Accumulation dynamics and cellular locations of Pb, Znand Cd in resident and transplanted Flavocetraria nivalislichens near a former Pb–Zn mine

Jens Søndergaard

Received: 10 January 2013 /Accepted: 25 June 2013# Springer Science+Business Media Dordrecht 2013

Abstract Accumulation dynamics and cellular loca-tions of lead (Pb), zinc (Zn) and cadmium (Cd) werestudied in Flavocetraria nivalis lichens near the formerBlack Angel Pb–Zn Mine in West Greenland. Naturalresident thalli were collected from four dust-contaminated sites near the mine. In addition, thalliwere taken from an uncontaminated reference siteand transplanted to the contaminated sites followedby a collection 1 year after. Total thalli metal contentswere determined, and thalli were subjected to a sequen-tial extraction procedure. After 1 year of transplanta-tion, total Pb thalli contents were significantly elevatedcompared with initial concentrations at all sites (for Znand Cd contents only at the two sites closest to themine). However, transplanted thalli contained signifi-cantly less Pb (26±12 %), Zn (64±13 %) and Cd (34±7 %) compared with resident thalli from these sites.Results from the sequential extraction procedureshowed marked differences among Pb, Zn andCd in the extracellular, intracellular and residual frac-tion. The lower total metal concentrations intransplanted compared with resident thalli at the con-taminated sites were mostly due to a larger metal contentbound in the residual fraction in resident thalli. In

contrast, the metal content bound in the extracellularfraction were not significantly different in transplantedand resident thalli. The results indicate that extracellular-bound Pb, Zn and Cd in F. nivalis can be used as a proxyfor recent (annual) atmospheric metal deposition where-as the large residual metal fraction in resident lichensindicate an accumulation of metal-containing particlesin the thalli over time that includes several years ofuptake.

Keywords Lichens .Flavocetraria nivalis . Metalaccumulation . Cellular location . Sequentialextraction . Greenland

Introduction

Lichens are widely used as bio-monitors of atmospher-ic element deposition because of their ability to accu-mulate elements from the air and their relative ease ofcollection and analyses (Cayir et al. 2007; Augustoet al. 2009; Conti et al. 2012). Given their widespreadabundance, lichens have been used to evaluate atmo-spheric deposition of elements from an extensive rangeof dust sources such as mines, smelters and cities(Carreras and Pignata 2002; Rusu et al. 2006; Naethand Wilkinson 2008).

In Greenland, the foliose–fruticose-type lichen Flavo-cetraria nivalis (L.) Kärnefelt and Thell (previously

Environ Monit AssessDOI 10.1007/s10661-013-3321-1

J. Søndergaard (*)Department of Bioscience, Aarhus University,Frederiksborgvej 399, 4000 Roskilde, Denmarke-mail: jens@dmu.dk

named Cetraria nivalis) is abundant in most areas andhas been used to assess metal dust deposition in severalstudies (Rigét et al. 2000; Søndergaard and Asmund2011; Søndergaard et al. 2011b; Søndergaard et al.2012). For environmental monitoring near mines inGreenland, sampling of F. nivalis lichens is typicallythe only method used for evaluating dust dispersionand subsequent deposition of elements from the air(mainly metals are of concern and are in focus in thefollowing). For this purpose, both resident lichens col-lected within the monitoring area and lichen transplantshave previously been used (Søndergaard and Asmund2011; Søndergaard et al. 2011b). Transplanted lichensare moved from an unpolluted area into the monitoringarea and typically collected 1 year after. Lichen trans-plants, as opposed to resident lichens, have the advantagethat the exposure period to any potential dust pollution isknown and that any change in the lichens element com-position relative to the original content can be related tothat period. Therefore, the method of lichen transplanta-tion is typically favoured to evaluate deposition during adefined period of time and has previously been used toassess temporal variations (between years) and spatialvariations in dust deposition of metals (Søndergaardet al. 2011b).

