Detoxification of olive mill wastewaters by Moroccan yeast isolates

A total of 105 yeast strains were isolated from Moroccan olive oil production plants and evaluated for their ability to grow in olive oil mill wastewaters (OMW). The 9 isolates that grew best on OMW were selected for further study to evaluate their effect on removal of organic pollutants and OMW phytotoxicity (barley seed germination test). The results showed that at least four yeast isolates effectively lowered the toxicity of this effluent in addition to providing very useful materials in terms of both yeast biomass (6 g/l DW) and an irrigation fluid. This group of yeast isolates significantly reduced the concentration of total phenols (44% removal) and Chemical Oxygen Demand, COD (63% removal). The best germination rate of 80% for undiluted OMW was obtained for strain Candida holstii that also increased the pH from 4.76 to 6.75. Principal component analysis of the results obtained for the best yeast strains confirmed the importance of COD and total phenol reduction along with increase of organic nitrogen and final pH for the improvement of germination rates and phytotoxic reduction. This study has highlighted the potential of indigenous yeasts in detoxification of olive mill wastewaters.     

Characterisation of Paenibacillus jamilae strains that produce exopolysaccharide during growth on and detoxification of olive mill wastewaters

A total of 10 bacterial strains were isolated from a compost of corn treated with olive mill wastewaters (OMW) and selected by their capacity to synthesize exopolysaccharides (EPS). Morphological, physiological, biochemical and nutritional tests were used for a phenotypic study. A numerical analysis showed that all strains were 90% similar to each other. A DNA–DNA hybridization assay confirmed that all the strains belonged to Paenibacillus jamilae species. All the characterized strains were able to produce EPS growing on OMW batch cultures. The strain which was able to produce the highest EPS yield was chosen to perform an assay for testing its putative detoxifying activity, and it showed to reduce more than half the toxic capacity of the OMW. The results presented in this study, indicated the possible perspectives for using these bacterial strains to produce EPS and contribute to the bioremediation of the waste waters that are produced in the olive oil elaboration process.     

The effect of olive mill wastewaters variability on xanthan production

AIMS: Xanthan production by Xanthomonas campestris from several olive mill wastewaters (OMW) was investigated. METHODS AND RESULTS: Maximum xanthan production of 4 g l(-1) was obtained in media with 50% OMW as sole source of nutrients. OMW storage decreased effluent quality for xanthan production. The range of effluent concentration for X. campestris growth and xanthan production varied depending on OMW extraction METHOD: Wastewaters from press and two-phase extraction methods required higher dilution rates (< 10%) than those from the three-phase extraction method (50%). Nitrogen supplementation improved xanthan production in press and two-phase OMW. CONCLUSION: Factors affecting wastewaters composition, namely, waste storage, time of olive harvesting, and method for oil extraction, were found to influence xanthan production in shake-flask cultures. SIGNIFICANCE AND IMPACT OF THE STUDY: Conditions for xanthan production from OMW should be optimized in accordance with the nature of the waste material.     

Use of olive mill wastewater compost for crop production

Vast amounts of olive mill wastewaters (OMW) are produced in Mediterranean countries, where their treatment and disposal are becoming a serious environmental problem. Increasing attention has been paid to discovering a use for OMW and a wide range of technological treatments are available nowadays for reducing their pollutant effects and for their transformation into valuable products, the most suitable procedures being found to involve recycling rather than the detoxication of these wastes. Direct application of OMW to soil has been considered as an inexpensive method of disposal and recovery of their mineral and organic components but, because of their organic acid and phenol contents, OMW are also a source of pollution. By using composting technologies, it is possible to transform either fresh OMW or sludge from pond-stored OMW mixed with appropriate plant waste waterials (carriers) into organic fertilizers (composts) with no phytotoxicity to improve soil fertility and plant production, the process involving the microbial degradation of the polluting load of the wastes. Results of field and pot experiments using OMW-composts to cultivate horticultural and other crops have shown that yields obtained with organic fertilization are similar, and sometimes higher, to those obtained with a balanced mineral fertilizer. A comparison between the macro and micronutrient contents of plants cultivated with organic or mineral fertilizers did not generally reveal important differences. However, the cases of iron and manganese are worth mentioning as their bio-availability may be linked to the soil humic complexes originated by the OMW organic fertilizers.     

