The objective of this work was to evaluate the semi-continuous post-treatment of anaerobically-pre-treated weak black liquor (anaerobic effluent, AnE) by aerobic post-treatment using hybrid pellets of Trametes versicolor. The latter consisted of fungus immobilized onto holm oak sawdust (mixed or double pellets) or a mixture of holm oak sawdust and powdered activated carbon (triple pellets). First, a semi-continuous experiment was run to compare the effectiveness of triple and mixed pellets in agitated flasks for 15 cycles of 7 days each. A second extended batch test was implemented with 500 mL AnE and triple pellets to give 400 mg fungal biomass; some units were spiked with protease inhibitor. In the first experiment, triple pellets displayed consistently higher removal efficiencies of pollutant parameters than double pellets (10 to 15% higher), although overall averages were moderate and no statistical significance to the difference could be set because of the noise of fluctuations. Periodic fluctuations of removal were characterized by three periods of approximately six cycles each with maximum removals occurring at cycles 3–4, 7–9, 13, and 14. Evaluating pooled removals of the latter cycles showed that triple pellets were significantly more effective than double pellets, with removal efficiencies as high as 47% of COD, colour, and absorbance at 254 nm (A254). In general, protease activity seemed to increase in the third period (last six cycles), whereas activities of MnP, LiP and Lac significantly decreased. In the second experiment, pollutant removals and enzymatic activities of triple pellets with protease inhibitor were significantly higher than those of units without added protease inhibitor. These results indicate that protease could be the main cause of periodic falls of pollutant removal efficiencies found in the first experiment.

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