Characterization of Hexachlorocyclohexane Isomer Dehydrochlorination by LinA1 and LinA2 Using Multi-element Compound-Specific Stable Isotope Analysis

Abstract

Dehydrochlorination is one of the main (thus far discovered) processes for aerobic microbial transformation of hexachlorocyclohexane (HCH) which is mainly catalyzed by LinA enzymes. In order to gain a better understanding of the reaction mechanisms, multi-element compound-specific stable isotope analysis was applied for evaluating αand γ-HCH transformations catalyzed by LinA1 and LinA2 enzymes. The isotopic fractionation (ε E ) values for particular elements of (), whereas the dual-isotope fractionation patterns were almost identical for both enantiomers (Λ C−Cl = 2.4 ± 0.4 and 2.5 ± 0.2, Λ H−C = 12.9 ± 2.4 and 14.9 ± 1.1). The ε E of γ-HCH transformation by LinA1 and LinA2 were −7.8 ± 1.0‰ and −7.5 ± 0.8‰ (ε C ), −2.7 ± 0.3‰ and −2.5 ± 0.4‰ (ε Cl ), −170 ± 25‰ and −150 ± 13‰ (ε H ), respectively. Similar Λ C−Cl values (2.7 ± 0.2 and 2.9 ± 0.2) were observed as well as similar Λ H−C values (20.1 ± 2.0 and 18.4 ± 1.9), indicating a similar reaction mechanism by both enzymes during γ-HCH transformation. This is the first data set on 3D isotope fractionation of αand γ-HCH enzymatic dehydrochlorination, which gave a more precise characterization of the bond cleavages, highlighting the potential of multi-element compound-specific stable isotope analysis to characterize different transformation processes (e.g., dehydrochlorination and reductive dehalogenation).

Uptake and Transformation of Hexachlorocyclohexane Isomers (HCHs) in Tree Growth Rings at a Contaminated Field Site

“…Dual-element isotope analysis (δ 13 C vs δ 37 Cl) was conducted to investigate the reaction pathways involved in the transformation of HCH in tree growth rings. The obtained results were then compared to previous Λ values acquired from aerobic α-HCH transformation by LinA enzymes (2.4 ± 0.4 to 2.5 ± 0.2), 44 anaerobic α-HCH transformation (1.7 ± 0.2 to 2.0 ± 0.3), 39 and aerobic α-HCH transformation in soil-plant systems (3.3 ± 0.3). 14 Among all middle ring samples, only one sample from T-1 was within the range of anaerobic microbial transformation.…”

“…The ε values for aerobic and anaerobic transformation could be used to estimate the order of transformation using the Rayleigh equation. The ε C and ε Cl values for aerobic (ranging from ε C = −1.0 ± 0.2 to −1.6 ± 0.3‰ for microbial cultures 48 44 The green area represents the Λ values obtained by the anaerobic microbial transformation of α-HCH. 44 The red area represents the Λ value obtained by the aerobic transformation of α-HCH in soil-plant systems.…”

“…The ε C and ε Cl values for aerobic (ranging from ε C = −1.0 ± 0.2 to −1.6 ± 0.3‰ for microbial cultures 48 44 The green area represents the Λ values obtained by the anaerobic microbial transformation of α-HCH. 44 The red area represents the Λ value obtained by the aerobic transformation of α-HCH in soil-plant systems. 14 The samples inside the blue cycle are the center samples of tree growth rings.…”

“…The gray line represents the correlation line between carbon and chlorine isotope fractionation of tree trunk samples (except the middle section) and the gray area represents the confidential bonds at level 95%. ± 0.3 to −2.0 ± 0.3‰ 44 for microbial cultures) were compiled to calculate the transformation extent. This method enables a rough estimation of the initial HCH uptake using the Rayleigh equation, which is described in detail in the Methods section (eq 2).…”

“…Dual-element isotope analysis (C−Cl) of α-HCH detected in different parts of tree trunks including the bark, outer, inner, and middle sections of growth rings of R. pseudoacacia. The purple area represents the Λ values obtained from the aerobic transformation of α-HCH by LinA enzymes 44. The green area represents the Λ values obtained by the anaerobic microbial transformation of α-HCH 44.…”