Lichens as Biomonitors of Depleted Uranium in Kosovo - Springer Link

Journal of Atmospheric Chemistry 49: 437–445, 2004. C 2004 Kluwer Academic Publishers. Printed in the Netherlands.

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Lichens as Biomonitors of Depleted Uranium in Kosovo S. LOPPI1 , L. A. DI LELLA1 , L. FRATI1 , G. PROTANO1 , S. A. PIRINTSOS2 and F. RICCOBONO1 1

Department of Environmental Science “G. Sarfatti”, University of Siena, Italy, e-mail: [email protected] 2 Department of Biology, University of Crete, Greece (Received: 23 April 2004; accepted: 5 May 2004) Abstract. This paper reports the results of a study using lichens as biomonitors to investigate the environmental distribution of depleted uranium (DU) at selected Kosovo sites as a consequence of the use of DU ordnance during the conflict of 1999. The results suggested that the use of DU ammunitions did not cause a diffuse environmental contamination in such a way to have caused a detectable U enrichment in lichens. Also isotopic 235 U/238 U measurements did not indicate the presence of DU particles in lichens, except for some samples at a heavily shelled site, which resulted contaminated by DU. Key words: biomonitoring, depleted uranium, Kosovo, lichens

1. Introduction The concern of some European governments about a series of leukemia cases among their soldiers who served in the Balkans gave rise to research addressed to the determination of the environmental impact and health risk as a consequence of the use of depleted uranium (DU) ordnance during the conflict of 1999 (Schiermeier, 2001). Apart from concern over the possible impacts of DU on military personnel, there has also been considerable concern over the possible impacts of DU on local populations and the field staff of international organizations. During the Kosovo conflict, DU antitank ammunitions were fired from NATO aircraft, and it has been reported that over 30,000 DU rounds were used, totalling about 10 tons of DU (UNEP, 2001). The DU dust eventually formed by the impact on hard targets, such as armored vehicles, dispersed into the environment contaminating the air and the ground. As a consequence of a widespread contamination of the ground surface by DU, there is the risk that some DU will become airborne through wind action and be subsequently inhaled by people. The risk of contamination of food (fruit, vegetables, meat, etc.) and drinking water must be also considered. Likewise natural uranium (natural uranium has a 235 U content of 0.7%, whereas in DU the 235 U content is depleted to about one third of its original amount), DU is an unstable, radioactive material that emits ionizing alpha, beta and gamma radiation.

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The effects of exposure to DU are both radiological (i.e. due to radiation) and chemical (i.e. as a result of biochemical effects in the human body of this toxic heavy metal). Corresponding health consequences may, depending upon the dose or intake, include cancer and malfunction of body organs, particularly kidneys (Priest, 2001). Biological monitoring can be very effective as an early warning system to detect environmental changes. This approach is based on the assumption that any changes taking place in the environment have a significant effect on the biota (Keddy, 1991). Sentinel organisms are species capable of accumulating persistent pollutants, such as heavy metals, and are used to measure the biologically available amount of a given pollutant in a given ecosystem (Beeby, 2001). Lichens are highly efficient accumulators of many elements because they are perennial, slow-growing organisms, that maintain a fairly uniform morphology in time, are highly dependent on the atmosphere for nutrients, and do not shed plant parts as readily as vascular plants (Hale, 1983). Elements acquired by lichens represent some fraction of the elements present in their immediate environment and as a consequence, lichens in an apparently healthy state must reflect a reasonable equilibrium between their requirements and the availability of elements in the environment (Brown et al., 1994). It has also been demonstrated that the concentrations of several trace elements in lichen thalli are directly correlated with the environmental levels of these elements (Bari et al., 2001; Sloof, 1995). With specific reference to uranium, lichens have been used to assess its distribution around known sources of pollution such as uranium mines and mills and yellow cake production plants (Boileau et al., 1982; Beckett et al., 1982; Fahselt et al., 1995; Jeran et al., 1995), but also coal-fired power plants (Gough and Erdman, 1977). Furthermore, lichens have been used for the evaluation of uranium mining impact on critical food chains, such as that of lichen-caribou-human (Thomas and Gates, 1999). In this study, the collection of lichen samples and the measurement of total uranium concentrations in their thalli, as well as the calculation of 235 U/238 U isotopic weight ratios, was considered as a means to get insight into the environmental consequences of the release and dispersal of uranium particles from the use of DU ammunitions during the Kosovo conflict. 2. Materials and Methods The study was designed in a multistep way, starting from a small-scale survey in an area of Kosovo heavy shelled with DU rounds, and adding later other Kosovo sites in a wider survey (Figure 1). 2.1.

HEAVY SHELLED SITE

The study was performed at the garrison of the former Yugoslavian Army (VJ) in Djakovica, used as an armored vehicle parking area and as an ammunition repository

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Figure 1. Study sites.

Figure 2. Schematic drawing of the garrison of the former Yugoslavian Army in Djakovica, with indication of sampling sites and penetrator impact holes.

