From the Ground Up: Global Nitrous Oxide Sources are Constrained by Stable Isotope Values

Rising concentrations of nitrous oxide (N₂O) in the atmosphere are causing widespread concern because this trace gas plays a key role in the destruction of stratospheric ozone and it is a strong greenhouse gas. The successful mitigation of N₂O emissions requires a solid understanding of the relative importance of all N₂O sources and sinks. Stable isotope ratio measurements (δ¹⁵N-N₂O and δ¹⁸O-N₂O), including the intramolecular distribution of ¹⁵N (site preference), are one way to track different sources if they are isotopically distinct. ‘Top-down’ isotope mass-balance studies have had limited success balancing the global N₂O budget thus far because the isotopic signatures of soil, freshwater, and marine sources are poorly constrained and a comprehensive analysis of global N₂O stable isotope measurements has not been done. Here we used a robust analysis of all available in situ measurements to define key global N₂O sources. We showed that the marine source is isotopically distinct from soil and freshwater N₂O (the continental source). Further, the global average source (sum of all natural and anthropogenic sources) is largely controlled by soils and freshwaters. These findings substantiate past modelling studies that relied on several assumptions about the global N₂O cycle. Finally, a two-box-model and a Bayesian isotope mixing model revealed marine and continental N₂O sources have relative contributions of 24–26% and 74–76% to the total, respectively. Further, the Bayesian modeling exercise indicated the N₂O flux from freshwaters may be much larger than currently thought.     

Tracing origins and migration of wildlife using stable isotopes: a review

To understand the ecology of migratory animals it is important to link geographic regions used by individuals including breeding, wintering, and intermediate stopover sites. Previous conventional approaches used to track animal movements have relied on extrinsic markers and typically the subsequent recovery of individuals. This approach has generally been inappropriate for most small, or non-game animals. The use of intrinsic markers such as fatty acid profiles, molecular DNA analyses, and the measurement of naturally occurring stable isotopes in animal tissues offer alternative approaches. This paper reviews the use of stable isotope analyses (primarily δ¹³C, δ¹⁵N, δ³⁴S, δD, δ⁸⁷Sr) to trace nutritional origin and migration in animals. This approach relies on the fact that foodweb isotopic signatures are reflected in the tissues of organisms and that such signatures can vary spatially based on a variety of biogeochemical processes. Organisms moving between isotopically distinct foodwebs can carry with them information on the location of previous feeding. Such an approach has been used to track animal use of inshore versus offshore, marine versus freshwater, terrestrial C₃ versus marine, terrestrial mesic versus xeric, and C₃ versus C₄ or Crassulacean acid metabolism foodwebs. More recently, the use of stable hydrogen isotope analyses (δD) to link organisms to broad geographic origin in North America is based on large-scale isotopic contours of growing-season average δD values in precipitation. This technique, especially when combined with the assay of other stable isotopes, will be extremely useful in helping to track migration and movement of a wide range of animals from insects to birds and mammals. Future research to refine our understanding of natural and anthropogenic-induced isotopic gradients in nature, and to explore the use of stable isotopes of other elements, is recommended. Received: 1 July 1998 / Accepted: 9 December 1998     

N<Subscript>2</Subscript>O production by denitrification in an urban river: evidence from isotopes,functional genes,and dissolved organic matter

Rivers are important sources of N₂O emissions into the atmosphere. Nevertheless, N₂O production processes in rivers are not well identified. We measured concentrations and isotopic ratios of N₂O, NH₄ ⁺, NO₂ ^?, and NO₃ ^? in surface water to identify the microbial processes of N₂O production along the Tama River in Japan. We also measured the functional gene abundance of nitrifiers and denitrifiers (amoA-bacteria, nirK, nirS, nosZ clade I, nosZ clade II) together with concentrations of dissolved organic carbon (DOC) and fluorescence intensities of protein and humic components of dissolved organic matter (DOM) to support the elucidation of N₂O production processes. The observed nitrogen (δ¹⁵N) and oxygen (δ¹⁸O) of N₂O were within the expected isotopic range of N₂O produced by nitrate reduction, indicating that N₂O was dominantly produced by denitrification. The positive significant correlation between N₂O_Net concentration and nirK gene abundance implied that nitrifiers and denitrifiers are contributors to N₂O production. Fluorescence intensities of protein and humic components of DOM and concentrations of DOC did not show significant correlations with N₂O concentrations, which suggests that DOC and abundance of DOM components do not control dissolved N₂O. Measurement of isotope ratios of N₂O and its substrates was found to be a useful tool to obtain evidence of denitrification as the main source of N₂O production along the Tama River.     

