Molecular variation between external and internal part of an archaeological sample TMT

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Botanical and geographical distribution

Botanical sources

The Burseraceae family includes over 20 genera and more than 600 species of trees and scrubs from tropical and subtropical regions from Asia, Africa and America. A characteristic of this family is to have resin products containing aromatic terpens and essential oils. Often called the balsam or torchwood family, Burseraceae are widely distributed in tropical and subtropical regions worldwide and are divided into three subfamilies: Bursereae, Canarieae and Protieae (Langenheim, 2003).
Burseras are related to the genus of frankincence (Boswellia) and myrrh (Commiphora) (figure 2). The genus Bursera comprises around 100 species in America that distribute between the Southwest of the United States and the North of Brasil, but reach their major diversity on the Pacific coast of Mexico (Rzedowski, 2004).
Burseras have been described as “typically low or medium si ze threes with colorfull trunks in blue, green, yellow, red or purple” (Langenheim, 20 03). The bark of some of them exfoliates in papery sheets. This genus is notable because its exudates are rich in terpenoids (Mooney and Emboden, 1968).
In Mexico, Bursera is one of the most important components of dry forest, in some parts of the Southern regions of Mexico the genre becomes dominant or co-dominant. In desert scrub and thorn scrub, they are also present and, occur in a lesser extent in lowland tropical rain forest and higher altitude woodland (Rzedowski, 2005).
Until now 82 species had been recensed by botanists. Nevertheless, its classification remains problematic and its phylogenetic relationships are not well understood (Rzedowski, 1979). Molecular studies (Becerra and Venable, 1999) supported the view that Bursera is monophyletic and more closely related to Commiphora than Boswellia. The figure 3 shows a simplified version of the DNA classification of Mexican Burseras made by Becerra and Venable to show the two sections and the five groups in which they were divided.
Concerning distribution, the southern pacific regions of Mexico concentrate the highest richness in Bursera species. Guerrero registers 48 species while Michoacán and Oaxaca 37. Although it is evident that each species presents different affinities and ecological tolerances, most of Mexican varieties grow in the good drained soil of slopes and mountains, in heights going from the sea level to 1800 m, where a long dryness season of 5 to 8 months is present, with no frost (Gregorio-Martinez, 2005). In the pacific slopes of Mexico where the maximum degree of variability of this genus is present, the ecosystem is classified as tropical non-evergreen forest, with an annual average temperature of 20-29 °C and a precipitation of 600-1200 mm. This landscape is founded continuously from the south of Sonora and Chihuahua regions to Chiapas and Central America (Campos-Osorno, 2006).

Historical context

The Maya civilization

Yucatan peninsula climate is characterized by recurring hurricanes, savannas pound to flooding, a cave system that drain rainwater, a lack of surface groundwater, and a 6 to 7 months of dry season (Faust, 2001).
Maya culture is one of the most important and sophisticated ancient cultures, from the Mesoamerican cultural area of pre-Columbian time (Vandenabeele et al, 2005). The first settlements of Maya civilization date, from 3500 B.C., in the Yucatan peninsula and, continued until 200 A.D. in a phase known as the Preclassic period. Considering Yucatán environment, water provision was a major concern to the Maya and that is why during the Preclassic phase cities as Edzná and Tikal were constructed with extensive systems for collecting water, using reservoirs and underground tanks. While other cities as Ek Balám, Mayapán and Loltún constructed basins and ditching channels for the management of water (Andrews et al, 1960). In the next map (figure 5) the southern classical sites are showed and the northern post classical sites, including Chichén Itzá.
Chichén Itzá extends near 15 kmand their constructions correspond to several styles related to the different stages of their evolution. The architectonic style that predominated in the first constructions was the Puuc, which is the style of southern and more ancient cities, like Uxmal and Sayil. Examples of this style in Chichén Itzá rea the buildings of the Nuns, the Annex, the Church, the House of the Deer, the Akabdzib, among others.
Then the assimilation of the culture of other areas of Mesoamerica, especially of the Gulf and the Plateau, induced Chichén Itzá to develop a new style in decoration, planning and iconography. This style has been denominated Mayan-tolteca of Chichén Iztá. Examples of this style are constructions as the Castle, the Thousand Columns, the Game of Ball, the Tzomplantli and the Temple of the Soldiers. This city was an important center of the Mayan culture between the years 900 and 1200 A.D. The most formidable constructions are conserved date from this period (Cano, 2002).
But undoubtedly the most remarkable building of all the city is the Kukulcán pyramid (Mayan equivalent of Quetzalcóatl God), also known as “the Castle”. In this building twice a year: in the spring equinox and in autumn solstice a game of shades on the building simulates a serpent that descends until loosing itself in the Earth.
The Castle as other important pyramids in Mesoamerica (like the Templo Mayor) hides an older pyramid inside itself. In its interior there are a Chac Mool (God associated to the rain cult) and a throne of jaguar painted in red with incrustations of jade simulating their skin. This building has been interpreted like the material expression of a calendar. In fact if the number of steps of the four sides and those of the entrance of the temple are added, they sum 365, the same number than the days of the year.