(Figure 2). On May 14, 1999, the garrison was attacked by NATO aircraft which fired about 300 DU rounds. Two sampling sites were selected according to the presence/absence of penetrator holes: one in an area heavily shelled with DU ammunition (71 holes were counted), and the other about 100 m away in an old field, used as a control site. 2.2.

OTHER KOSOVO SITES

The following localities were investigated (Figure 1): • Radonjiko. The site is located close to the dam on the southern shore of an artificial lake which provides drinking water to the southern part of Kosovo.

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Several artillery and tank positions were dug into the slope of a nearby ridge, SW of the dam; a radio station was located on the top of the ridge. On June 7, 1999 the site was attacked by NATO aircraft which fired 655 DU rounds. • Vranovac. The site is a hill, on the top of which the Serbian army placed 12 anti-aircraft artillery positions. On June 8, 1999 the site was attacked by NATO aircraft which fired 2320 DU rounds. • Rznic. The site was a arms depot of the former Yugoslavian Army (VJ). On June 7, 1999 the site was attacked by NATO aircraft which fired 530 DU rounds. 2.3.

LICHEN SAMPLING

Lichen samples were collected in June 2001 at Djakovica and in October 2001 at the other sites. For all the available lichen species, 3–10 thalli were harvested from tree trunks at a height of 1–2 m above the ground and from all sides of the trunk when possible. 2.4.

LICHEN ANALYSIS

In the laboratory, the lichen samples were air-dried to constant weight, carefully cleaned with nylon tweezers under a binocular microscope to remove dead or senescent tissue and as much extraneous material (adhering bark, mosses, other lichen species, soil particles, etc.) as possible. Samples were not washed to avoid loosing of particles trapped on the lichen surface and because there is evidence that the washing procedure can unpredictably alter the elemental composition of lichens (Bettinelli et al., 1996). The lichen samples were then immersed in liquid nitrogen until brittle and were then powdered and homogenized with a ceramic pestle and mortar. About 150 mg of lichen powder were mineralized with a 6:1 v.v. mixture of concentrated HNO3 and H2 O2 , at 280 ◦ C and a pressure of 55 bars, in a microwave digestion system (Milestone Ethos 900). Total U concentrations, expressed on a dry weight basis, were determined by ICP-MS, (Perkin-Elmer Sciex 6100). Analytical quality was checked by analyzing the Standard Reference Materials IAEA-336 “lichen” and GBW-07608 “bush branches and leaves.” Accuracy resulted within 7%. For the determination of DU, a rationale was adopted based on the assumption that if there was a substantial contribution of DU to the total U concentration, this could be detected by ascribing any excess U, with respect to normal levels in lichens, to DU. Otherwise, if the U in lichens was found at normal levels, then there had been no U/DU enrichment and thus no widespread environmental contamination by DU. For this reason, lichen samples with high U concentrations (close to 1 ppm or above) were used to measure the 235 U/238 U isotopic ratios. Samples were preconcentrated by ligand exchange sorption on the iminodiacetic chelating resin Chelex-100 (Bio Rad Laboratories). About 4 g of 100–200 mesh Na-form resin were

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then sealed in a polyethylene column and conditioned with a solution of Na-acetate trihydrate and glacial acetic acid, at pH 4.6. The sample solution was adjusted to pH 4 with a NaOH dilute solution. The sorbed U was finally eluted with 6 ml of a 2M HNO3 solution. The determination of uranium isotopic composition was performed by ICP-MS. Analytical quality was checked by analyzing the standard reference material NBS SRM U-005a, having a nominal U235 /U238 wt% ratio of 0.005025. Accuracy resulted within 2%. 3. Results 3.1.

HEAVY SHELLED SITE

The results (see also Di Lella et al., 2003) were extremely variable, with U concentrations ranging from 0.11 to 4.26 µg g−1 dw (Table I). The variability of Table I. Total U concentrations (µg g−1 dw) and 235 U/238 U isotopic weight ratios in lichen samples from selected Kosovo sites. S = heavily shelled site, C = control site, − = not determined Site

Lichen species

U

235

U/238 U

Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica S Djakovica C Djakovica C Djakovica C Radonjicko Radonjicko Vranovac Vranovac Rznic Rznic Rznic Rznic Rznic

Physcia biziana Xanthoria parietina Physcia adscendens Phaeophyscia orbicularis Xanthoria parietina Physcia biziana Phaeophyscia orbicularis Xanthoria parietina Physcia adscendens Physcia biziana Phaeophyscia orbicularis Xanthoria parietina Phaeophyscia orbicularis Physcia adscendens Xanthoria parietina Physcia adscendens Xanthoria parietina Parmelia tiliacea Xanthoria parietina Physcia aipolia Parmelia tiliacea Xanthoria fallax Physconia venusta

0.28 0.15 1.53 0.76 0.11 1.44 0.40 0.19 2.10 0.68 0.98 0.12 2.15 4.26 0.23 0.27 0.29 0.18 0.43 0.42 0.19 0.97 1.00

