Bone, Trace-Element Analysis

The use of certain chemical elements in bones and teeth of humans as tracers of
the source of foods. Bones and teeth consist of crystals of calcium phosphate
(apatite) intimately intergrown with an organic matrix, largely composed of
protein (collagen and minor amounts of noncollagenous protein). The mineral
component has a well-defined chemical composition, made up of calcium (the
chemical symbol for which is Ca), phosphate (PO4), hydroxyl (OH), and
carbonate (CO3). In addition, variable amounts of several trace elements can
sub- stitute for these major constituents. The amounts of these trace elements in
fossil bone depend, in part, on diet and have been used to reconstruct paleodiet.
Many of these trace elements, such as strontium (Sr), barium (Ba) and lead
(Pb), do not appear to be essential to the diet, but rather enter the body
passively and can be trapped at atomic sites in the bone mineral that are
normally occupied by Ca. The elements Sr and Ba resemble calcium in their
chemical behavior and are grouped together with Ca in the periodic table as
alkaline earth elements. Therefore, they have a particularly high affinity for bone
mineral. Usually, levels of trace elements in bones are reported as the ratio of
the concentration of the element to that of Ca (e.g., the Sr/Ca ratio).

Analyses of bone can be carried out on whole bone or bone powder using the
nondestructive analytical method of X-ray fluorescence (XRF) or, destructively,
by neutron activation analysis (NAA), atomic absorption (AA), or inductively
coupled plasma (ICP) spectrometry. These methods are widely available in
Departments of Geology where geo-chemical studies are done and, to a lesser
extent, in anthropology research laboratories. Concentrations of trace elements
are generally given in units of parts per million (ppm) (1 ppm=0.0001 percent).

All animal and human bodies tend to reject nonessential elements during
assimilation of nutrients. Therefore, the relative concentration of these elements
is less in the consumer than in the food consumed. For example, the Sr/Ca ratio
in an animal is about half of that in the food it consumes. This process of
exclusion of nonessential trace elements is referred to as biopurification. The
lowered interelement ratios are reflected in the composition of bone. In principle,
this phenomenon can be utilized in the study of the trophic level of an individual
—that is, where it lies on the gradient from herbivores (lowest trophic level) to
pure carnivores (highest). At each successively higher trophic level, a consumer’s
body (including its bones and teeth) should exhibit a lower Sr/Ca or Ba/Ca ratio
than that of its foods. There are a number of assumptions and interfering effects
that must be considered, however.

First, the Sr/Ca level in any organism in a given region is determined by the
average Sr/Ca ratio in the local environment, which can vary considerably as a
function of the local rock and soil types. Therefore, the Sr/Ca trophic gradient at
a site should always be referred to values for known herbivores and carnivores
for that locality. It is unsafe to assume an average, “standard” Sr/Ca ratio for a
site, since this ratio can vary by as much as a factor of 10, even over short
distances. On the other hand, values can be regionally uniform to within a few
percent, as in Britain, where Sr/Ca ratios do not deviate by more than 25 percent
from a regional value of 3×10−4 (molar ratio). Equally important, the Sr/Ca ratio
in a particular consumer depends on the average Sr/Ca ratio of the specific
mixture of foods consumed. Two species at the same trophic level living in the
same environment can, therefore, have different Sr/Ca ratios.

Another problem that affects all studies of trace elements in bone, is diagenesis.
During their period of burial in archaeological sites, bones do not remain inert.
Except in desert or permafrost areas where liquid water is absent, bones undergo
continuous chemical transformations, the extent and character of which critically
depend on their burial environment. The major and minor chemical constituents
of the bone mineral can change as a result of chemical interaction with soil
water, which is a dilute solution containing Sr, Ca, and many other elements at
significant levels. As a result, the Sr/Ca ratio of a fossil bone can become quite
different from that which was present at the time of death. New crystals of
phosphate mineral are formed during diagenesis, locking in the new trace-
element constituents so that the contamination cannot be easily removed.
Certain elements such as silicon, iron, manganese, and aluminum are enriched in
sediments but normally absent in living bone. The levels of these elements can
be used as indicators of the degree of diagenesis.

Various strategies have been attempted to counteract the effects of diagenesis.

Andrew Sillen has shown that one can partly dissolve (leach) the bone with
successive portions of dilute acid and observe gradual changes in the Sr/Ca ratio
in successive leachates. This ratio generally starts at some high (contaminated)
value and gradually drops to a constant “plateau” that is believed to be the initial
value for the living bone. This seems to yield satisfactory data in some cases but
has also given anomalous results. Another approach has been to analyze tooth
enamel. This material is less porous, denser, and contains less organic matter
than bone. It is believed to be more resistant to diagenesis and has also been
preferred for some stable-isotopic analyses. Most dental enamel (except for the
third molar) is, however, formed very early in the life of the animal or human,
and its composition is not representative of average lifetime diet. Other workers
have suggested the selective use of the interior part of cortical bone, noting that
compositional gradients can be detected for some elements such as uranium,
with higher concentrations occurring near the outer surface of the cortex.

Another possible interfering effect is the variation in trace element content

between different skeletal elements of the same individual. Limited studies of
some living and ancient populations have shown significant intraskeletal
differences. These may be partly a result of changes in diet through the lifetime
of the individual, coupled with bone remodeling, which affects different skeletal
elements to varying degrees. Likewise, intraskeletal variations seen in fossil or
ancient bones may be due to differential degrees of diagenesis as a result of
variation in initial bone density.

Sr/Ca has been used as a trophic-level indicator to evaluate the diet of early
hominids (Boaz and Hampel, 1978), as well as to study later hunter-gatherers,
pastoralists, and farming communities. Problems of diagenesis have generally
discouraged its use, although the leachate method shows some promise.
Barium (Ba), which is also chemically similar to calcium, is also potentially useful
for characterization of paleodiet, through the principle of biopurification. In fact,
the largest differences in Ba concentrations between populations are found where
there is a contrast between terrestrial and marine (coastal) populations. Barium
is extremely insoluble in solutions containing sulfate (SO 4), such as seawater.
The Ba/Sr ratio of seawater and foods that grow in it is much lower than that of
terrestrial waters and foods. James Burton and Theron Douglas Price (1990)
showed that even fossil bones preserved this great difference in Ba/Sr between
coastal and terrestrial populations, although the actual values gave only general
estimates of the percentage of terrestrial foods in the diet.

Many other trace elements can be easily analyzed in bone using AA, ICP, and
NAA, and concentrations of many of these have been studied in the hope of
obtaining further information about the diet of the individuals. In particular,
certain foods, such as shellfish and nuts, are notably enriched in certain trace
elements (vanadium, copper, zinc, manganese), and concentrations of these
elements in bone were believed to be indicators of the level of consumption of
certain foods. Studies of these elements in fossil bones (as well as other tissues,
where preserved, such as hair and skin) have made use of various statistical
procedures, including factor analysis, to search for key variables or combinations
of variables that could be used to characterize populations.

