Bone Diagenesis and Taphonomic History of the Paso Otero 1 Bone Bed, Pampas of Argentina

Samples of guanaco bone from an archaeological site in the Pampas of Argentina have been analysed to understand the diagenetic profile of the bone assemblages that characterized the taphonomic history of the site. Two archaeological occupations of Paso Otero 1 were investigated, encompassing similar landscape settings, climates, and depositional environments. The time span is a c. 2000 year period from c. 4800 to 2800 years . A total of 30 bone samples taken from both occupations were used to provide a preliminary characterization of the diagenetic pathways at the site. The parameters investigated provide a comprehensive account of how both mineral (hydroxyapatite) and bone protein (collagen) have been altered. In order to compare the two bone assemblages in terms of their diagenetic parameters, multivariate analyses were conducted. Results indicate two different diagenetic profiles in the site, % N being one of the variables that accounts for most of the variation in Paso Otero 1. The diagenetic analyses indicate that protein is less preserved in the bone assemblage from the middle stable landscape. Alternative interpretations of the diagenetic profiles are discussed in light of the taphonomic history of the site, and palaeoenvironmental information of the region. One hypothesis stresses the importance of the role of climate in defining the different diagenetic pathways, and the other the continued action of the combined diagenetic factors along time as the main explanation for the variability in the state of preservation of the bones in Paso Otero 1.     

Diagenesis and survival of osteocalcin in archaeological bone

It has been demonstrated that the protein osteocalcin can survive in bone in the archaeological record, and postulated that it has the potential to survive over geological time periods. The precise mechanism for this longevity of survival is not yet fully understood, and has not been extensively studied in comparison to other diagenetic aspects of archaeological bone. We report a comparison between osteocalcin survival and the state of preservation of more than 60 bones from 14 archaeological sites. The amount of osteocalcin, assayed immunologically, was compared with diagenetic parameters that measure: the amount of ‘collagen’ in the bone, the mineral changes, the porosity, and the histological preservation of the material. The findings indicate that microbial taphonomy and mineral alteration of bone have a profoundly damaging effect on the preservation of osteocalcin.     

The taphonomy of cooked bone: characterizing boiling and its physico–chemical effects

Cooking is perhaps the most common pre–burial taphonomic transformation that occurs to bone, yet it is still one of the least understood. Little progress has been made in determining a method of identifying cooked bone in the archaeological record, despite its import for various branches of archaeology. This paper attempts to describe boiling in terms of its physico–chemical effects on bone, and uses a suite of diagenetic indicators to do this. It is shown that cooking for brief periods of time has little distinguishable effect on bone in the short term, but that increased boiling times can mirror diagenetic effects observed in archaeological bone. The relationship between the loss of collagen and alterations to the bone mineral is explored through heating experiments, and the results compared with archaeological data.The possibility of boiling being used as an analogue for bone diagenesis in future studies is raised, and the key relationship between protein and mineral is once again highlighted as vital to our understanding of bone diagenesis.     

Pick the Right Pocket. Sub‐sampling of Bone Sections to Investigate Diagenesis and DNA Preservation

Many archaeological bones display a heterogeneous degradation pattern. Highly degraded bones could contain pockets of well‐preserved bone, harbouring good quality DNA. This dichotomy may explain why the relationships between global bone preservation parameters such as histological integrity, bone mineral crystallinity or collagen yield, and bulk DNA preservation/amplification success rate have been found to be at best, weak to moderate. In this pilot study, we explore whether or not a more localised approach will highlight a stronger relationship between diagenetic parameters and DNA preservation. This study includes a detailed histological characterisation of bone diagenesis in sub‐areas of three bone samples. Regions of the same bone, which displayed differential degrees of preservation or type of diagenesis were sampled for further analysis and both genetic (small scale Illumina MiSeq sequencing) and chemical (Fourier‐transform infrared spectrometric analysis) analyses were performed. The aim was to investigate how bone diagenetic processes relate to DNA preservation at a higher resolution than in previous studies. This is key in order to improve DNA analytical success rates. The expected relationship between bone and DNA preservation (retrieved endogenous DNA) was observed and the results corroborate previous work that DNA preservation is linked to the integrity of bone collagen and mineral. The results further suggest that non‐biological diagenetic alterations such as etching and the presence of mineral infiltrations and inclusions have a negative effect on DNA preservation/extraction. Copyright © 2016 John Wiley & Sons, Ltd.     

