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Journal of Archaeological Science 38 (2011) 1784e1797

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Journal of Archaeological Science

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Intra-skeletal variability in trace elemental content of Precolumbian Chupicuarohuman bones: the record of post-mortem alteration and a tool for palaeodietaryreconstruction

A.-F. Maurer a,*, M. Gerard b, A. Person a, I. Barrientos c, P. del Carmen Ruiz c, V. Darras d, C. Durlet e,V. Zeitoun f, M. Renard a, B. Faugère d

a Laboratoire Biominéralisations et Environnements sédimentaires UPMC, ISTeP, UMR 7193, 4 Place Jussieu, 75252 Paris cedex 05, Franceb Institut de Minéralogie et Physique des Milieux Condensés IRD, UMR CNRS 7590 UMPMC, 4 Place Jussieu, 75252 Paris cedex 05, FrancecCEMCA Sierra Leona 330 Lomas de Chapultepec, 11000 Mexico, D.F., MexicodUMR 8096 CNRS-Paris 1, Archéologie des Amériques, 21 allée de l’université, 92023 Nanterre, FranceeUniversité de Bourgogne, UMR CNRS 5561 Biogéosciences, 6 bd Gabriel, 21000 Dijon, FrancefUMR 9993 CNRS-Musée Guimet, 19 avenue d’Iéna, 75116 Paris, France

a r t i c l e i n f o

Article history:Received 10 December 2009Received in revised form7 March 2011Accepted 8 March 2011

Keywords:ChupicuaroApatiteGeochemistryIntra-skeletal variabilityDiagenesisDietHydrothermalism

* Corresponding author. Earth System Science ResApplied and Analytical Paleontology, Institute of GeosJohann-Joachim-Becher-Weg 21, 55128 Mainz, Germafax: þ49 6131 39 24768.

E-mail address: anmaurer@uni-mainz.de (A.-F. Ma

0305-4403/$ e see front matter � 2011 Elsevier Ltd.doi:10.1016/j.jas.2011.03.008

a b s t r a c t

This study applies an intra-skeletal sampling strategy to examine post-mortem alteration of archaeo-logical human bone fromwest Mexico, and to reconstruct ancient diet. Human bone from the Chupicuaroculture (Mexico, Preclassic period) constitutes an ideal material with which to examine subsistencestrategies because the specific hydrothermal environment in which the population lived would haveprovided certain food components (hydrothermal waters and carbonates) with distinct signature in Ca,Mg, F, Li, Sr, Mn, V and U values. Four to ten samples were taken from the long bones of six skeletons.Bone trace element content (Ca, P, F, Mn, Mg, Na, Li, V, Zn, Rb, Sr, Ba, Y, La, Ce, Nd, Th, U) and bonealteration parameters (crystallinity, organic matter and secondary calcite content) were analysed at theintra-skeletal level. Stable isotopic signatures (bone d13C and d18Ocarbonate) and histological analyses werealso performed on a single bone from each individual. Results indicate that all of the skeletons wereaffected by post-mortem mineralogical, structural and geochemical transformations. Biological bone d13Cvalues seem preserved for most of the individuals but an increase in crystallinity accompanies depletionin bone d18O values. The combination of bone alteration parameters with bone elemental content showsthat in this very specific context, a widespread dissolution-recrystallisation is unlikely. Of the hydro-thermal tracers, Sr, F and Li were of particular interest because their retention in living tissues is relatedto the amount ingested. The intra-skeletal Li content does not reveal any pattern but Li depletion is notexcluded. In contrast, Sr and F show a progressive intra-skeletal diagenetic enrichment likely due togradual diffusioneadsorption processes. The bones with the lowest concentrations in these elements areassumed to yield the best representative ante-mortem values. The signal extracted from each skeleton,a very unusually high bone Sr, F and Li content, is interpreted as reflecting the consumption of the localhydrothermal products, which are also enriched in these elements.

� 2011 Elsevier Ltd. All rights reserved.

1. Introduction

The Chupicuaro population was one of the most important ofthe Preclassic cultures (ca. 600 B.C.eA.D. 250) in Western Meso-america. The Chupicuaro settled approximately 170 km northwest

earch Center, Department ofciences, University of Mainz,ny. Tel.: þ49 6131 39 23429;

urer).

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of Mexico City, along the Lerma River in a unique environmentwhere the archaeological settlements are principally locatedaround hydrothermal springs (Darras et al., 1999) (Fig. 1). These hotsprings produce raw materials such as iron oxyhydroxides, andsome are bordered by voluminous hydrothermal carbonatedeposits. Hydrothermal products may have been used forconstruction and ceramic production, as suggested by the excava-tion of lime used for mortar and coating, and by the red and whitepigments characteristic of the Chupicuaro ceramics (Darras andFaugère, 2001, 2005; Mikrut, 2003). Hydrothermal products, suchas water and carbonates may also have been consumed. For

Fig. 1. Location of the study area. A) Map of Mexico showing the location of the Acambaro valley. B) Map of the Acambaro Valley indicating spatial distribution of the Preclassicarchaeological settlements. Digitization: Rodolfo Avila (CEMCA) C) Close-up of the hydrothermal area with the archaeological site JR24 studied.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1785

example, in Mexico today, carbonates are used in an alkaline pro-cessing method (nixtamalization) to prepare the traditional corndough (masa), to improve its nutritional quality (Bressani et al.,1958; Katz et al., 1974). This dough is then used for making torti-llas, which are cooked on a comal (ceramic griddles). Althoughcomal fragments have been discovered in the Basin of Mexico(Middle Preclassic) (Niederberger, 1976), it is unclear whether theChupicuaro populations used this method.

Therefore, the location of the Chupicuaro settlements, togetherwith the close proximity of raw materials available for construction,ceramic production and diet, raise questions about the settlementstrategy of this population. To investigate the importance of thehydrothermal springs in theeconomyof thesepre-Columbiangroups,an interdisciplinary study, The Chupicuaro Project, combiningarchaeology, geomorphology, sedimentology and geochemistry wasdesigned. Convergences between geochemical composition of somerawmaterials and archaeological artefacts (Darras, 2004) indicate theutilisation of carbonates and iron oxides in craft production. Thepresent study examines the geochemical composition of Chupicuaroskeletons to determinewhether they yield a signal attributable to theincorporation of some hydrothermal components in their diet.

Modern vertebrate bone geochemistry is directly related to thefood and water consumption. The bone signature is an average ofthe geochemical composition of the main foodstuffs ingestedduring the last 10e30 years before death, according to the turn-over rate of the anatomical part considered (Marshall et al., 1973;Tanaka et al., 1981). Major, minor and trace element content (Ca,P, F, Na, Mg, Sr, Ba, Zn, Mn, Li, V, U, Rb, Y, La, Ce, Nd, Th) of Chupi-cuaro bone mineral (called carbonate-hydroxylapatite or bioapatite

Ca10 [(PO4)6�x (CO3)x](OH)2, Chang et al., 1996) and stable isotopiccomposition (d13C and d18Ocarbonate) are examined in this study. Theelemental content of the hydrothermal carbonates and waters ofthe area is analysed for comparison with the skeletons.