Importance of “Sacred Cenote” at Chichén Itzá and their offerings

Cenotes are natural sinkholes in the limestone of rock that forms the geological base of the peninsule. Among Maya, cenotes were considered as sacred places, and offerings were common. Chichén Itzá is a Classic site, established on theNorthern lowlands of Yucatán (figure 6), “In the border of the well of the Itzá” (Andrews, 1990), undoubtedly a reference to the sacrifice cenote. The city had a height towards years 600 and 900 B.C. soon to be lefted and to be refunded by the “itzáes” in 500 A.D. It was the Mayan priest Lakin Chan, known also as Itzamná that brought new blossoming to this large city. From that moment, their inhabitants named themselves chanes or itzáes.
The Chichén Itzá complex counts with two cenotes:het “Xtoloc” in the west side, and the Sacred or Sacrificial on the east side.
Since the first explorations of Chichén Itzá, copalwas recovered from sacred Cenote. It is no surprising as it was a worship place, where not only Chichén Itza inhabitants celebrated important ceremonies, but also a peregrination site where Mayas from distant places deposed offerings as well (Morley, 1972).
Between 1904 and 1911, Edward H. Thompson explored and, drained Sacred Cenote, finding over 100 gold objects, bones, ceramic vessels, shells, a scepter with anthropomorphic form, 250 objects of jade, pectorals, lances of flint, textiles of cotton, pumpkins but according to their notes “the most abundant material is some pom-incense p ellets, often found into the bottom of blue painted vessels, where copal was “molded or poured before being deposed into the Cenote” (Coggings and Ladd, 1992).
Maya preparation of copal, was quite complex, the copal was worked until it became malleable, copal soot mixed with “Maya blue”, was applied to t he surface of ceramic pieces. In other occasions, different tree exudates were applied to the surface of the object to offer, like chicle (from Manilkara zapota) or rubber (from Castilla elastica). The pattern of the copal varied with round, rolled pellets, bars in a conglomerate and cones.
In the archaeobotanical collection of the INAH (National Institute of Anthropology and History of Mexico) over 200 pieces of copal extracted from the Sacred Cenote of Chichén Itza are preserved. Among these pieces there is one ball molded with ulli (rubber) incrustations, another ball with corn grained applications on its surface, and a human face (Montufar, 2007).

The Aztec civilization

The Mexicas or Aztecs were an ethnic group with nahua filiation, that after a long peregrination funded México-Tenochtitlan, a city that between theXVth and XVIth centuries became the center of one of the biggest empires of all the times in Mesoamerica. Aztecs developed rich and complex religious, political, cosmologic, astronomic, philosophic and artistic traditions. In a good amount Aztecs were the heirs of the cultures of many other ancient Mesoamerican civilizations as the Olmecs, the Teotihuacans and the Toltecs.

The foundation of Mexico Tenochtitlán

When Aztecs arrived to Mexico Valley the Tecpanecs from Azcapotzalco dominated the region, after a short period they established in Chapultepec where they were expelled. Then they settled in Tizapan a territory dominated by Culhuacan, they abandoned this settlement because of the hostile environment.
Finally they took the risk of resettle in a territory under the Tecpanecs control. A little island at Occident of Texcoco lake, in the year 1325 A.D. (year 2 Calli on Aztec calendar). This settlement implicated the subordination to Atzcapotzalco and the payment of tributes. This situation continued until the year 1428 A.D., when Aztec organized, along with Texcoco and Tlacolpan the “triple alliance”, which finally defeated Azcapotza lco.

