Monitoring chronic and acute PAH atmospheric pollution using transplants of the moss Hypnum cupressiforme and Robinia pseudacacia leaves

Atmospheric Environment 150 (2017) 45e54

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Monitoring chronic and acute PAH atmospheric pollution using transplants of the moss Hypnum cupressiforme and Robinia pseudacacia leaves F. Capozzi a, b, A. Di Palma b, c, P. Adamo c, V. Spagnuolo a, *, S. Giordano a
di Napoli Federico II, Via Cintia 4, 80126 Napoli, Italy Dipartimento di Biologia, Universita
di Napoli Federico II, Via Mezzocannone, 16, 80132 Napoli, Italy Centro Interdipartimentale di Ricerca Ambiente (CIRAM), Universita c
di Napoli Federico II, Via Universita
100, 80055 Portici (NA), Italy Dipartimento di Agraria, Universita a

b

h i g h l i g h t s
Relations between PAH contents in H. cupressiforme and R. pseudoacacia were studied.
R. pseudoacacia was able to accumulate LMW and HMW PAHs while moss prevalently HMW.
Locust tree combined chronic PAH inputs, while moss reflected PAH recently emitted.

a r t i c l e i n f o

a b s t r a c t

Article history: Received 1 July 2016 Received in revised form 16 November 2016 Accepted 17 November 2016 Available online 19 November 2016

Few studies are focused on correlations between the concentrations of PAHs in mosses and other bioindicator plant species. This study was carried out to investigate the potential of the joint use of devitalized H. cupressiforme transplants and R. pseudoacacia leaves as cost effective biomonitors for the assessment of PAHs in the air. The test was performed in a land historically devoted to agriculture, where recurrent waste burnings randomly occur, especially in the season we chose for the investigation. The presence of 20 PAHs was assessed following EPA 3550 C 2007 and EPA 8270 D 2014 protocols. R. pseudoacacia was able to accumulate both LMW and HMW PAHs, while moss prevalently collected the latter. It is suggested that R. pseudoacacia combined chronic pyrogenic and petrogenic PAH inputs, while moss transplants reflected PAH depositions from recent pyrogenic events. Our approach revealed long and short-term pollution footprints, with R. pseudoacacia recording the chronic input of PAH compounds loaded along its vegetative growth, and moss bags reflecting acute pollution inputs occurred during the exposure duration. © 2016 Elsevier Ltd. All rights reserved.

Keywords: Moss-bags Air biomonitoring Bioaccumulation Diagnostic ratios Locust tree

1. Introduction Polycyclic Aromatic Hydrocarbons (PAHs) are listed by US Environmental Protection Agency and the EU Directive 2004/107/ EC (US EPA, 1997; EU, 2005) among the contaminants of emerging environmental concern. These compounds consist of two to seven condensed aromatic rings; they can be of anthropogenic origin (mainly from fossil fuels burning), but they can also originate from natural sources such as volcanic eruptions and wild fires (Simonich and Hites, 1995). The transport pathways of PAHs in the

* Corresponding author. E-mail address: [email protected] (V. Spagnuolo). http://dx.doi.org/10.1016/j.atmosenv.2016.11.046 1352-2310/© 2016 Elsevier Ltd. All rights reserved.

atmosphere depend on their physical-chemical properties. The most volatile (with low molecular weight and number of rings LMW) remain in gaseous phase after their emission, while the heavier (high molecular weight and number of rings - HMW) are adsorbed on solid particulate matter. Due to the different partitioning between these two phases, lighter PAHs could be long range transported, while those with a higher molecular weight are generally deposited nearer to their emission sources (Thomas, 1986). Besides, environmental conditions such as temperature, UV radiation and wind dispersion can also affect their transport and persistence in the ambient air. To by-pass the difficulty of direct measures of PAHs in the air by conventional devices, many researchers proposed the use of vegetation to monitor the occurrence of such compounds (e.g.