Do centennial tree-ring and stable isotope trends of Larix gmelinii (Rupr.) Rupr. indicate increasing water shortage in the Siberian north?

Tree-ring width of Larix gmelinii (Rupr.) Rupr., ratios of stable isotopes of C (δ¹³C) and O (δ¹⁸O) of whole wood and cellulose chronologies were obtained for the northern part of central Siberia (Tura, Russia) for the period 1864–2006. A strong decrease in the isotope ratios of O and C (after atmospheric δ¹³C corrections) and tree-ring width was observed for the period 1967–2005, while weather station data show a decrease in July precipitation, along with increasing July air temperature and vapor pressure deficit (VPD). Temperature at the end of May and the whole month of June mainly determines tree radial growth and marks the beginning of the vegetation period in this region. A positive correlation between tree-ring width and July precipitation was found for the calibration period 1929–2005. Positive significant correlations between C isotope chronologies and temperatures of June and July were found for whole wood and cellulose and negative relationships with July precipitation. These relationships are strengthened when the likely physiological response of trees to increased CO₂ is taken into account (by applying a recently developed δ¹³C correction). For the O isotope ratios, positive relationships with annual temperature, VPD of July and a negative correlation with annual precipitation were observed. The δ¹⁸O in tree rings may reflect annual rather than summer temperatures, due to the late melting of the winter snow and its contribution to the tree water supply in summer. We observed a clear change in the isotope and climate trends after the 1960s, resulting in a drastic change in the relationship between C and O isotope ratios from a negative to a positive correlation. According to isotope fractionation models, this indicates reduced stomatal conductance at a relatively constant photosynthetic rate, as a response of trees to water deficit for the last half century in this permafrost region.     

Stable isotope analysis of modern human hair collected from Asia (China,India, Mongolia,and Pakistan)

We report isotopic data (δ²H, δ¹⁸O n = 196; δ¹³C, δ¹⁵N n = 142; δ³⁴S n = 85) from human hair and drinking water (δ²H, δ¹⁸O n = 67) collected across China, India, Mongolia, and Pakistan. Hair isotope ratios reflected the large environmental isotopic gradients and dietary differences. Geographic information was recorded in H and O and to a lesser extent, S isotopes. H and O data were entered into a recently developed model describing the relationship between the H and O isotope composition of human hair and drinking water in modern USA and pre‐globalized populations. This has anthropological and forensic applications including reconstructing environment and diet in modern and ancient human hair. However, it has not been applied to a modern population outside of the USA, where we expect different diet. Relationships between H and O isotope ratios in drinking water and hair of modern human populations in Asia were different to both modern USA and pre‐globalized populations. However, the Asian dataset was closer to the modern USA than to pre‐globalized populations. Model parameters suggested slightly higher consumption of locally producedfoods in our sampled population than modern USA residents, but lower than pre‐globalized populations. The degree of in vivo amino acid synthesis was comparable to both the modern USA and pre‐globalized populations. C isotope ratios reflected the predominantly C₃‐based regional agriculture and C₄ consumption in northernChina. C, N, and S isotope ratios supported marine food consumption in some coastal locales. N isotope ratios suggested a relatively low consumption of animal‐derived products compared to western populations. Am J Phys Anthropol 2010. © 2009 Wiley‐Liss, Inc.     

