Production and characterization of β-glucosidase from Aspergillus niger fermentation: Application for organic fraction of municipal solid waste hydrolysis and methane enhancement

The anaerobic digestion of the organic fraction of municipal solid waste (OFMSW) is currently an attractive treatment process with energy production in the form of biogas. Hydrolysis is the rate-limiting step for the anaerobic digestion of solid wastes. Thus, in the present study fungal enzymatic pretreatment of OFMSW was applied to enhance biogas production. Two enzyme cocktails rich on β-glucosidase were produced from submerged fermentation of Aspergillus niger on basal medium using OFMSW as carbon source and urea (Urea cocktail) and Ulva rigida as nitrogen source (Ulva cocktail). Ulva cocktail displayed an important effect on OFMSW solubilization. Therefore, an increase of reducing sugar concentration about 60% was obtained which was in correlation with chemical oxygen demand (COD) increase. The performance of enzymatic pretreatment on anaerobic digestion of OFMSW was studied by conducting biochemical methane potential tests. Results showed that the enzymatic pretreatment improved methane yield of OFMSW even at high solid concentration. High methane yield about 500 ml/g total volatile solid was obtained, which corresponds up to 68% enhancement over the control.     

Ultrasonic and Thermal Pretreatments on Anaerobic Digestion of Petrochemical Sludge: Dewaterability and Degradation of PAHs

Effects of different pretreatment methods on sludge dewaterability and polycyclic aromatic hydrocarbons (PAHs) degradation during petrochemical sludge anaerobic digestion were studied. Results showed that the total biogas production volume in the thermal pretreatment system was 4 and 5 times higher than that in the ultrasound pretreatment and in the control system, and the corresponding volatile solid removal efficiencies reached 28%, 15%, and 8%. Phenanthrene, paranaphthalene, fluoranthene, benzofluoranthene, and benzopyrene removal rates reached 43.3%, 55.5%, 30.6%, 42.9%, and 41.7%, respectively, in the thermal pretreatment system, which were much higher than those in the ultrasound pretreatment and in the control system. Moreover, capillary suction time (CST) of sludge increased after pretreatment, and then reduced after 20 days of anaerobic digestion, indicating that sludge dewaterability was greatly improved after anaerobic digestion. The decrease of protein and polysaccharide in the sludge could improve sludge dewaterability during petrochemical sludge anaerobic digestion. This study suggested that thermal pretreatment might be a promising enhancement method for petrochemical sludge solubilization, thus contributing to degradation of the PAHs, biogas production, and improvement of dewaterability during petrochemical sludge anaerobic digestion.     

Improvement of the kinetics of anaerobic digestion of molasses by the removal of phenolic compounds

Summary Removal of most of the phenolic compounds (88.5%) present in molasses by an aerobic pretreatment with Geotrichum candidum considerably increased the kinetics of anaerobic digestion of this wastewater. Inhibition phenomena were not observed in the range of COD studied (2.3–12.9 g/l). The yield coefficient of methane was 51 % higher than for untreated molasses.     

Two-stage biological treatment of olive mill wastewater with whey as co-substrate

Olive mill wastewater (OMW) is a highly polluting wastewater, caused by a high organic load and phenol content. These characteristics suggest that it may be suitable for aerobic treatment and anaerobic bacterial digestion. Aerobic treatment coupled with anaerobic bacterial digestion may be economically feasible as the methane produced is a valuable energy source while simultaneously purifying the OMW. In an attempt to improve the overall performance of the process, the addition of a co-substrate such as whey to the aerobic treatment pre-treatment of OMW by the yeast Candida tropicalis was studied.The two-stage system operated satisfactorily up to an organic loading rate (OLR) of 3.0 kg COD L⁻¹ day⁻¹ with a biogas production rate of 1.25 L_biogas L_reactor⁻¹ day⁻¹ and a total COD reduction in excess of 93% (62% COD reduction in aerobic pretreatment and 83% COD reduction in anaerobic digestion). Fifty-four percent of the phenol was biodegraded during the aerobic treatment stage, and biogas with between 68% and 75% methane was produced during anaerobic digestion.     