Patterns of Diagenesis in Bone II: Effects of Acetic Acid Treatment and the Removal of Diagenetic CO3

Archaeological bones of varying preservation have been treated with 0·1 M acetic acid in order to investigate the effect on structural and chemical alterations caused by diagenesis. Acetic acid is commonly used as a “cleaning agent” for removing diagenetic carbonate from bone and enamel, in an attempt to recover original, biogenic signals for use in dietary and¹⁴C dating studies.Diagenetic parameters were measured before and after treatment on a range of archaeological bones with good and bad preservation. Histological preservation defined the behaviour of the correlating parameters, where correlation coefficients between carbonate content and crystallinity, microporosity and macroporosity increased significantly after treatment. For histologically well preserved material, acetic acid is effective at returning carbonate content to around that of modern bone. Where bone is extensively damaged by micro-organisms, “loose” diagenetic material can be removed, but a fraction largely composed of hypermineralized bioapatite remains, which, we believe, cannot be reliably used to obtain accurate biological signals.     

Uranium and Vanadium Concentrations as a Trace Element Method for Identifying Diagenetically Altered Bone in the Inorganic Phase

Archaeologists have generally avoided analyzing inorganic isotopes in bone because of its high porosity, large crystalline lattice spacing, and small crystallite size, making it particularly susceptible to diagenetic alteration. Because the inorganic isotopes are left unstudied, we lose a significant portion of information pertaining to an individual’s life history, such as migration, health, and ranging behavior. Tooth enamel, which does not have the same susceptibility to diagenesis as bone, can be used to extract this information but this means that taxa lacking teeth, such as birds, some species of fish, and some reptiles, are excluded. Here, we present a method that can be used to identify diagenetic alternation in bone. This is done by focusing on abnormal concentrations of vanadium and uranium. Neither element is readily bio-precipitated into hydroxyapatite due to ionic radius, vibrational frequency, atomic mass, and ionic charge. This makes them an ideal marker for diagenetically altered bone. Vanadium occurs in very low concentrations in modern bone, while archaeological bone shows clear evidence of normal, non-diagenetically altered values alongside high concentrations of vanadium in diagenetically altered bone. Uranium also is a measure of diagenetic alteration, as modern bone has concentrations below detectable limits (0.017 ppb), while some archaeological bone contains uranium above detectable limits. The biogeochemistry of these elements in soil and bone are discussed with implications for enamel studies.     

Bone diagenesis in the European Holocene II: taphonomic and environmental considerations

We have applied cluster analysis to mercury intrusion porosimetry data from 219 archaeological bones (121 human and 98 animal) and soil chemistry data from 219 accompanying soil samples (1 per bone sample), to investigate the influence of soil chemistry on bone preservation. The samples chosen for the study were obtained from sites ranging in time from the pre-modern to the Mesolithic and were representative of burial environments across Europe (from the Baltic to the Mediterranean). These results represent the single largest database for archaeological bone preservation in the European Holocene to date and demonstrate the potential for large-scale diagenetic studies to help develop long term preservation strategies for our European heritage. Despite the variety of sites and environments, bones could be categorised into only four main diagenetic types. Furthermore, soil chemistry appears to significantly affect only one type of preservation, the pathway characterised by loss of mineral. In neutral to basic soils, taphonomy and in particular the differences between the treatment of human and animal remains, becomes the dominating factor in determining preservation. Using these results, strategies for heritage management of archaeological sites can be suggested; grouping sites into those requiring immediate excavation and those where in situ preservation is viable.     

