Fossils are time capsules, preserving snapshots of ancient life. For paleobotanists, fossil leaves are especially valuable, revealing clues about plant evolution, ancient ecosystems, and environmental conditions.
A groundbreaking study published in Scientific Reports has uncovered an extraordinary detail in fossil leaves from Germany’s Rott Fossil Lagerstätte: traces of calcium oxalate (CaOx), a biomineral common in modern plants. This discovery challenges assumptions about fossilization processes and opens new avenues for understanding plant biology across geological timescales.
The Role of Calcium Oxalate in Modern Plants
Calcium oxalate (CaOx) is a chemical compound composed of calcium ions bonded to oxalate, a organic acid. It is one of the most widespread biominerals in modern plants, serving multiple critical functions. Biominerals are minerals produced by living organisms to perform structural or functional roles. In plants, CaOx crystals form in specialized cells called idioblasts, which regulate their growth and distribution.
Firstly, calcium oxalate helps plants manage excess calcium, storing it in crystal form to prevent toxicity. Calcium is essential for cell wall structure, enzyme activity, and signaling, but excessive amounts disrupt cellular processes. By crystallizing surplus calcium, plants maintain metabolic balance.
Secondly, these crystals act as a defense mechanism. Their sharp edges deter herbivores like insects and grazing animals, making leaves less appealing to eat. For example, plants like Dieffenbachia (dumb cane) use needle-shaped CaOx crystals (called raphides) to irritate predators’ mouths.
Thirdly, in some species, CaOx provides structural support, reinforcing cell walls or adding weight to seeds for better dispersal. In living plants, calcium oxalate appears in two primary forms.
- The first is druses, spherical clusters of crystals ranging from 10 to 100 micrometers in diameter. These are commonly found in the soft tissues between leaf veins (called areoles).
- The second form is prismatic crystals—needle-like or brick-shaped structures that often line leaf veins or sit within specialized cells.
Despite their importance in modern plants, calcium oxalate crystals rarely survive the fossilization process, making their discovery in 25-million-year-old fossils a remarkable achievement.
Why Calcium Oxalate is Rare in Fossils
Fossilization is a complex and often destructive process where organic material transforms into mineralized remains. Soft tissues decay quickly, and minerals can dissolve or transform under heat, pressure, and chemical changes. Calcium-based minerals, in particular, face significant challenges.
Calcium carbonate, another common plant biomineral, dissolves rapidly even in mildly acidic conditions, such as rainwater. Calcium oxalate is slightly more stable but still vulnerable over geological timescales. Over millions of years, calcium oxalate can oxidize into calcium carbonate, which then dissolves, leaving no trace.
Even when calcium oxalate survives initial decay, traditional fossil preparation methods can erase it. For example, scientists often use acid treatments to extract silica phytoliths—tiny silica structures that fossilize easily—from plant remains. These harsh chemicals dissolve calcium-based minerals, further reducing the chances of preserving calcium oxalate.
However, the Rott fossils reveal an indirect way calcium oxalate leaves its mark: through “ghost crystals.” Ghost crystals are cavities or mineral replacements that retain the shape of the original biomineral. When the original crystals dissolve, they leave behind voids in the fossilized tissue.
These voids fill with sediment, organic matter, or secondary minerals, creating imprints that mirror the original structures. Identifying these ghost crystals requires advanced tools and meticulous analysis, as demonstrated by the research team.
The Rott Fossil Lagerstätte: A Window into the Oligocene
The Rott fossil site, located near Bonn, Germany, is a Lagerstätte—a sedimentary deposit with exceptionally preserved fossils. The term Lagerstätte (German for “storage place”) refers to sites where soft tissues and delicate structures fossilize due to unique conditions like rapid burial, low oxygen, or fine-grained sediments.
Around 23 to 24 million years ago, during the late Oligocene epoch (a geological period spanning 34–23 million years ago), a freshwater maar lake formed in a volcanic crater. Maar lakes are deep, steep-sided volcanic craters filled with water, often hosting unique ecosystems.
