Taphonomic experiments reveal authentic molecular signals for fossil melanins and verify preservation of phaeomelanin in fossils.

Melanin pigments play a critical role in physiological processes and shaping animal behaviour. Fossil melanin is a unique resource for understanding the functional evolution of melanin but the impact of fossilisation on molecular signatures for eumelanin and, especially, phaeomelanin is not fully understood. Here we present a model for the chemical taphonomy of fossil eumelanin and phaeomelanin based on thermal maturation experiments using feathers from extant birds. Our results reveal which molecular signatures are authentic signals for thermally matured eumelanin and phaeomelanin, which signatures are artefacts derived from the maturation of non-melanin molecules, and how these chemical data are impacted by sample preparation. Our model correctly predicts the molecular composition of eumelanins in diverse vertebrate fossils from the Miocene and Cretaceous and, critically, identifies direct molecular evidence for phaeomelanin in these fossils. This taphonomic framework adds to the geochemical toolbox that underpins reconstructions of melanin evolution and of melanin-based coloration in fossil vertebrates.

An ancestral hard-shelled sea turtle with a mosaic of soft skin and scutes.

The transition from terrestrial to marine environments by secondarily aquatic tetrapods necessitates a suite of adaptive changes associated with life in the sea, e.g., the scaleless skin in adult individuals of the extant leatherback turtle. A partial, yet exceptionally preserved hard-shelled (Pan-Cheloniidae) sea turtle with extensive soft-tissue remains, including epidermal scutes and a virtually complete flipper outline, was recently recovered from the Eocene Fur Formation of Denmark. Examination of the fossilized limb tissue revealed an originally soft, wrinkly skin devoid of scales, together with organic residues that contain remnant eumelanin pigment and inferred epidermal transformation products. Notably, this stem cheloniid-unlike its scaly living descendants-combined scaleless limbs with a bony carapace covered in scutes. Our findings show that the adaptive transition to neritic waters by the ancestral pan-chelonioids was more complex than hitherto appreciated, and included at least one evolutionary lineage with a mosaic of integumental features not seen in any living turtle.

Chemical Evaluation of Eumelanin Maturation by ToF-SIMS and Alkaline Peroxide Oxidation HPLC Analysis.

Residual melanins have been detected in multimillion-year-old animal body fossils; however, confident identification and characterization of these natural pigments remain challenging due to loss of chemical signatures during diagenesis. Here, we simulate this post-burial process through artificial maturation experiments using three synthetic and one natural eumelanin exposed to mild (100 °C/100 bar) and harsh (250 °C/200 bar) environmental conditions, followed by chemical analysis employing alkaline hydrogen peroxide oxidation (AHPO) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Our results show that AHPO is sensitive to changes in the melanin molecular structure already during mild heat and pressure treatment (resulting, e.g., in increased C-C cross-linking), whereas harsh maturation leads to extensive loss of eumelanin-specific chemical markers. In contrast, negative-ion ToF-SIMS spectra are considerably less affected by mild maturation conditions, and eumelanin-specific features remain even after harsh treatment. Detailed analysis of ToF-SIMS spectra acquired prior to experimental treatment revealed significant differences between the investigated eumelanins. However, systematic spectral changes upon maturation reduced these dissimilarities, indicating that intense heat and pressure treatment leads to the formation of a common, partially degraded, eumelanin molecular structure. Our findings elucidate the complementary nature of AHPO and ToF-SIMS during chemical characterization of eumelanin traces in fossilized organismal remains.

Exploring the light-induced dynamics in solvated metallogrid complexes with femtosecond pulses across the electromagnetic spectrum.

Oligonuclear complexes of d4-d7 transition metal ion centers that undergo spin-switching have long been developed for their practical role in molecular electronics. Recently, they also have appeared as promising photochemical reactants demonstrating improved stability. However, the lack of knowledge about their photophysical properties in the solution phase compared to mononuclear complexes is currently hampering their inclusion into advanced light-driven reactions. In the present study, the ultrafast photoinduced dynamics in a solvated [2 × 2] iron(II) metallogrid complex are characterized by combining measurements with transient optical-infrared absorption and x-ray emission spectroscopy on the femtosecond time scale. The analysis is supported by density functional theory calculations. The photocycle can be described in terms of intra-site transitions, where the FeII centers in the low-spin state are independently photoexcited. The Franck-Condon state decays via the formation of a vibrationally hot high-spin (HS) state that displays coherent behavior within a few picoseconds and thermalizes within tens of picoseconds to yield a metastable HS state living for several hundreds of nanoseconds. Systematic comparison with the closely related mononuclear complex [Fe(terpy)2]2+ reveals that nuclearity has a profound impact on the photoinduced dynamics. More generally, this work provides guidelines for expanding the integration of oligonuclear complexes into new photoconversion schemes that may be triggered by ultrafast spin-switching.