Restrictions on fluorine depth profiling for exposure age dating in archaeological bones

Archaeological fragments of bone and teeth that are exposed to a humid environment take up fluorine from the surrounding soil. The fluorine ion replaces the hydroxyl group in the mineral phase of the bone, forming chemically more stable fluorapatite. In cortical parts of the long bone diaphysis a fluorine concentration profile can be observed, which decreases from the outer surface and the marrow cavity towards the inner parts of the bone matrix. Geological time spans are needed for this process to reach equilibrium and for the distribution to become uniform. As the shape of the profile, which can be described by a diffusion model, contains information on the exposure duration of the fossil object, several attempts to use fluorine profiling as a dating method have been undertaken. The distribution of fluorine in an archaeological sample however is strongly influenced by environmentally induced processes of bone diagenesis, i.e. alteration in the structure and composition of the mineral phase and degradation of organic components that may make the time information indistinct. The primary chemical composition of bones can thus be obscured by diagenesis within tens, hundreds or thousands of years. This depends more on the diagenetic environment than on the geological age. To observe the impact of environmental influence on the profile shape, samples from several burial sites featuring various soil conditions have been analyzed for their fluorine distribution and preservation state. This paper provides an overview on the restrictions that have to be considered when attempts are undertaken to relate a fluorine diffusion pattern to the archaeological age of a bone specimen.     

Quantifying histological changes in archaeological bones using BSE–SEM image analysis

Recent years have seen rapid developments in the understanding of diagenetic changes to archaeological bones. In particular, the degradation or preservation of proteins and other biomolecules has been explored using an increasingly sophisticated battery of analytical techniques. Problems remain, however, in correlating these parameters with physical changes to bone that may be observed microscopically. This is due, in part, to the problems in reproducibly quantifying histological changes to archaeological bone. This paper introduces a novel method for the accurate quantification of these changes employing image analysis of SEM images. Self–consistency of results was tested using measurements of total calcium content at different magnifications. The term ‘bioerosion index’ is suggested for the measured parameter.     

Pre-screening techniques for identification of samples suitable for radiocarbon dating of poorly preserved bones

Under certain environmental conditions, post-depositional diagenetic loss of bone collagen can severely reduce the number of bones from a particular archaeological site that are suitable for stable isotopic analysis or radiocarbon dating. This study examined nearly 300 bones from 12 archaeological sites across southern England known to yield poor or variable preservation to try to identify one, or more, pre-screening technique(s) that would indicate suitable collagen preservation for radiocarbon dating. The most reliable method was shown to be the percent nitrogen (%N) of whole bone powder, which has an 84% chance of successfully predicting whether or not a bone will yield sufficient (i.e. >1% weight) collagen for dating.     

Bone preservation in human remains from the Terme del Sarno at Pompeii using light microscopy and scanning electron microscopy

Archaeological bone can show marked and complex alterations depending on the environment in which it was buried. In this study, the state of preservation of 27 femurs recovered from the archaeological site of Pompeii was evaluated by light microscopy and scanning electron microscopy. Most of the bone samples, prepared by the grinding method, showed good histological preservation, although they were characterized by microfissures (microcracking). Nine bone samples showed different states of histological preservation, including worst preservation (two femurs), due to diagenetic processes. Cryostat bone sections stained with thionin or 4′,6′-diamidino-phenylindole (DAPI) revealed the persistence of DNA within some osteocyte lacunae. Scanning electron microscopy analysis showed that ultrastructural characters, such as lamellae and collagen fibres, are recognizable only in unaltered bone. Our results reveal that most Pompeian samples are well preserved since they have a bone microstructure virtually indistinguishable from that of fresh bone. In methodological terms, although each of the various morphological methods used contributes information, histological and histochemical analyses are the most informative for studying the preservation state of bone and allow for rapid essential screening of archaeological bone.     

Bone diagenesis: an overview of processes

This overview is a summary of the state of understanding of processes and states in bone diagenesis, as seen from a chemical perspective. It deals with the significance and usefulness of the measurements of ‘diagenetic parameters’—that is, of measures of diagenetic alteration—and of the theories of physico–chemical processes which are considered to underlie the measured changes. In many ways these two aspects are seen to come together quite well, and some progress has been made in relating different burial environments to the observations of alteration. Such a framework also allows us to ask more penetrating questions, such as how characteristic differences in diagenetic alteration might arise, and how the pre–burial environment might influence the eventual course of diagenesis.     