The lake’s oxygen-poor (anoxic) bottom slowed decay, allowing leaves, insects, fish, and even delicate structures like feathers to fossilize in exceptional detail. The site’s fossil-rich layers include diatomite, a silica-rich sediment formed from the skeletons of algae called diatoms, and leafy coal beds, which are compressed layers of organic material from decaying plants.
These conditions preserved fossils with cellular-level details, making Rott a critical site for studying ancient plant life. Researchers examined 1,120 fossil leaf specimens from Rott, focusing on 64 that displayed mysterious granular structures. Earlier studies had interpreted these granules as algae colonies, pollen clumps, or damaged tissue.
However, the new study proposed a groundbreaking hypothesis: these granules were remnants of calcium oxalate biominerals. To test this idea, the team combined cutting-edge technology with comparative botany, bridging the gap between ancient fossils and modern plants.
Methods: Connecting Ancient Fossils to Modern Plants
The research team employed a multi-step approach to unravel the mystery of the granular structures. To begin, they analyzed modern plant leaves to understand how calcium oxalate crystals form and degrade.
Leaves from over 50 living species—including oaks (Quercus), walnuts (Juglans), ivy (Hedera helix), and lotus (Nelumbo nucifera)—were collected from the Bonn University Botanic Gardens. These species were chosen either because they were relatives of plants found in the Rott fossils or because they represented common biomineralization patterns.
One key experiment involved burning fresh leaves at high temperatures (600–650°C). Calcium oxalate crystals withstand burning, allowing researchers to study their size and distribution in the resulting ash. Ash analysis is a common method to study plant minerals, as organic material burns away, leaving behind inorganic residues like crystals.
Advanced imaging techniques, such as micro-computed tomography (micro-CT) and scanning electron microscopy (SEM), provided detailed 3D views of crystals inside fresh leaves without damaging the specimens. Micro-CT uses X-rays to create cross-sectional images of objects, while SEM scans surfaces with a focused electron beam to reveal nanoscale details.
For example, micro-CT scans of walnut leaves revealed large druses (50–70 micrometers) in the soft tissues between veins and smaller clusters along the veins themselves. SEM images of ivy leaves showed densely packed druses resembling grape-like clusters, while oak leaves displayed a mix of druses and prismatic crystals. These modern patterns became a reference for interpreting fossil structures.
Next, the team turned to the fossil leaves. Using light microscopy (LM), they identified granular structures on the fossil surfaces, noting their shapes, sizes, and locations. LM involves magnifying samples with visible light, making it ideal for initial observations.
Scanning electron microscopy (SEM) allowed them to examine these structures at magnifications up to 50,000 times, revealing intricate details invisible to the naked eye.
Energy-dispersive X-ray (EDX) spectroscopy, a technique that detects elemental composition by measuring X-rays emitted from a sample, mapped the chemical makeup of the granules, distinguishing between organic material (rich in carbon) and mineral deposits (rich in silicon or sulfur).
To rule out alternative explanations, the researchers compared the distribution and morphology of the granules to known structures like pollen, algae, or leaf hairs (trichomes).
Pollen grains, for instance, are typically scattered randomly on leaf surfaces, while calcium oxalate crystals align with veins or cluster in specific tissues. The granular structures in the fossils matched the latter pattern, supporting the calcium oxalate hypothesis.
Key Findings: Ghosts of an Ancient Mineral
The study yielded several groundbreaking discoveries. First, the shape and size of the fossil cavities closely matched calcium oxalate crystals in modern plants. For instance, globular voids in fossil oak leaves (assigned to Quercus nerifolia) mirrored druse clusters in living oaks (Quercus robur), both ranging from 40 to 50 micrometers in diameter.
Similarly, angular cavities along the veins of fossil willow leaves (Salix longa) resembled prismatic crystals in modern willows (Salix miyabeana). Chemical analysis provided further evidence. EDX spectroscopy revealed that many fossil “globules” consisted primarily of carbon and sulfur, with traces of iron—a signature of organic material, not the original calcium oxalate.