However, while lichens accumulate metals deposit-ed from the air, they cannot be regarded as simpleinert traps of metal-containing dust particles. Thelichen thalli are open complex systems in which metal-containing dust particles may undergo a series of trans-formations and the metals may be involved in cellmetabolic processes and subjected to excretion andwashed out by precipitation (Brown and Brown 1991).Consequently, the uptake and accumulation of metals inlichens are highly metal specific (Carreras et al. 2005).For metal accumulation in lichens, three main bindingmechanisms/locations are known: (1) trapping of solidmetal-containing particles on the thalli surface and in theintercellular spaces between them, (2) extracellularbinding of metals with exchange sites on the cell wallsof symbionts and (3) intracellular uptake of metals(Nash, 1996). The extracellular-bound metals are oftenregarded as a reflection of environmental conditionswhereas those located within the cell are more relatedto the effects on the physiological processes (Brown andBrown, 1991). Using various sequential extraction pro-cedures, studies of the cellular locations of metals in-cluding lead (Pb), zinc (Zn) and cadmium (Cd) in otherlichen species have previously been done (Branquinho

and Brown 1994; Mikhailova and Sharunova 2008).However, despite the extensive use of F. nivalis lichensfor environmental monitoring purposes in Greenland,little is known of the binding and cellular location ofmetals in the thalli of F. nivalis. Knowledge of thecellular location of metals in the lichen thalli is impor-tant to understand the metal-specific accumulation dy-namics and the use of F. nivalis as a monitoringorganism.

The purpose of this study was to investigate the totalaccumulation and cellular locations of Pb, Zn and Cdin both resident and transplanted F. nivalis lichensusing sequential extraction. Resident lichens weresampled at four dust contaminated sites near the formerBlack Angel Pb–Zn Mine in West Greenland and theresults were compared statistically to transplanted li-chens before and after transplantation to the same site.

Site description

The study area is located adjacent to the former BlackAngel Pb–Zn Mine in Maarmorilik in the inner part ofthe Uummannaq Fiord Complex in West Greenland(71°07′ N; 51°15′ W) (Fig. 1).

The climate is arctic with maximum summer tem-peratures around 10 °C and minimum winter tempera-tures falls below −30 °C. Precipitation in the area issparse, approximately 100 mm/year, and the wind re-gime is dominated by moderate to strong easterly windcoming from the Greenland ice sheet in the bottom ofthe Qaamarujuk Fiord. The nearest settlements areUkkussissat, 25 km to the west and Uummannaq,80 km to the south of Maarmorilik.

The Black Angel Mine operated between 1973 and1990, and a total of 5.9×105 tonnes Pb and 2.3×106

tonnes Zn concentrate were produced during the peri-od. The mining activity caused significant contamina-tion of mainly Pb and Zn but also other metals such asCd and to a lesser extent arsenic and mercury in thesurrounding environment (Asmund 1992; Elberlinget al. 2002; Perner et al. 2010). Mine-related metalcontamination can still be measured today (2012) morethan two decades after mine closure (Søndergaard et al.2011a; this study). At the time of writing (2012), there areplans to re-open the mine within the next years, partly toexploit the reserves of Pb and Zn left behind in pillarsin the old mine using new techniques, partly to exploitnew recently discovered ore bodies. At Maarmorilik,

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dispersion of metal-contaminated dust was, and still is,an important source of Pb, Zn and Cd contamination(Søndergaard et al. 2012). During the mining period,dust was generated from ore crushing, handling of con-centrates and from the waste rock dumps established onthe steep mountain slopes of the Black Angel Mountain.After the mining ceased, metal-contaminated dust isthought to originate mainly from remains of ore materialin the Maarmorilik mining town, especially near theformer ball mill and from the remaining waste rockdumps left on the steep mountain sides in the area(Fig. 1). In addition, in 2011, construction work tookplace at the mine site and the cable car connectionbetween the Maarmorilik mining town at sea level tothe mine entrance at 600-m altitude was re-established.This activity is likely to have contributed to the dustdispersion in the area during this period. The ore at theBlack Angel Mine contains approximately 4 % Pb,12 % Zn and 0.06 % Cd (Johansen et al. 2001). Therock material regarded as waste rock contained roughlythe same relative proportions of Pb and Zn (and likelyalso Cd) but at lower concentrations (0.1–0.8 % Pb and0.3–2.3 % Zn; Johansen et al. 2001). The dry climate inMaarmorilik, the scarce vegetation cover and prevalent-ly strong winds favours an effective mobilization anddispersion of dust in the area. A more detailed descrip-tion of the study area can be found in Søndergaard et al.(2011b).