In-depth analysis of N2O fluxes in tropical forest soils of the Congo Basin combining isotope and functional gene analysis

Primary tropical forests generally exhibit large gaseous nitrogen (N) losses, occurring as nitric oxide (NO), nitrous oxide (N₂O) or elemental nitrogen (N₂). The release of N₂O is of particular concern due to its high global warming potential and destruction of stratospheric ozone. Tropical forest soils are predicted to be among the largest natural sources of N₂O; however, despite being the world’s second-largest rainforest, measurements of gaseous N-losses from forest soils of the Congo Basin are scarce. In addition, long-term studies investigating N₂O fluxes from different forest ecosystem types (lowland and montane forests) are scarce. In this study we show that fluxes measured in the Congo Basin were lower than fluxes measured in the Neotropics, and in the tropical forests of Australia and South East Asia. In addition, we show that despite different climatic conditions, average annual N₂O fluxes in the Congo Basin’s lowland forests (0.97 ± 0.53 kg N ha⁻¹ year⁻¹) were comparable to those in its montane forest (0.88 ± 0.97 kg N ha⁻¹ year⁻¹). Measurements of soil pore air N₂O isotope data at multiple depths suggests that a microbial reduction of N₂O to N₂ within the soil may account for the observed low surface N₂O fluxes and low soil pore N₂O concentrations. The potential for microbial reduction is corroborated by a significant abundance and expression of the gene nosZ in soil samples from both study sites. Although isotopic and functional gene analyses indicate an enzymatic potential for complete denitrification, combined gaseous N-losses (N₂O, N₂) are unlikely to account for the missing N-sink in these forests. Other N-losses such as NO, N₂ via Feammox or hydrological particulate organic nitrogen export could play an important role in soils of the Congo Basin and should be the focus of future research.Subject terms: Microbiology, Biogeochemistry     

Isotopic signatures of N2O produced by ammonia-oxidizing archaea from soils

N₂O gas is involved in global warming and ozone depletion. The major sources of N₂O are soil microbial processes. Anthropogenic inputs into the nitrogen cycle have exacerbated these microbial processes, including nitrification. Ammonia-oxidizing archaea (AOA) are major members of the pool of soil ammonia-oxidizing microorganisms. This study investigated the isotopic signatures of N₂O produced by soil AOA and associated N₂O production processes. All five AOA strains (I.1a, I.1a-associated and I.1b clades of Thaumarchaeota) from soil produced N₂O and their yields were comparable to those of ammonia-oxidizing bacteria (AOB). The levels of site preference (SP), δ¹⁵N^bulk and δ¹⁸O -N₂O of soil AOA strains were 13–30%, −13 to −35% and 22–36%, respectively, and strains MY1–3 and other soil AOA strains had distinct isotopic signatures. A ¹⁵N-NH₄⁺-labeling experiment indicated that N₂O originated from two different production pathways (that is, ammonia oxidation and nitrifier denitrification), which suggests that the isotopic signatures of N₂O from AOA may be attributable to the relative contributions of these two processes. The highest N₂O production yield and lowest site preference of acidophilic strain CS may be related to enhanced nitrifier denitrification for detoxifying nitrite. Previously, it was not possible to detect N₂O from soil AOA because of similarities between its isotopic signatures and those from AOB. Given the predominance of AOA over AOB in most soils, a significant proportion of the total N₂O emissions from soil nitrification may be attributable to AOA.     

Variations in the natural abundance of oxygen-18 and deuterium in plant carbohydrates

Variations in the natural abundance of ¹⁸O and ²H in plant cellulose are influenced by the isotopic composition of the water directly involved in metabolism—the metabolic water fraction. The isotopic distinction between the metabolic source water and total tissue water must reflect the formation of isotopic gradients within the tissue that are influenced by the rate of water turnover, by properties of the water conducting system and by environmental conditions. It seems that the ¹⁸O abundance in the metabolic water is conserved in cellulose with a relatively constant isotope effect. The relationship of the ²H abundance between metabolic water and cellulose is more complex. Hydrogen incorporated into photosynthetic products during primary reduction steps is highly depleted in ²H. However, a large proportion of these hydrogens are subsequently replaced by exchange with water, leading to ²H enrichment during heterotrophic metabolism. Deciphering the oxygen isotope ratio of cellulose could help in providing insights into the carbon and oxygen fluxes exchanged between plants and the atmosphere. This is because the ¹⁸O abundance in cellulose records the ¹⁸O abundance in the metabolic water, which in turn, controls the oxygen isotopic signatures of the CO₂ and O₂ released by plants into the atmosphere. The hydrogen isotope effects associated with carbohydrate metabolism provide insights into the autotrophic state of a plant tissue. This is because the hydrogen isotope ratio of carbohydrates must reflect the net effects of the two opposing isotope effects associated with photosynthesis and heterotrophic metabolism.     