In some cases, the cavities had filled with secondary minerals like silica or calcium sulfate, creating mineral “ghosts” that replicated the crystals’ original shapes. For example, in one fossil leaf assigned to Sideroxylon salicites, dark organic globules were surrounded by shells of calcium sulfate, likely deposited during fossilization.
Perhaps the most surprising finding involved taxonomic revisions. Fossil leaves previously classified as water lilies (Nymphaea nymphaeoides) contained druse-like structures absent in modern water lilies but common in lotus plants (Nelumbo nucifera). This mismatch suggested that some fossils might have been misidentified, highlighting the potential of calcium oxalate patterns as a tool for accurate classification.
The Fossilization Process: From Crystal to Ghost
The researchers proposed a detailed model to explain how calcium oxalate crystals transformed into ghostly imprints over millions of years. The process began when leaves fell into the ancient lake and sank to its anoxic depths, where low oxygen levels slowed decay.
Soft tissues like mesophyll cells (the inner tissue of leaves where photosynthesis occurs) decomposed, but tougher structures—lignified veins (veins reinforced with lignin, a complex polymer) and cuticles (waxy outer layers of leaves)—persisted. Over time, calcium oxalate crystals dissolved, leaving behind empty cavities.
These cavities then filled with organic matter, such as degraded cellulose, or minerals like silica carried by groundwater. As the organic infillings aged, they often shrank, forming spherical globules due to surface tension.
In some cases, iron and sulfur seeped into the cavities, creating dark, mineral-rich granules. The result was a fossil record that, while devoid of original calcium oxalate, preserved detailed imprints of its former presence.
Implications for Science and Evolution
This discovery reshapes our understanding of plant evolution and fossilization in several ways. Firstly, it demonstrates that calcium oxalate biomineralization in angiosperms (flowering plants) dates back at least 25 million years, suggesting that this trait evolved much earlier than previously assumed.
This timeline parallels the rise of silica biomineralization in grasses, indicating that different plant groups developed unique strategies to cope with similar challenges, such as herbivory and nutrient management.
Secondly, the study offers a new tool for fossil identification. By analyzing the size, shape, and distribution of calcium oxalate ghosts, paleobotanists can distinguish between plant groups.
For example, druses in the soft tissues between veins might signal dicotyledonous plants (plants with two seed leaves, like oaks), while prismatic crystals along veins could help identify specific families or genera. This approach is particularly valuable for fragmentary fossils or those lacking diagnostic features like cuticles.
Thirdly, the findings provide insights into ancient ecosystems. The presence of calcium oxalate suggests that Oligocene plants faced herbivore pressure and soil conditions similar to those of today. By comparing fossil patterns to modern plants, scientists can infer past environmental factors, such as calcium availability or predation levels, enriching our understanding of prehistoric climates.
Challenges and Future Directions
While the study marks a significant advance, it also raises new questions. For instance, are calcium oxalate ghosts unique to the Rott fossils, or do they exist in other sites with exceptional preservation? Answering this requires re-examining fossil collections worldwide using similar methods.
Additionally, researchers must explore how fossilization conditions—such as sediment type, pH, and temperature—affect the preservation of calcium oxalate imprints.
The team also emphasized the need for a global database of plant biomineralization patterns. Such a resource would allow scientists to compare fossil structures with those of living plants, improving taxonomic accuracy and evolutionary studies. For example, if a fossil druse matches those in modern walnut leaves, it could strengthen evidence for the presence of walnuts in ancient ecosystems.
Conclusion
The Rott fossil leaves are a testament to the resilience of nature’s designs. Even delicate structures like calcium oxalate crystals, long thought to vanish without a trace, leave echoes across millions of years.
By combining advanced microscopy, chemical analysis, and comparative botany, this study bridges the gap between modern biology and deep time, offering fresh tools to decode plant evolution. As scientists revisit fossil collections with these methods, they may uncover more hidden biominerals—and with them, new chapters in the story of life on Earth.
