An almost complete skull of a proboscide was found in Tlaxcala in 2014. The remains were identified by Dra. Maria Teresa Alberdi as a gomphothere (Cuvieronius sp.) based on the morphology of the dentition and the spiral-shaped band of tusks enamel.

The cortical bone, better preserved than the trabecular bone, showed heterogeneity in color, density and porosity. Several analyses have been carried out to understand the bone conservation and the postmortem bone mineral transformations (diagenesis and possible fossilization processes) with the aim to propose better preservation methodologies for the cranium. This remain is now located in the Coordinacion Nacional de Conservacion del Patrimonio Cultural for its restoration and then it will be at display at the Museo Regional de Tlaxcala.

Pleistocene megafauna remains are quite frequent in Mexico (Ferrusquia-Villafranca et al., 2010). The primary interest in paleontological remains has been taxonomic (e.g. Garcia-Zepeda and Garduno-Monroy, 2006; Robles-Camacho et al., 2010), however, fossils were often recollected without following a systematic procedure, in which stratigraphic context and sedimentology were overlooked. The relationship between initial peopling and megafauna in the Mexican territory was also documented (Cruz-y-Cruz et al., 2015; Arroyo- Cabrales et al., 2006; Vinas-Vallverdu et al., 2015). More recently, the isotopic signatures and stratigraphy of these paleontological remains have been used as paleoenvironmental records (Pate et al., 1989; Wings, 2004; Robles-Camacho et al., 2010; Perez-Crespo et al., 2012; Cruz-y- Cruz et al., 2016; Perez-Crespo et al., 2016).

The in vivo bone structure is composed of an organic and inorganic fraction forming a complex material that provides flexibility and strength to the skeleton. The organic part is formed mainly by collagen. The inorganic, or mineralized, fraction is composed of microcrystalline bioapatite, always very similar in structure to hydroxylapatite but differing in composition by the substitution of carbonate for PO₄ ^3- (biopatite in fresh bone can contain 7% wt. CO₃ ^2-), the presence of minor elements into lattice, and water incorporation (Wopenka and Pasteris, 2005).

During early diagenetic processes, the soft tissues are removed, collagen is degraded and mineralization, including chemical changes like fluor (F) enrichment and structural changes (e.g. cell parameters) transform bioapatite into a more thermodynamically stable phase. Apatite can incorporate a big number of chemical elements and the stoichiometric and perfect hexagonal structure is only possible for the structure of fluorapatite (Sponheimer and Lee-Thorp, 1999; Hedges, 2002; Jans, 2008).

Apatite is a mineral group with complex solid solutions. In order to simplify, the International Mineralogical Association (IMA) accepted minerals considering three end members: hydroxylapatite, fluorapatite and chlorapatite (Chang et al., 1996; Back, 2014). The different size of monovalent anions (F^-, Cl^-, OH^-) determines a variation in cell parameters. The informal names francolite and dahllite have been applied for apatites enriched in CO₂ and fluorine, and apatites enriched only in CO2, respectively (Deer et al., 2013).

The study of fossil diagenetic processes from neoformed and precipitated minerals in the vascular cavities of the bone and in the fractures allows to stablish the physical and chemical conditions that existed in the lithosphere during lithification, permineralization, and other fossilization processes (Casal et al., 2017). Thus, it is important to evaluate the conservation state, post mortem alteration and approximately estimate the impact of the transformations in its geochemical and isotopic signatures.

On May 2014, workers of the Tepeticpac Archaeological Project (TAP) of the Instituto Nacional de Antropologia e Historia (INAH), accidentally uncovered around 10 centimeters of the distal section of a gomphothere tusk (Gomphotheriidae family). Afterwards, controlled excavations were undertaken using an archaeological exploration methodology in order to provide adequate stratigraphic information of local deposits. An area of four square meters was excavated using a horizontal grid made of four 1 x 1 m quadrants. Vertical depth was registered using a level bank placed over the deposits in the UTM coordinates: 0579830, 2138859, at an elevation of 2,298 meters above sea level (Figure 1). If we project the finding, coordinates in the interactive geological map published by Ferrari et al. (2018) we can verify that the location where the gomphotere cranium was found correspond to the unit described as: Volcano-Sedimentary and Lacustrine Deposits (Late Miocene to Pleistocene).