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F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54

Augusto et al., 2013; Capozzi et al., 2016a; Di Palma et al., 2016; Harmens et al., 2013; Kodnik et al., 2015). The use of biomonitors in fact can give valuable indications about their presence and fate in the atmosphere, and in some cases, it remains the only feasible approach together with the passive samplers. The accumulation of PAHs in plants can occur through the uptake in their lipophilic compartments and the entrapment of particles, depending on the chemical physical state of PAHs and the physiology of plants. Mosses and tree leaves are among the most used biomonitors of atmospheric contaminants, including PAHs (e.g. Augusto et al., 2010; Çelik et al., 2005; De Nicola et al., 2016; Gjorgieva et al., 2011; Kaya et al., 2010; Vingiani et al., 2015); these two groups of plants have different physiology and water relations and have been used in a variety of locations and surveys (Bargagli, 1998; Harmens et al., 2013; Vukovi c et al., 2015). In some instances, the insufficiency or even absence of suitable moss species living in the most impacted environs as urban, agricultural or industrial areas has encouraged the use of moss transplants to monitor airborne PAHs (Anicic et al., 2009; De Nicola et al., 2013a,b; Giordano et al., 2013). At the present, only few papers considered the use of moss transplants to monitor the atmospheric deposition of PAHs, also due to the lack of well-defined methodological procedures for moss bag ~a exposure (Capozzi et al., 2016b) and moss analyses (Concha Gran et al., 2015). In a recent review, Harmens et al. (2013) highlighted that even less studies concern the relations between concentrations of PAHs in mosses and vascular plants.

Table 1 Experimental design and geographical coordinates of exposure/collection sites. Site N

Municipality

Est-UTM 33T

Nord-UTM 33T

Exposure/presence

Site Site Site Site Site Site Site Site Site Site Site Site Site

Acerra

447868.83 442692.23 442737.94 443889.73 436395.58 425898.48 424814.67 440932.51 440441.03 435041.75 478028 478220 479954

4536987.28 4533502.09 4535595.45 4536913.49 4529609.49 4533193.47 4533282.63 4540446.14 4540552.11 4530032.13 4541711 4541873 4541495

M/Ra M/R M/R M/R M/R M/R M/R M M M/R R R R

a

1 2 3 4 5 6 7 8 9 10 RB1 RB2 RB3

Caivano Casandrino Giugliano Marcianise Melito Arpaise

M ¼ H. cupressiforme transplants, R ¼ naturally growing R. pseudoacacia.

The illegal waste burning is a deplorable practice often prevailing in peri-urban lands where the waste cycle is not properly concluded; it is known to release a large amount of gaseous and particulate PAHs, for its nature of uncontrolled and random phenomenon. It is hard to monitor the presence in the atmosphere of these pollutants due to their rapid diffusion, decomposition and transformation (Ravindra et al., 2008). In the present paper, in a jeopardized urban/rural area (di Gennaro, 2014) we investigated the joint use of two different biomonitoring approaches: the leaves of autochthonous Robinia pseudacacia L. trees, and the moss Hypnum cupressiforme Hedw. Transplanted in bags, with the aim of answering the following questions: 1) Do mosses and higher plant leaves provide similar information regarding airborne PAH levels? 2) Do the two biomonitors behave differently in the enrichment of specific PAHs? 3) Is it possible, analyzing the content of PAHs in mosses and leaves, to detect the transient occurrence of waste burning?

2. Materials and methods 2.1. Study area The study area falls in a land historically devoted to agriculture and located at north of the metropolitan area of Naples. Nowadays this area is characterized by a complex and intermingled mixture of urban, agricultural and industrial settlements and recently it has been under the media attention as the land of fires due to the effects of a waste crisis compromising the environmental quality. Recurrent waste burnings randomly occur in the area and sometimes are denounced by citizens. ARPAC (Regional agency for environment protection) has attempted to realize an inventory of the fires, which represents only a limited part of the phenomenon. For instance, in the first seven months of 2015, 115 fires were recorded, prevalently during the summer months (ARPAC, 2015). 2.2. Experimental design, bags preparation and exposure, and R. pseudoacacia leaves collection Following the exposure protocol adopted by Capozzi et al. (2016a), sub-spherical moss bags, filled with devitalized moss

P P Fig. 1. PAHs, grouped by number of rings, measured in H. cupressiforme moss-bags in pre-exposure (T0) and post-exposure (M1e10) materials. Pale-grey bars ¼ 2e3 rings, P P grey bars ¼ 4 rings, black bars ¼ 5e6 rings.

