Compound specific stable isotope analysis (CSIA) to characterize transformation mechanisms of α-hexachlorocyclohexane

Highlights

• Isotope fractionation to investigate the mode of chemical reactions of α-HCH.

• Characterization of the mode of reaction mechanisms using AKIE_C.

• Comparison of chemical and biological reaction mechanisms.

• ESIA is feasible for identifying chemical and biological in situ transformations.

Abstract

A systematic investigation of environmentally relevant transformation processes of alpha-hexachlorocyclohexane (α-HCH) was performed in order to explore the potential of compound specific stable isotope analysis (CSIA) to characterize reaction mechanisms. The carbon isotope enrichment factors (ɛ_C) for the chemical transformations of α-HCH via direct photolysis, indirect photolysis (UV/H₂O₂), hydrolysis, electro-reduction or reduction by Fe⁰ were quantified and compared to those previously published for biodegradation. Hydrogen abstraction by hydroxyl radicals generated by UV/H₂O₂ led to ɛ_C of −1.9 ± 0.2‰ with an apparent kinetic carbon isotope effect (AKIE_C) of 1.012 ± 0.001. Dehydrochlorination by alkaline hydrolysis yielded ɛ_C of −7.6 ± 0.4‰ with AKIE_C of 1.048 ± 0.003. Dechlorination either by homolytic bond cleavage in direct photolysis (ɛ_C = −2.8 ± 0.2‰) or single-electron transfer in electro-reduction (ɛ_C = −3.8 ± 0.4‰) corresponded to AKIE_C of 1.017 ± 0.001 and 1.023 ± 0.003, respectively. Dichloroelimination catalyzed by Fe⁰ via two-electron transfers resulted in ɛ_C of −4.9 ± 0.1‰. AKIE_C values assuming either a concerted or a stepwise mechanism were 1.030 ± 0.0006 and 1.015 ± 0.0003, respectively. Contrary to biodegradation, no enantioselectivity of α-HCH was observed in chemical reactions, which might be used to discriminate chemical and biological in situ transformations.

Introduction

Hexachlorocyclohexane (HCH) isomers have been listed in the Stockholm Convention on Persistent Organic Pollutants (POPs) [1], that aims to eliminate or restrict the production and use of POPs. Although production of HCH is mostly banned in countries signing the Stockholm Convention, residues from previous applications, improper waste disposal and storage require long-term monitoring in order to assess HCH sources and sinks. HCH isomers are ubiquitous in the environment due to their previously widespread global usage and physicochemical recalcitrance toward decomposition [2], [3]. Technical-grade HCH, an important insecticide formulation in agriculture, forestry, and wood preservative, consists mainly of five major isomers: α (60–70%), β (5–12%), γ (10–12%), δ (6–10%) and ɛ (3–4%) [4]. The intensive use of technical-grade HCH has released large amounts of α-HCH into the environment and its persistence led to its presence in environmental and biological samples until today [5]. Therefore, understanding of the fate of α-HCH is of great importance for long-term cleanup activities. Although HCH isomers are highly resistant to degradation under the conditions prevailing in the environment, biological and chemical transformation processes and related remediation strategies have been explored to clean up HCH-contaminated sites [6], [7], [8], such as aerobic and anaerobic microbial degradation [9], [10], hydrolysis [11], radical oxidation by photo-Fenton process [12] or by photocatalytic degradation [13], electrochemical reduction [14] or nanoscale zero-valent iron (Fe⁰) induced reduction [15], [16], [17], [18], [19], [20].

Transformation of HCH is typically initiated by hydrogen abstraction, dehydrochlorination, dechlorination or dichloroelimination (Scheme 1). Hydrogen abstraction can be found in radical oxidation processes via reactive hydroxyl radicals (·OH) formed by photo-induced H₂O₂ decomposition [21] or by TiO₂-enhanced photocatalysis [13]. Dehydrochlorination takes place during hydrolysis of γ-HCH under alkaline conditions [22], which might be a similar mechanism to aerobic degradation of α-HCH by Sphingomonas paucimobilis B90A [23]. Dechlorination initiated by direct photolysis of short-wavelength UV irradiation [24] or by single-electron transfer via electrodes [14] may directly transform HCH isomers into a pentachlorocyclohexyl radical by cleavage of a CCl bond. Two-electron transfers to the molecule initiate dichloroelimination in a stepwise or a concerted mode [25], [26]. Transformation of HCH by dichloroelimination can be induced by Fe⁰ nanoparticles [15], [16], [17]. Similarly, anaerobic degradation of α-HCH by Clostridium pasterianum proceeds via dichloroelimination whereby two CCl bonds are reductively cleaved leading to 3,4,5,6-tetrachlorocyclohexene (TeCCH) [27].

Information on the transformation mechanism is essential for assessing the fate of HCH isomers in the environment. However, it is difficult to distinguish among different reaction mechanisms solely based on analysis of the reaction intermediates. For instance, the first step in aerobic degradation of α-HCH is dehydrochlorination, a similar mechanism compared to alkaline hydrolysis. The pattern of intermediates alone often does not allow characterizing specific degradation pathways.

Compound specific stable isotope analysis (CSIA) is a promising tool for characterizing transformation pathways of pollutants in the environment [28]. When organic compounds are degraded, reactions have slightly different rates with respect to lighter and heavier isotopes, which can lead to the change of isotope ratios in the residual fraction, known as isotope fractionation. In order to gain insight into the reaction mechanism, isotope fractionation can be quantified as the kinetic isotope effect (KIE), which refers to a ratio of the rate constant for the reaction with the light isotope over the rate constant for the reaction with the heavy isotope. KIE is considered to be a useful tool for providing information on reaction mechanisms by determining the bond changes that occur during the rate limiting step of a reaction.

CSIA has been successfully applied to aerobic and anaerobic biotransformation of α- and γ-HCH and distinct isotope fractionation patterns for carbon were observed [27], [29], [30]. α-HCH exists in two enantiomeric forms. Enantiomer specific biodegradation of α-HCH results in enrichment of one enantiomer in the non-degraded residual phase which leads to changes in the enantiomeric fractions and thus can be used as an indicator for biodegradation [23]. The extent of enantiomer specific degradation is dependent on the degradation pathway and the environmental conditions [31], [32]. In contrast, a chemical reaction is not expected to preferentially transform the (+)-enantiomer or the (−)-enantiomer [33], [34], [35], [36], [37]. Therefore, enantiomer specific analysis and CSIA can be combined as enantiomer specific isotope analysis (ESIA) as a potential tool for distinguishing between biotic and abiotic degradation pathways. However, although CSIA and ESIA for HCH biodegradation has been a subject of recent studies, information regarding isotope fractionation upon chemical degradation of HCH is limited. There is only a single study on carbon isotope fractionation for the hydrolysis of α- and γ-HCH [38].

Due to the limited knowledge on KIEs for chemical degradation of α-HCH, we determined carbon isotope fractionation for environmentally relevant transformation processes of α-HCH in order to explore the potential of CSIA and ESIA for the identification of reaction mechanisms. Different mechanisms were investigated (Scheme 1) for chemical reactions including direct photolysis, photochemical oxidation by OH radicals, alkaline hydrolysis, electrochemical reduction and reduction by Fe⁰ nanoparticles. Apparent kinetic carbon isotope effects (AKIE_C) were calculated and compared with putative KIEs for the elucidation and distinction of α-HCH reaction mechanisms. Studies on the enantioselectivity of α-HCH were also performed to evaluate if various chemical reactions can be distinguished from biotransformation.