Comparison of Carbon Isotope Ratio Measurement of the Vanillin Methoxy Group by GC-IRMS and 13C-qNMR.

Site-specific carbon isotope ratio measurements by quantitative 13C NMR (13C-qNMR), Orbitrap-MS, and GC-IRMS offer a new dimension to conventional bulk carbon isotope ratio measurements used in food provenance, forensics, and a number of other applications. While the site-specific measurements of carbon isotope ratios in vanillin by 13C-qNMR or Orbitrap-MS are powerful new tools in food analysis, there are a limited number of studies regarding the validity of these measurement results. Here we present carbon site-specific measurements of vanillin by GC-IRMS and 13C-qNMR for methoxy carbon. Carbon isotope delta (δ13C) values obtained by these different measurement approaches demonstrate remarkable agreement; in five vanillin samples whose bulk δ13C values ranged from -31‰ to -26‰, their δ13C values of the methoxy carbon ranged from -62.4‰ to -30.6‰, yet the difference between the results of the two analytical approaches was within ±0.6‰. While the GC-IRMS approach afforded up to 9-fold lower uncertainties and required 100-fold less sample compared to the 13C-qNMR, the 13C-qNMR is able to assign δ13C values to all carbon atoms in the molecule, not just the cleavable methoxy group.

Sources and sinks of chloromethane in a salt marsh ecosystem: constraints from concentration and stable isotope measurements of laboratory incubation experiments.

Chloromethane (CH3Cl) is the most abundant long-lived chlorinated organic compound in the atmosphere and contributes significantly to natural stratospheric ozone depletion. Salt marsh ecosystems including halophyte plants are a known source of atmospheric CH3Cl but estimates of their total global source strength are highly uncertain and knowledge of the major production and consumption processes in the atmosphere-halophyte-soil system is yet incomplete. In this study we investigated the halophyte plant, Salicornia europaea, and soil samples from a coastal salt marsh site in Sardinia/Italy for their potential to emit and consume CH3Cl and using flux measurements, stable isotope techniques and Arrhenius plots differentiated between biotic and abiotic processes. Our laboratory approach clearly shows that at least 6 different production and consumption processes are active in controlling atmospheric CH3Cl fluxes of a salt marsh ecosystem. CH3Cl release by dried plant and soil material was substantially higher than that from the fresh material at temperatures ranging from 20 to 70 °C. Results of Arrhenius plots helped to distinguish between biotic and abiotic formation processes in plants and soils. Biotic CH3Cl consumption rates were highest at 30 °C for plants and 50 °C for soils, and microbial uptake was higher in soils with higher organic matter content. Stable isotope techniques helped to distinguish between formation and degradation processes and also provided a deeper insight into potential methyl moiety donor compounds, such as S-adenosyl-l-methionine, S-methylmethionine and pectin, that might be involved in the abiotic and biotic CH3Cl production processes. Our results clearly indicate that cycling of CH3Cl in salt marsh ecosystems is a result of several biotic and abiotic processes occurring simultaneously in the atmosphere-plant-soil system. Important precursor compounds for biotic and abiotic CH3Cl formation might be methionine derivatives and pectin. All formation and degradation processes are temperature dependent and thus environmental changes might affect the strength of each source and sink within salt marsh ecosystems and thus considerably alter total fluxes of CH3Cl from salt marsh ecosystems to the atmosphere.

Chlorine Isotope Fractionation of the Major Chloromethane Degradation Processes in the Environment.

Chloromethane (CH3Cl) is an important source of chlorine in the stratosphere, but detailed knowledge of the magnitude of its sources and sinks is missing. Here, we measured the stable chlorine isotope fractionation (εCl) associated with the major abiotic and biotic CH3Cl sinks in the environment, namely, CH3Cl degradation by hydroxyl (·OH) and chlorine (·Cl) radicals in the troposphere and by reference bacteria Methylorubrum extorquens CM4 and Leisingera methylohalidivorans MB2 from terrestrial and marine environments, respectively. No chlorine isotope fractionation was detected for reaction of CH3Cl with ·OH and ·Cl radicals, whereas a large chlorine isotope fractionation (εCl) of -10.9 ± 0.7‰ (n = 3) and -9.4 ± 0.9 (n = 3) was found for CH3Cl degradation by M. extorquens CM4 and L. methylohalidivorans MB2, respectively. The large difference in chlorine isotope fractionation observed between tropospheric and bacterial degradation of CH3Cl provides an effective isotopic tool to characterize and distinguish between major abiotic and biotic processes contributing to the CH3Cl sink in the environment. Our findings demonstrate the potential of emerging triple-element isotopic approaches including chlorine to carbon and hydrogen analysis for the assessment of global cycling of organochlorines.

