Determination of Heavy Metal Deposition in the County of Obrenovac (Serbia) Using Mosses As Bioindicators. I: Aluminum (Al), Arsenic (As), and Boron (B)

Arch. Biol. Sci., Belgrade, 57 (3), 205-212, 2005.

DETERMINATION OF HEAVY METAL DEPOSITION IN THE COUNTY OF OBRENOVAC (SERBIA) USING MOSSES AS BIOINDICATORS. I: ALUMINUM (AL), ARSENIC (AS), AND BORON (B)

M. SABOVLJEVIĆ1,4, V. VUKOJEVIĆ1, NEVENA MIHAJLOVIĆ2, GORDANA DRAŽIĆ2, andZ. VUČINIĆ3

1Institute of Botany and Jevremovac Botanical Garden, Faculty of Biology, University of Belgrade, 11000 Belgrade, Serbia and Montenegro 2INEP, 11080 Zemun, Serbia and Montenegro 3Center for Multidisciplinary Studies, University of Belgrade, 11000 Belgrade, Serbia and Montenegro 4Nees Institute of Plant Biodiversity, Rheinische Friedrich Wilhelms University of Bonn, 53115 Bonn, Germany

Abstract - In the present study, the deposition of three heavy metals (Al, As and B) in the county of Obrenovac (Serbia) is determined using four moss taxa (Bryum argenteum, Bryum capillare, Brachythecium sp., and Hypnum cupressiforme) as bioindicators. Distribution of average heavy metal content in all mosses in the county of Obrenovac is presented in maps, while long-term atmospheric deposition (in the mosses Bryum argenteum and B. capillare) and short-term atmospheric deposition (in the mosses Brachythecium sp. and Hypnum cupressiforme) are discussed and given in tables. Areas of the highest contaminations are highlighted.

Key words: Heavy metal deposition, mosses, bioindicators, Serbia

UDC 582.32(497.11Obrenovac):581.5

INTRODUCTION Mosses are better than other higher plants in scan- ning heavy metal deposition because: Surveillance of heavy metals in mosses was originally -they are perennial without deciduous periods; established in the Scandinavian countries in the 1980s. -they have high cation exchange capacity that allows However, the idea of using mosses to measure atmos- them to accumulate great amounts of heavy metals pheric heavy metal deposition was developed alredy in between the apoplast and symplast compartments the late 1960s (Rhülingand Tyler, 1968; Tyler, without damaging vital functions of the cells; 1970). It is based on the fact that mosses, especially the -mosses do not possess thick and strong protective carpet-forming species, obtain most of their nutrients layers like cuticles. directly from precipitation and dry deposition. In the present investigations, we decided to use two acrocar- Also, this time-integrated way of measuring patterns pous moss species (Bryum argenteum Hedw. and Bryum of heavy metal deposition from the atmosphere to terres- capillare Hedw.) that can give us an idea of long-term trial ecosystems, besides being spatially oriented, is easi- atmospheric deposition, inasmuch as they are attached to er and cheaper than conventional precipitation analyses, the substrate and also accumulate metals deposed during as it avoids the need for deploying large numbers of pre- the last few decades in the surface layers of the substrate. cipitation collectors. The higher trace element concentration in mosses compared to rain water makes analysis Two pleurocarpous taxa (Brachythecium sp. and more straightforward and less prone to contamination Hypnum cupressiforme Hedw.) were used to scan short- (Berg et al. 1997). term atmospheric deposition of heavy metals, consider- ing that these taxa are not strongly attached to the sub- Use of mosses to investigate heavy metal deposition strate and accumulate mostly from precipitation. shows transboundary heavy metal pollution and can indi-

