New data on food consumption in Pre-hispanic populations from Northwest Argentina (ca. 1000-1550 A.D.): the contribution of carbon and nitrogen isotopic composition of human bones.
The analyses of carbon and nitrogen stable isotopes were introduced to archaeology in the middle 1970s and have been used worldwide to assess human and animal diets of archaeological populations [1-5]. Their main potential is that they allow direct access to the average diet of an individual's life time before death which for bone samples is expected to reflect the last 7 to 10 years; while for hair samples, the value is expected to reflect a shorter time span [6, pages 137-138], complementing or broadening the interpretations made from traditional archaeological data, such as plant macro- and microremains, faunal remains, artifacts for food processing, or the osteological analysis of nutritional pathologies .
In the Andean area, the analyses of carbon and nitrogen isotopes have been used to assess the political implications of food consumption and distribution [8-10], the mobility and subsistence models of pre-Hispanic societies [11-13] or, the study of gendered food consumption in domestic contexts .
Following this line, we intend to approach the food consumption profiles of individuals from the archaeological sites of Tolomboon (Calchaqui Valley, Salta) and Esquina de Huajra (Quebrada de Humahuaca, Jujuy) located in NWA (Figure 1). The occupation of these sites encompasses a time span characterized by rapid social changes including a period of hostile conflict between communities and the annexation of the area to the Inca Empire, which probably affected these communities lifestyles--including which food was consumed. The Inca Empire (or Tawantinsuyu in Quechua) was acknowledged by considering maize as a staple, not only for daily consumption but also for chicha making, a traditional alcoholic beverage made from fermented maize which was consumed in feasts and celebrations [15-17]. Nevertheless, as recent isotopic studies demonstrate, maize was a staple in the Andes well before the Inca domination [18-21]but the increase of maize production seen in certain areas can be related to the empire's strategy of negotiation with local polities. In this regard, maize overproduction could have been used to support Tawantinsuyus populations, including the production of chicha for daily use and for sharing in festivities sponsored by the state .
What these studies suggest is that subsistence and diet were a complex matter that surely involved the interlocking of cultural predispositions on what was right to eat, the ecological settings, and the economic and political contexts that changed through time, especially as societies underwent rapid alterations in their social lives. In this regard, we intend to evaluate as many lines of evidence as possible to assess food consumption in two nuclear archaeological areas of NWA from ca. 1000 to 1550 A.D., including the results of recently performed analyses of carbon and nitrogen isotopic composition of human bone. Although we recognize that the sample is small in relation to the time span considered, these are the first results for theses core areas of archaeological development in NWA. In addition, we will consider information from vegetable macro- and microremains, written records, isotopic analysis on human and animal bones, and isotopic composition of edible plants from the area, including those published by other scholars as well as those recently performed in the context of our research agenda. As Tykot  mentions, where the consumed food might have included C4 or CAM plants other than maize, as in our case, it is important to test archaeological faunal remains and other sources of information such as ethnohistoric records to interpret the isotope results.
The theoretical and methodological background for carbon and nitrogen isotope studies in ecology and archaeology in particular have been revised at length in previous literature [19-23]. For the purpose of the present study, it is worth noting that plants that form the basis of the trophic network take up carbon from the atmosphere and fix it in their tissue through photosynthesis, following three different metabolic ways: [C.sub.3] (calvin), [C.sub.4] (hatch-slack) and CAM (crassulacean acid metabolism). [C.sub.3] plants take less atmospheric [sup.13]C[O.sub.2] and thus present values of [sup.13]C that range from -21[per thousand] to -26[per thousand]. Instead, [C.sub.4] plants take up more atmospheric [sup.13]C[O.sub.2] and their tissues are rich in this element, thus giving [sup.13]C values between -7[per thousand] and -12[per thousand] . CAM plants switch between [C.sub.3] and [C.sub.4] depending on their actual location and environmental circumstances , their values ranging from -10[per thousand] to -14[per thousand] if they grow by night following a CAM cycle or from -24[per thousand] to -30[per thousand] if they grow via a [C.sub.3] pathway . Generally speaking, it is assumed that an individual living on [C.sub.3] resources will show a [[delta].sup.13] C value of -21.5[per thousand] on collagen, while an individual living on [C.sub.4] resources will show a [[delta].sup.13] C value of -7.5[per thousand] on collagen. Isotopic fractionation (i.e., changes in the isotopic ratio due to biosynthetic chemical reactions) between diet and values reported in tissues are estimated in the order of 5[per thousand] for bone collagen and from 9.5[per thousand] to 12[per thousand] for bone apatite .
