ARTICLE IN PRESS Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
Contents lists available at ScienceDirect
Pesticide Biochemistry and Physiology j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / p e s t
Biodegradation of pesticide triclosan by A. versicolor in simulated wastewater and semi-synthetic media Burcu Ertit Taştan a,b,c,*, Gönül Dönmez a a b c
Department of Biology, Faculty of Science, Ankara University, 06100, Beşevler, Ankara, Turkey Life Sciences Application and Research Center, Gazi University, 06830, Gölbaşı, Ankara, Turkey Health Services Vocational School, Gazi University, 06830, Gölbaşı, Ankara, Turkey
A R T I C L E
I N F O
Article history: Received 24 June 2014 Accepted 7 November 2014 Available online Keywords: Fungus Triclosan 2,4-dichlorophenol Biodegradation Wastewater treatment
A B S T R A C T
Triclosan is known as an antimicrobial agent, a powerful bacteriostat and an important pesticide. In this paper biodegradation of triclosan by Aspergillus versicolor was investigated. Effects of simulated wastewater and semi-synthetic media on fungal triclosan degradation process were detected. HPLC analysis showed that fungal triclosan biodegradation yield was 71.91% at about 7.5 mg/L concentration in semisynthetic medium and was 37.47% in simulated wastewater. Fungus could be able to tolerate the highest triclosan concentration (15.69 mg/L). The biodegradation yield was 29.81% and qm was 2.22 mg/g at this concentration. Some of the parameters, such as pH, culture media, increasing triclosan and biomass concentrations were optimized in order to achieve the effective triclosan biodegradation process. The highest triclosan biodegradation yields of all microorganisms were achieved by A. versicolor. © 2014 Elsevier Inc. All rights reserved.
1. Introduction Triclosan, 5-chloro-2-(2,4-dichlorophenoxy)phenol or commercially known as Irgasan is an antimicrobial agent and a powerful bacteriostat. It has also been registered as a pesticide by the United States Environmental Protection Agency [1]. It blocks lipid biosynthesis in bacteria by inhibiting the enoyl reductase enzyme [2]. Thus, since its introduction in the 1960s, it has been widely used in soaps, shampoos, toothpastes, hand washers and disinfectants. Two tons of triclosan has been consumed per year only in Sweden [3]. However this fact seems to be a marketing strategy rather than a medical necessity. The safety profile of triclosan has been questioned recently after the observation that triclosan was determined not only in wastewater [4] but also in human breast milk [5]. Analyses performed by Ying and Kookana [6] showed that triclosan had a tendency to take part in the environment and it is a persistent pollutant, which does not degrade fast; its primary biodegradation half-life was given as “weeks” and ultimate biodegradation half-life was expected to take “months.” Due to its long half-life and lipophilic characteristics, it may potentially cause longterm health risks in human body. Animal studies suggest that triclosan can have a harmful effect on the reproductive system [7]. Therefore, it is necessary to develop an environmental friendly and
* Corresponding author. Life Sciences Application and Research Center, Gazi University, 06830, Gölbaşı, Ankara, Turkey. Fax: +90 312 484 62 71. E-mail address: [email protected] (B. Ertit Taştan).
fast method for triclosan biodegradation in order to reduce environmental pollution. Attention has been attracted to triclosan degradation processes in recent studies. Some of the triclosan degradation methods are sonoelectrochemical [8], photoelectrocatalytic [9], oxidative [10] and electro-fenton degradation [11]. Studies suggest that biodegradation is one of the most efficient methods for triclosan degradation. Different transformation metabolites could be detected during triclosan biodegradation and while degradation metabolites show androgenic responses, the end products do not [12]. It has been demonstrated that triclosan was highly toxic to organisms, especially to aquatic ones [13,14]. Therefore, in order to increase the triclosan removal efficiency, it is important to select the highly triclosan tolerant microorganisms. Fungi are often used and preferred in biological treatment studies due to their high pollutant removal yields even at their high concentrations as proven by several recent studies [15–17]. Hundt et al. [18] described some biotransformation reactions of triclosan by the white rot fungi Trametes versicolor and Pycnoporous cinnabarinus. T. versicolor produced 3 metabolites including 2,4-dichlorophenol. They explained that the conjugates showed a distinctly lower cytotoxic activity than triclosan. In an effort to address the biodegradation of increasing triclosan concentrations, this study aimed to examine several hypothesis: (i) whether triclosan blocks lipid biosynthesis in bacteria and whether fungal biomass might have higher resistance to its toxicity than bacteria; the fungal biomass could serve as an effective biomaterial, (ii) whether triclosan biodegradation realized not only in rich microbiological media
http://dx.doi.org/10.1016/j.pestbp.2014.11.002 0048-3575/© 2014 Elsevier Inc. All rights reserved.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
2
but also in simulated wastewater, it can be an original and useful study for real wastewater treatment systems. We also aimed to check the best triclosan biodegradation process by changing experimental parameters. 2. Materials and methods 2.1. Chemicals Triclosan 97% (Irgasan CAS: 3380-34-5) was purchased from Sigma; 2,4-Dichlorophenol 99% (CAS: 120-83-2) was supplied by Sigma-Aldrich. HPLC grade acetonitrile and ammonium acetate were purchased from Merck. The stock solutions were prepared in acetonitrile (80%)/ammonium acetate (20%) solution and were stored in a refrigerator at 4 °C. Appropriate volumes of the stock solutions were added to the media. 2.2. Culture conditions Aspergillus versicolor was isolated previously by Ertit Taştan et al. [17] from Batman, Turkey. Fungal biomass was inoculated into 250 mL Erlenmeyer flasks containing known triclosan concentrations in 100 mL culture media as described below. The flasks were incubated at 25 ± 2 °C on a rotary shaker (New Brunswick Scientific Innova 4230) at 100 rpm for 5 days. In order to obtain an equal initial biomass, 100 mL media was inoculated with 0.4 g/L dry weight biomass. 2.3. Growth medium Czapek Dox Medium (CDM) was selected as the first test media, which was a semisynthetic medium used for the cultivation of fungi and especially for Aspergillus spp. The ingredients (g/L) were NaNO2, 2; KCl, 0.5; MgSO4.7H2O, 0.5; K2HPO4, 1; FeSO4.7H2O, 0.01; Sucrose 30. Simulated Waste Water (SWW) was the second selected media. The composition of SWW was prepared according to Bracklow et al. [19] considering its resemblance to municipal wastewater and we slightly modified it for this study. The ingredients (mg/L) were as follows: peptone 25, yeast extract 80, starch 200, sunflower oil 35, ammonium acetate 150, KH2PO4 26, MgHPO4.3H2O 6, glucose 2000, K2HPO4 26, urea 50, FeSO4.7H2O 8, whey powder 160. Both of these media were autoclaved at 121 °C for 15 min before the inoculation step. Medial pH values were adjusted to appropriate volumes by adding diluted (0.01 M) and concentrated (1 M) sulfuric acid or sodium hydroxide solutions. 2.4. Analytical methods Triclosan biodegradation was determined on the samples (3 mL) taken daily during the incubation period. Uninoculated Erlenmeyer flasks containing triclosan were used as control samples to detect any reactions between media and triclosan. Growth of A. versicolor was determined by measuring the dry weight of biomass at the end of the incubation period. The concentration of triclosan in the supernatant was determined by high-performance liquid chromatography (Shimadzu, Kyoto, Japan), by using a C-18 column (250 mm × 4.6 mm inner diameter: 5 μm particle size) at 50 °C. Acetonitrile and 10 mM ammonium acetate were used as mobile phase to produce a binary elution gradient with a flow rate of 1 mL/min. Pure water was obtained from an apparatus (Neuberger, Germany), and it was used to prepare solutions and buffers, which were filtered through 0.45 μm Teknokroma membrane filters prior to use. The sample solutions were filtered through 0.20 μm Minisart RC syringe filters before the