205 206 M. SABOVLJEVIĆ et al. cate the paths by which atmospheric pollutants enter from the presence of all investigated species and yearly bio- other territories or reveal their sources within the investi- mass. More than 500 samples were analyzed. gated area. After collecting, samples were dried as soon as pos- Although 15 heavy metals were analyzed in all, only sible in a drying oven to constant dry weight (dw) at a deposition and distribution of aluminum, arsenic and constant temperature of 350C, then stored at -200C. Fol- boron are treated in the present study. lowing homogenization in porcelain, the samples were treated with 5+1 parts of nitric acid and perchloric acid MATERIAL AND METHODS (HNO3:HClO4 = 5:1) and left for 24 hours. After that, a Kjeldatherm digesting unit was used for digestion at 150- The acrocarpous mosses Bryum argenteum and 2000C for about one hour. Digested samples were filtered Bryum capillare were used to research long-term atmos- on qualitative filter paper to dispose of silicate remains. pheric deposition, while the pleurocarpous Brachytheci- Sample volume was then normated to 50 ml. Aluminum um sp. and Hypnum cupressiforme were used to scan (Al) was detected by AAS on a Pye Unican SP9 atomic short-term atmospheric deposition in the county of absorbance spectrophotometer from Philips using the Obrenovac (Serbia). Hypnum cupressiforme is one of the flame of acetylene/nitrogen-suboxide. Boron (B) was standard species used in Europe to survey heavy metal detected by ICP-ES, while arsenic (As) was determined deposition (Buse et al. 2003.), while the other three are by AAS using hydride techniques. standard in Europe, but do not grow in this region. In esti- mating which other species are eligible for use in heavy For explanation of the results and their presentation metal deposition monitoring, we relied on the experience on maps, the folloving statistical statistics parameters of Thöni (1996), Herpin et al. (1994), Siewers and Hair- were used: average values, standard deviation, minimum, pin (1998) Zechmeister (1994), and Ross (1990). maximum, and percent deviation. Map making and inter- polation of exact data were made using Agis software As far as possible, moss sampling was conducted (v1. 71 32 byte, © Agis Software, 2001). according to guidelines, set out in the experimental protocol for a survey carried out in 2000/2001 (UNECE, RESULTS AND DISCUSSION 2001). Details of the procedure are given in Rühling et al. (1998). Aluminum in Obrenovac County probably comes from heavy road traffic, intensive coal burning, and Each sampling site was located at least 300 m from incorrect waste disposal. The most contaminated areas main roads and populated areas and at least 100 m from according to deposition in mosses are the town of Obren- any road or single house. In forests or plantations, sam- ovac and the northernmost part of the county by the Sava ples were collected in small open spaces to preclude any River (>40 mg/g). A somewhat lower level of Al deposi- effect of canopy drip. Sampling and sample handling tion (20-30 mg/g) occurs in the central part of the coun- were carried out using plastic gloves and bags. About ty, where Al is probably spread by traffic on the main three moss samples were collected from each site. Dead roads. material and litter were removed from the samples. Green parts of mosses were used for the analyses. Aluminum represents 8% of the Earth's crust. It comes from bauxite (Merian, 1984), is very wide- The county of Obrenovac was chosen for this inves- spread in all forms of industry (machine construction, tigation because of its industry and location. Each sam- electro engineering, civil engineering, the food industry, pling site was GPS located with a precision of ±10 m, and agriculture), and is also present in aluminum foil, iron GPS dates (Germin) were digitalized on the maps using plates, and other widely used articles. Aluminum emis- the following softwares: OziExplorer 3.95.3b, © D&L sion into the atmosphere was great during the last centu- Software Pty Ltd, and WinDig 2.5 Shareware, © D.Lovy. ry. Only in the 70s it was 7.2 million tons per year All material was collected during November of 2002. Not (Lantzyand Mackenzie, 1979). It is known that more than one site was chosen in a 50x50 m square. the atmospheric concentration of aluminum is signifi- Seventy-five localities were chosen out of 129 for com- cantly elevated (>10mg/m3) in areas with a cement indu- parisons and further analyses. The selection was based on DETERMINATION OF HEAVY METAL DEPOSITION IN THE COUNTY OF OBRENOVAC (SERBIA) 207

