Simultaneous recovery of calcium phosphate granules (CaP granules) and methane from vacuum collected black water (BW), using an upflow anaerobic sludge blanket (UASB) reactor was previously investigated. It was calculated that only 2% of the total phosphorus (P) fed was present as CaP granules whereas 51% of the P accumulated dispersed in the reactor, limiting the applicability of this process for recovery of phosphate. This study proposes adding calcium to increase the P accumulation in the reactor and the production of CaP granules. Calcium was added in a lab-scale UASB reactor fed with BW. An identical UASB reactor was used as reference, to which no calcium was added. The treatment performance was evaluated by weekly monitoring of influent, effluent and produced biogas. Sludge bed development and CaP granulation were assessed through particle size analysis. The composition and structure of CaP granules were chemically and optically assessed. Calcium addition increased accumulation of P in the reactor and formation and growth of granules with size > 0.4 mm diameter (CaP granules). Moreover, with calcium addition, CaP granules contained 5.6 ± 1.5 wt% of P, while without calcium a lower P content was observed (3.7 ± 0.3 wt%). By adding Ca, 89% of the incoming P from BW accumulated in the reactor and 31% was sampled as CaP granules (> 0.4 mm diameter). Addition of 250 mgCa L of BW was the optimum loading found in this study. Furthermore, no significant reduction in COD removal (> 80%) and CH production (0.47 ± 0.10 gCOD-CH gCOD-BW) was observed. Therefore, adding calcium can significantly increase the CaP granulation without inhibiting the simultaneous CH recovery. This further indicates the potential of this process for phosphate recovery.

The present study focuses on predicting the concentration of intracellular storage polymers in enhanced biological phosphorus removal (EBPR) systems. For that purpose, quantitative image analysis techniques were developed for determining the intracellular concentrations of PHA (PHB and PHV) with Nile blue and glycogen with aniline blue staining. Partial least squares (PLS) were used to predict the standard analytical values of these polymers by the proposed methodology. Identification of the aerobic and anaerobic stages proved to be crucial for improving the assessment of PHA, PHB and PHV intracellular concentrations. Current Nile blue based methodology can be seen as a feasible starting point for further enhancement. Glycogen detection based on the developed aniline blue staining methodology combined with the image analysis data proved to be a promising technique, toward the elimination of the need for analytical off-line measurements.

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Simultaneous recovery of calcium phosphate granules (CaP granules) and methane in anaerobic treatment of source separated black water (BW) has been previously demonstrated. The exact mechanism behind the accumulation of calcium phosphate (Cax(PO4)y) in CaP granules during black water treatment was investigated in this study by examination of the interface between the outer anaerobic biofilm and the core of CaP granules. A key factor in this process is the pH profile in CaP granules, which increases from the edge (7.4) to the center (7.9). The pH increase enhances supersaturation for Cax(PO4)y phases, creating internal conditions preferable for Cax(PO4)y precipitation. The pH profile can be explained by measured bioconversion of acetate and H2, HCO3– and H+ into CH4 in the outer biofilm and eventual stripping of CO2 and CH4 (biogas) from the granule. Phosphorus content and Cax(PO4)y crystal mass quantity in the granules positively correlated with the granule size, in the reactor without Ca2+ addition, indicating that the phosphorus rich core matures with the granule growth. Adding Ca2+ increased the overall phosphorus content in granules >0.4 mm diameter, but not in fine particles (<0.4 mm). Additionally, H+ released from aqueous phosphate species during Cax(PO4)y crystallization were buffered by internal hydrogenotrophic methanogenesis and stripping of biogas from the granule. These insights into the formation and growth of CaP granules are important for process optimization, enabling simultaneous Cax(PO4)y and CH4 recovery in a single reactor. Moreover, the biological induction of Cax(PO4)y crystallization resulting from biological increase of pH is relevant for stimulation and control of (bio)crystallization and (bio)mineralization in real environmental conditions.

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