Hypnum cupressiforme Hedw. With a weight of moss/bag surface ratio of 10 mg/cm2 (Capozzi et al., 2016b), were exposed in ten sites (Site 1e10) for six weeks (see Table 1 for details). The moss was collected in a pristine area, the Taburno-Camposauro Regional Park (1000 m.a.s.l. - 464,013 m E 455,805 m N, 33T UTM). Three bags were exposed at each exposure site for a total of 30 samples. All the bags were exposed for six weeks starting from July 2015. Ten samples of devitalized, unexposed moss were analyzed to assess baseline PAH contents. This was necessary to determine the after exposure enrichment of moss tissue, as the algebraic sum of the specific PAH contents. Robinia pseudoacacia L., is a tree species naturalized in all temperate regions of North America, Europe and Southern Africa, and it is well adapted to dry and polluted soils (Tzvetkova and Petkova, 2015). Leaves of R. pseudoacacia were collected at each exposure site (within a range of 0e120 m from the moss bags), with the exceptions of the sites 8 and 9 where the target species did not grow; leaves from three background sites (RB-1, 2, 3) were also collected and analyzed to compare data from sites with different anthropogenic impact (Table 1). The leaves were collected at the end of their vegetative season (November 2015, after about seven growth months), each sample was composed of ten leaves collected all around the canopy at a height ranging between 4 and 5 m from ground. All the samples were stored in a zip aluminum bag, to avoid the direct contact with sunlight, and placed in a cool bag to reduce the loss of the lighter PAHs; once in the laboratory the samples were kept in the fridge at a temperature of 4  C until analyzed. 2.3. PAH analyses For PAH analyses we adopted the protocols EPA 3550 C 2007 and EPA 8270 D 2014. Two grams of moss samples (obtained combining 4 bags for each replica) and 2 grams of R. pseudoacacia (after homogenization and milling) were sonicated (Falc Sonicator) for two times, each in 25 mL of dichloromethane for 20 min each. The extracts were purified through activated silica gel and dried to a volume of 200 mL under a gentle nitrogen stream. Consecutively the samples were analyzed by GCeMSD (Agilent 5975C with a VF-17MS column) with helium as gas carrier at 1.3 mL min1. The oven temperature program started at 50  C, increased with ramp rate 30  C min1, to 350  C and held for 9 min. All analyses were performed in selected ion monitoring (SIM). The concentration of the following 20 PAHs were quantified by multi-point calibration curves and labelled internal standards; (2rings): naphtalene (Naph); (3-rings): acenaphthene (Ace), fluorene (Flu), phenanthrene (Phen), anthracene (Ant); (4-rings): fluoranthene (Flt), pyrene (Pyr), benz[a]anthracene (B[a]A); chrysene (Chrys), (5-rings): benzo[b]fluoranthene (B[b]F), benzo[k]fluoranthene (B[k]F), benzo[j]fluoranthene (B[j]F), benzo[a]pyrene (B [a]P), dibenz[a,h]anthracene (DB[a,h]A), dibenzo[a,i]pyrene (DB[a,i] P), dibenzo[a,h]pyrene (DB[a,h]P), dibenzo[a,e]pyrene (DB[a,e]P), dibenzo[a,l]pyrene (DB[a,l]P), (6-rings): benzo[g,h,i]perylene (B [g,h,i]P) indeno[1,2,3-c,d]pyrene (IP). For the quality control of the procedure, labelled PAHs (naphthalene D8, acenaphtene D10, phenanthrene D10, chrysene D12, perylene D12) were used as surrogates and the percentage recoveries (from 82% to 120%) were included to correct the concentration of each compound. The minimum detectable PAH concentration was 1 ng g1 d.w. for each compound. 2.4. Data analysis Basic statistics and figure elaborations were obtained by using Microsoft Office Excel 2010 and the software STATISTICA (StatSoft, 2008). The significance of PAH accumulation was evaluated by

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Fig. 2. Individual PAH contents, grouped by number of rings, in the pre (T0) and post exposure (M1e10) H. cupressiforme moss-bags.

