*3.1. PTEs and PAHs in the Soil*

The results of the soil analyses are summarized in Table 1. Descriptive statistics are detailed for each element (including PTEs and essential minerals) and PAHs analyzed. The soils closer (<5 km) to industrial areas contained higher content of PTEs and heavy weight PAHs than those located farther away (>5 km).


**Table 1.** Comparison of metals and polycyclic aromatic hydrocarbons in soils from farms located <5 or >5 km from industrial areas.

Max: maximum; min: minimum; *n*: number of farms; V: close-far variation; <0.1: not detected; RBSSL: risk-based soil-screening level.

The similitude between the mean and the median is a preliminary indicator of normal distribution. The variation (V%) revealed an enrichment of PTEs and PAHs in soils closer to industrial areas (Table 1), with the highest value for dibenzo(a,h)anthracene (89%). The enrichment of Zn, Cd, and Pb was consistent with the known metal emissions from current and past industrial activities in this region of northern Spain [30,31]. PAHs with a molecular weight higher than that of fluoranthene were also enriched in the soil closer to industrial areas, except for pyrene (38%), and these results are consistent with studies of soils near industrial areas in northern Spain [32,33]. The enrichment of these high molecular weight PAHs is concerning, as these are the most persistent PAHs in the environment. In addition, these data are consistent with previous studies on soils located near to the industrial areas [32–35].

To assess the risk that the observed levels of pollutants may pose for humans and the environment, we compared the measured levels to so-called "risk-based soil screening levels" (RBSSLs) [36], which are based on toxicity parameters for different uses of soil (Table 1). We applied the most restrictive values for "other uses" of soil, which include farming [36].

In the soils close to industries, Cu, Zn, As, Cd, Hg, and Pb exceeded the threshold limits by at least 100%. For instance, the mean concentration of Hg (0.97 mg kg−1), one of the most toxic elements, was close to its RBSSL (1 mg kg−1). In the case of soils located more than 5 km away from industrial areas, thresholds were occasionally exceeded only for Cu and Hg, and mean values were much lower than RBSSLs. In the case of PAHs, the concentration of benzo(a)pyrene in the soils closer to industries (45.4 μg kg−1) was more than twice the RBSSL (20 μg kg−1), while it was notably lower in the soils farther away (16.7 μg kg<sup>−</sup>1). More specifically, the soils from N1, N2, and N3 dairy farms showed levels of benzo(a)pyrene above their ML, with 85.9, 61.4, and 78.7 ug kg−1, respectively. These three farms are located less than 2 km from the steel industry and less than 5 km from the zinc industry. Similar enrichment in heavy-molecular-weighted PAHs has been previously reported in soils located less than 2 km from a Cu smelting industry [37]. These results suggest that livestock near industrial areas may be exposed to above-threshold levels of several pollutants when they feed on forage cultivated on local soils.

To identify potential pollution sources, principal component analysis was performed using all the samples, irrespective of their location (Table 2). Four principal components explained 83% of the initial variance with high communality values. PTEs such as As and Pb were quite well represented by principal component 1, which was also associated with high Fe and Se load, suggesting the presence of an anthropogenic source that was probably related to the steel industry (Fe) and/or coal-combustion (Fe and Se) power plants [27,38]. This component 1 was also associated with natural iron oxy-hydroxides, which may explain the presence of As. The elements with higher loads in the second principal component were Mg, Ca, and K, which were probably associated with natural sources, such as calcareous and clayey materials. In the third principal component, a remarkable association was observed among high concentrations of PAHs, Zn, and Cd, consistent with emissions from the Zn smelting industry [32]. The correlation between Zn and heavy molecular-weighted PAHs has been also observed in soils near Cu smelting industries [37]. The high levels of PAHs and the contribution of Pb in the third component, together with the absence of PAHs in the other two components, may indicate heavy-traffic pollution as another source [39]. The fourth principal component was linked to Na and Cr, both naturally occurring elements.


**Table 2.** Principal component data matrix (rotated) for potentially toxic elements and polycyclic aromatic hydrocarbons (PAHs) in soils.


**Table 2.** *Cont.*

VE: Variance explained (cumulative).

