Phenolic removal in olive oil mill wastewater by strains of Pleurotus spp. in respect to their phenol oxidase (laccase) activity

The ability of several Pleurotus spp. strains to remove phenolic compounds from an olive oil mill wastewater (OMW) was studied. All strains tested in this work were able to grow in OMW without any addition of nutrients and any pre-treatment, except sterilization. High laccase activity was measured in the growth medium, while 69-76% of the initial phenolic compounds were removed. The black color of OMW became yellow-brown and brighter as the strains grew. The lowest phenolic concentrations were reached after 12/15 days. A decrease of the phytotoxicity, as described by the parameter Germination Index, was noticed in the OMW treated with some Pleurotus spp strains, although this decrease was not proportional to the phenolic removal. A new parameter, namely Phenol-toxicity Index, was considered in the present paper. Using this parameter it was found that the remaining phenolics and/or some of the oxidation products of the laccase reaction in the treated OMW were more toxic than the original phenolic compounds.     

Antimicrobial activity of olive mill wastewaters (alpechin) and biotransformed olive oil mill wastewater

Olive oil extraction produces a great volume of residue. These olive mill wastes are known as alpechin. This wastewater is a powerful pollutant, resistant to degradation and presents a severe environmental problem related to its high organic content made up largely of simple phenolic compounds, that have been described as being both antimicrobial and phytotoxic. This paper reviews briefly the antimicrobial activity of olive mill wastewaters and provides evidence to show the potential of micro organisms (Bacillus pumilus) to reduce the phenol content of alpechin, and also that biotransformation depends on the dilution (v/v) of the alpechin. Furthermore, we sought to provide a real evaluation of the extent of alpechin biotransformation. This was achieved by means of an internal reference, i.e. in relative terms, the phenol content resulting from the biotransformation process. The phenol content was measured using HPLC techniques, and results were obtained showing that the bacterium had most effect in reducing the phenol content of alpechin at concentrations of between 40 and 100%. It was also observed that at concentrations of 80%, in addition to a slight reduction in total phenols, new phenolic compounds, not present in the original alpechin, were generated.     

Production of microalgae from olive mill wastewater

With olive mill wastewater (alpechin) used as a nutrient medium, the influence of the aeration level and the composition of the culture medium were analysed in relation to the concentration of alpechin and KNO₃ added in a batch culture of the microalgae Chlorella pyrenoidosa and Scenedesmus obliquus. The pH and temperature of the culture were previously fixed at 6.5 and 30°C. As kinetic parameters, the maximum specific growth rate (μ_m) and the biomass productivity (P_max) were determined. The biomass composition obtained was evaluated for chlorophyll and crude protein content as well as fatty-acid composition corresponding to the lipid fraction. To achieve a balanced biomass composition in terms of proteins and lipids, the most suitable conditions included an alpechin concentration of 10%, without the addition of nitrates and with an aeration level of 1 v/v min. Under these conditions, the μ_m values reached with both microalgae were around 0.03h⁻¹.     

Use of Yarrowia lipolytica strains for the treatment of olive mill wastewater

The principal aim of this research was to evaluate the ability of different Yarrowia lipolytica strains, having different origin, to grow in olive mill wastewater (OMW) and reduce its COD level. All the strains were able to grow in undiluted OMW; the comparison between the data obtained in a semi-synthetic medium and in OMW suggests that lipases with different specificity can be produced in relation to the medium composition. Under the adopted conditions, the reduction of the OMW COD values varied from 1.47% and 41.22% of the initial value. Some strains determined a significant reduction of polyphenol content, while other ones caused its apparent increase. Moreover, some Y. lipolytica strains, isolated from chilled foods, produced the highest citric acid concentrations. These results evidenced that some Y. lipolytica strains are good candidates for the reduction of the pollution potential of OMW and for the production of enzymes and metabolites such as lipase and citric acid.     

Oxirane-immobilized Lentinula edodes laccase: stability and phenolics removal efficiency in olive mill wastewater

Immobilization of Lentinula edodes laccase on Eupergit C increased pH, thermal and proteolytic stability with slight modifications in laccase oxidation efficiency. Immobilized laccase proved to be efficiently stable in removing olive mill wastewater phenolics.     

