ENZYMATIC ACTIVITY AND CONTENT OF POLYCYCLIC AROMATIC HYDROCARBONS (PAHS) IN SOILS UNDER LOW-STACK EMISSION IN LUBLIN

123 Abstract. Soils from the Lublin area exposed to low-stack emission from home heating and transport were investigated. Changes in soil enzymatic activity and polycyclic aromatic hydrocarbons (PAHs) content were analysed. Soil samples were collected in July and November 2016 and January 2017. Results of the study show that enzymatic activity and PAHs content depended on types of buildings, study period (intensification of pollutant emission from household heating), traffic volume and atmospheric air movement. Low enzymatic activities and high PAHs content in the soils were observed in the autumn-winter period in the areas with dense single-family housing and located in the vicinity of streets with intense road traffic.


INTRODUCTION
Polycyclic aromatic hydrocarbons (PAHs) are mutagenic and carcinogenic pollutants that pose a significant human health hazard (Wild and Jones 1995).PAHs identified as the strongest carcinogens include benzo[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenz[ah]anthracene, and indeno [1,2,3-cd]pyrene (Kanaly and Harayama 2000).Moreover, these xenobiotics disrupt homeostasis of soil environment (Caravaca andRodán 2003, Dębska et al. 2011), contributing to future ecological problems and consequences for the whole urban area.Low-stack emission (transport and home heating) is a significant source of PAH contamination in urban soils.Its environmental impact is further intensified by expansion of artificial surfaces (widely used to cover streets, squares, roofs, etc.) and impairment of complete air change in areas with high-density housing (Adamczewska et al. 2000, Bielińska et al. 2014).
Enzyme activity reflects anthropogenic changes in the soil environment caused by stressors.It is a precise measure of soil quality that includes both homeostatic capacity of an ecosystem, and a level of its pollution which endangers living organisms (Bielińska et al. 2009, Siwik-Ziomek and Lemanowicz 2016).In order to identify a risk of long-term adverse changes in urban soils, the biochemical indicators reflecting the metabolism of soil environment and describing the current eco-chemical soil condition were used.

MATERIALS AND METHODS
The study was conducted in the city of Lublin and encompassed the areas exposed to different levels of low-stack emission (transport, household furnaces).Soil samples were collected from the 0-25 cm layer in three time periods: in July and November 2016, and in January 2017.The soil sampling sites and potential sources of pollution are listed in Table 1.Representative surface areas on lawns were selected in each study site (Fig. 1).Then samples for further analysis were mixed to obtain an average of the original 5 soil subsamples taken from each surface area.Contents of 16 PAHs were analysed in the soil samples by HPLC with UV detection (254 nm) according to Baran and Oleszczuk (2001) methodology.The activity of four soil enzymes was determined: dehydrogenases (Thalmann 1968), acid and alkalic phosphatases (Tabatabai and Bremner 1969), and urease (Zantua and Bremner 1975).These enzymes play a significant role in transformation of soil organic matter and are highly sensitive to environmental factors.The statistical analysis of the results was performed in the Microsoft Office Excel 2003 spreadsheet and Statistica v. 10PL software.Statistical evaluation of the variability of the results was carried out with the two-way analysis of variance.The significance of the differences between the mean values was verified with Tukey's t-test at a significance level of p≤0.05.For some parameters, the value of the Pearson correlation coefficient (r) was calculated at p˂0.05.A maximum 5% dispersion of measurements in the biochemical and chemical analysis was assumed in the study.