Moreover, research suggests that bone collagen mainly derives from diet proteins and that bone apatite reflects all macronutrients included in the diets (i.e., lipids, carbohydrates, and proteins not used in the animal's own protein synthesis) [25, 26]. Thus, when protein and energy (i.e., carbohydrates and lipids) in an individual's diet derive from resources with different isotopic proportions, collagen may not signal the total isotopic composition of the diet but only its protein portion. Some researchers suggested that the whole diet may be inferred from the apatite bone isotopic composition or from the offset between [[delta].sup.13] C on collagen and [[delta].sup.13] C on apatite (i.e., [[DELTA].sup.13] [C.sub.CO-AP])[19, 20]. However, Kellner and Schoeninger  have recently suggested that most single measures of [[delta].sup.13]C either for collagen or apatite fall short of expectations for diet reconstruction. They suggest that the value of [[DELTA].sup.13][C.sub.CO-AP] is not specific to any combination of protein or to the whole diet. Instead, by plotting [[delta].sup.13][C.sub.CO] and [[delta].sup.13][C.sub.AP] values from organisms with different known diets, they presented a model of three regression lines which provides a way of distinguishing between diets with [C.sub.3] protein and those with C4 protein. Thus, according to the authors, the increase in [[delta].sup.13][C.sub.AP] in the three regression lines largely represents the increase of [[delta].sup.13]C values of diet energy (i.e., carbohydrate and lipid except in diets with excessively high protein levels). Taking this into account, we will plot data from [[delta].sup.13]C on collagen and apatite to assess the possible source of protein included in the diet of the individuals analyzed.
The atmosphere is nitrogen's biggest reservoir, and its stable isotope ([sup.15]N) is present as a gas ([N.sub.2]). Plants capture nitrogen in two different ways. Some of them do it through special nitrogen fixing bacteria attached to their roots that convert [N.sub.2] into forms that can be used by the plant, leading to little fractionation, which means that these plants have N values similar to that of the atmosphere (0[per thousand]). Other plants, known as nonfixing, capture N through the decomposition of organic matter present in their habitats. Generally, these plants possess significantly more positive values. The median value for fixing plants is +1[per thousand], with a typical range of-2[per thousand] to +2[per thousand]; whilst in the case of the nonfixing, it is +3[per thousand], with a typical range of 0[per thousand] to +6[per thousand] [2, 24].
It has been noted that organisms increase their [sup.15]N content in 3 or 4[per thousand] along the trophic chain, both for terrestrial and marine resources. For example, terrestrial mammals have [sup.15]N mean values of about 5.7[per thousand] while marine mammals have mean values of 15.6[per thousand] . Nitrogen has proved to be important for investigating topics like weaning or differentiating consumption of marine resources , and, unlike carbon, all nitrogen present in bone collagen derives from dietary proteins, representing trophic-level effects in protein intake associated with organisms' positions in food webs [21, 22]. However, several studies have also discussed how geographical and ecological factors such as aridity, humidity, cold, and latitude affect the values of [sup.15]N recorded in different tissues. Particularly, a negative relation has been observed between water availability and the values obtained from the collagen of herbivore bones . Research performed by Amundson et al.  noted that in drier ecosystems there seems to be a greater loss of nitrogen via lixiviation and its transformation (nitrification, denitrification, and volatility of ammonia) which would lead to the enrichment of [sup.15]N in the remaining nitrogen in the system. This fact implies that human tissues with enriched [[delta].sup.15]N could not reflect the consumption of animal protein, but the consumption of plants with enriched values .
2. Materials and Methods
2.1. The Study Area. The analyzed material came from the geographical area located in the northwestern corner of Argentina (Figure 1), specifically from two of its main valleys: Calchaqul Valley in Salta Province and Quebrada de Humahuaca in Jujuy Province. The former is characterized by a semiarid high altitude climate, as winds are depleted in humidity since they cross the mountain range. Mean annual precipitation is about 200 mm in Cafayate and rains are markedly distributed, as they occur more often in the western slopes of the valley during the summer months (December to March) which determines a lower water use due to evapotranspiration . However, there exists a relatively prolonged period free of frosts which is beneficial for agricultural production .
The Quebrada de Humahuaca is a deep narrow valley or ravine extending along 120 km and presenting two different climates, semi-arid and arid, according to the variation of precipitations . Mountainous relief influences the total annual precipitations, causing the geographical distribution of rains to be markedly irregular. For example, in the southern sector of the area, mean precipitation values of 391 mm and 199 mm were recorded for Volcaon and Tumbaya, respectively, While in Abra Pampa (northern sector) the mean value recorded is 282 mm . Although rains are torrential and are concentrated from November to March, they do not compensate for the marked water stress present in the area .
In a general sense, NWA is a geographical region defined by the presence of the Andean mountain range, running in a NS direction, as well as several intermountain valleys, quebradas, and a high altitude area (more than 3,500 masl) called puna that sustained the development of interconnected societies along its temporal occupation. The most important valleys and quebradas from North to South are Quebrada de Humahuaca, Quebrada del Toro, Calchaqul Valley, Santa Maria or Yokavil Valley, Capn Valley, and Hualfln Valley (Figure 1). The human occupation in the puna areas was concentrated in certain oasis such as Antofagasta de la Sierra in Catamarca Province, Huachichocana, Inca Cueva and Rinconada in Jujuy Province, and Pastos Grandes in Salta Province.