Fig. 1. Exact locations of moss sampling (left) and extrapolated maps (right) of selected heavy metal deposition in the county of Obrenovac (Serbia). stry (Merian, 1984). Arsenic and boron are probably spread in the county of Obrenovac by wind blowing from coal ash deposit In slightly acid and neutral soils, dissolving of alu- sites. According to the mosses analyzed, the center of minum is inconsiderable, while in acid soils (pH<5.0) the arsenic deposition is the southwestern part of the county concentration of Al3+ is significantly elevated. High con- around the village of Vučičevica. However, there is a centrations of Al3+ in the substrate exert toxic action on rather large area in the western part of county with aver- plants by damaging the roots and by lowering the uptake age arsenic deposition of 000.7-000.9 mg/g (dw of moss- of essential phosphorous, calcium, and magnesium es). Some smaller areas with high arsenic deposition are (Bergmann, 1988). Aluminum is also toxic for ani- also are present in the town of Obrenovac and around mals and man, especially if its concentration is high in Barič. The highest presence of boron is in the southwest- water and if air with a high concentration of it is inhaled ern part of Obrenovac County, where its concentra- over longer period. It can cause Alzheimer's disease tion is more than 1.2 mg/g. Some zones with medium (Merian, 1984). In drinking water, the maximal toler- concentrations (0.4-0.8 mg/g) are found in the town ated value of aluminum concentration is 0.2mg/l of Obrenovac and places by the Sava (Mladost and (Thöni and Seitler, 2004). Ušće). 208 M. SABOVLJEVIĆ et al.

Table 1. Deposition of Al, As, and B in the county of Obrenovac (Abbreviations: H - Hypnum cupressiforme, BRA - Brachythecium sp., Bc - Bryum capillare, Ba - Bryum argenteum. No. Locality Longitude Latitude Al As B mg/g mg/g mg/g 1Vinogradi - H20.16370244.3917583.24550.00100.4032 2Moštanica 1 - H 20.18367244.38424913.75340.00500.5001 3Iskra 1 - Bc20.15523544.39272215.75870.00520.2750 4Iskra 2 - Ba20.15282644.39328434.88130.0078<0.0261 5Iskra 1 - H20.15523544.3927229.57720.00520.3451 6Iskra 2 - Bc20.15282644.39328413.64240.0067<0.0111 7Rvati 1 - Bc20.11879644.39693034.67020.0152<0.0223 9Deponija B ulaz 1 - Bc20.02333144.38373535.10690.00870.0162 10Zabrežje 1 - Bc20.12127344.41124521.86410.00560.1374 11Ušće 2 - Bc20.06644144.41923514.58860.00580.0707 12Vinogradi - Bc20.16370244.3917587.54400.00350.3632 13Iskra 1 - Ba20.15523544.39272220.14200.0064<0.0080 14Ušće 2 - H20.06644144.41923544.84940.00630.0252 15Ušće 1 - Bc20.07034344.41473811.12720.00400.4919 16Urozv - BRA20.07977044.38904326.10970.0074<0.0167 17Zabrežje 2 - H20.13379644.40829312.84000.00410.0613 18Orašac 1 - H20.02181944.33671713.46940.00440.1429 19Hotel - Ba20.12745144.39404924.85990.00540.0781 20Moštanica 1 - H20.18367244.3842496.49350.00170.1623 21Grabovac 1 - H20.04693444.3599977.47000.00260.2916 22Šab.put nadv. - Bc20.09408544.39136731.27220.0104<0.0127 23Vranić - H20.15212244.34752913.60440.00410.1922 24Jasenak 2 - BRA20.15624644.36007124.29310.00780.0617 25Dren 1 - BRA20.02322444.35823826.95510.0055<0.0171 26Veliko Polje 1 - H20.10864844.36595417.79030.0023<0.0390 27Grabovac 1 - Bc20.04693444.35999710.74240.00320.0461 28Belo Polje 1 - Bc20.11806444.38278317.71770.0055<0.0250 29Brović 1 - Bc20.07220144.33510818.51530.00720.1608 30Ljubinić 2 - BRA20.02676244.33483212.15070.00210.1458 31Hotel - H20.12745144.39404924.85710.00550.0802 32Grabovac 1 - BRA20.04693444.35999726.50430.00810.0456 33Ljubinić 2 - Bc20.02676244.33483230.23260.00750.1525 34Veliko Polje 4 - H20.10905744.34190817.82960.0055<0.0070 35Zabran 3 - H20.13761544.3969057.08040.00230.1702 36Zabran 1 - H20.13939644.39826820.01400.0070<0.0117 37Orašac 3 - H20.01661244.3438555.31070.00531.8056 38Orašac 2 - H20.02086044.34063918.08900.00440.0162 39Zabran 2 - Ba20.14237744.4016729.99730.00270.1583 40Belo Polje 1 - Ba20.11806444.38278317.88830.00520.2356 41Orašac 2 - BRA20.02086044.3406398.68450.00350.1769 42Ljubinić 1 - BRA20.03763044.32213214.86200.00150.1696 43Grabovac nad. - Ba20.09278844.36516715.37910.00220.0918 44Joševa - H20.06054544.31074223.06460.00460.0688 45Brović 2 - BRA20.08892944.31853718.01710.00370.0487 46Jasenak 2 - Ba20.15624644.36007112.89430.00310.0904 47Garbovac nadv. - BRA20.09278844.36516725.81550.00790.0381 DETERMINATION OF HEAVY METAL DEPOSITION IN THE COUNTY OF OBRENOVAC (SERBIA) 209