Trace elements provided by the diet follow different metabolicpathways depending on whether they are essential (ETE) or not(NETE) (i.e. vital or non-vital) for the organism. ETE (Ca, Mn,Mg, Na,P, F, Zn.) are subject to homeostatic control mechanisms thatinclude regulation of absorption, excretion and tissue retention(Combs, 2005). Bone mineral plays a major role in those processesbecause of its non-stoichiometric properties (Weiner and Traub,1992), nano size of the crystals (Posner, 1987), high specificsurface, and carbonate content (NeumanandNeuman,1958;Weinerand Traub, 1992). The homeostatic mechanisms are maintained forvarying nutrient intakes as long as ETE are ingestedwithin adequateranges. When ETE ingestion is deficient or in excess, homeostaticregulation cannot be properly achieved (WHO/IPCS, 2002). Conse-quently, hypoorhyper concentrationof ETE leads to awidevarietyofclinical effects (Tapiero and Tew, 2003; Lindh, 2005). In contrast,NETE are not regulated. Instead, they accumulate in the organsdepending on the amount ingested. Among these elements, Sr, Baand Pb are mainly stored in the skeleton (Bauer et al., 1956) and aretherefore commonly used to reconstruct the diet of past populations(Toots and Voorhies, 1965; Sillen and Kavanagh, 1982; Balter et al.,2002; Sponheimer et al., 2005). Bone isotopic composition is alsodiet-related. The d18O value of biological carbonate-hydroxylapatitemainly records the d18O of drinkingwater (Longinelli, 1984; Luz andKolodny, 1985; D’Angela and Longinelli, 1990; Bryant et al., 1994;Delgado Huertas et al., 1995) whose isotopic composition is mainly

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e17971786

influenced by geoenvironmental factors (Dansgaard, 1964). Boneapatite d13C records the isotopic composition of all dietary compo-nents such as proteins, lipids and carbohydrates (Ambrose andNorr,1993; Tieszen and Fagre, 1993; Schwarcz, 2000). In continentalecosystems, d13C is driven by the isotopic variability in the photo-synthetic pathways of different plants: �13& (ranging from �15&to �10&) for C4 plants (some shrubs, tropical grasses) and �27&(ranging from �35& to �21&) for C3 plants (trees, herbs, majorshrubs, and shady grasses) (O’Leary, 1981; Ehleringer, 1989).

Unfortunately, the dietary signal may be modified by burial. Thefossilization process is complex and multiphase (Trueman andMartill, 2002). The degradation of the organic matrix exposesbone crystals to soil solutions with which chemical interactionsoccur. This results in loss, addition or exchange of some elementsacquired in vivo by the bone mineral. These geochemical trans-formations are accompanied by crystal maturation (improvementof the size of the homogeneous crystallites, elimination of defects)(Weiner and Bar-Yosef, 1990). In addition, diagenetic mineralsprecipitate in the pore spaces.

Paleodietary reconstructions using bone major and traceelement content are typically conducted on the diaphysis of longbones. However, long, compact bones may not always be preservedfrom alteration, as indicated by the highly variable intra-boneelemental content of archaeological skeletons (Keplinger et al.,1986). Here we propose to use the intra-skeletal variability intrace element content in order to control for post-mortem changes.Careful selection of the least altered bones enables us to reconstruct

Table 1Information for the six adult skeletons studied, S1, S3, S6, S7, S8 and S9. The bones collectulna; L: left, R: right). The archaeological period, position of the body, pathologies, sex aindicated.

Burial Number ofsamples

Bones sampled Period Depth(cm)

Burial fill

1 10 2F, 2T, 2H, 2R, 2U Mixtlan �195 Clay mixed withand occupationaresidues

3 4 R, HD, 2F Chupicuaro �570 Ashes and calcarocks

6 10 2F, 2T, 2H, 2R, 2U Chupicuaro �620 Forest soil and a

7 4 2H, 2F Chupicuaro �530 Forest soil

8 10 2F, 2T, 2H, 2R, 2U Chupicuaro �590 Forest soil

9 10 2F, 2T, 2H, 2R, 2U Chupicuaro �600 Forest soil

Chupicuaro dietary habits to determine whether or not they wereinfluenced by their hydrothermal environment.

2. Material and methods

2.1. The Chupicuaro setting: JR24 e La Tronera

The JR24 site is located on the right bank of the Lerma River inthe Acambaro Valley, near the modern hamlet of Puruagüita (stateof Guanajuato, Mexico). It is located on an eroded Pliocene lava flowand near three hot springs with hydrothermal carbonate deposits(Fig. 1c). The archaeological excavations performed in 2000/2001/2002 uncovered 27 strata and 21 occupation layers composed ofsands, silts and carbonates covering a consolidated pyroclasticsubstrate. The stratigraphic deposits (5 m deep) reveal that the areahad been occupied from Early Chupicuaro to Mixtlan phases(600e400 B.C. to A.D.1e250; Darras and Faugère, 2001, 2005). Nineprimary individual burials were uncovered, two of newborns, oneof a young child, and six of subadults or adults. The burials consistin simple pits digged in forest soil or anthropogenic sediment. Mostof the individuals were buried within small circular funerarystructures (Darras and Faugère, 2001) with the exception of indi-viduals S3 and S8. The bone samples used in this study werecollected from the adult specimens. In total 48 samples werecollected from six different skeletons (S1, S3, S6, S7, S8 and S9)(Table 1). One skeleton (S1) is dated from theMixtlan phase and theothers from the Late Chupicuaro phase (400e100 B.C.). The adult

ed from each individual are specified (F: femur, T: tibia, H: humerus, R: radius and U:nd age are given. The depth of the burial pit and the sediment used to fill it are also

Position of the body Pathologies Sex/age

gravell

Left lateral decubitusE-W cranial deformation

Caries, 3rd gradewearconsolidated fracture(L rib)slight exostose(lombar/cervicalcolumn)

Woman35e40 years

reous Extended dorsal decubitusE-W cranial deformation

Caries, 1st gradewearslight exostose(lombar column)

Woman30e45 years

shes Extended dorsal decubitusE-W cranial deformation

Caries, 3rd gradewear

Man25-30 years

Extended dorsal decubitusE-W cranial deformation

No wear, no carieinter vertebraldisc necrosisgrowth arrest lines

Woman18-20 years

Extended dorsal decubitusE-W cranial deformation

Caries, 3rd gradewearexostose (lombarcolumn)

Man30-35 years

Extended dorsal decubitusNE-SW cranial deformation

Caries, 3rd gradewearconsolidatedfractures (R clavicle,R coxal)exostoses (cervical/lombar column,R ulna and radius)Cifosisosteomyelitis(R ulna and radius)slight periostis(L tibia)dental fluorosis

Woman 35-40years

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1787

remains were oriented east/west, with the skeleton lying in dorsaldecubitus, and the upper limbs positioned alongside the thoraxwith the skull to the west or to the east. The exceptionwas skeletonS1 which was lying in dorsal decubitus with flexed legs on the leftside (Darras and Faugère, 2001).