Although NWA has been occupied for more than 10.000 years B.C., when the first mobile hunter-gatherers established in the area, we are interested in the four to five centuries before the Spanish conquest, a time span comprising important economic, political, demographic, and climatic changes that radically shaped the trajectories of these communities. From about 900 A.D., in some places of NWA, communities started to cluster in nucleated settlements, many of them located on landforms with difficult access, such as the slopes or tops of hills and small plateaus. Agricultural intensification played a key role in the development of these societies, which cleared vast extensions of land and built agricultural structures (e.g., irrigation systems, terraces) in Calchaqul Valley [32, 33], Quebrada de Humahuaca , and Antofagasta de la Sierra . Different ceramic styles emerged within specific geographical areas like Santamariano style in Calchaqul and Yocavil Valleys and Beien style in Hualfln and Abaucan Valleys  Figure 2).
While the traditional view of these societies posited that they were organized into chiefdoms that hierarchically ranked settlements levels [37-39], other scholars suggest that communities from the Regional Development Period (RDP, ca. 900-1450 A.D.) were organized into corporative or communal structures with flattened or no social hierarchies at all [40-43]. In this sense, regionalization of ceramic styles and material culture in general points to the existence of political fragmentation, which can also be seen in the replacement of previous dispersed hamlets for key nucleated settlements located in defensive locations. This scenario possibly resulted from the presence of interregional conflict, evidenced in the appearance of war related paraphernalia (e.g., weapons, rock art scenes, and defensive architecture) [44, 45]and violent trauma on human bones .
Staple goods in communities from NWA temperate valleys included maize (Zea mays), potatoes (Solanum spp.), grains such as quinoa (Chenopodium quinoa) and kiwicha (Amaranthus caudatus), squashes (Cucurbita spp.), peppers such as ajx (Capsicum baccatum), salt, and beans (Phaseolus spp.). Outstandingly, apart from maize, kiwicha (A. caudatus) is the only [C.sub.4] plant in the western hemisphere consumed by Precolumbian human populations. Besides, they relied on seasonal resources such as algarroba (Prosopis nigra or Prosopis alba) and chahar (Geoffroea decorticans) and meat from camelids and micromammals, among others [47-53].
2.2. The Archaeological Sites. Tolombon is a complex archaeological settlement located in Salta Province near the modern town of Cafayate (Figure 1). Although mentioned in the archaeological literature as early as 1894 by Ten Kate, it was later studied by different archaeological research teams . The settlement consists of architectural remains built on the slopes of a hill, a pattern shared with other sites from the Yocavil Valley (e.g., Rincon Chico, Cerro Pintado, Fuerte Quemado, and Quilmes). Research conducted by Dr. Williams [54,55] revealed the distribution of the architectural remains in the hill, covering 35 ha, in the following way: (1) a fort located at the top of the hill, a residential area on the slope of the hill, and possibly a public sector; (2) a residential sector located at the base of the hill and another one on the southeastern side of the hill spatially segregated and with agricultural structures between them; (3) more than 17 tombs located on the southeastern side of the hill; (4) staggered structures on the southern side of the hill, over the residential area (Figure 3).
Radiocarbon dating allows ascribing the occupation of the site to the years 1200 to 1600 A.D. (Table 1). The excavation of one of the residential enclosures located in the base sector (Building 6) yielded a number of cultural materials, including metal artifacts and debris, ceramic remains, lithic instruments and by-products, vegetables and faunal remains, and a hearth in the centre of the enclosure. The analysis of these materials allowed for the interpretation of this enclosure as a domestic space where quotidian activities were carried out . Local ceramic styles are abundant in this enclosure (i.e., Santamariano, Famabalasto negro grabado, and Belen styles), while foreign material is scarce, including only a minor percentage of Incaic pottery style. Many undecorated fragments show fire marks, possibly related to their use as cooking vessels . The vegetable macroremains found consistedoffive different racesorvariantsofmaize (Colorado, Chaucha, Socorro, Amarillo Chico, and Pisingallo); several caryopses were burnt, suggesting that they had probably been cooked (Table 2).
Faunal remains found in this enclosure were divided into three components. The first one (stratigraphic levels 7 to 10) is mainly composed of camelids (Lama sp., Lama glama and Lama vicuna), although fox (Lycalopex sp.), Chinchillidae, and bird bones were identified. The second component (stratigraphic levels 3 to 6) presents a variety of resources, including fox (Lycalopex sp.), armadillo (Dasypodidae), viscacha (Chinchillidae), weasels (Mustelidae), ostrich egg shells, and camelids (Lama sp.; Lama glama and Lama vicuna). In the third component (stratigraphic levels 1 and 2) the only osseous remains found were from camelids (Lama sp.). In general, faunal remains from this enclosure show the predominant consumption of camelids with a mortality profile of young individuals in two of the three defined components. Additionally, remains of an Andean deer (Hippocamelus antisensis, commonly known as taruca) were found. Anthropic modifications, like cut marks or thermal alterations, were recorded, suggesting the possible performance of human activities such as slaughter and cooking .