Emission of gaseous nitrogen oxides from an extensively managed grassland in NE Bavaria, Germany.

We analysed the stable isotope composition of emitted N₂O in a one-year field experiment (June 1998 to April 1999) in unfertilized controls, and after adding nitrogen by applying slurry or mineral N (calcium ammonium nitrate). Emitted N₂O was analysed every 2–4 weeks, with additional daily sampling for 10 days after each fertilizer application. In supplementary soil incubations, the isotopic composition of N₂O was measured under defined conditions, favouring either denitrification or nitrification. Soil incubated for 48 h under conditions favouring nitrification emitted very little N₂O (0.024 mol g_dw ^–1) and still produced N₂O from denitrification. Under denitrifying incubation conditions, much more N₂O was formed (0.91 mol g_dw ^–1 after 48 h). The isotope ratios of N₂O emitted from denitrification stabilized at ¹⁵N = –40.8 ± 5.7 and ¹⁸O = 2.7 ± 6.3. In the field experiment, the N₂O isotope data showed no clear seasonal trends or treatment effects. Annual means weighted by time and emission rate were ¹⁵N = –8.6 and ¹⁸O = 34.7 after slurry application, ¹⁵N = –4.6 and ¹⁸O = 24.0 after mineral fertilizer application and ¹⁵N = –6.4 and ¹⁸O = 35.6 in the control plots, respectively. So, in all treatments the emitted N₂O was ¹⁵N-depleted compared to ambient air N₂O (¹⁵N = 11.4 ± 11.6, ¹⁸O = 36.9 ± 10.7). Isotope analyses of the emitted N₂O under field conditions per se allowed no unequivocal identification of the main N₂O producing process. However, additional data on soil conditions and from laboratory experiments point to denitrification as the predominant N₂O source. We concluded (1) that the isotope ratios of N₂O emitted from the field soil were not only influenced by the source processes, but also by microbial reduction of N₂O to N₂ and (2) that N₂O emission rates had to exceed 3.4 mol N₂O m^–2 h^–1 to obtain reliable N₂O isotope data.     

Tracking Cats: Problems with Placing Feline Carnivores on ��18O, ��D Isoscapes

Background

Several felids are endangered and threatened by the illegal wildlife trade. Establishing geographic origin of tissues of endangered species is thus crucial for wildlife crime investigations and effective conservation strategies. As shown in other species, stable isotope analysis of hydrogen and oxygen in hair (δD_h, δ¹⁸O_h) can be used as a tool for provenance determination. However, reliably predicting the spatial distribution of δD_h and δ¹⁸O_h requires confirmation from animal tissues of known origin and a detailed understanding of the isotopic routing of dietary nutrients into felid hair.

Methodology/Findings

We used coupled δD_h and δ¹⁸O_h measurements from the North American bobcat (Lynx rufus) and puma (Puma concolor) with precipitation-based assignment isoscapes to test the feasibility of isotopic geo-location of felidae. Hairs of felid and rabbit museum specimens from 75 sites across the United States and Canada were analyzed. Bobcat and puma lacked a significant correlation between H/O isotopes in hair and local waters, and also exhibited an isotopic decoupling of δ¹⁸O_h and δD_h. Conversely, strong δD and δ¹⁸O coupling was found for key prey, eastern cottontail rabbit (Sylvilagus floridanus; hair) and white-tailed deer (Odocoileus virginianus; collagen, bone phosphate).