Power Terms
Calcium Oxalate (CaOx):
Calcium oxalate is a chemical compound made of calcium ions and oxalate, a natural acid found in plants. It forms crystals inside plant cells and serves two main roles: storing excess calcium to prevent toxicity and defending against herbivores through sharp, irritating crystals. For example, plants like spinach and rhubarb contain calcium oxalate crystals. These crystals are important because they help plants manage nutrients and survive in their environments. In the study, fossilized calcium oxalate traces revealed how ancient plants functioned similarly to modern ones.
Biomineralization:
Biomineralization is the process by which living organisms produce minerals, often to strengthen tissues or perform specific functions. In plants, this includes forming silica phytoliths or calcium oxalate crystals. These minerals help plants defend against predators, store nutrients, or support structures. For instance, grasses use silica to deter insects, while oak trees use calcium oxalate. Understanding biomineralization helps scientists study plant evolution and ecology, as seen in the Rott fossils, where biomineral traces provided clues about ancient plant life.
Fossilization:
Fossilization is the process by which organic material transforms into preserved remains over millions of years. It typically occurs when organisms are buried in sediment, protecting them from decay. For plants, this often involves the replacement of original tissues with minerals like silica or calcium carbonate. Fossilization is crucial because it allows us to study extinct species and past ecosystems. The Rott fossils, preserved in an oxygen-poor lake, show how delicate structures like calcium oxalate ghosts can survive under rare conditions.
Ghost Crystals:
Ghost crystals are mineral imprints left behind after original crystals dissolve during fossilization. These voids fill with sediment or organic matter, preserving the shape of the original mineral. In the Rott study, calcium oxalate crystals dissolved but left cavities that later filled with silica or organic material. Ghost crystals are important because they act as “footprints” of ancient biological processes, helping scientists infer the presence of minerals that no longer exist in the fossil.
Druses:
Druses are spherical clusters of calcium oxalate crystals found in plant tissues. They range from 10 to 100 micrometers in size and often form in soft leaf areas between veins. For example, oak leaves contain druses that store calcium. In the Rott fossils, globular cavities matching druse sizes and locations confirmed their ancient presence. Druses are key to understanding how plants manage minerals and adapt to their environments.
Prismatic Crystals:
Prismatic crystals are needle- or brick-shaped calcium oxalate structures that line leaf veins or specialized cells. Plants like Parrotia persica use them for defense or structural support. The Rott fossils showed angular cavities along veins, mirroring these crystals. Prismatic crystals help scientists identify plant groups in fossils, as their shapes and locations vary between species.
Anoxic:
Anoxic describes environments without oxygen, such as the bottom of deep lakes or swamps. These conditions slow decay, allowing organic material to fossilize. The Rott maar lake’s anoxic depths preserved leaves and insects in exceptional detail. Anoxic settings are vital for fossil preservation because they protect tissues from bacteria and scavengers.
Lagerstätte:
A Lagerstätte (German for “storage place”) is a fossil site with extraordinary preservation of soft tissues or delicate structures. Examples include the Burgess Shale and the Rott site. These deposits provide rare insights into ancient life, such as the calcium oxalate ghosts in Rott fossils. Lagerstätten are critical for studying organisms that rarely fossilize under normal conditions.
Diatomite:
Diatomite is a sedimentary rock made of silica skeletons from diatoms, a type of algae. It forms in lakes or oceans and preserves fine details in fossils. The Rott fossils were found in diatomite, which helped retain cellular structures. Diatomite is also used in filters, abrasives, and as a soil additive.
Light Microscopy (LM):
Light microscopy uses visible light and lenses to magnify small objects. Scientists used LM in the Rott study to observe granular structures on fossil surfaces. While limited in resolution compared to electron microscopy, LM is simple and effective for initial fossil examinations.
