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. 2019 Nov 4;9(1):15947.
doi: 10.1038/s41598-019-52318-y.

Chemical characterization of pterosaur melanin challenges color inferences in extinct animals

Affiliations

Chemical characterization of pterosaur melanin challenges color inferences in extinct animals

Felipe L Pinheiro et al. Sci Rep. .

Abstract

Melanosomes (melanin-bearing organelles) are common in the fossil record occurring as dense packs of globular microbodies. The organic component comprising the melanosome, melanin, is often preserved in fossils, allowing identification of the chemical nature of the constituent pigment. In present-day vertebrates, melanosome morphology correlates with their pigment content in selected melanin-containing structures, and this interdependency is employed in the color reconstruction of extinct animals. The lack of analyses integrating the morphology of fossil melanosomes with the chemical identification of pigments, however, makes these inferences tentative. Here, we chemically characterize the melanin content of the soft tissue headcrest of the pterosaur Tupandactylus imperator by alkaline hydrogen peroxide oxidation followed by high-performance liquid chromatography. Our results demonstrate the unequivocal presence of eumelanin in T. imperator headcrest. Scanning electron microscopy followed by statistical analyses, however, reveal that preserved melanosomes containing eumelanin are undistinguishable to pheomelanin-bearing organelles of extant vertebrates. Based on these new findings, straightforward color inferences based on melanosome morphology may not be valid for all fossil vertebrates, and color reconstructions based on ultrastructure alone should be regarded with caution.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Tupandactylus imperator (specimen CPCA 3590) from the Crato Formation and headcrest microbodies. (A) Partial skull with its lower jaw overlying the headcrest. (B) Interpretative drawings of the figure (A,C) according to skull position. (D) Headcrest tissue exhibiting the blocky calcite crystals, and (E) melanosomes amidst external molds of neomorphic crystals (arrowheads). (F,G) Densely packed microbodies with subspherical morphology. (H) Keratin-like structure overlying or surrounding particles. (I) Scattered pits (arrowheads) on the microbody surface. (E) Calcite crystals blocks located amongst microbodies, with several pigmentary particles scattered on their surface (arrowheads).
Figure 2
Figure 2
Raman spectra from T. imperator (CPCA 3590) headcrest and melanins. (A) The top spectrum is from the bony area of the headcrest showing peaks of CO32− (at 1086 cm−1) and PO43− (at 965 cm−1), which is consistent with a bioapatite variety. The second spectrum is that of the headcrest, and is similar to the two shown below, those of synthetic and Sepia melanins, respectively. (B) Raman spectra from dark bands of the headcrest exhibiting a signal variation between the two regions of measurement (red circles indicate the two points where four measurements were made) (C). Grey dashed lines in (B) represent the theoretical bands of eumelanin. (D) Fitted spectra using Gaussian function (R2 = 0.9852) from the Point 1 seen in (B), showing that multiple bands are also observed as predicted in other studies. (E) Microscopic image from the headcrest surface, showing the region where the fluorescence mapping was performed (white dashed lines). (F) Map of the 1085 cm−1 region that is diagnostic of calcite, from the area seen in (E), indicating a faint signal of the soft tissues, suggesting that calcite from matrix predominates.
Figure 3
Figure 3
Mass spectrum of CPCA 3590 headcrest melanin. (A) Mass spectrum from the degradation products of alkaline peroxide oxidation. (B) Hypothetic structural modifications of eumelanin from T. imperator (based on Ito et al.), according to the most abundant yielding recovered from the oxidation process (i.e. PTeCA). (C) Oxidation was performed on four separate occasions and the results were shown as mean ± SEM. Values for the sediment are close to the detection limits. Values are in ng/mg. Differences as evaluated by students’ t-Test (two-tailed) are P < 0.01 except for PDCA (P < 0.05).

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