Revealing Hot and Long-Lived Metastable Spin States in the Photoinduced Switching of Solvated Metallogrid Complexes with Femtosecond Optical and X-ray Spectroscopies.

An atomistic understanding of the photoinduced spin-state switching (PSS) within polynuclear systems of d4-d7 transition metal ion complexes is required for their rational integration into light-driven reactions of chemical and biological interest. However, in contrast to mononuclear systems, the multidimensional dynamics of the PSS in solvated molecular arrays have not yet been elucidated due to the expected complications associated with the connectivity between the metal centers and the strong interactions with the surroundings. In this work, the PSS in a solvated triiron(II) metallogrid complex is characterized using transient optical absorption and X-ray emission spectroscopies on the femtosecond time scale. The complementary measurements reveal the photoinduced creation of energy-rich (hot) and long-lived quintet states, whose dynamics differ critically from their mononuclear congeners. This finding opens major prospects for developing novel schemes in solution-phase spin chemistry that are driven by the dynamic PSS process in compact oligometallic arrays.

Excited State Dynamics of Bistridentate and Trisbidentate RuII Complexes of Quinoline-Pyrazole Ligands.

Three homoleptic ruthenium(II) complexes, [Ru(Q3PzH)3]2+, [Ru(Q1Pz)3]2+, and [Ru(DQPz)2]2+, based on the quinoline-pyrazole ligands, Q3PzH (8-(3-pyrazole)-quinoline), Q1Pz (8-(1-pyrazole)-quinoline), and DQPz (bis(quinolinyl)-1,3-pyrazole), have been spectroscopically and theoretically investigated. Spectral component analysis, transient absorption spectroscopy, density functional theory calculations, and ligand exchange reactions with different chlorination agents reveal that the excited state dynamics for Ru(II) complexes with these biheteroaromatic ligands differ significantly from that of traditional polypyridyl complexes. Despite the high energy and low reorganization energy of the excited state, nonradiative decay dominates even at liquid nitrogen temperatures, where triplet metal-to-ligand-charge-transfer emission quantum yields range from 0.7 to 3.8%, and microsecond excited state lifetimes are observed. In contrast to traditional polypyridyl complexes where ligand exchange is facilitated by expansion of the metal-ligand bonds to stabilize a metal-centered state, photoinduced ligand exchange occurs in the bidentate complexes despite no substantial MC state population, while the tridentate complex is extremely photostable despite an activated decay route, highlighting the versatile photochemistry of nonpolypyridine ligands.

Fossil insect eyes shed light on trilobite optics and the arthropod pigment screen.

Fossilized eyes permit inferences of the visual capacity of extinct arthropods1-3. However, structural and/or chemical modifications as a result of taphonomic and diagenetic processes can alter the original features, thereby necessitating comparisons with modern species. Here we report the detailed molecular composition and microanatomy of the eyes of 54-million-year-old crane-flies, which together provide a proxy for the interpretation of optical systems in some other ancient arthropods. These well-preserved visual organs comprise calcified corneal lenses that are separated by intervening spaces containing eumelanin pigment. We also show that eumelanin is present in the facet walls of living crane-flies, in which it forms the outermost ommatidial pigment shield in compound eyes incorporating a chitinous cornea. To our knowledge, this is the first record of melanic screening pigments in arthropods, and reveals a fossilization mode in insect eyes that involves a decay-resistant biochrome coupled with early diagenetic mineralization of the ommatidial lenses. The demonstrable secondary calcification of lens cuticle that was initially chitinous has implications for the proposed calcitic corneas of trilobites, which we posit are artefacts of preservation rather than a product of in vivo biomineralization4-7. Although trilobite eyes might have been partly mineralized for mechanical strength, a (more likely) organic composition would have enhanced function via gradient-index optics and increased control of lens shape.

