Simple extraction methods for pesticide compound-specific isotope analysis from environmental samples.

Compound-specific isotope analysis (CSIA) is a powerful approach to evaluate the transformation of organic pollutants in the environment. However, the application of CSIA to micropollutants, such as pesticides, remains limited because appropriate extraction methods are currently lacking. Such methods should address a wide range of pesticides and environmental matrices, while recovering sufficient mass for reliable CSIA without inducing stable isotope fractionation. Here, we present simple extraction methods for carbon and nitrogen CSIA for different environmental matrices and six commonly used herbicides, i.e., atrazine, terbutryn, acetochlor, alachlor, butachlor, and S-metolachlor, and three fungicides, i.e., dimethomorph, tebuconazole, and metalaxyl. We examined the potential of several extraction methods for four types of soils or sediments, three types of environmental waters and aerial and root plant samples for multielement (ME)-CSIA.•Pesticide extraction recoveries varied depending on the physical characteristics of the pesticides and matrix properties for environmental water (77 to 87%), soil and sediment (35 to 82%), and plant (40 to 59%) extraction.•The tested extraction methods did not significantly affect the carbon and nitrogen stable isotope signatures of pesticides (Δ(13C) <0.9‰ for Δ(15N) <1.0‰).

Do rainfall characteristics affect the export of copper, zinc and synthetic pesticides in surface runoff from headwater catchments?

Rainfall and runoff characteristics may influence off-site export of pesticides into downstream aquatic ecosystems. However, the relationship between rainfall characteristics and pesticide export from small headwater catchments remains elusive due to confounding factors including the application dose and timing and the variation of pesticide stocks in soil. Here we examined the impact of rainfall characteristics on the export of copper (Cu), zinc (Zn) and 12 legacy and currently used synthetic pesticides in surface runoff from a headwater vineyard catchment. Cluster analysis of rainfall intensity, depth and duration of 78 events revealed four distinct rainfall categories, i.e., Small, Long, Moderate and Intense (p < 0.001). Event mean concentrations of pesticides did not differ among rainfall categories (p > 0.05). In contrast, event loads of both dissolved and solid-bound Cu and Zn significantly differed among rainfall categories (p < 0.001). Rainfall depth and intensity significantly correlated with both Cu and Zn loads in runoff (ρs = 0.33 to 0.92, p < 0.002), and might be the main drivers of Cu and Zn export at the catchment scale. In contrast, rainfall depth, intensity or duration did not influence the loads of synthetic pesticides in runoff, even when weekly variations of pesticide stocks in the soil were accounted for. However, intense rainfall-runoff events, that can fragment soil, may control the export of persistent and hydrophobic legacy pesticides stocks in the soil, such as simazine and tetraconazole. Our results show that rainfall characteristics controlled the off-site export of Cu, Zn and legacy synthetic pesticides in a small headwater catchment, whereas the application timing drove the export of currently used synthetic pesticides in runoff. We anticipate our results to be a preliminary step to forecast the influence of regional rainfall patterns on the export of both metallic and synthetic pesticides by surface runoff from small agricultural headwater catchments.

Dissipation of S-metolachlor and butachlor in agricultural soils and responses of bacterial communities: Insights from compound-specific isotope and biomolecular analyses.

The soil dissipation of the widely used herbicides S-metolachlor (SM) and butachlor (BUT) was evaluated in laboratory microcosms at two environmentally relevant doses (15 and 150 µg/g) and for two agricultural soils (crop and paddy). Over 80% of SM and BUT were dissipated within 60 and 30 days, respectively, except in experiments with crop soil at 150 µg/g. Based on compound-specific isotope analysis (CSIA) and observed dissipation, biodegradation was the main process responsible for the observed decrease of SM and BUT in the paddy soil. For SM, biodegradation dominated over other dissipation processes, with changes of carbon isotope ratios (Δδ13C) of up to 6.5‰ after 60 days, and concomitant production of ethane sulfonic acid (ESA) and oxanilic acid (OXA) transformation products. In crop soil experiments, biodegradation of SM occurred to a lesser extent than in paddy soil, and sorption was the main driver of apparent BUT dissipation. Sequencing of the 16S rRNA gene showed that soil type and duration of herbicide exposure were the main determinants of bacterial community variation. In contrast, herbicide identity and spiking dose had no significant effect. In paddy soil experiments, a high (4:1, V/V) ESA to OXA ratio for SM was observed, and phylotypes assigned to anaerobic Clostridiales and sulfur reducers such as Desulfuromonadales and Syntrophobacterales were dominant for both herbicides. Crop soil microcosms, in contrast, were associated with a reverse, low (1:3, V/V) ratio of ESA to OXA for SM, and Alphaproteobacteria, Actinobacteria, and Bacillales dominated regardless of the herbicide. Our results emphasize the variability in the extent and modes of SM and BUT dissipation in agricultural soils, and in associated changes in bacterial communities.

Impact of rainfall patterns and frequency on the export of pesticides and heavy-metals from agricultural soils.

