Small and reproducible isotope effects during methylation with trimethylsulfonium hydroxide (TMSH): A convenient derivatization method for isotope analysis of negatively charged molecules.

Negatively charged analytes must be derivatized prior to gas chromatography-isotope ratio mass spectrometry (GC-IRMS), with stringent control of isotope fractionation. Current methods require offline sample preparation. This study tests for the first time trimethylsulfonium hydroxide (TMSH) as online methylation agent prior to isotope analysis, addressing the herbicides bentazone and MCPA. Fully automated derivatization was achieved in a temperature-programmable GC injector, where reactants were injected into a packed liner, solvents were removed by split flow, and subsequent flash heating triggered the derivatization, thereby transferring derivatives onto the chromatographic column. Stoichiometric addition of TMSH resulted in complete conversion giving accurate and reproducible nitrogen isotope values of bentazone. In contrast, reproducible carbon isotope analysis required TMSH in > or = 250-fold excess. Contrary to expectations, delta(13)C values became more negative at smaller TMSH excess. This indicates that elevated methyl group concentrations in the pore space of the injection liner facilitated close-to-equilibrium rather than kinetic isotope fractionation resulting in reproducible derivatization conditions. delta(13)C results under these conditions compared favorably with liquid chromatography-IRMS: low standard deviations (0.3 per thousand for GC-IRMS, 0.1 per thousand for LC-IRMS) and a comparable offset of 1 per thousand compared to elemental analyzer-IRMS demonstrate that both methods represent expedient ways for online isotope analysis of anionic target compounds

Carbon and nitrogen isotope analysis of atrazine and desethylatrazine at sub-microgram per liter concentrations in groundwater.

Environmental degradation of organic micropollutants is difficult to monitor due to their diffuse and ubiquitous input. Current approaches-concentration measurements over time, or daughter-to-parent compound ratios-may fall short, because they do not consider dilution, compound-specific sorption characteristics or alternative degradation pathways. Compound-specific isotope analysis (CSIA) offers an alternative approach based on evidence from isotope values. Until now, however, the relatively high limits for precise isotope analysis by gas chromatography-isotope ratio mass spectrometry (GC-IRMS) have impeded CSIA of sub-microgram-per-liter scale micropollutant concentrations in field samples. This study presents the first measurements of C and N isotope ratios of the herbicide atrazine and its metabolite desethylatrazine at concentrations of 100 to 1,000 ng/L in natural groundwater samples. Solid-phase extraction and preparative HPLC were tested and validated for preconcentration and cleanup of groundwater samples of up to 10 L without bias by isotope effects. Matrix interferences after solid-phase extraction could be greatly reduced by a preparative HPLC cleanup step prior to GC-IRMS analysis. Sensitivity was increased by a factor of 6 to 8 by changing the injection method from large-volume to cold-on-column injection on the GC-IRMS system. Carbon and nitrogen isotope values of field samples showed no obvious correlation with concentrations or desethylatrazine-to-atrazine ratios. Contrary to expectations, however, delta (13) C values of desethylatrazine were consistently less negative than those of atrazine from the same sites. Potentially, this line of evidence may contain information about further desethylatrazine degradation. In such a case, the common practice of using desethylatrazine-to-atrazine ratios would underestimate natural atrazine degradation

Gas chromatography/isotope ratio mass spectrometry of recalcitrant target compounds: Performance of different combustion reactors and strategies for standardization.

RATIONALE: Compound-specific isotope analysis (CSIA) relies on continuous flow combustion of organic substances to CO(2) and N(2) in a miniature reactor to measure (13)  C/(12)  C and (15)  N/(14)  N stable isotope ratios. Accurate analysis is well established for many volatile hydrocarbons. In contrast, compounds which contain hetero and halogen atoms are less volatile and may be more recalcitrant to combustion. METHODS: This study tested carbon and nitrogen isotope analysis of atrazine, desethylatrazine (DEA), dichlobenil and 2,6-dichlorobenzamide (BAM) by gas chromatography/isotope ratio mass spectrometry (GC/IRMS) with multiple reactor tubes of two different kinds (conventional CuO/NiO/Pt and a NiO tube/CuO-NiO reactor prototype). RESULTS: The advantages of the NiO tube/CuO-NiO reactor were the absence of an additional reduction reactor, the possibility of routine reoxidation in nitrogen isotope analysis, and reliable atrazine and DEA measurements over several hundred injections. In contrast, BAM analysis showed good accuracy for carbon, but notable variations in the trueness of nitrogen isotope ratios. Accurate carbon and nitrogen analysis was nevertheless possible by bracketing samples with external compound-specific standards and subsequent offset correction. CONCLUSIONS: We conclude that instrument data should never be taken at its 'face value', but must consistently be validated with compound-specific standards of the respective analytes

C and N isotope fractionation during biodegradation of the pesticide metabolite 2,6-dichlorobenzamide (BAM): Potential for environmental assessments.

2,6-Dichlorobenzamide (BAM) is a metabolite of the herbicide 2,6-dichlorobenzonitrile (dichlobenil), and a prominent groundwater contaminant. Observable compound-specific isotope fractionation during BAM formation-through transformation of dichlobenil by Rhodococcus erythropolis DSM 9685-was small. In contrast, isotope fractionation during BAM degradation-with Aminobacter sp. MSH1 and ASI1, the only known bacterial strains capable of mineralizing BAM-was large, with pronounced carbon (ε(C) = -7.5‰ to -7.8‰) and nitrogen (ε(N) = -10.7‰ to -13.5‰) isotopic enrichment factors. BAM isotope values in natural samples are therefore expected to be dominated by the effects of its degradation rather than formation. Dual isotope slopes Δ (=Δδ(15)N/Δδ(13)C ≈ ε(N)/ε(C)) showed only small differences for MSH1 (1.75 ± 0.03) and ASI1 (1.45 ± 0.03) suggesting similar transformation mechanisms of BAM hydrolysis. Observations are in agreement with either a tetrahedral intermediate promoted by OH(-) or H(3)O(+) catalysis, or a concerted reaction mechanism. Therefore, owing to consistent carbon isotopic fractionation, isotope shifts of BAM can be linked to BAM biodegradation, and may even be used to quantify degradation of this persistent metabolite. In contrast, nitrogen isotope values may be rather indicative of different sources. Our results delineate a new approach to assessing the fate of BAM in the environment