Methods and instrumentation

Sampling of lichens

F. nivalis lichens for transplantation were collected inAugust 2011 at Site L, 35 km fromMaarmorilik (Fig. 1).Site L is considered uncontaminated with respect tometal-containing dust dispersion from the mine. Onlyfresh living lichens growing on dead organic matterwere collected while lichens growing directly on soilor rocks were excluded. This selection was done tomaximise the probability that metals accumulating inthe lichens originated from deposition of particles fromthe air and not from underlying soil or rocks. A repre-sentative sample of the lichens was taken beforetransplantation to determine the initial metal concentra-tions and distributions of metals. Then lichens weretransplanted to four dust contaminated sites: sitesT22, T17B, T30 and T36, situated 1–12 km fromMaarmorilik (Fig. 1). The lichen transplants wereplaced on the ground on dead organic matter andcovered by a 1×1 cm mesh nylon net held in placeby small flat pieces of rock to avoid the removal of thelichens by the wind. Three, ca. 15×15-cm patches withlichens were made.

In August 2012, after 1 year of transplantation, thelichen transplants were collected at the four sites to-gether with resident F. nivalis lichens from the sites.

Greenland

St. T17B

St. L

Qaamarujuk

Ukkussissat

Maarmorilik+++

Affarlikassaa

St. T30

St. T22St. T36

Fig. 1 Map of Maarmorilikand the surrounding area inWest Greenland. The loca-tion of the mining town ad-jacent to the Black AngelPb–Zn mine (marked witha mining symbol), the sam-pling sites (sites L, T36, T30,17B and T22), the wasterock dumps (marked withcrosses) and the settlementUkkussissat is shown

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Polyethylene bags were used for collection and lichensfrom the three patches were pooled into one bag. Inthe laboratory, lichen samples were sorted using stain-less steel tweezers and only the green/yellow thalliwere kept. Samples were then kept frozen until furtheranalyses.

In addition to the lichen studies performed in 2011–2012, data were included for total Pb and Zn contents(Cd was not consistently measured) in transplanted andresident lichens sampled as part of the regular moni-toring program in Maarmorilik during the period1997–2012 (Søndergaard et al. 2011b). These lichenswere sampled as described above.

Chemical analyses

After returning to Denmark, lichens from all foursites were analysed for total metal concentrationsas described below (three replicate samples). In addi-tion, lichens from site T17B were analysed for metalcellular locations using a sequential extraction proce-dure modified from Branquinho et al. (1999). The li-chens from site T17B consist of three sample types: (1)resident lichens from site T17B, (2) transplanted lichensbefore transplantation and (3) transplanted lichens aftertransplantation.

The sequential extraction method includes the fol-lowing: before analyses, samples were stored in a highhumidity atmosphere for 24 h (over water in an air tightbox at room temperature) in order to reactivate thephysiological activity and ensure that cell membraneswere not leaky because of desiccation damage (Buckand Brown 1979). Five replicate samples (100–150 mgwet wt.) from each of the three sample types were takenout for sequential extraction. The following four ex-traction steps were applied: Step 1: the samples wereplaced in 14 ml polyethylene vials, 10 ml of MilliQwater was added and the samples shaken for 5 min on alaboratory shaker. The lichens were transferred to anew set of vials using polyethylene pipette tips. MerckSuprapure HNO3 was added to the solutions a concen-tration of 1 % (wt./wt.). This was to determine thewater soluble metal fraction. Step 2: the samples wereshaken for 40 min in 9 ml of 20 mM Na2-EDTA atpH 4.5 (SCP Science, ACS grade) followed by a sec-ond extraction with 5 ml of same solution for 30 min.The two extraction solutions were pooled. This stepwas made to determine the extracellular metal fractionusing Na2-EDTA at pH 4.5 as a metal-displacing agent

(Branquinho and Brown, 1994). Step 3: the lichenssamples were dried at 80 °C for 12 h and weighted.The oven drying process has previously been shown torupture cell membranes without changing the distribu-tion of the elements (Branquinho et al. 1999). Then, thelichens samples were shaken in 10 ml of 20 mM Na2-EDTA at pH 4.5 for 2 h to determine the intracellularmetal fraction. Step 4: the remaining metals not re-moved during the previous extraction steps, i.e. theresidual metal fraction was extracted using a digestionin 4 ml concentrated Merck Suprapure HNO3 and 4 mlMilliQ water in Teflon bombs under pressure in amicrowave oven (Anton Paar Multiwave 3000). Threeblank samples were prepared for each extraction stepsdescribed above to determine the blank values (meanof blanks) and detection limits (3 SD on blanks). Blankvalues were subtracted from the analytical results. As acontrol for the extraction steps, the total metal concen-trations were determined in five replicate samples(100–150 mg wet wt.) from each of the three differentlichen sample types. These samples were dried at60 °C, then weighted and digested in half-concentrated HNO3 in a microwave oven as describedabove in step 4 (but not subjected to steps 1–3). Thesuspensions were not filtered prior to analyses.