Methyl sulfates as methoxy isotopic reference materials for δ13 C and δ2 H measurements.

RATIONALE: Stable hydrogen and carbon isotope ratios of methoxy groups (OCH3 ) of plant organic matter have many potential applications in biogeochemical, atmospheric and food research. So far, most of the analyses of plant methoxy groups by isotope ratio mass spectrometry have employed liquid iodomethane (CH3 I) as the reference material to normalise stable isotope measurements of these moieties to isotope-δ scales. However, comparisons of measurements of stable hydrogen and carbon isotopes of plant methoxy groups are still hindered by the lack of suitable reference materials. METHODS: We have investigated two methyl sulfate salts (HUBG1 and HUBG2), which exclusively contain carbon and hydrogen from one methoxy group, for their suitability as methoxy reference materials. Firstly, the stable hydrogen and carbon isotope values of the bulk compounds were calibrated against international reference substances by high-temperature conversion- and elemental analyser isotope ratio mass spectrometry (HTC- and EA-IRMS). In a second step these values were compared with values obtained by measurements using gas chromatography/isotope ratio mass spectrometry (GC/IRMS) where prior to analysis the methoxy groups were converted into gaseous iodomethane. RESULTS: The 2 H- and 13 C isotopic abundances of HUBG1 measured by HTC- and EA-IRMS and expressed as δ-values on the usual international scales are -144.5 ± 1.2 mUr (n = 30) and -50.31 ± 0.16 mUr (n = 14), respectively. For HUBG2 we obtained -102.0 ± 1.3 mUr (n = 32) and +1.60 ± 0.12 mUr (n = 16). Furthermore, the values obtained by GC/IRMS were in good agreement with the HTC- and EA-IRMS values. CONCLUSIONS: We suggest that both methyl sulfates are suitable reference materials for normalisation of isotope measurements of carbon of plant methoxy groups to isotope-δ scales and for inter-laboratory calibration. For stable hydrogen isotope measurements, we suggest that in addition to HUBG1 and HUBG2 additional reference materials are required to cover the full range of plant methoxy groups reported so far.

Chloromethane Degradation in Soils: A Combined Microbial and Two-Dimensional Stable Isotope Approach.

Chloromethane (CHCl, methyl chloride) is the most abundant volatile halocarbon in the atmosphere and involved in stratospheric ozone depletion. The global CHCl budget, and especially the CHCl sink from microbial degradation in soil, still involves large uncertainties. These may potentially be resolved by a combination of stable isotope analysis and bacterial diversity studies. We determined the stable isotope fractionation of CHCl hydrogen and carbon and investigated bacterial diversity during CHCl degradation in three soils with different properties (forest, grassland, and agricultural soils) and at different temperatures and headspace mixing ratios of CHCl. The extent of chloromethane degradation decreased in the order forest > grassland > agricultural soil. Rates ranged from 0.7 to 2.5 µg g dry wt. d for forest soil, from 0.1 to 0.9 µg g dry wt. d for grassland soil, and from 0.1 to 0.4 µg g dry wt. d for agricultural soil and increased with increasing temperature and CHCl supplementation. The measured mean stable hydrogen enrichment factor of CHCl of -50 ± 13‰ was unaffected by temperature, mixing ratio, or soil type. In contrast, the stable carbon enrichment factor depended on CHCl degradation rates and ranged from -38 to -11‰. Bacterial community composition correlated with soil properties was independent from CHCl degradation or isotope enrichment. Nevertheless, increased abundance after CHCl incubation was observed in 21 bacterial operational taxonomical units (OTUs at the 97% 16S RNA sequence identity level). This suggests that some of these bacterial taxa, although not previously associated with CHCl degradation, may play a role in the microbial CHCl sink in soil.