In addition, the following samples were obtained for comple-mentary analyses:

- a modern cow bone discovered at the surface of JR24 and usedas a reference sample,

- hydrothermal and non-hydrothermal samples: water (7 hydro-thermal, 3 rivers, 4 springs) collected in the area of Puruagüita,hydrothermal carbonates sampled at Puruagüita (3 samples)and the carbonate fraction of hydrothermal deposits, 20 kmwestward, at Aguas Calientes hot springs (2 samples) (Fig. 1).

Fig. 2. Histological sections of the Chupicuaro bones (skeletons S1, S3, S6, S7, S

The cortical bone samples were cut using a drill equipped witha diamond saw. After a mechanical cleaning, they were divided intwo aliquots: one for histological and crystallinity examination andthe second for chemical analysis.

2.2. Histological analysis

Transverse bone sections were embedded into a polyesterresin under vacuum, before being cut with a diamond saw, gluedon glass slides and polished with cerium oxide. The final thick-ness of the thin sections is 30 mm. One thin section was examinedfor each skeleton (Fig. 2) using optical transmitted light micros-copy. The histological index (HI) of Hedges et al. (1995) was usedin order to estimate the quality of preservation of the fossilbones. HI varies from 5 for fresh bone (all structures are

8 and S9; F: femur, H: humerus; L: left and R: right) under polarized light.

Table 2NIST SRM 1400: recovery data for certified and non-certified analytes. All elementscertified in SRM were accurately determined by the proposed method. Concentra-tions in italics are below the quantification limit. Analyses were performed on 21samples at different times.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e17971788

distinguished) to 0 when less than 5% of the bone is intact. Whenthe thin sections showed differential histological preservation,a mean index was assigned, based on the percentage of intactbone.

Analyte Target Measured % Recovery

a Ca % 38.18 � 0.13 38.51 � 0.66 101a P % 17.91 � 0.19 18.01 � 0.17 101a Mg % 0.684 � 0.013 0.665 � 0.014 97b Na % 0.6 0.61 � 0.04 101b F % 0.125 0.114 � 0.006 91a Sr mg g�1 249 � 7 252 � 5 101c Ba mg g�1 240 � 10 241 � 7 100a Zn mg g�1 181 � 3 178 � 6 98b Mn mg g�1 17 15 � 1.0 86c V mg g�1 0.77 � 0.09 0.78 � 0.14 101c Li mg g�1 0.95 � 0.14 1.13 � 0.10 118c U mg g�1 0.066 � 0.003 0.060 � 0.005 92c Y mg g�1 0.29 � 0.03 0.23 � 0.02 80c La mg g�1 0.39 � 0.08 0.28 � 0.02 74c Ce mg g�1 0.82 � 0.10 0.56 � 0.07 68c Rb mg g�1 0.71 � 0.13 0.47 � 0.01 67c Nd mg g�1 0.32 � 0.02 0.30 � 0.02 94

2.3. Crystallinity analysis and the secondary calcite content of bonevoids

For bulk mineralogical analyses, bones samples were groundwith an agate mortar and pestle. The powder was pressed ina glass-aluminium sample-holder. A Siemens D500 diffractom-eter with Ni-filtered CuKa radiation at 40 kV and 30 mAwas usedfor X-ray diffraction analysis (XRD). Samples were scanned from2� to 70� (2q), with counting for 6 s every 0.02�, in a rotatingsample-holder. The CI (Crystallinity Index) was determined usingthe semi-quantitative method of Person et al. (1996). A semi-quantitative estimation of the calcite content (precipitatedwithin the pore structure) was calculated as Surfacecalcite[104]/Surfaceapatite[002].

c Th mg g�1 0.12 � 0.00 0.11 � 0.01 87

a- NIST certified concentrations, b- NIST informational value, c- Hinners et al., 1998.

2.4. Chemical analysis

2.4.1. Pretreatments and sample digestionBone powder was etched in dilute acetic acid (0.1 M) for 1 h.

The residual material was rinsed and then used for heat treat-ment in order to remove the bone organic matter (OM). A lowtemperature (300 �C) heat treatment was applied to maximisethe removal of OM while minimising microstructural and ultra-structural changes to the bone mineral. Bone powder, in a plat-inum beaker, was placed into a closed system furnace undervacuum for 6 h to distil the OM. This was followed by heattreatment in a muffle furnace at 300 �C for an additional 20 h tooxidize the OM carbon. This step was achieved by a pure O2 flowfor 1 h. This protocol ensures the complete destruction of boneOM and allows an estimation of the bone protein content (bone %weight loss).

A 100 mg sample of the resulting ash was dried overnight at80 �C and weighed into a Teflon beaker before being digested in5ml of 20% nitric acid (Merck, suprapur) and heated at 80 �C for 1 h.The resulting solution was then diluted with deionized water toa total volume of 50 ml, stocked in polypropylene or Teflon tubesand kept for analysis.

2.4.2. Trace element and stable isotope analysisBone chemical analyses were performed using a Varian Ultra-

Mass ICP-MS for bone Li, V, Zn, Rb, Sr, Ba, Y, La, Ce, Nd, Th and Ucontents. Bone Ca, Mn, Mg and Na contents were analysed witha Varian ICP-AES. In order to avoid matrix effects from majorelements (essentially Ca and P) on bone trace element content, weapplied a “standard” addition method. The standard matrix wasprepared with NIST SRM1400 “bone ash” digested with nitric acid,with a 200-fold dilution for Ca, and a 5-fold dilution for the otherelements. The standard calibration was prepared from Spex,1000 mg/ml certified stock. The quality control and method vali-dation were performed by analysis of NIST SRM1400 “bone ash”(Table 2). Fluoride concentrationsweremeasured by potentiometryusing a specific electrode. PO4 was measured photometrically.Isotopic analyses were performed using a mass spectrometer VGSIRA9. Samples were reacted under vacuum with orthophosphoricacid overnight at 50 �C.

All water and carbonate analyses were performed by ICP-MSand ICP-AES at the Institut de Recherche pour le Développement(Bondy, Paris) and the Centre de Recherches Pétrographiques etGéochimiques in Nancy (France), respectively.

3. Results

3.1. Hydrothermal products

Trace element data from analysis of hydrothermal waters andtheir associated recent carbonate caps, rivers and wells fromPuruagüita, as well as carbonates from Aguas Calientes, are pre-sented in Table 3. Puruagüita hydrothermal springs are slightlyacidic (mean pH 6.3) and warm (mean T�C 39.3 �C). The non-hydrothermal waters sampled from adjacent rivers and wells(referred to as “cold waters” below) are more basic (mean pH 7.5and 7, respectively) and thewells are colder (mean T�C 25.4 �C). Thehydrothermal waters are highly enriched in Cl, Na, Li, Sr and Rb(over 10 times) and enriched in HCO3

�, Ca, Mg, F, Ba, Mn, Cr, Cu, Y,Nd and La compared to the cold waters. In contrast, hydrothermalwaters are highly depleted in U and SO4

2� and slightly depleted inV compared to cold waters. Carbonates from Puruagüita hot springsare highly enriched in Sr but highly depleted in Rb, V, Y, Nd, La, U, Cecompared to the other hydrothermal deposits sampled at AguasCalientes. Mg, Ca, Ba, Li and Mn enrichment and Cu, Cr and Nadepletion in carbonates from Puruagüita are also significant.

