Environmental Pollution 196 (2015) 53e59
Contents lists available at ScienceDirect
Environmental Pollution journal homepage: www.elsevier.com/locate/envpol
Lipid-content-normalized polycyclic aromatic hydrocarbons (PAHs) in the xylem of conifers can indicate historical changes in regional airborne PAHs Yuan-wen Kuang*, Jiong Li, En-qing Hou Key Laboratory of Vegetation Restoration and Management of Degraded Ecosystems, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, PR China
a r t i c l e i n f o
a b s t r a c t
Article history: Received 11 July 2014 Received in revised form 7 September 2014 Accepted 13 September 2014 Available online
The temporal variation of polycyclic aromatic hydrocarbons (PAHs) concentrations as well as the lipid content in the xylem of Masson pine trees sampled from the same site were determined and compared with the days of haze occurrence and with the historical PAHs reported in sedimentary cores. The patterns of the lipid content as well as the PAH concentrations based on the xylem dry weight (PAHsDW) decreased from the heartwood to the sapwood. The trajectories of PAHs normalized by xylem lipid content (PAHs-LC) coincided well with the number of haze-occurred days and were partly similar with the historical changes in airborne PAHs recorded in the sedimentary cores. The results indicated that PAHs-LC in the xylem of conifers might reliably reflect the historical changes in airborne PAHs at a regional scale. The species-specificity should be addressed in the utility and application of dendrochemical monitoring on historical and comparative studies of airborne PAHs. © 2014 Elsevier Ltd. All rights reserved.
Keywords: Dendrochemistry Historical change Lipid-content-normalization Masson pine Polycyclic aromatic hydrocarbons (PAHs) Xylem
1. Introduction The drastic emissions from anthropogenic activities have led to the great abundance of polycyclic aromatic hydrocarbons (PAHs) in environments (Zhang and Tao, 2009). Monitoring of airborne PAHs has arisen a global concern for the carcinogenic, mutagenic, and € m et al., 2002). teratogenic effects of PAHs on human beings (Bostro Conventionally, routine PAHs were monitored using high-volume air samplers which were time-consuming and expensive. Alternatively, spatial and temporal PAHs were bio-monitored via “passive samplers”, which included vascular and non-vascular plants (Prajapati and Tripathi, 2008; Augusto et al., 2009). Needles of conifer species, especially pine species such as Pinus sylvestris (Holoubek et al., 2000), P. densiflora and P. maximartinezii (Hwang et al., 2003), P. massoniana (Tian et al., 2008), P. nigra, P. pinaster (Ratola et al., 2010, 2011) and P. halepensis (Ratola et al., 2012), have been frequently adopted to bio-monitor the current levels of
* Corresponding author. Present address: Xingke Road #723, Tianhe District, Guangzhou 510650, China. E-mail addresses: [email protected] (Y.-w. Kuang), [email protected] (J. Li), [email protected] (E.-q. Hou). http://dx.doi.org/10.1016/j.envpol.2014.09.018 0269-7491/© 2014 Elsevier Ltd. All rights reserved.
