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Revista mexicana de ciencias agrícolas
versión impresa ISSN 2007-0934
Rev. Mex. Cienc. Agríc vol.7 spe 14 Texcoco feb./mar. 2016
Articles
Use of principal component in melissopalynology classification of honey from Apis mellifera L.
1Colegio de Postgraduados-Campus Tabasco. Periférico Carlos A. Molina s/n, km 3.5. Carretera Cárdenas-Huimanguillo. C. P. 86500, Cárdenas, Tabasco. (selene. jacinto@colpos.mx; rodolfo@colpos.mx; sol@colpos.mx; luis@avanzavet.com).
2Universidad Politécnica del Centro. Carretera Federal Villahermosa-Teapa, km 22.5, R/a. Tumbulushal, C. P. 86290, Villahermosa, Tabasco. (iqreyes@hotmail.com).
38 honey samples from Apis mellifera L. were collected in six municipalities belonging to five subregions from the state of Tabasco, the melisopalinological analysis was performed, obtaining 37 important pollen types (>10%). Bursera simaruba L. (Burseraceae), Spondiasmombin (Anacardiaceae), Diphysa sp. (Fabaceae), Erythrina sp., (Fabaceae), Mimosa orthocarpa (Fabaceae), Pipersp1, sp2, sp3 (Piperaceae), Machaerium sp., (Fabaceae), Cecropia obtussifolia (Moraceae), Psidium guajava (Myrtaceae), Acalypha sp., (Euphorbiaceae), Celtis sp., (Ulmaceae), Heliocarpus appendiculatus (Tiliaceae), Conocarpus sp., (Combretaceae), Haematoxy lumcampechianum (Fabaceae), Mimosa albida (Fabaceae), Zea mays L. (Gramineae), Diphysacarthagenensis (Fabaceae), Quercus sp., (Fagaceae), Citrus sp., (Rutaceae), Cocos nucifera (Arecaceae), Mimosa sp., (Fabaceae), Mimosa pigra var. Berlandieri (Fabaceae), Eleocharis sp., (Cyperaceae), Rumex sp., (Polygonaceae), Eragrostis sp., (Poaceae), Spondiasradlkoferi (Anacardiaceae), Muntingiacalabura (Elaeocarpaceae), Asteraceae sp1, (Asteraceae), Borreriaverticillata (Rubiaceae), Coccolobaaff. Diversifolia (Polygonaceae), Quercusoleoides (Fagaceae). Overall honey samples from 22 multi-floral, 9 monofloral and 7 bifloral were classified. And when performing the PCA, two components that explain 65% of total variations were obtained, showing that Cardenas and Huimanguillo have affinity for Bursera simaruba L. (Burseraceae) and Cecropiaobtussifolia (Moraceae), instead Centro, Tacotalpa and Balancan share a preference for Mimosa albida (Fabaceae), Psidiumguajava (Myrtaceae) and Rumex sp. (Polygonaceae) and Centla showed a clear difference with the other 5 municipalities.