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Fig. 3. PCA for the pre exposure (T0) and exposure sites (M1e10) based on the PAH contents of H. cupressiforme moss-bags: a) projection of the cases (n ¼ 11); b) projection of the variables (n ¼ 14).

F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54

comparing pre-exposure to post-exposure values following Couto et al. (2004) as modified in Ares et al. (2015). Some of the investigated PAHs were below quantification limit (BQL) in pre-exposure moss samples as well as in some sites after exposure; to assess the enrichment for these compounds it was used, as T0 value, the QL/2. The correlations among the PAHs within each biomonitor, and between the PAH profiles of the two biomonitors was evaluated by Spearman test. The Wilcoxon matched pairs test was used P to assess the differences between the content of PAHs, grouped by number of rings, accumulated in the two biomonitors. 3. Results and discussion 3.1. PAHs accumulation in moss transplants The concentrations of the 16 PAHs (mean values ± standard deviations) found in the moss samples are reported in Table S1. The total PAH content in the unexposed moss was 176.75 ng g1, while in the post-exposure samples the total content ranged between 79.53 and 294 ng g1, sites 3 and 8 respectively (Fig. 1). The postexposure total content was in general lower than the preexposure one, except for site 8. However, after the exposure, the PAH profiles drastically changed; in the exposed moss we found a higher content of HMW PAHs (4e6 rings) and an overall decrement of the lighter PAHs (2e3 rings). Indeed, in the unexposed moss the P P P 2e3 rings, 4 rings and 5e6 rings PAHs, were respectively 1 142.8, 26.8 and 7.2 ng g , while in the post-exposure moss they ranged between 27.7 and 51.9 ng g1 (site 3 and 9), 40.7e169.7 ng g1 (site 3 and 8) and 10.1e81.6 ng g1 (site 10 and 8), respectively (Figs. 1 and 2). In particular, DB(a,i)P, DB(a,h)P, DB(a,e)P, DB(a,l)P, were BQL both in T0 and in all exposed samples. Benzo(a)antracene, B(a)P, DB(a,h)A were BQL in T0 samples; the concentration of Naph, Ace, Flu, Phen and Ant were above quantification limit (AQL) in T0 samples but were never enriched in the exposed mosses. Benzo(a) antracene was accumulated in all ten exposure sites, while the B(a) P only in five sites (2, 3, 5, 8, 9); the DB(a,h)A was AQL and enriched only in the moss exposed in the site 8. Fluoranthene was accumulated in sites 2, 4 and 8, B[k]F in four sites (2, 5, 8, 9), and IP was accumulated in six sites (1, 2, 5, 6, 8, 9). The carcinogenic B(a)P was absent in T0 samples, and was accumulated in sites 2, 3, 5, 8 and 9. The remaining PAHs (Chrys, Pyr, B(b)F, B(j)F, B(g,h,i)P) were always

