#An ancient penguin with yellow feathers

Heard Island emerged from the Indian Ocean millions of years ago and is now savaged by snow and battered by wind. The island is among the most remote places on Earth and our record of its existence only began in the mid 19^th century. Victorian-era access to Heard Island was by schooner or barque, from a harbour on the Kerguelen Islands 450 km away. In 1874 a collection of Heard Island wildlife was brought to the Kerguelen Islands and given to the crew of the visiting USS Swatara. The animals from this remote wilderness were passed along to Dr. Jerome Kidder, surgeon and naturalist for Swatara. Kidder prepared and studied the specimens, and 17,000 km away in Washington, DC, he stored the bounty at the Smithsonian Institution.

The specimens from Heard Island included this macaroni penguin:

Crest feathers from a macaroni penguin Eudyptes chrysolophus (USNM 533533). Photo credit D Thomas.

The penguin that Dr. Kidder brought back from Heard Island has recently helped us with an interesting feather colour puzzle (Thomas et al. 2013). Yellow is a very common feather colour and is achieved in a variety of ways. For example, a canary will eat seeds that contain yellow pigments, and those pigments will be deposited in feathers. A parrot will use biochemical pathways to make yellow pigments while the feather is growing. The yellow pigment in penguin feathers is… a mystery. By studying yellow penguin feathers we could not only learn about the pigment chemistry, but we could also discover when the pigment first evolved and which fossil penguins had yellow feathers.

Raman spectroscopy has given us some new insight into the chemistry of the yellow penguin pigment. As mentioned elsewhere in this blog, Raman spectroscopy describes interactions between atoms in molecules or minerals. A Raman spectrum is a set of bands (or peaks…), and each band describes the energy of a particular atomic interaction. For example, a Raman spectrum of bone contains a band at 960 cm^-1, and that band relates to an interaction between phosphorus and oxygen atoms. The Raman spectrum from the penguin pigment was brand new; it is not like anything that we have found in the published literature, and it is completely different from other feather pigments. The spectrum contains important bands at 1578, 1491, 1285 and 683 cm^-1, which are hallmarks for a nitrogen-bearing, heterocyclic aromatic ring.

Nitrogen-bearing aromatic heterocyclic rings, highlighted in orange and shown in a pterin (left) and a porphyrin (right). Our best guess at this stage is that the penguin pigment contains something similar. Images adapted from Wikimedia Commons.

We still need more information before we can completely solve the structure, but at this stage we know for certain that the pigment is unlike anything so far observed in nature, and it is not likely to be a pigment that the penguins are eating. This means that penguins have evolved a biochemical pathway for making the pigment themselves. Armed with this new knowledge, we can look at the evolutionary history for the yellow penguin pigment. We do this by lining up the relevant evidence. First, we know that ten species of living penguin make the yellow pigment, including king penguins, yellow-eyed penguins and macaroni penguins. Second, we have a good idea that all living penguins are the descendants of an ancestor that lived more than 13 years ago (Ksepka and Thomas, 2012). Third, only penguins have this yellow pigment, and it is absent from albatrosses, shearwaters or petrels. This means:

1)     Living penguins inherited the yellow pigment from the ancestor that lived 13 years ago.

2)     The yellow pigment evolved after penguins had separated from albatrosses, petrels and shearwaters, at least 62 million years ago (Slack et al. 2006).

So the yellow penguin pigment is ancient, evolving sometime between 62 and 13 million years ago. We can use this information to add colour to an ancient penguin from South America.

Madrynornis mirandus (the ‘wonderful bird of Madryn’) is a 10 million year old penguin from Argentina (Acosta Hospitaleche et al. 2007). Madrynornis is more closely related to crested-penguins, like Kidder’s macaroni penguin, than any other group of penguins that we know of. Yellow-eyed penguins are also close relatives of both Madrynornis and the crested penguins, and all three groups shared a common ancestor that probably lived between 10 and 13 million years ago. We know that two of the groups descended from this common ancestor now have yellow feathers (crested and yellow-eyed penguins), so this means that the common ancestor probably had yellow feathers. And, because Madrynornis is a descendant of a penguin with yellow feathers, it very likely had them as well.

So, using Raman spectroscopy of modern feathers, we can add yellow pigments to Madrynornis mirandus, a 10 million year old penguin.