During one of the field seasons in Tolombon, a burial place which had been disturbed was discovered in one of the ravines leading to the fort. The tomb was located under a big rock which stood on the sides of a ditch forming a kind of natural roof of 2 m wide and 2.6 m deep, 0.9 m tall in one extreme and 0.4 m in the other. Although the human remains were highly fragmented, osteological analysis allowed determining that at least six individuals were buried, including both adults and subadults . From this funerary context, five individuals were sampled for isotopic analysis (see details in Table 3) using bones that could be clearly associated to an individual.
Esquina de Huajra is located 45 km from the modern city of San Salvador de Jujuy in Tumbaya Department, Quebrada de Humahuaca (Figure 1). The site was built on the slopes of a landform elevated 90 m above the valley level which extends close to the alluvial plain of Rio Grande. In 2001, a salvage project was conducted at the site and six tombs were found, along with other archaeological materials . The tombs were located in what was called Terrace III, and five of them were excavated (tombs 1,2,3,4,and 6) (Figure 4). The laboratory analysis of the human bone remains allowed establishing the presence of at least 18 individuals, comprising adults of both sexes and unsexed subadults of different ages at death .
Radiocarbon analyses postulate an occupation span ranging from 1400 A.D. onwards (Table 1). This would imply that the site was occupied (and possibly built) in a latest age of the Inca Period, continuing this occupation in the initial stage of the Early Colonial Period, although the real control of the territory by the Spaniards was not exerted until 1600 A.D. approximately . This is consistent both with the findings of Inca material culture in the tombs and the absence of Spanish objects in the site. The analyses of the materials recovered suggest the performance of different activities in the sectors of the site. In Terrace I, a possible patio was excavated, which contained a hearth with carbon lenses, ceramic and bone fragments, a stone grinder, red pigment, and several ceramic remains. This occupation floor also contained abundant animal remains, two bone instruments, and a metal instrument [61, 62]. The ceramic assemblage contains some of the typical Inca forms present in the provinces of the Empire : footed ollas, small plates, plates and aribalos possibly related to chicha elaboration and consumption  and nonlocal ceramics (i.e., Chicha/Yavi, Inca Pacajes, and Casabindo Pintado). The lithic material obtained points to the performance of reduction activities directed to obtaining base forms . Faunal remains analyzed by Mengoni Gonalons [64, 65] came from this domestic context and correspond to members of the Camelidae family, including llama (Lama glama), vicuna (Lama vicugna), and guanaco (Lama guanicoe). As in Tolombon, remains from Cervidae were found, including a specimen of taruca (Hippocamelus antinensis), as well as rodents (undetermined) and birds (Cairina moschata).
In turn, Terrace II probably functioned as a circulation path. More than 1,000 ceramic fragments were found, half of them undecorated and corresponding to medium and large vessels and ollas. A few fragments were from the body of five small vessels, and also bowl fragments were recovered. Nonlocal pottery fragments corresponded to Chicha/Yavi Bowls and six polished bowls . As far as we know, most of the surface of Terrace III seems to have been used mainly as a funerary space , as six tombs containing the remains of several individuals were found and analyzed. Five of the six detected tombs could be excavated, and six samples of human bone were collected for isotopic analysis, including adult individuals from both sexes and unsexed subadults from different ages as described in Table 3. These bone elements were selected so as to have at least one sample from each tomb.
2.3. The Methods. The isotopic analyses were carried out in different laboratories, implying that, although results are reported relative to the same standard (VPDB for [[delta].sup.13]C and Air for [[delta].sup.15]N) some caution must be taken when comparing and combining data. A first set of human bone samples was sent to the Center for Applied Isotope of the University of Georgia (CAIS) and a second set to Geochron. The bone was cleaned with wire brush and washed using ultrasonic bath. After cleaning, the dried bone was gently crushed into small fragments. The crushed bone was treated with 1N acetic acid to remove any secondary carbonates at room temperature with periodic evacuation of C[O.sub.2] for 24 hours. The residue was filtered and rinsed with deionized water to remove any remains of acetic acid. Then, the sample was dried overnight at 60[degrees]C. The dried sample was evacuated in the flask and treated with diluted HCl at 4[degrees]C to recover C[O.sub.2] from bioapatite. The carbon dioxide was cryogenically purified and collected for stable isotope ratio analysis. The acid solution was collected and left overnight at 4[degrees]C. Then the solution was filtered on fiberglass filter, and precipitate was rinsed with deionized water. The precipitate was boiled in deionized water (pH = 3) for 6 hours to dissolve collagen and leave humic substances in the precipitate. The collagen solution was then filtered and dried out to isolate pure collagen. The dried collagen was combusted at 575[degrees] C in an evacuated/sealed Pyrex ampoule in the presence of CuO. The carbon dioxide and nitrogen were cryogenically separated and purified and collected in the flasks for analyses. The [[delta].sup.13]C sample was measured with respect to PDB, with an error of less than 0.1[per thousand]; the [[delta].sup.15]N sample was measured with respect to atmospheric air nitrogen, with an error of less than 0.2[per thousand]. At CAIS, stable isotope ratio analyses were conducted on Finnigan MAT252 mass spectrometer. The C/N ratio was measured using Perkin-Elmer 2400 CHN analyzer. At Geochron, a VG Micromass gas source stable isotope ratio mass spectrometer was used.