Organic matter humification in olive oil mill wastewater by abiotic catalysis with manganese(IV) oxide

The chemical changes occurring in an olive oil mill wastewater (OMW) sample digested catalytically with MnO(2) for 30 and 60 days were evaluated comparatively with those occurring in the same OMW left standing for the same time in an open-air lagoon. Both treatments increased the pH and electrical conductivity and decreased the contents of dry matter, total organic C and total N, and C/N ratio of OMW. The humic acid (HA)-like fraction isolated from the fresh OMW was characterized by a marked aliphatic character, small O and acidic functional group contents, marked presence of proteinaceous materials, partially modified lignin moieties and polysaccharides-like structures, extended molecular heterogeneity, and small degrees of aromatic ring polycondensation, polymerization and humification. With increasing the time of either lagooning or catalytic digestion, a loss of aliphatic materials and an increase of extraction yield, oxygenation, acidic functional groups, carbohydrates and aromaticity occurred in the HA-like fractions. The more evident changes measured for the HA-like fractions from catalytically-digested OMW, with respect to those from lagooned OMW, indicated that MnO(2) was able to catalyze organic matter humification in OMW.     

Modelling of the mesophilic anaerobic co-digestion of olive mill wastewater with olive mill solid waste using anaerobic digestion model No. 1 (ADM1)

The anaerobic digestion model No. 1 (ADM1), conceived by the international water association (IWA) task group for mathematical modelling of anaerobic digestion processes is a structured generic model which includes multiples steps describing biochemical and physicochemical processes encountered in the anaerobic degradation of complex organic substrates and a common platform for further model enhancement and validation of dynamic simulations for a variety of anaerobic processes. In this study the ADM1 model was modified and applied to simulate the mesophilic anaerobic co-digestion of olive mill wastewater (OMW) with olive mill solid waste (OMSW). The ADM1 equations were coded and implemented using the simulation software package MATLAB/Simulink. The most sensitive parameters were calibrated and validated using updated experimental data of our previous work. The results indicated that the ADM1 model could simulate with good accuracy: gas flows, methane and carbon-dioxide contents, pH and total volatile fatty acids (TVFA) concentrations of effluents for various feed concentrations digested at different hydraulic retention times (HRTs) and especially at HRTs of 36 and 24 days. Furthermore, effluent alkalinity and ammonium nitrogen were successfully predicted by the model at HRTs of 12 and 24 days for some feed concentrations.     

Evolution of the fulvic acid fractions during co-composting of olive oil mill wastewater sludge and tree cuttings

Fulvic acids (FAs) were isolated by a conventional procedure from two mixtures of the sludge residue obtained from olive oil mill wastewater (OMW) evaporated in open-air pond and tree cuttings (TC) at different stages of the co-composting process. The FAs were analyzed for elemental (C, H, N, S, O) and acidic functional group (carboxylic and phenolic) composition, and by ultraviolet/visible, Fourier transform infrared and fluorescence spectroscopies. At the initial stage of composting, FAs from the OMW sludge-TC mixtures were characterized by a prevalent aliphatic character, large contents of C, S-containing groups, proteinaceous materials and polysaccharide components, extended molecular heterogeneity, small O and acidic functional group contents, and small degrees of aromatic ring polycondensation, polymerization and humification. As composting proceeded, C, H and S contents, C/N ratio, and aliphaticity decreased, whereas N, O, COOH and phenolic OH contents, C/H and O/C ratios, and aromaticity increased. These results suggested that, with increasing the composting time, the chemical and structural properties of the FA components of the two OMW sludge-TC mixtures approached the characteristics typical of native soil FAs. Thus, co-composting of OMW sludge mixed with TC may represent a suitable treatment for enhancing the quality of organic matter in these materials when used as soil amendments.     

Determination of aerobic biodegradation kinetics of olive oil mill wastewater

Aerobic biological treatment was conducted for the treatment of high strength olive oil mill wastewater (OMW). Two different approaches were used for kinetic modeling of OMW biodegradation. TOC removal and CO₂–C evolution were monitored in an open and a closed bioreactor systems, respectively. Gompertz, Refractory organics plus first-order (RFO) and Chen–Hashimoto equations were applied to estimate the kinetic parameters by using the data from bioreactors. Furthermore, change in oxidation stage of carbon was monitored and temperature dependency of OMW biodegradation was investigated based on activation energy. At room temperature, 64% of TOC was removed in the open bioreactor while cumulative CO₂–C evolution was 6.32 g L⁻¹ in closed the bioreactor. Higher biodegradation efficiency and kinetic parameters were obtained at 25 °C rather than 10 °C. Gompertz and RFO equations provided better fitting with CO₂–C and TOC data, respectively. Experimental and kinetic estimations indicated that OMW constituted of approximately 30% refractory organics. The comparison of two different modeling approaches showed that kinetic modeling based on CO₂–C provided better correlation with the experimental data. Temperature coefficient indicated that biological degradation of OMW is slightly dependent on temperature.     

