Search

In the present study, a novel and efficient technology based on the use of an oxidative enzyme was developed to perform the continuous removal of estrogenic compounds from polluted wastewaters. A 2 L enzymatic membrane reactor (EMR) was successfully operated for 100 h with minimal requirements of laccase for the transformation of estrone (E1), 17β-estradiol (E2), and 17α-ethinylestradiol (EE2)from both buffer solution and real wastewater (filtered secondary effluent). When the experiments were performed at high and low concentrations of the target compounds, 4 mg/L and 100 μg/L, not only high removal yields (80-100%) but also outstanding reduction of estrogenicity (about 84-95%) were attained. When the EMR was applied for the treatment of municipal wastewaters with real environmental concentrations of the different compounds (0.29-1.52 ng/L), excellent results were also achieved indicating the high efficiency and potential of the enzymatic reactor system. A second goal of this study relied on the identification of the transformation products to elucidate the catalytic mechanism of estrogens' transformation by laccase. The formation of dimers and trimers of E1, E2, and EE2, as well as the decomposition of E2 into E1 by laccase-catalyzed treatment, has been demonstrated by liquid chromatography atmospheric pressure chemical ionization (LC-APCI) analysis and confirmed by determination of accurate masses through liquid chromatography electrospray time-of-flight mass spectrometry (LC-ESI-TOF). Dimeric products of E2 and EE2 were found even when operating at environmental concentrations. Moreover, the reaction pathways of laccase-catalyzed transformation of E2 were proposed.

A methodology that programs eukaryotic or bacterial cells to encapsulate proteins of any kind inside micro/nanospheres formed by muNS-Mi viral protein was developed in our laboratory. In the present study such "in cellulo" encapsulation technology is utilized for immobilizing a protein with an enzymatic activity of industrial interest, CotA laccase. The encapsulation facilitates its purification, resulting in a cost-effective, one-step way of producing immobilized enzymes for industrial use. In addition to the ability to be recycled without activity loss, the encapsulated protein showed an increased pH working range and high resistance to chemical inactivation. Also, its activity was almost unaffected after 30 min incubation at 90 °C and 15 min at the almost-boiling temperature of 95 °C. Furthermore, the encapsulated laccase was able to efficiently decolorate the recalcitrant dye RB19 at room temperature. [ABSTRACT FROM AUTHOR]

BACKGROUND: Organosolv pretreatment produces a hemicellulose‐rich stream with limited applications due to its toxicity. In this study, beech wood organosolv liquor was valorized through the production of ligninolytic enzymes, the extraction of ganoderic acid and the detoxification of this process stream. RESULTS: Laccase and manganese peroxidase (MnP) were produced in 30 L bioreactors by the white‐rot fungi Ganoderma lucidum and Irpex lacteus, respectively, using the hemicellulose‐rich liquor as carbon source. Both fungi caused a significant reduction in the hemicellulose stream toxicity (between 6.1 and 8.3 times lower). The extraction of ganoderic acid (24.3 mg g−1 of G. lucidum biomass) as a fungal metabolite with high pharmacological activity greatly increased the profitability of the process. Finally, the economic viability of the proposed valorization strategy was analyzed on the basis of the operational costs estimated for the process scale‐up. The specific costs of MnP and laccase production were €2.27 kU−1and €1.08 kU−1, respectively, which are 1109 and 78 times lower than the average price of commercially available enzymes. CONCLUSION: It was found that the integration of the proposed valorization strategy into an organosolv‐based biorefinery would help to increase the economic feasibility of this pretreatment technology. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A magnetic separation unit of toroidal magnets in alternate polarity was designed. • Laccase immobilized on magnetic nanoparticles was successfully recovered. • Methyl green and rutin were successfully transformed in the magnetic enzymatic reactor. • The scalability and economic feasibility of the process were investigated. Abstract In this research, a new sequential batch reactor (SBR) coupled to an internal magnetic separator was developed. The separator consists of a set of axially magnetized permanent toroidal magnets, distributed along a non-magnetic steel rod, uniformly spaced with alternate polarity, which provide an external magnetic field of up to 1.2 T. The feasibility of magnetic separation was assessed for the retention of the nanobiocatalyst based on laccase immobilized on silica-coated MNPs. The proof of concept was evaluated for the enzymatic decolorization of the methyl green (MG) dye and the oligomerization of rutin, with virtually complete recovery of the nanobiocatalyst (99%) and high biotransformation efficiency of both compunds. The characterization of the reaction products of MG by laccase was conducted by high performance liquid chromatography (HPLC) with a diode array detector (DAD) coupled to mass spectrometry (MS) using the electrospray ionization source (ESI) (HPLC-DAD-ESI-MS). Moreover, the biotransformation products showed less microtoxicity than the parent compound and increased biodegradability. Beyond laboratory scale experiments, the reactor proposed here was scaled-up to a volume of 100 L and the environmental performance and cost analysis were estimated. [ABSTRACT FROM AUTHOR]