Polarizing Microscopy Identification of Chemical Diagenesis in Archaeological Cementum

Cementum increment analysis can potentially retrieve relatively complete, high-precision seasonality and mortality profiles from archaeological mammalian tooth assemblages. However, cementum exhibits many similarities to bone in composition, histology, ultrastructure, and even microstructure. Consequently, the mineralized dental tissue may be prone to the same processes of post-depositional chemical alteration that affect bone. This article reviews the issues surrounding chemical diagenesis in cementum and presents a new application to archaeological ungulate cementum of a polarizing microscopy technique that has previously been utilized to identify the effects of chemical diagenesis in archaeological bone and human cementum (Geusa et al., 1999; Gilbert, 1989; Watson, 1975; Zeder, 1978). It is argued that the post mortem leaching of collagen and the diagenetic growth of apatite crystallites can develop into banded features that mimic seasonal cementum increments. This pattern of diagenesis is demonstrated in multiple locations on a macroscopically well-preserved Upper Pleistocene-age archaeological tooth, which was previously interpreted by the author to exhibit biogenic seasonal cementum increments (Stutz, 1993; Stutz et al., 1995). It is suggested that if researchers seek to retrieve seasonality and mortality data from archaeological cementum, the microscopy analysis protocol must include screening of a pilot sample for the frequency and extent of chemical diagenesis.     

Characterisation of microbial attack on archaeological bone

As part of an EU funded project to investigate the factors influencing bone preservation in the archaeological record, more than 250 bones from 41 archaeological sites in five countries spanning four climatic regions were studied for diagenetic alteration. Sites were selected to cover a range of environmental conditions and archaeological contexts. Microscopic and physical (mercury intrusion porosimetry) analyses of these bones revealed that the majority (68%) had suffered microbial attack. Furthermore, significant differences were found between animal and human bone in both the state of preservation and the type of microbial attack present. These differences in preservation might result from differences in early taphonomy of the bones.     

Cross-Cultural Approaches in Archaeology: Comparative Ethnology, Comparative Archaeology, and Archaeoethnology

Cross-cultural approaches have been used widely in archaeological research. Comparative ethnology has provided a number of archaeological indicators of behavior, but large segments of the archaeological record have not yet been subjected to extensive comparative analysis. Comparative archaeology has aided in exploring variation among societal types (such as chiefdoms) and categories within the archaeological record (such as settlements). Diachronic comparisons have been used frequently by archaeologists, but these have often been based on unique samples and only rarely have employed statistics to aid in the discovery or testing of hypotheses. Archaeoethnology, comparative analyses of archaeological cases employing valid samples and statistical evaluation of theories and hypotheses, is introduced.     

An assessment of the value of bone density measurements to archaeoichthyological studies

This paper reviews the meaning of the term ‘density’ and the problems associated with the methods of density determination for animal bones in archaeology. It has often been assumed that density is the intrinsic property of most influence in controlling the rate of a bone's decay. Values for whole bone density have been published only for large mammal bone, however. Fish bone appears to be particularly vulnerable to decay, and usually a restricted range of skeletal elements are recovered from archaeological sites. The object of this study was to examine the relationship between fish bone density and the ability of the bone to survive on occupation sites and in archaeological deposits. A set of ‘density’ measurements was established for the bones of cod (Gadus morhua). The usefulness of these measurements as a predictive tool in archaeoichthyological studies is assessed. It was found that ‘density’ as measured did not explain adequately the relative survival of skeletal elements after mechanical abrasion and weathering, or within archaeological deposits.     