RESULTS AND DISCUSSION
Elevated levels of PAHs (Table 2) were observed in the soils taken from the investigated parts of the city in all three sampling periods.According to the classification by Maliszewska-Kordybach (1996), the soils studied can be classified as polluted, with the exception of the soil from the urban park (Ogród Saski) that could be described as slightly contaminated.The highest sum of 16 PAHs was found in the soil sampled in the Kalinowszczyzna district (areas I and II), particularly from Stara Kalinowszczyzna (area II) with its tenement houses built in the early 1900s where household heating most often uses cheap coal of poor quality and, consequently, of low heating value.Another factor influencing the level of soil contamination with PAHs in area II is its location in a land depression (the Bystrzyca River valley) which, alongside the high-density housing, restricts air movement.Moreover, areas I and II are located in the immediate vicinity of a busy arterial road.The total content of PAHs in the soil from area I (Osiedle 40-lecia, a tower block housing estate) ranged between 2,669 and 3,219 µg•kg -1 and between 3,448 to 4,501 µg•kg -1 in the soil from area II (Stara Kalinowszczyzna), whereas the PAH content in the soil from the remaining areas examined was several times lower (Table 2).In the soil collected from area III, despite its heavy traffic and dense single-family housing, the total content of PAHs was about two-fold lower than in the soil sampled in areas I and II.This result can be explained by the fact that low-rise buildings that prevail in area III allow particulate matter and gases to be transported longer distances (Kluska andKroszczyński 2000, Futa et al. 2016).This conclusion is further supported by lower concentrations of benzo[a]pyrene and benzo[ghi]perylene found in the soil from this area compared with areas I and II (Table 2).The above PAHs are adsorbed and emitted into the air on very fine particulate matter and can travel considerable distances (Świetlik et al. 1998).Soil contamination with PAHs in area IV (Lublin-Zamość exit road) was similar to that found in the soil from area III.Some differences could result from the lower level of low-stack emission in area IV compared with that collected from area III.The high content of PAHs (1,010-1,203 μg•kg -1 , Table 2) observed in the soil from an urban park (area V), especially in the autumn-winter period, could be partly explained by the lack of protection provided by trees due to a seasonal loss of their foliage.Ogród Saski, the park located in the city centre, is exposed to serious urban air pollution, mainly due to vehicle traffic, while its stand of trees reduces air movement.
The analysis of the individual PAHs demonstrated that in all cases (areas, sampling dates) hydrocarbons composed of 2-4 aromatic rings were dominant: naphthalene, fluorene, phenanthrene, fluoranthene, and pyrene accounted for about 70% of all the determined PAHs (    1.6 1.7 1.9 1.8 2.0 2.2 0.8 0.9 1.0 0.7 0.9 1.0 1.6 2.8 2.8 DahA 0.8 0.9 1.0 0.7 0.9 1.0 1.1 1.5 1.5 1.0 1.4 1.6 2.7 3.9 3.8 BghiP 1.0 1.1 1.1 1.6 1.9 2.0 0.5 0.7 0.8 0.6 0.9 1.0 0.7 0.8 0.9 Ind 1.The level of soil contamination with PAHs in areas I and II was similar, however, differences were found in contents of particular hydrocarbons, i.e.: fluorene, phenanthrene, pyrene, and benz[a]anthracene (Table 3).Greater percentage of benz[a]anthracene in the soils from area II noted in sampling time 2 and 3 indicates increased emission from household heating where coal is burned.One method of identifying sources of soil contamination with PAHs is to calculate the ratio between selected hydrocarbons (Oleszczuk and Baran 2005).In order to determine likely PAH sources, phenanthrene/anthracene and fluoranthene/pyrene ratios are most often used (Gschwend and Hites 1981, Oleszczuk and Baran 2005).The data analysis showed that coal burning in the autumn-winter period is the main source of PAHs in the soils of area I, II, III and V, which is reflected in the values of phenanthrene/anthracene ratios.The values did not exceed 10 (they ranged from 5.3 to 9.9), and in the case of the fluoranthene/pyrene ratios were higher than 1.The concentrations of the most carcino-and mutagenic PAHs (i.e.benz[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenz[a,h]anthracene, indeno[1,2,3-cd]pyrene) did not exceed 20% of the total PAH content, ranging from 8.5% to 12.7% in the soil from areas III, IV and V, and from 14.8% to 19.0% in the soil from areas I and II (Table 5).1.6 1.7 1.9 1.8 2.0 2.2 0.8 0.9 1.0 0.7 0.9 1.0 1.6 2.8 2.8 DahA 0.8 0.9 1.0 0.7 0.9 1.0 1.1 1.5 1.5 1.0 1. 4  Enzymatic activity in the soil studied clearly varied depending upon the area and sampling time.The level and direction of the change was influenced by properties of the enzyme analyzed (Table 6).