Finding intersections between electronic excited state potential energy surfaces with simultaneous ultrafast X-ray scattering and spectroscopy.

Light-driven molecular reactions are dictated by the excited state potential energy landscape, depending critically on the location of conical intersections and intersystem crossing points between potential surfaces where non-adiabatic effects govern transition probabilities between distinct electronic states. While ultrafast studies have provided significant insight into electronic excited state reaction dynamics, experimental approaches for identifying and characterizing intersections and seams between electronic states remain highly system dependent. Here we show that for 3d transition metal systems simultaneously recorded X-ray diffuse scattering and X-ray emission spectroscopy at sub-70 femtosecond time-resolution provide a solid experimental foundation for determining the mechanistic details of excited state reactions. In modeling the mechanistic information retrieved from such experiments, it becomes possible to identify the dominant trajectory followed during the excited state cascade and to determine the relevant loci of intersections between states. We illustrate our approach by explicitly mapping parts of the potential energy landscape dictating the light driven low-to-high spin-state transition (spin crossover) of [Fe(2,2'-bipyridine)3]2+, where the strongly coupled nuclear and electronic dynamics have been a source of interest and controversy. We anticipate that simultaneous X-ray diffuse scattering and X-ray emission spectroscopy will provide a valuable approach for mapping the reactive trajectories of light-triggered molecular systems involving 3d transition metals.

Soft-tissue evidence for homeothermy and crypsis in a Jurassic ichthyosaur.

Ichthyosaurs are extinct marine reptiles that display a notable external similarity to modern toothed whales. Here we show that this resemblance is more than skin deep. We apply a multidisciplinary experimental approach to characterize the cellular and molecular composition of integumental tissues in an exceptionally preserved specimen of the Early Jurassic ichthyosaur Stenopterygius. Our analyses recovered still-flexible remnants of the original scaleless skin, which comprises morphologically distinct epidermal and dermal layers. These are underlain by insulating blubber that would have augmented streamlining, buoyancy and homeothermy. Additionally, we identify endogenous proteinaceous and lipid constituents, together with keratinocytes and branched melanophores that contain eumelanin pigment. Distributional variation of melanophores across the body suggests countershading, possibly enhanced by physiological adjustments of colour to enable photoprotection, concealment and/or thermoregulation. Convergence of ichthyosaurs with extant marine amniotes thus extends to the ultrastructural and molecular levels, reflecting the omnipresent constraints of their shared adaptation to pelagic life.

Chemical consequences of pyrazole orientation in Ru(II) complexes of unsymmetric quinoline-pyrazole ligands.

A series of homoleptic Ru(II) complexes including the tris-bidentate complexes of a new bidentate ligand 8-(1-pyrazol)-quinoline (Q1Pz) and bidentate 8-(3-pyrazol)-quinoline (Q3PzH), as well as the bis-tridentate complex of bis(quinolinyl)-1,3-pyrazole (DQPz) was studied. Together these complexes explore the orientation of the pyrazole relative to the quinoline. By examining the complexes structurally, photophysically, photochemically, electrochemically, and computationally by DFT and TD-DFT, it is shown that the pyrazole orientation has a significant influence on key properties. In particular, its orientation has noticeable effects on oxidation and reduction potentials, photostability and proton sensitivity, indicating that [Ru(Q3PzH)3](2+) is a particularly good local environment acidity-probe candidate.

Diastereomerization Dynamics of a Bistridentate Ru(II) Complex.

The unsymmetrical nature of a new tridentate ligand bis(quinolinyl)-1,3-pyrazole (DQPz) is exploited in a bistridentate Ru(II) complex [Ru(DQPz)2](2+) to elucidate an unexpected dynamic diastereomerism. Structural characterization based on a combination of nuclear magnetic resonance spectroscopy and density functional theory calculations reveals the first quantifiable diastereomerization dynamics for Ru complexes with fully conjugated tridentate heteroaromatic ligands. A mechanism that involves a large-scale twisting motion of the ligands is proposed to explain the dynamic interconversion between the observed diastereomers, and the analysis of both experiments and calculations reveals a potential energy landscape with a transition barrier for the diastereomerization of ∼70 kJ mol(-1). The structural flexibility demonstrated around the central transition metal ion has implications for integration of complexes into catalytic and photochemical applications.