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.     

Olive mill wastewater disposal in evaporation ponds in Sfax (Tunisia): moisture content effect on microbiological and physical chemical parameters

The study of the isotherms desorption of olive mill wastewater (OMW) was investigated to describe its water activity under different saturated environments. The microbial biodegradation of OMW during its storage in 5 evaporation ponds located in Agareb (Sfax-Tunisia) was carried out during the oil-harvesting year held 105 days in 2004. Gravimetric static method using saturated salt solutions was used and OMW as placed at 30°C and under different water activities ranging from 0.11 to 0.90. Eight models were taken from the literature to describe experimental desorption isotherms. During storage, the evolution of physico-chemical parameters including pH, temperature, evaporation, humidity, total phosphorus, chemical oxygen demand (COD), biological oxygen demand (BOD) and phenols and three microbiological flora (aerobic mesophilic bacteria, yeasts and moulds) were considered. At 30°C, when relative humidity increased in the experimented ponds of 69, 84 and 90%, the evaporation speed decreased from 1.24 × 10⁻⁵ to 5 × 10⁻⁶ cm³ s⁻¹, from 6 × 10⁻⁵ to 7 × 10⁻⁶ cm³ s⁻¹ and from 5 × 10⁻⁶ to 1.1 × 10⁻⁷ cm³ s⁻¹ respectively. The desorption isotherm exhibited a sigmoidal curve corresponding to type II, typical of many organic material. The GAB and Peleg models gave the best fit for describing the relationship between the equilibrium moisture content and water activity in OMW (R ² = 0.998). During the storage period, the analysis showed an increase of all the physico-chemical parameters studied, except phenols and total phosphorus concentrations. The microbiological study showed the predominance of yeasts and moulds and the decrease of bacteria population after 75 days reflecting both effect of recalcitrant compounds and the water activity on microbial growth.     

Potentials of anaerobic treatment for catalytically oxidized olive mill wastewater (OMW)

The catalytically oxidized olive mill wastewater (OMW) was subjected to continuous anaerobic treatment using two treatment schemes. The 1st step in both schemes was an up-flow anaerobic sludge blanket (UASB) reactor (2 0 l). The 2nd step was either a hybrid UASB reactor or a classical one (1 0 l, each). The 1st stage was operated at constant hydraulic retention time (HRT) of 24 h. The organic loading rate (OLR) varied from 3.4 to 4.8 kgCOD/m³ d depending on the quality of the pretreated wastewater. The results obtained indicated that, the 1st step UASB reactor achieved a COD percentage removal value of 53.9%. Corresponding total BOD₅ and TSS removal were 51.5% and 68.3%, respectively.The results obtained indicated that the hybrid UASB reactor as a 2nd step produced better quality effluent as compared to the classical one. This could be attributed to the presence of the packing curtain sponge with active biomass in the sedimentation part of hybrid UASB reactor which minimizes suspended solids washout, consequently enhancement of the efficiency of the reactor.Available data showed that a two stage system consisting of a classical and a hybrid UASB reactor operated at a total HRT of 48 h and OLR of 2.0 kgCOD/m³ d provided promising results. Removal values of COD_total, BOD_5 total, TOC, VFA, oil and grease were 83%, 84%, 81%, 93% and 81%, respectively. Based on the available data, the use of a two stage anaerobic system consisting of a classical UASB reactor followed by a hybrid UASB as a post-treatment step for catalytically oxidized OMW is recommended.     

Photochemical UV/TiO2 treatment of olive mill wastewater (OMW)

Olive mill wastewater (OMW) was treated by photocatalysis using TiO2 under UV irradiation on the laboratory scale. The chemical oxygen demand, the coloration at 330nm, and the level of phenols all showed decreases which, after a 24-h treatment, reached 22%, 57% and 94%, respectively. The differences between these three values indicate the persistence of colourless, non-phenolic compounds. Application of the novel Fictitious Atomic-Group Separation method showed an increase in carbon oxidation state and confirmed that the attack primarily concerns, aromatic moieties. A fine spectroscopic study revealed the occurrence of three successive phases during the degradation process, thought to correspond to three different categories of molecules in the OMW and the presence of pectin compounds.     

