also utilized to simulate the lattice geometries under compression loading conditions to investigate their deformation and stress distribution in relation to experimental compression testing. The results indicated that the 3d-printed biocarrier 13X/halloysite nanotubes was endowed with high specific surface area and extended mesoporous structure. Due to these assets, its bulk density was one of the lowest observed amongst the biocarriers derived from the various combinations of materials. The biocarriers based on the 13X zeolite exhibited the highest mechanical stability and appropriate morphological features. The 13X/halloysite nanotubes scaffold exhibits a moderate hardness value compared to the rest, while it presents the highest value of modulus of elasticity. In conclusion, Aluminosilicate zeolites and their combination with clays and inorganic nanotubes afford 3d-printed biocarriers of various textural and structural properties, which can be utilized to improve biological processes.