Intra-individual Sequential Carbon and Oxygen Isotope Analyses of Neolithic Livestock for Assessing the Early Husbandry Strategies in the Southern Caucasus

Masato Hirose

Department of Earth and Environmental Sciences at Nagoya University

By sequential sampling and analyzing tooth enamel, we can read the seasonal variations in stable isotope ratios recorded in teeth. Seasonal variations in the stable carbon (δ13C) and oxygen (δ18O) isotopes can be used to infer what the individual consumed, where, and in what season. In other words, this method is suitable for studying the husbandry strategies from livestock remains. However, in the analysis of an isolated tooth, the term of the obtained variation would be short, and difficult to capture the feature of the variation. Analyzing two teeth per individual can provide long-term variation, although this requires the use of well-preserved archaeological materials.

Excavations at Göytepe (Photo: Dr. Seiji Kadowaki)

In our new study, we used this intra-individual sequential isotope analysis with M2 and M3 teeth per individual as far as possible to investigate early husbandry strategies in the southern Caucasus. We analyzed the mandibular tooth enamel of goats, sheep, and cattle (only isolated teeth) from the Neolithic settlements, Göytepe (ca. 5650–5460 cal BC) and Hacı Elamxanlı Tepe (ca. 5950–5800 cal BC) located in western Azerbaijan. As reference samples, we also analyzed modern goat and sheep individuals that are known to have grazed in the vicinity of the sites.

Map of the southern Caucasus, showing the location of Göytepe, Hacı Elamxanlı Tepe, and other main Neolithic sites belonging to the Shomutepe-Shulaveri culture. The locations of the obsidian sources are only those where the use of the obsidian was recognized in Göytepe materials (Nishiaki et al., 2019).

In this region, the first full-fledged Neolithic agro-pastoral economy called the Shomutepe-Shulaveri culture, emerged suddenly around 6000 cal BC. Despite their geographical closeness to each other, this is about 2000 to 3000 years later than the emergence of agro-pastoral practices in the Fertile Crescent. It is important to study the delay in this phenomenon in detail in order to understand the process of diffusion of agro-pastoral economy in human society. Therefore, in this study, we focused on how early agro-pastoral economy was practiced in this region under the peculiar environmental conditions colder in winter, adjacent to mountainous regions, that are uncommon in the Fertile Crescent. Livestock management regarding mobility and migration is a key aspect in understanding agro-pastoral societies. Therefore, we attempted to provide isotopic evidence indicative of early husbandry practices in the southern Caucasus.

Sequential stable carbon and oxygen isotope values of the Neolithic livestock from Göytepe.

The obtained data showed several different patterns that may be explained by different modes of husbandry practices:

  1. Some of the goats and sheep exhibit large amplitudes in δ13C and δ18O variations (see figure above: part a). This was interpreted as a lowland pasturing pattern because the modern goat and sheep showed the same pattern.
  2. A Neolithic goat and three Neolithic cattle samples exhibited relatively small amplitudes and/or inverse cyclical variation patterns (figure b and d). While these patterns may have been caused by multiple factors, such as drinking water and food/fodder, vertical transhumance has also been proposed to result in the similar isotope patterns (e.g., Henton et al., 2010; Tornero et al., 2016). If animals experience seasonal vertical transhumance between lowlands in winter and highlands in summer, it is expected that the δ13C fluctuation range reduces. In addition, the δ13C value of C4 plant feeders is presumed to decrease if they spend summer in highlands where C4 plants are less.
  3. Some individuals illustrated a pattern with a larger amplitude of δ18O seasonal variations but a smaller amplitude of δ13C variations (c). It is likely that some factor reduced the variation of δ13C. We proposed a possibility of the use of fodder (C3 plants) collected in a short term and given to livestock thus dampening the seasonal variation in δ13C.

The factors contributing to the patterns of isotopic variation presented in our study may not be limited to transhumance or the use of fodder but are consistent with such possibilities. To verify these hypotheses, other analytical methods, such as strontium isotope analysis, are required to specify pasturing places in different seasons on the basis of a local and regional isoscape. In any case, at least, these various sequential isotopic patterns suggest that a variety of livestock breeding strategies were already adopted by Neolithic inhabitants in the southern Caucasus. Thus, long-term sequential isotope data from plural teeth per individual would provide us more specific seasonal variation patterns.