Chloromethane formation and degradation in the fern phyllosphere.

Chloromethane (CH3Cl) is the most abundant halogenated trace gas in the atmosphere. It plays an important role in natural stratospheric ozone destruction. Current estimates of the global CH3Cl budget are approximate. The strength of the CH3Cl global sink by microbial degradation in soils and plants is under discussion. Some plants, particularly ferns, have been identified as substantial emitters of CH3Cl. Their ability to degrade CH3Cl remains uncertain. In this study, we investigated the potential of leaves from 3 abundant ferns (Osmunda regalis, Cyathea cooperi, Dryopteris filix-mas) to produce and degrade CH3Cl by measuring their production and consumption rates and their stable carbon and hydrogen isotope signatures. Investigated ferns are able to degrade CH3Cl at rates from 2.1 to 17 and 0.3 to 0.9µggdw-1day-1 for C. cooperi and D. filix-mas respectively, depending on CH3Cl supplementation and temperature. The stable carbon isotope enrichment factor of remaining CH3Cl was -39±13‰, whereas negligible isotope fractionation was observed for hydrogen (-8±19‰). In contrast, O. regalis did not consume CH3Cl, but produced it at rates ranging from 0.6 to 128µggdw-1day-1, with stable isotope values of -97±8‰ for carbon and -202±10‰ for hydrogen, respectively. Even though the 3 ferns showed clearly different formation and consumption patterns, their leaf-associated bacterial diversity was not notably different. Moreover, we did not detect genes associated with the only known chloromethane utilization pathway "cmu" in the microbial phyllosphere of the investigated ferns. Our study suggests that still unknown CH3Cl biodegradation processes on plants play an important role in global cycling of atmospheric CH3Cl.

Methane oxidation in industrial biogas plants-Insights in a novel methanotrophic environment evidenced by pmoA gene analyses and stable isotope labelling studies.

A broad methanotrophic community consisting of 16 different operational taxonomic units (OTUs) was detected by particulate methane monooxygenase A (pmoA) gene analyses of reactor sludge samples obtained from an industrial biogas plant. Using a cloning-sequencing approach, 75% of the OTUs were affiliated to the group of type I methanotrophs (γ-Proteobacteria) and 25% to type II methanotrophs (α-Proteobacteria) with a distinct predominance of the genus Methylobacter. By database matching, half of the total OTUs may constitute entirely novel species. For evaluation of process conditions that support growth of methanotrophic bacteria, qPCR analyses of pmoA gene copy numbers were performed during a sampling period of 70 days at varying reactor feeding scenarios. During the investigation period, methanotrophic cell counts estimated by qPCR fluctuated between 3.4 × 104 and 2 × 105 cells/mL with no distinct correlation to the organic loading rate, the amount of CH4, O2 and NH4-N. Methanotrophic activity was proofed even at low O2 levels (1%) by using stable carbon isotope labelling experiments of CH4 in batch experiments inoculated with reactor sludge. Supplementation of 13C labelled CH4 in the headspace of the reaction vials unambiguously confirmed the formation of 13C labelled CO2. Thus, industrial biogas reactors can be considered as a further methanotrophic habitat that exhibits a unique methanotrophic community which is specifically adapted to high CH4 and low O2 concentrations. To the best of our knowledge, our study is the first accurate detection and quantification of methanotrophic bacteria in industrial biogas reactors.

Warm season precipitation signal in δ2 H values of wood lignin methoxyl groups from high elevation larch trees in Switzerland.

