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A Data Mining Approach to Improve Inorganic Characterization of Amanita ponderosa Mushrooms.

1. Introduction

Mushrooms are known from ancient times for their medicinal properties and gastronomic properties. Therefore the consumption of edible wild-growing mushrooms has been very popular. The demand for the commercialization of edible wild mushrooms has proved to be a widely expanding business with increasing economic importance in many rural areas of some countries. In recent years, the consumption of edible mushrooms has been increasing and gaining prominence due to their gastronomic potential, also for their both organoleptic properties (texture and pleasant aroma) [1-4], their chemical composition, mineral content, and nutraceutical value [4-8]. Mushrooms are an important source of proteins, dietary fibres, and vitamins (B, C, D, E) containing low levels of sugar and fats. They can assimilate large amounts of water and minerals such as phosphorus, iron, potassium, cadmium, magnesium, copper, and zinc, due to the large area of mycelium overgrowing the surface layer of soil [9]. This mycelia network is ideally suited to penetrate and access soil pore spaces and an extensive surface area of fungal hyphae and physiology enable for many species on effective absorption and bio concentration of various metallic elements, metalloids, and nonmetals [10]. Bioconcentration factor (BCF), the ratio of the element content in fruiting body to the content in underlying substrate, can express the ability of fungi to accumulate elements from substrate, and this capacity of the mushroom is affected by fungal lifestyle, age of fruiting body, specific species and element, and environment such as pH, organic matter, and pollution [9]. Moreover, the symbiotic relationships that some mushrooms species, namely, A. ponderosa, can establish with some plants of their habitats allowing the accumulation of high concentrations of some metals. Therefore, mineral content and organic composition of edible mushrooms are dependent on the species and the characteristics of the ecosystems in which they are inserted [11, 12]. Some minerals are essential elements for the correct human body function, although others may present toxicity [5, 6, 9, 11, 13, 14]. There are many species of edible mushrooms growing wild, and some of them are slightly characterized about minerals content and the potential of heavy metals bioaccumulation such as lead, mercury, cadmium, and silver [5, 13, 15-17]. On the other hand, due to the great diversity of wild mushrooms, the similarity between some species and the lack of knowledge of others may lead to intoxication when harvesting wild species, leading in some cases to death [18].

The genus Amanita is one of the best known from Agaricales order and comprises edible and poisonous mushrooms distributed worldwide, occupying mainly a mycorrhizal habitat and playing a significant role in forest ecosystems [19, 20]. This genus includes important species of edible mushrooms, as is the case of Amanita ponderosa [18, 20-24]. The south of Portugal, due to its Mediterranean characteristics and diversity of flora, is one of the regions of Europe with a greater predominance of A. ponderosa wild edible mushrooms [23]. These robust basidiomata grow during spring in mounted areas with acid soils, in forests of holm oaks and cork trees like Quercus ilex and Quercus suber, and with shrubs like Cistus ladanifer, Cistus laurifolius, and Lavandula stoechas establishing a symbiotic relationship with them (Figure 1). Gastronomically, this species is very relevant, not only due to the traditional consumption in the rural populations, but also due to its commercial value in the gourmet markets having high exportation potential in Portugal, thus, the chemical and mineral characterization of numerous species of edible mushrooms for certification purposes and further commercialization process becoming of extreme importance.

In recent years, some artificial intelligence based tools, namely, Data Mining, Artificial Neural Networks (ANNs), and Decision Trees (DTs) have been applied for fungal environment systems [18, 25, 26]. Data mining tools were used in latest studies about A. ponderosa mushrooms in order to establish a bridge of inorganic contents, molecular fingerprints, and geographical sites. In one study a segmentation model based on the molecular analysis was developed, which allowed relating the clusters obtained to the geographical site of sampling. There were also developed explanatory models of the segmentation, using Decision Trees, to relate the molecular and inorganic data, following two different strategies: one based on DNA profile and another based on the mineral composition [22]. Another study based on ANNs exposed the selected model and can predict molecular profile based on inorganic composition with a good match between the observed and predicted values [18].

The aim of this study was to determine the inorganic composition of fruiting bodies of A. ponderosa mushrooms from different sampling sites in the southwest of the Iberian Peninsula, analysing the variations in mineral content. Additionally, the mineral composition of the corresponding soil substrates was analysed in order to correlate with the mineral composition of the fruiting bodies. In the present work, the k-means clustering method was used to study the mineral composition of A. ponderosa fruiting bodies. To explain the segmentation model, that is, in order to obtain rules to assign a case to a cluster DTs were used.

2. Materials and Methods

2.1. Sampling Collection. Fruiting bodies of the Amanita ponderosa mushrooms were collected in spring, between February and April, from 24 different sampling sites, in the southwest of the Iberian Peninsula, namely, 11 samples collected from Alto Alentejo region and 10 from Baixo Alentejo, Portugal, 2 samples collected from Andalusia and 1 from Extremadura, Spain (Table 1). These mushrooms were collected in acid soils, in forests of Quercus suber, Q. ilex ssp. ballota, Cistus ladanifer, and Cistus laurifolius (Figure 1).

Three individuals in the same growth stage were selected per each sampling site to avoid the effect of size. Fruiting bodies were identified by a specialist, based on morphological features according to taxonomic description of A. ponderosa [27]. During the collection process fruiting bodies were placed in wicker baskets and samples were transported in refrigerated containers and deposited in the Biotechnology Laboratory of Chemistry Department of University of Evora (Portugal). In parallel, soil samples were randomly collected from the surrounding soil of fruiting bodies of each site.

Fruiting bodies samples were weighed and carefully washed with double distilled water and representative samples of each sampling site were catalogued, stored in sterile bags, and preserved at -20[degrees]C for its inorganic study. The sheath was eliminated during the pretreatment of fruiting bodies samples, since it is not usually consumed.

Soil substrates were sampled (0-15 cm), after removing some visible organisms, small stones, sticks, and leaves. Samples were air-dried in room temperature for 1-2 weeks and, subsequently, sieved through a pore size of 2 mm and stored at -4[degrees]C.

2.2. Inorganic Characterization. Mineral composition was determined in A. ponderosa fruiting bodies and respective soil collected in 24 different sampling sites described, using different analytical techniques: Flame Atomic Absorption Spectrometry (FAAS), Flame Emission Photometry (FEP), and UV-Vis molecular absorption spectrometry.

2.2.1. Treatment of Samples. Mineral composition analysis of fruiting bodies was performed by the dry mineralization method [28]. Three samples of mushrooms from each sampling site (25 g, fresh weight) were dried in a furnace at 100[degrees]C to constant weight (48 h) from which the moisture content was calculated. Samples were homogenized, transferred to porcelain crucibles, and incinerated in a muffle furnace (Termolab) at 460[degrees] C for 14 h. In order to determine the organic matter and mineral content in mushrooms samples, ashes were weighted at constant weight. Afterwards, ashes were bleached after cooling by adding 2 M nitric acid, drying them on thermostatic hotplates, and maintaining them at 460[degrees]C for 1 h. Ashes recovery was performed with 15 mL of 0.1 M nitric acid and stored at 4[degrees]C until analyses.

Three soil samples (5 g dw) collected from each sampling site were cold treated with an extracting solution of 25% nitric acid (HN[O.sub.3]) (40 ml) and shaking with orbital agitation for 24 h at room temperature. The extracts obtained after filtering through Whatman No. 42 filter paper were stored into polyethylene bottles and stored at 4[degrees]C until analyses [11].

2.2.2. Analytical Determinations. Aluminium (Al), barium (Ba), calcium (Ca), cadmium (Cd), lead (Pb), copper (Cu), chromium (Cr), iron (Fe), magnesium (Mg), manganese (Mn), silver (Ag), and zinc (Zn) contents were quantified by flame atomic absorption spectrometry (Perkin-Elmer 3100 model) with atomization in an air-acetylene flame and single element hollow cathode lamps and background correction with deuterium lamp for manganese. For the determinations of calcium and magnesium, strontium chloride (Sr[Cl.sub.2]) was added to make up a final concentration of 0.1125% of the sample, in order to prevent anionic interferences which might modify the result [29].

Sodium (Na) and potassium (K) were quantified by flame emission photometry with a Jenway model PFP7 flame emission photometer [30, 31]. Phosphorus (P) was quantified by vanadomolybdophosphoric acid colorimetric method, using a spectrophotometer (Hitachi, U-3010 model) [32].

All sample determinations were performed in triplicate for each one of the three independent ash extracts of mushroom samples from 24 sampling sites (n = 9) and for three extracts of each soil substrate (n = 9). Concentration values were calculated through the standard curves for each element expressed in mg/kg dw. The bioconcentration factor (BCF) value, which is the quotient of the concentration in fruiting bodies divided by the concentration in the soil substrates, was determined for all minerals in each sampling site.

2.3. Data Mining

2.3.1. Cluster Analysis. In the present study a cluster analysis was carried out. The technique applied in order to build up clusters was the k-means clustering method [33] and the software used was WEKA [34]. In WEKA Simple k-means algorithm the normalization of the numerical attributes is carried out automatically when the Euclidean distance is computed. A more detailed description of the mentioned algorithm can be found in Witten [35].

2.3.2. Decision Trees. In the k-means clustering method clusters were formed without information about the groups and their characteristics. Therefore, it is necessary to understand how the clusters were formed. To attain such a purpose Decision Trees (DTs) were used. In this study the algorithm used to generate DTs was the WEKA J48 [34], corresponding to the 8th revision of the C4.5 algorithm. The detailed description of the J48 algorithm can be found in Witten et al. [35].

2.4. Statistical Analysis. Data were evaluated statistically using the SPSS[R] 21.0 software Windows Copyright[c], (Microsoft Corporation), by descriptive parameters and by one-way ANOVA in order to determine statistically significant differences at the 95% confidence level (p < 0.05). The homogeneity of the population variances was confirmed by the Levene test and the multiple comparisons of media were evaluated by Tukey's test.

3. Results

3.1. Fruiting Bodies and Soil Substrates' Mineral Composition. Mineral analysis of A. ponderosa mushrooms samples collected from 24 sampling sites of Alentejo (Portugal), Andalusia, and Extremadura (Spain) regions was evaluated.

Table 2 shows the contents in moisture, organic content, and minerals present in the samples of A. ponderosa collected in the different places. Moisture content ranged from 89.5[+ or -] 0.0 to 93.8[+ or -]0.5% (Table 2). The analyses of variance (one-way ANOVA) showed that A. ponderosa fruiting bodies samples collected in Almendres (1) and Serpa (20) presented significantly higher values compared to the other samples, while the sample from Mina S. Domingos (11) showed moisture content significantly lower than the remaining samples. Dry weight values, for several samples, ranged between 6.2 [+ or -] 0.5 and 10.5 [+ or -] 0.1%, consisting in organic content values between 5.8 [+ or -] 0.3 and 9.8 [+ or -] 0.1% and minerals between 0.5 [+ or -] 0.0 and 1.4 [+ or -] 0.1%. Fruiting bodies samples collected in Almendres (1) and Serpa (20) presented values of organic content significantly lower and the sample collected in the Mina S. Domingos (11) significantly higher than all analysed samples (p < 0.05). The mineral content of the fruiting bodies collected in Cabezas Rubias (6) was 1.4 [+ or -] 0.1%, a value significantly higher than the remaining samples. However, the samples from Serpa (20) and Villanueva del Fresno (24) showed the lowest values, significantly different from the another samples (p < 0.05).

Figure 2 shows the average contents of water, dry weight, organic content, and minerals present in the 24 samples of A. ponderosa analysed. The fruiting bodies presented water content, corresponding to 90.3 to 93.1% of their fresh weight. The organic matter content ranged between 6.2 and 9% and mineral content between 0.5 and 0.9%. Thus, 100 g of edible A. ponderosa mushrooms corresponds to a maximum of 9 g of macronutrients, such as lipids, carbohydrates, and proteins, and less than 1 g of mineral content. These values were similar to those observed in a study with mushrooms harvested in some regions of Andalusia (water content 87.8%, organic content 11.8%, carbohydrates 6.6%, proteins 3.2%, lipids 0.5%, fibre 1.5%, and mineral content 0.4%) [24] and these fruiting bodies have a caloric content of 42kcal/100 g of mushroom, similar to other edible mushroom species, characterized as low calorie foods [8, 28, 36].

Tables 3-6 show the mineral content of 24 sampling sites of A. ponderosa fruiting bodies as well as their corresponding soil samples. The studied A. ponderosa fruiting bodies showed higher mineral content in macroelements calcium (Ca), magnesium (Mg), sodium (Na), potassium (K), phosphorus (P), and microelements aluminium (Al), copper (Cu), and iron (Fe) (Tables 3 and 4). Potassium was present in higher concentrations and was higher in Cabezas Rubias (6) fruiting bodies, 69565 [+ or -] 362 mg/kg dw. The samples showed lower values of the trace elements silver (Ag), barium (Ba), cadmium (Cd), chromium (Cr), manganese (Mn), lead (Pb), and zinc (Zn) (Tables 5 and 6).

Minerals contents of A. ponderosa fruiting bodies and their corresponding soil samples presented significant differences for all the studied elements (p < 0.05). Cabezas Rubias (6) presented significantly higher aluminium and calcium contents (p < 0.05). The cadmium content was significantly higher (p < 0.05) in the fruiting bodies collected in N. [S.sup.ra] Machede (16) and Almendres (1). Samples from Evora (7), Cabezas Rubias (6), and Villanueva del Fresno (24) showed similar chromium contents (p > 0.05), which were significantly higher than the others A. ponderosa fruiting bodies (p < 0.05). Results obtained for soil mineral content showed some significant differences between sampling sites (p < 0.05). The sample collected in N. [S.sup.ra] Machede (16) presented significant different contents of silver, aluminium, and magnesium (p < 0.05). The Montejuntos sample (14) presented significantly differences of aluminium, iron, phosphorus, and magnesium contents (p < 0.05) and Villanueva del Fresno (24) presented significant differences for barium and manganese (p < 0.05). Significant differences in copper content were observed for [Her.sup.de] da Mitra sample (9). Mertola sample (10) presented significant differences in zinc and Baleizao sample (3) in potassium contents (p < 0.05). Serpa samples (20), presented significant differences in calcium, chromium, iron, magnesium, sodium, and zinc (p < 0.05). Almendres (1), Evora (7), [M.sup.te] Novo (13) and Valverde (22) samples did not present significant differences for the iron content (p > 0.05). Samples of Cabezas Rubias (6), Serpa (20), and Rosal de la Frontera (17) did not present significant differences in lead content (p > 0.05). Concerning phosphorus content, two groups with no significant differences (p > 0.05) were observed, one including Baleizao (3) and Rosal de la Frontera (17) and the other group including [S.sup.to] Aleixo Restauracao (18), S. Miguel de Machede (19), and Vila Nueva del Fresno (24) samples. Finally, samples from Baleizao (3) and Valverde (22) did not present significant differences in sodium and potassium content (p > 0.05).

In fact, mineral content of A. ponderosa fruiting bodies and their soil samples (Figure 3) varied very similarly for almost all the analysed elements, indicating the influence of the substrate characteristics on the composition of mushrooms samples collected at the different sites.