Image credits: Yellow-eyed penguin photo from Christian Mehlfuhrer, Macaroni penguin photo from Liam Quinn, Madrynornis images from Acosta Hospitaleche et al. 2007.

 

Acosta Hospitaleche C, Tambussi C,  Donato M, Cozzuol M. 2007. A new Miocene penguin from Patagonia and its phylogenetic relationships. Acta Palaeontologica Polonica 52, 299-314.

Ksepka DT, Thomas DB. 2012 Multiple Cenozoic invasions of Africa by penguins (Aves, Sphenisciformes). Proceedings of the Royal Society B 279, 1027–1032.

Slack K, Jones C, Ando T, Harrison G, Fordyce RE, Arnason U, Penny D. 2006. Early penguin fossils, plus mitochondrial genomes, calibrate avian evolution. Molecular Biology and Evolution 23, 1144–1155.

Thomas, DB, McGoverin CM., McGraw KJ. James HF, Madden O. 2013. Vibrational spectroscopic analyses of unique yellow feather pigments (spheniscins) in penguins. Journal of the Royal Society Interface (doi: 10.1098/​rsif.2012.1065)

Image links: macaroni penguin, Madrynornis mirandus, porphyrin, pterin, yellow-eyed penguin

 

P.S. We called the Raman spectrum ‘spheniscin’, and eventually we adopted this as the name for the pigment. Unfortunately, this was a very embarrassing mistake. A quick google search will show that the name ‘spheniscin’ has already been taken, and I will make sure this mistake is formally corrected in a follow-up publication.

#Organic traces in dinosaur embryos

Robert Reisz and colleagues have described dinosaur embryos in a Letter published in Nature. The embryos are from a sauropodomorph dinosaur, “…probably Lufengosaurus…”, and were collected from Early Jurassic sediments (Sinemurian, 190–197 million years old) in the Yunnan Province of China. The tiny embryonic bones are impressive and the microstructural detail (i.e. histology) is astounding. The thing that caught my eye, though, was the evidence for organic molecules.

Thin sections of the tiny dinosaur bones were analysed at the National Synchrotron Radiation Research Center (NSRRC) in Taiwan. The researchers were interested in the wavelengths of infrared light that would be absorbed by the fossils (the synchrotron was their light source). Infrared absorbance is an excellent method for identifying molecules in a sample. Atoms bind together to form molecules – the type of atoms and the way they are bound controls the wavelengths of light that a molecule will absorb. More specifically, a molecule can be identified from the wavelengths of light it absorbs. The researchers presented the results from one bone: infrared wavelengths were directed at 120 points (150 × 180 µm, one spectrum collected every 15 µm), and the spectrum of wavelengths absorbed from each point was mapped.

Organic remnants in a dinosaur bone. Infrared absorption in the amide I and amide II regions provides evidence for a peptide bond, which are found in proteins. Light microscope images show section of fossil bone that was analysed (left), colored maps show the distribution of apatite, amide I and a carbonate (middle), and spectra were collected from points highlighted with a red cross (right). Reprinted by permission from Macmillan Publishers Ltd: Nature. RR Reisz et al. Nature 496, 210-214 (2013) doi:10.1038/nature11978, copyright (2013).

Some of wavelengths absorbed by the dinosaur bones would also be absorbed by the proteins of living animals. The basic structure of a protein involves a set of small molecules (amino acids) linking together to form a long chain (peptide). The ‘linking together’ forms a peptide bond, which has a characteristic infrared absorption. The characteristic absorptions of a peptide bond appear at very specific regions in an infrared absorption spectrum – two of those regions fall between 1500 and 1700 cm^-1 and are termed ‘amide I and amide II’. The embryonic dinosaur bones absorb infrared light in the amide I and amide II regions, suggesting the presence of a peptide. Bone is a mixture of mineral (bioapatite) and protein (collagen), so it might be possible that the peptide traces in the fossil are remnants of collagen. The authors state that “…Previous reports of preserved dinosaur organic compounds, or ‘dinosaurian soft tissues’, have been controversial because it was difficult to rule out bacterial biofilms or some other form of contamination as a possible source of the organics. Our results clearly indicate the presence of both apatite and amide peaks within woven embryonic bone tissue, which should not be susceptible to microbial contamination or other post-mortem artefacts….”