Figure 1 Figure 1 Location Site.

The discovery took place on an alluvial deposit exposed by a crosscutting road on the hillslopes of Tenextepetl hill, which is part of a mountain chain system that elevates around 300 meters above the Puebla-Tlaxcala valley and that separates the bottom of the valley to the south from the elevated Tlaxcala Block to the north.

The uppermost stratum was formed of 20 to 30 centimeters of loosened tepetate rock that had a steep north to south inclination, which matches the present day-slope surface. It contained no paleontological remains. Below the upper stratum lays Layer A, a colluvial stratum consisting of blocks of medium size blocks (10 to 20 cm) of calcareous rocks and fragments of tepetate rock (fragipan–indurated tephra material). The deposit also had a north to south slope and a thickness of around 1 meter; it was strongly compacted, although some sand lenticular deposits were observed on the north and west profiles.

Below Layer A, on the north portion of the excavation, we detected Layer B, a deposit formed by the superimposition of several thin layers of fine to coarse rounded sands, some of them forming diagonal bands. These seemed to be part of alluvial or lacustrine deposits. In some sections, the sands were interspersed with thin layers of carbonates. The sand layers had grayish tones (10YR 6/1) and covered more than half of the excavation.

Layer C was the bottommost sediment. It is an alluvial stratum made up of medium to coarse rounded sands, probably corresponding to old lacustrine sequence, and very similar in composition to Layer B, but without carbonates. This layer extended throughout the excavation and had a very gentle inclination from north to south, discordant with the steep present day slope. It included dispersed concretions and small intrusions of Layer A materials, mainly calcareous rocks and medium size (10 to 20 cm) blocks of tepetate rock, particularly near the east profile of the exploration.

Correlating the studied section with the complete Pliocene- Quaternary paleosol-sedimentary sequence of Tlaxcala block (Sedov et al., 2009) we relate the B and C layers with the uppermost part of the uplifted Pliocene lacustrine sedimentary unit. Colluvial Layer A incorporates materials of the calcareous Pliocene lake deposits as well as fragments of Pleistocene tepetate. Six to eight layers of tepetate (indurated tephra) interlayered with paleosols formed during the middle and late Pleistocene constitute a semi-continuous mantle over the Tlaxcala Block (Heine and Schonhals, 1973; Sedov et al., 2009). As far as colluvial Layer A includes redeposited materials derived from both Pliocene (lacustrine) and Pleistocene (tephra-paleosol) units, we suppose that its development post-dates these units. We associate this colluviation with the phase of high geomorphic activity and intensification of surface erosion/sedimentation processes which occurred in the Central Mexican Highlands in the Terminal Pleistocene (late Glacial) (Heine, 1984; Sedov et al. 2009). This chronostratigraphic interpretation supposes a major time gap (hiatus) between formation of the Layer A and layers B and C.

Excavation of the skeletal remains was extremely difficult due to the compactness of the stratum, and by intrusive remnants of Layer A. At around 50 to 60 centimeters of excavation, we uncovered the top portion of the gomphothere cranium, which was comprised of the parietal, frontal and zygomatic bones, along with its intact right tusk and a partially looted left one. The cranium had its frontal bones upwards, and had an east-west orientation with its tusks pointing towards the west. Part of the front portion of the skull had collapsed due to the weight exerted by the upper layers. The tusks, maxillae and teeth were well-preserved. The rest of the cranium bones, especially the frontal bone, were fragile and fragmented due to compression because medium sized rocks that laid on top heavily smashed them. Some sections of the bones were absent such as portions of the zygomatic. Also, parts of the cranium bones were partially covered with a fine layer of clayish material of Layers A and B. It appears that sediments of Layer A incorporated and sealed the upper half of the fossil, while the lower section intruded into the underlying B and C layers (Figure 2).