Mushrooms have a specialized mechanism to accumulate nutrients and minerals in their fruiting bodies. The age of the fruiting body or its size may contribute to mineral composition. Thus, A. ponderosa mushrooms analysed in this study were obtained at the same development stage, in order to eliminate possible size interferences in the comparison of their mineral content. Macroelements content, such as Ca, Mg, Na, K, and P, in fruiting bodies were similar to those reported in literature for A. ponderosa [28] and for other edible mushrooms species [5, 36-38]. The calcium and phosphorus contents for the different samples studied were 523 [+ or -] 143 and 294 [+ or -] 171 mg/kg dw. Potassium and sodium, minerals responsible for the hydroelectrolytic balance maintenance and important enzyme cofactors, have high RDIs (Recommended Daily Intakes), which are 4700 mg/day and 2000mg/day, respectively, for an adult [39]. Potassium and sodium levels of A. ponderosa mushrooms studied were 29648 [+ or -] 14908 and 1092 [+ or -] 620 mg/kgdw, respectively. Potassium content ranged from 18021 [+ or -] 1806 to 69565 [+ or -] 362 mg/Kg dw, presenting values similar to those described by Moreno-Rojas et al. [28], that report potassium levels ranging between 22410 [+ or -] 211 and 60890 [+ or -] 23950 mg/Kg dw. However, the low K levels observed for fruiting bodies from three sampling sites can be correlated with the large differences observed for potassium content in the surrounding substrates. Magnesium also plays an important role in large number of biological functions, particularly linked to energy metabolism, it is required for the proper function of certain enzymes as cofactor, and structural functions, the recommended magnesium intakes for adult (19-65 years), are 220-260 mg/day [40]. The mean magnesium content in the fruiting bodies was 738 [+ or -] 261 mg/kgdw, similar to that described in the literature [28].

A. ponderosa fruiting bodies presented values of trace minerals smaller than those found in other species of edible mushrooms [6, 11, 14, 38, 41]. Concentrations of trace elements in fruiting bodies are generally species-dependent [36]; however only one study of A. ponderosa mushrooms is reported [28]. The existence of higher metal concentrations in younger fruiting bodies can be explained by the transport of metals from the mycelium to the fruiting body during the beginning of fructification [36]. Trace elements like Fe, Cu, Zn, Cr, and Mn are essential metals since they play an important role in biological systems; however, these trace metals can also produce toxic effects when intake is excessively amount [38, 42, 43]. Copper is the third most abundant trace element found in the human body, and being an important element of several enzymes, it is also one of the agents involved in iron metabolism [42]. Iron is a component of haemoglobin and myoglobin, proteins responsible for transporting oxygen to tissues. It also participates in protein metabolism, energy production in cells and in various enzymatic reactions [44]. The copper content was 185 [+ or -] 125 mg/kg dw, and the highest value was found in A. ponderosa samples collected from Rosal de la Frontera (17) (584 [+ or -] 51 mg/kg dw). The iron content in A. ponderosa mushrooms was 92 [+ or -] 85 mg/kg dw. Copper levels were similar and iron levels are slightly lower than those described for Amanita spp. [5, 28]. The adult RDI is 2 mg/day for Cu and 18 mg/day for Fe [12, 39], so the concentrations of copper and iron present in these edible mushrooms are not considered to be a health risk. Manganese and zinc are important trace elements for the human organism, participating in macronutrients and nucleic acids metabolism and promoting several enzyme activity processes [14, 45, 46]. These elements can be accumulated by mushrooms and the recommended daily intakes were 2 mg/day and 15 mg/day, for manganese and zinc, respectively [12, 39]. In this study, mushrooms presented a mean value of 56 [+ or -] 27 mg/kg dw for manganese and 68 [+ or -] 25 mg/kg dw for zinc content, similar to values described in literature [5, 6, 11, 36, 47]. Chromium biological functions are not known precisely; it seems to participate in the metabolism of lipids and carbohydrates, as well as in the insulin action. The RDI for this metal is 120 [micro]g/day. The mean chromium content obtained was 1,19 [+ or -] 0,74 mg/kg dw, lower than those reported in the literature for other species of Amanita [5] and similar to some species of edible mushrooms described [11, 36]. Aluminium is one of the few abundant elements in nature but no significant biological function is known, although there are some evidences of toxicity when ingested in large quantities. Most acidic soils are saturated in aluminium instead of hydrogen ions, and this acidity is the result of aluminium compounds hydrolysis [48]. A. ponderosa fruiting bodies from the different sampling sites showed a high range of aluminium content with a medium value of 362 [+ or -] 204 mg/Kg dw. High aluminium levels were reported for some Amanita species, for example, A. rubescens that showed values around 262 mg/kg dw [15] and A. strobiliformis and A. verna presented aluminium levels of 72 and 343 mg/kg dw, respectively [5]. Other studies report different levels of aluminium in Amanita rubescens: 293, 75, and 512 mg/kg dw for the whole fruiting body, cap, and stipe, respectively [37]. The large range in aluminium content was also reported in a study of A. fulva that showed aluminium levels ranging from 40 to 500 mg/Kg dw in the stipe and 40 to 200 mg/Kg dw in cap [10]. Regarding cadmium and lead, these elements have the highest toxicological significance. Cadmium has probably been the most damaging metal found in mushrooms; some studies point out the existence of accumulating species, which, in polluted areas, accumulate this metal. The mean values of cadmium and lead were 1.00 [+ or -] 0.73 mg/kg dw and 2.41 [+ or -] 1.34 mg/kg dw, respectively, showing that the A. ponderosa had no toxicity due to the presence of these two elements [5,11]. The contents of heavy metals barium and silver in the studied mushrooms were 1.10 [+ or -] 0.59 mg/kg dw and 2.01 [+ or -] 1.56 mg/kg dw, respectively. These values are similar to those described in the literature for other species of edible mushrooms [11] and lower than those described for species of the genus Amanita [5, 13].

Bioconcentration factor (BCF) allows estimating the mushroom potential for the bioextraction of elements from the substratum (soil). Values of BCF of A. ponderosa fruiting bodies are summarized in Tables 7 and 8. The macroelements, K and Na, exhibited the highest values of BCF but Ca, Mg, and P also presented BCF > 1. Trace elements, Ag, Cd, Cu, and Zn, presented BCF > 1 with higher values for Cu. The remaining elements (Fe, Mn, Ba, Cr, and Pb) are bioexcluded showing lower values (BCF < 1). For copper, BCF values ranged from 9 to 248, showing bioaccumulation of this metal. The same heterogeneous behaviour is observed for Ag, Cd, and Zn with values ranging from 1-72, 1-38, to 2-25, respectively. Ag bioaccumulation was found in other Amanita species with much higher values of BCF [13]. The high levels of aluminium observed for some A. ponderosa fruiting bodies can be related to soil content and to their different ability to accumulate this mineral, with BCF > 1 for some stands. Some works with different species such as Leccinum scabrum, Amanita rubescens, and Xerocomus chrysenteron reported different aluminium accumulation [15]. Some species of Basidiomycetes can be useful for assessing the environmental pollution levels [38].

Metal concentrations were usually assumed to be species-dependent, but soil composition is also an important factor in mineral content [12, 36, 38, 49]. In order to clarify this association between inorganic composition of A. ponderosa mushrooms and soil, a data mining approach was developed.

3.2. Segmentation Models Based on Mushrooms and Soil Mineral Content: Interpretation and Assessment. The k-Means Clustering Method is a segmentation algorithm that uses unsupervised learning. The input variables used in the segmentation approach are macroelements content (Na, K, Ca, P, and Mg), trace elements content (Al, Fe, Cu, Zn, Cr, and Mn), and heavy metals (Ag, Ba, Cd, and Pb). The algorithm input parameter is the number of clusters, 3 in this study. Table 9 shows the clusters centre of gravity, in order to characterize the clusters formed.

The analysis of Table 9 shows that cluster 1 is characterized by high values of Ag, Ba, Cd, K, Mg, and P. Cluster 2 is characterized by high content of Cr, Fe, Mn, Na, Pb, and Zn and Cluster 3 showed lower mineral content. In order to evaluate the relationships between clusters and fruiting bodies sampling sites the graph presented Figure 4 was conceived.

The analysis of Figure 4 shows that cluster 1 is formed by the samples collected at Almendres (1), Baleizao (3), Cabeca Gorda (5), Evoramonte (8), Mina S. Domingos (11), Montejuntos (14), N. [S.sup.ra] Guadalupe (15), N. [S.sup.ra] Machede (16), Rosal de la Frontera (17), Serpa (20), and Villanueva del Fresno (24). Cluster 2 includes the samples collected at Beja (2), Evora (7), S. Miguel de Machede (19), and V. N. S. Bento (23). Finally, cluster 3 is composed by the samples collected at Azaruja (2), [Her.sup.de] da Mitra (9), Martola (10), [M.sup.te] da Borralha (12), [M.sup.te] Novo (13), [S.sup.to] Aleixo da Restauracao (18), [V.sup.e] de Rocins (21), and Valverde (22).

In order to generate an explanatory model of segmentation (i.e., seek to establish rules for assigning a new case to a cluster), Decisions Trees (DT) were used. Two different strategies were followed: one of them based on the mineral mushrooms content (strategy 1) and the other one based on the soil mineral composition (strategy 2).

To ensure statistical significance of the attained results, 25 (twenty-five) runs were applied in all tests, the accuracy estimates being achieved using the Holdout method [50]. In each simulation, the available data are randomly divided into two mutually exclusive partitions: the training set, with about 2/3 of the available data and used during the modelling phase, and the test set, with the remaining examples, being used after training, in order to compute the accuracy values.

The DT obtained using the strategy 1 is shown in Figure 5. The minerals that contribute to this explanatory model are Fe, Zn, and Ba and the rules to assign a case to a cluster are as follows:
(i) If Fe [less than or equal to] 118.76 mg/Kg
       and Zn > 51.454 mg/Kg
        and Ba > 0.523 mg/Kg
          Then [right arrow] Cluster 1
(ii) If Fe > 118.76 mg/Kg
        Then [right arrow] Cluster 2
(iii) If Fe [less than or equal to] 118.76 mg/Kg
        and Zn [less than or equal to] 51.454 mg/Kg
          Then [right arrow] Cluster 3
(iv) If Fe [less than or equal to] 118.76 mg/Kg
        and Zn > 51.454 mg/Kg
          and Ba [less than or equal to] 0.523 mg/Kg
            Then [right arrow] Cluster 3


A common tool for classification analysis is the coincidence matrix [51], a matrix of size L x L, where L denotes the number of possible classes. This matrix is created by matching the predicted and actual values. L was set to 3 (three) in the present case. Table 10 presents the coincidence matrix. The results reveal that the model accuracy is 100% both in the training set and in test set.

In order to relate the mineral mushrooms content to the soil mineral composition an explanatory model of the clusters formed was made, using the soil content of the same minerals (i.e., Fe, Zn, and Ba). Since the model accuracy is only 74% a new explanation model was built up. In this attempt all inorganic soil mineral content was available. The referred model is shown in Figure 6 and the respective coincidence matrix is presented in Table 11.

The model accuracy was 89.1% and 82.6%, respectively, for training and test sets. The soil minerals that contribute to this explanatory model are Cr, Ba, and Zn. The rules to assign the cases to each cluster are as follows:
 (i) If Cr (soil) > 3.765 mg/Kg
        Then [right arrow] Cluster 1
 (ii) If Cr (soil) [less than or equal to] 3.765 mg/Kg
        and Ba (soil) [less than or equal to] 3.012 mg/Kg
          and Zn (soil) [less than or equal to] 5.944 mg/Kg
            Then [right arrow] Cluster 1
(iii) If Cr (soil) [less than or equal to] 3.765 mg/Kg
        and Ba (soil) > 3.012 mg/Kg
          and Zn (soil) > 5.033 mg/Kg
            Then [right arrow] Cluster 2
 (iv) If Cr (soil) [less than or equal to] 3.765 mg/Kg
        and Ba (soil) [less than or equal to] 3.012 mg/Kg
          and Zn (soil) > 5.944 mg/Kg
            Then [right arrow] Cluster 3
  (v) If Cr (soil) [right arrow] 3.765 mg/Kg
        and Ba (soil) > 3.012 mg/Kg
          and Zn (soil) [less than or equal to] 5.033 mg/Kg
            Then [right arrow] Cluster 3


This study observed that the mineral content was influenced by the location area in which the mushroom samples were collected, possibly due to the soil composition and by environmental factors, such as medium temperature, vegetation, and rainfall. Indeed the inorganic composition of A. ponderosa allows group mushrooms according to the location area, based mainly in their Fe, Zn, and Ba content. On the other hand, it is possible to predict the same mushroom clustering taking into account the mineral soil content, based in Cr, Ba, and Zn soil composition although with a lower model accuracy.

Results of mineral composition do not reveal a direct correlation between inorganic composition of A. ponderosa mushrooms and their corresponding soil substrate; nevertheless, mushrooms are agents that play an important role in the continuous changes that occur in their habitats, and indeed they present a very effective mechanism for accumulating metals from the environment.

Moreno-Rojas et al. (2004), in the study of mineral content evaluation of A. ponderosa samples from Andalusia, also verified that variations occurred in the mineral composition according to the sample collection site, particularly in relation to Fe, K, and Na contents. Other authors also report that the main cause of variation of mineral composition between samples of different species of Amanita is the character and composition of the substrates (e.g., sand and wood) and may be influenced by the presence or absence of ability of the different species for a specific accumulation of some metals, namely, copper and zinc [5]. In a study carried out with a species of Boletaceae (Suillus grevillei), it is also mentioned that the variations in mineral composition of the different samples are related to the composition of the substrates and the geochemistry of the soils of each site [11].

4. Conclusions

A. ponderosa mushrooms collected from different sites showed fruiting bodies with water content of 90-93%, dry mass ranging from 6.9 to 9.7%, contents of organic matter between 6.2 and 9.0%, and minerals between 0.5 and 0.9%. Mineral composition revealed high content in macroelements, such as potassium, phosphorus, and magnesium. Copper, chromium, iron, manganese, and zinc, essential microelements in biological systems, can also be found in fruiting bodies of A. ponderosa, within the limits of RDI. Bioconcentration was observed for some macro- and microelements, such as K, Na, Cu, Zn, Mg, P, Ag, Ca, and Cd. The presence of heavy metals, such as barium, cadmium, lead and silver, was quite low, within the limits of RDI, and did not constitute a risk to human health.

Our results pointed out that it is possible to generate an explanatory model of segmentation performed with data based on the inorganic composition of mushrooms and soil mineral content, showing that it may be possible to relate these two types of data. The inorganic analysis provides evidence that mushrooms mineral composition variation is according to the collecting location, indicating the influence of the substrate characteristics in the fruiting bodies. The relationship between mineral elements in mushrooms and soils from the different sampling sites can be an important contribution to the certification process and seem to be related to the substrate effects from interindividual or interstrain differences.

https://doi.org/10.1155/2018/5265291

Conflicts of Interest

The authors declare that they have no conflicts of interest.

Acknowledgments

The authors would like to acknowledge the HIT3CH Project-HERCULES Interface for Technology Transfer and Teaming in Cultural Heritage (ALT20-03-0246-FEDER-000004), MEDUSA Project-Microorganisms Monitoring and Mitigation-Developing and Unlocking Novel Sustainable Approaches (ALT20-03-0145-FEDER-000015), cofinanced by the European Regional Development Fund (ERDF), and ALENTEJO 2020 (Alentejo Regional Operational Programme).