Recently, however, systematic feeding studies (Lambert and Weydert-Homer

1993) have shown that concentrations of many trace elements in bone bear no
relation to the content of these elements in diet. Indeed, the presence of various
organic constituents, such as indigestible fiber, in the diet seems to have a larger
effect on the concentration of these elements in bone than does their
concentration in the diet. On further reflection, this is not surprising, since all
animals tend to control their internal chemical state (homeostasis), and this
control might be expected to extend to the concentrations of trace elements.

We can distinguish among three types of chemical elements with respect to their
behavior in the body. Elements such as copper and vanadium, which are
biologically inactive and are chemically dissimilar to Ca, are assimilated from the
gut at very low levels and, therefore, are greatly depleted in the blood and
tissues with respect to their concentration in foods. On the other hand, calcium,
strontium, and barium resemble one another chemically, and the body is,
therefore, less effective at excluding these minor elements from bone mineral.
Their concentration in bone is proportional to their content in the diet. Finally,
some trace elements are biologically active, and their concentration is
maintained at a constant level by biochemical systems in the body; these include
cobalt, zinc, and iron. Within certain broad limits, concentrations of these
elements in bone should not depend on the level in diet. Extreme depletion of
these elements in the diet can lead to pathologic effects that might be coupled
with abnormally low levels of these elements in bone. Organic constituents of
certain foods are able to form stable complexes(chelates) with some essential
trace elements. For example, phytate, which is found in some grains, forms a
stable complex with zinc and can prevent absorption of this element by the gut.
Excessive consumption of some grains may lead to dietary deficiency in elements
that have been chelated in this way.

The bone concentrations of two other elements, lead (Pb) and uranium (U), have
been studied extensively. Although chemically dissimilar to the alkaline earth
elements, the mineral phase of bones and teeth has a strong affinity for both.
Living organisms take up lead in their bones in direct response to its
concentration in the environment. Lead serves no known biological function.
Rather, it is a highly toxic metal that can produce neurological disorders and
general morbidity. Numerous studies have followed the variation of Pb content in
ancient bones and shown that higher concentrations of Pb are found in members
of complex societies in which metal production is used, while individuals from
agricultural or hunter-gatherer societies have low levels of lead.

Unlike lead, the uranium content of living bone is generally almost undetectable
(less than 0.01 ppm), whereas ancient bone typically contains from 1 to 100
ppm of U. The higher concentration of U in older, buried bone is due to
diagenesis and the uptake of U from groundwater. Bone has a particularly high
affinity for this element (some uraniumore deposits are concentrated in bone-rich
sedimentary layers). Early studies showed that it was possible to obtain crude
estimates of the age of buried bone from the U content. More precise ages are
obtainable by measurement of the radioactive decay of uranium to its daughter
isotope, thorium-230. Uranium/thorium ages determined on bones or teeth have
to take into account the gradual uptake of U by the bone and the possibility of
periods of U loss as well as uptake.
Ancient Nubians Made Antibiotic Beer

Chemical analysis of the bones of ancient Sudanese Nubians who lived nearly 2000 years ago
shows they were ingesting the antibiotic tetracycline on a regular basis, likely from a special brew
of beer. The find is the strongest yet that antibiotics were previously discovered by humans
before Alexander Fleming discovered penicillin in 1928.
“I’m going to ask Alexander Fleming to hand back his Nobel Prize,” joked chemist Mark Nelson,
who works on developing new tetracyclines at Paratek Pharmaceuticals and is lead author of the
paper published June in the American Journal of Physical Anthropology.

Nelson found large amounts of tetracycline in the bones tested from the ancient population, which
lived in the Nubian kingdom (present day Sudan) between 250 A.D. and 550 A.D. and left no
written record.

“The bones of these ancient people were saturated with tetracycline, showing that they had been
taking it for a long time,” Nelson said in a press release August 30. “I’m convinced that they had
the science of fermentation under control and were purposely producing the drug.”
“This discovery will provide a whole new framework for understanding the relationship between
microbes and antibiotics,” said anthropologist Dennis Van Gerven of University of Colorado at
Boulder. “There might have been other populations that were also doing the same thing,
anywhere that there were these microbes. This is going to drive other scientists to start this
search, and that is incredibly important.”

Scientists have
suspected this population was ingesting tetracycline since they first noticed a florescent yellow-
green appearance of the bones under ultraviolet light, indicative of tetracycline.

“When we reported that in 1981, it was met with a lot of skepticism,” said anthropologist George
Armelagos of Emory University, who made the original discovery and is co-author of this new
study. “If you were unwrapping an Egyptian mummy and suddenly it had Ray-Ban sunglasses on
it, that’s what it was like with us.”

Tetracycline latches on to calcium and gets deposited in bones, which is how it can be detected it
in fossils. The ultraviolet light technique said little about how much tetracycline there was in the
bone, and it was hard to convince others it wasn’t simply a produced of microbial contamination
of the bones, or aone-time beer event.

Nelson was able to solve the problem by dissolving the bones in hydrogen flouride, the nastiest
acid on the planet, he said. He was able to clearly identify the amount and identity of the
tetracycline in the bones. It was in all the bones, including those of a four-year-old child.

Armelagos, who specializes in reconstructing ancient diets, proposed that the Nubians made the
tetracycline in their beer. There is evidence they knew how to make it, he says. Tetracycline is
produced by a soil bacteria called streptomyces, which is how it was discovered by modern
society in the 1940s. Streptomyces thrives in warm, arid regions such as that of ancient Nubia,
and likely contaminated a batch of beer.

They must have known how to propagate the beer because they were doing it to make wine,
Nelson says. There was also so much of it in their bones that it is near impossible that the
tetracycline-laced beer was a fluke event.
To make sure that making the antibiotic beer was possible, Armelagos had his graduate students
give it a try.

“What they were making wasn’t like a Bud Light but a cereal gruel,” Armelagos said. “My students
said that it was ‘not bad,’ but it is like a sour porridge substance. The ancient people would have
drained the liquid off and also eaten the gruel.”

The Nubians likely noticed the antibiotics cured them of bacterial infection. It may have had
negative effects as well: If taken in too large quantities the antibiotic can also cause iron
deficiency because it latches on to the iron in the body.

Streptomyces produces a golden-colored bacterial colony that would have floated on top of the
beer and likely encouraged its propagation. Gold was revered by the ancient cultures.

When and why the antibiotic beer secret was lost is a mystery. It is not the first technology to
disappear with the disappearance of cultures. Armelagos is continuing to look for the tetracycline
in the bones of different cultures. He says he has already found preliminary evidence it is in
bones of people who date to as late as 1300 to 1400 A.D.

Armelagos hopes this find might also help explain why animals have been found with antibiotic
resistance in Northern Africa where there is no previous evidence of antibiotics being used.

About a quarter of the grain in Africa is still made into beer, Armelagos says.

Diet and Dentition- A Chemical Analysis

With an anthropology and biochemistry double major, I have an interest in

the application of chemical analysis for forensic anthropology. For my senior
project in anthropology I will be using chemical analysis of bone and dental
remains to try to determine the diet of the humans in past cultures. Through
this chemical analysis I should be able to determine the main components of
the diet of the human I will be analyzing.

