Reduction of the phenolic components in olive-mill wastewater by an enzymatic treatment and its impact on durum wheat (Triticum durum Desf.) germinability

Olive-mill wastewater (OMW), an effluent of olive oil extraction process, is annually produced in huge amounts in olive growing areas. An interesting option for its disposal is the spreading on agricultural land, provided that phytotoxic effects are neutralized. The objective of the present investigation was to evaluate the potential of an enzyme-based treatment in removing OMW phytotoxicity. To this aim, germinability experiments on durum wheat (Triticum durum Desf. cv. Duilio) were conducted in the presence of different dilutions of raw or enzyme-treated OMW. OMW treatment with laccase resulted in a 65% and 86% reduction in total phenols and ortho-diphenols respectively, due their polymerization as revealed by size-exclusion chromatography. Raw OMW exerted a significant concentration-dependent inhibition on the germinability of durum wheat seeds which was evident up to a dilution rate of 1:8. When the effluent was treated with a fungal laccase, germinability was increased by 57% at a 1:8 dilution and by 94% at a 1:2 dilution, as compared to the same dilutions using untreated OMW. The treatment with laccase also decreased the mean germination time by about 1 day as compared to untreated controls. These results show that germinability inhibition due to OMW can be reduced effectively using fungal laccase, suggesting that phenols are the main determinants of its phytotoxicity.     

Enzyme and fungal treatments and a combination thereof reduce olive mill wastewater phytotoxicity on Zea mays L. seeds

The phytotoxicity of olive-mill wastewater (OMW) has been suggested to be mainly due to its phenolic components. This study investigated the impact of three different low-cost dephenolization treatments on the wastewater phytotoxicity. To this aim, germinability of maize (Zea mays L.) seeds sown on a sandy-loamy soil which had been spread with different volumes (from 40 to 160m(3)ha(-1)) of either biologically-treated OMW or relative incubation control was determined. Biological treatments included either Panus tigrinus liquid cultures or incubation with commercial laccase (1UIml(-1)) or an innovative sequential combination of laccase and P. tigrinus cultures. All treatments markedly reduced phytotoxicity and promising results were obtained with commercial laccase. In fact, germinability and mean germination times in soil spread with laccase-treated OMW, did not significantly differ from those observed in soil irrigated with tap water (control) up to OMW volumes of 120m(3)ha(-1). Although the highest phenol reduction (ca. 81%) was obtained by the sequential use of laccase and P. tigrinus, the feasibility of the enzyme treatment is undoubtedly more convincing under the technological point of view.     

Land spreading of olive mill wastewater: effects on soil microbial activity and potential phytotoxicity

Extremely high organic load and the toxic nature of olive mill wastewater (OMW) prevent their direct discharge into domestic wastewater treatment systems. In addition to the various treatment schemes designed for such wastewater, controlled land spreading of untreated OMW has been suggested as an alternative mean of disposal. A field study was conducted between October 2004 and September 2005 to assess possible effects of OMW on soil microbial activity and potential phytotoxicity. The experiment was carried out in an organic orchard located on a Vertisol-type soil (Jezre'el Valley, Israel) and included two application levels of OMW (36 and 72m(3)ha(-1)). Total microbial counts, and to less extent the hydrolytic activity and soil respiration were increased following the high OMW application level. A bench-scale lab experiment showed that the rate of OMW mineralization was mainly dependent on the general status of soil activity and was not related to previous acclimatization of the soil microflora to OMW. Soil phytotoxicity (% germination and root elongation) was assessed in soil extracts of samples collected before and after each OMW application, using germinating cress (Lepidium sativum L.) seeds. We found direct short-term effect of OMW application on soil phytotoxicity. However, the soil was partly or completely recovered between successive applications. No further phytotoxicity was observed in treated soils as compared with control soil, 3 months after OMW application. Such short-term phytotoxicity was not in correlation with measured EC and total polyphenols in the soil extracts. Overall, the results of this study further support a safe controlled OMW spreading on lands that are not associated with sensitive aquifers.     