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Catia Salvador, (1) M. Rosario Martins, (1,2) Henrique Vicente [ID], (2,3) and A. Teresa Caldeira [ID] (1,2)

(1) HERCULES Laboratory, Evora University, Largo Marques de Marialva 8, 7000-809 Evora, Portugal

(2) Departamento de Quimica, School of Sciences and Technology, Evora University, Rua Romao Ramalho 59, 7000-671 Evora, Portugal

(3) Centro ALGORITMI, Universidade do Minho, Braga, Portugal

Correspondence should be addressed to A. Teresa Caldeira; atc@uevora.pt

Received 30 April 2017; Revised 12 November 2017; Accepted 14 December 2017; Published 31 January 2018

Academic Editor: Gunther K. Bonn

Caption: Figure 1: Geographic representation of sampling location areas of the A. ponderosa fruiting bodies of forest area of the Alentejo, Andalusia, and Extremadura regions. In detail, we observe the sampling site of Evora region (Alentejo, Portugal), showing the surrounding vegetation of Quercus suber, Cistus ladanifer and a sample of the fruiting body collected at this local.

Caption: Figure 3: Concentration of mineral content of A. ponderosa fruiting bodies and soil substrates. The values are represented in logarithmic scale.

Caption: Figure 4: Relationships between clusters of mushrooms and different sampling sites.

Caption: Figure 5: The Decision Tree explanatory of segmentation model based in mineral content of mushrooms samples (strategy 1).

Caption: Figure 6: The Decision Tree explanatory of segmentation model based in mineral content of substrates.
Table 1: Sampling sites description of A. ponderosa mushrooms and
soil substrates.

Sampling sites                  GPS coordinates       Region/country

(1) Almendres                 38[degrees]33'57"N
                               8[degrees]02'40"O

(2) Azaruja                   38[degrees]42'10"N
                               7[degrees]45'58"O

(3) Baleizao                  38[degrees]01'34"N    Alentejo, Portugal
                               7[degrees]42'38"O

(4) Beja                      38[degrees]02'44"N
                               7[degrees]50'56"O

(5) Cabeca Gorda              37[degrees]55'19"N
                               7[degrees]48'47"O

(6) Cabezas Rubias            37[degrees]43'50"N     Andaluzia, Spain
                               7[degrees]05'11"O

(7) Evora                     38[degrees]35'01"N
                               7[degrees]51'46"O

(8) Evoramonte                38[degrees]46'22"N
                               7[degrees]42'45"O

(9) [Her.sup.de] da Mitra     38[degrees]31'35"N
                               8[degrees]00'51"O

(10) Mertola                  37[degrees]37'44"N
                               7[degrees]39'30"O

(11) Mina S. Domingos         37[degrees]41'02"N    Alentejo, Portugal
                              7[degrees]28'49"O

(12) [M.sup.te] da Borralha   37[degrees]58'31"N
                               7[degrees]37'22"O

(13) [M.sup.te] Novo          38[degrees]30'39"N
                               7[degrees]43'06"O

(14) Montejuntos              38[degrees]32'24"N
                               7[degrees]19'49"O

(15) N. [S.sup.ra] Guadalupe  38[degrees]33'47"N
                               8[degrees]01'23"O

(16) N. [S.sup.ra] Machede    38[degrees]35'21"N
                               7[degrees]48'19"O

(17) Rosal de la              37[degrees]57'21"N     Andaluzia, Spain
Frontera                       7[degrees]13'12"O

(18) [S.sup.to] Aleixo da     38[degrees]04'01"N
Restauracao                    7[degrees]09'46"O

(19) S. Miguel de             38[degrees]37'34"N
Machede                        7[degrees]42'33"O

(20) Serpa                    37[degrees]55'59"N    Alentejo, Portugal
                              7[degrees]35'05"O

(21) [V.sup.e] Rocins         37[degrees]52'15"N
                               7[degrees]44'41"O

(22) Valverde                 38[degrees]32'24"N
                               8[degrees]01'18"O

(23) V. N. S. Bento           37[degrees]56'27"N
                               7[degrees]23'51"O

(24) Villanueva del            38[degrees]22'49"    Extremadura, Spain
Fresno                        N7[degrees]11'35"O

Table 2: Moisture, organic content, and minerals contents of
A. ponderosa fruiting bodies from different sampling sites.

Fruiting bodies
Sampling sites                          Moisture (%)

(1) Almendres                   93.6 [+ or -] 0.3 (a)
(2) Azaruja                     92.4 [+ or -] 0.5 (a,b,c,d)
(3) Baleizao                    90.7 [+ or -] 0.3 (b,c,d,e)
(4) Beja                        91.0 [+ or -] 0.0 (a,b,c,d,e)
(5) Cabeca Gorda                91.6 [+ or -] 0.7 (a,b,c,d,e)
(6) Cabezas Rubias              91.8 [+ or -] 0.4 (a,b,c,d,e)
(7) Evora                       90.2 [+ or -] 1.6 (c,d,e)
(8) Evoramonte                  90.4 [+ or -] 0.4 (b,c,d,e)
(9) [Her.sup.de] da Mitra       90.5 [+ or -] 0.3 (b,c,d,e)
(10) Mertola                    93.2 [+ or -] 0.3 (a,b)
(11) Mina S. Domingos           89.5 [+ or -] 0.0 (e)
(12) [M.sup.te] da Borralha     91.5 [+ or -] 1.5 (a,b,c,d,e)
(13) [M.sup.te] Novo            91.7 [+ or -] 1.2 (a,b,c,d,e)
(14) Montejuntos                92.7 [+ or -] 0.4 (a,b,c,d)
(15) N. [S.sup.ra] Guadalupe    91.4 [+ or -] 1.1 (a,b,c,d,e)
(16) N. [S.sup.ra] Machede      91.4 [+ or -] 1.5 (a,b,c,d,e)
(17) Rosal de la Frontera       90.4 [+ or -] 1.7 (b,c,d,e)
(18) [S.sup.to] Aleixo da
  Restauracao                   91.1 [+ or -] 1.1 (a,b,c,d,e)
(19) S. Miguel de Machede       92.7 [+ or -] 0.4 (a,b,c,d)
(20) Serpa                      93.8 [+ or -] 0.5 (a)
(21) [V.sup.e] Rocins           93.0 [+ or -] 0.7 (a,b,c)
(22) Valverde                   92.0 [+ or -] 1.6 (a,b,c,d,e)
(23) V. N. S. Bento             90.0 [+ or -] 0.2 (d,e)
(24) Villanueva del Fresno      93.1 [+ or -] 0.6 (a,b)

Fruiting bodies                Composition in fresh weight
Sampling sites                     Organic content (%)

(1) Almendres                    5.8 [+ or -] 0.3 (a)
(2) Azaruja                      6.7 [+ or -] 0.5 (a,b,c)
(3) Baleizao                     8.6 [+ or -] 0.4 (b,c,d)
(4) Beja                         8.4 [+ or -] 0.1 (a,b,c,d)
(5) Cabeca Gorda                 7.8 [+ or -] 0.7 (a,b,c,d)
(6) Cabezas Rubias               6.8 [+ or -] 0.1 (a,b,c)
(7) Evora                        9.2 [+ or -] 1.4 (c,d)
(8) Evoramonte                   8.8 [+ or -] 0.7 (b,c,d)
(9) [Her.sup.de] da Mitra        8.9 [+ or -] 0.3 (c,d)
(10) Mertola                     6.0 [+ or -] 0.3 (a,b)
(11) Mina S. Domingos            9.8 [+ or -] 0.1 (d)
(12) [M.sup.te] da Borralha      7.8 [+ or -] 1.5 (a,b,c,d)
(13) [M.sup.te] Novo             7.6 [+ or -] 1.1 (a,b,c,d)
(14) Montejuntos                 6.7 [+ or -] 0.4 (a,b,c)
(15) N. [S.sup.ra] Guadalupe     7.9 [+ or -] 1.3 (a,b,c,d)
(16) N. [S.sup.ra] Machede       8.0 [+ or -] 1.4 (a,b,c,d)
(17) Rosal de la Frontera        8.8 [+ or -] 1.6 (b,c,d)
(18) [S.sup.to] Aleixo da
  Restauracao                    8.1 [+ or -] 1.1 (a,b,c,d)
(19) S. Miguel de Machede        6.5 [+ or -] 0.3 (a,b,c)
(20) Serpa                       5.8 [+ or -] 0.5 (a)
(21) [V.sup.e] Rocins            6.4 [+ or -] 0.7 (a,b,c)
(22) Valverde                    7.1 [+ or -] 1.6 (a,b,c,d)
(23) V. N. S. Bento              9.1 [+ or -] 0.2 (c,d)
(24) Villanueva del Fresno       6.4 [+ or -] 0.5 (a,b,c)

Fruiting bodies
Sampling sites                       Minerals (%)

(1) Almendres                   0.6 [+ or -] 0.1 (a,b,c)
(2) Azaruja                     0.9 [+ or -] 0.0 (b,c)
(3) Baleizao                    0.7 [+ or -] 0.1 (a,b,c)
(4) Beja                        0.6 [+ or -] 0.0 (a,b,c)
(5) Cabeca Gorda                0.5 [+ or -] 0.0 (a,b)
(6) Cabezas Rubias              1.4 [+ or -] 0.1 (d)
(7) Evora                       0.7 [+ or -] 0.2 (a,b,c)
(8) Evoramonte                  0.8 [+ or -] 0.3 (a,b,c)
(9) [Her.sup.de] da Mitra       0.6 [+ or -] 0.1 (a,b,c)
(10) Mertola                    0.8 [+ or -] 0,1 (a,b,c)
(11) Mina S. Domingos           0.8 [+ or -] 0.1 (a,b,c)
(12) [M.sup.te] da Borralha     0.7 [+ or -] 0.1 (a,b,c)
(13) [M.sup.te] Novo            0.7 [+ or -] 0.1 (a,b,c)
(14) Montejuntos                0.5 [+ or -] 0.1 (a,b)
(15) N. [S.sup.ra] Guadalupe    0.7 [+ or -] 0.2 (a,b,c)
(16) N. [S.sup.ra] Machede      0.7 [+ or -] 0.2 (a,b,c)
(17) Rosal de la Frontera       0.8 [+ or -] 0.1 (a,b,c)
(18) [S.sup.to] Aleixo da
  Restauracao                   0.8 [+ or -] 0.1 (a,b,c)
(19) S. Miguel de Machede       0.7 [+ or -] 0.1 (a,b,c)
(20) Serpa                      0.5 [+ or -] 0.0 (a)
(21) [V.sup.e] Rocins           0.6 [+ or -] 0.1 (a,b,c)
(22) Valverde                   0.9 [+ or -] 0.0 (c)
(23) V. N. S. Bento             0.8 [+ or -] 0.0 (a,b,c)
(24) Villanueva del Fresno      0.5 [+ or -] 0.1 (a)

Values of each determination represents mean [+ or -] SD (n = 9).
Different letters following the values indicate significant
differences (p < 0.05).

Table 3: Ca, Mg, Na, and K mineral content of A. ponderosa fruiting
bodies and corresponding soil substrates from different sampling
sites

                                         Minerals
                                     (mg/Kg dry weight)

Sampling site             Sample             Ca

(1) Almendres               FB      757 [+ or -] 156 (a)
                            SS      106 [+ or -] 11 (a,b)
(2) Azaruja                 FB      139 [+ or -] 13 (b)
                            SS      102 [+ or -] 12 (a,b)
(3) Baleizao                FB      606 [+ or -] 61 (a)
                            SS      162 [+ or -] 11 (c,d)
(4) Beja                    FB      615 [+ or -] 1 (a)
                            SS      189 [+ or -] 14 (d,e)
(5) Cabeca Gorda            FB      554 [+ or -] 21 (a)
                            SS      108 [+ or -] 6 (a,b)
(6) Cabezas Rubias          FB      352 [+ or -] 2 (c)
                            SS      118 [+ or -] 16 (a,b,c)
(7) Evora                   FB      605 [+ or -] 1 (a)
                            SS       91 [+ or -] 23 (a)
(8) Evoramonte              FB      733 [+ or -] 15 (a)
                            SS      102 [+ or -] 8 (a,b)
(9) [Her.sup.de] da         FB      579 [+ or -] 111 (a)
  Mitra                     SS      159 [+ or -] 8 (c,d)
(10) Mertola                FB      370 [+ or -] 57 (a,b)
                            SS      112 [+ or -] 8 (a,b,c)
(11) Mina                   FB      582 [+ or -] 1 (a)
  S. Domingos               SS      143 [+ or -] 25 (b,c,d)
(12) [M.sup.te] da          FB      448 [+ or -] 33 (a,b)
  Borralha                  SS       70 [+ or -] 14 (a)
(13) [M.sup.te] Novo        FB      436 [+ or -] 72 (a,b)
                            SS      148 [+ or -] 19 (b,c,d)
(14) Montejuntos            FB      412 [+ or -] 50 (a,b)
                            SS      224 [+ or -] 16 (e,f)
(15) N. [S.sup.ra]          FB      408 [+ or -] 11 (a,b)
  Guadalupe                 SS      148 [+ or -] 8 (b,c,d)
(16) N. [S.sup.ra] de       FB      632 [+ or -] 111 (a)
  Machede                   SS      143 [+ or -] 3 (b,c,d)
(17) Rosal de la            FB      584 [+ or -] 25 (a)
  Frontera                  SS       83 [+ or -] 9 (a)
(18) [S.sup.to] Aleixo      FB      366 [+ or -] 1 (a)
  da Restauracao            SS       79 [+ or -] 8 (a)
(19) S. Miguel de           FB      649 [+ or -] 74 (a)
  Machede                   SS      338 [+ or -] 35 (a,b,c)
(20) Serpa                  FB      698 [+ or -] 148 (a,b)
                            SS      119 [+ or -] 16 (g)
(21) [V.sup.e] Rocins       FB      448 [+ or -] 269 (a,b)
                            SS      113 [+ or -] 8 (a,b,c)
(22) Valverde               FB      521 [+ or -] 146 (a,b)
                            SS       93 [+ or -] 23 (a)
(23) V. N. S. Bento         FB      599 [+ or -] 1 (a)
                            SS      233 [+ or -] 11 (e,f)
(24) Villanueva del         FB      459 [+ or -] 13 (a,b)
  Fresno                    SS      240 [+ or -] 26 (f)

Total                       FB      523 [+ or -] 143
                            SS      143 [+ or -] 63

                                               Minerals
                                          (mg/Kg dry weight)

