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AMS Carbon Dating of Bone Samples

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• Bones are one of the most common materials sent to labs for radiocarbon dating.
• Bone samples have a time-width.
• Contaminants that affect the true age of a bone sample can be natural or artificial.
• Bones are 30% organic; this portion is often extracted for radiocarbon dating.
• There are several steps done during the physical and chemical pretreatment of bones prior to radiocarbon dating.

Bones are one of the most common materials sent to accelerator mass spectrometry (AMS) labs for radiocarbon dating. This is because bones of animals or humans are often subjects of archaeological studies.

A lot about the prehistoric era has been learned due to archaeological studies and radiocarbon dating of bones. More in-depth information about old civilizations is also available due to radiocarbon dating results on bones.

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Beta Analytic provides AMS dating of cremated and non-cremated bones, antlers, and teeth.

Sample size required: 2-10 grams of non-cremated bones and antlers, 1-2 teeth, and 4-40 grams of cremated bones.

For teeth, preferred samples are incisors, canines, and molars with all four of the roots still attached.

For bones, good cortical bone is best from the larger bones of the body (femur, tibia, upper arm bone, jaw, skull plate and sometimes the ribs). Spongy bones like ball and sockets, vertebra and the like do not tend to preserve well in harsh conditions and may not yield sufficient collagen for AMS dating.

For bird bones, please consult the lab for sufficient sample size. By necessity, bird bones are not very dense. As such, the amount of collagen per unit gram in a bird bone is much less than a mammal bone might be. Sample size of 2-4 grams may not be enough for AMS dating after pretreatment.

When submitting antlers, please consult the lab whether it's best to submit antler chips, shavings, or powder.

Bones that have been submerged in water or wet sediments usually have very little collagen remaining despite their appearance. Water is very effective in leaching the collagen proteins out of the bone, leaving only bone carbonate. Thus, it's possible that the bones have very limited collagen. Please consult the lab before sending these bone samples.

Recommended container: Ziplock Bags

Important Note on Pretreatment - It is important to understand the pretreatments which are going to be applied to samples since they directly affect the final result. You are welcome to contact us to discuss the pretreatment or request that we contact you after the pretreatment (and prior to dating) to discuss.

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AMS Dating of Different Types of Bones

Non-burnt Bones - A bone that has not been burned is pretreated by extraction of the collagen proteins. This is the most reliable material that can be dated for non-cremated bones.

Preservation and quality of the preserved collagen is very important. This can be assessed during pretreatment. If collagen quality is good, pretreatment continues (acid/alkali/acid washes). If collagen quality is poor, the lab consults with the client for cancellation of the analysis (partial charges are incurred).

After the initial evaluation of the collagen, it is dried for Carbon-13/Carbon-12 ratio analysis. If the result of this analysis is reasonable, the lab proceeds with AMS dating. If the 13C/12C analytical result is poor, AMS dating can be canceled with only partial charges incurred.

Burnt bones - Radiocarbon dating of burnt bones can be somewhat problematic depending on how they are burned and to what degree. Burnt bones fall into 3 categories:

1) Cremated bones - These are bones that have been heated in excess of 600C for sufficient time to burn away all of the collagen, fats, and proteins. These bones are typically white in color; and if broken into two, they are completely white on the inside as well.

When bones are heated to above 600C, the osteocalcin (apatite) in the bone is converted to structural carbonate. This is the bone carbonate that is dated. The structural carbonate is very resistant to change and not easily contaminated once cremation has occurred, therefore it has been shown to be a good substance for reliable AMS dating.

In the absence of any charred collagen, a method is available for dating the carbonate fraction in cremated bones. The method was published and accepted in 2000 at the 17th International Radiocarbon conference. Studies indicate good agreement between bone carbonate in highly heated bones with associated charcoal. This method should only be attempted in the absence of collagen or charred collagen.

Recent studies have also shown that carbonate yields from separate sections of bone may be indicative of incomplete cremation. To test this, two portions of the bone are tested for carbonate yield. If they are similar, the lab proceeds with AMS dating. If the carbonate yields are dissimilar, the client can cancel the analysis (incurring only partial charges) or continue with AMS dating of both portions to test for similar age (incurring the cost of two analyses).

2) Bones that have not been heated to greater than 600C for sufficient time show black, blue or gray patches on the exterior surfaces or in the interior, indicating that all the fats and proteins have not been burned away and thus the osteocalcin has not been completely converted to structural carbonate. These bones are typically not datable.

3) Charred Bones - These are bones that are completely charred inside and out and look like a chunk of charcoal. These are usually charred in very low oxygen, less than 600C environments over a long period of time. The osteocalcin has been burned away leaving only the charred fats and proteins (collagen) behind. These types of burned bone can usually be dated but the pretreatments may be limited to acid leaches to remove carbonates. Many times they are too fragile to allow for alkali extractions to remove humic acids that may be present in abundance in the area of collection.

Bones that have been heated in low temperatures present special considerations. Whether or not a charred bone will yield a radiocarbon date depends on the degree of charring. Bones with charred protein can be very good samples for AMS dating. In this case, the carbon is resilient to decay and can be fully pretreated in the laboratory. If the protein is partially charred, it is probably damaged and highly susceptible to decay. It usually cannot be fully pretreated (or identified as protein) in the laboratory.

Generally, if the bone is bleached white throughout, charred collagen is not available. If the bone is black or blue, there is some chance it can be dated using a charred collagen remnant. The only way to know is to do some pretreatment.

If you are unsure of which category your bone samples belong, please send them to our radiocarbon dating lab. We will examine them and advise if they are datable and by what technique.