Conclusions/Significance

Puma and bobcat hairs do not adhere to expected pattern of H and O isotopic variation predicted by precipitation isoscapes for North America. Thus, using bulk hair, felids cannot be placed on δ¹⁸O and δD isoscapes for use in forensic investigations. The effective application of isotopes to trace the provenance of feline carnivores is likely compromised by major controls of their diet, physiology and metabolism on hair δ¹⁸O and δD related to body water budgets. Controlled feeding experiments, combined with single amino acid isotope analysis of diets and hair, are needed to reveal mechanisms and physiological traits explaining why felid hair does not follow isotopic patterns demonstrated in many other taxa.     

The relationship between N isotopic fractionation within soybean and N2 fixation during soybean development

The contribution of N₂ fixation to overall soybean N uptake has most commonly been quantified by N isotope‐based methods, which rely on isotopic differences in plant N between legumes and non‐fixing reference plants. The choice of non‐fixing reference plants is critical for the accuracy of isotope‐based methods, and mismatched reference plants remain a potential source of error. Accurate estimates of soybean N₂ fixation also require information on N isotopic fractionation within soybean. On the basis of a previous observation of a close correlation between an expression of N fractionation within soybean and the proportion of plant N derived from atmosphere (%Ndfa) determined by ¹⁵N natural abundance, this field study aimed at assessing the relationship between various expressions describing intraplant ¹⁵N or N partitioning and %Ndfa during soybean development. Starting from a late vegetative stage until beginning senescence, the N content and N isotopic composition of shoots, roots and nodules of nodulated and non‐nodulated soybeans was determined at eight different developmental stages. Regression analysis showed that %Ndfa most closely correlated with the difference in the N isotopic composition of shoot N minus that of root including nodule N, and that this relationship was similar to that obtained in a previous multi‐site field study. We therefore consider this expression to hold promise as a means of quantifying %Ndfa independent of a reference plant, which would avoid some of the external sources of error introduced by the use of reference plants in determining %Ndfa.     

Strong seasonal disequilibrium measured between the oxygen isotope signals of leaf and soil CO2 exchange

The oxygen isotope composition (δ¹⁸O) of atmospheric CO₂ is among a very limited number of tools available to constrain estimates of the biospheric gross CO₂ fluxes, photosynthesis and respiration at large scales. However, the accuracy of the partitioning strongly depends on the extent of isotopic disequilibrium between the signals carried by these two gross fluxes. Chamber‐based field measurements of total CO₂ and CO¹⁸O fluxes from foliage and soil can help evaluate and refine our models of isotopic fractionation by plants and soils and validate the extent and pattern of isotopic disequilibrium within terrestrial ecosystems. Owing to sampling limitations in the past, such measurements have been very rare and covered only a few days. In this study, we coupled automated branch and soil chambers with tuneable diode laser absorption spectroscopy techniques to continuously capture the δ¹⁸O signals of foliage and soil CO₂ exchange in a Pinus pinaster Aït forest in France. Over the growing season, we observed a seasonally persistent isotopic disequilibrium between the δ¹⁸O signatures of net CO₂ fluxes from leaves and soils, except during rain events when the isotopic imbalance became temporarily weaker. Variations in the δ¹⁸O of CO₂ exchanged between leaves, soil and the atmosphere were well explained by theory describing changes in the oxygen isotope composition of ecosystem water pools in response to changes in leaf transpiration and soil evaporation.     

A review of the calibration methods for measuring the carbon and oxygen isotopes in CO2 based on isotope ratio infrared spectroscopy北大核心CSCD

With the development of isotope ratio infrared spectroscopy (IRIS) technology, it is now possible for the in situ high temporal resolution and high precision measurement of carbon isotopic composition (d13C) and oxygen isotopic composition (d18O) of atmospheric CO2, which overcomes the low temporal resolution and labor intensive shortcoming of traditional isotope ratio mass spectrometry (IRMS). The dependence of d13C and d18O on CO2 concentration (termed as concentration dependence) and the drift due to sensitivity to changing environmental conditions (termed as instrumental drift) are the two main sources of error affecting the IRIS measurements. Therefore, it is important to obtain precise measurements by constructing a proper calibration strategy to solve the concentration dependence and instrumental drift. In this study, we briefly discussed the definition and related theoretical principle of concentration dependence, and elaborated the theoretical and empirical calibration methods of concentration dependence. Moreover, we introduced the calibration methods of instrumental drift, and reviewed the state of the art of calibration methods and its application of IRIS technology. Additionally, we briefly discussed the definition and method of data traceability to the international standard, and reviewed its application of IRIS technology. Finally, we recommend that concentration dependence is corrected by using three standards or above with known CO2 concentration and its d13C and d18O, bracketing the CO2 concentration of samples. The instrumental drift is corrected by setting appropriate calibration frequency and all dataset are traceable to the international standard. In the future, the comparative study of different IRIS instruments and calibration methods should be enhanced, and the similar methods should be used for measuring CH4, N2O and H2O isotopes by IRIS technique. The IRIS technology combined with other technology will provide a new opportunity for ecological research. © 2018 Editorial Office of Chinese Journal of Plant Ecology. All rights reserved.     