Additionally, modern plants from Calchaqul Valley were isotopically characterized. They consisted of seeds from quinoa, amaranth, squashes, and maize collected from a farm located in Animanao (Salta). No industrial fertilizers were used in order to avoid any potential nitrogen enrichment . Samples were raw and frozen-dried and ground prior to characterization of [[delta].sup.13]C and [[delta].sup.15]N. These samples were sent to the Instituto Nacional de Geologoa Isotoopica of the Universidad de Buenos Aires. The specimens were washed by ultrasonic bath, dried for 24 hours at 60[degrees]C, and crushed into 1mm fragments. Three mg of each sample was weighed in tin capsules and processed in a Carlo Erba EA1108 elemental analyzer coupled to an isotope ratio mass spectrometer of continuous flow interfaces (Thermo Scientific Delta V Advantage ConFlo IV).
Modern plant samples were corrected for the Suess Effect; that is, the reduction by at least 1.5[per thousand] of [[delta].sup.13]C values in the atmosphere as a consequence of the use of fossil fuels during the recent industrial period. It is, therefore, necessary to correct these samples by adding 1.5% at the moment of using them to reconstruct paleodiets and to plot them along archaeological data .
We also developed a database of published data on carbon and nitrogen isotopic composition of archaeological and modern plants, including results from Argentina  and Peruo . Isotopic data from archaeological camelid bones were taken from Mengoni Gonalons [64, 65]and included four samples for Tolombon and five samples for Esquina de Huajra. Only data containing both carbon and nitrogen information were considered, and median values were presented for each type of data (i.e., maize, camelid, etc.). Carbon isotopic results for the modern plant samples were corrected for postindustrial enrichment of atmospheric [sup.12]C, in order to make these data comparable with archaeological llama and human [[delta].sup.13]C results.
The isotopic results for the Tolombon and Esquina de Huajra Human bone samples are summarized in Table 3. The results for analyzed food samples are summarized in Table 4. To compare the values, we first designed a graphic plotting [[delta].sup.13]C and [[delta].sup.15]N values from modern and archaeological plants and fauna against human bone samples for Esquina de Huajra (Figure 5) and Tolombon (Figure 6).
If we first consider the values for [[delta].sup.13][C.sub.col], it is observed that results both for Esquina de Huajra and Tolomboon are comfortably included in the range of C4 plant group (i.e., maize, amaranth). Interestingly, camelid diet from both sites is no far from that of humans, showing only subtle differences. As mentioned above, collagen reflects the protein portion of the diet consumed in the last 5 to 7 years of life. In this regard, the results obtained for both sites show that the ingested proteins came from a [C.sub.4] source mainly. Moreover, data from carbon apatite suggest that energy also came from a [C.sub.4] source, as we shall discuss later.
In the case of nitrogen ([delta][sup.15]N), the value reflects directly the kinds of protein incorporated to the diet, which may be animal, vegetal, leguminous or marine, in relation to the position of an organism in a food web. Assuming a roughly 3[per thousand] enrichment between food source and consumer, the [[delta].sup.15]N results obtained would signal the consumption of organisms with N values between 4[per thousand] and 8[per thousand] of [[delta].sup.15]N in their diets, agreeing with faunal references (camelids only) from both sites. There is an individual from Tolombon (T4C2L1) with a very low value of [[delta].sup.15]N (3.2[per thousand]) that is difficult to interpret and a person from Esquina de Huajra (T1G2MS) that shows the lowest [[delta].sup.15]N value of the sample from this site (6.2[per thousand]), although not as low as the one from Tolombon. Remarkably, values of [[DELTA].sup.15]N for human bone show a higher dispersion than [[delta].sup.13]C values, as nitrogen values for reference food sources do.
We contrasted our data to those provided by Kellner and Schoeninger , who have questioned the utility of the difference between the value of carbon on collagen and apatite ([[DELTA].sup.13][C.sub.CO-AP]) to assess the whole diet of an individual, and have indicated the necessity of considering the photosynthetic path in which protein and energy resources were separately inscribed. What we can see from the following plot (Figure 7) is that both subsets are form discrete clusters, except for one individual from Esquina de Huajra (T1G2MS) which is fully included into Tolombons cluster. Both clusters are located over the 100[per thousand] [C.sub.4] energy line, although Esquina de Huajra's individuals are slightly closer to the [C.sub.3] energy line.