RATIONALE: In this study, we tested stable hydrogen isotope ratios of wood lignin methoxyl groups (δ2 Hmethoxyl values) as a palaeoclimate proxy in dendrochronology. This is a quite new method in the field of dendrochronology and the sample preparation is much simpler than the methods used before to measure δ2 H values from wood. METHODS: We measured δ2 Hmethoxyl values in high elevation larch trees (Larix decidua Mill.) from Simplon Valley (southern Switzerland). Thirty-seven larch trees were sampled and five individuals analysed for their δ2 Hmethoxyl values at annual (1971-2009) and pentadal resolution (1746-2009). The δ2 Hmethoxyl values were measured as CH3 I released upon treatment of the dried wood samples with hydroiodic acid. 10-90 µL from the head-space were injected into the gas chromatography/high-temperature conversion/isotope ratio mass spectrometry (GC/HTC-IRMS) system. RESULTS: Testing the climate response of the δ2 Hmethoxyl values, the annually resolved series show a positive correlation of r = 0.60 with June/July precipitation. The pentadally resolved δ2 Hmethoxyl series do not show any significant correlation to climate parameters. CONCLUSIONS: Increased precipitation during June and July, which are on average warm and relatively dry months, results in higher δ2 H values of the xylem water and, therefore, higher δ2 H values in the lignin methoxyl groups. Therefore, we suggest that δ2 Hmethoxyl values of high elevation larch trees might serve as a summer precipitation proxy.

Chloromethane emissions in human breath.

Chloromethane (CH3Cl), currently the most abundant chlorinated organic compound in the atmosphere at around ~550 parts per trillion by volume (pptv), is considered responsible for approximately 16% of halogen-catalyzed stratospheric ozone destruction. Although emissions of CH3Cl are known to occur from animals such as cattle, formation and release of CH3Cl from humans has not yet been reported. In this study a pre-concentration unit coupled with a gas chromatograph directly linked to a mass spectrometer was used to precisely measure concentrations of CH3Cl at the pptv level in exhaled breath from 31 human subjects with ages ranging from 3 to 87years. We provide analytical evidence that all subjects exhaled CH3Cl in the range of 2.5 to 33 parts per billion by volume, levels which significantly exceed those of inhaled air by a factor of up to 60. If the mean of these emissions was typical for the world's population, then the global source of atmospheric CH3Cl from humans would be around 0.66Ggyr-1 (0.33 to 1.48Ggyr-1), which is less than 0.03% of the total annual global atmospheric source strength. The observed endogenous formation of a chlorinated methyl group in humans might be of interest to biochemists and medical scientists as CH3Cl is also known to be a potent methylating agent and thus, could be an important target compound in future medical research diagnostic programs.

Exogenous addition of H2 for an in situ biogas upgrading through biological reduction of carbon dioxide into methane.

Biological reduction of CO2 into CH4 by exogenous addition of H2 is a promising technology for upgrading biogas into higher CH4 content. The aim of this work was to study the feasibility of exogenous H2 addition for an in situ biogas upgrading through biological conversion of the biogas CO2 into CH4. Moreover, this study employed systematic study with isotope analysis for providing comprehensive evidence on the underlying pathways of CH4 production and upstream processes. Batch reactors were inoculated with digestate originating from a full-scale biogas plant and fed once with maize leaf substrate. Periodic addition of H2 into the headspace resulted in a completely consumption of CO2 and a concomitant increase in CH4 content up to 89%. The microbial community and isotope analysis shows an enrichment of hydrogenotrophic Methanobacterium and the key role of hydrogenotrophic methanogenesis for biogas upgrading to higher CH4 content. Excess H2 was also supplied to evaluate its effect on overall process performance. The results show that excess H2 addition resulted in accumulation of H2, depletion of CO2 and inhibition of the degradation of acetate and other volatile fatty acids (VFA). A systematic isotope analysis revealed that excess H2 supply led to an increase in dissolved H2 to the level that thermodynamically inhibit the degradation of VFA and stimulate homo-acetogens for production of acetate from CO2 and H2. The inhibition was a temporary effect and acetate degradation resumed when the excess H2 was removed as well as in the presence of stoichiometric amount of H2 and CO2. This inhibition mechanism underlines the importance of carefully regulating the H2 addition rate and gas retention time to the CO2 production rate, H2-uptake rate and growth of hydrogenotrophic methanogens in order to achieve higher CH4 content without the accumulation of acetate and other VFA.

Organic compounds in fluid inclusions of Archean quartz-Analogues of prebiotic chemistry on early Earth.