injection into the chromatograph. The separation of triclosan was achieved with the following linear mobile phase gradient program: at 6.85 min 80:20 (v/v) at 254 nm. The percentage biodegradation of triclosan and qm (the maximum specific triclosan uptake) was calculated from the relevant equations. In the study, qm represents the maximum amount of triclosan biodegradation per unit dry weight of fungal cells (mg/g), X maximum dried cell mass (g/L), and C0 the initial concentration of the triclosan (mg/L), respectively. The data were subjected to analysis of significant differences among treatment means were compared by descriptive statistics (±S.E.).
2.5. pH experiments The effect of pH on triclosan biodegradation process was investigated at pH 5, 6, 7 and 8 at about 7.5 mg/L triclosan concentration for 5 days of incubation period in both of the growth media. Each of these experiments was performed in triplicates. There was not a significant change after 3 days of incubation period; therefore these results were taken into consideration.
2.6. Triclosan concentration experiments In order to highlight the effect of initial triclosan concentrations, the experiments were conducted at 5, 7.5, 12.5 and 15.5 mg/L triclosan concentrations. Each of these experiments was performed in triplicates.
2.7. Fungal biomass experiments The effect of fungal biomass concentrations on triclosan biodegradation process was also examined at three different biomass concentrations by using the dry weight method. The experiments were performed at 7.5 mg/L triclosan concentration at pH 7 in CDM and at pH 5 in SWW. For these experiments 100 mL media was inoculated with 0.4, 0.8 and 1.1 g/L dry weight biomass.
3. Results and discussions 3.1. Effects of pH on triclosan biodegradation in different culture media Change in pH values is an important parameter affecting the dissociation of triclosan molecule. The dissociation constant of triclosan is given as pKa = 8.14, and when pH > pKa more than 50% of the triclosan is expected to be deprotonated [20]. As seen in Fig. 1, when pH was increased from 5 to 7 in CDM triclosan biodegradation increased. The maximum biodegradation was observed at pH 7 as 55.00%. The selected optimum pH rate was used at the next steps of experiments. Previous studies showed that pollutant biodegradation yield was really linked to culture media [21]. In order to make this work applicable in daily life and to obtain a better understanding of the triclosan biodegradation process, we also examined the effect of SWW, which is close to municipal wastewater. The aim was to reveal if triclosan biodegradation process efficiency could also be obtained in SWW. Therefore, the same concentrations with CDM were tested in SWW. The results are presented in Fig. 1. The biodegradation rates decreased when pH was increased from 5 to 8. The maximum biodegradation was observed at pH 5 as 50.63%. The selected optimum pH rate was used at the next step of experiments.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
3
Fig. 2. Effects of triclosan concentrations on the biodegradation yield (Y%) of A. versicolor during the incubation period in CDM (pH: 7; T, 25 ± 2 °C; stirring rate: 100 rpm). Fig. 1. Effect of pH on triclosan biodegradation (Y%) of A. versicolor at about 7.5 mg/L triclosan concentration in CDM and SWW (incubation period, 3 days; T, 25 ± 2 °C; stirring rate: 100 rpm).
3.2. Effect of increasing triclosan concentrations on biodegradation process Triclosan concentrations are important parameter, because it was proven that microorganisms were affected by increasing triclosan concentrations [22]. Taking into consideration of daily increasing triclosan concentrations, the effects of this parameter was tested at four different concentrations (5, 7.5, 12.5 and 15 mg/L) in the second series of the experiments. It was clearly seen in Fig. 2 that when concentration was increased up to 15.5 mg/L, the biodegradation rate decreased in CDM. The highest biodegradation rate was 61.81% at 4.195 mg/L triclosan concentration. It should be clearly stated that after the 3rd day the biodegradation rate reached a constant rate. For instance, 35.68% biodegradation was obtained on the 1st day of the experiments and this rate was kept constant in the following days. The highest biodegradation rate was achieved in the last day of the experiments at the highest concentration of 15.692 mg/L as 31.27%. On the other hand, the maximum amount of triclosan biodegradation per unit dry weight of fungal cells (qm) was 0.872 mg/g at 4.195 mg/L and it was 2.225 mg/L at 15.692 mg/L (Table 1). The reason why the qm values were higher at highest triclosan concentrations than at lowest concentrations was due to the existence of higher concentrations of triclosan in culture media, which was accumulated by fungal cells. There was also a clear correlation between the biodegradation rates. Triclosan biodegradation rates in CDM and SWW were similar at low concentrations. As seen in Table 1, the biodegradation rates
Table 1 Comparison of the biodegradation yields and the maximum specific triclosan uptake (qm) values at different triclosan concentrations (C0) in CDM of A. versicolor. Co (mg/L) CDM 4.19 7.07 12.51 15.69 SWW 2.5 6.88 11.32 13.23
Y%
qm (mg/g)
61.81 ± 4.61 55.00 ± 2.02 34.96 ± 2.09 29.81 ± 1.64
0.87 ± 0.12 1.26 ± 0.26 1.96 ± 0.04 2.22 ± 0.01
of lower concentrations were more remarkable than the rates at higher ones in SWW. The maximum biodegradation rate was 64.56% at 2.50 mg/L triclosan concentration, and this rate decreased to 37.47% when concentration increased to 6.88 mg/L. When compared, a 34.96% yield was obtained in CDM at 12.51 mg/L triclosan concentration, and 18.38% yield was obtained at 11.32 mg/L in SWW. In this medium the qm values decreased when triclosan concentration was increased. This is related with the biodegradation rates which are lower than CDM. Because lower degradation yields was achieved in SWW and thus qm values was lower. In an another study Chlorella pyrenoidosa was exposed to a series of triclosan concentrations ranging from 100 to 800 ng/mL, and it was observed that more than 50% of triclosan was eliminated by algal uptake from the culture media [23]. These triclosan concentrations were lower than the ones tested at the present work. It is important to detect the most triclosan tolerant microorganism, considering its increasing concentrations in the environment. 3.3. Effect of increasing fungal biomass concentrations on biodegradation process To detect the effect of increasing fungal biomass concentrations on biodegradation process 0.4, 0.8 and 1.1 g/L fungal biomass was inoculated into media. It is evident from the results presented in Table 2 that biodegradation rates were really linked to the amount of initial biomass concentrations. The highest biodegradation rate was 55.00% at the first biomass concentration within 3 days in CDM. On the other hand fungus removed 71.91% triclosan at 1.1 g/L biomass concentration. This rate was 1.3 times higher than the rate that was obtained at 0.4 g/L biomass concentration.
