Fate of carbamazepine and anthracene in soils watered with UV-LED treated wastewaters.

Water disinfection technologies based on ultraviolet (UV) radiations emitted by Light-Emitting Diodes (LED), as a wastewater tertiary treatment, have been shown to be promising for water reuse. Here, we assessed the fate of two ubiquitous pollutants, carbamazepine and anthracene, in soil watered with either UV-LED treated wastewaters or irrigation water. After 3 months, anthracene and carbamazepine were transformed two and three times faster respectively, in soils watered with UV-LED wastewater than in soils watered with tap water (probably because of the addition of organic matter by the effluent). Laccase activity was induced in the presence of the pollutants and anthraquinone was found as anthracene product of oxidation by laccases. Moreover, the addition of these pollutants into soil did not affect the functional diversity of autochthonous microbial communities assessed by Ecolog plates. Cellulase, protease and urease activities increased in soils watered with UV-LED treated wastewaters (UV-LED WW), showing transformation of organic matter from the effluent and lipase activity increased by anthracene addition, confirming the potential role of these enzymes as indicators of hydrocarbon contamination.

Impact of watering with UV-LED-treated wastewater on microbial and physico-chemical parameters of soil.

Advanced oxidation processes based on UV radiations have been shown to be a promising wastewater disinfection technology. The UV-LED system involves innovative materials and could be an advantageous alternative to mercury-vapor lamps. The use of the UV-LED system results in good water quality meeting the legislative requirements relating to wastewater reuse for irrigation. The aim of this study was to investigate the impact of watering with UV-LED treated wastewaters (UV-LED WW) on soil parameters. Solid-state ¹³C NMR shows that watering with UV-LED WW do not change the chemical composition of soil organic matter compared to soil watered with potable water. Regarding microbiological parameters, laccase, cellulase, protease and urease activities increase in soils watered with UV-LED WW which means that organic matter brought by the effluent is actively degraded by soil microorganisms. The functional diversity of soil microorganisms is not affected by watering with UV-LED WW when it is altered by 4 and 8 months of watering with wastewater (WW). After 12 months, functional diversity is similar regardless of the water used for watering. The persistence of faecal indicator bacteria (coliform and enterococci) was also determined and watering with UV-LED WW does not increase their number nor their diversity unlike soils irrigated with activated sludge wastewater. The study of watering-soil microcosms with UV-LED WW indicates that this system seems to be a promising alternative to the UV-lamp-treated wastewaters.

Effect of coupled UV-A and UV-C LEDs on both microbiological and chemical pollution of urban wastewaters.

Wastewater reuse for irrigation is an interesting alternative for many Mediterranean countries suffering from water shortages. The development of new technologies for water recycling is a priority for these countries. In this study we test the efficiency of UV-LEDs (Ultraviolet-Light-Emitting Diodes) emitting UV-A or UV-C radiations, used alone or coupled, on bacterial and chemical indicators. We monitored the survival of fecal bioindicators found in urban wastewaters and the oxidation of creatinine and phenol which represent either conventional organic matter or the aromatic part of pollution respectively. It appears that coupling UV-A/UV-C i) achieves microbial reduction in wastewater more efficiently than when a UV-LED is used alone, and ii) oxidizes up to 37% of creatinine and phenol, a result comparable to that commonly obtained with photoreactants such as TiO(2).

Alkylphenol oxidation with a laccase from a white-rot fungus: effects of culture induction and of ABTS used as a mediator.

We investigated the potential of the laccase from the white-rot fungus Marasmius quercophilus to transform certain alkylphenols (p-nonylphenol, p-octylphenol and p-t-octylphenol). We tested the reactivity of this enzyme under different conditions: in liquid cultures and using the partially purified laccase with and without 2,2'-azino-bis-3-ehtylbenzothiazoline-6-sulfonicacid (ABTS) as a mediator. The percentage of p-t-octylphenol disappearance in liquid cultures was 69.0 ± 1.5% and 81 ± 5% after a 8-d or 15-d incubation, respectively, with p-nonylphenol, these percentages were 62 ± 4% and 91 ± 6% and with p-octylphenol 37 ± 3% and 65 ± 1% after a 15-d and a 21-d incubations, respectively. Induced pre-cultures were also used to inoculate the liquid cultures to enhance p-octylphenol transformation: the percentages of disappearance were 91.0 ± 0.5% and 97 ± 1% after a 8-d and a 15-d incubation, respectively. Mass spectrometry analysis showed that the products of oxidation of p-octylphenol were dimers with a mass of 411 m/z. Furthermore, we identified a purple compound (m/z 476) formed when ABTS was added to the reaction medium with the purified laccase. This result confirms that, in complex environments such as soils or litters where many molecules can interact with the enzyme substrate or the product of oxidation, laccase activities and those of other phenoloxidases should not be measured with ABTS.

Solid-state 13C NMR to assess organic matter transformation in a subsurface wetland under cheese-dairy farm effluents.