49

AQL and found in all the exposure sites. In the moss after exposure, significant correlations (Spearman Rank Order Correlation, p < 0.05) were found mainly among PAHs with 4e6 rings suggesting for them a common origin and similar diffusion pathways (Table S2a). Fig. 3a shows the PCA of the exposure sites classified according to PAH load in moss transplants. The two principal components account for about 94% of the total variance (Factor 1: 70.5%; Factor 2: 23.5%). All post-exposed moss samples (sites 1e10) are evidently separated from the pre-exposed moss (T0) according to their different PAH load and profiles characterized by prevalence of high- and low-molecular weight compounds, respectively (Fig. 3b). Sites 8 and 2 are the most impacted, while all the others are grouped together. Many authors report that native mosses gathered in natural or rural environments contain mainly LMW PAHs (2e3 rings) (Gerdol et al., 2002; Otvos et al., 2004). In our experiment the pre-exposure PAH profile (referring to moss H. cupressiforme collected from an unpolluted area and devitalized) was dominated by LMW PAHs. After the exposure, PAH profile changed and the HMW PAHs accounted for the majority of the total load. Some authors hypothesize that the 2e3 ring PAHs can be produced by natural as well as anthropogenic sources (Liu et al., 2005; Ravindra et al., 2008 and references therein); in addition, since they are the most volatile among PAHs, they can also be found in sites far from emission sources, as pristine areas are (Harmens et al., 2013; Ravindra et al., 2008; Van Jaarsveld et al., 1997). Forest canopy cover, dim light and low temperature can affect the residence of 2e3 ringed PAHs in the natural environment where they accumulated in the mosses collected for transplants. Our findings, for instance, can be also due to the low temperature and high altitude characterizing the control site, and favoring LMW PAH persistence in the atmosphere by cold condensation (Kallenborn, 2006; Liu et al., 2005). On the contrary, the low content of the 2e3 ringed PAHs in moss transplants, can be likely explained by their degradation during exposure due to sun radiation and high temperature (Ravindra et al., 2008), two conditions affecting the moss transplanted in bags especially during the summer months, when the survey was carried out. The lack of accumulation of LMW PAHs during exposure is likely related to i) moss characteristics (i.e., single-cell layered leaves, absence of cuticle) hampering their preservation in moss tissues, and ii) moss devitalization, determining accumulation only by passive mechanisms. However, we recently tested

P Fig. 4. PAHs grouped by number of rings contained in R. pseudoacacia leaves from background sites (R-B ¼ average RB 1e3) and tested sites (R1e10, except sites 8 and 9 where P P P R. pseudoacacia was absent). White bars ¼ 2e3 rings, grey bars ¼ 4 rings, black bars ¼ 5e6 rings.

F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54

four living moss species for phenanthrene (a 3-ringed PAH) uptake in a laboratory trial. We found that phenanthrene never entered living cells and was uptaken as micron-sized particles only by passive mechanisms (see Augusto et al., 2015; Spagnuolo et al., 2016). The greater amount of HMW PAHs (4e6 rings) in moss after exposure can be justified by the following reasons: i) the continuous production and residence of these compounds in the tested sites; ii) their resistance to degradation and chemical reaction; iii) their association to particulate matter, whose interception and trapping is considered the dominant uptake mechanism in mosses (Boileau et al., 1982; Tretiach et al., 2011).

3.2. PAH accumulation in R. pseudoacacia leaves The concentrations of the 14 PAHs (mean values ± standard deviations) found in R. pseudoacacia leaves are reported in Table S3. Acenaphthene, DB(a,i)P, DB(a,h)P, DB(a,e)P, DB(a,l)P and DB(a,h)A were BQL in all samples. The mean total PAH content in the leaves collected from background sites (RB-1, 2, 3) was 118.1 ng g1, while it ranged between 112.8 and 157.5 ng g1 (sites 3 and 6 respectively) in the samples collected from all the other sites. The leaves from the background sites had in general a lower total PAH content than those harvested from the tested sites, except for sites 2 and 3 (Fig. 4). In background P P P sites the 2e3 rings, 4 rings and 5e6 rings PAHs, were 1 respectively 90.3, 21.4 and 6.4 ng g , while in the tested sites they ranged between 65.7 and 91.7 ng g1 (sites 7 and 10), 29.9e53.1 ng g1 (sites 3 and 5) and 8.2e22.1 ng g1 (sites 2 and 5), respectively. In R. pseudoacacia collected from the tested sites we found a general increment of HMW PAHs (4e6 rings) respect to the background sites. This was not observed for the light PAHs (2e3 rings), displaying a profile qualitatively and quantitatively similar for background and tested sites (Fig. 5). It is worth to note that B(a)P was present in all the samples, with content in leaves from background sites generally lower than in the other sites. Wild fires regularly affecting the forests with a Mediterranean climate could be the source of B(a)P in background sites (Olivella et al., 2006). In R. pseudoacacia, excluding background sites, significant correlations (Spearman Rank Order Correlation, p < 0.05) were found especially among HMW PAHs (Table S2b); as previously described for moss, this finding could be explained by their common origin and residence in the environment. The PCA of the sampling sites classified according to PAH load in R. pseudoacacia leaves is given in Fig. 6a. The first two factors account for about 78% of the total variance (Factor 1: 60.8%; Factor 2: 12.3%). All the tested sites (1e7 and 10) are separated from the background sites (R-Bk ¼ average of RB-1, 2, 3). Also in this biomonitor a prevalence of LMW compounds was found in the samples from background sites while HMW PAHs prevailed in the leaves from the tested sites (Fig. 6b). The most impacted sites were 5 and 6, being the others homogeneously enriched. The PAH profiles found in control and tested sites were similar; the 2e3 ringed PAHs account always for at least 50% of the total load; the 4e6 ring PAHs showed an increasing trend from background areas to tested sites. It is possible that the adsorbed LMW PAHs can enter the leaves due to their affinity for hydrophobic compounds of the leaf cuticle and be stored in the cells where they are protected by photo-degradation (Chen et al., 2005; Domingos et al., 2015; Wang et al., 2014).