Table 2 Comparison of the biodegradation yields and the maximum specific triclosan uptake (qm) values at different fungal biomass concentrations at about 7.5 mg/L triclosan concentrations in CDM and SWW. X0 (g/L)
CDM 64.56 ± 2.63 37.47 ± 1.09 18.38 ± 3.09 16.47 ± 3.60
T, 25 ± 2 °C; stirring rate, 100 rpm; incubation period, 3 days; pH 7.
0.59 ± 0.19 1.66 ± 0.36 1.50 ± 0.03 1.01 ± 0.02
SWW
Y% qm (mg/g) Y% qm (mg/g)
0.4
0.8
1.1
55.00 ± 2.02 1.26 ± 0.26 37.47 ± 1.09 1.66 ± 0.35
66.44 ± 5.62 1.49 ± 0.10 38.04 ± 4.23 1.00 ± 0.26
71.91 ± 4.60 1.99 ± 0.38 46.12 ± 2.75 0.78 ± 0.05
C0, 7.5 mg/L triclosan; X0, initial fungal biomass concentration; T, 25 ± 2 °C; stirring rate, 100 rpm; incubation period, 3 days; pH 7.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
4
As presented in Table 2, an increase in the initial biomass concentration from 0.4 g/L to 1.1 g/L resulted in a significant increase of the maximum triclosan biodegradation per unit dry weight of A. versicolor, which was 1.26, 1.49 and 1.99 mg/g, respectively. The qm value at the maximum initial biomass concentration was also found considerably higher than the minimum biomass concentration, due to higher biodegradation yields. At the maximum biomass concentration (1.1 g/ L) in CDM, qm increased nearly 1.6 times. As expected, when biomass concentrations were higher at a known triclosan concentration, the maximum triclosan quantity per unit dry weight of the fungus would be higher, due to higher biodegradation rates and the lowest level of residual triclosan concentrations in culture media. In contrast to biodegradation efficiency, qm decreased from 1.66 to 0.78 mg/g in SWW. The maximum triclosan biodegradation was recorded as 46.12% at maximum initial biomass concentration; however this biodegradation rate was not equal to the minimum biodegradation yield that was obtained in CDM. Therefore, it is concluded that increases in initial biomass concentration accompanied by decreasing qm values in SWW, contrary to the changes in CDM. In another study Sphingopyxis strain KCY1 was grown on complex nutrient medium containing 5 mg/L triclosan and qm was found as 0.13 mg-triclosan/mg-protein/day. They suggested a possible biodegradation pathway for triclosan by strain KCY1 [12]. Song et al. [10] investigated the triclosan degradation in the presence of H2O2 as oxidant and BiFeO3 magnetic nanoparticles as catalyst. HPLC measurements showed that 2,4-dichlorophenol was the major degradation intermediate of triclosan. They explained that except for the 2,4-dichlorophenol pathways, the direct dechlorination process also occurred like Ferrer et al. [24] detected in their triclosan study in wastewater samples. It was also explained that 2,4-dichlorophenol was less toxic to Sphingomonas sp. than triclosan [25]. This finding has also been supported in our present work for A. versicolor. Triclosan degradation is shown in Fig. 3 with HPLC data at different times as C0 (time zero) and 5th day. A second product was performed by A. versicolor was identified as 2,4-dichlorophenol by comparison its retention time with a standard. This product was
-
Cl + end products
triclosan
2,4-dichlorophenol
Fig. 4. A possible biodegradation stoichiometry for triclosan by A. versicolor.