The different diets that can be determined include primarily marine, meat, or
plant diet. The diets that consist primarily of vegetation can further be
separated into two types according to the photosynthetic pathway followed-
plants which follow the Calvin-Benson, or C3, cycle, and plants that follow the
Hatch-Slack, or C4, cycle. C3 plants are called such because they produce an
intermediate molecule with 3 carbons during photosynthesis. These plants
include trees, shrubs, flowering plants and temperate zone grasses. C4 plants
produce an intermediate molecule with 4 carbons. There are about 10 families,
these include tropical grasses, maize, cane sugar, and some types of millet,
sorghums, amaranths and chenopods. The carbon plants use in photosynthesis
is taken in from atmospheric carbon dioxide. It is therefore important to note
whether the remains are from a time before the 1850’s because the
atmospheric carbon dioxide content was relatively stable until the industrial
revolution, in the 1850’s, when it started to steadily increase.

In order to analyze the diets of the past, we must first know what to look for.
When using bone and dental samples we tend to look at the stable isotope
ratios of carbon and nitrogen. This is especially useful when working with
collagen from bones. Collagen is an organic component of the bone which is
generally well preserved in sufficient quantities. Collagen extract is generally
known as gelatin. The one possible downfall of using collagen for analysis is
that in life it takes up to ten years to replace. This means any short term or
seasonal changes in diet will not show up, however this can also prove to be
useful if the study is aimed at an average diet and not a seasonal diet.

Atmospheric carbon dioxide has a stable ratio of carbon-12 and carbon-13,

therefore plants have a stable ratio, which is incorporated into plant tissue.
These plants are then digested by animals, human or otherwise, a percent of
this is then used by the animal for tissue. If an animal, such as a human, eats
an herbivore they will then incorporate some of the C-12 and C-13 into their
own tissue, giving another ratio. Carbon ratios for samples are obtained
through the use of a mass spectrometer. This is a mechanical chemical
analyzer. When using the mass spectrometer the sample material must be
tuned into its gaseous form. In order to do this the sample usually collagen, is
heated at very high temperatures- often 900 degrees Celsius. The sample gas is
then injected into the mass spectrometer where the elements that make up the
gas are separated according to density. They are analyzed in the mass
spectrometer by nucleic mass, therefore isotopes can be separated and the
number of each one can be compared, giving a ratio.

The ratios for the samples are then compared to a standard using this
mathematical equation:

The results are expressed in parts per thousand. The more negative a result
indicates a lower amount of carbon- 13 in the sample. As this graph shows,
through experimentation we have found average ratios for diets of C3 plants
only, meat from herbivores on C3 plant diets, C4 plants only, meat from C4
herbivores, marine plankton only, meat from marine herbivores and, finally,
meat from marine carnivores.

Diet Average Carbon Ratio Expected Consumer Ratio

C3 plants only -26.5 -21.5
Meat from C3 herbivores -25.5 -20.5
C4 plants only -12.5 -7.5
Meat from C4 herbivores -11.5 -6.5
Marine Plankton only -19.5 -14.5
Meat form Marine -18.5 -13.5
Herbivores
Meat from Marine -17.5 -12.5
Carnivores

Using such information we can determine if the diet of a human was more
likely one of these or if it was a combination of these. For instance, if the
carbon ratio for a human was found to be in between C3 diet and the diet of
meat from C3 herbivores, the most logical would be that the human was an
omnivore.

Nitrogen isotopes are used in much the same way as carbon isotopes. Nitrogen
is taken up by plants through the nitrates and nitrites in the soil, as opposed to
the atmospheric source of carbon. The ratio of nitrogen- 15 and nitrogen- 14
of the sample is measured using a mass spectrometer. This is then compared to
the standard using the same mathematical equation as carbon isotopes. One of
the first applications of nitrogen isotopes was in identifying legumes in
terrestrial diets. Nitrogen was useful because legumes have a lower nitrogen-
14 to nitrogen-15 ratio due to nitrogen fixation. Nitrogen fixation by legumes
do not have to rely on the soil for nitrogen, therefore they will receive a lower
ratio of isotopes. The ratio of nitrogen isotopes is also used to determine
terrestrial or marine protein. In studies done on historic and prehistoric
populations there was a significantly higher nitrogen ratio in people with a
primarily marine diet. Not only can nitrogen isotopes help to determine
marine or terrestrial diets, it can help distinguish terrestrial carnivorous or
herbivore diets. Terrestrial carnivores average a higher nitrogen ratio than
terrestrial herbivores. Studying nitrogen isotope ratios can help identify some
major aspects of diet. However, one aspect of diet this method cannot
distinguish between as readily is terrestrial and freshwater diet.

Unfortunately, there are still some questions and problems with determining
diet using stable isotopes. Because this is a relatively new approach, there are
no set standards on the technique, such as heating time or temperature of the
sample before mass spectrography, or storage of samples. Long term storage
of non-gaseous samples is also important, improper storage could introduce
contaminants such as bacteria or mold. There has only been limited work on
standards of carbon from collagen, this means that individual labs may be
using their own standards. This can cause problems in that each lab, which
tests the same sample, can obtain a different ratio due to different standards.
When working with nitrogen isotopes, one aspect that researchers have to
remember is that there can be different levels of nitrates and nitrites in soil in
different areas. If the soil level is not tested, the data may be skewed.
Scientists are also looking at using tooth remains by identifying phytoliths,
silica impressions of plant cells which can be found on teeth. However, this
technique is still in its infancy.

Although chemical analysis of remains to determine diet is a relatively new

technique, it is quickly becoming useful in the field of anthropology. Since it’s
beginning in the 1970’s it has helped in proposing a tentative timeline for the
development of certain types of agriculture, specifically maize, and proposing
diets of past cultures. Due to some of the problems in the young field, it would
be safest to use archaeological evidence, such as food remains, in conjunction
with the chemical analysis. It would also be best to use carbon and nitrogen
isotopes in conjunction with each other- this could limit uncertainties because
the analysis would overlap in some areas. Once this technique is perfected, we
may be able to discern much knowledge from a single piece of bone.

Bibliography

Bower, B. Tooth analysis May Decipher Prehistoric Diets

Ed. Price, T. D. The chemistry of prehistoric Human Bone University Press,

Cambridge 1989.

Schoeninger, M.J., De Niro, M.J. Stable Nitrogen Isotope Ratios of bone

Collagen Reflect Marine and Terrestrial Components of Prehistoric Human
Diet Science Vol 220, No 4640, June 24, 1983.

Sillen, A., Sealy, J.C., van der Merwe, N. Chemistry and Paleodietary
Research: No More Easy Answers American Antiquity Vol 54, No 3, 1989.

Note: Subjects for the project have been changed from humans to animals.

Special thanks to :

Matthew T. Millard

Tisha Strachan

Author: Dena Connors-Millard

Hemijske analize kostiju

Čitav princip hemijske analize kostiju je zasnovan na jednostavnoj činjenici da ono što jedemo ostavlja traga u

našim kostima. Određeni elementi i hemijska jedinjenja, karakteristična za specifične namirnice kroz biohemijske

procese se trajno zadržavaju u koštanom tkivu.