Sampling site             Sample                   K

(1) Almendres               FB       26677 [+ or -] 915 (a,b,c,d,e)
                            SS         363 [+ or -] 9 (a,b)
(2) Azaruja                 FB        4589 [+ or -] 103 (a)
                            SS         981 [+ or -] 37 (g)
(3) Baleizao                FB       25758 [+ or -] 4728 (a,b,c,d,e)
                            SS        1099 [+ or -] 24 (h)
(4) Beja                    FB        6090 [+ or -] 34 (a,b)
                            SS         619 [+ or -] 19 (d,e)
(5) Cabeca Gorda            FB       38652 [+ or -] 1607 (c,d,e)
                            SS         384 [+ or -] 16 (a,b,c)
(6) Cabezas Rubias          FB       69565 [+ or -] 362 (f)
                            SS         443 [+ or -] 9 (c)
(7) Evora                   FB       34630 [+ or -] 186 (c,d,e)
                            SS         651 [+ or -] 9 (e)
(8) Evoramonte              FB       25148 [+ or -] 4556 (a,b,c,d,e)
                            SS         576 [+ or -] 16 (d)
(9) [Her.sup.de] da         FB       31852 [+ or -] 5364 (a,b,c,d,e)
  Mitra                     SS         229 [+ or -] 9 (j,k)
(10) Mertola                FB       18021 [+ or -] 1806 (a,b,c)
                            SS         731 [+ or -] 19 (f)
(11) Mina                   FB       33607 [+ or -] 173 (b,c,d,e)
  S. Domingos               SS         256 [+ or -] 16 (y)
(12) [M.sup.te] da          FB       26099 [+ or -] 1618 (a,b,c,d,e)
  Borralha                  SS        155 [+ or -] 91
(13) [M.sup.te] Novo        FB      25763 [+ or -] 1608 (a,b,c,d,e)
                            SS        603 [+ or -] 56 (d,e)
(14) Montejuntos            FB      41285 [+ or -] 982 (c,d,e)
                            SS        256 [+ or -] 16 (i,j)
(15) N. [S.sup.ra]          FB      34814 [+ or -] 1763 (c,d,e)
  Guadalupe                 SS        656 [+ or -] 16 (e)
(16) N. [S.sup.ra] de       FB      24717 [+ or -] 1881 (a,b,c,d,e)
  Machede                   SS        912 [+ or -] 48 (g)
(17) Rosal de la            FB      51996 [+ or -] 2660 (e,f)
  Frontera                  SS        160 [+ or -] 16 (k,l)
(18) [S.sup.to] Aleixo      FB      18586 [+ or -] 106 (d,e,f)
  da Restauracao            SS        773 [+ or -] 24 (f)
(19) S. Miguel de           FB      45011 [+ or -] 1152 (c,d,e,f)
  Machede                   SS        315 [+ or -] 9 (k,l)
(20) Serpa                  FB      47715 [+ or -] 5220 (a,b,c)
                            SS        171 [+ or -] 9 (a,l)
(21) [V.sup.e] Rocins       FB      21788 [+ or -] 1237 (a,b,c,d)
                            SS        320 [+ or -] 16 (a,l)
(22) Valverde               FB      27256 [+ or -] 2180 (a,b,c,d,e)
                            SS        405 [+ or -] 24 (b,c)
(23) V. N. S. Bento         FB       6228 [+ or -] 30 (a,b)
                            SS        192 [+ or -] 16 (j,k,l)
(24) Villanueva del         FB      25713 [+ or -] 5847 (a,b,c,d,e)
  Fresno                    SS        368 [+ or -] 16 (a,b)

Total                       FB      29648 [+ or -] 14908
                            SS        484 [+ or -] 273

                                             Minerals
                                        (mg/Kg dry weight)

Sampling site             Sample                Mg

(1) Almendres               FB       774 [+ or -] 130 (a,b,c)
                            SS       286 [+ or -] 14 (a,b,c,d)
(2) Azaruja                 FB       194 [+ or -] 10 (a)
                            SS       324 [+ or -] 29 (a,b)
(3) Baleizao                FB       856 [+ or -] 107 (a,b,c,d)
                            SS       312 [+ or -] 38 (a,b)
(4) Beja                    FB       734 [+ or -] 1 (a,b,c)
                            SS       277 [+ or -] 33 (a,b,c,d,e)
(5) Cabeca Gorda            FB       848 [+ or -] 159 (a,b,c)
                            SS       186 [+ or -] 8 (e,f,g)
(6) Cabezas Rubias          FB      1265 [+ or -] 1 (c,d)
                            SS       277 [+ or -] 19 (a,b,c,d,e)
(7) Evora                   FB       713 [+ or -] 1 (a,b,c)
                            SS       171 [+ or -] 25 (f,g)
(8) Evoramonte              FB       760 [+ or -] 101 (a,b,c)
                            SS       156 [+ or -] 11 (g)
(9) [Her.sup.de] da         FB       685 [+ or -] 89 (a,b,c)
  Mitra                     SS       298 [+ or -] 29 (a,b,c)
(10) Mertola                FB       658 [+ or -] 33 (a,b,c)
                            SS       193 [+ or -] 18 (d,e,f,g)
(11) Mina                   FB       737 [+ or -] 1 (a,b,c)
  S. Domingos               SS       365 [+ or -] 43 (a)
(12) [M.sup.te] da          FB       696 [+ or -] 29 (a,b,c)
  Borralha                  SS       172 [+ or -] 23 (f,g)
(13) [M.sup.te] Novo        FB       562 [+ or -] 114 (a,b)
                            SS       217 [+ or -] 34 (c,d,e,f,g)
(14) Montejuntos            FB       742 [+ or -] 53 (a,b,c)
                            SS       910 [+ or -] 25 (j)
(15) N. [S.sup.ra]          FB       930 [+ or -] 158 (b,c,d)
  Guadalupe                 SS       255 [+ or -] 14 (b,c,d,e,f)
(16) N. [S.sup.ra] de       FB       843 [+ or -] 120 (a,b,c)
  Machede                   SS       804 [+ or -] 48 (i)
(17) Rosal de la            FB      1540 [+ or -] 77 (d)
  Frontera                  SS       365 [+ or -] 11 (a)
(18) [S.sup.to] Aleixo      FB       481 [+ or -] 1 (a,b,c)
  da Restauracao            SS       477 [+ or -] 14 (h)
(19) S. Miguel de           FB       549 [+ or -] 140 (a,b)
  Machede                   SS      1466 [+ or -] 68 (a,b,c,d,e)
(20) Serpa                  FB       736 [+ or -] 86 (a,b)
                            SS       272 [+ or -] 15 (k)
(21) [V.sup.e] Rocins       FB       418 [+ or -] 203 (a,b)
                            SS       284 [+ or -] 14 (a,b,c,d)
(22) Valverde               FB       586 [+ or -] 35 (a,b,c)
                            SS       265 [+ or -] 41 (b,c,d,e,f)
(23) V. N. S. Bento         FB       657 [+ or -] 1 (a,b,c)
                            SS       536 [+ or -] 37 (h)
(24) Villanueva del         FB       738 [+ or -] 49 (a,b,c)
  Fresno                    SS       267 [+ or -] 27 (b,c,d,e)

Total                       FB       738 [+ or -] 261
                            SS       381 [+ or -] 296

                                              Minerals
                                         (mg/Kg dry weight)

Sampling site             Sample                 Na

(1) Almendres               FB         1408 [+ or -] 146 (a)
                            SS           70 [+ or -] 1 (a,b,c,d)
(2) Azaruja                 FB          216 [+ or -] 135 (f)
                            SS           75 [+ or -] 4 (a,b,h,i)
(3) Baleizao                FB         2249 [+ or -] 231 (g,h)
                            SS          109 [+ or -] 4 (k)
(4) Beja                    FB         1540 [+ or -] 17 (a,g)
                            SS           69 [+ or -] 2 (a,b,c,d,e)
(5) Cabeca Gorda            FB          889 [+ or -] 38 (a,b,c,d,e,f)
                            SS           69 [+ or -] 5 (a,b,c,d,e)
(6) Cabezas Rubias          FB         2645 [+ or -] 18 (h)
                            SS           62 [+ or -] 2 (c,d,e,f)
(7) Evora                   FB         1241 [+ or -] 16 (a,b)
                            SS           52 [+ or -] 3 (f,g)
(8) Evoramonte              FB          614 [+ or -] 131 (b,c,d,e,f)
                            SS           51 [+ or -] 2 (g)
(9) [Her.sup.de] da         FB          930 [+ or -] 58 (a,b,c,d,e,f)
  Mitra                     SS           51 [+ or -] 3 (g)
(10) Mertola                FB          484 [+ or -] 171 (c,d,e,f)
                            SS           82 [+ or -] 2h' (i,j)
(11) Mina                   FB         1097 [+ or -] 14 (a,b,c,d)
  S. Domingos               SS           59 [+ or -] 5 (e,f,g)
(12) [M.sup.te] da          FB          368 [+ or -] 70 (e,f)
  Borralha                  SS          67 [+ or -] 5 (b,c,d,e)
(13) [M.sup.te] Novo        FB         379 [+ or -] 25 (e,f)
                            SS          53 [+ or -] 4 (f,g)
(14) Montejuntos            FB        1146 [+ or -] 304 (a,b,c)
                            SS          85 [+ or -] 2 (i,j)
(15) N. [S.sup.ra]          FB         551 [+ or -] 52 (b,c,d,e,f)
  Guadalupe                 SS          73 [+ or -] 1 (a,b,h)
(16) N. [S.sup.ra] de       FB        2185 [+ or -] 438 (g,h)
  Machede                   SS          80 [+ or -] 2 (a,h,i,j)
(17) Rosal de la            FB         964 [+ or -] 453 (a,b,c,d,e)
  Frontera                  SS          80 [+ or -] 1 (a,h,i,j)
(18) [S.sup.to] Aleixo      FB        1098 [+ or -] 103 (a,b,c,d)
  da Restauracao            SS          70 [+ or -] 2 (a,b,c)
(19) S. Miguel de           FB         897 [+ or -] 11 (a,b,c,d,e,f)
  Machede                   SS         158 [+ or -] 5 (c,d,e,f,g)
(20) Serpa                  FB        1078 [+ or -] 174 (a,b,c,d,e)
                            SS          60 [+ or -] 5 (l)
(21) [V.sup.e] Rocins       FB         419 [+ or -] 212 (d,e,f)
                            SS          86 [+ or -] 2 (j)
(22) Valverde               FB         895 [+ or -] 65 (a,b,c,d,e,f)
                            SS         111 [+ or -] 3 (k)
(23) V. N. S. Bento         FB        1467 [+ or -] 15 (a)
                            SS          60 [+ or -] 2 (d,e,f,g)
(24) Villanueva del         FB        1451 [+ or -] 147 (a)
  Fresno                    SS          78 [+ or -] 4 (a,h,i,j)

Total                       FB        1092 [+ or -] 620
                            SS          75 [+ or -] 24

FB, fruiting bodies; SS, soil substrate. Mean values
(n = 9) [+ or -] SD. Different letters for each element indicate
significant differences with the confidence level of
p < 0.05 (ANOVA, Tukey's test).

Table 4: Al, Cu, Fe, and P mineral content of A. ponderosa fruiting
bodies and corresponding soil substrates from different
sampling sites.

                                         Minerals (mg/Kg dry weight)

Sampling site                  Sample                Al

(1) Almendres                    FB      349 [+ or -] 124 (b,c,d,e,f)
                                 SS      277 [+ or -] 8 (a,b)
(2) Azaruja                      FB       58 [+ or -] 8 (a)
                                 SS      243 [+ or -] 30 (a)
(3) Baleizao                     FB      357 [+ or -] 29 (b,c,d,e,f)
                                 SS      397 [+ or -] 4 (c,d,e)
(4) Beja                         FB      449 [+ or -] 13 (d,e,f,g)
                                 SS      254 [+ or -] 15 (a)
(5) Cabeca Gorda                 FB      151 [+ or -] 25 (a,b)
                                 SS      196 [+ or -] 4 (k)
(6) Cabezas Rubias               FB      932 [+ or -] 77 (h)
                                 SS      375 [+ or -] 8 (c,d)
(7) Evora                        FB      208 [+ or -] 11 (a,b,c,d)
                                 SS      368 [+ or -] 18 (c)
(8) Evoramonte                   FB      423 [+ or -] 49 (c,d,e,f,g)
                                 SS      311 [+ or -] 6 (b)
(9) [Her.sup.de] da Mitra        FB      373 [+ or -] 26 (b,c,d,e,f)
                                 SS      157 [+ or -] 6 (1)
(10) Mertola                     FB      333 [+ or -] 36 (b,c,d,e)
                                 SS      160 [+ or -] 81
(11) Mina S. Domingos            FB      299 [+ or -] 14 (a,b,c,d,e)
                                 SS      417 [+ or -] 6 (e,f)
(12) [M.sup.te] da Borralha      FB      209 [+ or -] 42 (a,b,c,d)
                                 SS      159 [+ or -] 6 (1)
(13) [M.sup.te] Novo             FB      423 [+ or -] 59 (c,d,e,f,g)
                                 SS      300 [+ or -] 15 (b)
(14) Montejuntos                 FB      545 [+ or -] 99 (e,f,g)
                                 SS      813 [+ or -] 11 (j)
(15) N. [S.sup.ra] Guadalupe     FB      209 [+ or -] 73 (a,b,c,d)
                                 SS      200 [+ or -] 6 (k)
(16) N. [S.sup.ra]               FB      313 [+ or -] 47 (a,b,c,d,e)
  de Machede                     SS      632 [+ or -] 19 (1)
(17) Rosal de la Frontera        FB      659 [+ or -] 36 (g)
                                 SS      381 [+ or -] 4 (c,d)
(18) [S.sup.to] Aleixo da        FB       65 [+ or -] 12 (d,e,f,g)
  Restauracao                    SS      449 [+ or -] 16 (f,g)
(19) S. Miguel de Machede        FB      610 [+ or -] 183 (f,g)
                                 SS      497 [+ or -] 6 (c,d)
(20) Serpa                       FB      453 [+ or -] 66 (a)
                                 SS      378 [+ or -] 4 (h)
(21) [V.sup.e] Rocins            FB      181 [+ or -] 24 (a,b,c)
                                 SS      391 [+ or -] 9 (c,d,e)
(22) Valverde                    FB      183 [+ or -] 54 (a,b,c)
                                 SS      453 [+ or -] 6 (g)
(23) V. N. S. Bento              FB      303 [+ or -] 7 (a,b,c,d,e)
                                 SS      409 [+ or -] 6 (d,e)
(24) Villanueva del Fresno       FB      602 [+ or -] 66 (f,g)
                                 SS      494 [+ or -] 4 (h)

Total                            FB      362 [+ or -] 204
                                 SS      363 [+ or -] 155

                                               Minerals
                                           (mg/Kg dry weight)