Remnant collagen from a 190 million year old dinosaur embryo? Might well be.

Reisz, R R et al. 2013. Embryology of Early Jurassic dinosaur from China with evidence of preserved organic remains. Nature 496, 210-214.

#African fossil penguins, behind the scenes

A diverse group of penguins lived in Africa 10-12 million years ago. Dr. Dan Ksepka and I recently co-wrote an article describing these ancient penguins, and Dan has a great summary on his blog. I thought I would take this opportunity to show some of the ‘behind the scenes’ work that helped with the article but was not included in the final cut.

One of the first things we wanted to know was the age of the fossil penguin bones. Eventually we would solve this problem with stratigraphy – one of our first moves, though, was to see if the burial environment of the fossils we had just found was similar to the burial environment of fossil penguins that had previously been described. The burial environment for these already-described penguins is around five million years old.

The chemistry of fossil bones can be useful for describing different burial environments. Water that flows between grains of sediment can have very different chemical compositions in different burial environments, and water can alter the chemistry of a fossil bone in a distinct way. If two fossils have very similar elemental compositions, then you can start thinking about how they might have come from the similar burial environments. Likewise, if fossil bones have distinct chemical compositions, then it might be telling you that the bones also have different ages. Of course, fossils with the same age can be buried in different burial environments, so checking for similarities in burial environment is just a preliminary step.

Analysing the chemistry of a fossil bone is easy to do when you have access to a handheld x-ray fluorescence spectrometer. Two of the penguin bones that I analysed are shown below – most of a humerus from a ~5 million year old Inguza predemersus, and the head of a humerus from one of the newly found penguins (Sphenisciformes B). I collected XRF data from these specimens and found the same proportion of calcium and phosphorus in each fossil. This isn’t surprising – these are the two major ingredients of bone.

Fossil bones from the Western Cape of South Africa.

Both of these fossil bones are orange-brown-ish, and both have roughly the same proportion of iron. Iron oxides (rust) can produce orange-brown colours in fossils. So, no great differences in calcium, phosphorus or iron. Strontium, however, was a large component of the newly discovered bone, and represented a smaller proportion of the Inguza bone. Strontium can be fairly mobile, however, and bones from the same locality can have different amounts of strontium.

Energy dispersive x-ray fluorescence spectra from fossil penguin bones.

The most surprising and interesting results were at the higher end of the energy scale. A peak that might represent uranium was very clear in the spectrum from the newly discovered fossil penguin, and comparatively weak in the spectrum from Inguza. Likewise, a peak that might represent yttrium is distinct in the spectrum from Inguza, and weak in Sphenisciformes B. These trace elemental differences, combined with the variation in strontium concentration, are telling us that the two fossil bones have been altered by groundwater in different environments. We took this to mean that the bones were from different burial environments….

….and sure enough, the Inguza fossil was buried around 5 million years ago in a sandy river channel, and Sphenisciformes B was buried between 10 and 12 million years ago in a gravelly estuary. Of course, this conclusion was brought to us by sedimentology and stratigraphy, but it is very nicely supported by spectroscopy.

Thomas DB and Ksepka, DT. 2013. A history of shifting fortunes for African penguins. Zoological Journal of the Linnean Society. DOI: 10.1111/zoj.12024

#Ancient plants record mountain growth

Plants that live at higher altitudes have higher concentrations of ‘sunblock’ compounds in their walls. Barry Lomax and colleagues consider the mountain building implications of the ‘sunblock’/altitude relationship. Photo from Wikimedia Commons. Graph reproduced after Lomax et al. 2012

How quickly do mountains rise? The Southern Alps have been rapidly rising for the last five million years (…stone giants?), and India started raising the Himalayas about 10 million years ago. We might imagine dramatically different landscapes in these parts of the world before the mountains grew. Mountains have a profound effect on local climate, vegetation and animals, so when we reconstruct ancient environments in mountainous places, we need to know how tall the mountains were back in the day.

Modeling the rate of mountain growth is a difficult business, and many models factor in climate change, which is a science all on its own. Fortunately, infrared spectroscopy might provide an easier method. Barry Lomax and colleagues have recently described a method for calculating paleoaltimetry, that is, how high something was in the past, and suggested it could be useful for figuring out rates of mountain growth. In essence, imagine you are standing next to a deposit of fossil plants, and you know the altitude you are standing at, and the age of the fossil deposit. If you calculate the altitude that those plants were growing at during ancient times, then you can calculate a rate of altitude change. So, how can we calculate paleoaltimetry from a fossil plant?