Table 1. Continued

No. Locality Longitude Latitude Al As B mg/g mg/g mg/g 48Baljevac 1 - Bc20.15204444.34074330.78460.00710.0291 49Joševa - Bc20.06054544.31074237.14640.00590.0190 50Joševa - BRA20.06054544.31074235.15560.00730.0593 51EPS - Bc20.12040144.3888457.59560.00180.0550 52Konatice II - BRA20.14892844.33741018.26880.00330.0631 53Zabran 1 - Ba20.13939644.39826810.16770.00110.0797 54Mislođin 1 - BRA20.13657944.38309615.26450.00230.2971 55Brović 1 - H20.07220144.33510811.43230.00160.2473 56Mislođin 4 - H 20.13406744.36961614.54180.00240.1195 57Stubline 2 - H20.09164944.34509576.91570.00620.1634 58Konatice 1 - Bc20.16215044.3162656.14090.00160.1317 59Zabran 3 - Ba20.13761544.3969056.50140.00210.0468 60Jasenak - H20.14380444.36573621.02740.00260.0697 61Konatice 2 - Ba20.15583144.32296024.92500.00410.0879 62Veliko Polje 4 - Bc20.10905744.34190818.25310.0047<0.0167 63Mislođin 1 - BRA20.13385744.38704126.43330.00280.0616 64Veliko Polje 3 - Bc20.10611744.3446707.09100.00620.0522 65Konatice II - Bc20.14892844.33741043.09220.00460.0817 66Mislođin 6 - Ba20.16467644.3710270.2029<0.00030.0330 67Stubline 1 - H20.08635344.35718512.33000.00360.1989 68Šab.put nadv. - Ba20.09408544.39136723.30580.00990.0286 69Dren 1 - H20.02322444.35823813.36790.00290.2303 70Zabran 2 - Bc20.14237744.40167213.83890.00350.3460 71Baljevac 2 - H20.12943244.34238319.11940.00440.0546 72Mislođin 5 - Ba20.15081344.36727413.89260.0034<0.0096 73Orašac 1 - BRA20.02181944.33671724.45050.00730.1282 74Konatice II - H20.14892844.3374105.90670.00310.1136 75Šab.put 1 - BRA20.04909444.39656620.00180.00220.2788 76TENT