Koncentracija različitih elemenata varira u životinjskim i biljnim vrstama što će dovesti do toga da se u sadržaju

čovekovih kostiju elementi detektuju u različitim koncentracijama u zavisnosti od hrane koja je unošena u

organizam. Npr. možemo analizirati nivo stroncijuma za koji znamo da je značajno prisutan kod morskih riba i

školjki, dok je mnogo niži kod rečnih riba.

Takođe, u rekonstrukciji ishrane mogu se analizirati i stabilni izotopi, najčešće ugljenika i to  najviše u

rekonstrukciji biljne ishrane. Odnosi stabilnog izotopa ugljenika u biljkama variraju u odnosu na vrstu fotosinteze

koju koriste biljke. Tako od vrste biljke zavisi i sam proces fotosinteze a od njega i odnosi stabilnog izotopa

ugljenika. Zbog toga je, u nekim slučajevima, moguće utvrditi koja vrsta biljne hrane je preovlađivala u ishrani

određene osobe.

Analiza drevne ishrane ne samo da nam uvid u privrede određenog društva već otvara široko polje mogućih

drugih istraživanja. Npr. možemo analizirati da li je ishrana u prošlosti zavisila od pola ili društvenog statusa

pokojnika, ali i to da li je vrsta hrane uticala na dužinu ili kvalitet života određene osobe.

Dakle, u kostima je prisutna gotovo čitava biografija - od toga kada je neki čovek živeo, koliko je godina imao,

kako je izgledao, koliko su mu bili aktivni mišići, od čega je bolovao, kako se hranio i kakva je bila njegova

genetska struktura. I upravo zbog toga, ako brižljivo analiziramo kosti, možemo čuti kako govore i pričaju

uzbudljive priče o ljudima i događajima iz prošlosti.

C3 and C4 Plants at Archaeological Sites

A second way carbon is useful is in dietary analysis, since not only do humans burn or cut down living objects
(trees, plants) as necessary, but they also consume carbon in different ways. One of the most investigated areas
in archaeology is the transition in diet, including the advent of agriculture. Within the plants around the world,
there are two many types: C3 and C4 plants. The C refers to the carbon chain in the plant. C3 plants are older
plants, while C4 plants are a more recent adapted plant type.

C3 plants are most of the plants growing around the world, as they are quite flexible in what type of environments
they need, as long as the temperatures and rainfall totals are moderate. C3 plants include potatoes, rice, and
wheat.

C4 plants include most grasses, sugar cane, maize, and millet. C4 plants have a different outer protection layer of
plant fibers. These help keep in moisture in times of dry spells. They also may have a higher starch content.

This is where archaeologists find carbon levels useful. Knowing the carbon fixation of a plant helps determine the
likely foods eaten in the past.

Archaeology and Strontium Isotopes

In diet reconstruction, strontium levels are also significant in archaeological research. Strontium levels vary across
the world, and in some areas, are distinctive to specific smaller regions. Strontium levels vary in nature, and end
up in humans who reside in the region.
The fact that the human has a similar isotopic signature to strontium levels of a geographic location means that
person very likely was born or lived in that area for a length of time and was exposed to the strontium there.
Strontium adheres to bones and teeth given its likeness to calcium.

As a result, the bones found at an excavation site may be taken to a laboratory and analyzed for strontium levels.
The area this person lived in for a long time may be discernible, although not all of the time. This information may
help in identifying someone as a longtime resident (or conversely, as a transient or visitor).

Although at first glance archaeology and chemistry seem very separated, with one being a social science and the
other being a physical science, there is a great deal of overlap when it comes to how humans interact with their
surroundings. Carbon studies help in determining relative age of a site, as well as possible diets. Strontium helps
determine place of origin or homeland of a long-deceased individual. All of this gleans more information about a
past society and its environment when it comes to the final archaeological site report.

Bone Chemistry
Principles of Stable IsotopeAnalysis 
Analytical Methods 
Recent Applications 
References 
Laboratory Staff and Students 

Principles of Stable Isotope Analysis

Carbon and nitrogen isotope ratios in human bone may be used to reconstruct prehistoric diet because of
differential fractionation, between certain plant groups, of atmospheric carbon dioxide during photosynthesis and
of nitrogen during fixation or absorption. There are two stable isotopes each of carbon (12C, 13C) and nitrogen
(14N, 15N), with 12C and 14N by far the most common in nature. Small differences in the ratios of these
isotopes (12C/13C, 15N /14N) can be measured by isotope ratio mass spectrometry using samples smaller than 1
milligram. High precision isotope measurements are reported using the delta notation (d13C, d15N) relative to
internationally recognized standards and are expressed in parts per thousand or per mil (‰).

Experimental data have indicated that different bone tissues reflect different components of the diet (Ambrose
and Norr 1993; Tieszen and Fagre 1993). In general, bone collagen is disproportionately produced from the
protein portion of the diet, while bone carbonate and tooth enamel carbonate (both a calcium hydroxyphosphate,
called apatite) are produced from a mixture of dietary protein, carbohydrates and fats. Stable isotope analysis of
both bone collagen and apatite thus permits quantitative estimates of several dietary components. Both bone
collagen and bone apatite are constantly being resorbed and replenished, so that their isotopic composition
reflects dietary averages over at least the last several years of an individual's life, while the composition of tooth
enamel will reflect diet during the age of crown formation.

Since teeth form at different ages (beginning in utero for deciduous teeth, and from 0 to 12 yrs of age for
permanent teeth), the analysis of multiple teeth from the same individual may reveal dietary shifts caused by
weaning (first the introduction of solid foods and later the cessation of breastfeeding) (Wright & Schwarcz
1998), while significant differences between tooth (juvenile diet) and bone (adult diet) values may be the result
of subsistence changes due to migration. Analysis of hair segments also may reveal short-term or seasonal
dietary changes (O'Connell & Hedges 1999). Overall, stable isotope analysis of multiple tissues can provide a
quantifiable dietary life history of an individual (Sealy et al. 1995), whereas subsistence data from faunal and
archaeobotanical remains typically derive from at least multiple individuals, often over multiple generations.
While collagen is rarely preserved in bones predating the Upper Paleolithic, and is often badly degraded in hot
and moist environments (e.g. lowland Mesoamerica), bone apatite has provided reliable results for the Holocene,
and tooth enamel for early hominids (e.g.Sponheimer & Lee-Thorp 1999) and into the Miocene.

Typically, grasses originally native to hot, arid environments follow the C4 (Hatch-Slack) photosynthetic
pathway, and will have d13C values averaging about -12.5‰; trees, shrubs, and grasses from temperate
regions,which follow the C3 (Calvin-Benson) photosynthetic pathway, will have d13C values averaging about
-26.5‰. In some forested areas, a canopy effect occurs due to incomplete atmospheric mixing, resulting in even
more negative carbon isotope ratios (van der Merwe & Medina 1991). Empirical data for large mammals
combined with experimental data for rats and mice indicate that bone collagen is enriched about +5‰ relative to
diet, although this value is affected by the proportion of protein in the total diet and differences in d13C values
between protein and energy sources (van der Merwe 1989; Ambrose & Norr 1993; Tieszen & Fagre 1993).
Experimental data on rats also convincingly demonstrates that bone apatite is enriched about +9.5‰ relative to
the whole diet, regardless of the mixture or isotopic composition of the foods consumed (Ambrose & Norr
1993), although empirical data for larger herbivores suggest the diet-apatite spacing is about +12‰.  Stable
carbon isotope analysis is particularly useful in New World dietary studies since maize is often the only C4 plant
contributing significantly to human diets; its contribution to bone collagen and to bone apatite may be estimated
by interpolation. Some caution is warranted, however, if succulent plants were present, since they utilize the
alternative CAM (crassulacean acid metabolism) photosynthetic pathway which results in carbon isotope ratios
similar to those of C4 plants. Nevertheless, CAM plants are unlikely to have been major sources of dietary
protein, whether consumed directly or indirectly through herbivorous faunal intermediaries.

