Tahar Mechichi

Decolourization and detoxification of textile industry wastewater by the laccase-mediator system

Decolourization and detoxification of a textile industry effluent by laccase from Trametes trogii in the presence and the absence of laccase mediators was investigated. Laccase alone was not able to decolourize the effluent efficiently even at the highest enzyme concentration tested: less than 10% decolourization was obtained with 9 U/mL reaction mixture. To enhance effluent decolourization, several potential laccase mediators were tested at concentrations ranging from 0 to 1mM. Most potential mediators enhanced decolourization of the effluent, with 1-hydroxybenzotriazol (HBT) being the most effective. The effect of several physico-chemical parameters that could influence enzyme activity, such as pH, temperature and dye concentration was tested. Optimal decolourization occurred with 20% effluent at pH 5, a temperature of 50 degrees C, and in the presence of 1mM HBT. The toxicities of crude, laccase-HBT treated and laccase-acetosyringone treated effluent were evaluated using the Microtox assay. Only laccase-acetosyringone treated effluent was not toxic; crude and laccase-HBT treated effluent retained toxicity.

Clostridium methoxybenzovorans sp. nov., a new aromatic o-demethylating homoacetogen from an olive mill wastewater treatment digester.

A strictly anaerobic, spore-forming bacterium (3.0-5.0 x 0.4-0.8 microns), designated strain SR3T (T = type strain), which stained Gram-positive and possessed a Gram-positive type cell wall was isolated from a methanogenic pilot-scale digester fed with olive mill wastewater (Sfax, Tunisia). It utilized a number of carbohydrates (glucose, fructose, sorbose, galactose, myo-inositol, sucrose, lactose, cellobiose), organic compounds (lactate, betaine, sarcosine, dimethylglycine, methanethiol, dimethylsulfide), alcohol (methanol) and all methoxylated aromatic compounds only in the presence of yeast extract (0.1%). The end products from carbohydrate fermentation were H2, CO2, formate, acetate and ethanol, that from lactate was methanol, those from methoxylated aromatics were acetate and butyrate, and that from betaine, sarcosine, dimethylglycine, methanethiol and dimethylsulfide was only acetate. Strain SR3T was non-motile, had a G+C content of 44 mol% and grew optimally at 37 degrees C and pH 7.4 on a glucose-containing medium. Phylogenetically, the closest relatives of strain SR3T were the non-methoxylated aromatic-degrading Clostridium xylanolyticum, Clostridium aerotolerans, Clostridium sphenoides and Clostridium celerecrescens (mean similarity of 98%). On the basis of the phenotypic, genotypic and phylogenetic characteristics of the isolate, it is proposed to designate strain SR3T as Clostridium methoxybenzovorans sp. nov. The type strain is SR3T (= DSM 12182T).

Co-composting of spent coffee ground with olive mill wastewater sludge and poultry manure and effect of Trametes versicolor inoculation on the compost maturity.

The co-composting of spent coffee grounds, olive mill wastewater sludge and poultry manure was investigated on a semi-industrial scale. In order to reduce the toxicity of the phenolic fraction and to improve the degree of composting humification, composts were inoculated with the white-rot fungus Trametes versicolor in the early stages of the maturation phase. During composting, a range of physico-chemical parameters (temperature and both organic matter and C/N reduction), total organic carbon, total nitrogen, elemental composition, lignin degradation and spectroscopic characteristics of the humic acids (HAs) were determined; impacts of the composting process on germination index of Hordeum vulgare and Lactuca sativa were assessed. The coffee waste proved to be a highly compostable feedstock, resulting in mature final compost with a germination index of 120% in less than 5 months composting. In addition, inoculation with T. versicolor led to a greater degree of aromatization of HA than in the control pile. Moreover, in the inoculated mixture, lignin degradation was three times greater and HA increased by 30% (P<0.05), compared to the control pile. In the T. versicolor inoculated mixture, the averages of C and N were significantly enhanced in the HA molecules (P<0.05), by 26% and 22%, respectively. This improvement in the degree of humification was confirmed by the ratio of optical densities of HA solutions at 465 and 665 nm which was lower for HA from the treated mixture (4.5) than that from the control pile (5.4).