### *3.2. PTEs and PAHs in Forage*

Table 3 shows the concentration (mean, median, minimum, and maximum) and the percentage of variation between mean (and median) concentration of inorganic elements and PAHs in the forages produced near to (<5 km) and farther from (>5 km) the pointsources of pollution. The concentrations of PTEs and PAHs in the forage were generally lower than those measured in the soils, suggesting limited transfer from soils to plants [27]. This could be explained by the low bioavailability of PTEs in the soils of the industrial areas [27], and perhaps by low deposition from the atmosphere.

**Table 3.** Comparison of potentially toxic metals and polycyclic aromatic hydrocarbons in feed from farms <5 or >5 km from industrial areas.


Max: maximum; min: minimum; *n*: number of farms; V: close–far variation; <0.1: not detected; ML: maximum level according to EU regulations; <sup>a</sup> (EU 2019/1869); <sup>b</sup> (EU 1275/2013).

The number of pollutants enriched closer to the industry was smaller in the forage than in the soil (Tables 1 and 3), although in both types of samples, Zn, Cd, and PAHs with at least four aromatic rings were enriched closer to industrial areas. This enrichment in high-molecular-weight PAHs in the soils and plants can be partially explained by the "distillation effect" [40]: high molecular weight PAHs in the atmosphere deposit onto surfaces closer to their source, whereas low-molecular weight PAHs diffuse farther before deposition. The levels of PAHs found in forage samples (1–20 μg kg−<sup>1</sup> dry weight) were lower than those reported in 2003 in grasslands near roads with high-traffic intensity [20], but they were similar to those in forages from urban and rural farms [19].

Forage (fresh forage or silage) is the primary source of essential mineral supply to cattle in sustainable farms [7]. The essential trace minerals are required in the diet of the animals, as they play fundamental roles in their organisms, such as the roles of enzyme cofactors, catalyzers of metabolic reactions, and so on [7]; however, they become potentially toxic at high concentrations, so the National Research Council (NRC, United States) has established tolerable limits for these elements in the cattle diet. Nearly all the essential trace minerals (Zn, Cu, Se, and Cr) were below the maximum tolerable limits that the NRC recommends for cattle [41]. The only exception was Fe, whose median concentration (920 mg kg−<sup>1</sup> dry weight) in farms near industries exceeded the tolerable level of 500 mg kg<sup>−</sup>1, which was much higher than the concentration found in farms far away from industrial areas.

Among the PTEs, Cd and Pb showed respective median concentrations of 0.115 and 1.10 mg kg−<sup>1</sup> in the forage, which were below the levels in forage produced near industrial activities in Romania [11] or India [42] but above the levels on commercial farms in England [43]. The maximal content of As (7.87 mg kg−1), Cd (1.31 mg kg−1), and Hg (0.47 mg kg<sup>−</sup>1) in the forage exceeded the maximum levels (ML) for animal feed based on European Union regulations [44,45]. More specifically, one farm exceeded the ML of Cd (N1, see Table A1) with 1.31 mg kg<sup>−</sup>1. In that sense, other farms near industries (N2 and N3) also had high levels of Cd (>0.6 mg kg<sup>−</sup>1), although they did not exceed its ML. In contrast, the As concentration in the forages was above its ML in farm F4 (7.87 mg kg−1), which is more than 20 km away from point pollution sources (Table A1), and in the close-to-industry farms N1 (3.94 mg kg−1) and N4 (2.19 mg kg−1). Again, the Hg concentration exceed its ML in farm F4 (0.47 mg kg<sup>−</sup>1) and in the close-to-industry farms N4 (1.13 mg kg−1) and N7 (1.14 mg kg<sup>−</sup>1). Thus, As and Hg might not be enriched near industrial facilities, which is similar to what we observed in the soil. These results suggest that As and Hg, in particular, may have a natural occurrence.

Regarding PAHs, EU legislation has not established a ML in animal feed for these compounds. However, the most concerning PAH is benzo(a)pyrene, whose concentrations were higher in the forages produced in farm N2 (48.7 μg kg−1), which is located 0.4 km from the steel industry (Table A1). Together with benzo(a)pyrene, the forage from this farm also contained higher concentrations of the rest of PAHs.