Keywords: honey; melissopalynology; PCA; Prinqual; Tabasco
Se colectaron 38 muestras de miel de Apis mellifera L. en seis municipios, pertenecientes a las cinco subregiones del estado de Tabasco, se realizó el análisis melisopalinologico obteniendo 37 tipos polínicos de importancia (>10%). Bursera simaruba L. (Burseraceae), Spondiasmombin (Anacardiaceae), Diphysa sp. (Fabaceae), Erythrina sp., (Fabaceae), Mimosa orthocarpa (Fabaceae), Pipersp1, sp2, sp3 (Piperaceae), Machaerium sp., (Fabaceae), Cecropia obtussifolia (Moraceae), Psidium guajava (Myrtaceae), Acalypha sp., (Euphorbiaceae), Celtis sp., (Ulmaceae), Heliocarpus appendiculatus (Tiliaceae), Conocarpus sp., (Combretaceae), Haematoxy lumcampechianum (Fabaceae), Mimosa albida (Fabaceae), Zea mays L. (Gramineae), Diphysacarthagenensis (Fabaceae), Quercus sp., (Fagaceae), Citrus sp., (Rutaceae), Cocos nucifera (Arecaceae), Mimosa sp., (Fabaceae), Mimosa pigra var. Berlandieri (Fabaceae), Eleocharis sp., (Cyperaceae), Rumex sp., (Polygonaceae), Eragrostis sp., (Poaceae), Spondiasradlkoferi (Anacardiaceae), Muntingiacalabura (Elaeocarpaceae), Asteraceae sp1, (Asteraceae), Borreriaverticillata (Rubiaceae), Coccolobaaff. Diversifolia (Polygonaceae), Quercusoleoides (Fagaceae). En general se clasificaron 22 muestras de miel multifloral, 9 monofloral y 7 bifloral. Y al realizar el ACP, se obtuvieron dos componentes que explican 65% de la variación total, mostrando que Cárdenas y Huimanguillo tienen afinidad por Bursera simaruba L. (Burseraceae) y Cecropiaobtussifolia (Moraceae), en cambio Centro, Tacotalpa y Balancán comparten preferencia por Mimosa albida (Fabaceae), Psidiumguajava (Myrtaceae) y Rumex sp. (Polygonaceae) y Centla en cambio mostró una clara diferencia con los 5 municipios restantes.
Palabras claves: ACP; miel; melisopalinológia; Prinqual; Tabasco
Introduction
The annual production of honey in Mexico is 55 thousand 500 t, which is enough to meet domestic and external demand. In the last decade, honey has had an increase of two thousand tons (SENASICA, 2013), increase that has not been accompanied by higher prices due to recurrent adulteration of the product offered and the lack of quality. Currently, the honey industry is facing new market horizons both within and outside the country, therefore melissopalynology characterization of honey and its geographical location, are fundamental tools in defining floral species from which it comes, complementing this information with their physicochemical and organoleptic characteristics (De la Serna-Ramos, 2009). Defining quality standards of honey, perfectly located geographically and differentiated based on their unique outstanding characteristics, will allow consumers to have the opportunity to define their preferences, increase demand and, consequently, improve its price.
To assess the commercial quality of honey is necessary to consider its flavor, aroma, texture, color, moisture and sugar content, among others. Thus, a mono-floral honey has a characteristic smell and taste, which is appreciated by consumers, and therefore has a better price. However those coming from different plant species without a predominant called multi-floral does not have the same preference (Telleria, 2001). Therefore, the determination of the botanical origin of honey, based on qualitative and quantitative analysis of the type of predominant pollen, locating the plants from where they come, it is essential for management and location of new apiaries and possibly the achievement of greater harvest (Schweitzer, 2009). Although, if the geographical areas of honey production overlap or are very close, the geographical identification of honey is difficult, since their variations are minimal, which are influenced by weather conditions and flowering season (Varis et al., 1982 and 1983).
The principal component analysis (PCA) is a multivariate statistical technique that has been used in exploratory studies of data from surveys, direct observation or regular information and allowed the description of the total variation of pollen samples and classifies the honey according to their area of origin (Krauze and Zelewski, 1991). Using information from the melissopalynological analysis, it is possible the classification or cluster formation in similar production units. The PCA facilitates the reduction of cluster data set, by transforming them in a new uncorrelated cluster (Herrera, 2010).
The objective of this research was to classify honey from different floral sources, using principal component analysis.
Materials and methods
Description of the study area
The state of Tabasco is located geographically southeastern Mexico between 17° 15' 00'' and 18° 39' 07'' north latitude, 90° 50' 23'' and 94° 07' 49'' west longitude. Limit to the north with the Gulf of Mexico, northwest with the state of Campeche, in the southeast with the Republic of Guatemala and south and west by the states of Chiapas and Veracruz, respectively. The state of Tabasco has five sub-regions characterized by different geographical, ecological and socioeconomic conditions: Chontalpa Centro, Sierra, Rivers and Swamps. In this research samples were collected in various apiaries located in the five sub-regions suitable for the development of apiculture according to SAGARPA (2004).
