Removal of micropollutants by fungal laccases in model solution and municipal wastewater: evaluation of estrogenic activity and ecotoxicity

Abstract

a b s t r a c tThis study describes a multidisciplinary approach that investigates the breakdown potential of a laccase mediated system from Trametes pubescens MUT 2400 against several micropollutants including already recognized endocrine disrupting chemicals at their natural residual concentrations (from mg/L up to ng/ L).In model solution, the chemical speciation focused on a mixture of 18 analytes and adopted stir bar sorptive extraction with directed in-situ derivatization followed by gas chromatography and mass spectroscopy analysis. The method's key performance parameters were evaluated in consideration of the chemical peculiarities and complexity of real wastewaters: precision, accuracy, estimated working range extended to a wide residual concentration interval (10 ng/L to100 mg/L) indicated its fitness for purpose. Laccases were extremely active towards all the target compounds, both in term of removal yields and rate. The maximal percentage of removal was obtained for 4-t-butylphenol, 2-hydroxybiphenyl, 4-noctylphenol, salicylic acid and estrone (percentage of removal above 90%). Enzymes concentration played a central role and in most of the cases, the catalyzed reactions were very fast: the initial concentration of 9 compounds was halved within the first 3 h.The laccase-mediated treatment was then applied to a municipal wastewater collected in a real wastewater treatment plant, containing at least 9 xenobiotics as drugs, pesticides, plasticizers and personal care products. Although the harsh chemical and biological conditions of the effluent influenced enzyme stability, the reaction took place, and above 70% transformation was obtained for most analytes during the 24 h experiment. Bioassays were carried out to estimate the estrogenic activity (the E-screen test and the MELN gene-reporter luciferase assay) and the ecotoxicity (Lepidium sativum, Pseudokirchneriella subcapitata and Vibrio fischeri), demonstrating the capability of laccases to mediate an effective detoxification of the wastewater and a decrease of the estrogenic activity. treatment plant; U/L, enzymatic activity expressed as International Unit per liter; PR, percentage of removal; MEC, minimal effective concentration; t 1/2 , analyte half-life; Ti, Target Ions; RSD%, relative standard deviation percentage; LOQ, limit of quantification; QCx, quality control samples; Rel.Err %, relative error percentage; COD, chemical oxygen demand; GI%, germination index; I%, inhibition percentage of the algal growth or of the bacterial bioluminescence; EEQ, 17-b-estradiol equivalent quantity; RPE %, relative proliferative effect; TRANS%, rate of luciferase gene expression.

Fungal Bioremediation of Emerging Micropollutants in Municipal Wastewaters

“…Most of the activity (54%) was lost after 24h-treatment of municipal wastewater (Spina et al 2015) and the same effect was observed by Tran and collaborators (2013) who reported that 30% of laccase activity was lost in only 4 hours.…”

“…Oxybenzone removal required 2 hours (Garcia et al 2011), whereas 20,000 U/l completely removed estrogens in a wastewater sample within a 1 hours (Auriol et al 2008). The half-life was lower than 4 hours for 10 up to 18 micropollutants, and most of these occurred in the first minutes of the treatment (Spina et al 2015).…”

“…With the exception of phthalate and pyrene, the micropollutants were concurrently almost completely (90%) removed (Sei et al 2008). Eighteen EDCs, pharmaceuticals and PCPs with known or suspected estrogenic activity were treated by laccases from Trametes pubescens: high removal efficiency (> 60%) was achieved for 13 compounds (Spina et al 2015).…”

“…With the only exception of 2,4-dichlorophenol and BPA, 7 emerging contaminants including biological active compounds as pharmaceutical, pesticides and herbicides, were converted at high extent (above 72%) by laccases from T. pubescens (Spina et al 2015). The toxicological analysis provides the final evidence about the enhanced water quality after the treatment.…”

“…Laccases of T. pubescens effectively treated a municipal wastewater containing 9 micropollutants at a concentration of 0.13 to 67.6 µg/l (Spina et al 2015).…”