airborne PAHs, due to their wide distribution and their high lipid contents. Lacking the long-term monitoring data in most regions, researchers have accessed to relatively scarce reports on regional and historical changes in airborne PAHs via the analysis on treerings of bark pockets (Wang et al., 2004), sedimentary cores (Liu et al., 2005; Guo et al., 2011), and ice cores (Wang et al., 2008). Dendrochemistry is a subject that traces the historical changes in environmental pollution based upon the chemical analysis of the growth rings of trees (xylem). It assumed that the pollutants “recorded” in the xylem were immobile, thus, the levels of the studied pollutants in the growth rings were indicative of their real status in the environments when the growth rings formed (Nabais et al., 1999). Despite the doubt of concerning the efficiency on dendrochemistry (Bellis et al., 2002), researchers have used it to trace the regional history of heavy-metal emissions (Punshon et al., 2005), nitrogen pollution (Savard et al., 2009;Doucet et al., 2012) and soil acidification (Kuang et al., 2008), which were highly consistent with some detailed history events. To our knowledge, there have been relatively few reports on dendrochemistry tracing the historical changes in airborne PAHs, except a study of Wang et al. (2004), who selected a broadleaved tree (Cinnamomum camphora, approximately 10e15 years old) to investigate the accumulation, distribution and translocation of PAHs in the xylem,
Y.-w. Kuang et al. / Environmental Pollution 196 (2015) 53e59
400 D1 D3 D10
300 200 100
2006-2010
2001-2005
1996-2000
1991-1995
1986-1990
1981-1985
1976-1980
1971-1975
1966-1970
0 1961-1965
Xylem of Masson pines used for PAHs and lipid detection were sampled from Dinghushan natural reserve, which located in the northwest of PRD of southern China. This reserve has a lowsubtropical monsoon climate and tropical/subtropical forest and has experienced long-term exposure to atmospheric pollutants transported by the southwest prevailing wind in the summer from the numerous industries and vehicles in PRD economic zone. The atmospheric PAHs in this reserve were reported mainly from the anthropogenic emissions in PRD (Qi et al., 2001). Taking the opportunities of biomass investigation conducted by Dinghushan Forest Ecosystem Research Station, Chinese Academy of Sciences, we sampled two (D1, D3) and one (D10) discs from the cut-down model trees of Masson pine at the end of 2005 and 2010, respectively. The trees (about 40 cm in diameter without visible injury in the needles or a reduction in crown density) were selected in the forest plots far away from any roads. In order to avoid the contamination of external PAHs, the disc (about 20 cm in thickness) was cut from the stem of the model tree at breast height above ground (~1.3 m) with man-powered saw and transported to the laboratory immediately for processing. In the laboratory, the disc was mounted and polished to reveal the annual growth rings and then dated with the Windendro™ system (V 6.1D, Canada). Fresh chips of the xylem representing fiveyear intervals were carefully sliced from the heartwood to the sapwood (one chip per time interval) with a clean, electric microchisel, to obtain sufficient mass for PAHs analysis. A total of 34 chips (11 from both D1 and D3, 12 from D10) representing about a 60-year history (1950e2010) was sliced from the disc. The chips
About 8 g powder of the xylem chips (dry weight, DW) was extracted and purified for PAHs analysis. The process of the extraction, purification and analysis was described elsewhere (Tian et al., 2008; Wang et al., 2004). A Dionex ASE 300 (Dionex Co., USA) was used for PAHs extraction. The powder was mixed with 5 g of anhydrous sodium sulfate (barked for 4 h at 600 C in oven before use) and packed in a 33 mL stainless extracting cell. The samples were spiked with a known aliquot of naphthalene-d8, acenaphthane-d10, phenanthrene-d10, chrysene-d12 and perylene-d12 as analyte surrogates. The extraction used Dichloromethane (DCM) and acetone (1:1, v/v) as solvent under the conditions of 1500 psi and 100 C. The ASE was conducted for two cycles, each lasting for 5 min of heating time and 5 min of static extraction time. Activated copper slices were added to the extract to remove elemental sulfur. The xylem extract was concentrated to 1 mL with a rotary evaporator at 30 C and then loaded on a multiple layer chromatographic column packed with 2 g anhydrous sodium sulfate, 5 g aluminum oxide (extracted by DCM for 72 h, heated for 12 h in 250 C), 5 g florisil (baked for 8 h at 450 C in an oven) and 10 g silica gel (extracted by DCM for 72 h, heated for 12 h in 150 C), and eluted with 60 mL of DCM. The eluent was evaporated and solventexchanged to 1 mL of hexane. A gel permeation chromatography (GPC) column (10 mm i.d., packed with 10 g of S-X3Biobeads, dipped by dichloromethane in advance, Accustandard Co., USA) was used to eliminate lipids. The GPC column was eluted with 80 mL n-hexane: DCM (1:1 v/v) at a flow rate of 0.5 mL min1. The first 35 mL eluent was discarded, and the following 45 mL which contained PAHs were collected and concentrated to a final volume of 200 mL under a gentle stream of nitrogen. PAHs compounds were separated on a 30 m 0.25 mm i.d. HP-5 capillary column (film thickness 0.25 mm) and quantified using a HewlettePackard 5890 gas chromatography and 5972 mass selective (GC-MSD) detector operated in the electron impact mode (70 eV). The instrumental conditions were as follows: injector temperature, 280 C; ion source temperature, 180 C; temperature program: 60 C (2min), 60e290 C at 3 C min1, 290 C (30 min). The carrier gas was helium at a constant flow rate of 1.5 mL 30 m1. 1 mL sample was injected in splitless mode. Mass range m/z 50e500 was used for quantitative determinations. Data acquisition and processing was
1956-1960
2.1. Xylem sampling and processing
2.2. PAHs analysis
1951-1955
2. Materials and methods
were freeze-dried for 72 h and finely ground to pass through a 0.2mm screen.