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The rise of Aztec empire

The importance of this alliance relies on the fact that in barely 100 years, it conquered a big territory: from Atlantic to Pacific Oceans of current territory of Mexico. After a couple of years Aztecs took control over their allies, dominating them thus giving rise to the Aztec empire (Obregon-Rodriguez, 2001).
This empire, as many other empires in history, excerpted an effective political, economic and ideological control over the dominated territory, Aztecs got a big city, as capital of the empire: Mexico-Tenochtitlan, where the proclamations of imperial ideology where marked and reflected in the architecture of the buildings (Smith, 2001). At the heart of this imperial ideology we find the “Templo Mayor”.

The importance of “Templo Mayor” site

The “Templo Mayor” was built at the geographical ce nter of the city it was a precinct surrounded by a staggered platform, where the main buildings of the city where located. It was the tributary center of the empire. Merchandises from a vast territory either sold in markets, or paid as tribute (Carrasco, 1999) were put together in offerings and funerary deposits.
At this temple took place all ritual celebrations: from the regular ones, marked in pre Columbian calendar to the most extraordinary ones corresponding to the enthronization and funerary ceremonies for the emperor (Lopez-Luján, 2010). Ineach of these events offerings were buried (Matos-Moctezuma, 2011).

The building stages in Templo Mayor construction

Although the position of Templo Mayor was known long time ago, it was believed to had been totally destroyed. Since at the moment of the defeat of the Aztecs, all the buildings of the city were destroyed and its materials employed in the construction of European style constructions. It was until 1978 that some excavations of the electricity company found the Coyolxhauqui (Moon Goddess) sculpture. This sculpture is important as documentary sources established its location at the foot of the stairs of Huitzilopoztli altar (Lopez Austin, 2001).
From this moment a program of excavation, conservation and preservation was designed (Renfrew, 1998). This archeological work, has shown that the construction of this ceremonial center was divided in 7 stages, corresponding more or less with each emperor reign, meaning that each ruler addressed a campaign of enlargement of the temple, that consisted in the construction of a superimposed pyramid onto the previous one, preserving the east-west orientation of the building (figure 7).
By the time the Spanish arrived to Tenochtitlan, Templo Mayor reached 60 m height (Matos-Moctezuma, 1987). It was a double Temple, the northern part was dedicated to Tlaloc (God of the rain), while the southern part was dedicated to Huitzilopochtli (God of War).
The Templo Mayor precinct measured 500 m on each side, having a total surface of 250 thousand square meters.
The first stage was probably made of wood, and to our days it is impossible to know if in the inside the remains of the pyramid there are still some remains of the first stage.
The second stage was constructed before 1428 A.D. and is visible now on, Mexico city downtown. There are some paintings preserved on the Tlaloc adoratory as well as a Chac-Mool. In the last staircase just before a sacrificial stone there is a glyph that identifies this stage with a date that in our calendar would correspond to 1390 A.D. This is the earliest fully exposed structure, so its orientation, dimensions and structural characteristics have been measured and described (Aveni, 1988).
The third stage dates from 1431 A.D. (year 4 cane in pre Columbian calendar) that corresponds to Itzcoatl reign. From this period only some stairs and part of the paved are preserved.
The fourth stage present two constructive periods: the first one (dating from around 1454 A.D.) corresponds to Moctezuma I government. From this period, were recovered a good number of ornamental elements such as braziers dedicated to the regent gods of the temple, and the Coyolxauqhi sculpture.
The second constructive period corresponds maybe to the Axayacatl reign, where a platform ornamented with undulating serpents, was added to the front of the pyramid.
From the fifth constructive stage, remain few elements: a stuccoed platform and, part of the ceremonial floor dated from 1470 A.D.

Table of contents :