Table 1. Continued

No. Locality Longitude Latitude Al As B mg/g mg/g mg/g 94TENT

Median17.38280.00450.1346

Arsenic is present in the Earth's crust with ca. 1.7 g for preservation of wood products. Use of arsenic in pro- per ton (Thöniand Seitler, 2004). It is often present duction of herbicides, insecticides, fungicides, and pig- in the form of arsenictrioxide (As2O3) as a side product in ments is strictly excluded due to its easy binding in production of copper, lead, zinc, and nickel. Because organisms. Arsenic is mainly present in the atmosphere arsenic induces cancer, it is used to only a limited extent as a side product of copper and nickel production and of in industry (production of screens and some other arti- brown coal burning. It can be present in brown coal in cles). However, it is still present in glass and substances concentrations up to 1500 mg/kg, although the average is DETERMINATION OF HEAVY METAL DEPOSITION IN THE COUNTY OF OBRENOVAC (SERBIA) 211

18 mg/kg (Thöniand Seitler, 2004). Further REFERENCES sources of arsenic are the metal industry, agriculture industry, and volcanic activity. Global emission of Berg, T., Steinnes, E. (1997). Use of mosses (Hylocomium splen- dens and Pleurozium schreberi) as biomonitors of heavy metal arsenic fell from 1983 (19000 t per year) to 1995 (5000 t deposition: form relative to absolute values. Environmental per year) (Pacynaand Pacyna, 2001). Normally, it Pollution 98, 61-71. . is present in the atmosphere with 2800 t (Lantzyand Bergmann, W. (ed.) (1988). Ernährunsstörungen bei Kulturpflanzen. Mackenzie, 1979). Gustav Fischer Verlag, Stuttgart, New York. 1-762. .

There are no data on the effects of arsenic on plants Buse, A., Norris, D., Harmens, H., Büker, P., Ashenden, T., Mills, G. (2003). Heavy metals in European mosses. Centre for Ecology

(Thöniand Seitler, 2004). For animals and man, and Hydrology, Bangor. . arsenic is known to be toxic, carcinogenic, teratogenic Herpin, U., Markert, B., Siewers, U., Lieth, H. (1994). Monitoring der and mutagenic. The chronic presence of arsenic pollution Schwermetallbelastung in der Bundesrepublik Deutschland mit can lead to serious hematological and neurological dis- Hilf von Moosanalysen. Bundesministerium für Umwelt, eases (Merian, 1984). Borderline values of arsenic Naturschutz und Reaktorsicherheit. Ökologie, Forschungbericht 180 02 087, 1-161. concentration are as follows: 1 mg/m3 in the air, 0.1 mg/l . in water, and 0.05 mg/l in drinking water (Thöniand Lantzy, R. J., Mackenzie, F. T. (1979). Atmospheric trace metals: glob- Seitler, 2004). al cycles and assessments of man´s impact. Geochemica et CosmochemicaActa 43, 511-525.

Since all of the sampled species could not be found Merian, E. (ed.) (1984). Metalle in der Umwelt, Verteilung, Analytik at each locality, average values of all specimens were und biologiche Relevanz. Verlag Chemie, Weinheim. 1-722. extrapolated on the maps to get an idea of heavy metal Pacyna, J. M., Pacyna, E. G. (2001). An assessment of global and deposition in the county of Obrenovac (Fig. 1). However, regional emissions of trace metals to the atmosphere from anthro- if one separates the values of deposition obtained from pogenic sources worldwide. Canadian Journal of Environmental Reviews (Environ. Rev.) 9, 269-298. pleurocarpous mosses (Brachythecium sp. and Hypnum cupressiforme) and ones obtained from acrocarpous Rühling, A. (1998). Atmospheric heavy metal deposition in Europe 1995-1996. NORD 1998: 15. Nordic Council of mosses (Bryum argenteum and Bryum capillare), it is Ministers, Copenhagen. readily discernible that the first two give us an idea of short-term and the last two of long-term deposition, .Siewers, U., Hairpin, U. (1998). Schwermetalleinträge in Deutschland- respectively (Tab. 1). Moosmonitoring 1995/96. Geologisches Jahrbuch, Sonderhefte, Reihe D, Heft SD 2, Stuttgart, Schweizerbarat, 1-199. .