-1
based on the physiological features of trunk tissues, the physicochemical properties of PAHs, and the correlations between PAHs concentrations in the tissues. They then reconstructed a 150-year airborne PAHs history using the concentrations of PAHs in the bark pockets of another broadleaved tree (Fagus longipetioleta, about 150 years old). Recommended by researchers, ringeporous species (i.e. conifers species) was more suitable for dendrochemical analysis than diffuseeporous ones (i.e. broadleaved species) (Bellis et al., 2002). Considering the differences in chemical properties between conifers and broadleaved species, we put forward the following questions: 1) Were the distribution patterns of PAHs in xylem related to the chemical properties of conifers? 2) Could the patterns of PAHs in conifer xylem indicate historical changes in regional airborne PAHs like those in tree-rings of bark pockets and in sedimentary cores? To address these questions, we analyzed 16 EPA PAHs and the lipid contents in the xylem of Masson pine (P. massoniana L.) trees, the most representative predominant species in southern China, from Dinghushan natural reserve in the Pearl River Delta (PRD). The number of haze days occurred in PRD and the historical data of airborne PAHs reconstructed from the analysis on the sedimentary cores were also used to analyses the correlation with the xylem PAHs. The specific objectives were: 1) to detect whether the temporal patterns of PAHs concentration and lipid content occurred synchronously in the xylem of Masson pines; 2) to reveal whether the concentration of PAHs was related to the content of lipids in the xylem; and 3) to determine the potential of lipid-content-normalized PAHs in the xylem of Masson pine to indicate the historical changes in airborne PAHs at a regional scale. The results were expected to improve the reliability of tracing historical changes in airborne PAHs using dendrochemistry.
Lipid content: mg. g
54
Period
Fig. 1. Temporal distribution of lipid in the xylem of the three Masson pines (D1, D3 and D10) from Dinghushan natural reserve. The xylem from trees D1 and D3 was formed from 1951 to 2005; the xylem of tree D10 was formed from 1951 to 2010.
Y.-w. Kuang et al. / Environmental Pollution 196 (2015) 53e59
controlled by HP Chemstation software. The concentrations of 16 EPA PAHs: naphthalene (Nap), acenaphthylene (Acpy), acenaphthene (Acp), fluorene(Flu), phenanthrene (PA), anthracene (Ant), fluoranthene (FL), pyrene (Pyr), benz[a]anthracene (BaA), chrysene (Chr), benzo[b]fluoranthene (BbF), benzo[k]fluoranthene (BkF), benzo[a]pyrene (BaP), dibenz[a,h]anthracene (DBA), indeno[1,2,3cd]-pyrene (IND) and benzo[g,h,i]-perylene (BghiP), in the chips were analyzed. Samples, including laboratory blanks only containing solvent and recovery standards (SRM 1649A, NIST, Gaitherbury, USA), were processed in the same way for quality control.