also detected in the study of Hundt et al. [18]. 2,4-dichlorophenol was formed in small amounts from triclosan by Trametes versicolor in their study. When compared data there is a 2,4dichlorophenol peak at time zero. This peak decreased after the fungal degradation at Peak 2. At the end of triclosan degradation experiments, a stoichiometric release of chloride ions was also observed in the previous studies after 2,4-dichlorophenol producted [12]. A third product detected after 2,4-dichlorophenol probably is Cl− according to stoichiometry of triclosan (Fig. 4). This is also an important data that fungus removed not only triclosan but also its product 2,4-dichlorophenol. The other peaks that occurred initially probably related with media content. According to HPLC analyses and previous reports it is suggested that A. versicolor showed biodegradation ability and dechlorination of triclosan based on stoichiometric release of chloride. 4. Conclusion Triclosan biodegradation by using an effective biomaterial and finding the cost effective method is still a challenging problem. The maximum triclosan biodegradation yield was achieved by A. versicolor as 71.91% at 7.5 mg/L triclosan concentration. To the best of our knowledge this is the best yield that was obtained by any of the microorganisms. The results showed that A. versicolor can be used as a potential bioaccumulator for triclosan biodegradation process
mAU 11
254nm,4nm (1.00)
Peak 1
10 9
R2=0.999983
Peak 2
8 7 6 5 4 3
2,4-dichlorophenol
2
Triclosan 7.040
1 0 -1
7.017 0.0
2.5
5.0
7.5
min
Fig. 3. Detection of the triclosan degradation process by comparing different HPLC data at different incubation time. Peak 1: The data at the time zero, (C0); Peak 2: The data at the end of the fungal incubation (5th day); R2 = 0.99983; Retention time of triclosan, 7 min; Retention time of 2,4-dichlorophenol, 4 min.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
up to high triclosan levels. The study is the first report that offered A. versicolor as an effective biomaterial in triclosan removal process. Acknowledgment The authors are grateful to the Scientific and Technological Research Council of Turkey (TÜBİTAK and TÜBİTAK-BİDEB) for financial support. References [1] United States Environmental Protection Agency. (EPA) Washington, D.C. 20460 Offıce of Prevention, Pestıcıdes and Toxıc Substances September 11, 2008. [2] L.M. McMurry, M. Oethinger, S.B. Levy, Triclosan targets lipid synthesis, Nature 394 (1998) 531–532. [3] A.B. Dann, A. Hontela, Triclosan: environmental exposure, toxicity and mechanisms of action, J. Appl. Toxicol. 31 (4) (2011) 285–311, doi:10.1002/ jat.1660. [4] J.L. Wu, N.P. Lam, D. Martens, A. Kettrup, Z. Cai, Triclosan determination in water related to wastewater treatment, Talanta 72 (2007) 1650–1654. [5] A.D. Dayan, Risk assessment of triclosan [Irgasan®] in human breast milk, Food Chem. Toxicol. 45 (2007) 125–129. [6] G.G. Ying, R.S. Kookana, Triclosan in wastewaters and biosolids from Australian wastewater treatment plants, Environ. Int. 33 (2007) 199–205. [7] Z. Lan, T. Hyung Kim, K. Shun Bi, X. Hui Chen, H. Sik Kim, Triclosan exhibits a tendency to accumulate in the epididymis and shows sperm toxicity in male sprague-dawley rats, Environ. Toxicol. (2013) doi:10.1002/tox.21897. [8] Y.Z. Ren, M. Franke, F. Anschuetz, B. Ondruschka, A. Ignaszak, P. Braeutigam, Sonoelectrochemical degradation of triclosan in water, Ultrason. Sonochem. 21 (6) (2014) 2020–2025. [9] H. Liu, X. Cao, G. Liu, Y. Wang, N. Zhang, T. Li, et al., Photoelectrocatalytic degradation of triclosan on TiO2 nanotube arrays and toxicity change, Chemosphere 93 (1) (2013) 160–165. [10] Z. Song, N. Wang, L. Zhu, A. Huang, X. Zhao, H. Tang, Efficient oxidative degradation of triclosan by using an enhanced Fenton-like process, Chem. Eng. J. 198–199 (2012) 379–387. [11] I. Sirés, N. Oturan, M.A. Oturan, R.M. Rodríguez, J.A. Garrido, E. Brillas, ElectroFenton degradation of antimicrobials triclosan and triclocarban, Electrochim. Acta 52 (17) (2007) 5493–5503.
5
[12] D.G. Lee, F. Zhao, Y.H. Rezenom, D.H. Russell, K.H. Chu, Biodegradation of triclosan by a wastewater microorganism, Water Res. 46 (13) (2012) 4226–4234. [13] Y. Gao, Y. Ji, G. Li, T. An, Mechanism, kinetics and toxicity assessment of OH-initiated transformation of triclosan in aquatic environments, Water Res. 49 (2014) 360–370. [14] C. Riva, S. Cristoni, A. Binelli, Effects of triclosan in the freshwater mussel Dreissena polymorpha: a proteomic investigation, Aquat. Toxicol. 118–119 (15) (2012) 62–71. [15] K. Malachova, Z. Rybkova, H. Sezimova, J. Cerven, C. Novotny, Biodegradation and detoxification potential of rotating biological contactor (RBC) with Irpex lacteus for remediation of dye-containing wastewater, Water Res. 47 (19) (2013) 7143–7148. [16] A. Mishra, A. Malik, Simultaneous bioaccumulation of multiple metals from electroplating effluent using Aspergillus lentulus, Water Res. 46 (16) (2012) 4991–4998. [17] B. Ertit Taştan, S. Ertuğrul, G. Dönmez, Effective bioremoval of reactive dye and heavy metals by Aspergillus versicolor, Bioresour. Technol. 101 (2010) 870– 876. [18] K. Hundt, D. Martin, E. Hammer, U. Jonas, K.K. Markus, F. Schauer, Transformation of triclosan by Trametes versicolor and Pycnoporus cinnabarinus, Appl. Environ. Microbiol. (2000) doi:10.1128/AEM.66.9.4157-4160.2000. [19] U. Bracklow, A. Drews, M. Vocks, M. Kraume, Comparison of nutrients degradation in small scale membrane bioreactors fed with synthetic/domestic wastewater, J. Hazard. Mater. 144 (2007) 620–626. [20] S.K. Behera, S.Y. Oh, H.S. Park, Sorption of triclosan onto activated carbon, kaolinite and montmorillonite: effects of pH, ionic strength, and humic acid, J. Hazard. Mater. 179 (2010) 684–691. [21] B. Ertit Taştan, E. Duygu, G. Dönmez, Boron bioremoval by a newly isolated Chlorella sp. and its stimulation by growth stimulators, Water Res. 46 (2012) 167–175. [22] M. Ricart, H. Guasch, M. Alberch, D. Barceló, C. Bonnineau, A. Geiszinger, et al., Triclosan persistence through wastewater treatment plants and its potential toxic effects on river biofilms, Aquat. Toxicol. 100 (2010) 346–353. [23] S. Wang, X. Wang, K. Poon, Y. Wang, S. Li, H. Liu, et al., Removal and reductive dechlorination of triclosan by Chlorella pyrenoidosa, Chemosphere 92 (11) (2013) 1498–1505. [24] I. Ferrer, M. Mezcua, M.J. Gómez, E.M. Thurman, A. Agüera, M.D. Hernando, et al., Liquid chromatography/time-of-flight mass spectrometric analyses for the elucidation of the photodegradation products of triclosan in wastewater samples, Rapid Commun. Mass Spectrom. 18 (2004) 443–450. [25] Y.M. Kim, K. Murugesan, S. Schmidt, V. Bokare, J.R. Jeon, E.J. Kim, et al., Triclosan susceptibility and co-metabolism – a comparison for three aerobic pollutantdegrading Bacteria, Bioresour. Technol. 102 (3) (2011) 2206–2212.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
Contents lists available at ScienceDirect
Pesticide Biochemistry and Physiology j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / p e s t
Biodegradation of pesticide triclosan by A. versicolor in simulated wastewater and semi-synthetic media Burcu Ertit Taştan a,b,c,*, Gönül Dönmez a a b c
Department of Biology, Faculty of Science, Ankara University, 06100, Beşevler, Ankara, Turkey Life Sciences Application and Research Center, Gazi University, 06830, Gölbaşı, Ankara, Turkey Health Services Vocational School, Gazi University, 06830, Gölbaşı, Ankara, Turkey
A R T I C L E
I N F O
Article history: Received 24 June 2014 Accepted 7 November 2014 Available online Keywords: Fungus Triclosan 2,4-dichlorophenol Biodegradation Wastewater treatment
A B S T R A C T
Triclosan is known as an antimicrobial agent, a powerful bacteriostat and an important pesticide. In this paper biodegradation of triclosan by Aspergillus versicolor was investigated. Effects of simulated wastewater and semi-synthetic media on fungal triclosan degradation process were detected. HPLC analysis showed that fungal triclosan biodegradation yield was 71.91% at about 7.5 mg/L concentration in semisynthetic medium and was 37.47% in simulated wastewater. Fungus could be able to tolerate the highest triclosan concentration (15.69 mg/L). The biodegradation yield was 29.81% and qm was 2.22 mg/g at this concentration. Some of the parameters, such as pH, culture media, increasing triclosan and biomass concentrations were optimized in order to achieve the effective triclosan biodegradation process. The highest triclosan biodegradation yields of all microorganisms were achieved by A. versicolor. © 2014 Elsevier Inc. All rights reserved.
