Low 13C-13C abundances in abiotic ethane.

Distinguishing biotic compounds from abiotic ones is important in resource geology, biogeochemistry, and the search for life in the universe. Stable isotopes have traditionally been used to discriminate the origins of organic materials, with particular focus on hydrocarbons. However, despite extensive efforts, unequivocal distinction of abiotic hydrocarbons remains challenging. Recent development of clumped-isotope analysis provides more robust information because it is independent of the stable isotopic composition of the starting material. Here, we report data from a 13C-13C clumped-isotope analysis of ethane and demonstrate that the abiotically-synthesized ethane shows distinctively low 13C-13C abundances compared to thermogenic ethane. A collision frequency model predicts the observed low 13C-13C abundances (anti-clumping) in ethane produced from methyl radical recombination. In contrast, thermogenic ethane presumably exhibits near stochastic 13C-13C distribution inherited from the biological precursor, which undergoes C-C bond cleavage/recombination during metabolism. Further, we find an exceptionally high 13C-13C signature in ethane remaining after microbial oxidation. In summary, the approach distinguishes between thermogenic, microbially altered, and abiotic hydrocarbons. The 13C-13C signature can provide an important step forward for discrimination of the origin of organic molecules on Earth and in extra-terrestrial environments.

Hydrocarbon Cycling in the Tokamachi Mud Volcano (Japan): Insights from Isotopologue and Metataxonomic Analyses.

Understanding hydrocarbon cycling in the subsurface is important in various disciplines including climate science, energy resources and astrobiology. Mud volcanoes provide insights into biogeochemical processes occurring in the subsurface. They are usually associated with natural gas reservoirs consisting mainly of methane and other hydrocarbons as well as CO2. Stable isotopes have been used to decipher the sources and sinks of hydrocarbons in the subsurface, although the interpretation can be ambiguous due to the numerous processes involved. Here we report new data for hydrocarbon isotope analysis, including position-specific isotope composition of propane, for samples from the Tokamachi mud volcano area, Japan. The data suggest that C2+ hydrocarbons are being biodegraded, with indirect production of methane ("secondary methanogenesis"). Data from chemical and isotopic composition are discussed with regard to 16S rRNA analysis, which exhibits the presence of hydrogenotrophic and acetoclastic methoanogens. Overall, the combination of isotopologue analysis with 16S rRNA gene data allows refining of our understanding of hydrocarbon cycling in subsurface environments.

Standardization for 13 C-13 C clumped isotope analysis by the fluorination method.

RATIONALE: The 13 C-13 C isotopologues of C2 molecules have recently been measured using a fluorination method. The C2 compound is first fluorinated into hexafluoroethane (C2 F6 ), and its 13 C-isotopologues are subsequently measured using a conventional isotope ratio mass spectrometer. Here, we present an approach for standardizing the fluorination method on an absolute reference scale by using isotopically enriched C2 F6 . METHODS: We prepared physical mixtures of 13 C-13 C-labeled ethanol and natural ethanol. The enriched ethanol samples were measured using the recently developed fluorination method. Based on the difference between the calculated and measured ∆13 C13 C values, we quantified the extent to which isotopologues were scrambled during dehydration, fluorination, and ionization in the ion source. RESULTS: The measured ∆13 C13 C value was approximately 20% lower than that expected from the amount of 13 C-13 C ethanol. The potential scrambling in the ion source was estimated to be 0.5-2%, which is lower than the observed isotopic reordering. Therefore, isotopic reordering may have occurred during either dehydration or fluorination. CONCLUSIONS: For typical analysis of natural samples, scrambling in the ion source can only change the ∆13 C13 C value by less than 0.04‰, which is lower than the current analytical precision (±0.07‰). Therefore, the observed isotopic reordering may have occurred during the fluorination of ethene through the scrambling of isotopologues of ethene but not in the ion source of the mass spectrometer or during the dehydration of ethanol, given the small amount of C1 and C3+ molecules. Thus, we obtained the empirical transfer function ∆13 C13 CCSC = λ × ∆13 C13 C with a λ value of 1.25 ± 0.01 for ethanol/ethene and 1.00 for ethane. Using the empirical transfer function, the developed fluorination method can provide actual differences in ∆ values.

A fluorination method for measuring the 13 C-13 C isotopologue of C2 molecules.

RATIONALE: Doubly substituted isotope species ("clumped" isotopes) can provide insights into the biogeochemical history of a molecule, including its temperature of formation and/or its (bio)synthetic pathway. Here, we propose a new fluorination method for the measurement of 13 C-13 C species in C2 molecules using a conventional isotope ratio mass spectrometer. Target molecules include ethane, ethene and ethanol. METHODS: 13 C-13 C isotope species in C2 molecules were measured as C2 F6 using a conventional isotope ratio mass spectrometer. Ethane and ethene are directly fluorinated to C2 F6 . Ethanol is measured after dehydration to ethene and subsequent fluorination of the latter. The method enables the measurement of the Δ13 C13 C values normalized against a reference working standard. RESULTS: The reproducibility of the whole protocol, including chemical modification steps and measurement of C2 F6 isotopologues, is better than ±0.14‰ for all the compounds. Ethane from natural gas samples and biologically derived ethanol show a narrow range of Δ13 C13 C values, varying from 0.72‰ to 0.90‰. In contrast, synthetic ethanol as well as putative abiotic ethane show Δ13 C13 C values significantly different from this range with values of 1.14‰ and 0.25‰, respectively. CONCLUSIONS: The method presented here provides alternative means of measuring 13 C-13 C species to that using high-resolution mass spectrometry. Preliminary data from natural and synthetic molecules re-emphasizes the potential of 13 C clumped isotope species as a (bio)marker.