Soils from areas I and II showed the lowest activity of dehydrogenases, acid phosphatase and alkaline phosphatase (Table 6).The decreased enzyme activities in both areas resulted from intensive low-stack emission, which was the source of PAHs in the soils (Table 2).The 16 PAHs content had a significantly negative correlation with the activity of most of the studied soil enzymes: dehydrogenases, acid phosphatase and alkaline phosphatase (Table 7).The data presented by numerous authors (Maliszewska-Kordybach and Smreczak 1997, Baran et al. 2004, Wyszkowska et al. 2006, Bielińska et al. 2014, Lipińska et al. 2014) confirm particularly high sensitivity of dehydrogenases and phosphatases to soil contamination with PAHs.Their activity may be used as an indicator of soil pollution with these xenobiotics (Bielińska et al. 2014).Areas III and IV, where PAH levels were within the same range, exhibited similar activities of dehydrogenases and phosphatases in the soil (Table 2).The highest activity of these enzymes was stated in the soil from the park (area V) and it was 1.5-2 times higher than in the soils collected in areas I and II (Table 6).Relatively high enzymatic activity of the soil from the park could be connected with the entry of fresh organic matter to soil environment.It should be emphasised that fresh organic matter not only activates metabolism of microorganisms but also is favourable for decomposition of organic pollutants (Junter et al. 2002).No clear link between the study area and urease activity was found in the study period (Table 6).Urease is resistant to external factors and can even increase its activity in extreme conditions (Stępniewska andSamborska 2002, Li et al. 2015).As an extracellular enzyme, urease is synthesised only in the presence of its substrate, urea, and its availability is the only factor limiting urease activity (Carbrera et al. 1994).Large urban agglomerations are particularly significant urea producers.The sources of urea in urban soils are, among others things, animal excreta, household food waste, fragments of tissues and cells of soil micro-, mesoand macrofauna, plant residues, and, in the case of parks, intentionally added organic fertilisers (Russel and Wyczółkowski 2005).Urease activity can be used to assess the level of soil anthropogenisation (Russel and Wyczółkowski 2005).
Changes of biochemical processes in the specified sampling periods differed depending on enzyme properties (Table 6).The lowest activities of dehydrogenases, acid phosphatase and alkalic phosphatase in all studied soils were observed in the 2 nd sampling period (the second half of November).Activities of these enzymes in winter (the 3 rd sampling period) were comparable to those stated in summer (the second half of July).Many authors (i.a.Burton andBeauchamp 1994, Díaz-Raviña et al. 1995) indicate relatively high soil enzymatic activity in winter and directly after thawing of the soil.The literature data describe a complex mechanism of sorption and desorption of enzymes following the freezing and thawing of the soil (Burton andBeauchamp 1994, Bielińska et al. 2014a).The increase in enzyme activity as a result of freezing and thawing of the soil may be linked to destruction of soil aggregates.Bacteria frequently form the core of soil micro-aggregates.Winter increase in soil enzymatic activity is attributed to accumulation of enzymes released by dying soil organisms and to desorption and reactivation of enzymes accumulated earlier in the soil (Bielińska et al. 2014a).The lowest activity of urease was observed in the winter time (Table 6).It could be connected with different mechanisms regulating immobilisation of this enzyme in soils (Januszek 1999), and reduced availability of the substrate for urease in this season.In the study on influence of soil freezing and thawing on activity of selected enzymes, Januszek (1999) showed decrease in urease activity as the result of its desorption.CONCLUSIONS 1.The results indicate that both enzyme activity and content of PAHs in soil depend on a type of housing, study period (intensification of pollutant emission from household heating), traffic volume and atmospheric air movement.
2. The level of soil contamination with PAHs within the studied areas of Lublin is high and dominance of 2-4 ring aromatic hydrocarbons proves that the low-stack emission is the main source of soil pollution with PAHs.
3. Coefficients calculated on the basis of phenanthrene/anthracene and fluoranthene/pyrene ratios indicate that carbon combustion is the main source of PAHs in the soil studied in the autumn-winter period.
4. Low enzymatic activities and high PAHs contents in the soils were noticed in the autumn-winter period in the areas with dense single-family housing and located in the vicinity of streets with intense road traffic.PAHs pollutants generated in open areas by low-stack emission are transported over longer distances, which was reflected by their lower contents in the soils.
5. The enzymatic activity measurements in the soils allow environmental risk assessment connected with low-stack emission.
6. PAHs durability in the soil contributes to their accumulation, indicating the need of monitoring the soils of Lublin in the areas exposed to low-stack emission.

TABLE 1 .
CHARACTERISTICS OF SAMPLING SITES

Table 3 )
. Observations of other authors (Sims and Overcash 1983, Wild and Jones 1995, Zerbe et al. 1995) suggest that

TABLE 2 .
THE CONTENT OF PAHS IN SOILS OF LUBLIN (µG•KG -1

TABLE 3 .
COMPOSITION (%) OF INDIVIDUAL PAHS IN THE SOILS UNDER INVESTIGATION

TABLE 7 .
CORRELATION COEFFICIENTS BETWEEN THE ACTIVITY OF THE EXAMINED ENZYMES AND CONTENT OF Σ16PAHS (N=30). *