Limonene bioconversion to high concentrations of a single and stable product, perillic acid, by a solvent-resistant Pseudomonas putida strain

The white-rot basidiomycete Phanerochaete chrysosporium BKM-F-1767 was tested for its capacity to degrade dehydroabietic acid (DHA). In anaerobic treatment, this molecule is the most recalcitrant member of the resin acid group, which is known to cause operational problems to anaerobic reactors treating pulp and paper industry wastewaters. In this study the effect of DHA on different parameters, such as growth, ligninolytic enzyme activity, extracellular protein production as well as both glycerol and ammonium consumption by the fungus, was determined. Although the above parameters were affected by the addition of DHA, the results show that the fungus could still produce significant titres of ligninolytic enzymes. The fungus removed 47% of the DHA initially present in the static culture, after 10 days of incubation. Anaerobic toxicity assays showed that the treatment of DHA with P. chrysosporium reduced the methanogenesis and acetogenesis inhibition caused by DHA and allowed improved methane production by the anaerobic bacteria. Received: 10 June 1997 / Received revision: 6 January 1998 / Accepted: 24 January 1998     

A new alternative process for Kraft E1 effluent treatment

Lentinus edodes (UEC-2019 strain) was selected after screening 51 ligninolytic strains of fungi for their ability to decolorize phenolic industrial effluent with high content of lignin peroxidases, Mn-peroxidases and beta-glucosidases. This strain removed 73 % of color in theEucalyptus Kraft E1 effluent in 5 days without any additional carbon sources. A 13% mycelial adsorption was found. Correlation between mass loss, COD, TOC and decolorization was observed. When an effluent pre-irradiated (10 min) in the presence of ZnO was treated withL. edodes, a marked enhancement of the decolorization at 48 h was obtained.L. edodes is an active fungus in this pre-treatment and biobleaching process. The combined photo-biological decolorization procedure appears to be an efficient decontamination method with great potential in industrial effluent treatment.Abbreviation COD Chemical oxygen demand - TOC Total organic carbon     

Enhanced anaerobic gas production of waste activated sludge pretreated by pulse power technique

An electric pulse-power reactor consisting of one coaxial electrode and multiple ring electrodes was developed to solubilize waste activated sludge (WAS) prior to anaerobic digestion. By pretreatment of WAS, the soluble chemical oxygen demand (SCOD)/total chemical oxygen demand (TCOD) ratio and exocelluar polymers (ECP) content of WAS increased 4.5 times and 6.5 times, respectively. SEM images clearly showed that pulse-power pretreatment of WAS was found to result in destruction of sludge cells. Batch-anaerobic digestion of pulse-power treated sludge showed 2.5 times higher gas production than that of untreated sludge. Solubilized sludge cells by pulse-power pretreatment would be readily utilized for anaerobic microorganisms to produce anaerobically-digested gas. Slow or lagged gas production in the initial anaerobic digestion stage of pulse-power pretreated sludge implied that the methane-forming stage of anaerobic digestion would be the rate-limiting step for anaerobic digestion of pulse-power pretreated sludge.     

Thermophilic anaerobic digestion of Lurgi coal gasification wastewater in a UASB reactor

Lurgi coal gasification wastewater (LCGW) is a refractory wastewater, whose anaerobic treatment has been a severe problem due to its toxicity and poor biodegradability. Using a mesophilic (35 ± 2 °C) reactor as a control, thermophilic anaerobic digestion (55 ± 2 °C) of LCGW was investigated in a UASB reactor. After 120 days of operation, the removal of COD and total phenols by the thermophilic reactor could reach 50-55% and 50-60% respectively, at an organic loading rate of 2.5 kg COD/(m³ d) and HRT of 24 h; the corresponding efficiencies were both only 20-30% in the mesophilic reactor. After thermophilic digestion, the wastewater concentrations of the aerobic effluent COD could reach below 200 mg/L compared with around 294 mg/L if mesophilic digestion was done and around 375 mg/L if sole aerobic pretreatment was done. The results suggested that thermophilic anaerobic digestion improved significantly both anaerobic and aerobic biodegradation of LCGW.     

Microbiological contamination of digested products from anaerobic co-digestion of bovine manure and agricultural by-products

Aims: This study was performed to investigate the microbiological contamination of digestate product (DP) obtained from the anaerobic co‐digestion of bovine manure and agricultural by‐products. Methods and results: Microbiological analyses were performed on bovine manure, fresh DP, liquid and solid fractions and stored liquid fraction of DP. A statistically significant reduction of faecal bacterial indicator was found after anaerobic digestion except for enterococci. After liquid/solid DP separation, bacteria tend to be concentrated in the solid fraction. Storage does not seem to influence the indicator parameters, except for enterococci. Escherichia coli O157:H7 and Yersinia were not found in any samples analysed. Salmonella was rarely detected in DP samples and its derivates, while Listeria monocytogenes was encountered in many samples. Conclusions: The results obtained indicate that the hygienic quality of DP is for almost all microbiological parameters better than that of the bovine manure (range of reduction 1.6–3.1 log₁₀) and suggest the need to identify specific pathogen indicators related to the hygienic characteristics of DPs. Significance and Impact of the Study: This study highlights that the anaerobic co‐digestion of bovine manure and agricultural by‐products in a field‐scale biogas plant does not increase human health risk with respect to the use of animal manure for agricultural fertilization.     