The carbon isotope ratios of marine and freshwater organisms are more variable, depending on local ecological
circumstances, and often overlap with those of terrestrial plants and their consumers. These foods typically have
much higher nitrogen isotope values, however, and their high protein content will contribute much more carbon
to bone collagen than will maize (about 10% protein) or other plant foods. The analysis of bone apatite,which is
derived from all food groups, should allow the identification of just a few percent maize or other C4 resources in
an otherwise C3-based diet.

The nitrogen isotope ratios for plants depend primarily on how they obtain their nitrogen - by symbiotic bacterial
fixation or directly from soil nitrates - and these values are similarly passed along through the food chain
accompanied by an approximately 2-3‰ positive shift for each trophic level, including between mother and
nursing infant. Human consumers of terrestrial plants and animals typically have d15N values inbone collagenof
about 6-10‰ whereas consumers of freshwater or marinefish, seals and sea lions may have d15N values of 15-
20‰ (Schoeninger and DeNiro 1983). While most plants follow either the C3 or C4 photosynthetic pathway and
have similar carbon isotope ratios in most ecological settings, nitrogen isotope ratios vary according to rainfall,
altitude and other factors (Ambrose1991), and both carbon and nitrogen isotope ratios vary considerably among
marine organisms (Schoeninger and DeNiro 1984).

It is critical, therefore, to establish a site-relevant isotopic baseline for interpreting human skeletal data. Analyses
of faunal remains provide a good estimate both of the animals themselves and the plants they consume.
Establishing an isotopic baseline is particularly important in coastal areas where marine and riverine resources,
as well as C4 and/or CAM plants may have been available for direct or indirect consumption by
humans.Atmospheric carbon isotope ratios have become depleted by about 1.5‰ since the industrial revolution,
so values obtained on modern terrestrial plants and animals must be adjusted accordingly for most archaeological
studies.

Other surveys and summaries of stable isotope analysis in archaeology may be found in Schwarcz &
Schoeninger (1991); van der Merwe (1992); Ambrose (1993); Schoeninger 1995; Katzenberg & Harrison
(1997); Larsen (1997); Tykot (2004), and Tykot (2006).
 

Analytical Methods
My experience in bone chemistry dates to 1990 when I became the manager of the Archaeometry Laboratories in
the Anthropology Department at Harvard University. Professor Nikolaas J. van der Merwe, the new director of
the laboratory, was the first archaeologist to use stable isotopes, and the methods I learned were adapted from
those in use in his laboratory in Cape Town, where he has recently returned as Professor of Natural History. In
my lab at USF, we extract bone collagen using well-established laboratory procedures (see also Ambrose 1990).
Whole bone is demineralized in 2% hydrochloric acid (72 hrs), base-soluble contaminants are removed using 0.1
M sodium hydroxide (24 hrs each before and after demineralization),and residual lipids are dissolved in a 2:1:0.8
mixture of methanol, chloroform,and water (24 hrs). Freeze-dried collagen pseudomorphs are analyzed for d13C
and d15N in continuous flow mode using a CHN analyzer coupled with a Finnigan MAT stable isotope ratio
mass spectrometer located at USF. Collagen yields and C:N ratios are used to confirm the integrity of collagen
samples.

  bone collagen pseudomorphs   the

archaeological science laboratory in SCA

Bone apatite and tooth enamel samples are prepared using procedures designed to remove non-biogenic carbon
without altering the biogenic carbon isotope values (cf. Koch et al. 1997). Powder samples are obtained by
drilling from the center of carefully cleaned bone samples,or from tooth enamel after the surface layer has been
removed. Approximately 10 mg of powder are immersed in 2% sodium hypochlorite to dissolve organic
components (24hrs for enamel, 72 hrs for bone apatite). Non-biogenic carbonates are then removed in 1.0 M
buffered acetic acid for 24 hours. The integrity of apatite and enamel samples is assessed through yields obtained
in each stage of the pretreatment process. Samples are analyzed on a second Finnigan MAT mass spectrometer
equipped with a Kiel III individual acid bath carbonate system.

The analytical precision for stable isotope ratio mass spectrometry is ± 0.1‰ for d13C and ± 0.2‰ for d15N. 
 

Recent Applications

I have been involved in a number of collaborative research projects around the world using stable isotope or
elemental analysis to assess ancient diets. Preliminary results have been presented at various conferences and
some have been published or are in press (see my list of 'publications' on this website). A brief outline of these
projects and the results obtained is given below.

Belize. Skeletal remains from the Preclassicsite of Cuello, excavated by Norman Hammond of Boston
University, have been analyzed for bone collagen, bone apatite, and tooth enamel (Tykot et al. 1996), as have
samples from Classic sites at La Milpa (Hammond excavations) and the Programme for Belize (Fred
Valdez/Richard Adams excavations). In conjunction with data published by others, a chronological trend
towards increasing maize reliance is evident both in bone collagen and bone apatite. At Cuello, and probably
other sites, some of the maize signature likely came from the consumption of maize-fed dogs. At La Milpa, diets
also seem to vary based on social status and residence location. 
 
Guatemala. At Iximché in highland Guatemala, a number of individuals dating to the Late Postclassic period
have been tested for bone collagen (by David Reed and Stephen Whittington) and tooth enamel. The results
reveal an extremely high dependence on maize, more so than most other Guatemalan populations at least of the
Late Classic period, and certainly more so than contemporary populations at Lamanai in Belize. This high
dependence may be related to a combination of local ecological as well as socioeconomic factors. Within the
group of individuals tested, most of whom were decapitated and may have formed a skull rack (tzompantli),
several individuals standout as having significantly different diets. We suggest that they may have been warrior-
elites from other communities who consumed different diets either because of differences in local ecology,
and/or had access to different foods because of their high status. A full report of our study ( Analysis of
Kaqchikel Skeletons: Iximché, Guatemala, S.L. Whittington and R. Tykot 2000), which was funded by the
Foundation for the Advancement of Mesoamerican Studies, Inc., is available on the web. 

 
Patagonia (Argentina and Chile). In southernmost South America, ethnographic observations of historic groups
of hunter-gatherers have revealed several discrete subsistence adaptations, including great dependence on marine
foods for those living on the coasts and inlets of Tierra del Fuego.Growing archaeological evidence, however,
has suggested at least that this historic pattern does not extend into antiquity. Isotopic analysis of bone collagen,
bone apatite, and tooth enamel from a large number of sites in central and southern Patagonia, provided through
collaborations with Julieta Gomez Otero, Juan Belardi, Ricardo Guichon and Luis Borrero, indicates that the
coastal populations had a broad range of diets, in many cases depending more on terrestrial than marine foods.
Some coastal groups clearly exploited high-trophic level marine resources such as sea lions and penguins, while
others had more mixed diets which included fish, mollusks, guanaco, and land birds. The result appears to be
that a continuum of subsistence combinations were employed, rather than the discrete selected diets proposed by
ethnographic observations. See Gomez Otero et al. 2000; Borrero et al. 2001.
 