This research derives from the Azerbaijani-Japanese Archaeological Mission directed by Prof. Yoshihiro Nishiaki (The University of Tokyo) and Dr. Farhad Guliyev (The National Academy of Science, Azerbaijan). The financial support for this study was provided by the JSPS KAKENHI (No. 17H04534), the MEXT KAKENHI (No. 16H06408 and 20H00026), and The Takanashi Foundation for Historical Science.

A consistent desire for animal proteins in the Bronze Age Xinjiang, China

Minghao Lin

Department of History, Shanghai Jiao Tong University, 200240 Shanghai, China

Location of Xinjiang (China)

The Xinjiang region is located in far northwestern China neighboring central Asia and Mongolian Steppe. Throughout antiquity it has been a nexus for exchange in ideas, technologies, livestock, or even populations. Although most of its land is covered by severe environments such as the Taklamakan Desert, diverse cultures have been recognised since the early Bronze Age. Near Eastern cultivars such as wheat and barley have been excavated and dated to by the beginning of the second millennium BC in this region. In spite of the appearance of wheat and barley at some sites, did the subsistence of ancient Xinjiang populations really rely on these novel C3 cultivars? The degree to which these Near Eastern cultivars contributed to human dietary intake and how these new food types shaped local subsistence strategies remains poorly understood.

The northern slope of the East Tian Shan Mountains in Balikun, Xinjiang (China). (Credit: Minghao Lin)

In this research, part of my Humboldt fellowship project hosted by Prof Cheryl Makarewicz at Kiel University, we conducted a regional scale meta-analyses of carbon and nitrogen isotope values measured from humans and animals including cattle, sheep/goats, and deer from sites dating from c. 2000 BC to AD 1000 within the Xinjiang region to examine the degree to which novel cultivars and animal proteins were incorporated into local subsistence system from the perspective of isotopic expression. We notice a narrow gap (1.7-2.7‰) in δ13C values but a wide space (4.5-6.5‰) in δ15N between humans and bovid samples. Meanwhile, we also record a consistent expression of high nitrogen isotope values visible in humans across time, indicating a persistent preference of the Xinjiang people for animal resources (e.g. meat, dairy proteins). This suggests novel cultivars of wheat and barley were not rapidly adopted in Xinjiang highlighting the food dispersal and acceptance across Eurasia during prehistory was a complex process.

Results of meta-analysis of stable carbon and nitrogen isotopic results from Xinjiang (China)
(Credit: Minghao Lin).

Current Methods and Best Practices in Strontium Isotope Mapping

Emily Holt, Jane A. Evans, and Richard Madgwick

Drawing on variations in bioavailable strontium in different environments to provenance biological materials has become increasingly common since its first application in archaeology almost four decades ago, and it is frequently applied to zooarchaeological materials. Provenancing biological materials, including faunal remains, using strontium isotope ratios generally requires a map of bioavailable strontium, commonly known as an isoscape, to compare results with. However, both producing the isoscape and using it to interpret results present methodological challenges that researchers must carefully consider.

To help researchers understand the complexities of strontium analysis, we reviewed current research to produce a critical synthesis and recommend best practices. We addressed sampling the archives needed to build an isoscape, applying the different mapping and modeling methods currently available, and interpreting the results of archaeological analyses against isoscapes. Our critical review is freely available to read and download from Earth-Science Reviews until May 8, 2021


Current research indicates that, while many archives can be analyzed to produce isoscapes, modern plant materials usually provide the best approximation of bioavailable strontium. Modern plants can be used alone or combined with other archives if applying a machine learning approach to mapping. In areas where erosion has significantly shifted the location of surface sediments, modern plants may not provide an appropriate archive for building an isoscape that is applicable to archaeological materials. In these areas, alternative archives such as archaeological rodent remains may be preferable. Archaeologists should also be aware that contemporary soil treatments like fertilizers may affect the bioavailable strontium in soils. Areas used for agriculture should be avoided when sampling.

Strontium pathways (adapted from Bataille et al. 2020, Fig. 1)

It is essential to collect appropriate metadata when sampling. Collecting metadata improves the legacy benefits of strontium data by making them more broadly applicable. These metadata and the results of the analyses should be archived in one of several online databases to maximize their usefulness.

Mapping and modeling

We found that domain mapping currently produces the most accurate, most interpretable isoscapes for most research questions. However, machine learning approaches are also powerful and promise to provide more accurate and geographically wide-ranging isoscapes over time. Machine learning is computationally intensive and may not currently be an option for many researchers, but will be more widely available in the future.