The samples' range in [[delta].sup.13]C encompasses much of the values found in C4 resources available for animal and plant data presented here and in other studies. Interestingly, camelid diet is quite similar to that of humans. Isotopic data from camelid bone from Esquina de Huajra suggest that their diet was quite homogeneous and that they would correspond to locally raised herds or to animals that lived freely in low altitude grasslands . The diet of camelids from Tolombon is more variable, suggesting that they obtained food from different ecological zones. Some of them, especially larger animals like llamas, were raised in lower areas (less than 4000 masl) while smaller animals, like vicunas, might have been brought from the Highlands (close to or over to 4,000 masl) . In spite of these differences, human diets from both sites seem quite homogeneous, at least for carbon values, as they cluster tightly. This would imply that diets from both adults and subadults and both males and females were quite similar; moreover, they came primarily from a [C.sub.4] source, as we can see in Figures 4 and 5.
In contrast, [[delta].sup.15]N values for human bone are more varied, ranging from 3[per thousand] to almost 13[per thousand] for the samples considered, which was also noted for camelid bones from both sites  and, as we can see in this study, modern vegetal resources show higher dispersion in nitrogen values. As we previously discussed, it is possible that in arid or semi-arid environments as the one we are dealing with, plants respond physiologically to water stress by showing higher nitrogen values, which would prevent or at least warn about interpreting results visa-vis protein consumption. In this regard, it is important to highlight that our sample has a tendency towards protein consumption both in carbon and nitrogen values.
Judging from the data presented, the origin of all dietary sources (protein and energy) came mainly from [C.sub.4] resources. It is interesting to consider that some written documents of the Colonial era (after 1540 A.D. approximately) point to a wider diet, quoting Governor of Tucumoan, Don Mercado de Villacorta: "the calchaqutes [are] more supplied than others, as they do not content with only maize, but wheat and barley and legumes and potatoes and quinoa and algarroba ..." . Regarding the consumption of the latter, this resource seems to have been especially important as several written sources mention quarrels over its exploitation. For example, Torreblanca mentions that "... the quilmes and other nations, [in] a year of famine, (...) had no resources and were going to perish if they did not become friends with the pacciocas, who had abundance and were owners of San Carlos, where a large quantity of algarroba existed, they made peace and the enemies' towns were depopulated to pick up the algarroba ...". The archaeological record of the regions points to a similar direction, as maize remains were found in Molinos I, La Paya, and El Churcal as well as gourd, squash, and beans in La Paya and El Churcal [69, 70]. In Valdez site, besides corn grains and cobs, remains of Chenopodium, tubers (Solanum), beans, and peppers were also found . In the northern sector of Calchaqui Valley, D'Altroy and coauthors mention the existence of maize, quinoa, and tubers in both Potrero de Payogasta and Cortaderas Bajo sites. In Los Graneros site, Tarragoo and Gonzoalez  mention the presence of maize, beans, gourd, and algarroba macro remains.
The source of [C.sub.4] energy and protein could have been provided by maize, amaranth, or animals that, in turn, consumed [C.sub.4] plants (as mentioned, camelid isotopic data points to this direction) [64, 65]. The overconsumption of maize in American pre-Hispanic populations has been traditionally linked to the presence of iron deficiency anemia, which, in turn, was linked to the appearance of porotic lesions in the skull (i.e., porotic hyperostosis) and the orbits (i.e., cribra orbitalia) . The absence of porotic lesions in relation to diets with a high component of maize was already noted [73,74]. Osteological markers of nutritional stress in the population of Esquina de Huajra are present in a low percentage (only one male adult showed mild signs of porotic hyperostosis ), and although the preservation of the remains from Tolomboon analyzed here prevents a thorough examination in this respect, the study of a collection of nine skulls from this site carried out by one of the authors  did not show positive results for the presence of porotic lesions in the skull or orbits. This observation may indicate two possibilities: whether the link between lesions traditionally associated with iron deficiency anemia in relation to high maize consumption is incorrect, as recently published studies suggest [76, 77], or populations may have achieved an adequate balance between maize consumption and other resources (e.g., grasses, legumes, tubers, and meat) that possibly counteracted the negative effects of the sole consumption of maize. As one of the reviewers of this paper suggested, the values obtained for [[delta].sup.13]C on apatite and collagen, although signaling the importance of [C.sub.4] plants consumption, leave room for the consumption of other resources as well.
Our results on carbon and nitrogen isotopes for modern amaranth do not allow us to rule out the consumption of this plant, even though no archaeological findings of this cultivar were made in the area of study. Medina et al.  mention that Amaranthaceae pollen remains found in some archaeological sites from Coordoba (Argentinean Central area) may correspond to the species Amaranthus caudatus. Babot  mentions the finding of amaranth remains in Penas Chicas and Punta de la Pena (Antofagasta de la Sierra, Catamarca), El Remate and Cueva de los Corrales (Tucumaon, Argentina), and Los Viscos (Beleon, Catamarca), but, as far as we are concerned, no remains of this plant have been found in Quebrada de Humahuaca nor in Calchaqui Valley. The [[delta].sup.15]N value reported here for this pseudocereal is substantially enriched and similar to the results presented by Turner et al. ; it is also slightly higher than nitrogen values obtained for the human bone samples in our study,r equiring further efforts in isotopically characterizing this cultivar.