It all comes down to sunlight. From Lomax et al. 2012 “…UV-B radiation flux increases with altitude…due to the physical properties of the atmosphere…” Consider then, that “…The vast majority of terrestrial land plants require sunlight to drive photosynthesis leading to exposure to high-energy short wavelength UV-B radiation, resulting in damage to plant proteins, membrane lipids and DNA. One mechanism by which plants can mitigate these effects is via the up-regulation of UV-B absorbing compounds (UACs)…” (Lomax et al. 2012). Plants that grow at higher altitudes have more UACs – more sunblock – and hence plants record the altitude of their growth.

The UV-B absorbing compounds include ferulic acid and p-coumaric acid, which very importantly, contain aromatic rings of carbon atoms. An aromatic ring is a group of atoms that are joined together in a closed loop, and which freely share electrons, which is what makes them useful for gobbling up UV light. An infrared spectrum of “…[s]poropollenin, the biopolymer that makes up the exine (outer wall) of spores and pollen…” includes a band at 1520 cm^-1, which is attributed to aromaticity (Lomax et al. 2012). Lomax and colleagues describe the relative abundance of UACs in sporopollenin by normalizing the 1520 cm^-1 band against the absorption of hydroxyl groups (a band around 3200 cm^-1). Lomax and colleagues then show a relationship between UAC abundance and altitude. Sure enough, plants that live at higher altitudes need more sunblock.

The next step is to include fossils. The method of Lomax and colleagues could be applied to well-preserved fossil plants found on a mountainside, to figure out how tall the mountain was when the plants lived.

Lomax BH, Fraser WT, Harrington G, Blackmore S, Sephton MA, Harris NBW. 2012. A novel palaeoaltimetry proxy based on spore and pollen wall chemistry. Earth and Planetary Science Letters 353-354: 22–28.

Top image modified from Wikimedia Commons

#A problem with sand dunes

The West Coast National Park in the Western Cape of South Africa is a great place for spotting wildlife. The Geelbek Dune System lies in the southern reaches of the National Park, which is the distant background of this photo. Fossils recovered from the Geelbek Dunes have provided great insight into the ancient wildlife of the region.

How old is a fossil from a sand dune? Fossils recovered from dune fields help us to reconstruct ancient environments. But how ancient is ancient? Thanks to Steno we know that the fossils are younger than the sand dunes they are buried in, and we know that the dune system is younger than the rocks underneath. But sand dunes are dynamic, shifting and reforming with every gust of wind and every drop of rain. Fossils can be winnowed out from depth and settle in newly formed layers, or simply be transported to the dune surface to sit alongside modern debris. Our usual methods for reading the age of a fossil are less reliable in dune environments – for an analogy, imagine pulling clothes out of a tumble dryer and trying to decide what order you first put them in.

We can try and solve this problem in a few different ways. One approach is to use  relative dating. When we don’t have the ability to place an absolute age on a fossil assemblage – fossil A is 12,000 years old and fossil B is 10,000 years old – we can instead place fossils in  an order of deposition – fossil A is older than fossil B. Relative ages are useful for tracking changes in ancient environments, and importantly, these ages can tell us about the natural disintegration of fossils over time.

Fossil ‘survivorship’ in the Geelbek Dune systems was the focus of a 2008 study by Nicholas Conard, Steven Walker and Andrew Kandel. Here the authors gathered an assemblage of fossils, placed them into size categories, and assigned a relative age to each one. Conard and colleagues observed that both dense and porous bones stood the test of time at Geelbek, in contrast to the standing paradigm, which states that dense fossil bones have a better chance of being preserved as fossils. Importantly, this means there is a good chance that tiny animals are also preserved from the ancient environment represented in the Geelbek Dunes assemblage. There is a big question looming over this conclusion though – how were the fossils assigned a relative age?

Conard and colleagues used colour, heft and other physical parameters to gauge the extent to which minerals had grown on and in the fossils. The authors reasoned that bones that had more minerals had been in the ground for longer, and hence where older. So, fossil bones were sorted into ‘mineralisation categories’ based on the perceived amounts of secondary minerals, and ‘mineralisation category’ became a proxy for burial duration.

