ACTIVITY CONCENTRATIONS OF 137 CS AND 40 K IN MOSSES FROM SPAS IN EASTERN SERBIA

With the aim of detecting the presence of radionuclides in moss samples in eastern Serbia, 129 samples of mosses were collected from 2000 to 2010 in the region of the spas: Sokobanja, Banja Jošanica and Gamzigradska banja. The data obtained show that 137Cs was present in all of the 129 samples from 3 sampling areas and 15 sampling localities and in 20 moss taxa. This indicates that pollution with this radionuclide resulting from the Chernobyl accident still exists. Changes in the average activity concentrations of 40K are within measurement error limits, while for 137Cs changes in the average activity concentrations are more noticeable, i.e. they have decreased with time.


INTRODUCTION
Evolutionarily, mosses were among the first plants to colonize land (Richardson, 1981) and face the challenges of terrestrial living.Thus, they have developed many adaptations for survival in harsh environments, some of which even nowadays are fascinating (e.g.anabiosis and poikilohidry, metal ion accumulations in the thallus with no cuticles etc.).The structure of mosses makes them suitable for monitoring the aerial metal burden.Mosses have a slow growth rate and a relatively large surface areato-biomass ratio; the lack of waxy cuticles and associated stomata means that many contaminants can be absorbed over the whole external surface; they do not have a rooting system; they obtain nutrients from dry and wet precipitation by absorption through their leaves which lack cuticle (Richardson, 1981).The use of mosses in biomonitoring has significantly increased since 1968, when Rühling and Tyler first used them as biomonitors of atmospheric quality (Rühling and Tyler, 1968).Mosses may be considered the most commonly applied organisms for biomonitoring studies of radioactive contamination (Chakrabortty and Paratkar, 2006;Batias and Siontorou, 2007;Berg and Steinnes, 1997;Basile et al., 2008;Čučulović and Veselinović, 2008;Čučulović and Veselinović, 2009;Čučulović et al., 2010).
Mosses are among the pioneering plant species on burnt land, secondary bare surfaces and in tundra.They prevent land erosion and represent miniecosystems due to the number of organisms that depend on them.Moss utilization is high in industry, medicine and pharmacology (Longton, 1992;Delfanti at al., 1999).
Air pollutants are deposited on mosses in three forms: as an aqueous solution, in gaseous form or as attached particles.The accumulation of pollutants in mosses occurs through a number of different mechanisms.
Potassium is a lithophilic element and is monovalent under natural conditions.Of the naturally occurring primordial radionuclides, 40 K (T 1/2 =1.28x10 9 y) is very abundant in soil as the molar fraction of 40 K is 0.0117 (Veselinović et al., 2004), and the average content in the Earth's core is 25.9 kg/t (Mason, 1996).In a living organism, potassium is evenly distributed.The greatest part of internal radiation coming from natural sources of radioactivity originates from potassium (United Nations Committee, 2000).Cs + and K + are competing ions.
Cesium-137 ( 137 Cs) is one of the most frequent artificial radionuclides in the environment, arising from different anthropogenic discharges such as atmospheric nuclear weapon testing, accidental release from nuclear power plants, chronic emissions from nuclear power plants and chronic emissions from nuclear reactors and fuel-reprocessing plants (Eisler, 2000.;Davis, 1963;Whicker, 1997).Cesium-137 is an emitter of gamma and beta radiation, with a long radioactive half-life (30.17 years).
The first nuclear weapons were detonated in New Mexico and Japan in 1945.The partial meltdown of the reactor at Chernobyl in April 1986 released high amounts of radionuclides into the environment, especially 137 Cs.Following the Chernobyl reactor accident, numerous studies involving the monitoring of the long-lived fission product 137 Cs were undertaken using mosses as biological indicators to update the inventory of this long-lived radionuclide (e.g.Boileau at al., 1982;Giovani at al., 1994;Papastefanou at al., 1992;Butkus and Konstatinova, 2005;Sawidis at al., 2009;Elstner et al., 1987;Papastefanou et al., 1989;Marović et al., 2008;Ilus et al., 1987;Elstner et al., 1987;Čučulović et al., 2002;Čučulović et al., 2008).As mosses lack a well-developed root system, the uptake of radionuclides occurs mainly through an ion exchange process directly from wet and dry deposition on the moss body (Delfanti et al., 1999;Mishev et al., 1996).
The expansion of knowledge about contamination paths by following bioindicators is important for radiological research and comparison with available data.