Anaerobic co-digestion of olive mill wastewater with olive mill solid waste in a tubular digester at mesophilic temperature

Anaerobic co-digestion is a well established process for treating many types of organic wastes, both solid and liquid. In this study we have investigated, on a laboratory scale, the anaerobic co-digestion of olive mill wastewater (OMW) with olive mill solid waste (OMSW) using semi-continuous, feeding, tubular digesters operated at mesophilic temperatures. Each digester was fed with an influent, composed of OMW and OMSW, at an organic loading rate (OLR) varying between 0.67 and 6.67 g COD/l/d. The hydraulic retention times (HRT) were 12, 24 and 36 days. The TCOD concentrations of OMW used as the main substrate were 24, 56 and 80 g COD/l; the amount of the dry OMSW used as a co-substrate was fixed to approximately 56 g/l of OMW. The results indicated that the best methane production was about 0.95 l/l/day obtained at an OLR = 4.67 g COD/l/d, corresponding to influent TCOD = 56 g COD/l at an HRT = 12d. In contrast, the maximum TCOD removal efficiency (89%) was achieved at an OLR = 0.67 g COD/l/d, corresponding to influent TCOD = 24 g COD/l at an HRT = 36 d. Moreover, the inhibition of biogas production was observed at the highest OLR studied.     

Olive orchard amended with two experimental olive mill wastes mixtures: effects on soil organic carbon, plant growth and yield

Amendments of olive orchard soil with two different preparations of olive mill solid waste (OMWMs) at the rate of 9tonha(-1) per year for five years in two different plots were compared with an industry standard soil amendment using urea. Both the OMWMs amendments showed significant increases in total organic carbon and humic substances in soil of approximately 40% and 58%, respectively, without negative effects on tree growth and yield. This work has shown that olive oil mill waste (OMW) can be recycled safely using the bioremediation system used in this study. We suggest that this system is particularly beneficial to organic farming and is an alternative solution to direct spreading of raw OMW on farm lands.     

Changes in microbial and soil properties following amendment with treated and untreated olive mill wastewater

We investigated the effect of untreated and biologically treated olive mill wastewater (OMW) spreading on the soil characteristics and the microbial communities. The water holding capacity, the salinity and the content of total organic carbon, humus, total nitrogen, phosphate and potassium increased when the spread amounts of the treated or untreated OMW increased. The OMW treated soil exhibited significantly higher respiration compared to the control soil. However, the C-CO2/C(tot) ratio decreased from 1.7 in the control soil to 0.5 in the soil amended with 100 m3 ha(-1) of untreated OMW. However, it slightly decreased to 1.15 in the soil amended with 400 m3 ha(-1) of treated OMW. The treated OMW increased the total mesophylic number while the number of fungi and nitrifiers decreased. Actinomycetes and spore-forming bacteria were neither sensitive to treated nor to untreated OMW. The total coliforms increased with higher doses of treated and untreated OMW. A toxic effect of the untreated OMW appeared from 100 m3 ha(-1). This toxicity was more significant with 200 m3 ha(-1), where microflora of total mesophilic, yeasts and moulds, actinomycetes, and nitrifiers were seriously inhibited except for total coliforms and spore-forming bacteria.     

Activity and elution profile of laccase during biological decolorization and dephenolization of olive mill wastewater

The performance and enzymatic strategy exhibited by basidiomycete Euc-1, a laccase producing strain, was investigated during the biodegradation of olive mill wastewater (OMW). This strain yielded better decolorization of solidified OMW than Phanerochaete chrysosporium and removed 90% of phenols (initial concentration=800 mg l(-1)), 73% of color (initial A465=4.4), and 45% of chemical oxygen demand in batch cultures containing OMW. Since partial phenol removal occurred before the detection of enzymatic activity, no plausible correlation could be established between them. In contrast, decolorization occurred only after the detection of laccase activity and coincided with its production over time. Two laccase fractions (Lac1 and Lac2) were separated by chromatography. OMW strongly induced Lac2 that was almost absent in defined liquid medium. Furthermore, Lac2 was the main laccase fraction in the presence of OMW. This study pointed out that basidiomycete Euc-1 and its ligninolytic system could be a useful tool for the bioremediation of wastewater generated in the process of olive oil extraction.     