A homoleptic trisbidentate Ru(II) complex of a novel bidentate biheteroaromatic ligand based on quinoline and pyrazole groups: structural, electrochemical, photophysical, and computational characterization.

We synthesized a new homoleptic, tris-bidentate complex [Ru(QPzH)3](2+) based on the novel biheteroaromatic, 8-(3-pyrazolyl)-quinoline ligand QPzH. The QPzH ligand was designed to reduce the distortions typically observed in complexes incorporating the 8-quinolinyl group into the ligand framework. This was indeed observed, and was also, as anticipated, found to facilitate the formation of tris-homoleptic Ru(II) complexes; [Ru(QPzH)3](2+) is the first reported tris-homoleptic complex with ligands based on the 8-quinolinyl group. The synthesis can either result in a statistical 3:1 mer/fac ratio of the complex, or, through controlled exposure to light, be tweaked to allow isolation of the pure mer isomer only. X-ray crystallography reveals three nonequivalent ligands, with significantly less strain than other quinoline-based bidentate ligands. The complex exhibits a nearly octahedral coordination geometry but shows large differences in bond lengths between the Ru core and the quinoline and pyrazoles, respectively. The Ru-N(pyrazole) bond distances are ∼2.04 Å, while the corresponding distances for Ru-N(quinoline) are ∼2.12 Å. Structural, photophysical, electrochemical, and theoretical characterization revealed a mer-Ru(II) complex with a low oxidation potential (0.57 V vs ferrocene(0/+)) attributed to the incorporation of the pyrazolyl group, a ground state absorption that is sensitive to the local environment of the complex, and a short-lived (3)MLCT excited state.

A dinuclear zinc(II) complex of a new unsymmetric ligand with an N(5)O(2) donor set: a structural and functional model for the active site of zinc phosphoesterases.

The dinuclear complex [Zn(2)(DPCPMP)(pivalate)](ClO4), where DPCPMP is the new unsymmetrical ligand [2-(N-(3-((bis((pyridin-2-yl)methyl)amino)methyl)-2-hydroxy-5-methylbenzyl)-N-((pyridin-2-yl)methyl)amino)acetic acid], has been synthesized and characterized. The complex is a functional model for zinc phosphoesterases with dinuclear active sites. The hydrolytic efficacy of the complex has been investigated using bis-(2,4-dinitrophenyl)phosphate (BDNPP), a DNA analog, as substrate. Speciation studies using potentiometric titrations have been performed for both the ligand and the corresponding dizinc complex to elucidate the formation of the active hydrolysis catalyst; they reveals that the dinuclear zinc(II) complexes, [Zn(2)(DPCPMP)](2+) and [Zn(2)(DPCPMP)(OH)](+) predominate the solution above pH4. The relatively high pK(a) of 8.38 for water deprotonation suggests that a terminal hydroxide complex is formed. Kinetic investigations of BDNPP hydrolysis over the pH range 5.5-11.0 and with varying metal to ligand ratio (metal salt:ligand=0.5:1 to 3:1) have been performed. Variable temperature studies gave the activation parameters ΔH(‡)=95.6kJmol(-1), ΔS(‡)=-44.8Jmol(-1)K(-1), and ΔG(‡)=108.0 kJmol(-1). The cumulative results indicate the hydroxido-bridged dinuclear Zn(II) complex [Zn(2)(DPCPMP)(µ-OH)](+) as the effective catalyst. The mechanism of hydrolysis has been probed by computational modeling using density functional theory (DFT). Calculations show that the reaction goes through one concerted step (S(N)2 type) in which the bridging hydroxide in the transition state becomes terminal and performs a nucleophilic attack on the BDNPP phosphorus; the leaving group dissociates simultaneously in an overall inner sphere type activation. The calculated free energy barrier is in good agreement with the experimentally determined activation parameters.

Synthesis, characterization, and reactivity studies of heterodinuclear complexes modeling active sites in purple acid phospatases.