This is where spectroscopy comes in. Professor Anusuya Chinsamy-Turan, my postdoc advisor at the University of Cape Town, spoke to Dr Kandel after he presented this work at a conference in Cape Town. Andrew and Anusuya agreed that studying the chemistry of the Geelbek fossils might make the ‘mineralisation categories’ (and the assessment of relative age in a dune system) a little more robust. Andrew brought the very precious fossils from Iziko South African Museum to the University of Cape Town, where I analysed the bones with a handheld x-ray fluorescence spectrometer. We were interested to see if the concentration of mineral forming elements agreed with the mineralisation category that had been assigned to each fossil.

Fossil bones from the Geelbek Dune system, grouped by mineralisation category. X-ray fluorescence was used to assess the elemental concentrations of mineral forming elements in these bones. The physically assessed mineralisation categories, and the chemical assessments from XRF, did not always agree. Image from Thomas et al. 2012. Permission for image use granted through Rightslink.

Well, we found a loose correlation between the mineralisation categories and the XRF data  (Thomas et al. 2012). To the original five categories we added a zeroth – modern bone. X-ray fluorescence spectra could distinguish the lower categories (zero to three) from the higher categories (four to five), but it couldn’t separate out individual categories. We were able to see a range of elemental concentrations, but the chemical data was not a strong match for the physically assessed ‘mineralisation categories’. From a chemical perspective, the ‘mineralisation categories’ did not reflect mineral accumulation.

So what does this mean? The original conclusion about bone ‘survivorship’ is mostly valid – there are both dense and porous fossils with very high elemental concentrations of mineral forming elements, and hence they have may been buried the longest. Unfortunately, I don’t think physically assessed ‘mineralisation categories’ can give accurate relative ages by themselves. Fortunately, x-ray fluorescence is non-destructive and portable, so we could easily supplement the physical assessments with chemical data. Together, these assessments can help assign relative ages to fossils recovered from sand dunes.

Conard NJ, Walker SJ, Kandel AW. 2008. How heating and cooling and wetting and drying can destroy dense faunal elements and lead to differential preservation. Palaeogeography, Palaeoclimatology, Palaeoecology 266: 236-245

Thomas DB, Chinsamy A, Conard NJ, Kandel AW. 2012 Chemical investigation of mineralisation categories used to assess taphonomy. Palaeogeography, Palaeoclimatology, Palaeoecology 361-362: 104-110

Behind the curtain at the Smithsonian

A week ago I was at the 72^nd Annual meeting of the Society of Vertebrate Paleontology. Some of the research presented there has been showing up online – like T. rex eating Triceratops, and descriptions of giant sea creatures. I presented research I am working on at the Smithsonian and it lead to some great conversations with some very interesting people – including two incredible paleoartists, Tyler Keilor and Julius Csotonyi. Before I talk about the conference, I just want to give a bit of background to my work, and mention the mind-blowing collections at the Smithsonian.

I am based in the Division of Birds, in the National Museum of Natural History. Without a doubt, the Natural History Museum is one of the greatest places on Earth. The specimens on display in the public galleries are a small fraction of the amazing things that have been collected over the years. I have the incredible privilege of being among the relatively few people that gets to step behind the curtain and see the wonders that are not on public display.

A large part of the Division of Birds is occupied with the study skin collection and this is what I want to talk about here. I have been working with these skins quite a bit lately and so it is only fair that I do my part to add to the collections. So, under the tutelage of Carla Dove, James Whatton, Chris Milensky, Brian Schmidt, and mostly, Christina Gebhard, I have been learning to prepare birds as study skins. The study skin collection receives many visiting researchers every year, from within the United States and around the world, so it is important to maintain and grow the collection. I am part of a small class of novices that are being trained in the sacred art of preparing study skins. How does this relate to the spectroscopy of fossils? All in good time. First, let me tell my story about how a simple lad from Thames, New Zealand has helped to grow one of the most important natural history collections in the world.