Coastal Ecuador and Peru . The diets associated with complex societies on the coasts of Ecuador and Peru
have been the subject of great debate because of the conceived importance of agriculture- especially maize - as
the basis for sedentary societies and the emergence of political hierarchies, and the reported presence of maize
pollen and phytoliths in the region by 5000 BC. The precocious Valdivia culture on the coast of Ecuador has
ceramics and ceremonial sites by 3000 BC, while monumental platform structures appear on the coasts of Peru
in the 3rd millennium BC and ceramics shortly afterwards. Analyses of a time series of individuals from coastal
Ecuador (vander Merwe et al. 1993; Tykot & Staller 2002, supplement), however, suggest that maize is not
noticeable in diets until the end of the Valdivia period (phases 7-8, about 2000 BC), long after the appearance of
settled villages with ceramics and ceremonial architecture,and only became a dietary staple in the second half of
the 1st millennium BC. In Peru, also, analyses of remains from the Lurin (Mina Perdida, Cardal, Tablada de
Lurin: Tykot et al. 2006) and Viru (Ericson et al. 1989) valleys suggests that maize was not a dietary staple until
well into the Initial Period. For interpreting coastal diets, where a combination of riverine, marine, terrestrial C3
and C4 resources could have been consumed, it is important to obtain stable isotope data from both human bone
collagen and apatite, as well as the available fauna and flora. 
 
Highland Ecuador and Peru. In highland Ecuador and Peru, collaborations with Richard Burger and J. Stephen
Athens have provided the opportunity to examine the introduction and increasing importance of maize in the
highland Andes. New collagen carbon and nitrogen isotope data from Pacopampa and La Chimba support earlier
results based on collagen carbon from Huaricoto and Chavin de Huantar (Burger and van der Merwe 1990)
which suggest modest importance of maize during the first millennium BC in the highland Andes. The apatite
data, however, suggest that while maize was a similar proportion of the whole diet in both regions during the 1st
millennium BC, it was a greater contributor to dietary protein in highland Ecuador. In later periods maize
became a dietary staple, with differential access (e.g. the consumption of maize beer) according to social status
at sites such as La Florida (Ubelaker et al. 1995). Although there appears to be an increase in d15N values over
time, which could be interpreted as due to greater consumption of animals (higher trophic level than plants) or
the availability of small quantities of fish by trade with people living at lower elevations, this is most likely an
effect of the differences in altitude of the sites in our study. 
New Mexico. Excavations in the La Plata valley by Debra Martin and colleagues uncovered a number of
Anasazi burials (1000-1200 AD), some which received different burial treatments. Stable isotope analysis of
bone collagen and bone apatite samples indicate all were heavily dependent on maize. Individuals in flexed,
twisted, or 'hurled' burial contexts exhibit gender-based diet differentiation. These results are significant for
current discussions of cultural practices and violence/warfare in this region. Two groups of males are suggested
by the isotope data, and at least one female appears to have been a recent immigrant to the La Plata valley.
 
Microsampling of  Teeth. Like hair, teeth are incremental growth structures, which presents both advantages
and limitations for isotopic analysis. In most cases, archaeologists attempt to be minimally destructive in their
analyses, so the removal of a small sample of tooth enamel is required (with the tooth mostly preserved). The
diet revealed by isotopic analysis of such a small sample, however, will be very short term, perhaps as little as a
few weeks. A series of analyses of sequentially formed enamel layers may then be used to examine short term or
seasonal variations in diet, and even to correlate dietary changes with pathologies such as enamel hypoplasia.
Preliminary results from our project in collaboration with Douglas Ubelaker shows potentially large isotopic
differences within a single human tooth (4 per mil). On the other hand, 'routine' studies in which comparisons are
made of  individuals (or of multiple teeth from a single individual) from which a single small enamel sample has
been tested must allow for the possibility of seasonal dietary variation obscuring variability which otherwise
would have been based on eruption age and weaning practices, migration, etc.
 microsampling molar with .75 mm bit

 ten d13C measurements on single incisor 

 
Elemental Analyses . The elemental composition of bones is also a function of diet, but often more difficult to
interpret in archaeological samples (Burton 1996). Elemental ratios of Sr/Ca are a function of trophic level, but
also face difficulties in interpretation (see Burton et al. 1999; Wilson 2000). At USF, elemental analyses are
obtained by Inductively Coupled Plasma (ICP) Spectroscopy, ICP Mass Spectrometry, DC Plasma
Spectroscopy,or Atomic Absorption Spectroscopy. One current project, in collaboration with Francis Thackeray
of the Transvaal Museum, is examining trophic levels in baboons and australopithecines from South Africa.
 Jason Wilson using the

DCP   Johan Schijf & the ICP-MS 

  
 
For more recent bone chemistry studies in Florida, Texas, New York, New Mexico, Mexico, Peru,
Bolivia, Argentina, Chile, Italy, Sardinia,  Malta, Greece, Sweden, China, and elsewhere, go to the full list of my
conference presentations and publications (in PDF) at  http://shell.cas.usf.edu/~rtykot/Pubs.html
  
 

References

Ambrose, S. H. 1990. Preparation and characterization of bone and tooth collagen for isotopic analysis. Journal
of Archaeological Science 17(4):431-451.

Ambrose, S. H. 1991. Effects of diet, climate and physiology on nitrogen isotope abundances in terrestrial
foodwebs. Journal of Archaeological Science 18: 293-317.

Ambrose, S. H. 1993. "Isotopic analysis of paleodiets: methodological and interpretive considerations," in

Investigations of Ancient Human Tissue: Chemical Analysis in Anthropology. Edited by M. K. Sandford, pp.
59-129. Langhorne, PA: Gordon and Breach Science Publishers.

Ambrose, S. H. and L. Norr. 1993. "Experimental evidence for the relationship of the carbon isotope ratios of
whole diet and dietary protein to those of  bone collagen and carbonate," in Prehistoric Human
Bone:Archaeology at the Molecular Level. Edited by J. B. Lambert and G. Grupe, pp. 1-37. New York:
Springer-Verlag.

Burger, R. L. and N. J. van der Merwe. 1990. Maize and the origin of highland Chavín civilization: an isotopic
perspective. American Anthropologist 92: 85-95.

Burton, J.H. 1996. Trace elements in bone as paleodietary indicators.  In M.V. Orna (ed.), Archaeological
Chemistry: Organic, Inorganic, and Biochemical Analysis.  ACS Symposium Series 625: 327-333. Washington:
American Chemical Society.
Burton, J.H., T.D. Price & W.D. Middleton. 1999. Correlation of bone Ba/Ca and Sr/Ca due to biological
purification of calcium. Journal of Archaeological Science 26: 609-616.