1.1 General presentation of Mexican copals
1.2 Botanical and geographical distribution
1.2.1 Botanical sources and collection
1.3 Historical Context
1.3.1 The Maya civilization
1.3.2 Chichén Itzá city
1.3.3 Importance of “Sacred Cenote” at Chichén Itzá and their offerings
1.4 The Aztec civilization
1.4.1 The foundation of Mexico-Tenochtitlán
1.4.2 The rise of Aztec empire
1.4.3 The importance of “Templo Mayor” site
1.4.4 The building stages in Templo Mayor construction
1.5 Uses and properties
1.5.1 Copal uses in ancient Mexico
1.5.2 Contemporary uses and applications of copal
1.6 Previous works on chemical composition of Mexican copals
1.7 Oleo-gum-resin: definitions
1.8 Gums
1.9 Gum composition of Bursera simaruba
1.10 Fats
1.11 Terpenic resins
1.12 Bio synthesis of terpenic precursors
1.13 The essential oils
1.13.1 Triterpens
1.13.2 The ursans
1.13.3 The oleanans
1.13.4 The lupans
1.13.5 Biological activity
2.1 Extraction of copal
2.2 Resins from botanical certified origin
2.2.1 Botanical References
2.2.2 Choice of studied species
2.3 Archaeological samples from Templo Mayor site
2.3.1 Description of the offerings from Templo Mayor
2.3.2 Macroambiental conditions
2.3.3 Microambiental conditions
2.4 Description of archeological Aztec offerings
2.4.1 Offering 120
2.4.2 Offering 125
2.4.3 Offering 126
2.4.4 Offering 128
2.5 Archaeological sample from the “Sacred Cenote” of Chichén Itzá
2.5.1 Macroambiental conditions
2.5.2 The Chichén Itzá Cenote: microambiental conditions
2.5.3 Underwater Chichén Itzá cenote explorations
2.6 Commercial Samples
2.7 Presentation of the Analytical Strategy
2.8 Study by optical microscopy
2.9 Conclusion
PART B Analytical Study of the resins and Discussion
3.1 Mathematical Treatment of Experimental data: chemometric strategies
3.1.1 Principal Component Analysis
3.1.2 Linear discriminant analysis (LDA)
3.2 Chemotaxonomy. Correlation of resin composition and botanical origin
3.2.1 Terpenoids and chemotaxonomy
3.3 Study by Fourier-transformed infrared spectroscopy (FTIR)
3.3.1 FTIR spectra of terpene and terpene like molecules
3.4 Sampling Methods: transmission methods
3.5 Reflectance Methods
3.6 Results and discussion
3.7 PCA
3.8 Linear discriminant analysis
3.9 Application to a botanical origin certified sample
3.10 Application to two commercial samples of copal
3.11 Deconvolution of the bands in the region between 1200 and 1300 cm -1 of the spectra. Application of the technique to two commercial resins
3.12 Application to archeological resins
3.13 Partial analysis of the surface of the samples
3.13.1 Comparison of partial analysis from commercial samples and a naturally aged sample of them.
3.13.2 Spectra variation between external and internal part of an archaeological sample TMT
3.14 Conclusions
4.1 High Performance Liquid Chromatography (HPLC)
4.2 Standard molecules
4.3 Quantification of triterpenoids: analytical calibration
4.4 Global analysis of certified resin samples
4.5 Comparison of global chromatograms from certified origin resins
4.6 Comparison of global chromatograms from commercial resins
4.7 Comparison of global chromatograms of archaeological resins
4.8 HPLC coupled to PCA analysis for botanical certified resins
4.9 Linear Discriminant Analysis
4.10 HPLC coupled to PCA analysis for archeological resins
4.11 Purification by Solid Phase Extraction (SPE)
4.12.1 General Methodology
4.13 Conclusion
5.1 Derivatization
5.2 The study of standard triterpenoic molecules
5.3 Results of the study on botanical certified samples
5.4 Comparison of global chromatograms from certified origin resins
5.5 Insight on triterpenic composition for samples of certificated origin
5.5.1 Comparison between B. bipinnata and B. stenophylla resins
5.5.2 B. excelsa triterpenic composition
5.5.3 B. grandifolia triterpenic composition
5.5.4 B. laxiflora triterpenic composition
5.5.5 B. penicillata triterpenic composition
5.5.6 B. simaruba triterpenic composition
5.5.7 B. copallifera triterpenic composition
5.6 Study of archeological samples
5.6.1 Molecular composition of archeological sample 26
5.6.2 Molecular composition of archeological sample 51
5.6.3 Molecular composition of archeological sample 52
5.6.4 Molecular composition of archeological sample 84
5.6.5 Molecular composition of archeological sample 173
5.6.7 Molecular composition of archeological sample TMT
5.6.8 Molecular composition of archeological sample 140
5.6.9 Molecular composition of archeological sample from Chichén Itzá
5.7 Study of commercial samples
5.7.1 Molecular composition of Sidral
5.7.2 Molecular composition of commercial sample SONB4
5.7.3 Comparison between molecular composition of ATZJ2 and SONB4
5.7.4 Molecular composition Oaxc2
5.7.5 Molecular composition IZUP1 and SONR1
5.8 Comparison of partial analysis from the surface and the inner part of the samples
5.8.1 Comparison between SONB4 and a sample naturally aged portion of it
5.8.2 Molecular variation between external and internal part of an archaeological sample TMT
5.9 Conclusions


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