Structural, physicochemical and antioxidant properties of sodium alginate isolated from a Tunisian brown seaweed

An original sodium alginate from Tunisian seaweed (Cystoseira barbata) was purified and characterized by circular dichroism (CD) and ATR-FTIR spectroscopies. ATR-FTIR spectrum of C. barbata sodium alginate (CBSA) showed the characteristic bands of mannuronic (M) and guluronic acids (G). The M/G ratio was estimated by CD (M/G = 0.59) indicating that CBSA was composed of 37% mannuronic acid and 63% guluronic acid. The analysis of viscosity of CBSA showed evidence of pseudoplastic fluid behaviour. The emulsifying capacity of CBSA was evaluated at different concentrations (0.25-3%), temperatures (25-100 °C) and pH (3.0-11.0). Compared to most commercial emulsifiers, the emulsion formulated by CBSA was found to be less sensitive to temperature changes and more stable at acidic pH. CBSA was examined for antioxidant properties using various antioxidant assays. CBSA exhibited important DPPH radical-scavenging activity (74% inhibition at a concentration of 0.5 mg/ml) and considerable ferric reducing potential. Effective hydroxyl-radical scavenging activity (82% at a concentration of 5 mg/ml) and potent protection activity against DNA breakage were also recorded for CBSA. However, in the linoleate-β-carotene system, CBSA exerted moderate antioxidant activity (60% at a concentration of 1.5 mg/ml). Therefore, CBSA can be used as a natural ingredient in food industry or in the pharmaceutical field.

Evolution of the fatty fraction during co-composting of olive oil industry wastes with animal manure: Maturity assessment of the end product

Olive mill wastewater sludge, resulting from the natural evaporation of olive oil processing effluent, was co-composted with poultry manure and changes in the lipid fraction investigated. Composting was achieved after approximately 9 months, leading to a compost with high stability and maturity (C/N ratio: 11.9; cation exchange capacity (CEC): 85.9 meq 100 g(-1) organic matter, CEC/total organic carbon: 4.2 meq g(-1); humic acids carbon/fulvic acids carbon: 2.2) useable directly in agriculture and having the same fertilizing capacity as farmyard manure. Composting led to a reduction in the lipid fraction by at least 95%. Unsaturated fatty acids, particularly polyunsaturated acids, were the most degraded (reduction of 55%) leading to an increase in saturated fatty acids. This change was confirmed by the relative increase in the peroxide index from 5 to 32.5 meq O(2)kg(-1) fats, and a decrease in the C(18:2)/C(16:0) ratio from 0.9 to 0.3. In addition, this study demonstrated that 1.2% of the humic acids component of the compost comprised fatty acids.

Purification and biochemical characterization of a halotolerant Staphylococcus sp. extracellular lipase

We have isolated a lipolytic halotolerant bacterium, designated as CJ3, that was identified as a Staphylococcus sp. Culture conditions were optimized and the highest extracellular lipase production amounting to 5 U/ml was achieved after 24 h of cultivation. The extracellular lipase was purified 24-fold by ammonium sulfate precipitation and a Sephacryl S-200 chromatography, and its molecular mass was found to be around 38 kDa, as revealed by SDS-PAGE and gel filtration. The lipase substrate specificity was investigated using short (tributyrin) and long (olive oil) chain triglyceride substrates. The lipase was inhibited by submicellar concentrations of Triton X-100, and maximum specific activities were found to be 802 U/mg on tributyrin and 260 U/mg on olive oil at pH 8.0 and 45 °C. The lipase was fairly stable in the pH range from 6.0 to 9.0, and about 69% of its activity was retained after incubation at 45 °C for 60 min. The enzyme showed a high tolerance to a wide range of salt concentration and a good stability in organic solvents, especially in long-chain alcohols.

Sporobacterium olearium gen. nov., sp. nov., a new methanethiol-producing bacterium that degrades aromatic compounds, isolated from an olive mill wastewater treatment digester.

A strictly chemo-organotrophic, anaerobic bacterium was isolated from an olive mill wastewater treatment digester on syringate and designated strain SR1T. The cells were slightly curved rods, stained Gram-positive and possessed terminal spores. Strain SR1T utilized crotonate, methanol and a wide range of aromatic compounds including 3,4,5-trimethoxybenzoate (TMB), 3,4,5-trimethoxycinnamate (TMC), syringate, 3,4,5-trimethoxyphenylacetate (TMPA), 3,4,5-trimethoxyphenylpropionate (TMPP), ferulate, sinapate, vanillate, 3,4-dimethoxybenzoate, 2,3-dimethoxybenzoate, gallate, 2,4,6-trihydroxybenzoate (THB), pyrogallol, phloroglucinol and quercetin as carbon and energy sources. Acetate and butyrate were produced from aromatic compounds, methanol and crotonate whereas methanethiol (MT) was produced from methoxylated aromatic compounds and methanol. Strain SR1T had a G + C content of 38 mol% and grew optimally between 37 and 40 degrees C at pH 7.2 on a crotonate-containing medium. Phylogenetically, strain SR1T was a member of cluster XIVa of the Clostridiales group and shared a sequence similarity of 90% with Clostridum aminovalericum and Eubacterium fissicatena. Consequently, its precise neighbourliness to any one of them depended on the selection of strains of the cluster. On the basis of the phylogenetic and phenotypic evidence presented in this paper, the designation of strain SR1T as Sporobacterium olearium gen. nov., sp. nov. is proposed. The type strain is SR1T (= DSM 12504T).