Infrared and Isotopic Evidence for Diagenesis of Bone Apatite at Dos Pilas, Guatemala: Palaeodietary Implications

The use of stable carbon isotopic analysis of bone apatite to reconstruct prehistoric diets is hindered by the possibility of diagenetic alteration of carbonate during burial. We examine apatite preservation in Classic Period Maya skeletal remains from Dos Pilas, Guatemala, using Fourier transform infrared spectroscopy (FTIR). We use weight % CO₂evolved from apatite, FTIR carbonate/phosphate absorbance ratios, phosphate peak splitting crystallinity indices, fluoride peaks, and stable oxygen isotopic ratios to identify diagenetic change in apatite chemistry. Isotopically light carbon taken up from burial soil is adequately removed from most Dos Pilas bone by treatment with dilute acetic acid, but more severe alteration cannot be reversed by standard preparation methods. Infrared criteria identify recrystallized apatite in a subset of Dos Pilas burials, that is accompanied by isotopic exchange, and that no longer preserves biogenic δ¹³C. These results illustrate that comparatively recent bone may be diagenetically altered and demonstrate a need for systematic evaluation of mineral integrity in all archaeological bone prior to interpreting paleodiets with apatite δ¹³C.     

Calcium–iron–phosphate features in archaeological sediments: characterization through microfocus synchrotron X-ray scattering analyses

The occurrence of amorphous calcium (Ca)–iron (Fe)–phosphate infilling features in thin-section samples from archaeological stratigraphies is increasingly being reported and used in the cultural interpretations of sites. In some contexts, these materials are the product of dissolution and recrystallization of bone material within pores of the soil or sediment matrix. This study uses transmitted microfocus X-ray scattering to characterize and measure features of known cod fish bone (Gadus morhua) materials, and compare them to archaeological samples of amorphous Ca-Fe-phosphate infilling material found in thin section from early fishing community sites. The analyses characterize the structure of these features for the first time, and allow discussion of the diagenetic processes that lead to their formation.     

Re-examining the chemical evaluation of diagenesis in human bone apatite

Isotopic analysis of human bone is becoming an increasingly important tool for the archaeologist in divining past life-ways. The isotopic ratios within bone are often assumed to be preserved as in life, but diagenetic change can alter the ratios, invalidating the results of isotopic analysis. Diagenesis can be evaluated in a number of ways, but most often spectroscopic techniques are utilised as the most efficient and easiest to understand methods for the archaeologist. Many isotopic studies do not report the possibility of diagenetic change, and if it is reported it has often been quantified using a single method of chemical analysis, FTIR spectroscopy. This study set out to test the value of FTIR analysis using human remains from the prehistoric site of Ban Non Wat, Northeast Thailand, and to compare the results with the non-destructive technique of FT-Raman spectroscopy. The study shows that FTIR spectroscopic analysis gives far less detail on the condition of bone than Raman spectroscopy, which does not merely indicate recrystallisation has occurred, but also shows clearly whether or not collagen is present, allows identification of ionic substitions which have occurred and identification of secondary minerals which have formed. Raman spectroscopy, combined with LA-ICP-MS analysis also revealed that soil composition and groundwater flow are the conditions which most affect diagenesis at Ban Non Wat.     

Serum Proteins in Archaeological Human Bone

Research on the decomposition of bone collagen offers the key to a wealth of hidden information, for instance, to understanding palaeoclimatic conditions; habitat-specific parameters such as altitude; and for the reconstruction of palaeodiet and subsistence patterns. Radiocarbon dating, one of the most commonly used geochronological techniques is also preferentially carried out on bone collagen. Because negatively charged ions, and especially phosphate groups, are responsible for the tight bonding between organic molecules and bone, soluble serum proteins need not necessarily be leached from buried bone. The authors' hypothesis is that intruding minerals, aided by recrystallization of the bone mineral matrix, and colloid formation, aided by humic substances, form protective layers that preserve serum proteins in bone even after long periods of burial. We have used electrophoresis and Western blotting to recover protein fractions from about 150 archaeological bones from various sites and epochs (up to 5500 BC ) and have succeeded in recovering, purifying all, and identifying some of the proteins. Molecular weight bands corresponding to albumin, transferrin and a-2HS-glycoprotein (A2HS) were frequently recovered. This paper presents the method for separating serum proteins from archaeological bone and for their identification. Twenty-five per cent of samples with the respective molecular weight band still gave a positive immunological reaction with antibodies. We conclude that non-collagenous proteins, especially serum proteins, may be well preserved in bone.