Lentinula edodes removes phenols from olive-mill wastewater: impact on durum wheat (Triticum durum Desf.) germinability

Olive-mill wastewater (OMW) exhibits highly phytotoxic properties, mainly due to phenols. A valuable option for OMW disposal is its agricultural use provided that phytotoxic effects are removed. The present investigation was aimed at evaluating the efficacy of the lignin-degrading fungus Lentinula edodes in achieving OMW detoxification. Germinability experiments on durum wheat showed that OMW phytotoxicity was significantly reduced by L. edodes cultures. Germinability on undiluted and twofold diluted OMW from fungal cultures was 34+/-5% and 57+/-6%, respectively, while on related incubation controls it was almost completely suppressed.These results suggest that fungal cultures of L. edodes would decrease the phytotoxicity of this waste.     

Release of substituents from phenolic compounds during oxidative coupling reactions

Phenolic compounds originating from plant residue decomposition or microbial metabolism form humic-like polymers during oxidative coupling reactions mediated by various phenoloxidases or metal oxides. Xenobiotic phenols participating in these reactions undergo either polymerization or binding to soil organic matter. Another effect of oxidative coupling is dehalogenation, decarboxylation or demethoxylation of the substrates. To investigate these phenomena, several naturally occurring and xenobiotic phenols were incubated with various phenoloxidases (peroxidase, laccase, tyrosinase) or with birnessite (delta-MnO(2)), and monitored for chloride release, CO(2) evolution, and methanol or methane production. The release of chloride ions during polymerization and binding ranged between 0.2% and 41.4%. Using the test compounds labeled with 14C in three different locations (carboxyl group, aromatic ring, or aliphatic chain), it was demonstrated that 14CO(2) evolution was mainly associated with the release of carboxyl groups (17.8-54.8% of the initial radioactivity). Little mineralization of 14C-labeled aromatic rings or aliphatic carbons occurred in catechol, ferulic or p-coumaric acids (0.1-0.7%). Demethoxylation ranged from 0.5% to 13.9% for 2,6-dimethoxyphenol and syringic acid, respectively. Methylphenols showed no demethylation. In conclusion, dehalogenation, decarboxylation and demethoxylation of phenolic substrates appear to be controlled by a common mechanism, in which various substituents are released if they are attached to carbon atoms involved in coupling. Electron-withdrawing substituents, such as -COOH and -Cl, are more susceptible to release than electron-donating ones, such as -OCH(3) and -CH(3). The release of organic substituents during polymerization and binding of phenols may add to CO(2) production in soil.     

Valorization of treated olive mill wastewater in fertigation practice

Olive mill wastewater (OMW) brings about a major environmental problem in Tunisia as well as in the other Mediterranean countries. Its strong organic load and its toxicity due to the presence of complex phenolic compounds have dire effects when applied to soil. To overcome this difficulty, the OMW pretreatment was investigated in the present work using the Fenton oxidation reaction with zero-valent iron. Then, this pretreated wastewater was valorized in fertigation practice. The effects of the addition of different concentrations of both treated and raw OMW on soil and cropping system were investigated. The treatment by Fenton oxidation with zero-valent iron could reduce 50 % of COD and decrease 53 % of phenolic compounds. OMW application had a temporary effect on the soil pH and EC. The results showed that the evolution of soil pH and EC was related to the organic matter of the soil which depends on the spread concentrations of raw or treated OMW. After 15-day incubation period, the soil pH and EC tended to stabilize and return to the control level. Moreover, this stabilization is faster in treated OMW than that in raw OMW especially for concentrations as high as 3 and 4 %. Plants cultivated with treated OMW showed an increase in their germination. The results pointed an improvement in the stem length of plants which is almost similar to that of the control for both pea and tomato, especially for high concentrations of 3 and 4 %.     