Removal of low molecular weight phenols from olive oil mill wastewater using microalgae

The treatment of olive oil mill wastewater (OMW) with two phenol resistant algae, Ankistrodesmus braunii and Scenedesmus quadricauda, showed a limited reduction of phenol content after 5 d of treatment, irrespective of algal concentration. Otherwise, cultures of both algae, grown in the dark, degraded over 50% of the low molecular weight phenols contained in OMW, but they were not completely removed, but were biotransformed into other non-identified, aromatic compounds.     

Lipase production by Aspergillus ibericus using olive mill wastewater

Olive mill wastewater (OMW) characteristics make it a suitable resource to be used as a microbial culture media to produce value-added compounds, such as enzymes. In this work, the ability of the novel species Aspergillus ibericus to discolor OMW and produce lipase was studied. An initial screening on plates containing an OMW-based agar medium and an emulsified olive oil/rhodamine-B agar medium was employed to select the strain A. ibericus MUM 03.49. Then, experiments in conical flasks with liquid OMW-based media showed that the fungus could growth on undiluted OMW, with a chemical oxygen demand (COD) of 97 ± 2 g/L, and to produce up to 2,927 ± 54 U/L of lipase. When pure OMW was used in the media, the maximum COD and color reduction achieved were 45 and 97 %, respectively. When OMW diluted to 10 % was used, A. ibericus was able to reduce phenolic and aromatic compounds by 37 and 39 %, respectively. Additionally, lipase production was found to be promoted by the addition of mineral nutrients. When the fermentations were scaled up to a 2-L bioreactor, A. ibericus produced up to 8,319 ± 33 U/L of lipase, and the maximum COD and color reduction were 57 and 24 %, respectively.     

Use of Geotrichum candidum for olive mill wastewater treatment in submerged and static culture

The production of lignin peroxidase (LiP), manganese peroxidase (MnP) and lipases by Geotrichum candidum were performed in order to control the decolourisation and biodegradation of olive mill wastewater (OMW). Optimisation of different factors showed that dilution, carbon and ammonium concentrations significantly affected decolourisation and activities of ligniolytic peroxidases (LiP and MnP) on OMW. Moreover, addition of olive oil and agitation improved the lipase production. Batch and continuous OMW treatments in settler or bubble column bioreactors showed high COD and colour removal efficiencies of 60% and 50%, respectively. Lipolytic activity was greater in the batch bubble column whereas, LiP and MnP productions were improved in the settler. The performance of the continuous processes decreased with the decrease of hydraulic retention time (HRT). It has been shown that decolourisation and biodegradation decreased with an average of 40% and 45%, respectively, by decreasing the HRT from 4 d to 1.7 d.     

The use of hydroxyl-radical-generating systems for the treatment of olive mill wastewaters

Three hydroxyl-radical producing biomimetic systems, composed of CuII, hydrogen peroxide and pyridine, glucaric or succinic acid, were able to perform decolorization of olive mill wastewaters (OMW) >85 % within 3 d combined with a significant removal of total phenols (>75 %). The systems consisting of 50 mmol/L succinic acid, 5-10 mmol/L CuSO4 and 100 mmol/L H2O2 were the most effective at OMW treatment, and led to the reduction of phenol contents to <1 % along with high decolorization (>88 %) and acceptable values of chemical oxygen demand.     

Phenolic removal in olive oil mill wastewater using loofah-immobilized Phanerochaete chrysosporium

Summary Olive oil mill wastewater (OMW) has a high organic load, and this is a serious concern of the olive industry. Conventional biological wastewater treatments, despite their simplicity and suitable performance are ineffective for OMW treatment since phenolics possess antimicrobial activity. In order to carry out a proper treatment of OMW, use of a microorganism able to degrade the phenolics is thus necessary. In this study the ability of Phanerochaete chrysosporium to degrade the phenolic compounds of OMW and to decrease the chemical oxygen demand (COD) using cells immobilized on loofah was examined. The basal mineral salt solution along with glucose, ammonium sulfate and yeast extract was used to dilute the OMW appropriately. The fungus did not grow on the concentrated OMW. The extent of removal in this bio-treatment, of total phenols (TP) and the COD were 90 and 50%, respectively, while the color and aromaticity decreased by 60 and 95%, respectively. The kinetic behavior of the loofah-immobilized fungus was found to follow the Monod equation. The maximum growth rate μ_max was 0.045 h⁻¹ while the Monod constant based on the consumed TP and COD were (mg/l) 370 and 6900, respectively.     