Fig. 5. Direct comparison, based on individual PAH contents grouped by number of rings, among the R. pseudoacacia leaves from background sites (R-B ¼ average RB 1e3) and tested sites (R1e10, except sites 8 and 9 where R. pseudoacacia was absent).

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F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54 Fig. 6. e PCA for the collection sites of the R. pseudoacacia leaves based on the PAH contents: a) projection of the cases (n ¼ 9); b) projection of the variables (n ¼ 14). (R-B ¼ average RB 1e3 and R1e10, collection sites; in sites 8 and 9 R. pseudoacacia was absent).

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F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54

3.3. PAH source apportionment: H. cupressiforme transplants vs. R. pseudoacacia leaves Many factors can contribute at different extent to PAH accumulation depending on the specific compound and the biomonitors. Fig. 7 shows the total content of PAHs, grouped by number of rings, measured in H. cupressiforme moss-bags and R. pseudoacacia leaves. Light PAH (2e3 rings) load in R. pseudoacacia was about two times that observed in H. cupressiforme transplants, p < 0.05 (Fig. 7a); this trend reverts considering the 4 rings PAHs (Fig. 7b) for which, in all sites, the contents in moss were higher than in R. pseudoacacia, p < 0.05. A more heterogeneous situation occurs when considering the 5e6 rings PAHs whose accumulation was in some cases higher in moss and in some others in leaves, p > 0.05. Indeed, the PAH profiles in the two biomonitors are never correlated (p > 0.05), confirming the different accumulation capability of the two biological matrices. The different abilities of the two biomonitors to uptake PAHs reflect their morphological and physiological traits: R. pseudoacacia can incorporate LMW PAHs and store them inside the cuticle and multi-cell layered leaves, avoiding their degradation, while devitalized moss prevalently entraps and accumulates, by passive mechanisms, HMW PAHs bound to particulate matter. The fate of PAH compounds in plants is ruled by their presence in the atmosphere, uptake and storage mechanisms inside leaf cells and by the environmental conditions such as temperature and radiation affecting their photodegradation (Wild et al., 2005). To look for the possible PAH sources, the diagnostic ratio method is often used (a review in Tobiszewski and Namiesnik, 2012). Although this method is potentially very helpful, it has to be applied with caution; indeed, the ratios can be altered by different reactivity of some PAH species with other compounds; in addition, the degradation, chemical reactivity, volatility, and solubility of the different PAHs can affect the results (Tsapakis and Stephanou, 2003). Moreover, some of these ratios were developed on a specific matrix and consequently are not always applicable to other media. Finally, considering the PAH markers, some degree of overlap between the profiles from different emission source categories is possible (Ravindra et al., 2008). To overcome some of the above-mentioned limitations, the diagnostic ratios calculated with PAHs having similar physicochemical properties should be preferred (Ravindra et al., 2008). Accordingly, we applied only diagnostic ratios considered appropriate for our experimental design and matrices. P P Zhang et al. (2008) proposed the ratio LMW/ HMW as an appropriate marker to distinguish between pyrogenic and petrogenic sources. In our study, for moss this ratio is above 1 only in the unexposed material (T0), while it is always below 1 in all the tested sites, suggesting petrogenic derivation of PAHs found in unexposed moss and pyrogenic derivation of PAHs measured in the exposed moss. Accordingly, the ratio Flt/Pyr þ Flt (Yunker et al., 2002) in T0 indicates petrogenic derivation of PAHs, while in all the exposed moss a pirolytic derivation of PAHs (Fig. 8a). To reinforce our hypothesis of recent burning events mainly affecting PAH profile in moss bags, we found a joint presence of B(a) P and IP in sites 2, 5, 8 and 9, probably deriving from illegal wastetire fires (Chen et al., 2007), a dreadful common practice in our study areas, known as the land of fires; moreover, in the same sites, the dominance of chrysene and benzo[k]fluoranthene might be indicative of coal combustion (Ravindra et al., 2008). The presence of Chry is also associated with biomass burning (Simoneit, 2002). This PAH was always accumulated in both plant matrices, with an increasing load in the tested sites compared with the reference areas. P P In R. pseudoacacia the ratio LMW/ HMW was always >1,