Field work
During 2006, 2007 and 2008 the fieldwork was conducted in six apiaries located in five regions, sampling only six municipalities within them. In each apiary collected sedimented honey samples in plastic bottles and stainless steel, using plastic bottles with a capacity of 200 ml. The honey contained in the bottles was previously centrifuged, settled and filtered so that it is free of impurities, according to the technique from apiarist. Each sample was identified with a label with the name of the municipality, apiarist, date of collection and region of origin (Table 1).
The samples were stored in the Laboratory of Food Science Campus Tabasco from the Postgraduate College in Agricultural Sciences, until analysis and identification.
Laboratory analysis
Analysis melisopalinological
The melisopalinological analysis was performed on the Palynology Laboratory from the Institute of Geology from the National Autonomous University of Mexico (UNAM) (IGLUNAM) under the direction of Dr. Elia Ramirez Arriaga, in order to have better representativeness from the pollen content of the honey samples, the conventional technique was modified from Louveaux et al. (1978), later proceeded to acetolysis following the method from Erdtman (1969), 4 platelets per sample were prepared, which were recorded and incorporated into the palynological collection of UNAM.
These analyzes included the processing of honey samples to know pollen morphology of pollen grains and to describe them, to identify to the possible level and count them to know its representativeness, this analysis has two stages: qualitative and quantitative.
Qualitative analysis
Preparations were observed under a transmitted light microscope Carl Zeiss phase contrast and under 100 x objective. The pollen types found at a frequency higher than 10% were described.
This allowed the identification, by comparison with the reference collection from the same laboratory, besides Palynological Catalogues were consulte as the "Atlas of plants and pollen used by the five main species producing honey bees in the region from Tacana, Chiapas, Mexico" (1993) and "palynological flora from Sian Ka'an biosphere reserve in Quintana Roo, Mexico" (1991), in addition to the advice of specialists in melissopalynology.
Determinations of pollen grains were made at family, genus and / or species level. In cases where could not be determined, this were assigned a number corresponding to a morphological type.
Quantitative analysis
Once described and differentiated the pollen types, proceeded to count 500 grains randomly, according to Louveaux and Vorwhol (1977) and independently recording Lycopodium clavatum spores. Subsequently, percentages of each species were calculated to know its representativeness in each sample, species with percentages >10% were considered significant.
Absolute number of pollen grains
To calculate the total amount of pollen grains per gram of sample, Stockmarr (1971) and Maher (1981) formula was used:
Where: Pconc is pollen per gram; R is counted pollen grains / number of spores counted; M is the number of spores added in each tablet; V is the total weight of honey sample. Pollen concentrations were classified following the scheme from Maurizio (1939).
Botanical characterization of honey
Once the absolute number of pollen grains, honey was characterized as "mononofloral" when in its composition had a species with a percentage greater than or equal to 45% and multifloral when three or more species were with percentages ≥ 10% (Louveax and Vorwhol, 1977; Ramírez-Arriaga and Martinez-Hernández, 2009). The latter were divided into "bifloral" when two species were of importance, though it has not been formally accepted, but in this research were considered for the classification honey samples.
Statistic analysis
The results from the melisopalinological analysis of honey samples from six municipalities, were analyzed using the Statistical Analysis System (SAS) version 9.3 program using a preference of pollen analysis through PRINQUAL procedure, which performs principal component analysis (PCA) from qualitative, quantitative and mixed data, finding linear and nonlinear transformations of the variables, using partial least square, optimizing the correlation or covariance matrix properties of the transformed variables. Of all the samples taken from each municipality, recurrent species were added forming a single percentage, thus distinguishing preferences for certain species and then to the principal components analysis with qualitative data.
PROC PRINQUAL was used to perform a non-metric multidimensional preference (MDPREF) analysis. MDPREF analysis is a principal component analysis of a matrix with columns that correspond to municipalities and rows corresponding to pollen types. The percentages are the preferences of the municipalities for each pollen type. The end result of this analysis is a biplot; that is, a graphical representation of municipalities preferences for certain species.
Results and discussion
Melisopalinological analysis
The analysis from 38 honey samples from Apis mellifera L. of Tabasco state, showed a total of 37 pollen types of importance, which were identified at genus, species and family level, the rest were assigned to morphological types (Table 2, 3, 4, 5, 6, 7).