55
The relative percentage difference for individual PAH identified in paired duplicate samples (n ¼ 2) was always
Contents lists available at ScienceDirect
Environmental Pollution journal homepage: www.elsevier.com/locate/envpol
Lipid-content-normalized polycyclic aromatic hydrocarbons (PAHs) in the xylem of conifers can indicate historical changes in regional airborne PAHs Yuan-wen Kuang*, Jiong Li, En-qing Hou Key Laboratory of Vegetation Restoration and Management of Degraded Ecosystems, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, PR China
a r t i c l e i n f o
a b s t r a c t
Article history: Received 11 July 2014 Received in revised form 7 September 2014 Accepted 13 September 2014 Available online
The temporal variation of polycyclic aromatic hydrocarbons (PAHs) concentrations as well as the lipid content in the xylem of Masson pine trees sampled from the same site were determined and compared with the days of haze occurrence and with the historical PAHs reported in sedimentary cores. The patterns of the lipid content as well as the PAH concentrations based on the xylem dry weight (PAHsDW) decreased from the heartwood to the sapwood. The trajectories of PAHs normalized by xylem lipid content (PAHs-LC) coincided well with the number of haze-occurred days and were partly similar with the historical changes in airborne PAHs recorded in the sedimentary cores. The results indicated that PAHs-LC in the xylem of conifers might reliably reflect the historical changes in airborne PAHs at a regional scale. The species-specificity should be addressed in the utility and application of dendrochemical monitoring on historical and comparative studies of airborne PAHs. © 2014 Elsevier Ltd. All rights reserved.
Keywords: Dendrochemistry Historical change Lipid-content-normalization Masson pine Polycyclic aromatic hydrocarbons (PAHs) Xylem
1. Introduction The drastic emissions from anthropogenic activities have led to the great abundance of polycyclic aromatic hydrocarbons (PAHs) in environments (Zhang and Tao, 2009). Monitoring of airborne PAHs has arisen a global concern for the carcinogenic, mutagenic, and € m et al., 2002). teratogenic effects of PAHs on human beings (Bostro Conventionally, routine PAHs were monitored using high-volume air samplers which were time-consuming and expensive. Alternatively, spatial and temporal PAHs were bio-monitored via “passive samplers”, which included vascular and non-vascular plants (Prajapati and Tripathi, 2008; Augusto et al., 2009). Needles of conifer species, especially pine species such as Pinus sylvestris (Holoubek et al., 2000), P. densiflora and P. maximartinezii (Hwang et al., 2003), P. massoniana (Tian et al., 2008), P. nigra, P. pinaster (Ratola et al., 2010, 2011) and P. halepensis (Ratola et al., 2012), have been frequently adopted to bio-monitor the current levels of
* Corresponding author. Present address: Xingke Road #723, Tianhe District, Guangzhou 510650, China. E-mail addresses: [email protected] (Y.-w. Kuang), [email protected] (J. Li), [email protected] (E.-q. Hou). http://dx.doi.org/10.1016/j.envpol.2014.09.018 0269-7491/© 2014 Elsevier Ltd. All rights reserved.