Engineered tobacco and microalgae secreting the fungal laccase POXA1b reduce phenol content in olive oil mill wastewater

Olive oil mill wastewaters (OMWs) are characterised by low pH and a high content of mono- and polyaromatic compounds that exert microbial and phytotoxic activity. The laccase cDNA of the poxA1b gene from Pleurotus ostreatus, carrying a signal peptide sequence for enzyme secretion and driven by the CaMV 35S promoter, was cloned into a plant expression vector. Nuclear genetic transformation was carried out by co-cultivation of Agrobacterium tumefaciens with tobacco cv Samsun NN leaves and cells of five different microalgae accessions belonging to the genera Chlamydomonas, Chlorella and Ankistrodesmus. Transgenic plants and microalgae were able to express and secrete the recombinant laccase in the root exudates and the culture medium, respectively. In comparison to untransformed controls, the ability to reduce phenol content in OMW solution was enhanced up to 2.8-fold in transgenic tobacco lines and by up to about 40% in two microalgae accessions. The present work provides new evidence for metabolic improvement of green organisms through the transgenic approach to remediation.     

Engineered tobacco and microalgae secreting the fungal laccase POXA1b reduce phenol content in olive oil mill wastewater

Olive oil mill wastewaters (OMWs) are characterised by low pH and a high content of mono- and polyaromatic compounds that exert microbial and phytotoxic activity. The laccase cDNA of the poxA1b gene from Pleurotus ostreatus, carrying a signal peptide sequence for enzyme secretion and driven by the CaMV 35S promoter, was cloned into a plant expression vector. Nuclear genetic transformation was carried out by co-cultivation of Agrobacterium tumefaciens with tobacco cv Samsun NN leaves and cells of five different microalgae accessions belonging to the genera Chlamydomonas, Chlorella and Ankistrodesmus. Transgenic plants and microalgae were able to express and secrete the recombinant laccase in the root exudates and the culture medium, respectively. In comparison to untransformed controls, the ability to reduce phenol content in OMW solution was enhanced up to 2.8-fold in transgenic tobacco lines and by up to about 40% in two microalgae accessions. The present work provides new evidence for metabolic improvement of green organisms through the transgenic approach to remediation.     

Conversion of xylose to ethanol by a novel phenol-tolerant strain of Enterobacteriaceae isolated from olive mill wastewater

A xylose-fermenting bacterium of the family Enterobacteriaceae was isolated from olive mill wastewater. It converted xylose to ethanol with a yield of 0.19 g ethanol g^–1 xylose. Although phenolic compounds normally inhibit pentose-utilizing microorganisms, this isolate was tolerant to phenol. Both the yield and the productivity of xylose fermentation decreased by 30% when phenol was added at a final concentration of 0.8 g phenol l^–1. Xylose (23 g l^–1) was totally fermented to ethanol (4.3 g l^–1) within 48 h in the absence of phenol; however, in the presence of 0.8 g phenol l^–1, only 3.3 g ethanol l^–1 was obtained from the same starting concentration of xylose after 70 h.     

中国栓孔菌属漆酶的筛选与应用

漆酶是一种含铜的多酚氧化酶[1],广泛分布于植物、真菌、少数昆虫和细菌中,可用于纸浆造纸、生物合成、食品、能源、木材加工、环保、改善纤维特性、生物检测等多个领域。鉴于其重要的应用价值,漆酶正日益受到重视。     

Composting of the solid fraction of olive mill wastewater with olive leaves: organic matter degradation and biological activity