suggesting a petrogenic derivation of the PAH content, except for the site 5 (ratio <1); the prevalence of LMW-PAHs was suggested as indicative of petrogenic sources (Wu et al., 2014; Dias et al., 2016). In contrast, the ratio Flt/Pyr þ Flt was always >0.5 indicating pyrogenic derivation (Fig. 8b). This apparent contradiction might depend by: i) R. pseudoacacia reflects PAH depositions over a long time interval corresponding to its vegetative season (about 7 months), combining pyrogenic and petrogenic emissions; ii) R. pseudoacacia is able to accumulate and protect from degradation the LMW PAHs affecting the ratio LMW/ HMW. On the contrary, the uptake in moss transplants refers only to the exposure duration (6 weeks in this experiment), reflecting PAH depositions from recent pollution events, accumulating, storing and mirroring mainly the HMW PAHs, chemically more stable. A similar outcome was observed in a previous paper (De Nicola et al., 2013a), in which the authors reported that leaves of evergreen holm oak (Quercus ilex L.) and moss transplants (H. cupressiforme) provided different PAH profiles in a survey devoted to evaluate PAH distribution in an urban street canyon. The different PAH profile of H. cupressiforme collected in background site (T0) compared to that observed in the exposed moss can be explained by their different time scale, probably reflecting in the native moss a diffuse and chronic pollution mainly of longrange petrogenic origin, according to the two calculated ratios (Fig. 8a). The use of autochthonous mosses as suitable organisms to monitor spatial patterns and temporal trends of atmospheric concentrations or deposition of POPs was recently studied in deep (Harmens et al., 2013). The diffuse and chronic pollution by PAHs, P P emerging from the ratio LMW/ HMW interpretation, might induce to an erroneous perception of the quality of the site chosen as background, thus biasing the interpretation of the results of this

P Fig. 7. Comparison between the PAHs content (grouped by number of rings) measured in H. cupressiforme moss-bags and R. pseudoacacia leaves from exposure/ tested sites.

F. Capozzi et al. / Atmospheric Environment 150 (2017) 45e54

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P P P P Fig. 8. PAH cross plots for the ratio of low molecular weight/ high molecular weight PAHs ( LMW/ HMW) to fluoranthene/fluoranthene pyrene (Flt/Pyr þ Flt), for (a) H. cupressiforme moss-bags and (b) R. pseudoacacia leaves.

study. However, when we calculate ratios, it must be always taken into account that similar ratios can stem from different absolute values. In our case PAH values for the background sites were always lower than those measured in the exposure sites, proving their character of low impacted reference areas. Therefore, the PAH profiles in R. pseudoacacia describe the chronic input of these pollutants over the investigated area of both pyrogenic (e.g. waste burning, open agricultural burning) and petrogenic origin (e.g. domestic heating, vehicular traffic, power plants), while moss transplants reflect recent acute pollution inputs, mainly of pyrogenic derivation and particularly frequent during the summer months (i.e. during the exposure period) (Fig. 8).