The origin of life is still an unsolved mystery in science. Hypothetically, prebiotic chemistry and the formation of protocells may have evolved in the hydrothermal environment of tectonic fault zones in the upper continental crust, an environment where sensitive molecules are protected against degradation induced e.g. by UV radiation. The composition of fluid inclusions in minerals such as quartz crystals which have grown in this environment during the Archean period might provide important information about the first organic molecules formed by hydrothermal synthesis. Here we present evidence for organic compounds which were preserved in fluid inclusions of Archean quartz minerals from Western Australia. We found a variety of organic compounds such as alkanes, halocarbons, alcohols and aldehydes which unambiguously show that simple and even more complex prebiotic organic molecules have been formed by hydrothermal processes. Stable-isotope analysis confirms that the methane found in the inclusions has most likely been formed from abiotic sources by hydrothermal chemistry. Obviously, the liquid phase in the continental Archean crust provided an interesting choice of functional organic molecules. We conclude that organic substances such as these could have made an important contribution to prebiotic chemistry which might eventually have led to the formation of living cells.

Stable hydrogen isotope values of lignin methoxyl groups of four tree species across Germany and their implication for temperature reconstruction.

Stable hydrogen isotope ratios of lignin methoxyl groups (δ2HLM values) in wood have been shown to mirror the δ2H signatures of precipitation (δ2Hprecip values). Thus, δ2HLM values were suggested to serve as a potential paleotemperature proxy since δ2Hprecip values are dominantly controlled by air temperature in the mid-latitudes. A recent study where a significant δ2HLM-temperature relationship was found for a European transect with mean annual temperatures ranging from -4 to 17°C strengthened this assumption. However, using δ2HLM values as a paleotemperature proxy requires quantification of noise from site-, species- and biosynthetic-specific influences to determine the significance of recording smaller temperature changes. Here, we measured δ2HLM values of tree-ring sections covering 1981-1990 and 1991-2011 of four different tree species (European beech, English oak, Scots pine, Norway spruce) at 15 sampling sites across Germany. The maximum difference in mean annual temperature between sample sites was 5°C and all sites showed small temperature increases from 1981 to 1990 to 1991-2011 (mean Δ=0.7°C). For all species investigated, the maximum difference of δ2HLM within the tree was <10mUr or ‰ (median values) and between trees at a single site was ≤28mUr (median values). The general pattern of the spatial δ2HLM-temperature relationship found for the European transect was confirmed here although a significant correlation was lacking. This can be explained by the lower spatial δ2Hprecip-temperature correlation (R2=0.39) found for sampling sites in this study and the δ2HLM differences between trees. Nevertheless, the temporal changes in δ2HLM values of European beech trees correctly reflected within ±2°C the temperature change at every sampling site. Therefore, we suggest that δ2HLM values of European beech trees have considerable potential for reconstructing temperature changes when applied on tree-ring chronologies and consider this approach particularly suited for Late Holocene climate studies.

Mean annual temperatures of mid-latitude regions derived from δ2H values of wood lignin methoxyl groups and its implications for paleoclimate studies.

Tree-rings are widely used climate archives providing annual resolutions on centennial to millennial timescales. Stable isotope ratios of tree-rings have been applied to assist with the delineation of climate parameters such as temperature and precipitation. Here, we investigated stable hydrogen isotope ratios (expressed as δ2H values) of lignin methoxyl groups of wood from various tree species collected along a ~3500km north-south transect across Europe with mean annual temperatures (MAT) ranging from -4 to +17°C. We found a strong linear relationship between MATs and δ2H values of wood lignin methoxyl groups. We used this relationship to predict MATs from randomly collected wood samples and found general agreement between predicted and observed MATs for the mid-latitudes on a global scale. Our results are discussed in context of their paleoclimate relevance and suggest that δ2H values of lignin methoxyl groups might have the potential to reconstruct MATs when applied on mid-latitudinal tree-ring chronologies of the Late Holocene.

Stable isotope and high precision concentration measurements confirm that all humans produce and exhale methane.