Enhanced reduction of phenol content and toxicity in olive mill wastewaters by a newly isolated strain of Coriolopsis gallica

The search for novel microorganisms able to degrade olive mill wastewaters (OMW) and withstand the toxic effects of the initially high phenolic concentrations is of great scientific and industrial interest. In this work, the possibility of reducing the phenolic content of OMW using new isolates of fungal strains (Coriolopsis gallica, Bjerkandera adusta, Trametes versicolor, Trichoderma citrinoviride, Phanerochaete chrysosporium, Gloeophyllum trabeum, Trametes trogii, and Fusarium solani) was investigated. In vitro, all fungal isolates tested caused an outstanding decolorization of OMW. However, C. gallica gave the highest decolorization and dephenolization rates at 30 % v/v OMW dilution in water. Fungal growth in OMW medium was affected by several parameters including phenolic compound concentration, nitrogen source, and inoculum size. The optimal OMW medium for the removal of phenolics and color was with the OMW concentration (in percent)/[(NH₄)₂SO₄]/inoculum ratio of 30:6:3. Under these conditions, 90 and 85 % of the initial phenolic compounds and color were removed, respectively. High-pressure liquid chromatography analysis of extracts from treated and untreated OMW showed a clear and substantial reduction in phenolic compound concentrations. Phytotoxicity, assessed using radish (Raphanus sativus) seeds, indicated an increase in germination index of 23–92 % when a 30 % OMW concentration was treated with C. gallica in different dilutions (1/2, 1/4, and 1/8).

Reusing ethyl acetate and aqueous exhausted fractions of dry olive mill residue by saprobe fungi

Some saprobe fungi (Phlebia radiata, Trametes versicolor, Coriolopsis rigida, Pycnoporus cinnabarinus, Fomes sclerodermus or Pleurotus pulmonarius) were able to bioconvert the ethyl acetate fraction (DEAF) and the corresponding aqueous exhausted fraction (EAF) of dry olive mill residue (DOR), reducing their phytotoxicity on Lepidium sativum seeds. Large amount of hydroxytyrosol together with other eight monomeric phenols were found in the native DEAF fraction, which represents a good source of antioxidants. P. radiata, T. versicolor and F. sclerodermus caused an effective phytotoxicity reduction of EAF in the concentration range of 25-3 gl(-1). In particular, in the range between 12.5 and 3 gl(-1), the EAF samples inoculated with P. radiata and F. sclerodermus surprisingly stimulated the germinability of L. sativum, suggesting their use as a potential biofertilizer. This is the first report which showed the bioconversion of the above fractions in shorter time with respect to the previous findings concerning DOR. The possible implications of laccase in the decrease of DEAF and EAF phytotoxicity was also discussed.     

Ozone oxidation and aerobic biodegradation with spent mushroom compost for detoxification and benzo(a)pyrene removal from contaminated soil

The combination of ozonation and spent mushroom compost (SMC)-mediated aerobic biological treatment was investigated in the removal of benzo(a)pyrene from contaminated soil. The performances of the process alone and combined were evaluated in terms of benzo(a)pyrene removal efficiency, mineralization efficiency (as total organic carbon removal), and soil residual toxicity (phytotoxicity to Lepidium Sativum and toxicity to Vibrio fischeri). In spite of the removal efficiency (35%) obtained by SMC-mediated biological process as a stand-alone treatment, the combined process showed a benzo(a)pyrene concentration reduction higher than 75%; the best removal (82%) was observed after 10 min pre-ozonation treatment. In particular, ozonation improved the biodegradability of the contaminant, as confirmed by the increase of CO(2) production (close to 70% compared to the control), mineralization (greater than 60%) and bacterial density (which increased by two orders of magnitude). Moreover, according to phytotoxicity tests on L. Sativum, the aerobic biological process of pre-ozonated soil decreased toxicity. According to the results achieved in the present study, ozonation pre-treatment showed an high potential to overcome the limitation of bioremediation of recalcitrant compound, but it should be carefully operated in order to maximize PAH removal efficiency as well as to minimize soil residual toxicity which can result from the formation of the oxidation intermediates.     