Metal analyses were performed on suspensions usingan Agilent 7500ce ICP-MS at the trace element labora-tory in Roskilde, Denmark. The analytical quality (bothdigestion and analyses) was checked by analysingblanks, duplicates and the certified lichen referencematerial BCR-482 from the European Commission,Joint Research Centre (www.irmm.jrc.be). The resultson BCR-482 are shown in Table 1 and the detectionlimits for all extraction steps are shown in Table 2.

Table 1 Analytical results on the certified lichen referencematerial BCR-482 from the European Commission, Joint Re-search Centre

Zn Cd Pb

BCR-482 (n=4)

Measured

Mean 96.6 0.56 39.5

SD 5.6 0.02 2.9

Certificate

Mean 100.6 0.56 40.9

2 SD 2.2 0.02 1.4

The unit is milligrams per kilogram dry weight

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Table 2 Metal concentrations measured in each sequential extraction fraction (water soluble (S), extracellular (E), intracellular (I) andresidual (R)) and total concentrations measured in lichens not subjected to the sequential extraction procedure (Control; n=5)

Concentrations (mg kg−1 dry wt.) % of total (S+E+I+R)

S E I R Control S E I R Control

D.l. (3 SD on 3 blanks) 0.01 0.02 0.01 0.17 0.17 – – – – –

Before t.

Pb

Mean 0.14 0.39 0.10 0.27 0.95 15 43 11 31 106

SD 0.12 0.16 0.04 0.14 0.21 14 10 2 17 23

After t.

Pb

Mean 0.49 7.68 1.52 2.14 17.5 4 65 13 18 148

SD 0.15 1.26 0.35 0.82 4.1 1 4 1 7 35

Resident

Pb

Mean 2.74 11.2 4.83 33.3 54.0 5 22 9 63 104

SD 0.97 5.4 0.13 9.5 7.4 2 12 1 15 14

D.l. (3 SD on 3 blanks) 0.14 0.22 0.04 0.24 0.24 – – – – –

Before t.

Zn

Mean 0.47 8.46 12.3 1.61 19.5 2 36 54 7 86

SD 0.14 3.18 1.3 0.33 3.0 0 8 6 2 13

After t.

Zn

Mean 0.56 14.5 14.5 4.79 41.2 2 43 41 14 124

SD 0.14 4.5 0.7 1.70 2.2 0 6 6 4 7

Resident

Zn

Mean 1.93 17.5 18.3 12.6 56.1 4 35 37 25 111

SD 0.53 3.8 1.4 4.3 2.4 1 5 3 6 5

D.l. (3 SD on 3 blanks) 0.001 0.011 0.004 0.046 0.046 – – – –

Before t.

Cd

Mean 0.010 0.105 0.057 0.060 0.268 4 44 27 25 115

SD 0.007 0.062 0.015 0.033 0.059 2 10 7 11 25

After t.

Cd

Mean 0.008 0.117 0.117 0.090 0.324 2 35 35 28 97

SD 0.004 0.032 0.060 0.030 0.034 1 6 13 9 10

Resident

Cd

Mean 0.029 0.104 0.188 0.717 0.827 3 10 21 66 80

SD 0.016 0.059 0.013 0.418 0.085 0 1 10 9 8

The percentages of each sequential extraction fraction and for the control samples relatively to the calculated total content (S+E+I+R) areshown. Results are given for lichens before and after transplantation to site T17B and for natural resident lichens at site T17B

D.l. detection limit

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As a control of the sequential extraction procedure,the sum of all fractions was evaluated against the meanof five control samples (not subjected to sequential ex-traction but analysed for total metal contents). As shownin Table 2, the concentrations in the control sampleswere close to 100 % of the sum of all fractions (80±8to 148±35 %). Furthermore, the analytical results on thecertified lichen reference material BCR-482 (Table 1)was close to the certificate values (96±6 to 100±3 %).