Ericson, J. E., M. West, C. H. Sullivan and H.W. Krueger.1989. "The development of maize agriculture in the
Viru valley, Peru," in The Chemistry of Prehistoric Human Bone. Edited by T. D. Price, pp. 68-104. Cambridge
University Press.

Katzenberg, M.A. & R.G. Harrison. 1997. What's in a bone? Recent advances in archaeological bone chemistry.
Journal of Archaeological Research 5(3): 265-293.

Koch, P. L., N. Tuross and M. L. Fogel. 1997. The effects of sample treatment and diagenesis on the isotopic
integrity of carbonate in biogenic hyroxylapatite. Journal of Archaeological Science 24:417- 429.

Kosiba, S., R.H. Tykot, & D. Carlsson. 2007. Stable Isotopes as Indicators of Change in the Food Procurement
and Food Preference of Viking Age and Early Christian Populations on Gotland (Sweden). Journal of
Anthropological Archaeology 26: 394-411.

Larsen, C. S. 1997. Bioarchaeology: Interpreting Behavior from the Human Skeleton. Cambridge: Cambridge
University Press.

O'Connell, T.C. & R.E.M. Hedges. 1999. Investigations into the effect of diet on modern human hair isotopic
values. American Journal of Physical Anthropology 108: 409-425.

Schoeninger, M. J. 1995. Stable isotope studies in human evolution. Evolutionary Anthropology 4(3): 83-98.

Schoeninger, M. J. and DeNiro, M. J. 1983. Stable nitrogen isotope ratios of bone collagen reflect marine and
terrestrial components of prehistoric human diet. Science 220:1381-1383.

Schoeninger, M. J. and DeNiro, M. J. 1984. Nitrogen and carbon isotopic composition of bone collagen from
marine and terrestrial animals. Geochimica et Cosmochimica Acta 48: 625-639.

Schwarcz, H. P. and M. J. Schoeninger. 1991. Stable isotope analysis in human nutritional ecology. Yearbook of
Physical Anthropology 34: 283-321.

Sealy, J., R. Armstrong & C. Schrire. 1995. Beyond lifetime averages: tracing life histories through isotopic
analysis of different calcified tissues from archaeological human skeletons. Antiquity 69: 290-300.

Sponheimer, M & J.A. Lee-Thorp. 1999. Isotopic Evidence for the Diet of an Early Hominid, Australopithecus
africanus. Science 283(5400): 368.

Tieszen, L. L. and T. Fagre. 1993. Effect of diet quality and composition on the isotopic composition of
respiratory CO2, bone collagen, bioapatite, and soft tissues. In Prehistoric Human Bone: Archaeology at the
Molecular Level. Edited by J. B. Lambert and G. Grupe, pp. 121-155. NewYork: Springer-Verlag.

Tykot, R.H. 2004. Stable Isotopes and Diet: You Are What You Eat. In M. Martini, M. Milazzo & M. Piacentini
(eds.), Physics Methods in Archaeometry. Proceedings of the International School of Physics “Enrico Fermi”
Course CLIV, 433-444. Bologna, Italy: Società Italiana di Fisica. 

Tykot, R.H. 2006. Isotope Analyses and the Histories of Maize. In J.E. Staller, R.H. Tykot & B.F. Benz (eds.),
Histories of Maize: Multidisciplinary Approaches to the Prehistory, Linguistics, Biogeography, Domestication,
and Evolution of Maize, 131-142. Academic Press (Elsevier).

Tykot, R.H. & J.E. Staller. 2002. On the earliest introduction of maize agriculture in coastal Ecuador: new data
from La Emerenciana. Current Anthropology 43(4): 666-677. 

Tykot, R. H., N. J. van der Merwe and N. Hammond. 1996. "Stable isotope analysis of bone collagen and apatite
in the reconstruction of human diet: a case study from Cuello, Belize," in Archaeological Chemistry.Organic,
Inorganic and Biochemical Analysis. Edited by M.V. Orna, pp. 355-365. ACS Symposium Series 625,
Washington, DC: American Chemical Society.

Ubelaker, D. H., M. A. Katzenberg and L. G. Doyon.1995. Status and diet in precontact highland Ecuador.
American Journal of Physical Anthropology 97: 403-411.

van der Merwe, N. J. 1992. Light stable isotopes and the reconstruction of prehistoric diets. Proceedings of the
British Academy 77: 247-264.

van der Merwe, N.J. 1989. Natural variation in C13 concentration and its effect on environmental reconstruction
using C12/C13 ratios in animal bones. In T.D. Price (ed.), The Chemistry of Prehistoric Human Bone, 105-125.
Cambridge University Press.

van der Merwe, N. J., J. A. Lee-Thorp and J. S.Raymond. 1993. "Light, stable isotopes and the subsistence base
of Formative cultures at Valdivia, Ecuador," in Prehistoric Human Bone: Archaeology at the Molecular Level.
Edited by J.B. Lambert and G. Grupe, pp. 63-97. Berlin: Springer-Verlag.

Whittington, S.L., R.H. Tykot & D.M. Reed. nd. Diet, demography, and death at Iximché, Guatemala: stable
isotope analyses of  Late Postclassic human skeletal remains from the Kaqchikel Maya capital. Ancient
Mesoamerica. Submitted.

Wilson, J.W. 2000. Strontium-Calcium Ratio Analysis and Dietary Adaptations of Early Hominids and
Hominoids from South Africa. Senior Honors Thesis, Department of Anthropology, University of South Florida.

Wright, L.E. & H.P. Schwarcz. 1998. Stable carbon and oxygen isotopes in human tooth enamel: identifying
breastfeeding and weaning in prehistory. American Journal of Physical Anthropology 106: 1-18. 
 