Biodegradation and detoxification of olive mill wastewater by selected strains of the mushroom genera Ganoderma and Pleurotus

Thirty-nine white-rot fungi belonging to nine species of Agaricomycotina (Basidiomycota) were initially screened for their ability to decrease olive-mill wastewater (OMW) phenolics. Four strains of Ganoderma australe, Ganoderma carnosum, Pleurotus eryngii and Pleurotus ostreatus, were selected and further examined for key-aspects of the OMW biodegradation process. Fungal growth in OMW-containing batch cultures resulted in significant decolorization (by 40-46% and 60-65% for Ganoderma and Pleurotus spp. respectively) and reduction of phenolics (by 64-67% and 74-81% for Ganoderma and Pleurotus spp. respectively). COD decrease was less pronounced (12-29%). Cress-seeds germination increased by 30-40% when OMW was treated by Pleurotus strains. Toxicity expressed as inhibition of Aliivibrio fischeri luminescence was reduced in fungal-treated OMW samples by approximately 5-15 times compared to the control. As regards the pertinent enzyme activities, laccase and Mn-independent peroxidase were detected for Ganoderma spp. during the entire incubation period. In contrast, Pleurotus spp. did not exhibit any enzyme activities at early growth stages; instead, high laccase (five times greater than those of Ganoderma spp.) and Mn peroxidases activities were determined at the end of treatment. OMW decolorization by Ganoderma strains was strongly correlated to the reduction of phenolics, whereas P. eryngii laccase activity was correlated with the effluent’s decolorization.     

Laccase activity in soils: considerations for the measurement of enzyme activity

Laccases (benzenediol: oxygen oxidoreductases, EC 1.10.3.2) are copper-containing enzymes that catalyze the oxidative conversion of a variety of chemicals, such as mono-, oligo-, and polyphenols and aromatic amines. Laccases have been proposed to participate in the transformation of organic matter and xenobiotics as well as microbial interactions. Several laccase assays have been proposed and used in soils. Here, we show that the optimal pH conditions for the laccase substrates 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS, pH 3-5), 2,6-dimethoxyphenol (4-5.5), L-3,4-dihydroxyphenylalanine (DOPA; 4-6), guaiacol (3.5-5), 4-methylcatechol (3.5-5), and syringaldazine (5.5-7.0) are similar between purified laccases from Trametes versicolor and Pyricularia sp. and soil extracts; the substrate affinities of purified enzymes (K(M)) and soil extracts were also similar. The laccase assays showed specificity overlap with tyrosinase and ligninolytic peroxidases when hydrogen peroxide is present. The ABTS oxidation assay is able to reliably detect the presence of 13.5 pg mL(-1) or 0.199×10(-12) mol mL(-1) of T. versicolor laccase, which is three times more sensitive than the 2,6-dimethoxyphenol-based assay and more than 40 times more sensitive than any of the other assays. The low molecular mass soil-derived compounds and the isolated fulvic and humic acids influence the laccase assays and should be removed from the soil extracts before measurements of the enzyme activity are performed.     

Decolourization and removal of some organic compounds from olive mill wastewater by advanced oxidation processes and lime treatment