None of the hydrothermal products (water and carbonate) ana-lysed exceeds the Tolerable Upper Intake Levels (Food and NutritionBoard, Institute of Medicine, 1997, 2001, 2004) considered fora normal diet (i.e. 1 or 2 L of water and several g of carbonatesingested per day).

According to the composition of hydrothermal products (watersand carbonates), Ca, Mg, F, Li, Sr, Ba, Mn, V and U contents can beused as tracers of hydrothermalism in the area of Puruagüita. Theircontent in Chupicuaro bones is therefore of particular interest. Bahowever will not be discussed further. Its incorporation into bone,like Sr, is related to the amount of calcium ingested (Elias et al.,1982) and although Ba/Ca differentiates hydrothermal depositsfrom Puruagüita and Aguas Calientes, it is not a good tracer for thehydrothermal waters analysed (Table 3). Conversely, Sr/Ca issignificantly higher in these potential dietary components.

3.2. Human bones from La Tronera

Trace elements, OM, calcite contents, stable isotope composi-tion, CI and HI of Chupicuaro human bones are summarized in

Table 3Composition of hydrothermal products (waters and carbonates, grey zones). The average value and the standard deviation of the samples are given. The number of samplesanalysed is given in brackets. “Acceptable” concentrations in water as well as recommended or tolerable intakes are presented for most elements/species. Font style andsymbols (bold, italic, * and **) correspond to individual references listed below the table.

ELEMENTS

in water

RDA AI ULmean pH

mean T°C

µg.l-1 µg.l-1 µg.l-1 µg.l-1 µg.j-1 µg.j-1 µg.g-1 µg.g-1

HCO3- 989 ±52 .103 209 ±34 .103 94 ±48 .103

Cl- 730 ±46 .103 16 ±6 .103 5 ±5 .103 250 .103 2300 .103 3600 .103

Na 669 ±47 .103 44 ±11 .103 18 ±7 .103 200 .103 1500 .103 2300 .103 9 ±5 .103 15 ±0.7 .103

Ca 113 ±12 .103 24 ±2 .103 13 ±9 .103 1000 .103 2500 .103 338 ±6 .103 109 ±13 .103

Mg 14 ±4 .103 8 ±1 .103 4 ±2 .103 310-420 .103 350 .103 11 ±2 .103 3 ±0.8 .103

F- 3.6 ±0.4 .103 1.8 .103 0.7 ±0.6 .103 4-1.5 .103 3 4 .103 10 .103

NO3- 2.4 ±0.2 .103 2.5 ±1.6 .103 7 ±3.5 .103 10 .103

SO42- 0.6 .103 8 ±11 .103 11 ±9 .103 500 .103 ND .103

Li 3366 ±62 54 ±56 11 ±8 *1000 **500000 184 ±103 37 ±1

Sr 2428 ±142 252 ±28 127 ±94 5163 ±694 140 ±30

Rb 347 ±13 30 ±7 31 ±15 13 ±6 122 ±4

Ba 338 ±62 110 ±83 36 ±35 2000 1000 902 ±211 133 ±29

Mn 205 ±31 63 ±94 0.5 ±0.3 50 1800-2300 11000 0.5 ±0.2

Cr 9 ±3.5 2 ±1.3 1 ±0.5 100-50 25-35 ND 2 ±2.2 13 ±3

V 1.9 ±0.9 5.2 ±4.9 5.1 ±3.4 unregulated 1800 1 ±0.9 22 ±6

Cu 1.5 ±0.8 0.2 ±0.1 0.1 ±0.1 1300 1000 900 10000

Y 63 ±26 .10-3 26 ±14 .10-3 9 ±3 .10-3 2000 * .10-3 703 ±166 .10-3 22550 ±780 .10-3

Nd 14 ±10 .10-3 2 ±1 .10-3 1 ±1 .10-3 1050000 * .10-3 411 ±163 .10-3 10800 ±0 .10-3

La 9 ±5 .10-3 4 ±2 .10-3 2 ±1 .10-3 2000 * .10-3 499 ±210 .10-3 16350 ±1202 .10-3

U 8 ±8 .10-3 444 ±419 .10-3 394 ±301 .10-3 30000-20000 .10-3 34 ±30 .10-3 3825 ±21 .10-3

Ce 5 ±3 .10-3 10 ±5 .10-3 12 ±11 .10-3 2000 * .10-3 1022 ±197 .10-3 34500 ±2263 .10-3

MCL: Maximum Contaminant Level (US EPA, 2009) RDA: Recommended Dietary Allowances; AI: Adequate Intakes

MAC: Maximum Acceptable Concentrations or OG: Operational Guidance value(FPT Committee on Drinking Water, 2008)

UL: Tolerable Upper Intake Levels

* iAC: indicative Admissible drinking water Concentrations (de Boer et al., 1996)

from Food and Nutrition Board, Institute of Medicine, National Academies, 1997, 2001, 2004

* from Schrauzer, 2002** from Anke, 1993

(2)

GUIDELINES FOR HEALTHY LEVELS HYDROTHERMAL DEPOSITS

PuruagüitaAguas

Calientes

6.339.3

7.5 725.4

intake

DRI: Dietary Reference Intakes

hydrothermal rivers wells(7) (3) (4)

WATERS

(3)

2.4 ±2 22 ±7

Ba/Ca 0.0030 ±0.0004 0.0045 ±0.0033 0.0025 ±0.0013 0.0027 ±0.0006 0.0012 ±0.0004

Sr/Ca 0.022 ±0.001 0.010 ±0.001 0.009 ±0.002 0.015 ±0.0022 0.0013 ±0.0004

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1789

Table 4 (see Appendix A for more detail). Complementary data froma modern cow bone are also presented.

3.2.1. Mineralogical and structural parametersSkeletal Ca/P ratios vary between 2.19 (S8) and 2.28 (S9) with an

average value of 2.24 � 0.03. CI ranges from 0.07 (S1) to 0.28 (S9)with an average value of 0.20 � 0.07. Skeletal OM content variesbetween 10% (S7) and 27% (S1), with a mean value of 17 � 6%. Thethin sections show that all skeletons are affected bymicro-bacterialattacks with HI ranging from 1 to 3 (Fig. 2). Secondary calcite ispresent in all bones except one (S1).