Table 4 Results of the melisopalinological analysis of samples from the municipality of Huimanguillo.
![](/img/revistas/remexca/v7nspe14//2007-0934-remexca-7-spe14-2831-gt4.gif)
In the municipality of Cárdenas was reported a monofloral honey sample of Cecropia obtussifolia and three from multi-floral.
For the municipality of Tacotalpa two monofloral honey samples of Diphysa carthagenensis and Mimosa albida, two bifloral samples and eight multifloral samples were reported.
For the municipality of Huimanguillo reported three multifloral honey samples.
For the municipality of Centla reported one sample of monofloral honey sample from Cocos nucifera, a bifloral sample and a multifloral sample.
For the municipality Centro reported a monofloral honey sample from mimosa albida, two bifloral samples and five multifloral samples.
For the municipality Balancan reported five monofloral honey samples, four from Rumex sp., and one from Mimosa albida, two bifloral samples and one multifloral sample.
S. mombin, (jobo roñoso), C. nucifera, B. simaruba (palo mulato), M. albida, M. pigra var. Berlandieri, M. sp., Zea mays, C. aff. diversifolia y Rumex sp., Psidium guajava, Piper sp1, sp2, sp3, Celtis, Q. oleoides, C. obtusifolia (Guarumo), H. appendiculatus (jolotzin), H. campechianum (Palo de tinto), B. verticillata, Muntingia calabura, Erythrina, D. carthagenensis, have been reported as part of the honey flora from Mexico (Espina and Ordetx 1983), Valley of Mexico (Stones and Quiroz 2007), the state of Tabasco (SAGARPA, 1998, 2002, 2003 and 2004; Cardenas, 1985; Villegas et al., 2004); Oaxaca (Santos Ramos, 2008; Navarro, 2008; Ramírez-Arriaga et al., 2011); Puebla (Ramírez-Arriaga and MartínezHernández 2007); Yucatán (Villanueva et al., 1998, 2002); Veracruz (Villegas et al., 2003); Campeche (Porter et al., 2001); South America (Sodre et al., 2007) and it is part of the bee flora by field observations in tropical regions (Espina and Ordetx, 1983).
Principal component analysis
The results from the PROC Prinqual analysis shows eigenvalues and explained variance, observing separation of the first two components explaining 65% of the variability and the rest of it (Figure 2).
Since the MDPREF analysis is based on a model of principal components, the biplot, which is a graphical representation showing the relationship between the matrix data (columns and rows); the biplot dimensions are the first two components (Figure 3). The first component is one that has a greater dimension of the biplot, is the global preference of municipalities. The second component is orthogonal to the first, being the second most preferred. In the biplot, shows points that observe the pollen types and each municipality by a vector is. The points are closely grouped in a region of the graph represent the pollen types that have the same preference patterns through the municipalities. The vectors points in more or less to same direction represent the municipalities with similar preferences patterns.
On the upper left side can be seen the preference from M1 (Cárdenas) and M3 (Huimanguillo) by type of pollen: Cec (Cecropia obtussifolia), Bur (Bursera simaruba L.), Leg. (Leguminosae), Com. (Compositae) and Que (Quercus sp.). On the right side the preference from M2 (Tacotalpa), M5 (centro) and M6 (Balancán) by pollen type: Mia (Mimosa albida), Psi (Psidium guajava) amd Rum (Rumex sp.), and finally the preference from M4 (Centla) by Coc (Cocos nucifera) and Gra (Gramineae). This result is explained by the percentages of pollen type present in the samples taken from each municipality.
Conclusions
The analysis of 38 samples of honey from Apis mellifera L. Tabasco state, showed a total of 37 pollen types of importance and make the ACP, two components that explain 65 % of the total variation were obtained, showing that Cardenas Huimanguillo have affinity for Bursera simaruba L. (Burseraceae) and Cecropia obtussifolia (Moraceae), however Centro, Tacotalpa and preference share Balancán Mimosa albida (Fabaceae), Psidium guajava (Myrtaceae) and Rumex sp. (Polygonaceae) and Centla instead showed a clear difference with the other 5 municipalities.
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Received: October 2015; Accepted: January 2016