Strontium isoscapes that are both appropriate and sufficiently high resolution to answer specific research questions do not exist for most parts of the world. Researchers intending to incorporate strontium analysis into their research designs should expect to conduct primary sampling and analysis to create appropriate isoscapes or refine existing ones, which should themselves not be utilized uncritically.


Strontium isotope analysis is currently the best developed and understood provenance tool. However, because isotope data can only exclude options of possible origin, the development of multi-isotope methods provides the route to a more powerful future approach. Using strontium isotope analysis for provenancing is most successful when combined with other isotopes and/or trace elements as part of a likelihood approach.

Future directions

In the future, we expect that increasing amounts of primary data and the increasing application of machine learning approaches to mapping will mean that strontium analysis continues to improve as a method of provenancing.

This research was apart of the ZANBA – Zooarchaeology of the Nuragic Bronze Age, a Marie Sklodowska Curie fellowship awarded to Emily Holt.

Forest pasturing and foddering: stable isotope prespectives

The use of forests for pasturing and fodder resources remains globally an important, preventing over-grazing and providing complementary fodder in times of poor pasture. Traditionally, forests in Europe were a rich source of collected forage (leafy hay) in the form of branches and leaves. At the same time, herd animals can have a negative impact on forests, where grazing is unchecked. The first herders of Central and Northern Europe experienced a landscape ‘Bristling with forests and foul swamps’ as described by Tacitus in 98AD [1]. The impact of animals on forests has been a focus of archaeologists for over 40 years. For example, Iversen [2] highlighted the importance of cattle in the expansion of the initial Neolithic settlements in his landam theory. The elm decline was initial contributed to increased use of leafy hay as animal forage.

How can we study forest pasture and foddering via stable carbon isotopes?

The canopy effect is where plants growing under dense forest canopies will exhibit depleted δ13C values. This is due to a combination of carbon -13 depletion of atmospheric CO2 under the canopy caused by CO2 respired by decaying organic matter and low light intensity at the forest floor decreasing photosynthesis efficiency. Consequently, animals browsing and grazing under heavy forest canopies or being fed leafy hay from these environments will exhibit low carbon isotope values in their tissues. Using this principal, researchers such as Dorothee Drucker and Rhiannon Stevens have explored the use of forests by wild ruminants past and present.

Schematic of the Canopy effect on δ13C values of plants growing under different canopy densities and its relationship to different ruminant tissues (cone collagen/enamel bioapatite)

A cautionary note

The issue of equifinality can arise with the interpretation of stable isotopic values because different growing environments can produce similar effects on the stable isotopic ratios of plants. These continue up the food chain. For example, waterlogged environments can have a similar impact on δ13C values of plant communities as a dense forest canopy. Lynch and colleagues interpreted depleted δ13C values observed in British aurochs as a reflection of animals feeding on plants from waterlogged environments. Whereas, a similar study by Noe-Nygaard and colleagues of Scandinavian aurochs suggested these animals were forest dwellers. This is why it is key prior to the interpretation of stable isotopic results that robust interpretative frameworks using paleoenvironmental are created for testing hypotheses.

Independent methods for determining forest foddering

Compound-specific stable nitrogen isotope analysis of collagen amino acids provides an independent means for identifying consumption of woody plants, such as leafy hay. Developed by researchers at University of Bristol, direct evidence of the plant composition of animal fodder (woody/herbaceous) can be uncovered using the dietary β values based on δ15N CSIA of amino acids from incremental samples of dentine from cattle molars. These values represent the Δ15NGlx-Phe values of the plants at the base of the food web, using a known trophic offset of −4.0‰ between cattle and their diet. The dietary β values are then be compared with established ranges of Δ15NGlx-Phe values expected for herbaceous (−5.4±2.1‰) and woody plants (−9.3±1.6‰), based on modern references. Combining incremental analysis of enamel bioapatite and CSIA-AA of dentine of the same tooth provides a powerful method to identify forest pasturing and seasonal use of leafy-hay.

Look out for upcoming papers by myself and colleagues from University of Bristol, and European institutions from Hungary, Poland, France and Germany, discussing the role of forests in LBK cattle husbandry uncovered during the NeoMilk project (ERC-advance awarded to Prof. Richard Evershed).


1. Bogucki, P., 1988. Forest farmers and stockholders. Early agriculture and its consequences in North-Central Europe. Cambridge: Cambridge press.

2. Iversen, J., 1969. The influence of prehistoric man on vegetation, in The Neolithisation of Denmark: 150 years of debate., A. Fischer and K. Kristiansen, Editors. Sheffield Archaeological Monographs: Sheffield. p. 105-16.