This paper presents the results of fungal treatment of a real industrial wastewater (WW), providing insight into the main mechanisms involved and clarifying some ambiguities and uncertainties in the previous reports. In this regard, the mycoremediation potentials of four strains of white rot fungi (WRF): Phanerochaete chrysosporium, Trametes versicolor, Pleurotus ostreatus and Pleurotus pulmonarius were tested to remove humic acids (HA) from a real humic-rich industrial treated WW of a food-processing plant. The HA removal was assessed by color measurement and size-exclusion chromatography (SEC) analysis. T. versicolor showed the best decolorization efficiency of 90% and yielded more than 45% degradation of HA, which was the highest among the tested fungal strains. The nitrogen limitation was studied and results showed that it affected the fungal extracellular laccase and manganese peroxidase (MnP) activities. The results of the SEC analysis revealed that the mechanism of HA removal by WRF involves degradation of large HA molecules to smaller molecules, conversion of HA to fulvic acid-like molecules and also biosorption of HA by fungal mycelia. The effect of HS on the growth of WRF was investigated and results showed that the inhibition or stimulation of growth differs among the fungal strains.@en

In order to address the ambiguities and knowledge gaps in the HA removal by WRF, mechanisms of removal were explored in chapter 2. It was shown that the mechanisms of HA removal by WRF involves the degradation of HA to smaller molecules, conversion to FA and also biosorption of HA by fungal mycelia. Furthermore, the ability of laccase to effectively degrade HA was demonstrated and it was shown that the presence of suitable mediators, such as ABTS, have a crucial effect on the degradation process. Also, the contribution of the membrane-bond CYP 450 enzymes to the rate of the HA degradation by fungi was demonstrated. In addition, addressing the ambiguities in the measurement techniques mentioned in chapter 1, it was shown that decolorization was not necessarily representing the degradation of HA, but removal of HA in the general context. This removal could be due to biosorption, in-complete or complete degradation of HA to smaller molecules. Also, it was shown that the degradation or depolymerization of the HA is not necessarily reflected by a change in the average MW of the HA in the wastewater. The reason is that the depolymerization and/or degradation of HA could basically result in three products, smaller HA molecules, smaller non-aromatic molecules, and FA. Except for the first group, the other products from the degradation of HA could not be detected by the SEC analysis (at 254 nm) and were therefore not considered in the calculation of the average MW. Also, it was shown (chapter 2 and 3) that the biosorption of HA to fungal mycelia was not uniform among large, medium and small size HA. The large HA molecules were shown to be more susceptible to biosorption. Therefore, sole biosorption of HA to fungal mycelia could result in a change in MW distribution of the HA, even when no degradation or polymerization have occurred...@en

In this study, we tested a new approach to facilitate the application of white rot fungi (WRF) under non-sterile conditions, by introducing grain sorghum as carrier and sole carbon and nutrient source for WRF. To this end, Trametes versicolor was immobilized on sorghum, and its ability to remove humic acid (HA) from synthetic and real industrial wastewater was studied. HA removal was measured as colour reduction and also analysed via size exclusion chromatography (SEC). Under sterile conditions, 80% colour removal was achieved for both synthetic and real wastewater using immobilized WRF on sorghum, without adding any additional carbon or nutrient sources. Under non-sterile conditions, immobilized fungi could again remove 80% of the colour and reached a maximum of 40 U/L laccase activity. In contrast, non-immobilized fungi cultivated in non-sterile wastewater supplemented with additional nutrients, reached only 10% decolourization and maximum 5 U/L laccase activity. SEC analysis showed that bioremoval of HA by WRF was associated with degradation of HA. Finally, immobilized fungi were used to treat real wastewater, under non-sterile conditions, in a sequential batch order without renewing the immobilized fungi. Four batch feedings were conducted and 80%, 70%, 50% and 40% colour removal was achieved for each batch, respectively, over a total incubation period of 19 days.@en