Application of response surface methodology to optimize decolourization of dyes by the laccase-mediator system.

Response surface methodology (RSM) was applied to optimize the decolourization of 3 dyes belonging to 3 dye families such as reactive black 5 (diazoic), indigo carmine (indigoid) and aniline blue (anthraquinonic). Crude laccase from Trametes trogii and the laccase-mediator 1-hydroxybenzotriazole (HBT) were used in this study. Box-Behnken design using RSM with six variables namely pH, temperature, enzyme concentration, HBT concentration, dye concentration and incubation time was used in this study to optimize significant correlation between the effects of these variables on the decolourization of reactive black 5 (RB5), indigo carmine (IC) and aniline blue (AB). The optimum of pH, temperature, laccase, HBT, RB5 and reaction time were 4.5, 0.5 U ml(-1), 0.5 mM, 100 mg ml(-1) and 150 min respectively, for a maximum decolourization of RB5 (about 92.92% ± 7.21). Whereas, the optimum decolourization conditions of both IC (99.76% ± 7.75) and AB (98.44% ± 10) were: pH 4.5, temperature of 45 °C, enzyme concentration of 0.1 U ml(-1) and 0.5 U ml(-1), HBT concentration of 0.9 mM and 0.5 mM, dye concentration of 60 mg l(-1) and reaction time of 150 and 90 min, respectively. The experimental values were in good agreement with the predicted ones and the models were highly significant, the correlation coefficient (R(2)) being 0.864, 0.663 and 0.776 for RB5, IC and AB, respectively. In addition, when the kinetic parameters for the three dyes decolourization were calculated according to Hannes-Wolf plot, the following values were obtained: Km of 268.4, 47.94 and 44.64 mg l(-1) then V(max) of 35.58, 10.43 and 9.23 mg l(-1) min for the RB5, IC and AB decolourizations by laccase, respectively.

Effect of Natural Mediators on the Stability of Trametes trogii Laccase during the Decolourization of Textile Wastewaters

The purpose of the present study was to determine the effect of natural mediators on the stability of the Trametes trogii crude laccase in the process of decolourization of textile effluents. Acetosyringone allowed the highest wastewaters decolourization rate of 25%. At higher concentrations of acetosyringone, the relative activity of laccase decreased approximately by between 38% and 88% after 5 days of incubation. T. trogii laccase was strongly inactivated at 3 mM syringaldehyde, after 3 days of incubation. However, laccase activity is more stable in the presence of the vanillin and m-coumarate. The T. trogii growth on solid effluent-based-medium was examined and evaluated by measuring the colony diameter in cm. T. trogii was completely inhibited on 100:0 and 80:20 effluent:water solid medium, however, colony diameter reached 5 cm on 60:40 effluent:water solid medium after 13-14 days incubation. When the textile effluent was pre-treated with laccase and laccase-acetosyringone system, the colony diameter of 2 cm of T. trogii on 80:20 effluent:water solid medium was reached after 14 and 10 days of incubation respectively.

Clostridium peptidivorans sp. nov., a peptide-fermenting bacterium from an olive mill wastewater treatment digester

A new peptide-degrading, strictly anaerobic bacterium, designated strain TMC4T, was isolated from an olive mill wastewater treatment digester. Cells of strain TMC4T were motile, rod-shaped (5-10 x 0.6-1.2 microm), stained Gram-positive and formed terminal to subterminal spores that distended the cells. Optimal growth occurred at 37 degrees C and pH 7 in an anaerobic basal medium containing 0.5% Casamino acids. Arginine, lysine, cysteine, methionine, histidine, serine, isoleucine, yeast extract, peptone, Biotrypcase, gelatin and crotonate also supported growth, but not carbohydrates, organic acids or alcohols. The end-products of degradation were: acetate and butyrate from lysine and crotonate; acetate, butyrate, H2 and CO2 from Biotrypcase, gelatin and peptone; acetate, alanine, H2 and CO2 from cysteine; acetate, H2 and CO2 from serine, cysteine and yeast extract; acetate and formate from histidine; propionate from methionine; methyl 2-butyrate, H2 and CO2 from isoleucine; acetate and ethanol from arginine; and acetate, propionate, butyrate, methyl 2-butyrate, H2 and CO2 from Casamino acids. The DNA G+C content of strain TMC4T was 31 mol%. Phylogeny based on 16S rRNA sequence analysis showed that strain TMC4T was a member of the low-G+C-content Gram-positive genus Clostridium, with the closest relative being Clostridium pascui (sequence similarity of 96 %). Due to considerable differences in genomic and phenotypic properties between strain TMC4T and those of its nearest relative, strain TMC4T is proposed as a new species of the genus Clostridium, Clostridium peptidivorans sp. nov. Strain TMC4T has been deposited in the DSMZ as strain DSM 12505T.