Degradation of 4-nitrophenol by the lignin-degrading basidiomycete <Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis>

The fungal metabolism of 4-nitrophenol (4-NP) was investigated using the lignin-degrading basidiomycete, Phanerochaete chrysosporium. Despite its phenolic feature, 4-NP was not oxidized by extracellular ligninolytic peroxidases. However, 4-NP was converted to 1,2-dimethoxy-4-nitrobenzene via intermediate formation of 4-nitroanisole by the fungus only under ligninolytic conditions. The metabolism proceeded via hydroxylation of the aromatic ring and methylation of phenolic hydroxyl groups. Although the involvement of nitroreductase in the metabolism of 2,4-dinitrotoluene by many aerobic and anaerobic microorganisms including P. chrysosporium has been reported, no formation of 4-aminophenol was observed during 4-NP metabolism. The formation of 1,2-dimethoxy-4-nitrobenzene was effectively inhibited by exogenously added piperonyl butoxide, a cytochrome P450 inhibitor, suggesting that cytochrome P450 is involved in the hydroxylation reaction. Thus, P. chrysosporium seems to utilize hydroxylation and methylation reactions to produce a more susceptible structure for an oxidative metabolic system.     

Using Contaminated Plants Involved in Phytoremediation for Anaerobic Digestion

This study investigated the anaerobic digestion capability of five plants and the effects of copper (Cu) and S,S’-ethylenediaminedisuccinic acid (EDDS, a chelator widely used in chelant-assisted phytoremediation) on biogas production to determine a feasible disposal method for plants used in remediation. The results showed that in addition to Phytolacca americana L., plants such as Zea mays L., Brassica napus L., Elsholtzia splendens Nakai ex F. Maekawa, and Oenothera biennis L. performed well in biogas production. Among these, O. biennis required the shortest period to finish anaerobic digestion. Compared to normal plants with low Cu content, the plants used in remediation with increased Cu levels (100 mg kg^?1) not only promoted anaerobic digestion and required a shorter anaerobic digestion time, but also increased the methane content in biogas. When the Cu content in plants increased to 500, 1000, and 5000 mg kg^?1, the cumulative biogas production decreased by 12.3%, 14.6%, and 41.2%, respectively. Studies also found that EDDS conspicuously restrained biogas production from anaerobic digestion. The results suggest that anaerobic digestion has great potential for the disposal of contaminated plants and may provide a solution for the resource utilization of plants used in remediation.     

Effect of biological pretreatment with Trametes hirsuta yj9 on enzymatic hydrolysis of corn stover

In this study, a newly isolated Trametes hirsuta yj9 was used to pretreat corn stover in order to enhance enzymatic digestibility. T. hirsuta yj9 preferentially degraded lignin to be as high as 71.49% after 42-day pretreatment. Laccase and xylanase was the major ligninolytic and hydrolytic enzyme, respectively and filter paper activity (FPA) increased gradually with prolonged pretreatment time. Sugar yields increased significantly after pretreatment with T. hirsuta yj9, reaching an enzymatic digestibility of 73.99% after 42 days of pretreatment. Scanning electron microscopy (SEM) showed significant structural changes in pretreated corn stover, the surface of pretreated corn stover became increasingly coarse, the gaps between cellulose fibers were visible, and many pores were developed. Correlation analysis showed that sugar yields were inversely proportional to the lignin contents, less related to cellulose and hemicellulose contents.     

Low temperature thermo-chemical pretreatment of dairy waste activated sludge for anaerobic digestion process

An investigation into the influence of low temperature thermo-chemical pretreatment on sludge reduction in a semi-continuous anaerobic reactor was performed. Firstly, effect of sludge pretreatment was evaluated by COD solubilization, suspended solids reduction and biogas production. At optimized condition (60 °C with pH 12), COD solubilization, suspended solids, reduction and biogas production was 23%, 22% and 51% higher than the control, respectively. Secondly, semi-continuous process performance was studied in a lab-scale semi-continuous anaerobic reactor (5 L), with 4 L working volume. With three operated SRTs, the SRT of 15 days was found to be most appropriate for economic operation of the reactor. Combining pretreatment with anaerobic digestion led to 80.5%, 117% and 90.4% of TS, SS and VS reduction respectively, with an improvement of 103% in biogas production. Thus, low temperature thermo-chemical can play an important role in reducing sludge production.     