Sampling site                  Sample              Cu

(1) Almendres                    FB       80 [+ or -] 1 (a,b)
                                 SS        1 [+ or -] 0 (a)
(2) Azaruja                      FB       30 [+ or -] 2 (a)
                                 SS        2 [+ or -] 0 (c)
(3) Baleizao                     FB      356 [+ or -] 37 (a,b,c)
                                 SS        7 [+ or -] 0 (k)
(4) Beja                         FB      373 [+ or -] 1 (b,c)
                                 SS        9 [+ or -] 1 (1)
(5) Cabeca Gorda                 FB      161 [+ or -] 10 (a,b)
                                 SS        1 [+ or -] 0 (a)
(6) Cabezas Rubias               FB      140 [+ or -] 1 (a,b)
                                 SS        5 [+ or -] 0 (g)
(7) Evora                        FB      198 [+ or -] 1 (a,b)
                                 SS        2 [+ or -] 0 (b,c)
(8) Evoramonte                   FB      124 [+ or -] 20 (a,b)
                                 SS        2 [+ or -] 0 (c,d)
(9) [Her.sup.de] da Mitra        FB      193 [+ or -] 19 (a,b)
                                 SS       10 [+ or -] 0 (m)
(10) Mertola                     FB       43 [+ or -] 15 (a,b)
                                 SS        5 [+ or -] 0 (f,g)
(11) Mina S. Domingos            FB      334 [+ or -] 1 (a,b,c)
                                 SS        3 [+ or -] 0 (d,e)
(12) [M.sup.te] da Borralha      FB      232 [+ or -] 63 (a,b)
                                 SS        2 [+ or -] 0 (c)
(13) [M.sup.te] Novo             FB      121 [+ or -] 16 (a,b)
                                 SS        3 [+ or -] 0 (c,d)
(14) Montejuntos                 FB      170 [+ or -] 21 (a,b)
                                 SS        7 [+ or -] 0 (j,k)
(15) N. [S.sup.ra] Guadalupe     FB      264 [+ or -] 23 (a,b,c)
                                 SS        8 [+ or -] 0 (1)
(16) N. [S.sup.ra]               FB      148 [+ or -] 26 (a,b)
  de Machede                     SS        4 [+ or -] 0 (e,f)
(17) Rosal de la Frontera        FB      584 [+ or -] 51 (c)
                                 SS        9 [+ or -] 0 (1)
(18) [S.sup.to] Aleixo da        FB      129 [+ or -] 1 (a,b)
  Restauracao                    SS        6 [+ or -] 0 (g,h,i)
(19) S. Miguel de Machede        FB      207 [+ or -] 39 (a,b)
                                 SS        5 [+ or -] 0 (g,h)
(20) Serpa                       FB      143 [+ or -] 14 (a,b)
                                 SS        5 [+ or -] 0 (f,g)
(21) [V.sup.e] Rocins            FB       84 [+ or -] 7 (a,b)
                                 SS        9 [+ or -] 0 (1)
(22) Valverde                    FB       62 [+ or -] 3 (a,b)
                                 SS        1 [+ or -] 0 (a,b)
(23) V. N. S. Bento              FB      172 [+ or -] 1 (a,b)
                                 SS        6 [+ or -] 0 (h,i,j)
(24) Villanueva del Fresno       FB      100 [+ or -] 9 (a,b)
                                 SS        6 [+ or -] 0 (i,j)

Total                            FB      185 [+ or -] 125
                                 SS        5 [+ or -] 3

                                                 Minerals
                                            (mg/Kg dry weight)

Sampling site                  Sample               Fe

(1) Almendres                    FB          66 [+ or -] 14 (a,b,c)
                                 SS        1547 [+ or -] 92 (a)
(2) Azaruja                      FB          18 [+ or -] 4 (a)
                                 SS        2240 [+ or -] 160 (b)
(3) Baleizao                     FB          27 [+ or -] 8 (a)
                                 SS        3093 [+ or -] 92 (d,e,f)
(4) Beja                         FB         227 [+ or -] 1 (f,g)
                                 SS        4107 [+ or -] 92 (i,j)
(5) Cabeca Gorda                 FB          51 [+ or -] 12 (a,b,c)
                                 SS        5067 [+ or -] 244 (k)
(6) Cabezas Rubias               FB         193 [+ or -] 1 (e,f)
                                 SS        3520 [+ or -] 160 (e,f,g,h)
(7) Evora                        FB         292 [+ or -] 1 (g,h)
                                 SS        1653 [+ or -] 92 (a)
(8) Evoramonte                   FB          62 [+ or -] 7 (a,b,c)
                                 SS        2400 [+ or -] 160 (b,c)
(9) [Her.sup.de] da Mitra        FB          35 [+ or -] 10 (a,b)
                                 SS        4747 [+ or -] 92 (k)
(10) Mertola                     FB          22 [+ or -] 3 (a)
                                 SS        4853 [+ or -] 244 (k)
(11) Mina S. Domingos            FB          62 [+ or -] 1 (a,b,c)
                                 SS        3573 [+ or -] 92 (f,g,h)
(12) [M.sup.te] da Borralha      FB          41 [+ or -] 38 (a,b)
                                 SS        3840 [+ or -] 160 (h,i)
(13) [M.sup.te] Novo             FB        29.3 [+ or -] 1 (a)
                                 SS        1653 [+ or -] 244 (a)
(14) Montejuntos                 FB          62 [+ or -] 10 (a,b,c)
                                 SS        8099 [+ or -] 251 (1)
(15) N. [S.sup.ra] Guadalupe     FB         110 [+ or -] 27 (b,c,d)
                                 SS        4960 [+ or -] 160 (k)
(16) N. [S.sup.ra]               FB          48 [+ or -] 13 (a,b,c)
  de Machede                     SS       3738 [+ or -] 258 (g,h,i)
(17) Rosal de la Frontera        FB        119 [+ or -] 97 (c,d,e)
                                 SS       3360 [+ or -] 160 (e,f,g,h)
(18) [S.sup.to] Aleixo da        FB         51 [+ or -] 1 (a,b,c)
  Restauracao                    SS       2827 [+ or -] 92 (c,d)
(19) S. Miguel de Machede        FB        311 [+ or -] 10 (h)
                                 SS       9040 [+ or -] 139 (d,e,f,g)
(20) Serpa                       FB         43 [+ or -] 18 (a,b,c)
                                 SS       3253 [+ or -] 92 (m)
(21) [V.sup.e] Rocins            FB         36 [+ or -] 24 (a,b)
                                 SS       3040 [+ or -] 160 (d,e)
(22) Valverde                    FB         57 [+ or -] 21 (a,b,c)
                                 SS       1440 [+ or -] 160 (a)
(23) V. N. S. Bento              FB        182 [+ or -] 1 (d,e,f)
                                 SS       4587 [+ or -] 244 (j,k)
(24) Villanueva del Fresno       FB         53 [+ or -] 17 (a,b,c)
                                 SS       2347 [+ or -] 92 (b,c)

Total                            FB         92 [+ or -] 85
                                 SS       3708 [+ or -] 1870

                                                  Minerals
                                              (mg/Kg dry weight)

Sampling site                  Sample                 P

(1) Almendres                    FB       319 [+ or -] 37 (a,b,c,d,e)
                                 SS        72 [+ or -] 2 (a,b)
(2) Azaruja                      FB        44 [+ or -] 18 (h,i)
                                 SS        74 [+ or -] 4 (a,b)
(3) Baleizao                     FB       525 [+ or -] 62 (j,k)
                                 SS       156 [+ or -] 4 (g)
(4) Beja                         FB       212 [+ or -] 1 (d,e,f,g)
                                 SS       133 [+ or -] 8 (f)
(5) Cabeca Gorda                 FB       447 [+ or -] 2 (a,j)
                                 SS       130 [+ or -] 3 (e,f)
(6) Cabezas Rubias               FB        73 [+ or -] 1 (g,h,i)
                                 SS        93 [+ or -] 4 (c)
(7) Evora                        FB       201 [+ or -] 1 (d,e,f,g,h)
                                 SS        82 [+ or -] 5 (b,c)
(8) Evoramonte                   FB       469 [+ or -] 83 (a,j)
                                 SS        94 [+ or -] 1 (c)
(9) [Her.sup.de] da Mitra        FB       277 [+ or -] 90 (b,c,d,e,f)
                                 SS       115 [+ or -] 3 (d,e)
(10) Mertola                     FB        61 [+ or -] 21 (g,h,i)
                                 SS       119 [+ or -] 2 (d,e,f)
(11) Mina S. Domingos            FB       149 [+ or -] 1 (f,g,h,i)
                                 SS        74 [+ or -] 2 (a,b)
(12) [M.sup.te] da Borralha      FB       417 [+ or -] 65 (a,b,c,j)
                                 SS        75 [+ or -] 5 (a,b)
(13) [M.sup.te] Novo             FB       316 [+ or -] 76 (a,b,c,d,e,f)
                                 SS        84 [+ or -] 4 (b,c)
(14) Montejuntos                 FB       442 [+ or -] 2 (a,b,j)
                                 SS       294 [+ or -] 11 (i)
(15) N. [S.sup.ra] Guadalupe     FB       344 [+ or -] 74 (a,b,c,d)
                                 SS       131 [+ or -] 5 (e,f)
(16) N. [S.sup.ra]               FB       542 [+ or -] 48 (j,k)
  de Machede                     SS        78 [+ or -] 1 (a,b,c)
(17) Rosal de la Frontera        FB       641 [+ or -] 98 (k)
                                 SS       154 [+ or -] 4 (g)
(18) [S.sup.to] Aleixo da        FB        34 [+ or -] 1 (a,j)
  Restauracao                    SS       185 [+ or -] 1 (h)
(19) S. Miguel de Machede        FB       270 [+ or -] 60 (c,d,ef)
                                 SS       111 [+ or -]2 (h)
(20) Serpa                       FB       460 [+ or -] 16 (i)
                                 SS       179 [+ or -] 7 (d)
(21) [V.sup.e] Rocins            FB       165 [+ or -] 85 (e,f,g,h,i)
                                 SS        75 [+ or -] 5 (a,b)
(22) Valverde                    FB       152 [+ or -] 34 (f,g,h,i)
                                 SS        65 [+ or -] 2 (a)
(23) V. N. S. Bento              FB       197 [+ or -] 1 (d,e,f,g,h,i)
                                 SS       116 [+ or -] 5 (d,e)
(24) Villanueva del Fresno       FB       310 [+ or -] 83 (a,b,c,d,e,f)
                                 SS       179 [+ or -] 12 (h)

Total                            FB       294 [+ or -] 171
                                 SS       120 [+ or -] 53

FB, fruiting bodies; SS, soil substrate. Mean values
(n = 9) [+ or -] SD. Different letters for each element indicate
significant differences with the confidence level of
p < 0.05 (ANOVA, Tukey's test).

Table 5: Ag, Ba, Cd, and Cr mineral content of A. ponderosa fruiting
bodies and corresponding soil substrates from different sampling
sites.

                                       Minerals (mg/kg dry weight)

Sampling site               Sample                  Ag

(1) Almendres                 FB        2.6 [+ or -] 0.5 (a,b,c,d,e)
                              SS        0.1 [+ or -] 0.0 (a)
(2) Azaruja                   FB        6.6 [+ or -] 2.2 (f)
                              SS        0.1 [+ or -] 0.0 (a, b,c,d)
(3) Baleizao                  FB        5.1 [+ or -] 1.8 (a, b,c,d)
                              SS        0.1 [+ or -] 0.0 (a, b,c,d)
(4) Beja                      FB        1.1 [+ or -] 0.2 (a,b)
                              SS        0.1 [+ or -] 0.0 (a, b,c,d,e,f)
(5) Cabeca Gorda              FB        2.1 [+ or -] 0.7 (a, b,c)
                              SS        0.2 [+ or -] 0.0 (b,c,d,e,f)
(6) Cabezas Rubias            FB        1.5 [+ or -] 0.2 (a, b,c)
                              SS        0.2 [+ or -] 0.0 (c,d,e,f,g)
(7) Evora                     FB        0.5 [+ or -] 0.3 (a, b,c,d)
                              SS        0.2 [+ or -] 0.0 (g,h,i)
(8) Evoramonte                FB        1.0 [+ or -] 0.1 (d,e,f)
                              SS        0.1 [+ or -] 0.0 (a, b,c,d,e,f)
(9) [Her.sup.de] da Mitra     FB        0.7 [+ or -] 0.1 (a, b,c,d)
                              SS        0.2 [+ or -] 0.0 (e,f,g,h)
(10) Mertola                  FB        4.3 [+ or -] 0.8 (a,b)
                              SS        0.1 [+ or -] 0.0 (a,b)
(11) Mina S. Domingos         FB        0.6 [+ or -] 0.1 (a, b,c)
                              SS        0.3 [+ or -] 0.0 (i)
(12) [M.sup.te] da            FB        1.5 [+ or -] 0.1 (a, b,c)
  Borralha                    SS        0.2 [+ or -] 0.0 (d,e,f,g)
(13) [M.sup.te] Novo          FB        0.6 [+ or -] 0.1 (a)
                              SS        0.1 [+ or -] 0.0 (a, b,c)
(14) Montejuntos              FB        1.5 [+ or -] 0.2 (a, b,c)
                              SS        0.2 [+ or -] 0.1 (h,i)
(15) N. [S.sup.ra]            FB        2.5 [+ or -] 0.3 (e,f)
  Guadalupe                   SS        0.2 [+ or -] 0.0 (b,c,d,e,f,g)
(16) N. [S.sup.ra]            FB        0.4 [+ or -] 0.2 (a,b,c,d,e)
  Machede                     SS        0.4 [+ or -] 0.0 (j)
(17) Rosal de la Frontera     FB        1.6 [+ or -] 0.3 (c,d,e)
                              SS        0.2 [+ or -] 0.0 (c,d,e,f,g)
(18) [S.sup.to] Aleixo da     FB        1.3 [+ or -] 0.3 (a, b,c)
  Restauracao                 SS        0.2 [+ or -] 0.0 (e,f,g,h)
(19) S. Miguel de Machede     FB        2.6 [+ or -] 0.3 (b,c,d,e)
                              SS        0.2 [+ or -] 0.0 (e,f,g)
(20) Serpa                    FB        1.7 [+ or -] 0.3 (a, b,c)
                              SS        0.2 [+ or -] 0.0 (f,g,h)
(21) [V.sup.e] Rocins         FB        3.3 [+ or -] 2.2 (a)
                              SS        0.1 [+ or -] 0.0 (a,b,c,d,e)
(22) Valverde                 FB        1.8 [+ or -] 0.6 (a,b)
                              SS        0.2 [+ or -] 0.0 (c,d,e,f,g)
(23) V. N. S. Bento           FB        0.4 [+ or -] 0.3 (a, b,c)
                              SS        0.1 [+ or -] 0.0 (a, b,c,d,e,f)
(24) Villanueva del           FB        3.0 [+ or -] 0.3 (a)
  Fresno                      SS        0.1 [+ or -] 0.0 (a,b,c,d,e)

Total                         FB        2.0 [+ or -] 1.6
                              SS        0.2 [+ or -] 0.1

                                       Minerals (mg/kg dry weight)