Mammalian formation of methane (methanogenesis) is widely considered to occur exclusively by anaerobic microbial activity in the gastrointestinal tract. Approximately one third of humans, depending on colonization of the gut by methanogenic archaea, are considered methane producers based on the classification terminology of high and low emitters. In this study laser absorption spectroscopy was used to precisely measure concentrations and stable carbon isotope signatures of exhaled methane in breath samples from 112 volunteers with an age range from 1 to 80 years. Here we provide analytical evidence that volunteers exhaled methane levels were significantly above background (inhaled) air. Furthermore, stable carbon isotope values of the exhaled methane unambiguously confirmed that this gas was produced by all of the human subjects studied. Based on the emission and stable carbon isotope patterns of various age groups we hypothesize that next to microbial sources in the gastrointestinal tracts there might be other, as yet unidentified, processes involved in methane formation supporting the idea that humans might also produce methane endogenously in cells. Finally we suggest that stable isotope measurements of volatile organic compounds such as methane might become a useful tool in future medical research diagnostic programs.

δ²H, δ¹³C and δ¹8O from whole wood, α-cellulose and lignin methoxyl groups in Pinus sylvestris: a multi-parameter approach.

Novel tree ring parameters - δ(13)C and δ(2)H from methoxyl groups - have been developed to reconstruct palaeoclimate. Tests with δ(13)C and δ(18)O derived from whole wood and cellulose samples, however, indicated differences in the isotopic composition and climate signal, depending on the extracted wood component. We assess this signal dependency by analysing (i) δ(13)C and δ(18)O from whole wood and cellulose and (ii) δ(13)C and δ(2)H from methoxyl groups, using Pinus sylvestris L. growing near Altenkirchen (Germany). Results indicate significant correlations among the time series derived from whole wood, cellulose, and lignin methoxyl groups. Compared with the whole wood samples, δ(13)C from methoxyl groups showed a different and overall lower response to climate parameters. On the other hand, δ(2)H from methoxyl groups showed high correlations with temperature and was also correlated with ring width, indicating its potential as a temperature proxy. Isotope time series with the highest correlation with climatic parameter were: (i) whole wood and cellulose δ(13)C with growing season precipitation and summer temperature; (ii) methoxyl groups with spring precipitation; (iii) whole wood and cellulose δ(18)O correlates with annual evapotranspiration and water balance; and (iv) methoxyl group δ(2)H with spring temperatures. These findings reveal that multiple climate elements can be reconstructed from different wood components and that whole wood proxies perform comparably to cellulose time series.

A stable isotope approach to assessing water loss in fruits and vegetables during storage.

Plant tissue water is the source of oxygen and hydrogen in organic biomatter. Recently, we demonstrated that the stable hydrogen isotope value (δ(2)H) of plant methoxyl groups is a very reliable and easily available archive for the δ(2)H value of this tissue water. Here we show in a model experiment that the δ(2)H values of methoxyl groups remain unchanged after water loss during storage of fruits and vegetables under controlled conditions, while δ(2)H and δ(18)O values of tissue water increase. This enhancement is plant-dependent, and the correlation differs from the meteoric water line. The δ(18)O value is better correlated to the weight decrease of the samples. Therefore, we postulate that the δ(2)H value of methoxyl groups and the δ(18)O value of tissue water are suitable parameters for checking postharvest alterations of tissue water, either addition or loss.

Stable hydrogen and carbon isotope ratios of methoxyl groups during plant litter degradation.

Stable hydrogen and carbon isotope ratios of methoxyl groups (δ(2)Hmethoxyl and δ(13)Cmethoxyl values, respectively) in plant material have been shown to possess characteristic signatures. These isotopic signatures can be used for a variety of applications such as constraining the geographical origin and authenticity of biomaterials. Recently, it has also been suggested that δ(2)Hmethoxyl values of sedimentary organic matter of geological archives might serve as a palaeoclimate/-hydrology proxy. However, deposited organic matter is subject to both biotic and abiotic degradation processes, and therefore an evaluation of their potential impact on the δ(2)Hmethoxyl and δ(13)Cmethoxyl values would allow more reliable interpretations of both isotopic signatures. Here, we investigated this potential influence by exposing foliar litter of five different tree species (Sycamore maple, Mountain ash, European beech, Norway spruce and Scots pine) to natural degradation. The foliar litter was sampled at nine intervals over a 27-month period, and the bulk methoxyl content as well as the δ(2)Hmethoxyl and δ(13)Cmethoxyl values were measured. At the end of the experiment, a loss of the bulk methoxyl in the range of ∼40-70% was measured. Linear regression analysis showed no dependence of δ(2)Hmethoxyl values with methoxyl content for four out of five foliar litter samples studied (R(2) in the range of 0.03 and 0.36, p > .05). On the contrary, the δ(13)Cmethoxyl values showed significant linear correlations for the great majority of the foliar litter samples (R(2) in the range of 0.51 and 0.73, p < .05). The litter species with the greatest methoxyl loss (Mountain ash, Scots pine and Norway spruce) showed the strongest (13)C enrichment, by up to ∼5‰. Since δ(2)Hmethoxyl shows no systematic overall change during the course of degradation, we propose that there is considerable potential for its use as a palaeoclimate proxy for a wide range of geological archives containing, for instance, fossil wood or sedimentary organic matter. Care would need to be taken if δ(13)Cmethoxyl values of degraded organic matter are used for palaeoclimate/-environmental investigations.