The combined influence of soil characteristics, pollutant aging and rainfall patterns on the export of pollutants from topsoils is poorly understood. We used laboratory experiments and parsimonious modeling to evaluate the impact of rainfall characteristics on the ponding and the leaching of a pollutant mixture from topsoils. The mixture included the fungicide metalaxyl, the herbicide S-metolachlor, as well as copper (Cu) and zinc (Zn). Four rainfall patterns, which differed in their durations and intensities, were applied twice successively with a 7days interval on each soil type. To evaluate the influence of soil type and aging, experiments included crop and vineyard soils and two stages of pollutant aging (0 and 10days). The global export of pollutants was significantly controlled by the rainfall duration and frequency (P<0.01). During the first rainfall event, the longest and most intense rainfall pattern yielded the largest export of metalaxyl (44.5±21.5% of the initial mass spiked in the soils), S-metolachlor (8.1±3.1%) and Cu (3.1±0.3%). Soil compaction caused by the first rainfall reduced in the second rainfall the leaching of remaining metalaxyl, S-metolachlor, Cu and Zn by 2.4-, 2.9-, 30- and 50-fold, respectively. In contrast, soil characteristics and aging had less influence on pollutant mass export. The soil type significantly influenced the leaching of Zn, while short-term aging impacted Cu leaching. Our results suggest that rainfall characteristics predominantly control export patterns of metalaxyl and S-metolachlor, in particular when the aging period is short. We anticipate our study to be a starting point for more systematic evaluation of the dissolved pollutant ponding/leaching partitioning and the export of pollutant mixtures from different soil types in relation to rainfall patterns.

Carbon and chlorine isotope fractionation during microbial degradation of tetra- and trichloroethene.

Two-dimensional compound-specific isotope analysis (2D-CSIA), combining stable carbon and chlorine isotopes, holds potential for monitoring of natural attenuation of chlorinated ethenes (CEs) in contaminated soil and groundwater. However, interpretation of 2D-CSIA data sets is challenged by a shortage of experimental Cl isotope enrichment factors. Here, isotope enrichments factors for C and Cl (i.e., εC and εCl) were determined for biodegradation of tetrachloroethene (PCE) and trichloroethene (TCE) using microbial enrichment cultures from a heavily CE-contaminated aquifer. The obtained values were εC = -5.6 ± 0.7‰ (95% CI) and εCl = -2.0 ± 0.5‰ for PCE degradation and εC = -8.8 ± 0.2‰ and εCl = -3.5 ± 0.5‰ for TCE degradation. Combining the values for both εC and εCl yielded mechanism-diagnostic εCl/εC ratios of 0.35 ± 0.11 and 0.37 ± 0.11 for the degradation of PCE and TCE, respectively. Application of the obtained εC and εCl values to a previously investigated field site gave similar estimates for the fraction of degraded contaminant as in the previous study, but with a reduced uncertainty in assessment of the natural attenuation. Furthermore, 16S rRNA gene clone library analyses were performed on three samples from the PCE degradation experiments. A species closely related to Desulfitobacterium aromaticivorans UKTL dominated the reductive dechlorination process. This study contributes to the development of 2D-CSIA as a tool for evaluating remediation strategies of CEs at contaminated sites.

Dual carbon-chlorine stable isotope investigation of sources and fate of chlorinated ethenes in contaminated groundwater.

Chlorinated ethenes (CEs) are ubiquitous groundwater contaminants, yet there remains a need for a method to efficiently monitor their in situ degradation. We report here the first field application of combined stable carbon and chlorine isotope analysis of tetrachloroethene (PCE) and trichloroethene (TCE) to investigate their biodegradation in a heavily contaminated aquifer. The two-dimensional Compound Specific Isotope Analysis (2D-CSIA) approach was facilitated by a recently developed gas chromatography-quadrupole mass spectrometry (GCqMS) method for δ(37)Cl determination. Both C and Cl isotopes showed evidence of ongoing PCE transformation. Applying published C isotope enrichment factors (ε(C)) enabled evaluation of the extent of in situ PCE degradation (11-78%). We interpreted C and Cl isotopes using a numerical reactive transport model along a 60-m flow path. It revealed that combined PCE and TCE mass load was dechlorinated by less than 10%, and that cis-dichloroethene was not further dechlorinated. Furthermore, the 2D-CSIA approach allowed estimation of Cl isotope enrichment factors ε(Cl) (-7.8 to -0.8‰) and characteristic ε(Cl)/ε(C) values (0.42-1.12) for reductive PCE dechlorination at this field site. This investigation demonstrates the benefit of 2D-CSIA to assess in situ degradation of CEs and the applicability of Cl isotope fractionation to evaluate PCE and TCE dechlorination.

Stereochemical and conformational exchanges in N,N'-di(2-pyridyl)formamidines: An X-ray and (1)H NMR study.

The solid state structure of N,N'-di(2-pyridyl)formamidine displays a four-hydrogen-bonded dimer. In solution, two isomers are observed, one of which is selected and amplified either by crystallization or by adding protons. Solution state analysis of N,N'-di(2-pyridyl)formamidines reveals the presence of the uncommon Z formamidine isomer, which equilibrates with the E-isomer with an activation energy of 90 kJ mol(-1) in CDCl(3).

Protease-catalysed coupling of N-protected amino acids and peptides with 4-aminoantipyrine.

The enzymatic synthesis of N-protected L-aminoacyl- and L-peptidyl-antipyrine amides was accomplished by proteases from different classes. Serine and cysteine proteases proved to be suitable tools for the production of amino acids and peptides conjugated to 4-aminoantipyrine, whereas metalloproteases do not seem to be very qualified for accepting this nucleophile. The product yields were optimised by applying ample opportunities of medium engineering, e.g. aqueous-organic, biphasic, suspension and solid-to-solid reaction systems. Thus, yields up to 100% could be obtained. The products were purified and characterised by polarimetry and NMR spectroscopy. These results broaden the common knowledge of the catalytic potential of proteases, in particular with regard to the suitability of a special heterocyclic 1,2-amino ketone as a nucleophile for the biocatalytic amidation of amino acids and peptides.