airborne PAHs, due to their wide distribution and their high lipid contents. Lacking the long-term monitoring data in most regions, researchers have accessed to relatively scarce reports on regional and historical changes in airborne PAHs via the analysis on treerings of bark pockets (Wang et al., 2004), sedimentary cores (Liu et al., 2005; Guo et al., 2011), and ice cores (Wang et al., 2008). Dendrochemistry is a subject that traces the historical changes in environmental pollution based upon the chemical analysis of the growth rings of trees (xylem). It assumed that the pollutants “recorded” in the xylem were immobile, thus, the levels of the studied pollutants in the growth rings were indicative of their real status in the environments when the growth rings formed (Nabais et al., 1999). Despite the doubt of concerning the efficiency on dendrochemistry (Bellis et al., 2002), researchers have used it to trace the regional history of heavy-metal emissions (Punshon et al., 2005), nitrogen pollution (Savard et al., 2009;Doucet et al., 2012) and soil acidification (Kuang et al., 2008), which were highly consistent with some detailed history events. To our knowledge, there have been relatively few reports on dendrochemistry tracing the historical changes in airborne PAHs, except a study of Wang et al. (2004), who selected a broadleaved tree (Cinnamomum camphora, approximately 10e15 years old) to investigate the accumulation, distribution and translocation of PAHs in the xylem,
Y.-w. Kuang et al. / Environmental Pollution 196 (2015) 53e59
400 D1 D3 D10
300 200 100
2006-2010
2001-2005
1996-2000
1991-1995
1986-1990
1981-1985
1976-1980
1971-1975
1966-1970
0 1961-1965
Xylem of Masson pines used for PAHs and lipid detection were sampled from Dinghushan natural reserve, which located in the northwest of PRD of southern China. This reserve has a lowsubtropical monsoon climate and tropical/subtropical forest and has experienced long-term exposure to atmospheric pollutants transported by the southwest prevailing wind in the summer from the numerous industries and vehicles in PRD economic zone. The atmospheric PAHs in this reserve were reported mainly from the anthropogenic emissions in PRD (Qi et al., 2001). Taking the opportunities of biomass investigation conducted by Dinghushan Forest Ecosystem Research Station, Chinese Academy of Sciences, we sampled two (D1, D3) and one (D10) discs from the cut-down model trees of Masson pine at the end of 2005 and 2010, respectively. The trees (about 40 cm in diameter without visible injury in the needles or a reduction in crown density) were selected in the forest plots far away from any roads. In order to avoid the contamination of external PAHs, the disc (about 20 cm in thickness) was cut from the stem of the model tree at breast height above ground (~1.3 m) with man-powered saw and transported to the laboratory immediately for processing. In the laboratory, the disc was mounted and polished to reveal the annual growth rings and then dated with the Windendro™ system (V 6.1D, Canada). Fresh chips of the xylem representing fiveyear intervals were carefully sliced from the heartwood to the sapwood (one chip per time interval) with a clean, electric microchisel, to obtain sufficient mass for PAHs analysis. A total of 34 chips (11 from both D1 and D3, 12 from D10) representing about a 60-year history (1950e2010) was sliced from the disc. The chips
About 8 g powder of the xylem chips (dry weight, DW) was extracted and purified for PAHs analysis. The process of the extraction, purification and analysis was described elsewhere (Tian et al., 2008; Wang et al., 2004). A Dionex ASE 300 (Dionex Co., USA) was used for PAHs extraction. The powder was mixed with 5 g of anhydrous sodium sulfate (barked for 4 h at 600 C in oven before use) and packed in a 33 mL stainless extracting cell. The samples were spiked with a known aliquot of naphthalene-d8, acenaphthane-d10, phenanthrene-d10, chrysene-d12 and perylene-d12 as analyte surrogates. The extraction used Dichloromethane (DCM) and acetone (1:1, v/v) as solvent under the conditions of 1500 psi and 100 C. The ASE was conducted for two cycles, each lasting for 5 min of heating time and 5 min of static extraction time. Activated copper slices were added to the extract to remove elemental sulfur. The xylem extract was concentrated to 1 mL with a rotary evaporator at 30 C and then loaded on a multiple layer chromatographic column packed with 2 g anhydrous sodium sulfate, 5 g aluminum oxide (extracted by DCM for 72 h, heated for 12 h in 250 C), 5 g florisil (baked for 8 h at 450 C in an oven) and 10 g silica gel (extracted by DCM for 72 h, heated for 12 h in 150 C), and eluted with 60 mL of DCM. The eluent was evaporated and solventexchanged to 1 mL of hexane. A gel permeation chromatography (GPC) column (10 mm i.d., packed with 10 g of S-X3Biobeads, dipped by dichloromethane in advance, Accustandard Co., USA) was used to eliminate lipids. The GPC column was eluted with 80 mL n-hexane: DCM (1:1 v/v) at a flow rate of 0.5 mL min1. The first 35 mL eluent was discarded, and the following 45 mL which contained PAHs were collected and concentrated to a final volume of 200 mL under a gentle stream of nitrogen. PAHs compounds were separated on a 30 m 0.25 mm i.d. HP-5 capillary column (film thickness 0.25 mm) and quantified using a HewlettePackard 5890 gas chromatography and 5972 mass selective (GC-MSD) detector operated in the electron impact mode (70 eV). The instrumental conditions were as follows: injector temperature, 280 C; ion source temperature, 180 C; temperature program: 60 C (2min), 60e290 C at 3 C min1, 290 C (30 min). The carrier gas was helium at a constant flow rate of 1.5 mL 30 m1. 1 mL sample was injected in splitless mode. Mass range m/z 50e500 was used for quantitative determinations. Data acquisition and processing was
1956-1960
2.1. Xylem sampling and processing
2.2. PAHs analysis
1951-1955
2. Materials and methods
were freeze-dried for 72 h and finely ground to pass through a 0.2mm screen.