This study has provided the first data on paleodiet of small scale complex societies from Northwest Argentina through analyzing the variation of carbon and nitrogen isotopes in a sample of individuals from archaeological sites of temperate valleys. Some advance has been made in starting to construct an isotopic ecology of the area. Although the sample is small, both for human bones and modern plants, the contribution lies in the originality of the results, especially for a nuclear area of archaeological research in NW Argentina.
In this regard, our results serve to posit the importance of [C.sub.4] plants for the diet, which could include maize and amaranth, before the arrival of the Inca Empire to the region, as Tolomboon results show an important component of [C.sub.4] plants in the individuals' diet. Apparently, this importance continued in the Inca period, as results from Esquina de Huajra reveal a high consumption of [C.sub.4] plants. Research in Central Chile reports the first enriched [[delta].sup.13]C on collagen and apatite from 1800 B.P. in Llolleo populations, signaling the importance of maize for these communities . Interestingly, during the Inca period, [[delta].sup.13]C values decrease both in collagen (mean [[delta].sup.13]C-13.1[per thousand]) and apatite (mean [[delta].sup.13]C 5.9[per thousand]), which do not coincide with expectations regarding the archaeological and bioarchaeological record . In a study that compared isotopic information from puna, valleys, and quebradas from NWA, Killian Galvan and Samec  did not find a temporal tendency towards more enriched [[delta].sup.13]C, although this was expected from previous research which emphasized that the consumption of maize would have progressively increased through time. Also, the research conducted by Gil et al.  with samples from Mendoza (Central Western Argentina) has not allowed for the postulation of a clear tendency towards increasing maize consumption in this region of South America.
Different results were reported from Peruvian populations, where maize consumption seems to have been intensified during the Inca domination. Burger et al.  report that, although maize was consumed with different intensity in farmer communities (Waman Wain), in communities with elite members (Jauja), and in Machu Picchu, with retainers serving the Inca nobility (yanaconas), it seems to have been a staple. This information not only supports the politics fostered by the Incas of replacing local cultivars but also the idea of intensification and expansion of agricultural lands under their dominion . In a similar vein, Hastorf and Johannessen  argued that during the Wanka III period (Inca) in Mantaro Valley, stable isotopes results suggest that maize consumption was increased but probably in the form of chicha drinking in connection to feasts sponsored by the state. However, Finucane et al. indicatethatmaize wasa staple well before the Inca Period in Conchopata and even suggest that it might have been the resource that supported one of the first urban development of the Peruvian Sierra and its associated social complexity.
This study, therefore, illustrates the potential of using multiple lines of evidence, despite noted limitations in sample size, for interpreting food consumption in prehistory. In this regard, the archaeological record of the study areas shows a variety of resources not entirely reflected in the isotopic values obtained for the human bone samples. Thus, this implies improving the studies on the isotopic ecology of the areas, considering resources not yet included (e.g., peppers, tubers, algarroba, chanar, and micromammals). Finally, we consider it imperative to develop a model for the contribution of carbon and nitrogen to the protein and nonprotein portions of bones in cases of mixed diets.
Correspondence should be addressed to Maria Soledad Gheggi; firstname.lastname@example.org
Received 29 April 2013; Revised 18 July 2013; Accepted 22 August 2013
Academic Editor: Maryna Steyn
The authors thank Randy Culp and Andrew Cherskinky from the Center for Applied Isotope Studies (CAIS) for their patience and permanent advice on the interpretation of the results. They are grateful to Beatriz Cremonte, who allowed access to the Esquina de Huajra human collection. They thank Veronica Vasvari, Mabel Mamani, and their families for the collection of modern plants for their analyses. The authors acknowledge Violeta Killian Galvaon's bibliography and her kindly given advice and the Instituto de Geocronologia y
Geologia Isotopica (INGEIS) for the performance of the analysis on modern samples. They also thank the anonymous reviewers, whose careful comments helped to strengthen the manuscript. This research was supported by grants from Consejo Nacional de Investigaciones Cientificas y Tecnicas (CONICET) (PIP 5361) and Agencia Nacional de Promotion Cientifica y Tecnologica (ANPCyT) (PICT 1550) awarded to Veronica Williams.
