Biogeologie

Methodological approaches

In a nutshell

Our research combines classical palaeontological and archaeological approaches with modern geochemical and microscopic methods to reconstruct past terrestrial ecosystems and their dynamics. We apply taxonomic and comparative anatomical analyses to identify vertebrate fossils and to assess their evolutionary and ecological significance. Fossil surfaces are examined for bioerosion and taphonomic alteration, particularly in Mesozoic, Paleogene, and Neogene contexts, providing insights into depositional environments, post-depositional processes, and preservation pathways. To reconstruct past environments and vegetation, we integrate palaeobotanical methods, including pollen analysis, microfossil studies, and sediment-based proxies. These data are complemented by microscopic and SEM analyses, enabling detailed investigation of plant remains, sediments, and micro-scale preservation features. Geochemical approaches, especially stable isotope analyses of collagen, tooth enamel, and individual amino acids, are used to quantify diet, trophic relationships, mobility, and climatic conditions. Resulting datasets are analysed using Bayesian statistical frameworks and trophic niche modelling, allowing robust ecological interpretations even for complex or limited datasets. Together, these methods provide a multi-proxy framework enabling integrated reconstructions of terrestrial ecosystems from the Mesozoic through the present.
 

Field work

Palaeobotanic sample preparation

Tooth enamel preparation and analysis of stable carbon (C)/ oxygen (O) isotopes

Documentation

Each tooth is carefully documented prior to sampling, including photographs and metric descriptions. High-resolution silicone casts are taken of the enamel surface to preserve morphological details and allow dental microwear analysis. After serial sampling, additional casts of the sampled enamel surface are produced. These casts enable reverse modelling of the original tooth surface and precise spatial control of sampled areas, following established protocols for sequential enamel studies.

Serial sampling

Stable isotope signals in tooth enamel record dietary and environmental information from the period of enamel formation. Serial sampling is primarily applied to high-crowned (hypsodont) teeth, such as those of horses, rhinoceroses, and mammoths. In these taxa, enamel formation extends over several years, allowing isotope signals to capture more than a short juvenile phase. To access this temporal information, enamel is sampled sequentially along the growth axis of the tooth. Multiple narrow sampling bands are drilled at regular intervals (typically 2–3 mm), moving from the occlusal surface toward the enamel–root junction. This approach enables the reconstruction of intra-tooth isotopic variation and provides insights into seasonal and environmental changes during tooth mineralization.

Carbonate pretreatment

Prior to isotope analysis, enamel powder undergoes chemical pretreatment to remove potential contaminants. Organic matter is eliminated using diluted sodium hypochlorite, followed by rinsing with ultrapure water. Secondary carbonates are removed using a weak acetic acid buffer. This procedure ensures that the measured isotopic signal reflects biogenic enamel carbonate rather than post-depositional alteration.

Stable isotope analysis

Pretreated enamel carbonate is analyzed for stable carbon (δ¹³C) and oxygen (δ¹⁸O) isotopes. Carbonate samples are reacted with highly concentrated phosphoric acid (H₃PO₄) at 70 °C using a MultiFlow-Geo system interfaced with an Elementar IsoPrime 100 isotope ratio mass spectrometer (IRMS).

Bone collagen preparation and analysis of stable carbon (C)/ nitrogen (N)/ sulfur (S) isotopes

Documentation

All specimens are documented prior to sampling using a combination of photography, measurements, and, where appropriate, 3D surface scanning. This documentation captures morphology, preservation state, and existing damage, and ensures that sampling decisions are transparent and reproducible. Detailed documentation also allows specimens to be revisited digitally and supports integration with complementary analyses such as morphometrics or imaging studies

Sampling

Sampling follows standardized procedures designed to minimize damage while ensuring sufficient material for downstream analyses. Bones and teeth are sampled from anatomically and taxonomically non-diagnostic areas whenever possible, taking into account preservation, collagen yield, and planned analytical workflows. Sample size is estimated in advance to avoid repeated sampling, and all steps, from cutting to cleaning and storage, are fully recorded to ensure traceability and compatibility with future studies.

Collagen extraction

Collagen is extracted from pretreated bone or dentin samples using an Acid-Base-Acid (ABA) protocol that removes mineral components and secondary contaminants while preserving the original biogenic signal. The procedure includes demineralization, removal of humic substances, gelatinization, and freeze-drying, resulting in purified collagen suitable for stable isotope analysis, radiocarbon dating, and proteomic applications. Collagen quality is routinely assessed to ensure analytical reliability and reproducibility.

Stable isotope analysis

Stable isotope analyses are carried out at the Geoecology Stable Isotope Platform, University of Tübingen, using a Vario Isotope Cube elemental analyzer coupled to an IsoPrime Vision isotope ratio mass spectrometer (IRMS). This setup allows the simultaneous measurement of stable carbon (δ¹³C), nitrogen (δ¹⁵N), and sulfur (δ³⁴S) isotope ratios together with elemental composition.