Solid-state (13)C NMR were used to follow organic matter transformation in a subsurface wetland under the effluent of a small cheese-dairy farm under a Mediterranean climate. The results showed that the ratios commonly used to quantify humification, (aromaticity and Alkyl-C/O-Alkyl-C ratios) can be considered as relevant chemical indicators of organic matter transformation. Polysaccharides were transformed throughout the subsurface wetland whereas aromatic, phenolic and alkyl compounds accumulated. Furthermore, Phenolic-C signal and O-Alkyl-C signal were negatively correlated to proteases and beta-galactosidase activities showing that recalcitrant molecules actually accumulated. These results were correlated with high purification yields: the average decrease in chemical demand in oxygen was 90.75% and that in Total Kjeldahl Nitrogen was 75.65%. Thus subsurface wetlands can be considered as an efficient technology to purify effluents with high organic matter contents, such as cheese-dairy effluent, under drastic climate conditions. Furthermore this study highlights the fact that solid-state (13)C NMR is a suitable tool to follow organic matter transformation.

A method to quantify transesterification activities of lipases in litters.

Lipases are glycerol ester hydrolases (EC 3.1.1.3) produced by a wide range of microorganisms. They catalyse the hydrolysis of different esters but this reaction is reversible, depending on the water content of the reaction medium, via esterification and transesterification. The synthetic activity of lipases can be of major importance in natural ecosystems since it can be involved in carbon stockage in soils or litters. Here, the detection of transesterification activities of lipases in litter is reported for the first time. We used two different litters: litter of Quercus pubescens (QP) and litter of both Q. pubescens and Q. ilex. Different p-nitrophenyl esters and pentanol were used to test transesterification in a reaction medium with an organic solvent (heptane). We showed that these activities were proportional to the amount of litter, the incubation time and the substrate concentration and that they increased with temperature. Furthermore, the lipases from the litters studied were very thermostable since they were still active after 2 h at 70 degrees C. These activities showed common properties of lipases: the highest activities were obtained with a medium acyl-chain substrate p-nitrophenyl caprylate and transesterification activities were correlated to water activity, a(w). The following parameters are recommended to quantify transesterification activities in litter: 10 mM of p-nitrophenyl caprylate, 1 g of litter, 500 microL of pentanol, q.s.p. 4 mL of heptane incubated at 30 degrees C for 2 h.

Soil microbial activities in a constructed soil reed-bed under cheese-dairy farm effluents.

Soil microbial activities in a reed-bed used for effluent purification of a small cheese-dairy farm under a Mediterranean climate were described and studied. This work aims to demonstrate (i) whether certain enzyme activities used as bioindicators of dairy waste degradation (beta-galactosidase and protease) vary over time, which might influence organic matter degradation and (ii) whether specific microbial communities are selected through contact with the discarded effluent using community level catabolic profiles (CLCPs). beta-galactosidase and protease activities were followed in a 14-month monitoring experiment. These enzyme activities were strongly expressed during the whey-discarding period from February to May. CLCPs using Biolog Ecoplate showed great microbial diversity, as described by Shannon-Weaver index, and no difference was observed in microbial diversity between areas at the receiving end of the reed-bed (where effluent was discarded) and those at the opposite end. This may be explained by successive environmental factors which made enzyme activities vary: whey discarded from February to May and Mediterranean climate conditions (drying-rewetting effects on summer). Microbial enumeration using epifluorescence microscopy also showed a pattern linked to Mediterranean conditions with a drastic decrease in biomass during summer drought. These results on functional biodiversity were correlated with high purification yields: the minimum decrease in Biological Demand in Oxygen was 84% and that in suspended solids was 75%.

Effects of pollutants on laccase activities of Marasmius quercophilus, a white-rot fungus isolated from a Mediterranean schlerophyllous litter.

Marasmius quercophilus is a white-rot fungus involved in carbon recycling in Mediterranean ecosystems because of its laccase production. Here we described the effect of metal ions and halide salts, on laccase activity in order to point out the action of such environmental pollutants on this enzyme of major importance. Furthermore we tested organic solvent effects on laccase reaction since reaction mixture including solvent can be used in the transformation of xenobiotics. In the case of metal ions, we found that chloride ions were responsible for inhibition while CuSO(4) and MnSO(4) enhanced laccase activity. When halides were tested, we showed the following degree of inhibition: F(-)>Cl(-)>Br(-). Furthermore we found that I(-) was oxidized by laccase with I(2) as the product of the reaction. With ABTS, 50% of the laccase activity remains for solvent concentration ranging from 40% to 60% depending on the solvent used while with syringaldazine solvent concentration ranged from 50% to 70%. The organic solvent effects observed were probably a result of enzyme denaturation and of both enhancement of oxidised product solubilisation and of substrate solubilisation (for syringaldazine). These results show that laccase from M. quercophilus is not rapidly inhibited by certain environmental pollutants which sustains its role in carbon turnover under pertubation. However the strong effect of chloride ion on laccase activity should be further investigated with in situ studies since this could drastically influence carbon recycling in litters from Mediterranean littoral locations.

Quercus suber cork extract displays a tensor and smoothing effect on human skin: an in vivo study.