Data treatment

Differences in total metal concentrations in lichens be-fore and after transplantation and between transplantedand resident lichens were investigated statistically usingMicrosoft Excel 2010 software using two-tailed t testsand a significance level of 5 %. Prior to the t tests, datawere tested for equal variances with an F test.

Results and discussion

Total metal concentrations in resident and transplantedlichens before and after transplantation

Total concentrations of Pb, Zn and Cd in thalli ofF. nivalis before and after 1 year of transplantation tofour contaminated sites in Maarmorilik and in resident F.nivalis from the same sites are shown in Fig. 2. Thesesites are situated 1–12 km from the mine. After trans-plantation, total Pb concentrations were significantlyelevated (p<0.05) in thalli (up to 23±3 mg Pb kg−1)compared with initial concentrations (1.0±0.2 mgPb kg−1) at all sites. Total Zn and Cd concentrationswere only significantly elevated in transplants at thetwo sites closest to the mine (1–2 km away; up to47±1 mg Zn kg−1 and 0.33±0.03 mg Cd kg−1) relativeto the initial concentrations (19±3 mg Zn kg−1 and0.25±0.06 mg Cd kg−1). Elevated metal concentra-tions in lichen transplants indicate a recent disper-sion and subsequent deposition of Pb-, Zn- and Cd-containing dust adjacent to the mine. It is important tonote that the smaller area apparently affected by Zn andCd contamination as compared with Pb as indicated bythe statistical tests does not necessarily mean that con-tamination with Zn and Cd does not occur in a wider areasimilar to that of Pb. The natural background concentra-tions of Zn in lichens in the area is much higher for Znthan for Pb (19±3 mg Zn kg−1 vs. 1.0±0.2 mg Pb kg−1)

and given the Zn/Pb ratio of typically 2:3 in ore/wasterock (Johansen et al. 2001), it is simply to possibly todetect a similar change in Zn concentration. For Cd, theconcentration in ore/waste rock (Cd/Pb∼0.02) is alsorelatively low compared with the natural backgroundconcentration in lichens (0.25±0.06 mg Cd kg−1)resulting in a similar detection problem.

After 1 year of transplantation, transplanted thallicontained significantly less Pb (26±12 %), Zn (64±13 %)and Cd (34±7 %) compared with resident thalli fromthe same sites. Since there has been an on-going dis-persion and deposition of dust in the area since the mineactivity started in 1973 (Søndergaard et al. 2011b),differences in metal contents in transplanted versus

0.1

1

10

100

1000

T36 T30 T17B T22

mg

kg-1

dry

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Pb

1

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100

1000

T36 T30 T17B T22

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kg-1

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Zn

0.1

1

10

T36 T30 T17B T22

mg

kg-1

dry

wt.

Cd

(a)

(b)

(c)

Sites

Fig. 2 Total Pb, Zn and Cd concentrations in F. nivalis lichensbefore transplantation (1. column), after 1 year of exposure (2.column) and in resident lichens (3. column) collected at the fourcontaminated sites in Maarmorilik (mean+SD)

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resident thalli indicate that there has been a net accumu-lation of Pb, Zn and Cd in resident F. nivalis over timethat includes several years of uptake. This is in contrastto Spiro et al. (2004) who found that Pb in the lichenHypogymnia physodes near a copper smelter in Russiawas largely replaced after a period of 3 months. Waltheret al. (1990) reported a residence time of 2–5 yearsfor many elements in lichen thalli of Parmotremapraesorediosum and Ramalina stenospora. The lattertime period will at least be the residence time for Pb,Zn and Cd in F. nivalis at Maarmorilik. Consequently,the observed metal accumulation over time in F. nivalismay not apply to other lichen species and will depend onfactors such as growth rates, age of the lichens, climatic

conditions, the specific metals of concern and metaldeposition rates. The following sequential extraction ofPb, Zn and Cd in F. nivalis thalli was done to study thecellular location of these metals in both transplanted andresident lichens to further characterise the accumulationdynamics.