Comparative Removal of Bentazon by Ganoderma lucidum in Liquid and Solid State Cultures

Bentazon removal by Ganoderma lucidum cultured in liquid and solid state conditions was compared in this work. In solid state cultures, the fungus produced both ligninolytic enzymes, namely laccase and Mn peroxidase. In liquid cultures, the main ligninolytic enzyme produced was laccase. In both types of cultures bentazon improved the production of laccase without significant alteration in the production of Mn peroxidase. In solid state cultures, where high levels of both laccase and Mn peroxidase activities were found, the fungus was more resistant to the action of the herbicide (50 mM in solid state cultures against 20 mM in liquid cultures) and more efficient in removing bentazon (90% removal against 55% in liquid cultures after 10 days of cultivation). Furthermore, the solid state culture filtrates were more efficient in the in vitro degradation of bentazon than the liquid culture filtrates. These observations suggest that both enzymes, laccase and Mn peroxidase, are involved in bentazon degradation. The results further suggest that solid state cultures of Ganoderma lucidum could be useful in strategies designed to reduce environmental contamination by bentazon.     

Enhancement on biodegradation and anaerobic digestion efficiency of activated sludge using a dual irradiation process

A dual irradiation process involving aerobic thermophilic irradiation pretreatment (ATIP) and intermittent irradiation anaerobic digestion was developed to improve the digestion of waste-activated sludge. First, the effect of ATIP on further anaerobic digestion of activated sludge in batch mode was investigated. When exposed to ATIP for 24 h, the digestion reactor gave the highest methane yield, removed the most dissolved organic carbon (DOC) and showed the most effective reduction of VS compared to other irradiation times. This process was further enhanced by using an anaerobic fluidised-bed reactor packed with carbon felt in semi-continuous mode for digesting the pretreated activated sludge under intermittent irradiation conditions. Dual irradiation for 24 h followed by 60 min of anaerobic irradiation processing per day turned out to be optimal. This resulted in 65.3% of VS reduction, 83.9% of DOC removal ratio and 538 ml/g-VS of methane yield.     

Effect of pre-aeration and inoculum on the start-up of batch thermophilic anaerobic digestion of municipal solid waste

In this study, a short pre-aeration step was investigated as pre-treatment for thermophilic anaerobic digestion of the organic fraction of municipal solid waste (OFMSW). It was found that pre-aeration of 48 h generated enough biological heat to increase the temperature of bulk OFMSW to 60 °C. This was sufficient self-heating of the bulk OFMSW for the start-up of thermophilic anaerobic digestion without the need for an external heat source. Pre-aeration also reduced excess easily degradable organic compounds in OFMSW, which were the common cause of acidification during the start-up of the batch system. Careful consideration however must be taken to avoid over aeration as this consumes substrate, which would otherwise be available to methanogens to produce biogas. To accelerate methane production and volatile solids destruction, the anaerobic digestion in this study was operated as a wet process with the anaerobic liquid recycled through the OFMSW. Appropriate anaerobic liquid inoculum was found to be particularly beneficial. It provided high buffer capacity as well as suitable microbial inoculum. As a result, acidification during start-up was kept to a minimum. With volatile fatty acids (VFAs-acetate in particular) and H₂ accumulation typical of hydrolysis and fermentation of the easily degradable substrates during start-up, inoculum with high numbers of hydrogenotrophic methanogens was critical to not only maximise CH₄ production but also reduce H₂ partial pressure in the system to allow VFAs degradation. In a lab-scale bioreactor, the combined pre-aeration and wet thermophilic anaerobic digestion was able to stabilise the OFMSW within a period of only 12 days. The stabilised inert residual material can be used as a soil amendment product.     