Chloromethane release from carbonaceous meteorite affords new insight into Mars lander findings.

Controversy continues as to whether chloromethane (CH3Cl) detected during pyrolysis of Martian soils by the Viking and Curiosity Mars landers is indicative of organic matter indigenous to Mars. Here we demonstrate CH3Cl release (up to 8 µg/g) during low temperature (150-400°C) pyrolysis of the carbonaceous chondrite Murchison with chloride or perchlorate as chlorine source and confirm unequivocally by stable isotope analysis the extraterrestrial origin of the methyl group (δ(2)H +800 to +1100‰, δ(13)C -19.2 to +10‰,). In the terrestrial environment CH3Cl released during pyrolysis of organic matter derives from the methoxyl pool. The methoxyl pool in Murchison is consistent both in magnitude (0.044%) and isotope signature (δ(2)H +1054 ± 626‰, δ(13)C +43.2 ± 38.8‰,) with that of the CH3Cl released on pyrolysis. Thus CH3Cl emissions recorded by Mars lander experiments may be attributed to methoxyl groups in undegraded organic matter in meteoritic debris reaching the Martian surface being converted to CH3Cl with perchlorate or chloride in Martian soil. However we cannot discount emissions arising additionally from organic matter of indigenous origin. The stable isotope signatures of CH3Cl detected on Mars could potentially be utilized to determine its origin by distinguishing between terrestrial contamination, meteoritic infall and indigenous Martian sources.

Probing the diversity of chloromethane-degrading bacteria by comparative genomics and isotopic fractionation.

Chloromethane (CH3Cl) is produced on earth by a variety of abiotic and biological processes. It is the most important halogenated trace gas in the atmosphere, where it contributes to ozone destruction. Current estimates of the global CH3Cl budget are uncertain and suggest that microorganisms might play a more important role in degrading atmospheric CH3Cl than previously thought. Its degradation by bacteria has been demonstrated in marine, terrestrial, and phyllospheric environments. Improving our knowledge of these degradation processes and their magnitude is thus highly relevant for a better understanding of the global budget of CH3Cl. The cmu pathway, for chloromethane utilisation, is the only microbial pathway for CH3Cl degradation elucidated so far, and was characterized in detail in aerobic methylotrophic Alphaproteobacteria. Here, we reveal the potential of using a two-pronged approach involving a combination of comparative genomics and isotopic fractionation during CH3Cl degradation to newly address the question of the diversity of chloromethane-degrading bacteria in the environment. Analysis of available bacterial genome sequences reveals that several bacteria not yet known to degrade CH3Cl contain part or all of the complement of cmu genes required for CH3Cl degradation. These organisms, unlike bacteria shown to grow with CH3Cl using the cmu pathway, are obligate anaerobes. On the other hand, analysis of the complete genome of the chloromethane-degrading bacterium Leisingera methylohalidivorans MB2 showed that this bacterium does not contain cmu genes. Isotope fractionation experiments with L. methylohalidivorans MB2 suggest that the unknown pathway used by this bacterium for growth with CH3Cl can be differentiated from the cmu pathway. This result opens the prospect that contributions from bacteria with the cmu and Leisingera-type pathways to the atmospheric CH3Cl budget may be teased apart in the future.