Archaeological Science Laboratory Staff and Students

Robert H. Tykot, Director 

Lori Adornato, Research Assistant (1999-2000)  
Marie Archaembeault, Graduate Research Assistant (2001-2004) . MA Thesis: Sources of Marble Used for
Sculptures and Mosaics at Antioch (2004)
Jonathan Auth, Senior Honors Student (2004-2005).  Senior Honors Thesis: Migration and Dietary
Change in South America: Stable Isotope Analysis of 33 Locations in Central Chile (2005)
Shannon Baker, Undergraduate Research Assistant (spring 2002)  
Malinda Berman, University Graduate Fellow (1999-2000)  
John Bernal, Undergraduate Research Assistant (2000-2002). Senior Honors Thesis: A Macroscopic Dental
Analysis of Skeletal Remains from the Main Burial Mound  
   at the Crystal River (Citrus County, Florida) Archaeological Site  
Lisa Beyer, Graduate Research Assistant (2000-2008) 
Julie Bliss, Research Assistant (2000-2001) 
Robert D. Bowers, Senior Honors Student and Graduate Research Assistasnt (2005-). Senior Honors
Thesis: Phytolith Analysis of Pottery from Liangchengzhen, Shandong, China (2006)
William Boynton, Senior Honors Student (2005-2006). Senior Honors Thesis: Practical Evaluation of
the Manufacturing Processes Involved in Blade Technology (2006)
Catherine Carvin, Graduate Research Assistant (fall 1996)
Kara Casto, Graduate Assistant (2008-)
Leya Collins, Senior Honors Student (2006-2007). 
Travis Doering, Graduate Research Assistant (2002-2007). PhD Dissertation: An Unexplored Realm in
the Heartland of the Southern Gulf Olmec: Investigations at El Marquesillo, Veracruz, Mexico (2007)
Jennifer Dukes, Research Assistant (1999-2001) 
Brian Eels, Research Assistant (2000-2001)  
Ted Fassler, Graduate Research Assistant (fall 1998) 
Kyle Freund, Graduate Research Assistant (2008-)
Kristine Gary, Research Assistant (1998) 
Teryn Gilbertson, PhD program (2003-)
Rebecca Glazer, Research Assistant (summer 2000)  
Scott Grammer, Graduate Research Assistant (1997-98). MA Thesis: Isotopic Evidence for Reconstructing
Prehistoric Subsistence Patterns 
    in the Central Coast of Northern Patagonia, Argentina (2005)
Jason Hicks, Research Assistant and Honors Student (1996-98)  
Peter Karellas, Senior Honors Student (2005-2006)
Jennifer Kelly, Honors and Graduate Student, Intern (1999-2005). Senior Honors Thesis: Stable Isotope Analysis
and Prehistoric Human Diet in Southern Patagonia (2000); 
    MA Thesis:  Stable Isotope Evidence for Maize Consumption and Other Dietary Practices at Bayshore Homes
(8Pi41) and Other Prehistoric Sites in Peninsular Florida (2004)  
Steven Kosiba, Research Assistant (2000-2001) 
Kathryn Lafrenz, University Graduate Fellow and Graduate Research Assistant (2001-2004) . MA Thesis:
Tracing the Source of the Elephant and Hippopotamus Ivory from the 14th Century B.C. Uluburun
Shipwreck: 
    The Archaeological, Historical, and Isotopic Evidence (2004)
Luca Lai, Graduate Research Assistant (2002-2008). Dissertation:  The Interplay of Economic, Climatic and
Cultural Change Investigated through Isotopic Analyses of Bone Tissue: the Case of Sardinia 4000-1900 BC
(2008)
Rheta Lanehart, PhD program (2003-)
Eugenia Brown Mansell, Graduate Research Assistant, Intern (1996, 1999-2004). 
Scott McCready, Research Assistant (2000-2001) 
Anne Metroka, Senior Honors Student (2006-2007). Senior Honors Thesis: Early Historic Diets in Latin
America
Melissa Miller, Honors Student (2001-2002). Senior Honors Thesis:  A Quantitative and Qualitative Analysis
Aimed Toward Discovering the Sex and Population Affinity of Five Unknown Individuals
Stacey Nott Hopper, Graduate Research Assistant (spring 1997)  
Michael Pateman, Graduate Research Assistant (2004-2006)
Shannon Peck Janssen, Graduate Research Assistant (2002-2006). MA Thesis: Animal Husbandry in
the Early Bronze Age Levant (2006)
Frederick Pirone, PhD program (2005-)
Theresa Pritchard, Undergraduate Research Assistant (spring 2002)  
Stephanie Roberts, Research Assistant (2000-2001) 
Teddi Setzer, Graduate Research Assistant (2000-2009) . MA thesis: Use-Wear Experiments with Sardinian
Obsidian: Determining its Function in the Neolithic (2004); PhD dissertation:  Malaria and the Bronze
Age in Sardinia (Italy) 
Alex Seufert, Senior Honors Student (2007-2008). Senior Honors Thesis: 
Nicole Shelnut, Graduate Research Assistant (2003-2007) . MA thesis: Prehistoric Subsistence
Patterns in Western Argentina (2006)
Charles Summers, Research Assistant (spring 2001) 
Dolores Tedesco, Research Assistant (1997-1999) 
Lisa Tucker, Graduate Research Assistant (spring 1999)  
Barbara Vargo, Graduate Research Assistant and Intern (1999-2000). MA Thesis: The Characterization of
Obsidian from Pantelleria (Italy):  
   The Archaeological Significance of Multiple Island Sources.  
Gail Walton, University Honors Student (2000-2001). 
Robert Whalen, Undergraduate Research Assistant (2000-2002).  
Michelle Whitaker, Honors Student (1999-2000). Senior Honors Thesis: Anasazi Subsistence in La Plata, New
Mexico: Carbon, Corn and Culture. 
Jason Wilson, Research Assistant (1996-2000). Senior Honors Thesis: Strontium-Calcium Ratio Analysis and
Dietary Adaptations 
   of Early Hominids and Hominoids from South Africa.  
Justin Winkler, Graduate Research Assistant, Intern (2000-2004). MA Thesis: Technology, Provenance, and
Economy of Etruscan Pottery from Poggio Colla, Italy: An  
   Archaeometric Investigation. 
 

Strontium in bones
ASPARN, AUSTRIA

Researchers use stronium in bones to distinguish between population groups

Strontium (Sr) is a chemical element

that builds up in the bones and tooth
enamel as food is digested. Through
its precise determination, researchers
are now able to distinguish between
friend and foe on ancient battlefields.

This is possible because in nature, Sr

occurs in a variety of forms (isotopes)
that differ according to the number of
particles in the atomic nucleus. The
87Sr und 86Sr isotopes are the ones
that are important for the classification
of bone material, and the ratio
between the two varies and changes
over time in accordance with
geological conditions. As the strontium in the bones is constantly exchanged with the Sr ingested with food, the
isotope ratio in bone material reflects that in the surrounding area. Because of this the Sr isotope ratio provides
clues about a person's origins.

However measuring the ratio accurately is a challenging task. A team under Professor Thomas Prohaska of the
Institute of Chemistry at the University of Natural Resources and Applied Life Sciences in Vienna has now
succeeded in achieving hitherto unknown levels of accuracy with this form of analysis. The interdisciplinary
project, supported by the Austrian Science Fund (FWF), has opened the way for fascinating reconstructions of
prehistoric events.

The scientific usefulness of the method was demonstrated at an excavation in cooperation with a team under
Prof. Maria Teschler-Nicola of the Vienna Museum of Natural History. Over 100 human skeletons, dating to 7,000
years ago, had been found very close together at the site of a Neolithic village in Asparn, Lower Austria.

However, to work out whether fighting had occurred at the village, the archaeologists could not use traditional
methods, which depend on comparisons of morphological characteristics, and require that the skeletons be in an
excellent state of preservation. It was only when the results of anthropological research were complemented by
determination of the Sr isotope ratio that it was possible to unambiguously identify a group of skeletons as those
of villagers who had been defeated by intruders.

The applications of this technique are myriad. Another FWF funded project aims to work out the migratory
behaviour of individuals by comparing the Sr Isotope ratio in the bones to that in the molars (a technique used to
identify the origin of Oetzi the Iceman, as reported in CWA 2). For, in contrast to the bones, Sr is no longer
exchanged in tooth enamel after the age of four, allowing researchers to tell where the person lived as a child.
Different isotope ratios in the bones and teeth are therefore proof of migration after the age of four.

This research project is an impressive example of the ways in which analytical chemistry, archaeology and
anthropology can join forces to produce new insights.