Enzymatic treatments based on oxidative enzymes, such as peroxidases, laccases and tyrosinases, have been proposed as an alternative to conventional methods to remove a broad range of contaminants present in wastewater. The aim of this study is to discuss existing technologies for the removal of pollutants based on the use of oxidative enzymes, including a discussion on the most important factors affecting the efficiency of the proposed systems. Factors involved in the catalytic cycle of the enzyme (biocatalyst, substrates and mediators), the addition of certain components to the reaction medium (additives, surfactants or solvents) as well as operational parameters (temperature, pH or agitation) will be discussed. Finally, two types of reactors: one-stage and two-stage enzymatic membrane reactors, especially designed for the treatment of micropollutants present in secondary effluents, will be described in detail. [ABSTRACT FROM AUTHOR]

Oxidative biocatalysis by laccase arises as a promising alternative in the development of advanced oxidation processes for the removal of xenobiotics. The aim of this work is to develop various types of nanobiocatalysts based on laccase immobilized on different superparamagnetic and non-magnetic nanoparticles to improve the stability of the biocatalysts. Several techniques of enzyme immobilization were evaluated based on ionic exchange and covalent bonding. The highest yields of laccase immobilization were achieved for the covalent laccase nanoconjugates of silica-coated magnetic nanoparticles (2.66 U mg-1 NPs), formed by the covalent attachment of the enzyme between the aldehyde groups of the glutaraldehyde-functionalized nanoparticle and the amino groups of the enzyme. Moreover, its application in the biotransformation of phenol as a model recalcitrant compound was tested at different pH and successfully achieved at pH 6 for 24 h. A sequential batch operation was carried out, with complete recovery of the nanobiocatalyst and minimal deactivation of the enzyme after four cycles of phenol oxidation. The major drawback associated with the use of the nanoparticles relies on the energy consumption required for their production and the use of chemicals, that account for a major contribution in the normalized index of 5.28 x 10-3. The reduction of cyclohexane (used in the synthesis of silica-coated magnetic nanoparticles) led to a significant lower index (3.62 x 10-3); however, the immobilization was negatively affected, which discouraged this alternative. [ABSTRACT FROM AUTHOR]

A three-level Box-Behnken factorial design combined with response surface methodology (RSM) was applied as a tool to study the laccase-catalyzed removal of three estrogenic compounds: estrone (E1), estradiol (E2), and ethinylestradiol (EE2), in a continuous enzymatic membrane reactor (EMR). Three main factors affecting the treatment efficiency were considered: enzyme activity, hydraulic residence time (HRT) and oxygenation rate. As expected, laccase activity and HRT showed large effects and, interestingly, the relevance of oxygen in improving the oxidation kinetics through raising the dissolved oxygen above saturation levels was demonstrated. When considering elimination rates as the response, optimal conditions were: 1,000 U/L of laccase, 1 h HRT and 60 mgO2/(L·h) of oxygenation rate, predicting 2.82-3.24 mg eliminated/(L·h), (71-81% of oxidation). These optimum conditions were successfully validated, and 75% of estrogenicity reduction was achieved. On the other hand, only 100 U/L were found as optimal to maximize the efficacy of the enzyme: E1 was oxidized by 0.06 mg/(L·h·U), although the removal of estrogenicity decreased to 60%. The methodology was also applied to maximize the reduction of estrogenic activity: the highest values assayed [1,000 U/L, HRT 4 h and 60 mgO2/(L·h)] provided 99% detoxification. [ABSTRACT FROM AUTHOR]

Fractionation of lignocellulosic biomass with solvents (organosolv process) generates a hemicellulose-rich liquor with a high content of phenolics which is particularly toxic. This work addresses the utilization of this stream as a potential carbon source for the production of ligninolytic enzymes (LE). Among six basidiomycetes species, Irpex lacteus and Ganoderma lucidum presented the highest activities of manganese peroxidase (646 ± 122 U L −1 ) and laccase (1497 ± 161 U L −1 ), respectively, growing on a medium composed mainly of cellulose fibers, lignin and hemicellulose. The influence of each lignocellulosic fraction on the LE production mechanisms was studied in more detail. The high concentration of phenolic compounds in the hemicellulose-rich stream acted as inducer of LE production, with levels even greater than those of xylose. Acute toxicity tests on Vibrio fischeri revealed a substantial reduction of the toxicity after the fungal treatment (by ca. 12–21 times). The proposed valorization and detoxification of this currently non-exploited and abundant by-product stream is a promising strategy to enhance the industrial feasibility of the organosolv fractionation process. [ABSTRACT FROM AUTHOR]