It began in earnest with a male house sparrow, Passer domesticus. I will skip the prologue and move to the part where the sparrow is sitting in front of me on a long table in the Museum prep lab. Our small group of trainees, each equipped with a house sparrow, is clustered either around Brian or Christina, who are also seated with a bird. Brian and Christina, and Chris, working at the high bench on the other side of the room, tell us the number of birds they have prepared and the time it would take them to produce a study skin from an inert sparrow. Thousands and 15 to 30 minutes. We would spend three hours that morning moving through the first set of steps, converting our deceased seed eater into a feather pelt. A quick break for lunch – a true testament of appetite after our mornings activities – and we were ready for the final steps. Cotton wool to fill, cotton thread to seal. For the second time that day the sparrows looked like silently sleeping woodland creatures. The finishing steps involved carefully pinning the bird to a spongy plastic board, making sure the tail was evenly fanned and the head and wings were perfectly straight. After a few days the pins were removed and the sparrow was set in an immortal pose.

Anne Wiley and I have just received the house sparrows that will be prepared as study skins. Anne is a Peter Buck postdoctoral fellow and studies isotopes from seabird tissues. Photo credit: Christina Gebhard.

On the left, foreground to background, are myself, Brian Schmidt and Megan Spitzer (just visible behind Brian). Anne Wiley is sitting opposite me and Hanneke Meijer is sitting across from Megan. It is not a coincidence that I am sitting near Brian, our tutor – preparing study skins is crazy challenging. Photo credit: Christina Gebhard.

The study skins are filled with cotton. I don’t think I need to go into the details. Photo credit: Christina Gebhard.

A final and critical step involves pinning the specimens. This controls the final appearance of the study skin. Photo credit: Christina Gebhard.

These study skins are eternal with proper care, and many tens of thousands of birds have been prepared in the museum’s long history. You might think this is morbid – I have no comment either way, but I will leave you with the following thought. Below is a bird that I have used in my studies. My methods are non-destructive and they use instruments that were built inside of the last decade. This bird was collected in 1883, decades before the technique I use was even discovered, let alone designed into a computerised instrument. This specimen was available to researchers 100 years before I was born and I expect it will outlive me by at least as long. So, there is a very long term, possibly eternal value to these specimens. I find it easy to see the value of these historic collections in a modern world that is pushing species to extinction.

Some birds in the collection are really old – this red-capped robin was collected from Australia in 1883.

Thanks Christina for inspiring this post!

What will be our final trace?

I met a traveller from an antique land
Who said: Two vast and trunkless legs of stone
Stand in the desert. Near them, on the sand,
Half sunk, a shattered visage lies, whose frown,
And wrinkled lip, and sneer of cold command,
Tell that its sculptor well those passions read
Which yet survive, stamped on these lifeless things,
The hand that mocked them and the heart that fed:
And on the pedestal these words appear:
“My name is Ozymandias, king of kings:
Look on my works, ye Mighty, and despair!”
Nothing beside remains. Round the decay
Of that colossal wreck, boundless and bare
The lone and level sands stretch far away.

- Percy Bysshe Shelley

 The legacy of Ozymandias provides an interesting metaphor for the fossil record. It is easy to recognise a living entity while it is still alive. It can be a little difficult to say that something was alive if we only ever saw it dead, and it becomes even harder to claim that something was alive if there is very little of it left. Time will eventually reclaim everything. So, what are the minimum traces that life leaves behind?

Biochemicals.

One group of biochemicals of particular interest are carotenoids. As the name suggests, carotenoids can be found in carrots – the carrot colour is ‘β-carotene’. Carotenoids are found elsewhere as well, and critically, they are only made by plants, algae, bacteria, fungi and one animal, an aphid. Carotenoids are not made by geological processes, but are instead evidence for life. Like all traces of life, carotenoids are eventually broken down and recycled by Earth processes. A plant rich with carotenoids will die and settle into the Earth, and even without the aid of bacteria, the atoms in the carotenoid will disassociate and break the biochemical apart. So, eventually these minimal traces of life will eventually fade away. These biochemicals break down into smaller and less complex molecules. So, carotenes in the fossil record, or carotanes – the breakdown products – are unambiguous signals that life once existed, even if those remnant molecules are all that remain. Like Ozymandias’ “…vast and trunkless legs of stone…”

Pigments like β-carotene are evidence for life, but these biochemicals can be altered during diagenesis.