-1
based on the physiological features of trunk tissues, the physicochemical properties of PAHs, and the correlations between PAHs concentrations in the tissues. They then reconstructed a 150-year airborne PAHs history using the concentrations of PAHs in the bark pockets of another broadleaved tree (Fagus longipetioleta, about 150 years old). Recommended by researchers, ringeporous species (i.e. conifers species) was more suitable for dendrochemical analysis than diffuseeporous ones (i.e. broadleaved species) (Bellis et al., 2002). Considering the differences in chemical properties between conifers and broadleaved species, we put forward the following questions: 1) Were the distribution patterns of PAHs in xylem related to the chemical properties of conifers? 2) Could the patterns of PAHs in conifer xylem indicate historical changes in regional airborne PAHs like those in tree-rings of bark pockets and in sedimentary cores? To address these questions, we analyzed 16 EPA PAHs and the lipid contents in the xylem of Masson pine (P. massoniana L.) trees, the most representative predominant species in southern China, from Dinghushan natural reserve in the Pearl River Delta (PRD). The number of haze days occurred in PRD and the historical data of airborne PAHs reconstructed from the analysis on the sedimentary cores were also used to analyses the correlation with the xylem PAHs. The specific objectives were: 1) to detect whether the temporal patterns of PAHs concentration and lipid content occurred synchronously in the xylem of Masson pines; 2) to reveal whether the concentration of PAHs was related to the content of lipids in the xylem; and 3) to determine the potential of lipid-content-normalized PAHs in the xylem of Masson pine to indicate the historical changes in airborne PAHs at a regional scale. The results were expected to improve the reliability of tracing historical changes in airborne PAHs using dendrochemistry.
Lipid content: mg. g
54
Period
Fig. 1. Temporal distribution of lipid in the xylem of the three Masson pines (D1, D3 and D10) from Dinghushan natural reserve. The xylem from trees D1 and D3 was formed from 1951 to 2005; the xylem of tree D10 was formed from 1951 to 2010.
Y.-w. Kuang et al. / Environmental Pollution 196 (2015) 53e59
controlled by HP Chemstation software. The concentrations of 16 EPA PAHs: naphthalene (Nap), acenaphthylene (Acpy), acenaphthene (Acp), fluorene(Flu), phenanthrene (PA), anthracene (Ant), fluoranthene (FL), pyrene (Pyr), benz[a]anthracene (BaA), chrysene (Chr), benzo[b]fluoranthene (BbF), benzo[k]fluoranthene (BkF), benzo[a]pyrene (BaP), dibenz[a,h]anthracene (DBA), indeno[1,2,3cd]-pyrene (IND) and benzo[g,h,i]-perylene (BghiP), in the chips were analyzed. Samples, including laboratory blanks only containing solvent and recovery standards (SRM 1649A, NIST, Gaitherbury, USA), were processed in the same way for quality control.
55
The relative percentage difference for individual PAH identified in paired duplicate samples (n ¼ 2) was always