Impact of altering the water table height of an acidic fen on N2O and NO fluxes and soil concentrations

The impact of experimentally intensified summer drought and precipitation on N₂O and NO turnover and fluxes was investigated in a minerotrophic fen over a 2‐year period. On three treatment plots, drought was induced for 6 and 10 weeks by means of roofs and drainage and decreased water table levels by 0.1–0.3 m compared with three nonmanipulated control plots. When averaged over the three treatment plots, both N₂O and NO emission showed only little response to the drought. On the single plot scale, however, a clear impact of the treatment on N₂O and NO fluxes could be identified. On the plot with the weakest water table reduction hardly any response could be observed, while on the plot with the greatest drainage effect, N₂O and NO fluxes increased by 530% and 270%, respectively. Rewetting reduced NO emissions to background levels (0.05–0.15 μmol m^?2 h^?1), but heavily enhanced N₂O emission (18–36 μmol m^?2 h^?1) for several days in the plots with largest water table reduction. These peaks contributed up to 40% to the cumulative N₂O fluxes and were caused by rapid N₂O production according to isotope abundance data. According to N₂O concentrations and isotope abundance analysis N₂O was mostly produced at depths between 0.3 and 0.5 m. During water table reduction net N₂O production in 0.1 m depth steadily increased in the most effectively dried plot from 2 up to 44 pmol cm^?3 day^?1. Rewetting immediately increased net N₂O production in the topsoil of the drought plots, showing rates of 18–174 pmol cm^?3 day^?1. This study demonstrates that drought and rewetting can temporarily increase N₂O emission to levels that have to date only been reported from nutrient rich and degraded fens that have been drained for agricultural purposes.     

Fine-fraction carbonate stable isotopes as indicators of seasonal shallow mixed-layer paleohydrography

We have measured stable isotopic compositions of Miocene pelagic fine-fraction (<63 μm) carbonates from oligotrophic deep-sea sites in the Pacific and Atlantic oceans and compared them with those of coexisting foraminifers to test their utility as near sea-surface indicators. Fine-fraction carbonates (primarily polyspecific nannofossils) and surface-dwelling planktic foraminiferal (Globigerinoides) stable isotopes both have been considered to reflect surface-water hydrographic conditions. However, our results indicate that fine-fraction stable isotopes are offset from and do not correlate well with those of Globigerinoides. In contrast, stable isotopic records of the deep-dwelling planktic foraminifer Globoquadrina are in good correspondence with the fine-fraction records in terms of long-term (ca. >1 m.y.) trends and temporal variability. On the basis of a time-series hydrography and flux study site in the oligotrophic subtropical North Atlantic, we interpret the isotopic discrepancies between fine-fraction and Globigerinoides as resulting primarily from season of calcification, as well as possible vital effects. We suggest that fine-fraction stable isotope values from oligotrophic waters reflect late winter–early spring relatively cool, nutrient-rich shallow mixed-layer conditions during the time of deep mixing (i.e., spring bloom), whereas Globigerinoides stable isotope values record conditions that prevailed in the stratified surface waters in the warmer late spring–fall. This implies that paired analyses of fine-fraction and surface-dwelling planktic foraminiferal δ¹⁸O could be applied to reconstruct paleoseasonality of the open oceans. However, because the fine-fraction δ¹³C values are not representative of the annual mean surface-water δ¹³C, we recommend use of near surface-dwelling planktic foraminiferal δ¹³C as a proxy for δ¹³C of stratified surface waters that are more or less in equilibrium with the atmosphere with respect to pCO₂.     