So, when we investigate the time worn sediments from an aeons-old Earth, or Mars, we can look for carotenoid degradation products as vestiges of life. This was the focus of a 2010 study by Craig Marshall and Alison Olcott Marshall. Diagenetic alteration can result in “…hydrogenation of the polyene chain…”, the long carbon backbone of the carotenoid that gives the molecule it’s colour. In essence, the unaltered carotenoid has many carbon atoms bound to each other with ‘double’ bonds, but in the altered carotenoid, these carbon atoms are only bound with ‘single’ bonds. Marshall and Olcott Marshall (2010) describe the Raman spectra β-carotane and lycopane and show us what to look for in the fossil record. These altered carotenoids could be the only traces of a once teeming ecosystem, and we might otherwise never know about them if not for spectroscopy.

Marshall P, Olcott Marshall A. 2010. The potential of Raman spectroscopy for the analysis of diagenetically transformed carotenoids. Philosophical Transactions of the Royal Society A 368: 3137–3144.

Images compiled from Wikimedia Commons, here and here

Smithsonian Associates presentation July 10

I have found many stories recently that I want to share, including a great study on fossil evidence for pigments. I look forward to sharing these stories after July 10….after my evening seminar:

From The Smithsonian Associates website:

Stories in Stone: Illuminating Animal Fossils

Evening Seminar

Tuesday, July 10 – 6:45 p.m. to 8:45 p.m.

How can we know the color of the birds that filled the ancient skies, the daily diet of extinct creatures, or when the ancestors of today’s penguins became used to the cold? Clues to these mysteries—and many more—are buried deep in the chemical labels that outlast the lives of animals and persist into their fossil records.

Using spectroscopy, Smithsonian scientist Daniel B. Thomas becomes a time-travelling detective, uncovering the chemical secrets that tell long-lost animal stories. Tonight, he explores how the chemistry locked in fossils can reveal the diet, lifestyle, habitat, physiology, and even the color of creatures, allowing us to reconstruct ancient life in exquisite and fascinating detail.

6:45 to 7:45 p.m. Atoms, Isotopes, and Molecules

Extracting chemical evidence from fossils reveals a wide range of ecological information.

8 to 8:45 p.m. The Color of Birds

Birds use color—probably inherited from their dinosaur ancestors—for many functions, from the covert disguise of nightjars, whose feathers are patterned like leaf litter, to the overt and outlandish display of the peacock, whose patterns are a lure for females.

Thomas is a research fellow at the Natural History Museum

Red blooded Ötzi

Raman spectroscopy has revealed that red blood cells preserved in the Iceman still contain oxygen-transporting porphyrin molecules.

Ötzi the Iceman was an important member of his ancient Tyrolean tribe. He planned to return to his village, but he took an arrow to the… shoulder… and instead was mummified in a glacier until discovery in 1991. His body has been so well preserved over the last 5300 years that the collagen in his skin is still intact. Ötzi is still contributing to science, and a recent study from Marek Janko and colleagues analysed the blood in Ötzi’s veins (Janko et al. 2012).

Small tissue samples were taken from wounds on Ötzi’s right hand and left shoulder. The Ötzi samples and fresh human tissues (from a volunteer) were prepared so that the red blood cells could be studied. Three analytical techniques were applied to the tissues, including Raman spectroscopy. Raman spectra from the ancient tissues were very similar to the spectrum of modern human blood. The peaks in each Raman spectrum were characteristic of a porphyrin – heme – which proved two things. First, there is still blood frozen in Ötzi’s arteries and veins. Second, the molecule heme, which has the important role of transporting oxygen around a living body, can be preserved for 5300 years. The heme in Ötzi’s tissues has degraded slightly, but it is still the red blood pigment we are all familiar with.

Janko M, Stark RW, Zink A. 2012. Preservation of 5300 year old red blood cells in the Iceman. Journal of the Royal Society Interface.doi:10.1098/rsif.2012.0174

Images compiled from Wikimedia Commons ( here, here and here)

#Archives: Ancient plant pigments

Absorbance spectrum of a chlorophyll degradation product compared to a pigment from a 44 million year old leaf. This spectrum is good evidence that complex molecules like plant pigments can survive intact in the fossil record. Figure images from Dilcher et al. (1970) and Matile et al. (1999)