3.2.2. Geochemical composition3.2.2.1. Trace element content. Trace element content of the humanskeletons differs from freshmodern cow bone, with higher F, Sr, Ba,Zn and Li. The Chupicuaro bones also display higher Mn, V, U values

compared to themodern cowbone for which these elements do notexceed the quantification limit. Although around the quantificationlimit, Rb, Y, Ce, La, Nd andTh content of the humanbones are slightlyhigher (except Rb) than that of the modern reference sample. Incontrast, Na andMg content of the archaeological human bones arelower than the content analysed in the modern animal bone.

Most elements display an intra-skeletal coefficient of variationof between 10% and 25% (F, Na, Mg, Sr, Ba, Zn, Li, V, U). This vari-ability is higher (27%e63%) for Rb, Y, La, Ce, Nd and Th content,which are around the quantification limit. Bone Mn contentdisplays the highest intra-skeletal variability (80%). The intra-skeletal variability of the elements that are used as tracers forhydrothermalism is mainly driven by the upper limb bones thattypically display higher F, Sr and Mn concentrations (Fig. 3) andlower Mg content (except for S8). For individual S9, F and Srenrichment is higher in the tibias. No general intra-skeletal pattern

Table

4Geo

chem

ical

composition(trace

elem

ents,stableisotop

es),mineralog

ical

andstructuralp

aram

eters(organ

icmatterOM,C

rystallin

ityIndex

CI,calciteco

ntenta

ndHistologicalIndex

HI)of

thestudiedhuman

skeleton

s.Th

emea

nskeletal

values,aswella

stheintraan

dinter-skeletal

coefficien

tof

variations(cv)

aregive

n.D

atafrom

amod

ernco

wbo

ne(non

-pretrea

tedwithacetic

acid)from

site

JR24

arepresentedforco

mparison

.Datain

italicsarebe

low

thequ

antification

limit.T

hebo

ldwas

usedto

highlig

httheskeleton

numbe

r.

Skeleton

sJR24

FNa

Mg

SrBa

ZnMn

LiV

URb

YLa

Ce

Nd

Thd1

3Cd1

8OCa/P

CI

OM

Calcite

HI

%%

%mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1mgg�

1&

&%

Mea

nS1

0.51

0.61

0.26

3271

1492

186

630

1517

0.23

0.46

0.28

0.46

0.31

0.08

L3.2

L4.4

2.22

0.07

270

2cv

(%)

237

1310

2013

5322

1738

2434

3534

1621

138

11Mea

nS3

0.49

0.45

0.18

2133

1065

213

2338

2654

0.17

0.32

0.14

0.30

0.19

0.05

L1.3

L5.6

2.27

0.22

170.15

1cv

(%)

185

69

1313

7216

1620

4493

3778

3828

118

188

Mea

nS6

0.61

0.84

0.14

1848

664

293

3924

3257

0.16

0.77

0.22

0.32

0.27

0.07

L1.7

L5.0

2.22

0.16

210.45

1cv

(%)

4392

810

2033

103

1519

2323

108

5555

4824

227

1311

4Mea

nS7

0.73

0.46

0.19

2572

1352

175

1445

2748

0.33

0.13

0.10

0.14

0.13

0.06

L1.6

L4.8

2.24

0.23

100.41

3cv

(%)

187

86

1317

128

48

2061

5342

5938

401

2610

21Mea

nS8

0.39

0.43

0.19

2023

766

209

2034

2436

0.11

0.30

0.11

0.21

0.15

0.04

L1.1

L5.2

2.19

0.25

140.43

2cv

(%)

198

1313

1819

649

1515

1033

2137

1919

219

1668

Mea

nS9

0.99

0.48

0.18

2242

1120

193

4023

3928

0.33

1.51

0.44

0.90

0.48

0.10

L2.3

L5.3

2.28

0.28

160.31

2cv

(%)

1418

127

1439

7711

1518

7756

2526

2632

211

1672

Mea

n0.62

0.54

0.19

2348

1076

212

2432

2740

0.22

0.58

0.21

0.39

0.26

0.06

L1.9

L5.0

2.24

0.20

170.29

2Intra-skeletal

cv(%)

2323

109

1622

8313

1523

4063

3648

3127

223

1457

Inter-skeletal

cv(%)

3529

2122

3020

5826

3039

4187

6070

5133

419

137

3462

41Modernco

wbone

0.00

0.79

0.45

832

156

132

1.7

10

00.46

0.04

0.03

0.04

0.05

0.02

�3.41

�2.91

2.19

037

05a

aHed

geset

al.,19

95.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e17971790

is observed for Li (except in S6), V (except in S6 and S1) and Ucontent. Inter-individual variability varies between 20 and 30% forNa, Mg, Sr, Zn and Li, 30 and 40% for F, Ba, V, U and Th, between 40and 50% for Rb and by more than 50% for Mn, Nd, La, Ce and Y.

3.2.2.2. Stable isotope composition. Bone d13C varies between�3.2&(S1) and �1.1& (S8) with a mean skeletal value of �1.9 � 0.8&.Bone d18O ranges from �5.6& (S3) to �4.4& (S1) with an averagevalue of �5.0 � 0.4&.

4. Discussion

4.1. Patterns of bone alteration

The increase in CI, the precipitation of secondary calcite and thedegradation of OM are common in archaeological bones (Personet al., 1995; Saliège et al., 1995; Nielsen-Marsh and Hedges, 2000;Trueman and Tuross, 2002; Reiche et al., 2003; Smith et al.,2007). The precipitation of calcite indicates contact of bones withgroundwater. The increase in crystallinity is often referred to asdissolutionerecrystallisation process (Trueman and Tuross, 2002)which would delete the dietary signal. However, bones submittedto a heat treatment to monitor diagenetic effects (Person et al.,1996; Munro et al., 2007, 2008; Zazzo et al., 2009) show that lossof CO3 radicals in the carbonate-hydroxylapatite also improves thecrystallinity without necessarily implying a dissolution-recrystal-lisation process (Person et al., 1996).

The intra and inter-skeletal variability of these parameters(Fig. 4ab) showgradual changes of bone tissues during burial withinthe same archaeological site for contemporaneous (except S1)human skeletons. Bacterial attack (Fig. 2), which is an early diage-netic process that starts soon after death (Yoshino et al., 1991; Bellet al., 1996) appears to be independent of these changes (age, crys-tallinity, OM and calcite content). The analysis of these bone struc-tural changes gives a first estimation of the skeletons’ state ofpreservation. Individual S1 seems to be the least altered of theskeletons, with a high OM content, a low CI and no calcite. Thisskeleton is more recent (Mixtlan phase) than the other Chupicuaroskeletons. A relationship between crystallinity increase and age isnot usually observed except perhaps during the early diagenesisphase (Person et al., 1996; Sillen and Parkington, 1996). Skeleton S1,together with skeletons S3, S8 and S9 represent the general diage-netic trajectories. The slight deviance of skeletons S6 and S7 fromthese general trends may indicate different taphonomic histories.Skeleton S6 shows a high calcite content in spite of a fairly high OMcontent, suggesting an important contact with groundwater. Incontrast, the increase in CI of skeleton S7 (except for the righthumerus) seems tohavebeendelayed, in spiteof its lowOMcontent.

