# Phytochemical Screening of 45‐Million‐Year‐Old Colored Angiosperm Leaves Reveals Distinctive Chlorophyll‐Derived and Polyphenolic Pigments

## TL;DR

Scientists found preserved plant pigments in 45-million-year-old leaves, revealing insights into ancient plant chemistry and fossilization.

## Contribution

Discovery of specific chlorophyll derivatives and polyphenolic pigments in ancient leaves using sensitive chemical analysis.

## Key findings

- Chlorophyll derivatives and polyphenolic pigments were preserved in 45-million-year-old angiosperm leaves.
- Dihydro-132,173-cyclopheophorbide a-enol was identified, a compound previously only known in modern microorganisms.
- Flavonoids apigenin and luteolin were detected, indicating stable preservation of polyphenolic compounds.

## Abstract

Present‐day angiosperm plants produce a plethora of metabolites including pigments that serve for important functions such as photosynthesis, protection against light, attraction of pollinators, and defense against microbes and herbivores. However, little is known about phytochemical constituents of ancient angiosperms, their distribution in the fossil record, their stability in deep time, and diagenesis. Outstanding preservation of ancient angiosperms, including exceptional color preservation, has been reported, but chemical analyses of such valuable specimens are limited by the rarity of the fossil material and the small amounts of potentially preserved metabolites. Here we use highly sensitive targeted liquid chromatography–tandem mass spectrometry in multiple reaction monitoring mode to screen for nanogram quantities of intact ancient phytochemical metabolites and their products in exceptionally well‐preserved, about 45‐Ma‐old leaves from the Eocene Geiseltal fossil Lagerstätte, Germany. We show that diverse chlorophyll derivatives and degradation products as well as polyphenolic pigments are preserved in green to yellow colored angiosperm leaves and the brown coal matrix from Geiseltal. Most interesting is the fossil occurrence of the “unstable” green chlorophyll derivative dihydro‐132,173‐cyclopheophorbide a‐enol, since cyclopheophorbide‐enols are otherwise known as unique non‐fluorescent chlorophyll catabolites of microorganisms in modern aquatic environments. The monopyrrole hematinic acid is interpreted as a stable product of chlorophyll catabolism via linear tetrapyrroles. Moreover, polyphenolic compounds in the fossil angiosperms are represented by the flavonoid pigments apigenin and luteolin. Our results demonstrate the potential of paleometabolomic‐like screening of individual plant fossils to trace the fate of phytochemical constituents and to understand the processes of fossilization at the molecular level.