1. Introduction Triclosan, 5-chloro-2-(2,4-dichlorophenoxy)phenol or commercially known as Irgasan is an antimicrobial agent and a powerful bacteriostat. It has also been registered as a pesticide by the United States Environmental Protection Agency [1]. It blocks lipid biosynthesis in bacteria by inhibiting the enoyl reductase enzyme [2]. Thus, since its introduction in the 1960s, it has been widely used in soaps, shampoos, toothpastes, hand washers and disinfectants. Two tons of triclosan has been consumed per year only in Sweden [3]. However this fact seems to be a marketing strategy rather than a medical necessity. The safety profile of triclosan has been questioned recently after the observation that triclosan was determined not only in wastewater [4] but also in human breast milk [5]. Analyses performed by Ying and Kookana [6] showed that triclosan had a tendency to take part in the environment and it is a persistent pollutant, which does not degrade fast; its primary biodegradation half-life was given as “weeks” and ultimate biodegradation half-life was expected to take “months.” Due to its long half-life and lipophilic characteristics, it may potentially cause longterm health risks in human body. Animal studies suggest that triclosan can have a harmful effect on the reproductive system [7]. Therefore, it is necessary to develop an environmental friendly and
* Corresponding author. Life Sciences Application and Research Center, Gazi University, 06830, Gölbaşı, Ankara, Turkey. Fax: +90 312 484 62 71. E-mail address: [email protected] (B. Ertit Taştan).
fast method for triclosan biodegradation in order to reduce environmental pollution. Attention has been attracted to triclosan degradation processes in recent studies. Some of the triclosan degradation methods are sonoelectrochemical [8], photoelectrocatalytic [9], oxidative [10] and electro-fenton degradation [11]. Studies suggest that biodegradation is one of the most efficient methods for triclosan degradation. Different transformation metabolites could be detected during triclosan biodegradation and while degradation metabolites show androgenic responses, the end products do not [12]. It has been demonstrated that triclosan was highly toxic to organisms, especially to aquatic ones [13,14]. Therefore, in order to increase the triclosan removal efficiency, it is important to select the highly triclosan tolerant microorganisms. Fungi are often used and preferred in biological treatment studies due to their high pollutant removal yields even at their high concentrations as proven by several recent studies [15–17]. Hundt et al. [18] described some biotransformation reactions of triclosan by the white rot fungi Trametes versicolor and Pycnoporous cinnabarinus. T. versicolor produced 3 metabolites including 2,4-dichlorophenol. They explained that the conjugates showed a distinctly lower cytotoxic activity than triclosan. In an effort to address the biodegradation of increasing triclosan concentrations, this study aimed to examine several hypothesis: (i) whether triclosan blocks lipid biosynthesis in bacteria and whether fungal biomass might have higher resistance to its toxicity than bacteria; the fungal biomass could serve as an effective biomaterial, (ii) whether triclosan biodegradation realized not only in rich microbiological media
http://dx.doi.org/10.1016/j.pestbp.2014.11.002 0048-3575/© 2014 Elsevier Inc. All rights reserved.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
2
but also in simulated wastewater, it can be an original and useful study for real wastewater treatment systems. We also aimed to check the best triclosan biodegradation process by changing experimental parameters. 2. Materials and methods 2.1. Chemicals Triclosan 97% (Irgasan CAS: 3380-34-5) was purchased from Sigma; 2,4-Dichlorophenol 99% (CAS: 120-83-2) was supplied by Sigma-Aldrich. HPLC grade acetonitrile and ammonium acetate were purchased from Merck. The stock solutions were prepared in acetonitrile (80%)/ammonium acetate (20%) solution and were stored in a refrigerator at 4 °C. Appropriate volumes of the stock solutions were added to the media. 2.2. Culture conditions Aspergillus versicolor was isolated previously by Ertit Taştan et al. [17] from Batman, Turkey. Fungal biomass was inoculated into 250 mL Erlenmeyer flasks containing known triclosan concentrations in 100 mL culture media as described below. The flasks were incubated at 25 ± 2 °C on a rotary shaker (New Brunswick Scientific Innova 4230) at 100 rpm for 5 days. In order to obtain an equal initial biomass, 100 mL media was inoculated with 0.4 g/L dry weight biomass. 2.3. Growth medium Czapek Dox Medium (CDM) was selected as the first test media, which was a semisynthetic medium used for the cultivation of fungi and especially for Aspergillus spp. The ingredients (g/L) were NaNO2, 2; KCl, 0.5; MgSO4.7H2O, 0.5; K2HPO4, 1; FeSO4.7H2O, 0.01; Sucrose 30. Simulated Waste Water (SWW) was the second selected media. The composition of SWW was prepared according to Bracklow et al. [19] considering its resemblance to municipal wastewater and we slightly modified it for this study. The ingredients (mg/L) were as follows: peptone 25, yeast extract 80, starch 200, sunflower oil 35, ammonium acetate 150, KH2PO4 26, MgHPO4.3H2O 6, glucose 2000, K2HPO4 26, urea 50, FeSO4.7H2O 8, whey powder 160. Both of these media were autoclaved at 121 °C for 15 min before the inoculation step. Medial pH values were adjusted to appropriate volumes by adding diluted (0.01 M) and concentrated (1 M) sulfuric acid or sodium hydroxide solutions. 2.4. Analytical methods Triclosan biodegradation was determined on the samples (3 mL) taken daily during the incubation period. Uninoculated Erlenmeyer flasks containing triclosan were used as control samples to detect any reactions between media and triclosan. Growth of A. versicolor was determined by measuring the dry weight of biomass at the end of the incubation period. The concentration of triclosan in the supernatant was determined by high-performance liquid chromatography (Shimadzu, Kyoto, Japan), by using a C-18 column (250 mm × 4.6 mm inner diameter: 5 μm particle size) at 50 °C. Acetonitrile and 10 mM ammonium acetate were used as mobile phase to produce a binary elution gradient with a flow rate of 1 mL/min. Pure water was obtained from an apparatus (Neuberger, Germany), and it was used to prepare solutions and buffers, which were filtered through 0.45 μm Teknokroma membrane filters prior to use. The sample solutions were filtered through 0.20 μm Minisart RC syringe filters before the
injection into the chromatograph. The separation of triclosan was achieved with the following linear mobile phase gradient program: at 6.85 min 80:20 (v/v) at 254 nm. The percentage biodegradation of triclosan and qm (the maximum specific triclosan uptake) was calculated from the relevant equations. In the study, qm represents the maximum amount of triclosan biodegradation per unit dry weight of fungal cells (mg/g), X maximum dried cell mass (g/L), and C0 the initial concentration of the triclosan (mg/L), respectively. The data were subjected to analysis of significant differences among treatment means were compared by descriptive statistics (±S.E.).