The analysis of bone d13C and d18Ocarbonate was conducted inorder to further investigate Chupicuaro diet. However, bone CI iscorrelated with bone d18Ocarbonate. In contrast, bone d13C does notshow any apparent correlation with bone CI (Fig. 5). An increase incrystallinity associated with a severe depletion of bone d18Ocarbonatewithout any significant modification of bone d13C, was alsoobserved during experimental heating (Munro et al., 2008). Theabrupt shift in bone d13C (w1.5&) at around CI 0.10e0.14 separatesthe skeletons studied into two main groups 1) containing only twobones from the individual S1 and 2) containing bones from indi-viduals S6, S7, S8 and S3, whose d13C values are very similar(average �1.42 � 0.24&). The bone d13C value of individual S9 liesbetween the two groups (d13C ¼ �2.33&) and has the highest CI(0.28). It is difficult to know whether or not it results from diage-netic exchange with carbonates of the diagenetic calcite.

Bone trace element content varies at the intra and inter-skeletallevel. Oxides may be responsible for the large intra (83%) and inter-

Fig. 3. Intra-skeletal trace element cartography. Content of the hydrothermal tracers in bones (Mg, F, Sr, Li, V, U and Mn) are reported for each part of the skeleton. The grey zoneshighlight mostly the upper limb bones (S1, S6 and S8) and the tibias in the case of S9.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1791

individual (58%) variability in bone Mn content. They arecommonly found in archaeological bone pore structure (Williamsand Potts, 1988). Mg is an essential trace element and is thereforeregulated by homeostatic mechanisms. However, although theintra and inter-skeletal variability in bone Mg content is fairly low(10% and 21%, respectively), all of the human skeletons studieddisplay a Mg depletion, compared to modern bones, which isprobably related to the decay of the collagen matrix (Fig. 4c). S6 isparticularly affected as it shows a significant Mg depletion despitesubstantial OM content. This depletion, together with this skele-ton’s calcite content, argues for an important leaching mechanismthat mainly affected the upper limbs. The S6 upper limb bones alsodisplay the minimum intra-skeletal Li content, although no generalintra-individual pattern is observed for this element (Fig. 3). Lidepletion during burial is therefore not excluded when consideringa unidirectional diagenetic trajectory for this element (i.e. notpreceded by early post-mortem enrichment). In that case, the highLi values found in Chupicuaro bones would not be attributable todiagenetic enrichment. Bone Mg leaching is also accompanied bya diagenetic enrichment in F, attested by the very high values of thehuman skeletons compared to the modern cow bone, the very highintra-skeletal variability of individual S6’s F concentrations (43%),the correlation between bone F and calcite content and the corre-lation between bone F and V content (Fig. 6a and b). S8 and S1 alsodisplay correlations between F and calcite and between F and Vcontent respectively (Fig. 6c and d). All of the skeletons show post-mortem F addition regardless of their skeletal F content and intra-skeletal variability. As Sr concentrations are related to those of F foreach skeleton (Fig. 7), bone Sr content also results from a post-mortem enrichment process, in spite of the generally low intra-skeletal variability (mean value 9%). In most cases, the upper limbsyield the highest Sr and F content.

Trace element uptakemechanisms remain obscure (Kohn, 2008)and it is therefore difficult to explain the exact mechanism for suchSr, F and V enrichment in the Chupicuaro human bones. It is worth

noting that with only two exceptions (S9 left humerus and righttibia), all of the skeletons display a Ca/P ratio that is in the biologicalrange 2e2.3 (Trueman and Tuross, 2002). This result strengthensthe idea that this parameter is not a relevant diagenetic proxy(Hubert et al., 1996; Fabig and Herrmann, 2002). However, otherparameters do show that dissolutionerecrystallisation processesseem unlikely:

i) Chupicuaro bones REE content is close to 1 ppm, typicalconcentrations for in vivo bone (Trueman and Tuross, 2002);

ii) Bone F, Sr and V content are not associatedwith an increase incrystallinity at the intra-skeletal scale, nor at the populationlevel;

iii) The U diffusioneadsorption process operating on bone crys-tallites (Badone and Farquhar, 1982; Millard and Hedges,1995, 1996; Simon et al., 2008) may explain the relativelyhomogeneous intra-skeletal U content, the significant humanbone U content, and the absence of any relationship betweenU and other elements or mineralogical parameters.

The intra-skeletal variability observed in this study cannot beattributed to a biological repartition, which would imply similarpatterns for the skeletons studied. It is therefore due to taphonomicprocesses and probably related to differential kinetic diffusionparameters depending on the chemical elements, the part of theskeleton and the individuals. The skeletal parts least affected by theprogressive enrichment of Sr, F and V concentrations (generallylower limb bones) should therefore be considered the mostrepresentative with regard to the in vivo signature of the Chupi-cuaro bone.

4.2. Consumption of hydrothermal products

Of the tracers of Puruagüita hydrothermalism (cf. 3.1.), Li, F and Srare of particular interest because their retention in living tissues

modern cow bone

S9S8S7S6S3S1

a

b

c

JR24

5 10 15 20 25 30 35 40

OM (%)

CI

CI = 0.43 - 0.01 * OM R = 0.86

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.00

0.40

1.20

1.60

2.00

0.80Bon

eca

lcit

e co

nten

t

0.05

0.15

0.25

0.35

0.45M

g (%

)

Fig. 4. Relationships between bone organic matter content (OM) and (a) bone crys-tallinity (CI), (b), bone calcite content, and (c) bone apatite magnesium content. Theresults are given for all of the bones analysed from each human skeleton from site JR24Data from a modern cow bone are given for comparison. The magnesium content of allof the bone samples (archaeological and modern) was obtained after acetic acid pre-treatment.

-6

-5.6

-5.2

-4.8

-4.4

-4

0 0.05 0.10 0.15 0.20 0.25 0.30

CI 18

Oc

‰

18O= -4.08 - 4.84 * CI R = 0.89

-3.5

-3

-2.5

-2

-1.5

-1

13 C

‰

S9S8S7S6S3S1

JR24

Fig. 5. Relationships between bone crystallinity (CI) and bone isotopic composition(d18Ocarbonate and d13C). Two samples were analysed from skeleton S1 (right humerusand left tibia) and one sample was analysed for each of the other skeletons: the leftfemur of S7 and S3, the left humerus of S6 and S8 and the left tibia of S9.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e17971792

(and especially in bones for F and Sr) is determined by the amountingested. The concentration of these elements in hydrothermalproducts is therefore compared to the data extracted from eachskeleton (i.e. the best representative concentrations of the in vivosignature).