Figure 2 Figure 2 North and east profiles with a top view of the cranium.

Three samples of bone where used for thin sections: a) Cortical bone and trabecular bone with crystalline minerals in the walls of the trabecula partially filled; b) Trabecular bone totally filled with sediments; c) Fractured trabecular bone cemented with the sediments of the filling (Figure 3). A total of six thin sections, two per sample were obtained, however, only one uncovered thin section from sample 2 was analyzed in SEM-EDS.

Figure 3 Figure 3 a) Cortical bone and trabecular bone with crystalline minerals in the walls of the trabecula partially filled. b) Trabecular bone totally filled with sediments. c) Fractured trabecular bone cemented with the sediments of the filling. We obtained two thin sections of each sample.

Samples of detached cortical bone, that was better conserved than the trabecular, were classified in three categories according to their anatomical position, color and porosity: 1- frontal bone with a white yellowish color in the anterior face, the internal side of the cortical bone has a brownish color, they both have low porosity and diagenetic alterations such as pitting and delamination; the pittings and surfaces losses were filled with a white material; 2- temporal bone with a brownish yellow color with more porosity and irregularities in the superficial area of the bone, the pores are filled with white and opaque mineral and with a translucent mineral. The internal face is black; 3- alveolus, the bone has a darker color and has more pores and other cavities which are filled with a white mineral. The internal face is black (Figure 4).

Figure 4 Figure 4 Cortical bone used for DRX, the sample sizes are no more than 3 cm long. 1) Less porous samples, 2) samples with medium porosity, 3) samples with the most porosity.

A detailed mineralogical study, using Powder X-ray Diffraction (PXRD) and Rietveld refinement, allow to characterized the differences between the tree categories to understand the permineralization of the bones and to identify the presence of diagenetic processes. For comparison purposes (Figure 5), we also use a pure inorganic well crystalized apatite (Cerro de Mercado, Durango fluorapatite), an actual bone of low crystallinity (cow bone composed by hydroxilapatite). We also measure with PDRX three samples associated with the sediments that covers the skeleton.

Figure 5 Figure 5 X-ray patterns of the Gomphoterium analyzed samples (blue) with intermediate crystallinity and the pure inorganic (Cerro de Mercado fluorapatite) and pure organic (cow bone hydroxilapatite) end members.

Scanning Electron Microscopy along with an Energy Dispersive X-ray Spectroscopy (SEM - EDS) were used to relate the micromorphology of the bones and the chemical variability of the samples including some diagenetic features, such as tunneling, in the bone fragments surface (Hubert et al., 1996; Reiche et al., 2003; Guido et al., 2012).

Thin section of an uncovered sample 2 along with a non-treated sample in a transversal section of cortical bone were considered for this analysis. A JEOL JSM6060LV with an INCAEnergy 250 EDSLKIE250 were used, under the following parameters: 20 kv, vacuum, back scattered signal, and diverse magnifications.

The transformation in the bone minerals has proceeded in such manner that the paleohistological structure is well preserved. Hubert et al. (1996) proposed that when collagen is removed from the structure the bone does not turn into a disorganized mass but the apatite grows on crystal seed preserving the orientation of original crystals and thus preserving micron scale features. Apatite growth stops when the pores and surface of the bone is sealed, in this case, with the calcite layers and clay coatings, and calcite and detrital minerals infillings. The biogenic hydroxylapatite reacted easily with soils and sediments because of its small size and large surface area.

Despite the well-preserved microstructure of the bone, changes in the mineral lattice has been noted on the XRD analysis: the mineralized bone has changed from bioapatite (carbonated form of a non-stoichiometric hydroxylapatite) to carbonate-apatite (e.g. card ICDD 01-073-9696) and exhibits higher “crystallinity index”, -smaller a cell parameter than the bioapatite phase characteristic of bones in vivo-. Person et al. (1995) stablished that the increase in crystallinity in fossil bones is more related to taphonomic conditions than to the geological age, and that the same variations of crystallinity are found in archaeological and in paleontological bones, which suggests that the increase in crystallinity occurs in the earliest stages of diagenesis.