2.5. pH experiments The effect of pH on triclosan biodegradation process was investigated at pH 5, 6, 7 and 8 at about 7.5 mg/L triclosan concentration for 5 days of incubation period in both of the growth media. Each of these experiments was performed in triplicates. There was not a significant change after 3 days of incubation period; therefore these results were taken into consideration.
2.6. Triclosan concentration experiments In order to highlight the effect of initial triclosan concentrations, the experiments were conducted at 5, 7.5, 12.5 and 15.5 mg/L triclosan concentrations. Each of these experiments was performed in triplicates.
2.7. Fungal biomass experiments The effect of fungal biomass concentrations on triclosan biodegradation process was also examined at three different biomass concentrations by using the dry weight method. The experiments were performed at 7.5 mg/L triclosan concentration at pH 7 in CDM and at pH 5 in SWW. For these experiments 100 mL media was inoculated with 0.4, 0.8 and 1.1 g/L dry weight biomass.
3. Results and discussions 3.1. Effects of pH on triclosan biodegradation in different culture media Change in pH values is an important parameter affecting the dissociation of triclosan molecule. The dissociation constant of triclosan is given as pKa = 8.14, and when pH > pKa more than 50% of the triclosan is expected to be deprotonated [20]. As seen in Fig. 1, when pH was increased from 5 to 7 in CDM triclosan biodegradation increased. The maximum biodegradation was observed at pH 7 as 55.00%. The selected optimum pH rate was used at the next steps of experiments. Previous studies showed that pollutant biodegradation yield was really linked to culture media [21]. In order to make this work applicable in daily life and to obtain a better understanding of the triclosan biodegradation process, we also examined the effect of SWW, which is close to municipal wastewater. The aim was to reveal if triclosan biodegradation process efficiency could also be obtained in SWW. Therefore, the same concentrations with CDM were tested in SWW. The results are presented in Fig. 1. The biodegradation rates decreased when pH was increased from 5 to 8. The maximum biodegradation was observed at pH 5 as 50.63%. The selected optimum pH rate was used at the next step of experiments.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
3
Fig. 2. Effects of triclosan concentrations on the biodegradation yield (Y%) of A. versicolor during the incubation period in CDM (pH: 7; T, 25 ± 2 °C; stirring rate: 100 rpm). Fig. 1. Effect of pH on triclosan biodegradation (Y%) of A. versicolor at about 7.5 mg/L triclosan concentration in CDM and SWW (incubation period, 3 days; T, 25 ± 2 °C; stirring rate: 100 rpm).
3.2. Effect of increasing triclosan concentrations on biodegradation process Triclosan concentrations are important parameter, because it was proven that microorganisms were affected by increasing triclosan concentrations [22]. Taking into consideration of daily increasing triclosan concentrations, the effects of this parameter was tested at four different concentrations (5, 7.5, 12.5 and 15 mg/L) in the second series of the experiments. It was clearly seen in Fig. 2 that when concentration was increased up to 15.5 mg/L, the biodegradation rate decreased in CDM. The highest biodegradation rate was 61.81% at 4.195 mg/L triclosan concentration. It should be clearly stated that after the 3rd day the biodegradation rate reached a constant rate. For instance, 35.68% biodegradation was obtained on the 1st day of the experiments and this rate was kept constant in the following days. The highest biodegradation rate was achieved in the last day of the experiments at the highest concentration of 15.692 mg/L as 31.27%. On the other hand, the maximum amount of triclosan biodegradation per unit dry weight of fungal cells (qm) was 0.872 mg/g at 4.195 mg/L and it was 2.225 mg/L at 15.692 mg/L (Table 1). The reason why the qm values were higher at highest triclosan concentrations than at lowest concentrations was due to the existence of higher concentrations of triclosan in culture media, which was accumulated by fungal cells. There was also a clear correlation between the biodegradation rates. Triclosan biodegradation rates in CDM and SWW were similar at low concentrations. As seen in Table 1, the biodegradation rates
Table 1 Comparison of the biodegradation yields and the maximum specific triclosan uptake (qm) values at different triclosan concentrations (C0) in CDM of A. versicolor. Co (mg/L) CDM 4.19 7.07 12.51 15.69 SWW 2.5 6.88 11.32 13.23
Y%
qm (mg/g)
61.81 ± 4.61 55.00 ± 2.02 34.96 ± 2.09 29.81 ± 1.64
0.87 ± 0.12 1.26 ± 0.26 1.96 ± 0.04 2.22 ± 0.01
of lower concentrations were more remarkable than the rates at higher ones in SWW. The maximum biodegradation rate was 64.56% at 2.50 mg/L triclosan concentration, and this rate decreased to 37.47% when concentration increased to 6.88 mg/L. When compared, a 34.96% yield was obtained in CDM at 12.51 mg/L triclosan concentration, and 18.38% yield was obtained at 11.32 mg/L in SWW. In this medium the qm values decreased when triclosan concentration was increased. This is related with the biodegradation rates which are lower than CDM. Because lower degradation yields was achieved in SWW and thus qm values was lower. In an another study Chlorella pyrenoidosa was exposed to a series of triclosan concentrations ranging from 100 to 800 ng/mL, and it was observed that more than 50% of triclosan was eliminated by algal uptake from the culture media [23]. These triclosan concentrations were lower than the ones tested at the present work. It is important to detect the most triclosan tolerant microorganism, considering its increasing concentrations in the environment. 3.3. Effect of increasing fungal biomass concentrations on biodegradation process To detect the effect of increasing fungal biomass concentrations on biodegradation process 0.4, 0.8 and 1.1 g/L fungal biomass was inoculated into media. It is evident from the results presented in Table 2 that biodegradation rates were really linked to the amount of initial biomass concentrations. The highest biodegradation rate was 55.00% at the first biomass concentration within 3 days in CDM. On the other hand fungus removed 71.91% triclosan at 1.1 g/L biomass concentration. This rate was 1.3 times higher than the rate that was obtained at 0.4 g/L biomass concentration.