Distributions of metals in resident and transplantedlichens as assessed by sequential extraction

Results from the sequential extraction on lichens fromsite 17B are shown in Fig. 3 and Table 2. Detectionlimits for each extraction step (3 SD on three blanksamples) are included in Table 2. As shown, all results

0

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100

Before t. After t. Resident

% o

f tot

al

Cd

Residual

Intracellular

Extracellular

Water soluble

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% o

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Residual

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(a)

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0.8

1

1.2

Before t. After t. Resident

mg

kg-1

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Extracellular

Water soluble

Fig. 3 Distribution of Pb, Zn and Cd in F. nivalis lichens before and after transplantation and in resident lichens from Site T17B asdetermined by the sequential extraction procedure (a–c in milligrams per kilogram dry weight; d–f in per cent)

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from the four extraction steps were above detectionlimits for Pb, Zn and Cd.

The first extraction showed that only a minor fraction(typically 2–5 %) of the Pb, Zn and Cd in resident andtransplanted F. nivalis was water extractable. F. nivalislichens collected for monitoring purposes near mines inGreenland are typically not rinsed with water prior toanalyses, and the results indicate that a rinse of the thalliwith water is not likely to make an important differencewith regards to the thalli Pb, Zn and Cd contents.

The second extraction revealed that for Pb and Zn,the extracellular concentrations were significantlyhigher after 1 year of transplantation (not significantfor Cd). In addition, extracellular concentrations of Pb,Zn and Cd were not significantly (p<0.05) different intransplanted and resident F. nivalis as opposed to totalconcentrations (Fig. 3a–c). This indicates a relativelyfast equilibrium between metal input and exclusion ofextracellular-bound metals in F. nivalis and agrees withthe concept that extracellular-bound metals in lichenthalli mainly reflect the current/recent level of atmo-spheric deposition (Bargagli and Mikhailova 2002).Similarly, a study by Mikhailova and Sharunova(2008) on the lichen H. physodes from a polluted areain Russia showed that the extracellular-bound metalcontent were near similar in resident lichens and intransplants after 1 year of exposure. Given the lowertotal Pb, Zn and Cd concentrations in transplantedlichens after 1 year as compared with resident lichens,the fraction of extracellular-bound metals were signif-icantly higher in the transplanted lichens (Fig. 3d–f).

The third extraction showed that for both transplantedand resident F. nivalis, the intracellular bound fractionwere higher for Zn (37–54%) than for Cd (21–35%) andPb (9–13 %) (Fig. 3d–f). A similar trend was observedby Mikhailova and Sharunova (2008) for the lichen H.physodes. A higher intracellular fraction of Zn as com-pared with Pb and Cd can be explained by Zn being anessential element in cell metabolic processes in contrastto Pb and Cd (Nagajyoti et al. 2010). The intracellularconcentrations of Pb, Zn and Cd in transplantedF. nivalisafter 1 year were above the initial concentrations prior totransplantation but lower than those of resident F. nivalis(significant for Pb and Zn but not for Cd). Thus, theresults indicate that long-term impact of metal pollutionon resident F. nivalis has an impact on the intracellularcontents at least for Pb and Zn that cannot be attributed tothe last year of exposure. This may be due a decreasedability of the resident lichens to exclude metals from the

cells (Bačkor and Loppi 2009) due to an overall highermetal content in the resident lichens as compared withthe lichen transplants.

The fourth and final extraction revealed that a majorpart of the Pb and Cd (63±15 and 66±9 %, respectively)and to a lesser degree Zn (25±6 %) in resident F. nivaliswere bound in a residual fraction not extractable withNa2-EDTA. In transplanted F. nivalis, the residual frac-tion of Pb, Zn and Cd was significantly lower than inresidentF. nivalis and the larger residual fraction was themain reason for the differences in total metal concentra-tion between resident and transplanted F. nivalis. Pre-sumably, the metals in the residual fraction are mainlybound as nearly insoluble particles near the thalli surfaceor in the intercellular spaces (Nash 1996; Cuny et al.2004). Previous studies have demonstrated the presenceof metal rich particulates in intercellular spaces of themedulla and on thallus surfaces using SEM and Electronmicroprobe techniques (Prithiviraj et al. 2011). Howev-er, further studies are needed to characterise how thesemetals are bound in the residual fraction of F. nivalis.