Derivatization and compound-specific analysis of stable carbon (C)/nitrogen (N) isotopes

Derivatization

For compound-specific isotope analysis, amino acids must first be chemically modified to make them suitable for gas chromatography. Collagen samples are hydrolysed to release individual amino acids, which are then derivatized using established protocols (e.g. NAIP). This process increases volatility and thermal stability while preserving the original carbon and nitrogen isotopic signatures of the amino acids.

Stable isotope analysis

Compound-specific stable carbon (δ¹³C) and nitrogen (δ¹⁵N) isotope ratios are measured on individual amino acids using gas chromatography coupled to isotope ratio mass spectrometry (GC-IRMS). After chromatographic separation, each amino acid is combusted and its isotopic composition is determined independently.

Data processing

Statistical methods

All statistical analyses are carried out in the R statistical environment. Raw isotope data are quality-checked, standardized, and combined with contextual information (e.g. taxonomy, site, chronology) before further analysis. Data visualization plays an important role throughout the workflow and is used both for exploratory analyses and for communicating results in a clear and transparent way. We apply a range of descriptive and inferential statistical methods to explore variability within and between samples, taxa, and sites, with a strong emphasis on Bayesian statistical approaches. Depending on the research question and dataset, analyses include summary statistics, correlation analyses, and multivariate methods implemented within a probabilistic framework. Bayesian methods allow us to explicitly incorporate uncertainty, prior information, and small sample sizes, which are common in archaeological and palaeoecological datasets. All analyses are fully reproducible, and scripts can be shared with collaborators to ensure transparency and methodological consistency.

Niche modeling

To investigate dietary composition and ecological niches, we use Bayesian niche and mixing models implemented in R. MixSIAR is applied to estimate the relative contribution of different food resources to consumer diets based on stable isotope data. This approach explicitly accounts for uncertainty in source values and trophic discrimination factors and allows probabilistic interpretation of dietary scenarios. For comparisons of dietary breadth and niche overlap between individuals, populations, or species, we use SIBER (Stable Isotope Bayesian Ellipses in R). SIBER quantifies isotopic niche width and overlap in a robust Bayesian framework, making it particularly suitable for datasets with limited sample sizes. Together, these modelling approaches allow us to move beyond simple isotope plots and provide quantitative, statistically grounded reconstructions of trophic relationships and ecological interactions.

Sample and data curation

Paleobotanic samples
Collagen collection

Extracted collagen is stored in drying cabinets at stable room temperature and under controlled low humidity (<1%). These conditions allow long-term preservation and ensure the availability of collagen for future analyses, including radiocarbon dating (¹⁴C), ZooMS, and compound-specific amino acid isotope analysis. Maintaining a collagen collection reduces the need for repeated sampling of rare or fragile archaeological specimens and safeguards valuable material for future analytical approaches.

Carbonate collection

Carbonate samples derived from tooth enamel or other mineralized tissues are stored dry at room temperature. This storage strategy enables potential re-sampling of mineral components and supports future analytical applications, such as trace element studies or calcium and zinc isotope analyses. Carbonate collections therefore represent an important resource for expanding research questions without additional destructive sampling.

Open access repositories 

Following publication, all relevant raw and processed data are made publicly available through open-access repositories (e.g. Zenodo, Pandora). Open data sharing ensures transparency, reproducibility, and long-term accessibility, and facilitates reuse of datasets by the wider scientific community.

  • CEMP SITE (Collection of European Mesolithic and Paleolithic stable Isotopic data of terrestrial ecosystems)

PhD, M.Sc. and B.Sc. students

Ongoing

2025

  • Nathanael Drüeke (M.Sc. ASHE):
    Isotopic spacing in bone and dentin of mammoth. (D. Drucker, S. Greif)

2024

2023

2022

2021

2020

  • Samuel Stern (M.Sc. Geowissenschaften): 
    Isotope tracking of Messel (Eocene) and Schöningen Pleistocene fauna. (H. Bocherens,  D. Drucker)

See all current and past PhD students and B.sc./M.sc. students.

Recent publications

2025

  • Baumann, C., Larbey, C., Ebner, M., Bocherens, H., Hardy, K., 2025. The effect of plant food treatment on stable isotopes and their relevance for archaeological studies: A methodological pilot study. Journal of Archaeological Method and Theory 32: 14.  https://doi.org/10.1007/s10816-024-09690-5
  • Krajcarz, M., Baumann, C., Bocherens, H., Presslee, S., Krajcarz, M.T., 2025. Dietary preferences and collagen to collagen prey-predator trophic discrimination factors (Δ13C, Δ15N) in Late Pleistocene cave hyena. Quaternary Research 123: 27-40. https://doi.org/10.1017/qua.2024.43   
  • Krajcarz, M.T., Krajcarz, M., Kowalczyk, R., Tung, P., Bocherens, H., 2025. New model for estimating trophic position in mammalian carnivores based on bone collagen individual amino acids nitrogen stable isotopes. Palaeogeography-Palaeoclimatology-Palaeoecology 674: 113040. https://doi.org/10.1016/j.palaeo.2025.113040