Sampling site               Sample                  Ba

(1) Almendres                 FB        2.9 [+ or -] 1.1 (a)
                              SS        2.0 [+ or -] 0.3 (a,b)
(2) Azaruja                   FB        0.9 [+ or -] 0.2 (d,e,f,g)
                              SS        1.6 [+ or -] 0.2 (a)
(3) Baleizao                  FB        0.6 [+ or -] 0.3 (d,e,f,g)
                              SS        5.3 [+ or -] 0.9 (i)
(4) Beja                      FB        0.8 [+ or -] 0.0 (b,c,d,e,f,g)
                              SS        2.6 [+ or -] 0.7 (a, b,c,d)
(5) Cabeca Gorda              FB        0.3 [+ or -] 0.0 (d,e,f,g)
                              SS        2.3 [+ or -] 0.1 (a, b,c,d)
(6) Cabezas Rubias            FB        2.0 [+ or -] 0.3 (d,e,f,g)
                              SS        2.6 [+ or -] 0.4 (a, b,c,d)
(7) Evora                     FB        0.8 [+ or -] 0.4 (f,g)
                              SS        4.3 [+ or -] 1.1 (f,g,h,i)
(8) Evoramonte                FB        0.9 [+ or -] 0.1 (b,c,d,e)
                              SS        2.8 [+ or -] 0.1 (a,b,c,d,e)
(9) [Her.sup.de] da Mitra     FB        1.2 [+ or -] 0.1 (g)
                              SS        2.3 [+ or -] 0.1 (a, b,c)
(10) Mertola                  FB        1.5 [+ or -] 0.2 (b,c,d,e,f)
                              SS        3.0 [+ or -] 0.4 (b,c,d,e,f)
(11) Mina S. Domingos         FB        0.6 [+ or -] 0.0 (c,d,e,f,g)
                              SS        1.9 [+ or -] 0.0 (a,b)
(12) [M.sup.te] da            FB        0.9 [+ or -] 0.1 (f,g)
  Borralha                    SS        2.3 [+ or -] 0.0 (a, b,c,d)
(13) [M.sup.te] Novo          FB        1.3 [+ or -] 0.2 (e,f,g)
                              SS        3.4 [+ or -] 0.2 (c,d,e,f,g)
(14) Montejuntos              FB        1.1 [+ or -] 0.1 (b,c,d)
                              SS        3.6 [+ or -] 0.3 (d,e,f,g,h)
(15) N. [S.sup.ra]            FB        0.7 [+ or -] 0.2 (d,e,f,g)
  Guadalupe                   SS        2.4 [+ or -] 0.1 (a, b,c,d)
(16) N. [S.sup.ra]            FB        0.6 [+ or -] 0.1 (b,c,d,e,f,g)
  Machede                     SS        4.0 [+ or -] 0.2 (e,f,g,h,i)
(17) Rosal de la Frontera     FB        1.5 [+ or -] 0.1 (b,c,d,e,f)
                              SS        2.3 [+ or -] 0.1 (a, b,c,d)
(18) [S.sup.to] Aleixo da     FB        1.1 [+ or -] 0.1 (d,e,f,g)
  Restauracao                 SS        2.8 [+ or -] 0.2 (a,b,c,d,e)
(19) S. Miguel de Machede     FB        1.3 [+ or -] 0.3 (b,c,d,e,f,g)
                              SS        3.4 [+ or -] 0.1 (c,d,e,f,g)
(20) Serpa                    FB        0.4 [+ or -] 0.1 (a, b,c)
                              SS        2.4 [+ or -] 0.3 (a, b,c,d)
(21) [V.sup.e] Rocins         FB        1.3 [+ or -] 0.5 (d,e,f,g)
                              SS        4.8 [+ or -] 0.2 (h,i)
(22) Valverde                 FB        0.5 [+ or -] 0.0 (d,e,f,g)
                              SS        1.7 [+ or -] 0.3 (a,b)
(23) V. N. S. Bento           FB        2.1 [+ or -] 0.2 (c,d,e,f,g)
                              SS        4.5 [+ or -] 0.4 (g,h,i)
(24) Villanueva del           FB        1.1 [+ or -] 0.2 (a,b)
  Fresno                      SS       11.3 [+ or -] 1.0 (j)

Total                         FB        1.1 [+ or -] 0.6
                              SS        3.3 [+ or -] 2.0

                                     Minerals (mg/kg dry weight)

Sampling site               Sample               Cd

(1) Almendres                 FB        2.2 [+ or -] 0.8 (a)
                              SS        0.3 [+ or -] 0.0 (a,b)
(2) Azaruja                   FB        0.3 [+ or -] 0.0 (b)
                              SS        0.3 [+ or -] 0.1 (a)
(3) Baleizao                  FB        1.1 [+ or -] 0.3 (b,c,d,e)
                              SS        0.2 [+ or -] 0.0 (e)
(4) Beja                      FB        0.4 [+ or -] 0.0 (b,c,d,e)
                              SS        0.1 [+ or -] 0.0 (f)
(5) Cabeca Gorda              FB        0.7 [+ or -] 0.1 (c,d,e)
                              SS        0.2 [+ or -] 0.0 (c,d,e)
(6) Cabezas Rubias            FB        0.8 [+ or -] 0.1 (c,d,e)
                              SS        0.1 [+ or -] 0.0 (e,f)
(7) Evora                     FB        0.4 [+ or -] 0.3 (b,c,d,e)
                              SS        0.1 [+ or -] 0.0 (e,f)
(8) Evoramonte                FB        1.2 [+ or -] 0.2 (b,c,d,e)
                              SS        0.1 [+ or -] 0.0 (e,f)
(9) [Her.sup.de] da Mitra     FB        1.0 [+ or -] 0.0 (b,c,d,e)
                              SS        0.1 [+ or -] 0.0 (e,f)
(10) Mertola                  FB        1.0 [+ or -] 0.3 (b,c,d)
                              SS        0.1 [+ or -] 0.0 (e,f)
(11) Mina S. Domingos         FB        1.4 [+ or -] 0.2 (d,e)
                              SS        0.1 [+ or -] 0.0 (e,f)
(12) [M.sup.te] da            FB       1.1 [+ or -] 0.2 (b,c)
  Borralha                    SS       0.1 [+ or -] 0.0 (e,f)
(13) [M.sup.te] Novo          FB       0.6 [+ or -] 0.1 (b,c,d,e)
                              SS       0.2 [+ or -] 0.0 (b,c,d,e)
(14) Montejuntos              FB       1.3 [+ or -] 0.1 (b,c,d,e)
                              SS       0.1 [+ or -] 0.0 (e,f)
(15) N. [S.sup.ra]            FB       0.9 [+ or -] 0.2 (b,c,d,e)
  Guadalupe                   SS       0.2 [+ or -] 0.0 (c,d,e)
(16) N. [S.sup.ra]            FB       0.7 [+ or -] 0.1 (f)
  Machede                     SS       0.1 [+ or -] 0.0 (e,f)
(17) Rosal de la Frontera     FB       1.0 [+ or -] 0.1 (b,c)
                              SS       0.1 [+ or -] 0.0 (e,f)
(18) [S.sup.to] Aleixo da     FB       0.7 [+ or -] 0.1 (b,c)
  Restauracao                 SS       0.3 [+ or -] 0.0 (a, b,c,d)
(19) S. Miguel de Machede     FB       3.9 [+ or -] 0.5 (b,c,d,e)
                              SS       0.1 [+ or -] 0.0 (e,f)
(20) Serpa                    FB       0.4 [+ or -] 0.1 (b,c,d,e)
                              SS       0.0 [+ or -] 0.0 (f)
(21) [V.sup.e] Rocins         FB       0.5 [+ or -] 0.2 (b,c)
                              SS       0.2 [+ or -] 0.0 (d,e)
(22) Valverde                 FB       1.0 [+ or -] 0.1 (e)
                              SS       0.2 [+ or -] 0.0 (b,c,d,e)
(23) V. N. S. Bento           FB       0.7 [+ or -] 0.1 (b,c,d,e)
                              SS       0.3 [+ or -] 0.1 (a, b,c)
(24) Villanueva del           FB       0.8 [+ or -] 0.3 (b,c,d,e)
  Fresno                      SS       0.3 [+ or -] 0.0 (a, b,c)

Total                         FB       1.0 [+ or -] 0.7
                              SS       0.2 [+ or -] 0.1

                                       Minerals (mg/kg dry weight)

Sampling site               Sample                 Cr

(1) Almendres                 FB        1.3 [+ or -] 0.6 (a, b,c)
                              SS        1.0 [+ or -] 0.2 (a)
(2) Azaruja                   FB        0.6 [+ or -] 0.1 (c,d,e)
                              SS        0.9 [+ or -] 0.1 (a)
(3) Baleizao                  FB        1.0 [+ or -] 0.2 (d,e)
                              SS        2.4 [+ or -] 0.1 (b,c,d)
(4) Beja                      FB        0.8 [+ or -] 0.0 (a)
                              SS        4.4 [+ or -] 0.2 (i,j)
(5) Cabeca Gorda              FB        0.8 [+ or -] 0.0 (b,c,d,e)
                              SS        4.2 [+ or -] 0.0 (g,h,i,j)
(6) Cabezas Rubias            FB        0.9 [+ or -] 0.1 (f)
                              SS        3.1 [+ or -] 0.5 (c,d,e,f,g,h)
(7) Evora                     FB        0.9 [+ or -] 0.5 (f)
                              SS        2.8 [+ or -] 0.5 (c,d,e,f)
(8) Evoramonte                FB        2.9 [+ or -] 0.1 (b,c,d,e)
                              SS        2.6 [+ or -] 0.0 (c,d,e,f)
(9) [Her.sup.de] da Mitra     FB        1.8 [+ or -] 0.1 (b,c,d,e)
                              SS        3.8 [+ or -] 0.1 (f,g,h,i,j)
(10) Mertola                  FB        0.9 [+ or -] 0.3 (d,e)
                              SS        3.0 [+ or -] 0.2 (c,d,e,f,g)
(11) Mina S. Domingos         FB        0.7 [+ or -] 0.1 (a,b,c,d,e)
                              SS        5.1 [+ or -] 0.7 (j,k)
(12) [M.sup.te] da            FB        0.9 [+ or -] 0.1 (c,d,e)
  Borralha                    SS        3.7 [+ or -] 0.0 (d,e,f,g,h,i)
(13) [M.sup.te] Novo          FB        0.5 [+ or -] 0.1 (e)
                              SS        2.6 [+ or -] 0.2 (c,d,e,f)
(14) Montejuntos              FB        1.0 [+ or -] 0.1 (a, b,c,d)
                              SS        5.7 [+ or -] 0.2 (k)
(15) N. [S.sup.ra]            FB        0.5 [+ or -] 0.1 (a,b,c,d,e)
  Guadalupe                   SS        4.3 [+ or -] 0.1 (h,i,j)
(16) N. [S.sup.ra]            FB        0.4 [+ or -] 0.1 (b,c,d,e)
  Machede                     SS        3.8 [+ or -] 0.3 (e,f,g,h,i,j)
(17) Rosal de la Frontera     FB        1.2 [+ or -] 0.0 (a)
                              SS        2.9 [+ or -] 0.2 (c,d,e,f)
(18) [S.sup.to] Aleixo da     FB        0.9 [+ or -] 0.0 (b,c,d,e)
  Restauracao                 SS        2.8 [+ or -] 0.9 (c,d,e,f)
(19) S. Miguel de Machede     FB        0.9 [+ or -] 0.1 (a,b)
                              SS        2.3 [+ or -] 0.2 (b,c)
(20) Serpa                    FB        0.6 [+ or -] 0.1 (b,c,d,e)
                              SS        7.9 [+ or -] 0.4 (1)
(21) [V.sup.e] Rocins         FB        1.8 [+ or -] 0.6 (b,c,d,e)
                              SS        2.5 [+ or -] 0.1 (b,c,d,e)
(22) Valverde                 FB        2.9 [+ or -] 0.1 (b,c,d,e)
                              SS        1.2 [+ or -] 0.1 (a,b)
(23) V. N. S. Bento           FB        2.8 [+ or -] 0.4 (b,c,d,e)
                              SS        1.8 [+ or -] 1.3 (a, b,c)
(24) Villanueva del           FB        1.5 [+ or -] 0.4 (f)
  Fresno                      SS        4.9 [+ or -] 0.6 (i,j,k)

Total                         FB        1.2 [+ or -] 0.7
                              SS        3.3 [+ or -] 1.6

FB, fruiting bodies; SS, soil substrate. Mean values
(n = 9) [+ or -] SD. Different letters for each element indicate
significant differences with the confidence level of
p < 0.05 (ANOVA, Tukey's test).

Table 6: Mn, Pb, and Zn mineral content of A. ponderosa fruiting
bodies and corresponding soil substrates from different sampling
sites.

                                          Minerals (mg/kg dry weight)

                               Sample                 Mn

(1) Almendres                    FB          28 [+ or -] 6 (a,b,c,d)
                                 SS          22 [+ or -] 1 (a)
(2) Azaruja                      FB           6 [+ or -] 1 (a)
                                 SS         131 [+ or -] 9 (a,b)
(3) Baleizao                     FB          91 [+ or -] 14 (g,h)
                                 SS         893 [+ or -] 17 (f,g)
(4) Beja                         FB          84 [+ or -] 1 (f,g,h)
                                 SS         507 [+ or -] 45 (c)
(5) Cabeca Gorda                 FB          66 [+ or -] 6 (c,d,e,f,g,h)
                                 SS         695 [+ or -] 62 (d,e)
(6) Cabezas Rubias               FB          81 [+ or -] 1 (e,f,g,h)
                                 SS         735 [+ or -] 30 (d,e,f)
(7) Evora                        FB          59 [+ or -] 1 (b,c,d,e,f,g,h)
                                 SS         175 [+ or -] 3 (a,b)
(8) Evoramonte                   FB          79 [+ or -] 11 (e,f,g,h)
                                 SS         212 [+ or -] 12 (b)
(9) [Her.sup.de] da Mitra        FB          47 [+ or -] 9 (a,b,c,d,e,f,g)
                                 SS         725 [+ or -] 17 (d,e)
(10) Mertola                     FB          29 [+ or -] 6 (a,b,c,d)
                                 SS         725 [+ or -] 17 (d,e)
(11) Mina S. Domingos            FB         104 [+ or -] 1 (h)
                                 SS         290 [+ or -] 6 (b)
(12) [M.sup.te] da Borralha      FB          39 [+ or -] 16 (a,b,c,d,e,f)
                                 SS         199 [+ or -] 9 (b)
(13) [M.sup.te] Novo             FB          34 [+ or -] 4 (a,b,c,d)
                                 SS         139 [+ or -] 3 (a,b)
(14) Montejuntos                 FB          49 [+ or -] 9 (a,b,c,d,e,f,g)
                                 SS         649 [+ or -] 27 (c,d,e)
(15) N. [S.sup.ra] Guadalupe     FB          38 [+ or -] 5 (a,b,c,d,e)
                                 SS         794 [+ or -] 232 (e,f)
(16) N. [S.sup.ra] Machede       FB          50 [+ or -] 13 (a,b,c,d,e,f,g)
                                 SS         123 [+ or -] 4 (a,b)
(17) Rosal de la Frontera        FB          90 [+ or -] 56 (g,h)
                                 SS         725 [+ or -] 17 (d,e)
(18) [S.sup.to] Aleixo da        FB          47 [+ or -] 15 (c,d,e,f,g,h)
  Restauracao                    SS         193 [+ or -] 7 (b)
(19) S. Miguel de Machede        FB          91 [+ or -] 1 (g,h)
                                 SS         576 [+ or -] 17 (g)
(20) Serpa                       FB          69 [+ or -] 19 (a,b,c,d,e,f,g)
                                 SS        1002 [+ or -] 30 (c,d)
(21) Ve Rocins                   FB          15 [+ or -] 10 (a,b)
                                 SS         893 [+ or -] 17 (f,g)
(22) Valverde                    FB          26 [+ or -] 7 (a,b,c)
                                 SS          21 [+ or -] 1 (a)
(23) V. N. S. Bento              FB          72 [+ or -] 1 (d,e,f,g,h)
                                 SS         745 [+ or -] 45 (e,f)
(24) Villanueva del Fresno       FB          39 [+ or -] 6 (a,b,c,d,e,f)
                                 SS        1518 [+ or -] 45 (h)