Oxygen and carbon isotope variations in Chamelea gallina shells: Environmental influences and vital effects

Stable isotopes in mollusc shells, together with variable growth rates and other geochemical properties, can register different environmental clues, including seawater temperature, salinity and primary productivity. However, the strict biological control over the construction of biominerals exerted by many calcifying organisms can constrain the use of these organisms for paleoenvironmental reconstructions. Biologically controlled calcification is responsible for the so called vital effects that cause a departure from isotopic equilibrium during shell formation, resulting in lower shell oxygen and carbon compared to the equilibrium value. We investigated shell oxygen and carbon isotopic composition of the bivalve Chamelea gallina in six sites along with a latitudinal gradient on the Adriatic Sea (NE Mediterranean Sea). Seawater δ¹⁸O and δ¹³C_DIC varied from North to South, reflecting variations in seawater temperature, salinity, and chlorophyll concentration among sites. Shell δ¹⁸O and δ¹³C differed among sites and exhibited a wide range of values along with the ~400 km latitudinal gradient, away from isotopic equilibrium for both isotopes. These results hampered the utilization of this bivalve as a proxy for environmental reconstructions, in spite of C. gallina showing promise as a warm temperature proxy. Rigorous calibration studies with a precise insight of environment and shell growth are crucial prior to considering this bivalve as a reliable paleoclimatic archive.     

Hydrogen and Oxygen Stable Isotope Fractionation in Body Fluid Compartments of Dairy Cattle According to Season,Farm, Breed,and Reproductive Stage

Environmental temperature affects water turnover and isotope fractionation by causing water evaporation from the body in mammals. This may lead to rearrangement of the water stable isotope equilibrium in body fluids. We propose an approach to detect possible variations in the isotope ratio in different body fluids on the basis of different homoeothermic adaptations in varying reproductive stages. Three different reproductive stages (pregnant heifer, primiparous lactating cow, and pluriparous lactating cow) of two dairy cattle breeds (Italian Friesian and Modenese) were studied in winter and summer. Blood plasma, urine, faecal water, and milk were sampled and the isotope ratios of H (²H/¹H) and O (¹⁸O/¹⁶O) were determined. Deuterium excess and isotope-fractionation factors were calculated for each passage from plasma to faeces, urine and milk. The effects of the season, reproductive stages and breed on δ²H and δ¹⁸O were significant in all the fluids, with few exceptions. Deuterium excess was affected by season in all the analysed fluids. The correlations between water isotope measurements in bovine body fluids ranged between 0.6936 (urine-milk) and 0.7848 (urine-plasma) for δ²H, and between 0.8705 (urine-milk) and 0.9602 (plasma-milk) for δ¹⁸O. The increase in both isotopic δ values in all body fluids during summer is representative of a condition in which fractionation took place as a consequence of a different ratio between ingested and excreted water, which leads to an increased presence of the heavy isotopes. The different body water turnover between adult lactating cattle and non-lactating heifers was confirmed by the higher isotopic δ for the latter, with a shift in the isotopic equilibrium towards values more distant from those of drinking water.     

Quantifying Inter-Laboratory Variability in Stable Isotope Analysis of Ancient Skeletal Remains