Evaluation of the white-rot fungi Ganoderma australe and Ceriporiopsis subvermispora in biotechnological applications

Ganoderma australe is a white-rot fungus that causes a selective wood biodelignification in some hardwoods found in the Chilean rainforest. Ceriporiopsis subvermispora is also a lignin-degrading fungus used in several biopulping studies. The enzymatic system responsible for lignin degradation in wood can also be used to degrade recalcitrant organic pollutants in liquid effluents. In this work, two strains of G. australe and one strain of C. subvermipora were comparatively evaluated in the biodegradation of ABTS and the dye Poly R-478 in liquid medium, and in the pretreatment of Eucalyptus globulus wood chips for further kraft biopulping. Laccase was detected in liquid and wood cultures with G. australe. Ceriporiopsis subvermispora produce laccase and manganese peroxidase when grown in liquid medium and only manganese peroxidase was detected during wood decay. ABTS was totally depleted by all strains after 8 days of incubation while Poly R-478 was degraded up to 40% with G. australe strains and up to 62% by C. subvermispora after 22 days of incubation. Eucalyptus globulus wood chips decayed for 15 days presented 1–6% of lignin loss and less than 2% of glucan loss. Kraft pulps with kappa number 15 were produced from biotreated wood chips with 2% less active alkali, with up to 3% increase in pulp yield and up to 20% less hexenuronic acids than pulps from undecayed control. Results showed that G. australe strains evaluated were not as efficient as C. subvermispora for dye and wood biodegradation, but could be used as a feasible alternative in biotechnological processes such as bioremediation and biopulping.     

Fungal pretreatment: An alternative in second-generation ethanol from wheat straw

The potential of a fungal pretreatment combined with a mild alkali treatment to replace or complement current physico-chemical methods for ethanol production from wheat straw has been investigated. Changes in substrate composition, secretion of ligninolytic enzymes, enzymatic hydrolysis efficiency and ethanol yield after 7, 14 and 21 days of solid-state fermentation were evaluated. Most fungi degraded lignin with variable selectivity degrees, although only eight of them improved sugar recovery compared to untreated samples. Glucose yield after 21 days of pretreatment with Poria subvermispora and Irpex lacteus reached 69% and 66% of cellulose available in the wheat straw, respectively, with an ethanol yield of 62% in both cases. Conversions from glucose to ethanol reached around 90%, showing that no inhibitors were generated during this pretreatment. No close correlations were found between ligninolytic enzymes production and sugar yields.     

Molecular weight distribution of Pinus radiata kraft mill wastewater treated by anaerobic digestion

Kraft mill is responsible for massive discharge of highly polluted effluents. The main characteristics of this effluent are high toxicity and low biodegradability due to tannin, lignin and chlorophenol compounds. The composition may vary dramatically depending, for instance, on the utilised feedstock and process. The purpose of this work was to investigate the molecular weight distribution of Pinus radiata kraft pulping wastewater treated by anaerobic digestion by using two types of anaerobic reactors: fixed bed and sludge blanket. Anaerobic sludge blanket (UASB) and anaerobic filter (AF) were operated. In both reactors, the total alkalinity ranged between 1.0 and 1.5 g CaCO3/l, while the organic load rate (OLR) was increasing during operation from 1.2 to 3.3 gCOD/l d. COD and total phenolic compounds (UV215) removal ranged between 30-50% and 13-20%, respectively, while the BOD5 removal ranged 60-90%. However only a partial biodegradation (10-43%) of tannin and lignin was observed. Results from ultrafiltration analyses indicated that the fraction with a molecular weight (MW) < 1000, COD and colour decreased after anaerobic treatment, but the total phenolic compounds increased. In the 1000 < MW < 10,000 fraction, there was no change in COD, UV215 and colour. In the > 10,000 MW fraction, colour and COD fraction increased by 14% and 5%, respectively, after anaerobic treatment. It can be concluded from this study, that treatment with UASB or AF reactors is not enough, under the conditions tested, for a large COD removal from Pinus radiata wastewater.     

Anaerobic Digestion of Dairy Manure in a Hybrid Reactor with Biogas Recirculation

Summary A novel anaerobic hybrid reactor (AHR) configuration incorporating floating support media for biomass immobilization and biogas recirculation for enhanced mixing was used for anaerobic digestion of dairy manure. No pretreatment or solid liquid separation was applied. The reactor was operated at high influent volatile solids (VS) and organic loading rates (OLR) of up to 9.87% and 7.30 g VS/l day, respectively. After 149 days of continuous operation the results revealed that a high amount (38.1 g VSS) of biomass was able to attach itself to the support medium being used. The investigated AHR configuration achieved COD, BOD, TS, and VS removal efficiencies of 48–63, 64–78, 55–65, and 59–68%, respectively, at a hydraulic retention time (HRT) of 15 days. The corresponding average methane production value obtained in this study was 0.191 l/g VS added.