Time-width of Bone Samples

The time-width of any given sample reflects the total growth of the original organism and the span of time that organism interacted with the biosphere. For most organisms that have bones, the time of their death is contemporaneous with their cessation of exchange with the biosphere. Hence, these organisms’ radiocarbon age at death is zero.

Radiocarbon dating results on bones need not be subjected to an age offset but bone samples have time-width. Literature suggests that a bone does not cease to assimilate carbon from the biosphere until death; there is a turnover time of about 30 years for human bone and a shorter period for animal bone.

Time-width data is necessary because they affect calibration of radiocarbon results and, consequently, the way radiocarbon age is converted to calendar years.

Bone Sample Contamination

Any carbon-containing material that may affect the carbon 14 content of bones is considered a contaminant. Considering that bones are often found surrounded by different kinds of organic matter, bones are arguably one of the most highly contaminated samples submitted to AMS labs for radiocarbon dating.

The common contaminants are humic and fulvic acids, which are organic acids present in soil that are produced by the microbial degradation of plant or animal tissues. According to literature, other organic compounds that can contaminate bone samples are polyphenols, polysaccharides, lignins, and degraded collagen. Depending on the location of the excavation, bones can also be contaminated by limestone. These contaminants are considered natural because they came in contact with the bones due to natural occurrences.

Artificial contaminants, on the other hand, are those that were introduced by man during the collection, conservation, or packaging of the bone samples. When bones are applied with animal glue during labeling, a contaminant has already been introduced to the sample. This is because animal glue is chemically identical to the bone sample. AMS lab results with this sample will be inaccurate.

Other potential contaminants that can be introduced to bone samples after excavation include biocides, polyvinyl acetate and polyethylene glycol (conservation chemicals), cigarette ash, and labels or wrappers that are made of paper.

Effect of Contaminants on AMS Carbon Dating Results

The effect of contamination on bone samples that were subjected to AMS dating is dependent on these factors: type of contaminant, degree of contamination, and the relative age of the bones and the contaminant.

If limestone has not been removed prior to AMS carbon dating, the radiocarbon age will be much older than the sample’s true age. Limestone is of geological origin and will therefore be much older than any archaeological samples.

The presence of humic and fulvic acids during AMS radiocarbon dating will lead to inaccurate results as well. Depending on the age of the organism that produced the organic acids, the AMS lab’s result might reflect a radiocarbon age younger or older than the bone sample’s true age.

Bones can also be exposed to modern sources of carbon due to plant rootlet intrusions. Modern sources of carbon can make the AMS carbon dating result of a bone younger than its true age.

In general, infinite-age contaminants add considerable number of years to the true age of a bone sample, making it older than it is. Modern carbon, on the other hand, makes the bone sample significantly younger than its true age.

To prevent these inaccuracies, AMS labs perform pretreatment on all bone samples before subjecting them to AMS radiocarbon dating.

Components of a Bone

A bone is 30% organic and 70% inorganic. The organic portion is protein; the inorganic portion is the mineral hydroxyapatite, which is a combination of calcium phosphate, calcium carbonate, calcium fluoride, calcium hydroxide, and citrate. The protein, which is mostly collagen, provides strength and flexibility to the bone whereas the hydroxyapatite gives the bone its rigidity and solid structure.

In theory, both organic and inorganic components can be dated. However, the open lattice structure of the hydroxyapatite makes it highly contaminated with carbonates from ground water. Removal of carbonate contaminants through dilute acid washing is also not applicable because hydroxyapatite is acid soluble.

AMS labs use the protein component of bone samples in radiocarbon dating because it is relatively acid insoluble and, therefore, can be easily isolated from the hydroxyapatite component and other carbonates.

In cases when the protein portion of the bone sample is not well preserved and have already degraded due to warm conditions and fungal or bacterial attack, AMS labs carbon date individual amino acids to check if several of them give the same radiocarbon age. This process is doable in AMS labs because only small samples are required. However, this process is costly and time consuming. Radiocarbon dating individual amino acids is not recommended by AMS labs unless necessary as in the case of old bone samples where the presence of even small levels of contaminants produce a large error.

Physical Pretreatment of Bones in AMS Labs

Physical pretreatment refers to processes done on the bone samples for carbon dating without using chemicals. Examples of physical pretreatment done on bones in AMS labs are removal of plant rootlets and reduction of sample size by crushing.

AMS lab personnel visually examine bone sample submissions for obvious contaminants.

Rootlets are removed using a pair of tweezers or forceps. A surgical scalpel or a dental grill is used to scrape off contaminated exterior layers of bone samples.

AMS labs would also check the bone’s hardness. Softness indicates the potential absence of collagen, which is needed for AMS carbon 14 dating.

After initial removal of visible contaminants, AMS lab personnel crush bone samples in a mortar and pestle. Size reduction is done to increase the surface area of the sample during succeeding pretreatment methods.

Chemical Pretreatment of Bones in AMS Labs

Different AMS labs may have slight variations in the procedure of chemical pretreatment, but they often use the same chemicals in treating bone samples.

The crushed bone sample is washed with dilute, cold hydrochloric acid (HCl) repeatedly until hydroxyapatite is eliminated and the collagen is isolated. Rootlets, if present, are further removed from the collagen.

To ensure the complete removal of organic acids, collagen is washed with an alkali solution, usually sodium hydroxide (NaOH). AMS labs, however, skip alkali washing when the collagen sample is not well preserved and the washing may remove the remaining organic materials that can still be dated.