Total                            FB          56 [+ or -] 27
                                 SS         529 [+ or -] 378

                                          Minerals (mg/kg dry weight)

                               Sample                 Pb

(1) Almendres                    FB         0.7 [+ or -] 0.5 (a,b)
                                 SS         4.0 [+ or -] 0.4 (a)
(2) Azaruja                      FB         0.4 [+ or -] 0.1 (a)
                                 SS         7.0 [+ or -] 1.0 (a,b,c,d,e)
(3) Baleizao                     FB         1.7 [+ or -] 0.5 (c,d,e,f)
                                 SS         9.2 [+ or -] 1.1 (d,e,f)
(4) Beja                         FB         2.8 [+ or -] 0.1 (a,b,c,d)
                                 SS         6.9 [+ or -] 0.6 (a,b,c,d,e)
(5) Cabeca Gorda                 FB         1.9 [+ or -] 0.2 (c,d,e,f)
                                 SS         4.5 [+ or -] 0.2 (a,b)
(6) Cabezas Rubias               FB         5.1 [+ or -] 0.8 (h,i)
                                 SS        16.5 [+ or -] 0.6 (g)
(7) Evora                        FB         1.9 [+ or -] 0.9 (i)
                                 SS        10.9 [+ or -] 0.2 (f)
(8) Evoramonte                   FB         4.2 [+ or -] 0.2 (g,h,i)
                                 SS         7.4 [+ or -] 0.3 (a,b,c,d,e,f)
(9) [Her.sup.de] da Mitra        FB         1.4 [+ or -] 0.2 (b,c,d,e,f)
                                 SS         5.4 [+ or -] 0.3 (a,b,c)
(10) Mertola                     FB         3.7 [+ or -] 0.9 (a,b,c,d,e)
                                 SS         8.0 [+ or -] 0.7 (b,c,d,e,f)
(11) Mina S. Domingos            FB         3.1 [+ or -] 0.7 (e,f,g,h)
                                 SS        10.3 [+ or -] 0.4 (e,f)
(12) [M.sup.te] da Borralha      FB         2.2 [+ or -] 0.3 (a,b,c)
                                 SS         5.4 [+ or -] 0.3 (a,b,c)
(13) [M.sup.te] Novo             FB         1.6 [+ or -] 0.3 (a)
                                 SS         8.2 [+ or -] 0.7 (c,d,e,f)
(14) Montejuntos                 FB         3.0 [+ or -] 0.1 (e,f,g,h)
                                 SS        10.2 [+ or -] 1.5 (e,f)
(15) N. [S.sup.ra] Guadalupe     FB         2.1 [+ or -] 0.3 (a,b,c,d,e,f)
                                 SS         4.5 [+ or -] 0.2 (a,b)
(16) N. [S.sup.ra] Machede       FB         0.4 [+ or -] 0.1 (a,b,c)
                                 SS         8.4 [+ or -] 0.1 (c,d,e,f)
(17) Rosal de la Frontera        FB         3.0 [+ or -] 0.1 (c,d,e,f,g)
                                 SS        16.5 [+ or -] 0.6 (g)
(18) [S.sup.to] Aleixo da        FB         3.9 [+ or -] 0.3 (i)
  Restauracao                    SS         6.4 [+ or -] 0.9 (a,b,c,d)
(19) S. Miguel de Machede        FB         1.4 [+ or -] 0.1 (f,g,h)
                                 SS         5.8 [+ or -] 0.2 (a,b,c,d)
(20) Serpa                       FB         1.0 [+ or -] 0.2 (d,e,f,g,h)
                                 SS        19.5 [+ or -] 3.7 (g)
(21) Ve Rocins                   FB         2.3 [+ or -] 0.8 (b,c,d,e,f)
                                 SS         9.2 [+ or -] 1.1 (d,e,f)
(22) Valverde                    FB         5.1 [+ or -] 0.2 (f,g,h)
                                 SS         4.4 [+ or -] 0.4 (a,b)
(23) V. N. S. Bento              FB         1.6 [+ or -] 0.1 (h,i)
                                 SS         8.4 [+ or -] 3.0 (c,d,e,f)
(24) Villanueva del Fresno       FB         3.1 [+ or -] 0.6 (a,b,c,d,e)
                                 SS         7.4 [+ or -] 0.5 (a,b,c,d,e,f)

Total                            FB         2.4 [+ or -] 1.3
                                 SS         8.5 [+ or -] 4.0

                                                Minerals
                                           (mg/kg dry weight)

                               Sample              Zn

(1) Almendres                    FB        59 [+ or -] 1 (a,b,c,d)
                                 SS         4 [+ or -] 0 (a,b)
(2) Azaruja                      FB        16 [+ or -] 2 (a)
                                 SS         5 [+ or -] 1 (a,b,c)
(3) Baleizao                     FB        97 [+ or -] 2 (c,d,e)
                                 SS         9 [+ or -] 0 (d)
(4) Beja                         FB       104 [+ or -] 1 (d,e)
                                 SS        13 [+ or -] 0 (g)
(5) Cabeca Gorda                 FB        81 [+ or -] 1 (b,c,d,e)
                                 SS        10 [+ or -] 0 (d)
(6) Cabezas Rubias               FB       132 [+ or -] 1 (e)
                                 SS        11 [+ or -] 1 (d,e)
(7) Evora                        FB        75 [+ or -] 1 (b,c,d,e)
                                 SS         5 [+ or -] 1 (b,c)
(8) Evoramonte                   FB        59 [+ or -] 7 (a,b,c,d)
                                 SS         6 [+ or -] 0 (c)
(9) [Her.sup.de] da Mitra        FB        61 [+ or -] 2 (a,b,c,d)
                                 SS        11 [+ or -] 0 (e,f)
(10) Mertola                     FB        52 [+ or -] 8 (a,b,c,d)
                                 SS        15 [+ or -] 1 (h)
(11) Mina S. Domingos            FB        91 [+ or -] 1 (c,d,e)
                                 SS         4 [+ or -] 1 (a)
(12) [M.sup.te] da Borralha      FB        62 [+ or -] 4 (a,b,c,d)
                                 SS        10 [+ or -] 1 (d,e)
(13) [M.sup.te] Novo             FB        52 [+ or -] 4 (a,b,c,d)
                                 SS         5 [+ or -] 0 (a,b,c)
(14) Montejuntos                 FB        60 [+ or -] 3 (a,b,c,d)
                                 SS        10 [+ or -] 0 (d,e)
(15) N. [S.sup.ra] Guadalupe     FB        63 [+ or -] 10 (a,b,c,d)
                                 SS        12 [+ or -] 0 (f,g)
(16) N. [S.sup.ra] Machede       FB        70 [+ or -] 2 (a,b,c,d)
                                 SS         9 [+ or -] 0 (d)
(17) Rosal de la Frontera        FB        95 [+ or -] 6 (c,d,e)
                                 SS         6 [+ or -] 0 (b,c)
(18) [S.sup.to] Aleixo da        FB        47 [+ or -] 1 (a,b,c,d)
  Restauracao                    SS        11 [+ or -] 0 (d,e)
(19) S. Miguel de Machede        FB        58 [+ or -] 8 (a,b,c,d)
                                 SS         9 [+ or -] 0 (d)
(20) Serpa                       FB        61 [+ or -] 9 (a,b,c,d)
                                 SS        30 [+ or -] 1 (j)
(21) Ve Rocins                   FB        31 [+ or -] 2 (a,b)
                                 SS         5 [+ or -] 1 (a,b,c)
(22) Valverde                    FB        42 [+ or -] 2 (a,b,c)
                                 SS         5 [+ or -] 0 (a,b,c)
(23) V. N. S. Bento              FB        86 [+ or -] 1 (b,c,d,e)
                                 SS        27 [+ or -] 1 (I)
(24) Villanueva del Fresno       FB        68 [+ or -] 4 (a,b,c,d)
                                 SS         5 [+ or -] 1 (a,b,c)

Total                            FB        68 [+ or -] 25
                                 SS        10 [+ or -] 6

FB, fruiting bodies; SS, soil substrate. Mean values
(n = 9) [+ or -] SD. Different letters for each element indicate
significant differences with the confidence level of
p < 0.05 (ANOVA, Tukey's test).

Table 7: Bioconcentration factor (BCF) values for A. ponderosa
fruiting bodies from different sampling sites.

Sampling site                         Al                 Ca

(1) Almendres                  1.3 [+ or -] 0.4   7.1 [+ or -] 0.7
(2) Azaruja                    0.2 [+ or -] 0.0   1.4 [+ or -] 0.0
(3) Baleizao                   0.9 [+ or -] 0.1   3.7 [+ or -] 0.1
(4) Beja                       1.8 [+ or -] 0.1   3.3 [+ or -] 0.2
(5) Cabeca Gorda               0.8 [+ or -] 0.1   5.1 [+ or -] 0.1
(6) Cabezas Rubias             2.5 [+ or -] 0.2   3.0 [+ or -] 0.4
(7) Evora                      0.6 [+ or -] 0.0   7.0 [+ or -] 1.8
(8) Evoramonte                 1.4 [+ or -] 0.1   7.2 [+ or -] 0.4
(9) [Her.sup.de] da Mitra      2.4 [+ or -] 0.1   3.6 [+ or -] 0.5
(10) Mertola                   2.1 [+ or -] 0.1   3.3 [+ or -] 0.3
(11) Mina S. Domingos          0.7 [+ or -] 0.0   4.2 [+ or -] 0.7
(12) [M.sup.te] da Borralha    1.3 [+ or -] 0.2   6.5 [+ or -] 0.9
(13) [M.sup.te] Novo           1.4 [+ or -] 0.1   2.9 [+ or -] 0.1
(14) Montejuntos               0.7 [+ or -] 0.1   1.8 [+ or -] 0.1
(15) N. [S.sup.ra] Guadalupe   1.0 [+ or -] 0.3   2.8 [+ or -] 0.1
(16) N. [S.sup.ra] Machede     0.5 [+ or -] 0.1   4.4 [+ or -] 0.7
(17) Rosal de la Frontera      1.7 [+ or -] 0.1   7.1 [+ or -] 0.5
(18) [S.sup.to] Aleixo da
  Restauracao                  0.1 [+ or -] 0.0   4.7 [+ or -] 0.5
(19) S. Miguel de Machede      1.2 [+ or -] 0.4   1.9 [+ or -] 0.0
(20) Serpa                     1.2 [+ or -] 0.2   5.8 [+ or -] 0.5
(21) [V.sup.e] Rocins          0.5 [+ or -] 0.1   3.9 [+ or -] 2.1
(22) Valverde                  0.4 [+ or -] 0.1   5.6 [+ or -] 0.2
(23) V. N. S. Bento            0.7 [+ or -] 0.0   2.6 [+ or -] 0.1
(24) Villanueva del Fresno     1.2 [+ or -] 0.1   1.9 [+ or -] 0.2

Sampling site                        Cu                  Fe

(1) Almendres                  123 [+ or -] 2    0.04 [+ or -] 0.01
(2) Azaruja                     16 [+ or -] 1    0.01 [+ or -] 0.00
(3) Baleizao                    48 [+ or -] 3    0.01 [+ or -] 0.00
(4) Beja                        43 [+ or -] 2    0.06 [+ or -] 0.00
(5) Cabeca Gorda               248 [+ or -] 16   0.01 [+ or -] 0.00
(6) Cabezas Rubias              28 [+ or -] 1    0.05 [+ or -] 0.00
(7) Evora                      122 [+ or -] 19   0.18 [+ or -] 0.01
(8) Evoramonte                  52 [+ or -] 1    0.03 [+ or -] 0.00
(9) [Her.sup.de] da Mitra       20 [+ or -] 1    0.01 [+ or -] 0.00
(10) Mertola                    9 [+ or -] 3     0.00 [+ or -] 0.00
(11) Mina S. Domingos          114 [+ or -] 10   0.02 [+ or -] 0.00
(12) [M.sup.te] da Borralha    119 [+ or -] 32   0.01 [+ or -] 0.01
(13) [M.sup.te] Novo            48 [+ or -] 2    0.02 [+ or -] 0.00
(14) Montejuntos                25 [+ or -] 1    0.01 [+ or -] 0.00
(15) N. [S.sup.ra] Guadalupe    31 [+ or -] 1    0.02 [+ or -] 0.00
(16) N. [S.sup.ra] Machede      39 [+ or -] 7    0.01 [+ or -] 0.00
(17) Rosal de la Frontera       68 [+ or -] 4    0.03 [+ or -] 0.03
(18) [S.sup.to] Aleixo da
  Restauracao                   23 [+ or -] 1    0.02 [+ or -] 0.00
(19) S. Miguel de Machede       38 [+ or -] 4    0.03 [+ or -] 0.00
(20) Serpa                      31 [+ or -] 1    0.01 [+ or -] 0.01
(21) [V.sup.e] Rocins           10 [+ or -] 1    0.01 [+ or -] 0.01
(22) Valverde                   83 [+ or -] 24   0.04 [+ or -] 0.01
(23) V. N. S. Bento             28 [+ or -] 1    0.04 [+ or -] 0.00
(24) Villanueva del Fresno      16 [+ or -] 1    0.02 [+ or -] 0.01

Sampling site                         K                 Mg

(1) Almendres                   74 [+ or -] 1    2.7 [+ or -] 0.3
(2) Azaruja                      5 [+ or -] 1    0.6 [+ or -] 0.0
(3) Baleizao                    23 [+ or -] 4    2.7 [+ or -] 0.0
(4) Beja                        10 [+ or -] 0    2.7 [+ or -] 0.3
(5) Cabeca Gorda               101 [+ or -] 0    4.5 [+ or -] 0.7
(6) Cabezas Rubias             157 [+ or -] 2    4.6 [+ or -] 0.3
(7) Evora                       53 [+ or -] 0    4.2 [+ or -] 0.6
(8) Evoramonte                  44 [+ or -] 7    4.9 [+ or -] 0.3
(9) [Her.sup.de] da Mitra      139 [+ or -] 18   2.3 [+ or -] 0.1
(10) Mertola                    25 [+ or -] 2    3.4 [+ or -] 0.2
(11) Mina S. Domingos          132 [+ or -] 8    2.0 [+ or -] 0.2
(12) [M.sup.te] da Borralha    168 [+ or -] 1    4.1 [+ or -] 0.4
(13) [M.sup.te] Novo            43 [+ or -] 1    2.6 [+ or -] 0.1
(14) Montejuntos               162 [+ or -] 6    0.8 [+ or -] 0.0
(15) N. [S.sup.ra] Guadalupe    53 [+ or -] 10   3.6 [+ or -] 0.4
(16) N. [S.sup.ra] Machede      27 [+ or -] 1    1.1 [+ or -] 0.1
(17) Rosal de la Frontera      326 [+ or -] 16   4.2 [+ or -] 0.1
(18) [S.sup.to] Aleixo da
  Restauracao                   24 [+ or -] 1    1.0 [+ or -] 0.0
(19) S. Miguel de Machede      142 [+ or -] 33   0.4 [+ or -] 0.1
(20) Serpa                     278 [+ or -] 16   2.7 [+ or -] 0.2
(21) [V.sup.e] Rocins           67 [+ or -] 34   1.5 [+ or -] 0.6
(22) Valverde                   65 [+ or -] 50   2.2 [+ or -] 0.2
(23) V. N. S. Bento             33 [+ or -] 3    1.2 [+ or -] 0.1
(24) Villanueva del Fresno      70 [+ or -] 13   2.8 [+ or -] 0.1