The fundamental difference between animals and plants is energy. Our source of energy is biotic – we consume things that were alive. The source of energy for plants is abiotic – they consume that which is not alive, namely, light. Plants harvest light and use it for photosynthesis, allowing them to convert CO₂ from the atmosphere into something nutritious. Chlorophyll is the key to photosynthesis on Earth, and is the fundamental difference between animals and plants. Chlorophyll is a green pigment that strongly absorbs blue and red light. Chlorophyll, like heme, is a porphyrin, which is based on four pyrrole subunits bound together into a macrocycle. The macrocycle is a basic support frame for a whole world of chemistry – side chains can attach to the outside of the macrocycle, and the space in the middle can house a metal ion. Chlorophyll a, for example, has a specific combination of side chains and houses a magnesium ion; chlorophyll b and chlorophyll d have slightly different side chains. The macrocycle gives chlorophyll its green colouration, and the chemistry of chlorophyll is critical to life on Earth as we know it. Chlorophyll is a sophisticated molecule that is expensive to produce, and rapidly disappears when the tissue dies. Imagine, then, how exciting it would be to discover green fossil leaves in a 44±4 million year old coal deposit.

“Green-colored angiosperm leaves were reported in 1931 by Weigelt and Noack from middle Eocene brown coals of the Geisel valley near Halle, East Germany…” This is the opening line from a report by David Dilcher and colleagues, published in 1970. Dilcher and colleagues describe how “…Weigelt and Noack identified several chlorophyll derivatives from crude extracts of these green fossil leaves and their associated brown coals…”, and “…[a] reinvestigation of this material by modem techniques of chromatography and spectrophotometry has made possible a more precise separation and identification of the green pigmentation of this material…” So, green fossil leaves were found in a German coal deposit, and initial studies suggested that the green pigment was chemically similar to chlorophyll. Building on this initial work, Dilcher and colleagues set out to discover whether the remarkable green of these ancient leaves is the same pigment we find in leaves today.

Dilcher and colleagues sought information from a suite of analytical techniques, including paper chromatography, Molisch phase test, HCl number test, mass spectroscopy, and absorbance spectroscopy. I will focus on the last of these, seeing as light absorption is the biological imperative of green plant pigments. The green pigment was extracted and the absorbance spectrum was measured. Light absorption is based on a simple concept that I will over-explain. Light is part of the electromagnetic spectrum, which can be described as a continuous range of wavelengths, from high energy x-rays, to low energy radio waves (and beyond). Light is the small part of the electromagnetic spectrum that we can detect. We perceive higher energy wavelengths as blue, and lower energy wavelengths as red, but that is as far as we go. Wavelengths with slightly higher energy than blue are called ultraviolet, which we cannot see but birds can, and wavelengths with slightly lower energy than red are called infrared, which are invisible to us and birds. Now, different materials absorb different wavelengths, which give rise to different colours through a complimentary arrangement. The absorption of different wavelengths is characteristic of a material, and can be used as a spectral fingerprint. Dilcher and colleagues shined ultraviolet, visible and infrared light at their mystery green compound from the leaf fossil, and measured the wavelengths that were absorbed.

The absorbance spectrum of the green pigment was an almost perfect match with methyl phaeophorbide a, which is a degradation product of chlorophyll a. The key difference between chlorophyll and phaeophorbide is a long phytol chain, which is lost when chlorophyll begins to break down. Phaeophorbide is still green, but it is a slightly different green to chlorophyll, and the slight differences can be quantified with an absorbance spectrum. Which is exactly what Dilcher and colleagues did, allowing them to show that methyl phaeophorbide a is preserved in a 44±4 million year old coal deposit.

From Dilcher et al. (1970): “…In the Eocene the Geisel valley was a poorly drained shallow basin, receiving organic sediments from plants and animals living along its swampy margins…The preservation of such fossils as frog epidermal cells containing nuclei and bacteria in the Geisel brown coal suggests an anaerobic environment with relatively rapid deposition, in which organic decay was slow and organic accumulation extensive. The rapid burial of plant material in an anaerobic environment and the fact that brown coal has a history of low temperatures may account for the preservation of this phorbin from the middle Eocene…”

Dilcher DL, Pavlick RJ, Mitchell J. 1970. Chlorophyll Derivatives in Middle Eocene Sediments. Science 168 1447–1449.

Matile P, Hörtensteiner S, Thomas H. 1999. Chlorophyll degredation. Annual Review of Plant Physiology and Plant Molecular Biology 50:67–95

Weigelt J, Noack K. 1931. Nova Acta Leopoldina 1, 87.