Over the past forty years, stable isotope analysis of bone (and tooth) collagen and hydroxyapatite has become a mainstay of archaeological and paleoanthropological reconstructions of paleodiet and paleoenvironment. Despite this method''s frequent use across anthropological subdisciplines (and beyond), the present work represents the first attempt at gauging the effects of inter-laboratory variability engendered by differences in a) sample preparation, and b) analysis (instrumentation, working standards, and data calibration). Replicate analyses of a ¹⁴C-dated ancient human bone by twenty-one archaeological and paleoecological stable isotope laboratories revealed significant inter-laboratory isotopic variation for both collagen and carbonate. For bone collagen, we found a sizeable range of 1.8‰ for δ¹³C_col and 1.9‰ for δ¹⁵N_col among laboratories, but an interpretatively insignificant average pairwise difference of 0.2‰ and 0.4‰ for δ¹³C_col and δ¹⁵N_col respectively. For bone hydroxyapatite the observed range increased to a troublingly large 3.5‰ for δ¹³C_ap and 6.7‰ for δ¹⁸O_ap, with average pairwise differences of 0.6‰ for δ¹³C_ap and a disquieting 2.0‰ for δ¹⁸O_ap. In order to assess the effects of preparation versus analysis on isotopic variability among laboratories, a subset of the samples prepared by the participating laboratories were analyzed a second time on the same instrument. Based on this duplicate analysis, it was determined that roughly half of the isotopic variability among laboratories could be attributed to differences in sample preparation, with the other half resulting from differences in analysis (instrumentation, working standards, and data calibration). These findings have serious implications for choices made in the preparation and extraction of target biomolecules, the comparison of results obtained from different laboratories, and the interpretation of small differences in bone collagen and hydroxyapatite isotope values. To address the issues arising from inter-laboratory comparisons, we devise a novel measure we term the Minimum Meaningful Difference (MMD), and demonstrate its application.     

Effects of growth and tissue type on the kinetics of <Superscript>13</Superscript>C and <Superscript>15</Superscript>N incorporation in a rapidly growing ectotherm

The use of stable isotopes to investigate animal diets, habitat use, and trophic level requires understanding the rate at which animals incorporate the ¹³C and ¹⁵N from their diets and the factors that determine the magnitude of the difference in isotopic composition between the animal’s diet and that of its tissues. We determined the contribution of growth and catabolic turnover to the rate of ¹³C and ¹⁵N incorporation into several tissues that can be sampled non-invasively (skin, scute, whole blood, red blood cells, and plasma solutes) in two age classes of a rapidly growing ectotherm (loggerhead turtles, Caretta caretta). We found significant differences in C and N incorporation rates and isotopic discrimination factors (Δ¹³C = δ¹³C_tissues − δ¹³C_diet and Δ¹⁵N = δ¹⁵N_tissues − δ¹⁵N_diet) among tissues and between age classes. Growth explained from 26 to 100% of the total rate of incorporation in hatchling turtles and from 15 to 52% of the total rate of incorporation in juvenile turtles. Because growth contributed significantly to the rate of isotopic incorporation, variation in rates among tissues was lower than reported in previous studies. The contribution of growth can homogenize the rate of isotopic incorporation and limit the application of stable isotopes to identify dietary changes at contrasting time scales and to determine the timing of diet shifts. The isotopic discrimination factor of nitrogen ranged from −0.64 to 1.77‰ in the turtles’ tissues. These values are lower than the commonly assumed average 3.4‰ discrimination factors reported for whole body and muscle isotopic analyses. The increasing reliance on non-invasive and non-destructive sampling in animal isotopic ecology requires that we recognize and understand why different tissues differ in isotopic discrimination factors.     

Oxygen-18 Content of Atmospheric Oxygen Does Not Affect the Oxygen Isotope Relationship between Environmental Water and Cellulose in a Submerged Aquatic Plant, Egeria densa Planch

We determined that the oxygen isotopic composition of cellulose synthesized by a submerged plant, Egeria densa Planch., is related to the isotopic composition of environmental water by a linear function, δ¹⁸O cellulose = 0.48 δ¹⁸O water + 24.1%‰. The observation of a slope of less than 1 indicates that a portion of cellulose oxygen is derived from an isotopically constant source other than water. We tested whether this source might be molecular oxygen by growing plants in the presence of high concentrations of ¹⁸O in the form of O₂ bubbled into the bottom of an aquarium. Cellulose synthesized during this experiment did not have significantly different oxygen isotope ratios than that synthesized by control plants exposed to O₂ of normal ¹⁸O abundance. We propose that oxygen in organic matter recycled from senescent portions of the plant is incorporated into cellulose. Our findings indicate that paleoclimatic models linking the oxygen isotope composition of environmental water to cellulose from fossil plants will have to be modified to account for contributions of oxygen from this or other sources besides water.