Background Olive mill wastewater (OMW) generated by the olive oil extracting industry is a major pollutant, because of its high organic load and phytotoxic and antibacterial phenolic compounds which resist biological degradation. Mediterranean countries are mostly affected by this serious environmental problem since they are responsible for 95% of the worldwide oliveoil production. There are many methods used for OMW treatment, such as adsorption, electro coagulation, electro-oxidation, biological degradation, advanced oxidation processes (AOPs), chemical coagulation, flocculation, filtration, lagoons of evaporation and burning systems, etc. Currently, there is no such economical and easy solution. The aim of this study was to evaluate the feasibility of decolourization and removal of phenol, lignin, TOC and TIC in OMW by UV/H₂O₂ (AOPs). The operating parameters, such as hydrogen peroxide dosage, times, pH, effect of UV and natural sunlight were determined to find the suitable operating conditions for the best removal. Moreover, there is no study reported in the literature related to the use of UV/H₂O₂ and lime together in OMW treatment. Methods OMW was obtained from an olive-oil producing plant (Muğla area of Turkey) which uses a modern production process. No chemical additives are used during olive oil production. This study was realised by using two different UV sources, while taking the time and energy consumption into consideration. These two sources were mercury lamps and natural sunlight. Before starting AOPs experiments, one litre of OMW was treated by adding lime until a pH of 7.00. Then, 100 ml was taken from each sample, and 1 to 10 ml of a 30% H₂O₂ (Riedel-deHaen) solution was added. These solutions in closed vessels were laid in the natural sunlight for a week and their compositions and colour changes were analysed daily by UV-Vis spectrophotometer. At the end of the one-week period, they were treated with lime. In this study, the effect of changes in the initial pH, times and H₂O₂ concentrations on removal was investigated. At the end of all experiments, changes in colour, phenol, lignin, TOC and TIC concentrations were analysed according to standard methods. Results and Discussion In the samples exposed to natural sunlight and having an H₂O₂/OMW ratio of 3 ml/100 ml, a significant colour removal was achieved approximately 90% of the time at the end of 7 days. When the same samples were treated with lime (pH: up to 7), 99% efficiency was achieved. When phenol and lignin removals were examined in the same concentration, phenol and lignin removal were found 99.5%, 35%, respectively. However, for maximum lignin removal, more use of H₂O₂ (10 ml H₂O₂/100 ml OMW) was found to be necessary. Under these conditions, it was found that lignin can be removed by 70%, but to 90% with lime, at the end of a seven-day period. Rate constants obtained in the experiments performed with direct UV were found to be much higher than those of the samples exposed to natural sunlight (k^a _lignin = 0.3883 ≫ k^b _lignin = 0.0078; k^a _phenol = 0.5187 ≫ k^b _phenol = 0.0146). Moreover, it should be remembered in this process that energy consumption may induce extra financial burden for organisations. Conclusions It was found, in general, that colour, lignin, total organic carbon and phenol were removed more efficiently from OMW by using H₂O₂ UV and lime OMW. Moreover, in the study, lime was found to contribute, both initially and after radical reactions, to the efficiency to a great extent. Recommendations and Perspectives Another result obtained from the study is that pre-purification carried out with hydrogen peroxide and lime may constitute an important step for further purification processes such as adsorption, membrane processes, etc.     

Validation of germination rate and root elongation as indicator to assess phytotoxicity with Cucumis sativus

Germination rate and root elongation, as a rapid phytotoxicity test method, possess several advantages, such as sensitivity, simplicity, low cost and suitability for unstable chemicals or samples. These advantages made them suitable for developing a large-scale phytotoxicity database and especially applicable for developing quantitative structure–activity relationship (QSAR) to study mechanisms of phytotoxicity. In this paper, the comparative inhibition of germination rate and root elongation of Cucumis sativus by selected halogen-substituted phenols and anilines were determined. The suitability of germination rate and root elongation as phytotoxicity endpoints was evaluated. Excellent reproducibility and stability of germination rate and root elongation in the control test, relatively greater sensitivity and similar dose–response relations for all tested compounds were observed. These results together with those of a 2-day test were used to demonstrate the suitability of this phytotoxicity test method. A QSAR was developed for the phytotoxicity mode of action of the tested compounds to C. sativus seeds. Models that combined the logarithm of 1-octanol/water partition coefficient (log K_ow) and the energy of the lowest unoccupied molecular orbital (E_lumo) were developed for both germination rate inhibition and root elongation inhibition. The results of these studies indicate that phytotoxicity of substituted phenols and anilines to C. sativus seeds could be explained by a polar narcosis mechanism. This paper will promote the application of germination rate and root elongation method and the development of large-scale phytotoxicity database, which will provide the fundamental data for QSAR and ecological risk assessment of organic pollutants.     