Table 2 Comparison of the biodegradation yields and the maximum specific triclosan uptake (qm) values at different fungal biomass concentrations at about 7.5 mg/L triclosan concentrations in CDM and SWW. X0 (g/L)
CDM 64.56 ± 2.63 37.47 ± 1.09 18.38 ± 3.09 16.47 ± 3.60
T, 25 ± 2 °C; stirring rate, 100 rpm; incubation period, 3 days; pH 7.
0.59 ± 0.19 1.66 ± 0.36 1.50 ± 0.03 1.01 ± 0.02
SWW
Y% qm (mg/g) Y% qm (mg/g)
0.4
0.8
1.1
55.00 ± 2.02 1.26 ± 0.26 37.47 ± 1.09 1.66 ± 0.35
66.44 ± 5.62 1.49 ± 0.10 38.04 ± 4.23 1.00 ± 0.26
71.91 ± 4.60 1.99 ± 0.38 46.12 ± 2.75 0.78 ± 0.05
C0, 7.5 mg/L triclosan; X0, initial fungal biomass concentration; T, 25 ± 2 °C; stirring rate, 100 rpm; incubation period, 3 days; pH 7.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
4
As presented in Table 2, an increase in the initial biomass concentration from 0.4 g/L to 1.1 g/L resulted in a significant increase of the maximum triclosan biodegradation per unit dry weight of A. versicolor, which was 1.26, 1.49 and 1.99 mg/g, respectively. The qm value at the maximum initial biomass concentration was also found considerably higher than the minimum biomass concentration, due to higher biodegradation yields. At the maximum biomass concentration (1.1 g/ L) in CDM, qm increased nearly 1.6 times. As expected, when biomass concentrations were higher at a known triclosan concentration, the maximum triclosan quantity per unit dry weight of the fungus would be higher, due to higher biodegradation rates and the lowest level of residual triclosan concentrations in culture media. In contrast to biodegradation efficiency, qm decreased from 1.66 to 0.78 mg/g in SWW. The maximum triclosan biodegradation was recorded as 46.12% at maximum initial biomass concentration; however this biodegradation rate was not equal to the minimum biodegradation yield that was obtained in CDM. Therefore, it is concluded that increases in initial biomass concentration accompanied by decreasing qm values in SWW, contrary to the changes in CDM. In another study Sphingopyxis strain KCY1 was grown on complex nutrient medium containing 5 mg/L triclosan and qm was found as 0.13 mg-triclosan/mg-protein/day. They suggested a possible biodegradation pathway for triclosan by strain KCY1 [12]. Song et al. [10] investigated the triclosan degradation in the presence of H2O2 as oxidant and BiFeO3 magnetic nanoparticles as catalyst. HPLC measurements showed that 2,4-dichlorophenol was the major degradation intermediate of triclosan. They explained that except for the 2,4-dichlorophenol pathways, the direct dechlorination process also occurred like Ferrer et al. [24] detected in their triclosan study in wastewater samples. It was also explained that 2,4-dichlorophenol was less toxic to Sphingomonas sp. than triclosan [25]. This finding has also been supported in our present work for A. versicolor. Triclosan degradation is shown in Fig. 3 with HPLC data at different times as C0 (time zero) and 5th day. A second product was performed by A. versicolor was identified as 2,4-dichlorophenol by comparison its retention time with a standard. This product was
-
Cl + end products
triclosan
2,4-dichlorophenol
Fig. 4. A possible biodegradation stoichiometry for triclosan by A. versicolor.