4. Conclusions The experiment carried out in this study demonstrated the potential of devitalized H. cupressiforme transplants and R. pseudoacacia leaves as cost-effective biomonitors for the evaluation of PAH presence in the air. Our approach suggests long- and short-term pollution footprints, with R. pseudoacacia recording the chronic input of PAH compounds loaded along its vegetative growth, and moss transplants reflecting acute pollution inputs during the exposure duration. In future studies, a similar trial, comparing the two biomonitors with passive samplers would be useful to substantiate these outcomes. Robinia pseudoacacia was able to accumulate and store both LMW and HMW PAHs, while moss prevalently accumulated the latter. In agreement with their specific morphological and physiological characteristics, and according to PAH profile and specific diagnostic ratios, R. pseudoacacia combined chronic pyrogenic and petrogenic PAH inputs, while moss transplants reflected PAH depositions from recent pollution events, pyrogenic in our survey. Moss bags exposed for 6 weeks confirm to be suitable biomonitors to detect pollution inputs, even considering PAHs. Although their use represents the only option in biomonitoring when suitable autochthonous species are lacking, longer or subsequent exposures would be tried to attempt the reconstruction of chronic PAH pollution. Therefore, at the state, the joint use of

autochthonous trees and moss transplants should be preferred to obtain a more comprehensive environmental information. Further studies are needed to understand the different PAH accumulation dynamics of higher plants and moss transplants (devitalized and alive) and to explore the possibilities (e.g. different plant species, seasonality, land use) of their joint use in view of the proposal of a reproducible biomonitoring protocol. Acknowledgments The present study was financed by the EU Commission, Project: LIFE/11/ENV/IT/275ECOREMED “Implementation of ecocompatible protocols for agricultural soil remediation in Litorale Domizio-Agro Aversano NIPS”. Appendix A. Supplementary data Supplementary data related to this article can be found at http:// dx.doi.org/10.1016/j.atmosenv.2016.11.046. References Ani ci c, M., Tomasevi c, M., Tasi c, M., Rajsi c, S., Popovi c, A., Frontasyeva, M.V., et al., 2009. Monitoring of trace element atmospheric deposition using dry and wet moss bags accumulation capacity versus exposure time. J. Hazard. Mater. 171, 182e188. Ares, A., Aboal, J.R., Carballeira, A., Fernandez, J.A., 2015. Do moss bags containing devitalized Sphagnum denticulatum reflect heavy metal concentrations in bulk deposition? Ecol. Indic. 50, 90e98. Augusto, S., Maguas, C., Matos, J., Pereira, M.J., Branquinho, C., 2010. Lichens as an integrating tool for monitoring PAH atmospheric deposition: a comparison with soil, air and pine needles. Environ. Pollut. 158, 483e489. Augusto, S., Pereira, M.J., Maguas, C., Branquinho, C., 2013. A step towards the use of biomonitors as estimators of atmospheric PAHs for regulatory purposes. Chemosphere 92, 626e632. Augusto, S., Sierra, J., Nadal, M., Schuhmacher, M., 2015. Tracking polycyclic aromatic hydrocarbonsin lichens: It’s all about the algae. Environ. Pollut. 207, 441e445. Bargagli, 1998. Trace Elements in Terrestrial Plants. An Ecophysiological Approach to Biomonitoring and Biorecovery. Springer-Verlag, Berlin, p. 324. Boileau, L.J.R., Beckett, P., Lavoie, P., Richardson, D.H.S., 1982. Lichens and mosses as monitors of industrial activity associated with uranium mining in Northern Ontario, Canada. Part 1. Environ. Pollut. 4, 69e84. Capozzi, F., Giordano, S., Di Palma, A., Spagnuolo, V., De Nicola, F., Adamo, P., 2016a. Biomonitoring of atmospheric pollution by moss bags: discriminating urban-

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