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Maria Soledad Gheggi and Veronica Isabel Williams
CONICET-Instituto de Arqueologta, Facultad de Filosofta yLetras, Universidad de Buenos Aires, 25 d eMayo 217, 3rd floor, C1002ABD Buenos Aires, Argentina
Table 1: Radiocarbon dating for Tolombon and Esquina de Huajra. Site Code Material Chronology B.P. Tolombon GX-29252 Charcoal 720 [+ or -] 60 GX-29251 Charcoal 500 [+ or -] 60 Beta 168672 Charcoal 440 [+ or -] 50 GX-29663 Charcoal 350 [+ or -] 60 Beta 171425 Charcoal 460 [+ or -] 60 Beta 171426 Charcoal 440 [+ or -] 60 Esquina Beta 193319 Charcoal 340 [+ or -] 55 de Huajra Beta 206919 Charcoal 280 [+ or -] 50 AA88375 Charcoal 393 [+ or -] 82 UGA16200 Human bone 550 [+ or -] 40 GX32577 Human bone 450 [+ or -] 60 GX 32576 Human bone 320 [+ or -] 50 Site Code Cal. A.D. Reference 2 sigmas Tolombon GX-29252 1221-1400  GX-29251 1327-1612 Beta 168672 1410-1520 GX-29663 1441-1793 Beta 171425 1400-1515 Beta 171426 1405-1525 Esquina Beta 193319 1455-1796  de Huajra Beta 206919 1496-1952 AA88375 1400-1664 UGA16200 1318-1463 GX32577 1419-1627 GX 32576 1460-1799 Table 2: Vegetable macro remains possibly used as food in Tolombon. Family Species Plant or reproductive Reference structure Poaceae Zea mays Fruit (caryopsis) Fabaceae Prosopis sp. Seeds and fruits (pods)  Fabaceae Prosopis ferox Fruits (pods) Table 3: Carbon and nitrogen results for human bone samples. Lab Code Site/sample code Chronology UGA16200 Esquina de Huajra T1G2MS 550 [+ or -] 40 AP UGA 2087 Esquina de Huajra T1G1MS Geochron 32577 Esquina de Huajra T2 I4 450 [+ or -] 60 AP UGA 2088 Esquina de Huajra T2I1 Geochron 32576 Esquina de Huajra T3I1 320 [+ or -] 50 AP UGA 2089 Esquina de Huajra T4I1 UGA 2090 Esquina de Huajra T6I1 Geochron 32578 Tolombon T4C1L1 UGA 16199 Tolombon T4C2L1 UGA 16201 Tolombon T4C1L1 UGA 2085 Tolombon T4C4L1 UGA 2086 Tolombon T4L5 Lab Code Sex/age Bone UGA16200 Male Adult Left humerus fragment UGA 2087 Male Adult Left humerus fragment Geochron 32577 Indeterminate Left radius fragment 8 [+ or -] 2 years UGA 2088 Indeterminate Right tibia fragment 6 [+ or -] 2 years Geochron 32576 Female 40 [+ or -] 5years Left femur fragment UGA 2089 Indeterminate Right femur fragment 7 [+ or -] 2years UGA 2090 Posibly Female 17-21 years Right tibia fragment Geochron 32578 Indeterminate Indeterminate Left humerus fragment UGA 16199 Indeterminate Indeterminate Left humerus fragment UGA 16201 Indeterminate Indeterminate Left humerus fragment UGA 2085 Indeterminate Indeterminate Right humerus fragment UGA 2086 Indeterminate Indeterminate Left humerus fragment Lab Code [delta] [delta] [delta] [sup.13]C [sup.13]C [sup.15]N [per thousand] [per thousand] [per thousand] Apatite Collagen UGA16200 -4.8 -9.63 6.7 UGA 2087 -6.2 -11.0 12.6 Geochron 32577 -6.5 -11.1 9.7 UGA 2088 -5.2 -11.3 11.7 Geochron 32576 -6 -11.3 12.2 UGA 2089 -6.6 -12.0 11.3 UGA 2090 -6.7 -11.5 12.1 Geochron 32578 -4.0 -9.30 9.0 UGA 16199 -4.8 -9.03 3.2 UGA 16201 -5.0 -9.63 11.1 UGA 2085 -4.7 -9.9 10.1 UGA 2086 -4.9 -9.3 11.7 Lab Code [DELTA] C/N ratio [sup.15]N [C.sub.CO-AP] UGA16200 4.83 2.66 UGA 2087 4.8 2.7 Geochron 32577 4.6 Not performed UGA 2088 6.1 2.68 Geochron 32576 5.3 Not performed UGA 2089 5.4 2.72 UGA 2090 4.8 7.95 Geochron 32578 5.3 Not performed UGA 16199 4.23 2.66 UGA 16201 4.63 2.7 UGA 2085 5.2 3.05 UGA 2086 4.4 2.74 Table 4: [delta][sup.13]C and [delta][sup.15]N for modern plants in the Calchaqularea. Carbon results were corrected for postindustrial enrichment. Code lab Material [delta] [delta] C/Nratio [sup.13]C [sup.15]N 27626 Quinoa -22.7 9 21.9 27627 Gourd -21.5 7.1 11.4 27628 Squash -24.5 6.1 16.7 27629 Sweet gourd -26 8.2 11.3 27630 Red maize -10 4.2 44.9 27631 Kiwicha -10.8 14 18.7
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|Title Annotation:||Research Article|
|Author:||Gheggi, Maria Soledad; Williams, Veronica Isabel|
|Publication:||Journal of Anthropology|
|Date:||Jan 1, 2013|
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