Sampling site                       Na                P

(1) Almendres                  20 [+ or -] 2     4.4 [+ or -] 0.4
(2) Azaruja                     3 [+ or -] 2     0.6 [+ or -] 0.2
(3) Baleizao                   21 [+ or -] 1     3.4 [+ or -] 0.3
(4) Beja                       22 [+ or -] 0     1.6 [+ or -] 0.1
(5) Cabeca Gorda               13 [+ or -] 0     3.4 [+ or -] 0.1
(6) Cabezas Rubias             43 [+ or -] 1     0.8 [+ or -] 0.0
(7) Evora                      24 [+ or -] 1     2.5 [+ or -] 0.1
(8) Evoramonte                 12 [+ or -] 2     5.0 [+ or -] 0.8
(9) [Her.sup.de] da Mitra      18 [+ or -] 0     2.4 [+ or -] 0.7
(10) Mertola                    6 [+ or -] 2     0.5 [+ or -] 0.2
(11) Mina S. Domingos          19 [+ or -] 1     2.0 [+ or -] 0.0
(12) [M.sup.te] da Borralha     5 [+ or -] 1     5.5 [+ or -] 0.5
(13) [M.sup.te] Novo            7 [+ or -] 0     3.7 [+ or -] 0.7
(14) Montejuntos               13 [+ or -] 3     1.5 [+ or -] 0.1
(15) N. [S.sup.ra] Guadalupe    8 [+ or -] 1     2.6 [+ or -] 0.5
(16) N. [S.sup.ra] Machede     27 [+ or -] 5     6.9 [+ or -] 0.5
(17) Rosal de la Frontera      12 [+ or -] 6     4.2 [+ or -] 0.5
(18) [S.sup.to] Aleixo da
  Restauracao                  16 [+ or -] 1     0.2 [+ or -] 0.0
(19) S. Miguel de Machede       6 [+ or -] 0     2.4 [+ or -] 0.5
(20) Serpa                     18 [+ or -] 1     2.6 [+ or -] 0.0
(21) [V.sup.e] Rocins           5 [+ or -] 2     2.2 [+ or -] 1.0
(22) Valverde                   8 [+ or -] 0     2.3 [+ or -] 0.5
(23) V. N. S. Bento            24 [+ or -] 1     1.7 [+ or -] 0.1
(24) Villanueva del Fresno     19 [+ or -] 1     1.7 [+ or -] 0.4

Values of each determination represents mean [+ or -] SD (n = 3).

Table 8: Bioconcentration factor (BCF) values for A. ponderosa
fruiting bodies from different sampling sites.

Sampling site                        Ag                 Ba

(1) Almendres                  38 [+ or -] 4    1.41 [+ or -] 0.33
(2) Azaruja                    72 [+ or -] 8    0.57 [+ or -] 0.04
(3) Baleizao                   55 [+ or -] 8    0.12 [+ or -] 0.04
(4) Beja                        8 [+ or -] 1    0.33 [+ or -] 0.08
(5) Cabeca Gorda               14 [+ or -] 4    0.14 [+ or -] 0.01
(6) Cabezas Rubias             10 [+ or -] 1    0.77 [+ or -] 0.01
(7) Evora                       2 [+ or -] 1    0.19 [+ or -] 0.05
(8) Evoramonte                  7 [+ or -] 0    0.32 [+ or -] 0.02
(9) [Her.sup.de] da Mitra       4 [+ or -] 0    0.53 [+ or -] 0.02
(10) Mertola                   61 [+ or -] 2    0.50 [+ or -] 0.01
(11) Mina S. Domingos           2 [+ or -] 0    0.30 [+ or -] 0.01
(12) [M.sup.te] da Borralha     9 [+ or -] 0    0.40 [+ or -] 0.04
(13) [M.sup.te] Novo            6 [+ or -] 0    0.39 [+ or -] 0.05
(14) Montejuntos                6 [+ or -] 1    0.29 [+ or -] 0.00
(15) N. [S.sup.ra] Guadalupe   16 [+ or -] 1    0.31 [+ or -] 0.08
(16) N. [S.sup.ra] Machede      1 [+ or -] 1    0.14 [+ or -] 0.02
(17) Rosal de la Frontera      11 [+ or -] 2    0.65 [+ or -] 0.01
(18) [S.sup.to] Aleixo da
  Restauracao                   7 [+ or -] 0    0.39 [+ or -] 0.00
(19) S. Miguel de Machede      16 [+ or -] 1    0.38 [+ or -] 0.08
(20) Serpa                      9 [+ or -] 0    0.18 [+ or -] 0.02
(21) [V.sup.e] Rocins          35 [+ or -] 21   0.27 [+ or -] 0.10
(22) Valverde                  12 [+ or -] 2    0.31 [+ or -] 0.04
(23) V. N. S. Bento             4 [+ or -] 2    0.46 [+ or -] 0.01
(24) Villanueva del Fresno     32 [+ or -] 6    0.10 [+ or -] 0.01

Sampling site                       Cd                 Cr

(1) Almendres                  7 [+ or -] 2    1.30 [+ or -] 0.39
(2) Azaruja                    1 [+ or -] 0    0.68 [+ or -] 0.03
(3) Baleizao                   7 [+ or -] 0    0.42 [+ or -] 0.07
(4) Beja                       15 [+ or -] 1   0.19 [+ or -] 0.00
(5) Cabeca Gorda               4 [+ or -] 0    0.18 [+ or -] 0.01
(6) Cabezas Rubias             8 [+ or -] 1    0.29 [+ or -] 0.00
(7) Evora                      4 [+ or -] 2    0.30 [+ or -] 0.13
(8) Evoramonte                 9 [+ or -] 1    1.14 [+ or -] 0.02
(9) [Her.sup.de] da Mitra      11 [+ or -] 2   0.47 [+ or -] 0.00
(10) Mertola                   8 [+ or -] 1    0.32 [+ or -] 0.07
(11) Mina S. Domingos          13 [+ or -] 1   0.13 [+ or -] 0.00
(12) [M.sup.te] da Borralha    12 [+ or -] 1   0.24 [+ or -] 0.02
(13) [M.sup.te] Novo           3 [+ or -] 0    0.20 [+ or -] 0.02
(14) Montejuntos               10 [+ or -] 2   0.18 [+ or -] 0.01
(15) N. [S.sup.ra] Guadalupe   5 [+ or -] 1    0.12 [+ or -] 0.01
(16) N. [S.sup.ra] Machede     8 [+ or -] 1    0.10 [+ or -] 0.00
(17) Rosal de la Frontera      9 [+ or -] 1    0.42 [+ or -] 0.02
(18) [S.sup.to] Aleixo da
  Restauracao                  3 [+ or -] 0    0.36 [+ or -] 0.10
(19) S. Miguel de Machede      38 [+ or -] 1   0.41 [+ or -] 0.03
(20) Serpa                     14 [+ or -] 3   0.07 [+ or -] 0.01
(21) [V.sup.e] Rocins          3 [+ or -] 1    0.71 [+ or -] 0.24
(22) Valverde                  5 [+ or -] 0    2.38 [+ or -] 0.03
(23) V. N. S. Bento            2 [+ or -] 0    2.46 [+ or -] 2.01
(24) Villanueva del Fresno     3 [+ or -] 1    0.30 [+ or -] 0.04

Sampling site                          Mn                   Pb

(1) Almendres                  1.27 [+ or -] 0.22   0.15 [+ or -] 0.10
(2) Azaruja                    0.05 [+ or -] 0.00   0.06 [+ or -] 0.00
(3) Baleizao                   0.10 [+ or -] 0.01   0.19 [+ or -] 0.03
(4) Beja                       0.17 [+ or -] 0.01   0.41 [+ or -] 0.02
(5) Cabeca Gorda               0.09 [+ or -] 0.00   0.41 [+ or -] 0.03
(6) Cabezas Rubias             0.11 [+ or -] 0.00   0.31 [+ or -] 0.04
(7) Evora                      0.34 [+ or -] 0.00   0.17 [+ or -] 0.08
(8) Evoramonte                 0.37 [+ or -] 0.03   0.56 [+ or -] 0.00
(9) [Her.sup.de] da Mitra      0.06 [+ or -] 0.01   0.26 [+ or -] 0.03
(10) Mertola                   0.04 [+ or -] 0.01   0.46 [+ or -] 0.07
(11) Mina S. Domingos          0.36 [+ or -] 0.00   0.30 [+ or -] 0.06
(12) [M.sup.te] da Borralha    0.19 [+ or -] 0.07   0.41 [+ or -] 0.03
(13) [M.sup.te] Novo           0.24 [+ or -] 0.02   0.20 [+ or -] 0.02
(14) Montejuntos               0.08 [+ or -] 0.01   0.30 [+ or -] 0.04
(15) N. [S.sup.ra] Guadalupe   0.05 [+ or -] 0.01   0.48 [+ or -] 0.06
(16) N. [S.sup.ra] Machede     0.40 [+ or -] 0.09   0.05 [+ or -] 0.02
(17) Rosal de la Frontera      0.12 [+ or -] 0.07   0.18 [+ or -] 0.00
(18) [S.sup.to] Aleixo da
  Restauracao                  0.24 [+ or -] 0.07   0.62 [+ or -] 0.04
(19) S. Miguel de Machede      0.16 [+ or -] 0.00   0.23 [+ or -] 0.01
(20) Serpa                     0.07 [+ or -] 0.02   0.05 [+ or -] 0.00
(21) [V.sup.e] Rocins          0.02 [+ or -] 0.01   0.25 [+ or -] 0.06
(22) Valverde                  1.23 [+ or -] 0.28   1.16 [+ or -] 0.05
(23) V. N. S. Bento            0.10 [+ or -] 0.00   0.21 [+ or -] 0.07
(24) Villanueva del Fresno     0.03 [+ or -] 0.00   0.42 [+ or -] 0.06

Sampling site                       Zn

(1) Almendres                  13 [+ or -] 0
(2) Azaruja                    3 [+ or -] 0
(3) Baleizao                   10 [+ or -] 0
(4) Beja                       8 [+ or -] 0
(5) Cabeca Gorda               8 [+ or -] 0
(6) Cabezas Rubias             13 [+ or -] 1
(7) Evora                      15 [+ or -] 2
(8) Evoramonte                 10 [+ or -] 1
(9) [Her.sup.de] da Mitra      5 [+ or -] 0
(10) Mertola                   3 [+ or -] 0
(11) Mina S. Domingos          25 [+ or -] 3
(12) [M.sup.te] da Borralha    6 [+ or -] 0
(13) [M.sup.te] Novo           10 [+ or -] 0
(14) Montejuntos               6 [+ or -] 0
(15) N. [S.sup.ra] Guadalupe   5 [+ or -] 1
(16) N. [S.sup.ra] Machede     7 [+ or -] 0
(17) Rosal de la Frontera      16 [+ or -] 1
(18) [S.sup.to] Aleixo da
  Restauracao                  4 [+ or -] 0
(19) S. Miguel de Machede      6 [+ or -] 1
(20) Serpa                     2 [+ or -] 0
(21) [V.sup.e] Rocins          6 [+ or -] 0
(22) Valverde                  9 [+ or -] 0
(23) V. N. S. Bento            3 [+ or -] 0
(24) Villanueva del Fresno     14 [+ or -] 1

Values of each determination represents mean [+ or -] SD (n = 3).

Table 9: Clusters center of gravity.

Variable             Cluster 1

Ag             2.433 [+ or -] 1.593
Al           396.503 [+ or -] 210.361
Ba             1.442 [+ or -] 0.962
Ca           583.148 [+ or -] 246.929
Cd             1.316 [+ or -] 1.120
Cr             1.213 [+ or -] 0.933
Cu           224.094 [+ or -] 168.207
Fe            63.880 [+ or -] 41.865
K          34192.173 [+ or -] 13478.898
Mg           864.049 [+ or -] 419.246
Mn            64.046 [+ or -] 29.957
Na           1240.238 [+ or -] 624.901
P            422.333 [+ or -] 167.110
Pb             2.440 [+ or -] 1.468
Zn            72.936 [+ or -] 29.664

Variable             Cluster 2

Ag             1.690 [+ or -] 1.248
Al           392.625 [+ or -] 226.842
Ba             0.996 [+ or -] 0.616
Ca           617.110 [+ or -] 151.838
Cd             0.854 [+ or -] 0.322
Cr             1.753 [+ or -] 1.122
Cu           238.065 [+ or -] 108.827
Fe            252.718 [+ or -] 74.423
K          23008.328 [+ or -] 23670.094
Mg           664.112 [+ or -] 268.732
Mn            76.347 [+ or -] 18.841
Na           1286.973 [+ or -] 354.879
P             219.741 [+ or -] 66.174
Pb             3.389 [+ or -] 1.805
Zn            80.972 [+ or -] 33.836

Variable             Cluster 3

Ag             1.690 [+ or -] 2.309
Al           227.812 [+ or -] 187.093
Ba             0.719 [+ or -] 0.616
Ca           413.341 [+ or -] 260.514
Cd             0.630 [+ or -] 0.526
Cr             0.656 [+ or -] 0.486
Cu            112.320 [+ or -] 90.198
Fe            36.283 [+ or -] 27.911
K          21750.972 [+ or -] 12089.189
Mg           533.816 [+ or -] 354.098
Mn            30.254 [+ or -] 19.582
Na           598.937 [+ or -] 391.228
P            182.971 [+ or -] 173.598
Pb             1.640 [+ or -] 1.256
Zn            45.118 [+ or -] 29.973

Table 10: The coincidence matrix for Decision Tree
model presented in Figure 5.

                        Training set

Variables   Cluster 1    Cluster 2     Cluster 3

Cluster 1      24            0             0
Cluster 2       0            8             0
Cluster 3       0            0            14

                        Test set

Variables   Cluster 1   Cluster 2   Cluster 3

Cluster 1       9           0           0
Cluster 2       0           4           0
Cluster 3       0           0          10

Table 11: The coincidence matrix for Decision Tree model
presented in Figure 6.

                        Training Set

Variables   Cluster 1   Cluster 2   Cluster 3

Cluster 1      22           2           0
Cluster 2       1           7           0
Cluster 3       2           0          12

                        Test Set

Variables   Cluster 1   Cluster 2   Cluster 3

Cluster 1       8           1           0
Cluster 2       1           3           0
Cluster 3       2           0           8

Figure 2: Moisture, dry weight, and organic and minerals content
of A. ponderosa fruiting bodies from 24 different sampling sites.

Organic content    7.6 [+ or -] 1.4%
Minerals           0.7 [+ or -] 0.2%

dry weight         8.3 [+ or -] 1.4%
Moisture          91.7 [+ or -] 1.4%

Note: Table made from pie chart.
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Title Annotation:Research Article
Author:Salvador, Catia; Martins, M. Rosario; Vicente, Henrique; Caldeira, A. Teresa
Publication:International Journal of Analytical Chemistry
Date:Jan 1, 2018
Words:18492
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