Elsevier

New Biotechnology

Volume 29, Issue 1, 15 December 2011, Pages 107-115
New Biotechnology

Research paper
Biotransformation of the organochlorine pesticide trans-chlordane by wood-rot fungi

https://doi.org/10.1016/j.nbt.2011.06.013Get rights and content

There is very limited information on the biotransformation of organochlorine pesticide chlordane by microorganisms, and no systematic study on the metabolic products and pathways for chlordane transformation by wood-rot fungi has been conducted. In this study, trans-chlordane was metabolized with the wood-rot fungi species Phlebia lindtneri, Phlebia brevispora and Phlebia aurea, which are capable of degrading polychlorinated dibenzo-p-dioxin and heptachlor epoxide. At the end of 42 days of incubation, over 50% of trans-chlordane was degraded by the fungal treatments in pure cultures. These fungi transformed trans-chlordane to at least eleven metabolites including a large amount of hydroxylated products such as 3-hydroxychlordane, chlordene chlorohydrin, heptachlor diol, monohydroxychlordene and dihydroxychlordene. P. lindtneri particularly can metabolize oxychlordane, a recalcitrant epoxide product of chlordane, into a hydroxylated product through substitution of chlorine atom by hydroxyl group. The present results suggest that hydroxylation reactions play an important role in the metabolism of trans-chlordane by these Phlebia species. Additionally, transformation of trans-chlordane and production of hydroxylated metabolites were efficiently inhibited by the addition of cytochrome P450 inhibitors, piperonyl butoxide and 1-aminobenzotriazole, demonstrating that fungal cytochrome P450 enzymes are involved in some steps of trans-chlordane metabolism, particularly in the hydroxylation process.

Introduction

Chlordane is an organochlorine pesticide that was at one time extensively used for both agricultural and residential applications around the world. An estimated 70,000 metric tons of technical chlordane were produced between 1948 and 1988, and nearly 20% of the total production still exists unaltered in the environment because of the environmental half-life of the pesticide, which is estimated to be between 5 and 15 years. Chlordane was heavily and widely used in Japan for household termite control, especially during the early 1980s [1]. Although the use of commercial chlordane was banned in Japan in 1986 and worldwide in 2004, chlordane and related compounds are still commonly found in biota, human and environmental samples because of chlordane's high partition coefficient (log KOW = 6.0), and there is concern about the adverse effects of these compounds for humans and ecosystems 1, 2, 3, 4. Because of its persistence and toxicity 1, 2, technical chlordane has also been listed as one of the 12 persistent organic pollutants (POPs) subject to global treaty restrictions in the UNER Stockholm Convention on POPs signed in 2001. Technical chlordane is known to consist of 147 distinct components, with trans-chlordane (13.2%) and cis-chlordane (11.3%) present in the highest amounts in the technical mixture [4].

Because contamination with POPs is still a serious problem in the agricultural environment, an efficient method for remediation is needed. Thermal desorption, thus for the only effective removal option of these pesticides from soil, is quite costly and leaves soil sterile [5]. Therefore, another method of ensuring complete restoration of all soil functions, like bioremediation, needs to be developed. Microbial degradation is regarded as an important process for the removal of POPs from the environment, and many studies on the biotransformation of POPs by microorganisms have been conducted. However, to data, little fundamental research on the biotransformation of chlordane has been carried out 6, 7, 8. To our knowledge, only one published study has examined the metabolic products of chlordane in microorganisms, and heptachlor, heptachlor epoxide, dichlorochlordene, oxychlordane, chlordene chlorohydrin, and 3-hydroxychlordane were the products of the microbial transformation of chlordane [7]. Wood-rot fungi can degrade lignin, a complex high-molecular weight aromatic polymer, and have been demonstrated as capable of transforming and/or mineralizing a wide range of organopollutants [9]. Therefore, wood-rot fungi represent a prospective tool in environmental bioremediation. However, there is a surprising lack of data on the metabolic products and metabolic pathway for the biotransformation of chlordane in wood-rot fungi.

In our previous study, we chose several wood-rot fungi for examination based on their ability to degrade dioxin, and found that Phlebia lindtneri and Phlebia brevispora could hydroxylate polychlorinated dibenzo-p-dioxins (PCDDs), such as 2,7-diCDD, 2,3,7-triCDD, 1,2,8,9-tetraCDD, 1,2,6,7-tetraCDD, 1,3,6,8-tetraCDD and 2,8-dichlorobenzofuran 10, 11, 12. It was also reported that chloronaphthalene, PCBs and dieldrin were metabolized by Phlebia species 13, 14, 15. It has been shown that hydroxylation is important as the first reaction in the transformation of these xenobiotics. In addition, our previous work evaluated the ability of 18 Phlebia species to degrade heptachlor and recalcitrant heptachlor epoxide, and showed that P. lindtneri, P. brevispora, and Phlebia aurea can transform heptachlor epoxide to hydroxylated products via hydrolysis and hydroxylation [16]. These studies suggest that Phlebia species have specific activity for the biotransformation of organohalogen compounds such as PCDDs and organochlorine pesticides, and led us to investigate the ability of Phlebia species to metabolize chlordane.

In this experiment, we evaluated the ability of several Phlebia species to degrade trans-chlordane and its metabolite oxychlordane, and describe newly identified metabolites obtained by fungal treatment. We also investigated the effects of cytochrome P450 inhibitors on the transformation process of trans-chlordane by P. lindtneri.

Section snippets

Chemicals

trans-Chlordane (99% pure), heptachlor, heptachlor epoxide, N,N-dimethylformamide, acetic anhydride, pyridine piperonyl butoxide (PB), 1-aminobenzotriazole (ABT) and all organic solvents were purchased from Wako Pure Chemical Industries (Osaka, Japan). Oxychlordane was purchased from Otsuka Pharmaceutical Co., Ltd. (Tokushima, Japan). Trimethylsilyldiazomethane was purchased from Tokyo Kasei Chemical Industry (Tokyo, Japan).

Fungi and culture conditions

Three species belonging to the genus Phlebia were used for

Biodegradation of trans-chlordane

The time courses for the degradation of trans-chlordane by the treatment with P. lindtneri, P. brevispora and P. aurea are shown in Fig. 1. In LN medium, the three fungi have a similar degradation curve throughout the incubation period. This pesticide was degraded approximately 21%, 25%, and 30% by P. lindtneri, P. brevispora and P. aurea during 14 days of incubation, respectively, while about 50% of trans-chlordane was degraded after 42 days of incubation by the three fungi. On the contrary,

Discussion

In contrast to the great number of studies on the metabolism of chlordane in mammals and fish, reports on the metabolism of chlordane in microorganisms are scarce. There is only one published study that examined the metabolic products of chlordane by microorganisms [7]. An actinomycete (Nocardiopsis sp.) isolated from soil was capable of extensively degrading cis- and trans-chlordane in pure culture, and Nocardiopsis sp. metabolized trans-chlordane to at least eight metabolic products including

Acknowledgements

This work was supported by a grant from Research project for ensuring food safety from farm to table, Ministry of Agriculture, Forestry and Fisheries, Japan (PO-3216).

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