Scanning Electron Microscopy (SEM):
SEM scans samples with a focused electron beam to create high-resolution images. It magnifies objects up to 50,000 times, revealing details like crystal shapes in fossil cavities. The Rott team used SEM to study the microstructure of fossil granules, comparing them to modern calcium oxalate crystals.
Energy-Dispersive X-ray Spectroscopy (EDX):
EDX detects elements in a sample by measuring X-rays emitted when electrons hit the material. In the Rott study, EDX showed fossil “globules” contained carbon and sulfur, confirming their organic origin. This technique helps distinguish minerals from organic remains in fossils.
Micro-Computed Tomography (Micro-CT):
Micro-CT uses X-rays to create 3D images of internal structures without damaging the sample. The Rott team scanned fresh leaves to map calcium oxalate crystals. This non-invasive method is ideal for studying delicate fossils or modern plant tissues.
Idioblasts:
Idioblasts are specialized plant cells that store unique substances like calcium oxalate crystals. They differ from surrounding cells and help plants manage toxins or deter herbivores. For example, idioblasts in ivy leaves produce druses. Studying idioblasts in fossils helps identify ancient plant functions.
Phytoliths:
Phytoliths are microscopic silica structures formed in plant cells. They fossilize easily and are used to study ancient diets and ecosystems. Unlike calcium oxalate, phytoliths are common in grasses. The Rott study compared phytoliths and CaOx preservation to understand fossilization biases.
Cuticle:
The cuticle is a waxy layer on plant surfaces that reduces water loss and protects against pests. Fossil cuticles often preserve leaf shapes and cell patterns. In the Rott fossils, cuticle remnants helped identify plant species and study their adaptations.
Lignified Veins:
Lignified veins are leaf veins reinforced with lignin, a tough polymer. Lignin resists decay, making veins more likely to fossilize. The Rott fossils’ preserved veins provided frameworks for studying calcium oxalate distribution.
Mesophyll Cells:
Mesophyll cells are the inner leaf tissues where photosynthesis occurs. They often degrade during fossilization, but their impressions can remain. In the Rott study, mesophyll cavities held clues about calcium oxalate crystal locations.
Maar Lake:
A maar lake forms in a volcanic crater and has steep, deep walls. The Rott maar lake’s quiet, anoxic waters preserved fossils exceptionally well. Such lakes are rare but valuable for studying ancient ecosystems.
Oligocene Epoch:
The Oligocene (34–23 million years ago) was a geologic period marked by cooling climates and expanding grasslands. The Rott fossils date to the late Oligocene, offering insights into plant life before modern ecosystems emerged.
Angiosperms:
Angiosperms are flowering plants, the most diverse plant group. They include oaks, ivy, and grasses. The Rott fossils confirmed that angiosperms used calcium oxalate for defense and nutrient storage millions of years ago.
Dicotyledonous Plants:
Dicotyledons (dicots) are plants with two seed leaves, broad leaves, and branching veins. Examples include oaks and roses. The Rott study used dicot CaOx patterns to identify fossil species and compare them to modern relatives.
Raphides:
Raphides are needle-shaped calcium oxalate crystals that irritate herbivores. Plants like pineapples and dumb cane use them for defense. Though not found in Rott fossils, raphides show how crystal shapes relate to function.
Silica:
Silica (SiO₂) is a mineral found in rocks and plant structures like phytoliths. It is highly resistant to decay, making it common in fossils. The Rott fossils’ diatomite sediment preserved silica-based algae skeletons.
Taxonomy:
Taxonomy is the science of classifying organisms based on shared traits. The Rott team revised fossil classifications by comparing calcium oxalate patterns to modern plants. Accurate taxonomy helps reconstruct evolutionary relationships.
Reference:
Malekhosseini, M., Ensikat, H. J., McCoy, V. E., Wappler, T., Weigend, M., Kunzmann, L., & Rust, J. (2022). Traces of calcium oxalate biomineralization in fossil leaves from late Oligocene maar deposits from Germany. Scientific Reports, 12(1), 15959.