Considerations for using of F. nivalis to monitoratmospheric metal deposition

The results from this study show that in order to use F.nivalis to evaluate recent atmospheric metal deposition

y = 1.333x + 16.806R² = 0.9175

y = 0.4728x + 20.808R² = 0.8267

0

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300

400

500

600

0 100 200 300 400 500 600

Zn

conc

entr

atio

ns (

mg

kg-1

dry

wt.)

Pb concentrations (mg kg-1dry wt.)

Transplanted lichens

Resident lichens

Ore/waste rock

Fig. 4 Total Zn versus total Pb concentrations in transplanted F.nivalis lichens after 1 year of exposure and in resident F. nivalislichens collected from 18 sites near Maarmorilik during theperiod 1997–2012. The trend lines for transplanted and residentlichens are shown with line equations. The hyphenated lineshows the average Zn/Pb ratio in ore and waste rock inMaarmorilik (from Johansen et al. 2001)

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within a long-term-polluted area, it is adequate totransplant F. nivalis from an unpolluted area and collectit after a period of time. A 1-year transplantation periodis typically used near mines in Greenland and thelichens are considered to accumulate metals duringthe entire snow-free period (Søndergaard et al. 2012).Resident lichens from a sampling area may be used toevaluate recent deposition if the pollution has justrecently occurred (e.g. the first year of a mining oper-ation) provided that there is enough background dataavailable. In this study, resident F. nivalis within along-term polluted area contained significantly highertotal Pb, Zn and Cd concentrations than transplanted F.nivalis after 1 year of exposure and a major part of thismetal was shown to be strongly bound in the thalli. Incontrast, the extracellular-bound metal concentrationswere near similar in transplanted and resident lichensindicating a relatively fast adjustment of this fraction tothe current level of metal deposition. Consequently, ifresident lichens within a long-term polluted area haveto be used to evaluate recent metal deposition, it will bemore adequate to isolate the extracellular fractionrather than the measuring the total metal concen-tration. The net accumulation in resident F. nivalisof Zn was relatively lower than for Pb and Cd indicat-ing a mechanism for F. nivalis to exclude more Znrelative to Pb and Cd. This conclusion is in line withresults reported in Søndergaard et al. (2012) showing alower net accumulation rate of Zn relative to total atmo-spheric deposition compared with Pb and Cd. Exclusionof metals is a relatively well-known detoxificationmechanism in lichens (Bačkor and Loppi 2009). Acontinuous exclusion of Zn over time in F. nivalis rela-tive to Pb is indicated in Fig. 4, which shows total Znand Pb concentrations in resident and transplanted thallisampled during the period 1997–2012 from 18 sitesin Maarmorilik (Cd was not measured). As shown,transplanted lichens contained less Zn compared withPb than the average Zn/Pb ratio in ore and waste rockconsidered to be the main source for metal dust deposi-tion in Maarmorilik (from Johansen et al. 2001). Inaddition, resident lichens from Maarmorilik that havebeen exposed to metal deposition for many yearscontained less Zn compared with Pb than transplantedlichens after 1 year of exposure. Consequently, this ithas to be into account if F. nivalis is used as a relativeproxy to evaluate atmospheric metal deposition as Zndeposition will otherwise be underestimated relative todeposition of Pb.

Conclusions

The present study shows that F. nivalis lichens areuseful bio-indicators of metal dust deposition nearmines in the Arctic. However, the results also showsthat F. nivalis cannot be regarded as simple inert trapsof metals and factors like metal-specific exclusiondynamics has to be taken into account with using theselichens for evaluating metal deposition. Specifically,Zn in F. nivalis was shown to be excluded at a fasterrate than Pb. Furthermore, total metal contents in res-ident lichens from a long-term polluted area cannot beused to evaluate recent uptake. This is due to an ob-served net accumulation of strongly bound metals in F.nivalis over time that includes several years of deposi-tion. Consequently, transplantation of lichens from anunpolluted area into the monitoring sites following bya collection, e.g. 1 year is generally considered themost adequate method of evaluating temporal and spa-tial variations in atmospheric metal deposition. Alter-natively, the results indicate that the extracellular metalfraction in resident lichens can be used as a proxy forrecent atmospheric deposition.

Acknowledgments The author wishes to thank Kim Gustavsonand Lis Bach for logistical support and inspiration and GertAsmund for chemical advice, all are from Department of Biosci-ence, Aarhus University in Roskilde. In addition, the miningcompany Angel Mining PLC is thanked for providing the logicalfacilities on site.

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