This mineral transformation can be described as a dissolution-recrystallization process related with decrease in the hydroxyl-ion content (Newesly, 1989). The transformation that preserves the external appearance of the bone was described as a stepwise (isomorphous exchange) non equilibrium transition (Lucas and Prevot, 1991) which is more efficient in porous bone like the one observed in our sample 3. Smaller a cell values in sample 3 were related with further transformation of biogenic hydroxylapatite. The end member of this transformation in a complete fossilization process would be an almost pure fluorapatite as observed in Figure 11 -taken from ICSD (Inorganic Crystal Structure Database) and ICDD (International Center for Diffraction Data) XRD database for a comparative purpose-.

This reduction of the . lattice parameter can be observed (Figure 11) for all the analyzed samples by comparison with power diffraction files (ICSD and ICDD) of pure hydroxylapatite.

Apatite has a hexagonal (P63/m) structure (a= 9.4–9.6, c= 6.8 to 6.9 A) with two metal cation sites, a tetrahedral (T) site (PO₄ ^3-) and an anion column where it is possible to find OH^-, F^- and Cl^-. The carbonate ion can substitute two places in the mineral structure: it can be found at anion column or can substitute phosphate ion at the T site (Hughes and Rakovan, 2015; Wopenka and Pasteris, 2005).

The carbonate in the apatite lattice structure could proceed from neoformed calcite -the main filling mineral in the vascular system of bones and in the voids of the trabecular bone. The calcitic permineralization of bone by micritic precipitation is relatively frequent and it is an usual phenomenon in the fossil record (Fernandez Lopez, 1999); field observations in archaeological sites show that bones are generally preserved when they co-exist with calcite in the sediments (Berna et al., 2004).

The carbonate-apatite mineral phase in the skull bone has a negative correlation between “crystallinity index” and CO₂ amount and a cell parameter (Figures 12 and 13). Carbonate has the effect of making the apatite less crystalline (e.g. Nemliher et al., 2004). In the studied samples it can be prove this as an indirect relationship between CO₂ content and “crystallinity index” (Figure 13). Sample 3 is enriched in CO₂ and because of that it is less crystalline and has a smaller crystallite size value. Carbonate in apatite constrains the crystallinity to nanometric scale and gomfotherium fragments have quite big size values of crystallite. Therefore, we suggested a partial isomorphous exchange of carbonate by fluorine in apatite as a result of the diagenetic processes.

Higher crystallinity and stability in the samples with low content of CO₃ ^2-, is related to the fact that Ca^–PO₄ ^3- bond is stronger than the Ca^–CO₃ ^2- bond, determining higher solubility of carbonate-containing apatite compared to carbonate-free apatite, thus making the carbonated apatite more susceptible to acid dissolution (Wopenka and Pasteris, 2005).

Bone paleohistological structure is well preserved; however, the cortical bone is more preserved that the cancellous bone because the clays that fill the trabecula macroporosity generated internal stresses that fragmented the bone walls.

Nevertheless, the bone composition changed; there is no collagen and there is exogenous C; the mineral part of the bone also was transformed from carbonate non-stoichiometric hydroxylapatite to more crystalline apatite- probably fluorine-carbonate apatite. These changes imply a more stable and less soluble mineral than that of the original bone.

The exogenous carbon detected with isotopical analysis and the carbonate phase in the carbonate-apatite could proceed from the calcite, the main filling mineral. The permineralization of bone by calcite is frequent and enhance the bone preservation.

Biological activity was detected by dissolution and structure modification of the bone surface and the transformation of micrite to needle fiber calcite.

With the calcite infilling we also observed clays, pure smectite and kaolinite, which could be the first minerals intruding the bone macroporosity and deposited in the bone surface. Furthermore, we found sand filling composed of quarts, feldspars, amphiboles and volcanic glass, with a clear different origin if compared with the clays and the calcite.