– – 0.006 – – 0.007 – – 0.007 – 0.004 – 0.007 0.007 – – – – – – – 0.007 0.007

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the concentrations was mainly related to differences among lichen species, with Physcia adscendens and Phaeophyscia orbicularis generally showing the highest values, Xanthoria parietina the lowest values and Physcia biziana intermediate values. These differences are probably related to differences in the exposed surface area of the lichens. Owing to these differences, comparison between the heavily-shelled site and the control site was only possible within the same lichen species. Uranium concentrations in X. parietina, the only species found at both sites, agree with the values found in several species from the Balkans (Loppi et al., 2003) and in X. parietina from Puglia, southern Italy (unpublished data). In general, the U values were fairly similar at both study sites. Phaeophyscia orbicularis is the other species found at both sites, and in this species uranium was present at concentrations one order of magnitude higher than in X. parietina. Interestingly, all values were higher at the control site. Although Phaeophyscia orbicularis and Physcia adscendens at the control site showed the highest U concentrations (2.15 and 4.26 µg g−1 dw respectively), their isotopic ratios did not indicate any DU contamination (Table I). Of the four lichen samples from the heavily-shelled site for which the isotopic ratio was measured, only one Physcia adscendens and especially one Phaeophyscia orbicularis sample showed some contamination by DU (Table I). 3.2.

OTHER KOSOVO SITES

Uranium concentrations spanned over about one order of magnitude (Table I). This variability was related both to differences across localities and across different lichen species. As already suggested, these latter differences can be explained in terms of different exposed surface area of the lichens. Differences across localities showed that Rznic had the highest mean U concentrations. However, owing to species-specific differences, comparison between localities was only possible within the same lichen species. Xanthoria parietina was the only species found at all sites, and comparison using only this species showed that the samples Rznic had about two-fold U concentrations. However, the U content of X. parietina agreed with the values found in several species from the Balkans (Loppi et al., 2003) and in X. parietina from Puglia, southern Italy (unpublished data). The 235 U/238 U ratios (Table I), did not indicate any DU contamination. 4. Discussion Our results agree with the findings of UNEP (2001), which reported very high variability of U concentrations in lichens and soils close to penetrator impact sites or penetrator holes. The high variability of total U concentrations in lichens could be due to contamination of thalli by terrigenous material (Loppi et al., 1999). In


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fact, it has been shown that in unwashed samples of lichens from unpolluted areas, where much of the material suspended in the air is simply soil or rock dust, the concentrations of several elements increase in proportion to soil contamination (Bargagli, 1995). Since much of the metal burden of lichens occurs as particulate matter trapped in the intercellular spaces of the medulla (Garty et al., 1979), trapped soil particles are probably not evenly distributed in the thalli. Replicates would thus show high variability of total concentrations. In rocks, uranium is usually concentrated, in variable amounts, in accessory minerals such as uraninite, coffinite, monazite, thorite and zircon (Finch and Murakami, 1999). As a consequence, if dust particles, even very small ones, are captured by lichens, they can strongly increase the total U content of these organisms. Even if contamination by the DU oxide dust that formed at the moment of the penetrator’s impact with hard targets cannot be excluded, the most likely source of DU in lichens is soil resuspension (Di Lella et al., 2003), which is known to be an important phenomenon owing to the small size of DU oxide particles (Danesi et al., 2003). Natural uranium consists of 99.3% 238 U and 0.7% fissionable 235 U, the relative proportions of the isotopes being determined by the respective decay rates. Most measured values of the 235 U/238 U weight ratio show a remarkably constant figure of 0.007 (Greenwood and Earnshaw, 1984). In the case of DU used in ammunition, this value drops to about 0.002 (Harley et al., 1999). Allowing for some variation in measurement, any observed value below 0.007 is regarded as a mixture containing DU. It is important to remark that unlike total element concentrations, isotopic measurements are not subject to possible uncertainty concerning standardization of lichen species and soil contamination of samples, and thus give an accurate estimate of the contamination by DU. The results for Vranovac and Rznic are at variance with the data of Sansone et al. (2001), which found some DU contamination in lichen samples collected there. However, since eventual DU contamination takes place as dust, this variability is likely caused by randomly trapped soil particles within lichen thalli. In fact, it has been shown that in soil samples collected at Kosovo sites where DU ammunition was used, several hundred thousand particles of DU can be present in a few milligrams of contaminated soil (Danesi et al., 2003). 5. Conclusions The results of the present survey suggested that the use of DU ammunitions in Kosovo did not cause a diffuse environmental contamination in such a way to have caused a detectable U enrichment in lichens. Also isotopic 235 U/238 U measurements did not indicate the presence of DU particles in lichens, except for some samples at a heavily-shelled site, which resulted contaminated by DU. Although it might be questioned if lichens are best suited for this kind of monitoring purpose, it should be remembered that all studies carried out by UNEP in Kosovo (UNEP, 2001), Serbia

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