Except for its therapeutic role in treating bipolar affectivedisorders (Schou, 1968), Li does not appear to be an essentialelement for life (Léonard et al., 1995) and its biochemical functionin vital components of the body is not known (Anke et al., 2005).Plasma Li concentrations vary according to the Li intakes (Zaldivar,1980; Schrauzer, 2002). Following ingestion, non-excreted Li rea-ches different target organs such as cerebellum, cerebrum, kidneys,hair and liver (Schrauzer, 2002; Anke et al., 2005). Li is also retainedand released by bone (Schrauzer, 2002; Kosla and Anke, 2005). Thelack of human bone Li concentration data associated with knowningested concentrations makes it impossible to calculate the Liingested from the measured Li content of the Chupicuaro bones.However, Li content has been measured in modern Japanese ribs asless than 0.08 mg g�1 (Yoshinaga et al., 1995). Assuming that this isrelated to a daily dietary Li intake of around 0.06 mg in Japan(Shiraishi, 2005), the very high Li content of the Chupicuaro skel-etons (mean 32 mg g�1) can be hypothesized to be the result of the

ingestion of hydrothermal waters, which would supply at least3.3 mg of Li to the Chupicuaro per day from water intake alone.

F is considered essential, or at least potentially essential, in traceamounts (Lindh, 2005). It is mainly incorporated in the skeleton(Cerklewski, 1997). Homeostasis occurs particularly during thegrowth period, through the combined effects of skeletal uptake andurinary excretion (Cerklewski, 1997). However, F absorption andplasma F concentrations are not regulated (Whitford, 1996;Cerklewski, 1997). Plasma F content, and in turn bone, is partlyrelated to the amount of F ingested (Ekstrand et al., 1977). A studyconductedon the iliac crestofmodernAmericanswhowereexposedto variable water F content (Zipkin et al., 1958) illustrates this rela-tionship (Fig. 8). The Chupicuaro skeletal F concentrations are inagreementwith the intakeof drinkingwater reachinganF contentofbetween 2.6 and 4 ppm. These concentrations are compatible withthose of the hydrothermal waters (3.6 ppm), close to the maximumcontaminant level (MCL) defined by the Environmental ProtectionAgency (cf. Table 3). The MCL was established in order to protectagainst crippling skeletal fluorosis, the most severe stage of thedisease. Fluorosis affects teeth and boneswhen the F intake exceedsa toxic level,which isnotparticularlywell-defined. Skeletalfluorosisis composed of four stages (preclinic, I, II, III or crippling), whichrange from weak symptoms to a significant alteration of mobility.Bone F concentrations and the levels at which skeletal fluorosisoccurs varywidely (Fig. 8).Male individuals S6 and S8 areunlikely tohave been affected by the disease. Conversely, female individuals 1,3, and 7mayhave experienced fluorosis, especially the early stage ofthedisease. The female individual S9 is theonlyonewhowouldhavebeen strongly affected by the disease, falling within stages II and III.Dental fluorosis was detected on S9’s teeth but no skeletal fluorosiswas observed (Barrientos and del CarmenRuiz, 2009). The very highskeletal F content of S9 can therefore be attributed to a post-mortemenrichment. The numerous pathologies (periostite, osteomyelite,enthesopathie) and fractures (Garcia, 2002) that affected this indi-vidual during her life very likely enhanced post-mortem F addition.

0

15.052.0

R = 0.83S8

Bone fluoride content (%)

0

21.12.0

R = 0.89S6 LU

femurstibias

humerusradius

ulnas

Bon

eca

lcit

e co

nten

t

a

bc

d

11

20

57.03.0

R = 0.85S1

22

44

1.12.0

R = 0.85S6

Bon

e V

µg/

g

Fig. 6. Intra-skeletal relationships between bone fluoride and bone calcite content (a and d) and bone fluoride and vanadium content (b and c) for three skeletons S1 (c), S6 (a andb) and S8 (d). The correlation coefficient, R, between bone fluoride and bone calcite content of skeleton S6 was calculated after exclusion of the left ulna (LU).

2700

4100

0.3 0.75

R = 0.71

1900

2400

0.38 0.58

R = 0.96

1400

2200

0.2 1.1

R = 0.93

2350

2800

0.55 0.9

R = 0.97

1500

2500

0.25 0.51900

2700

0.7 1.25

R = 0.70

Bon

e S

r co

nten

t µg

/g

Bone fluoride content (%)

femurstibiashumerusradius

ulnas

R = 0.75

RT

S8S9

S3

S7S6

S1

Fig. 7. Intra-skeletal relationships between bone fluoride and strontium content for all skeletons S1, S3, S6, S7, S8 and S9. The correlation coefficient, R, of skeleton S8 was obtainedafter exclusion of the right tibia (RT).

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1793

0

0.2

0.4

0.6

0.8

1

1.2

1.4

% F

hum

an b

ones

0

0.2

0.4

0.6

0.8

1

1.2

1.4

% F

hum

an b

ones

preclinic I II III

actual populationUSA

Zipkin et al., 1958

ppm F drinking water0 1 2 3 4

moderncowbone

JR24

Chupicuaro population skeletal fluorosis stages

S1S3

S6

S7

S8

S9

post

mor

tem

enr

ichm

ent

ante

mor

tem

val

ues

min

max

mean

cold waters hot waters

Fig. 8. Comparison of the fluoride content of Chupicuaro bones and modern bones (USA). Grey circles represent fluoride content from individuals whose drinking water’s fluoridecontent and time residence are known (Zipkin et al., 1958). Fluoride content of the waters of Puruagüita (cold and hot) is indicated. Skeletal fluorosis stages associated with iliaccrest or pelvis fluoride content are also shown (see a review of the Committee on Fluoride in Drinking Water/National Research Council, 2006). It is worth noting that the lowestvalue in stage III was known to be associated to hypocalcemia or secondary hyperparathyroidism.

0.0030

0.0040

0.0050

0.0020

0.0060

bone

Sr

/ Ca

0.0070

0.0074

0.0077

0.0108

0.0003/4

S1

S3

S6

S7

S8

S9

modern cow bone

post

mor

tem

enr

ichm

ent

ante

mor

tem

val

ues

min

max

mean

hydr

othe

rmal

wat

erri

ver

wat

erw

ells

hydr

othe

rmal

ca

rbon

ates

Pur

uagu

ita

0.3

0.2

0.3

0.2

0.3

0.2

bone Sr/Capure

hydrothermal product

consumer calculated for

0.2< ORSr <0.3

hydr

othe

rmal

de

posi

tsA

guas

Cal

ient

es

bone Sr/Cameasured in the

human skeletons and a modern cow bone

from site JR24

Fig. 9. Comparison of Sr/Ca ratios from Chupicuaro bones and a known pure hydrothermal product consumer. Ratios have been calculated for ORSr (Observed Ratio) valuescomprised between 0.2 and 0.3 and taking into account the average strontium and calcium content of each hydrothermal product (see Table 3).