These differences are easily explained in terms of the Thöni, L. (1996). Vergleich der Elementkonzentrazionen in drei Biomonitormoosen untereinander und mit Depositionfrachten im life forms of these mosses and their uptake of heavy met- Bulksammler nach Bergerhoff. BUWAL (ed.), Bern, 1-89. als. Pleurocarps are not closely attached to the substrate and so get the bulk of deposited heavy metals directly Thöni, L., Seitler, E. (2004). Deposition von Luftschadstoffen in der Schweiz, Moosanalysen 1990-2000. BUWAL (ed.), Bern, 1-139. from the atmosphere (during their pauciennial life period), while acrocarps are strictly attached to the substrate Tyler, G. (1970). Moss analyses -a method for surveying heavy metal and receive the bulk of deposited heavy metals from the deposition. In: England, H. H. and Berry, W. T. (eds.): Proceedings of the Second International Clean Air Congress. substrate solution (metals deposited over a period that is Academic Press, New York. . longer than their pauciennial life span). UNECE (2001). ICP Vegetation experimental protocol for the 2001 sea- sion. ICP Vegetation Coordination Centre, CEH Bangor, UK. Acknowledgements. The study was financially supported by the Serbian Ministry of Science and Environmental Protection. We are also thankful to two Zechmeister, H. (1994). Biomonitornig der Schwermetalldeposition anonymous reviewers who helped us by improving an early version of the man- mittels Moosen on Österreich. Monografien des uscript. Umweltbundesamtes, Wien, B. 94. 1-147. 212 M. SABOVLJEVIĆ et al.

ДЕПОЗИЦИЈА ТЕШКИХ МЕТАЛА НА ПОДРУЧЈУ ОПШТИНЕ ОБРЕНОВАЦ (СРБИЈА) ПРАЋЕНА ПРЕКО МАХОВИНА КАО БИОИНДИКАТОРА I: АЛУМИНИЈУМ (AL), АРСЕН (AS), И БОР (B)

М. САБОВЉЕВИЋ1,4, В. ВУКОЈЕВИЋ]1, НЕВЕНА МИХАЈЛОВИЋ2, ГОРДАНА ДРАЖИЋ2 И Ж. ВУЧИНИЋ3

1 Институт за ботанику и Ботаничка башта”Јевремовац”, Биолошки факултет, Универзитет у Београду, 11000 Београд, Србија и Црна Гора 2 ИНЕП, 11080 Земун, Србија и Црна Гора 3 Центар за мултидисциплинарне студије, Универзитет у Београду, 11000 Београд, Србија и Црна Гора 4Nees Institute of Plant Biodiversity, Rheinische Friedrich Wilhelms University of Bonn, 53115 Бон, Немачка

У овом раду представљена је депозиција три екстраполирано, док се дугорочна (вредности код тешка метала (aлуминијум, aрсен и бор) на подручју маховина Bryum argenteum и B. capillare) и општине Обреновац (Србија) праћена преко четири краткорочна (вредности код маховина Brachythecium маховине (Bryum argenteum, Bryum capillare, sp. и Hypnum cupressiforme) депозиција тешких Brachythecium sp. и Hypnum cupressiforme). Распоред метала може видети из табела. Подручја територије просечног садржаја ових метала у маховинама општине Обреновац са највишим степеном представљен је на картама са тачним подацима и контаминације су посебно означена.