Recently, it has become indispensable for anti-aging active ingredients to provide a visible and immediate smoothing antiwrinkle effect. In Quercus suber, suberin is the most important structural component of cork cell walls. Studies have shown that suberin is made up mostly of hydroxycarboxylic acids and that it is endowed with many special mechanical and chemical properties that evoke a possible smoothing effect on the surface of the skin. Therefore, we were interested in investigating the effect of this cork extract on the skin's surface in a double-blind clinical study. The study was conducted in 15 healthy volunteers, aged 22 to 52 years. The volunteers applied a gel formula with 3% of cork extract, or placebo gel, on each forearm. Skin surface roughness was evaluated visually by pictures and by silicone replicas 1 and 2 h after application, followed by statistical analysis using the matched-pairs McNemar statistical test. McNemar analysis of the pictures revealed that application of cork extract on the skin resulted in a highly significant reduction of roughness 1 h after application. This effect was observed in 73.3% of volunteers. Two hours after cork extract application, a highly significant improvement of skin roughness was found in 78.6% of volunteers. Moreover, silicone replica treatment confirmed significant improvement in average of roughness at 2 h. These results demonstrate that cork extract provides a remarkable and highly significant tensor and smoothing effect on the skin, which could be of great use in anti-aging skin care products.

Variations of lignocellulosic activities in dual cultures of Pleurotus ostreatus and Trichoderma longibrachiatum on unsterilized wheat straw.

Trichoderma spp., soil filamentous fungi, are antagonists that can cause great losses in mushroom production. We have investigated the influence of T. longibrachiatum on the production of lignocellulolytic enzymes by Pleurotus ostreatus during its vegetative growth on a straw-based cultivation substrate that either had been sterilized, pasteurized or not heat treated. The variations in the lignocellulolytic activities and the electrophoretic patterns in single and dual cultures were used as a tool for perturbation assessment. The various heat treatments of the wheat straw before inoculation affected both the bacterial populations and the abilities of T. longibrachiatum and P. ostreatus to colonize the substrate and to produce extracellar lignocellulolytic enzymes. Interactions between T. longibrachiatum and the microflora of the substrate led to a great decrease of hydrolytic activities due to reduced colonization of the substrate. Pleurotus ostreatus also was affected but it was less sensitive than T. longibrachiatum. As a consequence, in dual cultures with P. ostreatus, the competitive ability of T. longibrachiatum was reduced by bacteria in the substrates. The presence of total microflora or thermotolerant microflora increased the production of phenoloxidase activities by P. ostreatus, despite reduced colonization of the substrate. This contributed to the improvement of the competitive ability of P. ostreatus in the pasteurized substrate. Furthermore, a direct effect of bacteria on T. longibrachiatum also was observed. In sterilized substrate, both laccase and Mn-peroxydase activities were increased dramatically in dual cultures due to a faster production of a laccase isoform, which was stimulated by T. longibrachiatum.

Purification of a new isoform of laccase from a Marasmius quercophilus strain isolated from a cork oak litter (Quercus suber L).

A new isoform of laccase from Marasmius quercophilus is described in this study. The strain of this white-rot fungus was isolated for the first time on a cork oak litter. This isoform exhibited certain common properties of laccases (a molecular weight of 65 Kda, an optimum pH of 6.2 with syringaldazine). But this laccase has also particularly novel features: the best activity measured was observed at high temperatures (80 C) and this isoform was not inhibited with EDTA. Furthermore, this induced laccase was able to transform most of the aromatic compounds tested without the addition of mediators to the reaction mixture, and the transformation of certain chlorophenols (2-chlorophenol and 2,4-dichlorophenol) by a laccase isoform from M. quercophilus is reported here for the first time. We also demonstrate the importance of 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonate) (ABTS) as a mediator since it allowed veratryl alcohol and p-hydroxybenzoic acid transformation. Moreover, new products of transformation were observed using the combination of ABTS with this isoform of laccase.

Purification, partial characterization, and reactivity with aromatic compounds of two laccases from Marasmius quercophilus strain 17.

Two isozymes of laccase were obtained from an induced liquid culture of Marasmius quercophilus with p-hydroxybenzoic acid as the inducer. Both the constitutive and the induced isozyme have a molecular mass of 60 kDa as determined by polyacrylamide gel electrophoresis. Using isoelectric focusing, we found three isozymes with the constitutive enzyme (pI 4, 4.2, 4.4) and four of the induced form (pI 4.75, 4.85, 4.95, 5.1). We observed certain differences between these two isozymes; the specific activity of the induced isozyme was twice as high, and two optimum pH levels (5 and 6) were observed with the induced isozyme (only one, pH 5, for the constitutive isozyme). However, both of these enzymes have the same thermal stability and the same temperature for their highest activity (80 degrees C). Furthermore, the reactivity of both these enzymes with aromatic compounds was similar. The use of mediators extended the oxidized substrate range of the laccases studied. Various products of degradation were observed, depending on the mediator used. When laccase was used alone, the decrease of the signal corresponding to the aromatic cycle, without any formations of other peaks at different wavelengths, suggested polymerisation of aromatic compounds.