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e1797 1795

Sr is a non-essential element that is also mainly stored in theskeleton (Staub et al., 2003). With chemical properties similar tothose of Ca, it tends to follow the same biological pathways. Thepreferential absorption and retention of Ca compared to Sr throughthe gastro-intestinal tract and the kidneys (Comar et al., 1957)results in the reduction of bone Sr/Ca through the trophic chain ateach tier (Burton et al., 1999). This reduction is quantified by anObserved Ratio ORSr (Sr/Cabone/Sr/Cadiet). Using ORSr it is possibleto predict the bone Sr/Ca content for a consumer of “pure” hydro-thermal products, taking into account the average Sr and Ca value ofeach ingredient (Fig. 9). ORSr has been determined for modernmammals and in laboratory controlled experiments, where itgenerally ranges between 0.20 and 0.30 (Comar et al., 1957; Sillenand Kavanagh, 1982; Price et al., 1985, 1986; Balter, 2004). Repor-ted values in humans are typically between 0.25/0.23 (Bryant andLoutit, 1961; Burton and Mercer, 1962), 0.18 (Alexander et al.,1956; Rivera and Harley, 1965; Sillen and Kavanagh, 1982) and0.12 (Tanaka et al., 1981). Using ORSr between 0.2e0.3, most of theChupicuaro skeletons display bones Sr/Ca ratios compatible withthe consumption of hydrothermal products from Puruagüita (waterand carbonates) (Fig. 9). Conversely, hydrothermal deposits fromAguas Calientes and “cold waters” are completely excluded fromthe Chupicuaro diet. A distinct behaviour is observed in skeletonsS7 and S1, whose higher Sr/Ca ratios do not appear to be related tothe intake of hydrothermal products.

4.3. Variations within the Chupicuaro population

Despite the limited number of individuals studied, three groupscan be distinguished: the two males S8 and S6; the females S3 andS9 and the two other females S1 and S7. Males S6 and S8 display thelowest bone Sr and F values (Figs. 8 and 9). This is likely due tophysiological differences between the sexes rather than diet. Sr andF are mainly stored in bones and the heavier skeleton weight of the

Fig. 10. Two horizontal lines on the femoral diaphysis of the individual S7 thatevidence stress event or metabolic disequilibrium.

males (Warren and Maples, 1997) could explain the lower F and Srvalues. Such differences (for F) have already been observed ina modern population (Arnala et al., 1985). Furthermore, pregnancyand breastfeeding can cause ORSr variations (Kostial et al., 1969;Blanusa et al., 1970; Blakely, 1989). No sex related differenceswere observed in the Li content of the bones.

Among the females, the skeletons S1 and S7 display particularlyinteresting patterns. Bones from individual S1 yield the highest Sr/Ca ratios (Fig. 9) and the lowest d13C value (Table 4). A temporo-cultural or individual dietary preference is inferred fromS1, dated tothe more recent Mixtlan phase. Its carbon isotope signature(�3.2&) suggests a moderate addition of C3 plant material in thediet compared to the other individuals (�1.6& in average), whorelied almost exclusively on C4 plants when considering a meanvalue of �13& for C4 plants and an isotopic spacing between boneand diet of around 12e11& (Hare et al., 1991; Howland et al., 2003).A more important consumption of the common bean could havelowered S1 bone d13C value compared to the other individuals. Thisplant is known to have been present for at least 2300 years atTehuacan (Kaplan and Lynch, 1999). The more importantconsumption of C3-eating animals, as suggested by the numerousfaunal remains, worked bone industry and obsidian projectilepoints in the Mixtlan archaeological levels (Darras and Faugère,2000, 2001, 2010), would also decrease bone d13C value. However,as S1 is the only skeleton available for this period, the evolution ofdietary habits from the Chupicuaro to Mixtlan times cannot beconclusively determined. The S7 skeleton is also characterized byhigh F, Sr and Li content (Figs. 8 and 9, Table 4), with a similar boned13C to that of the other individuals. Since diagenesis does not seemto have significantly affected this skeleton, it is difficult to explainthis pattern. Lines of arrested growth were detected on the tibias(Fig. 10). Episodic consumption of hydrothermal products mayaccount for this signature, assuming there is an association betweenthe growth arrest lines and the bone elemental content. However,bones record the average of the overall dietary signal during the lastyears of life, “diluting” individual dietary episodes. Furthermore, theChupicuarowere constantly exposed to hydrothermal products andif they included these products in their diet, it was very likely ona daily basis. If this is the case, the lines of arrested growth wouldtherefore be independent of the bone elemental content of thisskeleton, whose composition remains enigmatic.

5. Conclusion

Archaeochemistry is useful to recognize human practices andway of life of past populations. However, before exploring suchinterests, post-mortem chemical changes must be estimated. Bonemineralogical and structural analyses (CI, Ca/P, OM content,secondary calcite and histology) are essential for understandingpost-mortem changes, although these proxies alone, are not suffi-cient to evaluate a skeleton’s state of geochemical preservation. Allof the individuals analysed in this study were affected by miner-alogical and structural post-mortem changes which vary at the intraand inter-skeletal scales. All of the skeletons were also affected bygeochemical post-mortem modifications, but a widespread disso-lution-recrystallisation process seems unlikely in this context. Athorough study of the diagenetic skeletal histories of the hydro-thermal tracers, F, Sr and Li (whose retention in living tissues isdetermined by the amount ingested) shows that the progressiveintra-skeletal post-mortem Sr and F enrichment is probably due todiffusion-adsorption. We assume that some skeletal parts arebetter preserved than others. Therefore, the lowest F and Sr intra-skeletal concentrations, best represent the biological values. Incontrast, it is unlikely that the high Li concentrations found are dueto diagenetic addition.

A.-F. Maurer et al. / Journal of Archaeological Science 38 (2011) 1784e17971796

This intra-skeletal study is useful to reconstruct the individualhistory, ante and post-mortem of individuals. Furthermore, thecomparison of values extracted from the skeletons with theelemental content of hydrothermal waters and carbonates availablein the Chupicuaro environment, demonstrates that the Chupicuarohuman skeletons record a hydrothermal signal very likely to be dueto the consumption of some of these products. However, it isimpossible to know whether this “hydrothermal diet” wasconsumed directly or indirectly and if this diet was a result ofcultural habits or environmental resource availability.

Acknowledgements

This work was funded by the ACI 67053 and APN 2000 (SHS-CNRS). The Chupicuaro Archaeological Project was supported bythe French Ministry of Foreign and European Affairs (MAEE), theCentre d’études mexicaines et centraméricaines (CEMCA) inMexico, and the Centre National de la Recherche Scientifique(CNRS). We wish to thank the Institut de Recherche pour leDéveloppement (IRD, Bondy Paris) for the use of their facilities.Many thanks are also due to N. Labourdette (Paris VI), F. Delbes(Paris VI) and F. Lecornec (IRD Bondy) for their technical assistanceand to J-F. Saliège (Paris VI) and Miranda Jans (VU Amsterdam) forthe fruitful discussions about thermal pre-treatment and histology,respectively. A. Zazzo (Muséum National d’Histoire Naturelle,MNHN, Paris), E. Nunn (University of Mainz) and M. Elliott (MAE,Nanterre) are also greatly acknowledged for their constructivecomments. Lastly, we thank an anonymous reviewer for sugges-tions for improvement of the manuscript.

Appendix A

Supplementary data related to this article can be found online atdoi:10.1016/j.jas.2011.03.008.

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