Roles of two white-rot basidiomycete fungi in decolorisation and detoxification of olive mill waste water

A Phanerochaete chrysosporium strain was isolated from Moroccan olive mill waste water (OMW) and its ability to degrade OMW in different culture conditions was investigated and compared to that of Pleurotus ostreatus. The results indicated that Ph. chrysosporium isolate is more efficient than Pl. ostreatus in decolorising and detoxifying OMW in the presence of added nutrients. Ph. chrysosporium is able to remove more than 50% of the colour and phenols from OMW within 6 days of incubation, whereas Pl. ostreatus needs more than 12 days to reach similar results in the same conditions. Many factors affecting the treatment of diluted OMW (20%) by Ph. chrysosporium were studied, including the effects of added nutrients, initial pH, temperature and inoculated biomass. Once the optimisation of 20% OMW biodegradation process had been set up, higher OMW concentrations (50%) were tested. The results show that the fungus is capable of reducing all parameters analysed (colour A395, phenol content and chemical oxygen demand) by at least 60%, after only 9 days of growth.     

Biological treatment of olive mill wastewater by non-conventional yeasts

The ability of lipolytic yeasts to grow on olive mill wastewater (OMW)-based medium and to produce high-value compounds while degrading this waste, was tested. OMW collected from three-phase olive mills from the North region of Portugal were characterized and used. OMW with COD ranging from 100 g L⁻¹ to 200 g L⁻¹ were supplemented with yeast extract and ammonium chloride. Studies of OMW consumption were carried out in batch cultures of Candida rugosa, Candida cylindracea and Yarrowia lipolytica. All strains were able to grow in the OMW-based media, without dilution, to consume reducing sugars and to reduce COD. C. cylindracea was the best strain concerning the lipase production and the reduction of phenolic compounds and COD. For all strains, the phenols degradation was quite difficult, mostly when more easily degradable carbon source is still present in the medium. Among the phenolic compounds tested catechol is the most inhibitory to the cells.     

Effects of agronomic application of olive mill wastewater in a field of olive trees on carbohydrate profiles, chlorophyll a fluorescence and mineral nutrient content

Olive mill wastewater (OMW) management is a serious environmental issue for the Mediterranean area where there is the most production of olive oil. OMW contains a high organic load, substantial amounts of plant nutrients but also several compounds with recognized toxicity towards living organisms. Moreover, OMW may represent a low cost source of water. We studied the influence of irrigation with OMW (amounts applied: 30, 60, 100 and 150 m³ h⁻¹) in a field of olive trees on root colonization, photosynthesis, chlorophyll fluorescence, leaf nutrient concentration and soluble carbohydrate. The soil fatty acid methyl ester (FAME) 16:1ω5 was used to quantify biomass of arbuscular mycorrhizal (AM) fungi and the root FAME 16:1ω5 analysis was used as index for the development of colonization in the roots. Agronomic application of OMW decreased significantly the abundance of the soil FAME 16:1ω5 and the root FAME 16:1ω5 in the soil amended with 60, 100 and 150 m³ ha⁻¹ OMW. Decreased root FAME 16:1ω5 due to OMW amendment was associated with a significant reduction of tissue nutrient concentrations in the olive trees. The highest application of OMW to the soil reduced significantly the olive trees uptake of N, P, K, Ca, Mg, Fe, Cu, Mn and Zn. Land spreading of OMW increased concentration of soluble carbohydrate in the olive leaves, mostly due to decreased sink demand for carbon by the root. In the olive trees amended with 150 m³ ha⁻¹ OMW, net CO₂ uptake rate (A), quantum yield of photosystem II electron transport (Φ_PSII), maximal photochemical efficiency of photosystem II (Fv/Fm), photochemical quenching (q_p) and the electron transport rate (ETR) were significantly depressed, whereas non-photochemical quenching (NPQ) was found to increase. Taken with data from experiments in field conditions, our results suggest that agronomic application of OMW alters the functioning of arbuscular mycorrhizas and can even disrupt the relationship between AM fungi and olive trees.