To model the heterodinuclear active sites in plant purple acid phosphatases, a mononuclear synthon, [Fe(III)(H(2)IPCPMP)(Cl(2))][PF(6)] (1), has been generated in situ from the ligand 2-(N-isopropyl-N-((2-pyridyl)methyl)aminomethyl)-6-(N-(carboxylmethyl)-N-((2-pyridyl)methyl)amino methyl)-4-methylphenol (IPCPMP) and used to synthesize heterodinuclear complexes of the formulas [Fe(III)M(II)(IPCPMP)(OAc)(2)(CH(3)OH)][PF(6)] (M = Zn (2), Co (3), Ni (4), Mn (5)), [Fe(III)Zn(II)(IPCPMP)(mpdp)][PF(6)] (6) (mpdp = meta-phenylene-dipropionate), and [Fe(III)Cu(II)(IPCPMP) (OAc)}(2)(µ-O)][PF(6)] (7). Complexes 2-4, 6, and 7 have been crystallographically characterized. The structure of 6 is a solid state coordination polymer with heterodinuclear monomeric units, and 7 is a tetranuclear complex consisting of two heterodinuclear phenolate-bridged Fe(III)Cu(II) units bridged through a µ-oxido group between the two Fe(III) ions. Mössbauer spectra confirm the presence of high spin Fe(III) in an octahedral environment for 1, 3, and 5 while 2 and 4 display relaxation effects. Magnetic susceptibility measurements indicate weak antiferromagnetic coupling for 3, 4, and 5 and confirm the assignment of the metal centers in 2-5 as high spin Fe(III)-M(II) (M = Zn, Co (high spin), Ni (high spin), Mn (high spin)). Complexes 2-5 are intact in acetonitrile solution as indicated by IR spectroscopy (for 2-4) and electrospray ionization mass spectrometry (ESI-MS) but partly dissociate to hydroxide species and a mononuclear complex in water/acetonitrile solutions. UV-vis spectroscopy reveal pH-dependent behavior, and species that form upon increasing the pH have been assigned to µ-hydroxido-bridged Fe(III)M(II) complexes for 2-5 although 2 and 3 is further transformed into what is propsed to be a µ-oxido-bridged tetranuclear complex similar to 7. Complexes 2-5 enhance phosphodiester cleavage of 2-hydroxy-propyl-p-nitrophenyl phosphate (HPNP) and bis(2,4-dinitrophenyl)phosphate (BDNPP), but the reactivities are different for different complexes and generally show strong pH dependence.

Unsymmetrical dizinc complexes as models for the active sites of phosphohydrolases.

The unsymmetrical dinucleating ligand 2-(N-isopropyl-N-((2-pyridyl)methyl)aminomethyl)-6-(N-(carboxylmethyl)-N-((2-pyridyl)methyl)aminomethyl)-4-methylphenol (IPCPMP or L) has been synthesized to model the active site environment of dinuclear metallohydrolases. It has been isolated as the hexafluorophosphate salt H(4)IPCPMP(PF(6))(2) x 2 H(2)O (H(4)L), which has been structurally characterized, and has been used to form two different Zn(II) complexes, [{Zn(2)(IPCPMP)(OAc)}(2)][PF(6)](2) (2) and [{Zn(2)(IPCPMP)(Piv)}(2)][PF(6)](2) (3) (OAc = acetate; Piv = pivalate). The crystal structures of and show that they consist of tetranuclear complexes with very similar structures. Infrared spectroscopy and mass spectrometry indicate that the tetranuclear complexes dissociate into dinuclear complexes in solution. Potentiometric studies of the Zn(II):IPCPMP system in aqueous solution reveal that a mononuclear complex is surprisingly stable at low pH, even at a 2:1 Zn(II):L ratio, but a dinuclear complex dominates at high pH and transforms into a dihydroxido complex by a cooperative deprotonation of two, probably terminally coordinated, water molecules. A kinetic investigation indicates that one of these hydroxides is the active nucleophile in the hydrolysis of bis(2,4-dinitrophenyl)phosphate (BDNPP) enhanced by complex 2, and mechanistic proposals are presented for this reaction as well as the previously reported transesterification of 2-hydroxypropyl p-nitrophenyl phosphate (HPNP) promoted by Zn(II) complexes of IPCPMP.

Symmetrical and unsymmetrical dizinc complexes as models for the active sites of hydrolytic enzymes.

Dinuclear carboxylate-bridged zinc complexes of one symmetric and one asymmetric phenolate-based ligand catalyse the transesterification of 2-hydroxypropyl-p-nitrophenyl phosphate (HPNP) at different rates, with an unsymmetrical complex being more active than a symmetric one.