The flocculated solid fraction of olive mill wastewaters, obtained from two different olive oil extraction systems (FOMW1 and FOMW2) was composted, with olive leaves (OL) as bulking agent, by the static pile system (Rutgers). The dynamic of organic matter (OM) degradation during composting and its relationship with the basal respiration and fluorescein diacetate (FDA) hydrolytic activity, as indicators of biological activity, were studied. Two mixtures were prepared: C1, from 65% FOMW1 plus 35% OL; and C2, from 74% FOMW2 plus 25% OL and 1% urea. The biooxidative phase of composting in C1, which had a high initial C/N ratio, was long, leading to a high OM degradation, mainly of the lignocellulosic compounds. The water-soluble organic carbon content, C/N ratio and the urea supplied as a N source for the C2 compost make this mixture more adequate for composting, as it had a shorter composting time than C1, and developed a microbial population with a high metabolic activity. The results for basal respiration in C1 and C2 were correlated at a high probability level with those of FDA hydrolysis, and both parameters can be used for establishing the degree of biological stability of the composting material.     

Drug resistance in Salmonella strains isolated from domestic wastewater before and after treatment in stabilization ponds in an arid region (Marrakech,Morocco)

Some 118 Salmonella strains isolated before and after treatment in stabilization ponds were tested for antimicrobial resistance. In the treatment plant, which decreases the abundance of Salmonella by 99%, a significantly lower level of antibiotic resistance (P<0.01) was identified at the system's inflow point (19%) than at its outflow (29%). The serotypes most frequently identified as having multiple antibiotic resistance were Salmonella paratyphi B and S. typhimurium. High tetracycline resistance was observed at all sampling points, followed by resistance to ampicillin and streptomycin. Antibiotic resistance can be transferred from Salmonella to other members of the Enterobacteriaceae family, such as Escherichia coli K12; transfer frequencies in nutrient broth and filtered sewage water were 4.5×10^-4 and 7×10^-7, respectively.The authors are with the Université CADI AYYAD, Faculté des Sciences—Semlalia, Département de Biologie, Laboratoire de Microbiologie, Bd Le Prince My Abdallah, BP. S.15, Marrakech, Morocco.     

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.     

Effect of clay pretreatment on photofermentative hydrogen production from olive mill wastewater

The aim of this paper was to gain further insight into the effect of the clay pretreatment process on photofermentative hydrogen production. This two-stage process involved a clay pretreatment step followed by photofermentation which was performed under anaerobic conditions with the illumination by Tungsten lamps. Rhodobacter sphaeroides O.U.001 was used for photofermentation. Higher amounts of color (65%), total phenol (81%) and chemical oxygen demand (31%) removal efficiencies were achieved after clay pretreatment process. During photofermentative hydrogen production with the effluent of clay pretreatment process, the main organic compounds resulting higher hydrogen production rates were found to be acetic, lactic, propionic, and butyric acids. Compared to photofermentation using raw olive mill wastewater ( 16LH2/LOMW), the amount of photofermentative hydrogen production was doubled by using the effluent of the clay pretreatment process (31.5LH2/LOMW). The reasons for the improvement of hydrogen production by clay treatment can be attributed to the high removal of the hardly biodegradable compounds such as phenols; minor removal of organic acids, sugars and amino acids that are known to enhance photofermentative hydrogen production; and the color depletion of raw OMW which might cause a shadowing effect on the photosynthetic bacteria.     

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.     

Optimisation of biodegradation conditions for the treatment of olive mill wastewater

The aim of the present paper was to optimise the conditions of aerobic treatment of olive mill wastewater. To do so, the waste was treated following the experimental optimal design methodology studying the set of factors susceptible to influence the treatment (pH, C/N ratio, aeration and temperature). The results of a first series of experiments showed a strong correlation between the reduction in the levels of polyphenols and three of the parameters studied, i.e. the C/N ratio, aeration and temperature. Optimised conditions led to a 94% drop in polyphenols.

Then, for a finer study of the conditions, just two parameters were varied, the pH and the C/N ratio. The results showed that the conditions of pH modification (addition of lime or sodium hydroxide) and the C/N ratio (urea or ammonium nitrate) allowed the microbiological activity to be very significantly improved. This led to polyphenol reductions of 51% and 76%.