also detected in the study of Hundt et al. [18]. 2,4-dichlorophenol was formed in small amounts from triclosan by Trametes versicolor in their study. When compared data there is a 2,4dichlorophenol peak at time zero. This peak decreased after the fungal degradation at Peak 2. At the end of triclosan degradation experiments, a stoichiometric release of chloride ions was also observed in the previous studies after 2,4-dichlorophenol producted [12]. A third product detected after 2,4-dichlorophenol probably is Cl− according to stoichiometry of triclosan (Fig. 4). This is also an important data that fungus removed not only triclosan but also its product 2,4-dichlorophenol. The other peaks that occurred initially probably related with media content. According to HPLC analyses and previous reports it is suggested that A. versicolor showed biodegradation ability and dechlorination of triclosan based on stoichiometric release of chloride. 4. Conclusion Triclosan biodegradation by using an effective biomaterial and finding the cost effective method is still a challenging problem. The maximum triclosan biodegradation yield was achieved by A. versicolor as 71.91% at 7.5 mg/L triclosan concentration. To the best of our knowledge this is the best yield that was obtained by any of the microorganisms. The results showed that A. versicolor can be used as a potential bioaccumulator for triclosan biodegradation process
mAU 11
254nm,4nm (1.00)
Peak 1
10 9
R2=0.999983
Peak 2
8 7 6 5 4 3
2,4-dichlorophenol
2
Triclosan 7.040
1 0 -1
7.017 0.0
2.5
5.0
7.5
min
Fig. 3. Detection of the triclosan degradation process by comparing different HPLC data at different incubation time. Peak 1: The data at the time zero, (C0); Peak 2: The data at the end of the fungal incubation (5th day); R2 = 0.99983; Retention time of triclosan, 7 min; Retention time of 2,4-dichlorophenol, 4 min.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
ARTICLE IN PRESS B. Ertit Taştan, G. Dönmez/Pesticide Biochemistry and Physiology ■■ (2014) ■■–■■
up to high triclosan levels. The study is the first report that offered A. versicolor as an effective biomaterial in triclosan removal process. Acknowledgment The authors are grateful to the Scientific and Technological Research Council of Turkey (TÜBİTAK and TÜBİTAK-BİDEB) for financial support. References [1] United States Environmental Protection Agency. (EPA) Washington, D.C. 20460 Offıce of Prevention, Pestıcıdes and Toxıc Substances September 11, 2008. [2] L.M. McMurry, M. Oethinger, S.B. Levy, Triclosan targets lipid synthesis, Nature 394 (1998) 531–532. [3] A.B. Dann, A. Hontela, Triclosan: environmental exposure, toxicity and mechanisms of action, J. Appl. Toxicol. 31 (4) (2011) 285–311, doi:10.1002/ jat.1660. [4] J.L. Wu, N.P. Lam, D. Martens, A. Kettrup, Z. Cai, Triclosan determination in water related to wastewater treatment, Talanta 72 (2007) 1650–1654. [5] A.D. Dayan, Risk assessment of triclosan [Irgasan®] in human breast milk, Food Chem. Toxicol. 45 (2007) 125–129. [6] G.G. Ying, R.S. Kookana, Triclosan in wastewaters and biosolids from Australian wastewater treatment plants, Environ. Int. 33 (2007) 199–205. [7] Z. Lan, T. Hyung Kim, K. Shun Bi, X. Hui Chen, H. Sik Kim, Triclosan exhibits a tendency to accumulate in the epididymis and shows sperm toxicity in male sprague-dawley rats, Environ. Toxicol. (2013) doi:10.1002/tox.21897. [8] Y.Z. Ren, M. Franke, F. Anschuetz, B. Ondruschka, A. Ignaszak, P. Braeutigam, Sonoelectrochemical degradation of triclosan in water, Ultrason. Sonochem. 21 (6) (2014) 2020–2025. [9] H. Liu, X. Cao, G. Liu, Y. Wang, N. Zhang, T. Li, et al., Photoelectrocatalytic degradation of triclosan on TiO2 nanotube arrays and toxicity change, Chemosphere 93 (1) (2013) 160–165. [10] Z. Song, N. Wang, L. Zhu, A. Huang, X. Zhao, H. Tang, Efficient oxidative degradation of triclosan by using an enhanced Fenton-like process, Chem. Eng. J. 198–199 (2012) 379–387. [11] I. Sirés, N. Oturan, M.A. Oturan, R.M. Rodríguez, J.A. Garrido, E. Brillas, ElectroFenton degradation of antimicrobials triclosan and triclocarban, Electrochim. Acta 52 (17) (2007) 5493–5503.
5
[12] D.G. Lee, F. Zhao, Y.H. Rezenom, D.H. Russell, K.H. Chu, Biodegradation of triclosan by a wastewater microorganism, Water Res. 46 (13) (2012) 4226–4234. [13] Y. Gao, Y. Ji, G. Li, T. An, Mechanism, kinetics and toxicity assessment of OH-initiated transformation of triclosan in aquatic environments, Water Res. 49 (2014) 360–370. [14] C. Riva, S. Cristoni, A. Binelli, Effects of triclosan in the freshwater mussel Dreissena polymorpha: a proteomic investigation, Aquat. Toxicol. 118–119 (15) (2012) 62–71. [15] K. Malachova, Z. Rybkova, H. Sezimova, J. Cerven, C. Novotny, Biodegradation and detoxification potential of rotating biological contactor (RBC) with Irpex lacteus for remediation of dye-containing wastewater, Water Res. 47 (19) (2013) 7143–7148. [16] A. Mishra, A. Malik, Simultaneous bioaccumulation of multiple metals from electroplating effluent using Aspergillus lentulus, Water Res. 46 (16) (2012) 4991–4998. [17] B. Ertit Taştan, S. Ertuğrul, G. Dönmez, Effective bioremoval of reactive dye and heavy metals by Aspergillus versicolor, Bioresour. Technol. 101 (2010) 870– 876. [18] K. Hundt, D. Martin, E. Hammer, U. Jonas, K.K. Markus, F. Schauer, Transformation of triclosan by Trametes versicolor and Pycnoporus cinnabarinus, Appl. Environ. Microbiol. (2000) doi:10.1128/AEM.66.9.4157-4160.2000. [19] U. Bracklow, A. Drews, M. Vocks, M. Kraume, Comparison of nutrients degradation in small scale membrane bioreactors fed with synthetic/domestic wastewater, J. Hazard. Mater. 144 (2007) 620–626. [20] S.K. Behera, S.Y. Oh, H.S. Park, Sorption of triclosan onto activated carbon, kaolinite and montmorillonite: effects of pH, ionic strength, and humic acid, J. Hazard. Mater. 179 (2010) 684–691. [21] B. Ertit Taştan, E. Duygu, G. Dönmez, Boron bioremoval by a newly isolated Chlorella sp. and its stimulation by growth stimulators, Water Res. 46 (2012) 167–175. [22] M. Ricart, H. Guasch, M. Alberch, D. Barceló, C. Bonnineau, A. Geiszinger, et al., Triclosan persistence through wastewater treatment plants and its potential toxic effects on river biofilms, Aquat. Toxicol. 100 (2010) 346–353. [23] S. Wang, X. Wang, K. Poon, Y. Wang, S. Li, H. Liu, et al., Removal and reductive dechlorination of triclosan by Chlorella pyrenoidosa, Chemosphere 92 (11) (2013) 1498–1505. [24] I. Ferrer, M. Mezcua, M.J. Gómez, E.M. Thurman, A. Agüera, M.D. Hernando, et al., Liquid chromatography/time-of-flight mass spectrometric analyses for the elucidation of the photodegradation products of triclosan in wastewater samples, Rapid Commun. Mass Spectrom. 18 (2004) 443–450. [25] Y.M. Kim, K. Murugesan, S. Schmidt, V. Bokare, J.R. Jeon, E.J. Kim, et al., Triclosan susceptibility and co-metabolism – a comparison for three aerobic pollutantdegrading Bacteria, Bioresour. Technol. 102 (3) (2011) 2206–2212.
Please cite this article in press as: Burcu Ertit Tas¸tan, Gönül Dönmez, Biodegradation of pesticide triclosan by A.versicolor in simulated wastewater and semi-synthetic media, Pesticide Biochemistry and Physiology (2014), doi: 10.1016/j.pestbp.2014.11.002