Models for the field-based toxicity of copper and zinc salts to wheat in 11 Australian soils and comparison to laboratory-based models

Laboratory-based relationships that model the phytotoxicity of metals using soil properties have been developed. This paper presents the first field-based phytotoxicity relationships. Wheat (Triticum aestivum L.) was grown at 11 Australian field sites at which soil was spiked with copper (Cu) and zinc (Zn) salts. Toxicity was measured as inhibition of plant growth at 8 weeks and grain yield at harvest. The added Cu and Zn EC10 values for both endpoints ranged from approximately 3 to 4760 mg/kg. There were no relationships between field-based 8-week biomass and grain yield toxicity values for either metal. Cu toxicity was best modelled using pH and organic carbon content while Zn toxicity was best modelled using pH and the cation exchange capacity. The best relationships estimated toxicity within a factor of two of measured values. Laboratory-based phytotoxicity relationships could not accurately predict field-based phytotoxicity responses.     

Effect of aging on catalytic properties in mechanochemical degradation of pentachlorophenol by birnessite

Mechanochemistry, a technique concerning with milling contaminated samples for prolonged times, induces massive degradation of pollutants by grinding them in ball mills with different soil components or additives. In the present study, laboratory experiments were conducted to evaluate the effect of aging on the mechanochemical efficiency of the Mn-oxide birnessite in degrading pentachlorophenol (PCP). A comparative study on an aged birnessite (KBiA), used after 3 years from synthesis, and a fresh birnessite (KBiF), employed immediately after synthesis, was carried out. The differences between the two birnessites, evidenced by spectroscopic and diffractometric techniques, are mainly relative to reduction of the Mn(IV) centered at the MnO₆ octahedra layers from the birnessite structure, which represent the most reactive sites for PCP degradation. The long term air drying at room temperature, by favouring reduction of Mn(IV) to Mn(III), produces an inorganic substrate that offers paucity of the less reactive sites for PCP degradation, thus reducing the oxidative potential of the KBiA. Accordingly, the more reactive fresh birnessite was employed in the experiment with a polluted soil. Adding a small amount of KBiF to soil only induces a light increase in PCP removal, probably due to the mechanically induced PCP adsorption and transformation onto clay minerals present in the soil. Besides, adding a higher dose of birnessite causes a stronger degradation of PCP.     

Quantitative structure-activity relationships for the inhibition toxicity to root elongation of Cucumis sativus of selected phenols and interspecies correlation with Tetrahymena pyriformis

The comparative toxicities of selected phenols to higher plants Cucumis sativus were measured and the negative logarithm molar concentration of the root elongation median inhibition (IRC50) were derived. Quantitative structure-activity relationships (QSARs) were developed to explore the toxicity influencing factors and for predictive purpose. The toxicity data, fell into two classes: polar narcosis and bio-reactive. For polar narcotic phenols, a highly significant two-parameter QSAR based on 1-octanol/water partition coefficient (logKow) and energy of the lowest unoccupied orbital (E(lumo)) was derived (IRC50 = 0.77 log Kow - 0.39E(lumo) + 2.36 n = 22 r2 = 0.89). The five bio-reactive chemicals proved to show elevated toxicity due to their typical substructure involved diverse reactive mechanisms. In an effort to model all chemicals, a robust multiple-variable QSAR combining logKow, E(lumo) and Qmax, the most negative net atomic charge, was developed (IRC50 = 0.65 logKow - 0.72E(lumo) + 0.23Qmax + 2.81 n = 27 r2 = 0.94), indicating that hydrophobicity, electrophilicity and hydrogen bond interaction contribute mainly to the phytotoxicity. The toxicological data was compared with Tetrahymena pyriformis 2-d population growth inhibition toxicity (IGC50) and excellent interspecies correlations were observed both for the polar narcotics and for five reactive chemicals (for polar narcotics: IRC50 = 0.95IGC50 + 1.07 n = 16 r2 = 0.89; for bio-reactive chemicals: IRC50 = 0.98IGC50 + 2.19 n = 5 r2 = 0.97; and for all: IRC50 = 0.93IGC50 + 1.63 n = 21 r2 = 0.87). This suggested that T pyriformis toxicity could serve as a surrogate of C. sativus toxicity for phenols and interspecies correlation also could be established for reactive chemicals.     

