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Variability of oxygen isotopes in environmental water is recorded in tooth enamel, providing a record of seasonal change, dietary variability, and mobility. Physiology dampens this variability, however, as oxygen passes from environmental sources into blood and forming teeth. We showcase two methods of high resolution, 2-dimensional enamel sampling, and conduct modeling, to report why and how environmental oxygen isotope variability is reduced in animal bodies and teeth. First, using two modern experimental sheep, we introduce a sampling method, die-saw dicing, that provides high-resolution physical samples (n = 109 and 111 sample locations per tooth) for use in conventional stable isotope and molecular measurement protocols. Second, we use an ion microprobe to sample innermost enamel in an experimental sheep (n = 156 measurements), and in a Pleistocene orangutan (n = 176 measurements). Synchrotron and conventional μCT scans reveal innermost enamel thicknesses averaging 18 and 21 μm in width. Experimental data in sheep show that compared to drinking water, oxygen isotope variability in blood is reduced to 70–90 %; inner and innermost enamel retain between 36 and 48 % of likely drinking water stable isotope range, but this recovery declines to 28–34 % in outer enamel. 2D isotope sampling suggests that declines in isotopic variability, and shifted isotopic oscillations throughout enamel, result from the angle of secretory hydroxyapatite deposition and its overprinting by maturation. This overprinting occurs at all locations including innermost enamel, and is greatest in outer enamel. These findings confirm that all regions of enamel undergo maturation to varying degrees and confirm that inner and innermost enamel preserve more environmental variability than other regions. We further show how the resolution of isotope sampling — not only the spatial resolution within teeth, but also the temporal resolution of water in the environment — impacts our estimate of how much variation teeth recover from the environment. We suggest inverse methods, or multiplication by standard factors determined by ecology, taxon, and sampling strategy, to reconstruct the full scale of seasonal environmental variability. We advocate for combined inverse modeling and high-resolution sampling informed by the spatiotemporal pattern of enamel formation, and at the inner or innermost enamel when possible, to recover seasonal records from teeth. 

Clumped isotope studies on CO₂, Δ₄₇, that is the excess in the isotopologue containing both¹³C and¹⁸O at mass 47, can be very useful since they can give temperature estimates independent of the bulk isotopic composition. The measurement itself can be affected by a number of items. Here we develop a data processing model to examine the effects different interferences might have on the final calculated value. It incorporates known issues, for example, pressure baseline,¹⁷O excess, and shifts in absolute ratios for primary reference materials and parameters used for¹⁷O correction. We also included linearity effects as well as differences in isotopologue absolute abundances at a givenm/z. What normally would be considered acceptable mass spectrometer⁴⁵Rand⁴⁶Rlinearity can skew Δ₄₇results. That is 0.04‰/V and −0.04‰/V linearity on⁴⁵Rand⁴⁶Rrespectively would also cause an apparent shift in the parameters used for¹⁷O corrections. Measurements were made on CO₂(g) equilibrated with water, and we were able to match up the effects seen with model results. Linearity and small differences in amplitude between sample and working reference gas affected Δ₄₇determination, as did apparent shifts in isotopologue abundances under different conditions. This may (partially) account for discrepancies seen in Δ₄₇‐temperature calibrations curves between laboratories. We also developed an easy way to precisely calculate the δ¹³C and δ¹⁸O that works well in spreadsheets without the need for multiple iterations.

 

Blood water oxygen isotope compositions can provide valuable insights into physiological processes and ecological patterns. While blood samples are commonly drawn for medical or scientific purposes, blood fractions are infrequently measured for oxygen isotopic compositions (δ¹⁸O) because such measurements are time consuming and expensive.

We sampled blood from sheep, goats, and iguanas raised in field and animal laboratories into serum, EDTA, heparin, and uncoated plastic vials commonly used in medical and scientific research, then separated red blood cell (RBC) and plasma or serum blood fractions. These were injected into helium‐flushed Exetainer tubes where they naturally outgassed endogenous CO₂(goat blood), or into He‐ and CO₂‐flushed tubes (iguana blood). The CO₂gas was sampled on a GasBench II system, and δ¹⁸O was measured by an isotope ratio mass spectrometer (IRMS).

Repeated δ¹⁸O measurements were stable over multiple days. The addition of desiccated blood solids to water standards had little impact on their δ¹⁸O measurements, suggesting that organic molecular constituents within blood serum and plasma do not interfere with blood water δ¹⁸O values. We observed slight but statistically significant δ¹⁸O offsets between plasma, serum and RBC fractions. Mass‐dependent body water turnover times for iguanas were derived from the data.

We demonstrate that a simple blood‐CO₂equilibration method using the GasBench can quickly, reliably and accurately characterize water δ¹⁸O in the plasma, RBC, and whole blood fractions of mammalian and reptilian blood samples (precision ≤ 0.1‰). This method will expand the application of blood stable isotope analysis in physiological and medical research.

 

Many explanations for Eocene climate change focus on the Southern Ocean—where tectonics influenced oceanic gateways, ocean circulation reduced heat transport, and greenhouse gas declines prompted glaciation. To date, few studies focus on marine vertebrates at high latitudes to discern paleoecological and paleoenvironmental impacts of this climate transition. The Tertiary Eocene La Meseta (TELM) Formation has a rich fossil assemblage to characterize these impacts;Striatolamia macrota, an extinct (†) sand tiger shark, is abundant throughout the La Meseta Formation. Body size is often tracked to characterize and integrate across multiple ecological dimensions. †S. macrotabody size distributions indicate limited changes during TELMs 2–5 based on anterior tooth crown height (n = 450, mean = 19.6 ± 6.4 mm). Similarly, environmental conditions remained stable through this period based on δ¹⁸O_PO4values from tooth enameloid (n = 42; 21.5 ± 1.6‰), which corresponds to a mean temperature of 22.0 ± 4.0°C. Our preliminaryε_Nd(n = 4) results indicate an early Drake Passage opening with Pacific inputs during TELM 2–3 (45–43 Ma) based on single unit variation with an overall radiogenic trend. Two possible hypotheses to explain these observations are (1) †S. macrotamodified its migration behavior to ameliorate environmental changes related to the Drake Passage opening, or (2) the local climate change was small and gateway opening had little impact. While we cannot rule out an ecological explanation, a comparison with climate model results suggests that increased CO₂produces warm conditions that also parsimoniously explain the observations.