Inactivation of enzyme laccase and role of cosubstrate oxygen in enzymatic removal of phenol from water.

Research was conducted to evaluate the potential use of laccase and its susceptibility to inactivation in an alternative enzyme-based treatment technology to remove parent phenol from buffered distilled water. Enzymatic oxidative polymerization of phenol with laccase was carried out in continuously stirred batch reactors. The reaction products were insoluble polymers, which precipitated out of the solution once their solubility limits were exceeded. The findings demonstrated that the polymeric products had significant effects on enzyme activity consumption and subsequent phenol removal. Enzyme species present in the reaction vessel were classified into enzyme remaining in the solution (type 1) and enzyme adhering to the precipitate polymers (type 2). Type 1 enzyme was more efficient in removal of phenol from solution compared with type 2. Subsequent filtration enhanced the phenol removal by removing type 2 enzyme adhering to the polymer particles and decelerating enzyme inactivation. The study also investigated the effects of available dissolved oxygen, provided through aeration and hydrogen peroxide addition, on phenol removal. Aeration and hydrogen peroxide addition increased the dissolved oxygen concentration, but had no effect on the progress curve for phenol removal.     

Potential applications of immobilized bitter gourd (Momordica charantia) peroxidase in the removal of phenols from polluted water

The potential applications of immobilized bitter gourd peroxidase in the treatment of model wastewater contaminated with phenols have been investigated. The synthetic water was treated with soluble and immobilized enzyme preparations under various experimental conditions. Maximum removal of phenols was found in the buffers of pH values 5.0-6.0 and at 40 degrees C in the presence of 0.75 mM H(2)O(2). Fourteen different phenols were independently treated with soluble and immobilized bitter gourd peroxidase in the buffer of pH 5.6 at 37 degrees C. Chlorinated phenols and native phenol were significantly removed while other substituted phenols were marginally removed by the treatment. Phloroglucinol and pyrogallol were recalcitrant to the action of bitter gourd peroxidase. Immobilized bitter gourd peroxidase preparation was capable of removing remarkably high percentage of phenols from the phenolic mixtures. Significantly higher level of total organic carbon was removed from the model wastewater containing individual phenol or complex mixture of phenols by immobilized bitter gourd peroxidase as compared to the soluble enzyme. 2,4-dichlorophenol and a phenolic mixture were also treated in a stirred batch reactor with fixed quantity of enzyme for longer duration. The soluble bitter gourd peroxidase ceased to function after 3h while the immobilized enzyme was active even after 6h of incubation with phenolic solutions.     

Membrane filtration of olive mill wastewater and exploitation of its fractions.

Olive mill wastewater (OMW) produced from small units scattered in rural areas of Southern Europe is a major source of pollution of surface and subsurface water. In the present work, a treatment scheme based on physical separation methods is presented. The investigation was carried out using a pilot-plant unit equipped with ultrafiltration, nanofiltration, and reverse osmosis membranes. Approximately 80% of the total volume of wastewater treated by the membrane units was sufficiently cleaned to meet the standards for irrigation water. The concentrated fractions collected in the treatment concentrates were characterized by high organic load and high content of phenolic compounds. The concentrates were tested in hydroponic systems to examine their toxicity towards undesired herbs. The calculations of the cost of the overall process showed that fixed and operational costs could be recovered from the exploitation of OMW byproducts as water for irrigation and/or as bioherbicides.