MATERIALS AND METHODS
Samples of mosses were collected from 2000 to 2010 in the territories of the following spas: Sokobanja, Banja Jošanica and Gamzigradska banja.
Sokobanja is located in the central part of eastern Serbia and is part of the Zaječar region.Most of the municipality is part of the extensive Carpathian-Balkan mountain range.A smaller western part is in the zone of the old Rhodopes mass.It is located 43° 38' 29" latitude north and 21° 52' 37" longitude east at 400 m above sea level.The spa is located in a west-east direction and slopes westward.The climate is mild continental.Samples from the territory of the Sokobanja municipality were taken at the following locations: in the city of Sokobanja (hotel "Sunce") (locality SB1); Lepterija (1.5 km east from the center of Sokobanja on the left bank of the Moravica river) (SB2); in Soko grad (400 m from Lepterija, upstream the Moravica) (SB3); Ozren (about 4.5 km from the center of Sokobanja in the "Ozren" Special Hospital Complex) (SB4); Ophthalmology hospital (on the Ozren road, 5 km from Sokobanja Southeast, close to the Ozren Special hospital for treating eye problems) (SB5).In the period 2000-2010, samples were taken as follows: from locality SB1 5 samples, and one sample from each of the localities SB2 23, SB3 1, SB4 57 and SB5, giving a total of 87 samples.
Banja Jošanica is located in the northwestern part of the Sokobanja valley, between the western part of Mt.Rtanj and the eastern slopes of Mt.Bukovik, close to Sokobanja.In the territory of Banja Jošanica, samples were taken in the following locations: in the park (BJ1); close to the mineral water springs (BJ2); and on the Jošanica River banks (BJ3).In the period 2009-2010, from each location (BJ1, BJ2 3 and BJ3) 3 samples were taken, giving a total of 9 samples.Gamzigradska banja is located close to Zaječar, in the meandering valley of the Crni Timok River, at the outermost southwestern part of the Romanian basin close to the Carpathian Mountains of eastern Serbia.This spa is one of the oldest balneological centers in Serbia.In the territory of Gamzigradska banja samples were taken on the left (GB1) and right bank of the Crni Timok river (GB2), near the Gamzigrad Rehabilitation Center (GB3), near Hotel Kastrum (GB4), from the hotel annexes (GB5), hydroelectric power station (HE) Gamzigrad (GB6) and in the archeological locality of Felix Romuliana (GB7).In the period 2006-2010, from locality GB1 7 samples were taken, from GB2 1, GB3 5, GB4 3, GB5 3, GB6 12 and GB7 2 samples were taken, in total 33 samples.The moss samples were dried in air, homogenized, and the radionuclide activities measured gamma-spectrometrically.The specific activity of radionuclides was measured using an HPGe gamma-ray spectrometer (ORTEC-AMETEK, with 8192 channels, resolution 1.65 keV, relative efficiency 34% at 1.33 MeV for 60 Co).Samples were measured in Marinelli vessels.The sample weight was about 0.1 kg.The counting time for each sample was 60000 s.The relative error for sample preparation and measurement was 10%.Gamma Vision 32, MCA emulation software, was used to analyze the measured gamma-ray spectra.The specific activity of the artificially produced radionuclide 137 Cs was measured via the γ-line at energy of 661.6 keV.The specific activity of the 40 K radionuclide was determined from its 1460.8keV gamma-ray line.Nuclides were identified using a library driven search routine and quantitative analyses were carried out using the appropriate detector calibration.Radionuclide results were reported in Bq/kg on a dry weight basis.

RESULTS
Activity values of 137 Cs and 40 K (Bq/kg) in mosses collected on the territory of Sokobanja, Banja Jošanica and Gamzigradska banja in the period from 2000 to 2010 are given in Table 1.
Maximal and minimal values of 137 Cs and 40 K were noted on the territory of the Sokobanja municipality in the locality SB4.The minimal activity value of 137 Cs was 1.12 Bq/kg (locality SB4, 2010, sample 7) and 40  On the territory of Banja Jošanica, the minimal activity concentration of 137 Cs was 72 Bq/kg (locality The average activity concentrations of 137 Cs and their standard deviations in moss samples collected on localities SB2 and SB4 in the period 2000-2010 are shown in Fig. 1.The average activity concentrations of 137 Cs in mosses collected on locality SB2 in most cases are higher than the average activity concentrations of 137 Cs in moss samples collected in locality SB4.
The average activity concentrations of 40 K and their standard deviations in moss samples collected on localities SB2 and SB4 are shown in Fig. 2.

DISCUSSION
The presence of the artificially produced radionuclide 137 Cs and the natural radionuclide 40 K was noted in all the investigated moss samples collected from spas in eastern Serbia in the period from 2000 to 2010.
Previous research has shown that mosses C. molluscum (4), L. sciuroides (11), H. splendens (13) and H. cupressiforme (14), are good bioindicators and biomonitors of radioactive pollution (Delfanti et al., 1999;Hanssen et al., 1980;Sawidis et al., 2009;Giovani et al., 1994).These mosses grow on the territory of Sokobanja, Banja Jošanica and Gamzigradska banja and they were randomly selected and used in this investigation.The results of this research confirm  that besides these, other moss species can be used as bioindicators of 137 Cs and 40 K radionuclide pollution (Table 1).
The data obtained (Table 1) shows that 137 Cs was present in all of the 129 samples from the 3 sampling areas and 15 sampling localities, and in 20 moss taxa.This definitely indicates that pollution by this radionuclide as a consequence of the Chernobyl accident still exists.The different activity concentrations of 137 Cs in mosses from spas in eastern Serbia are the consequence of different contents of 137 Cs in the fallout after the Chernobyl accident, moss species, substrate on which they grow, altitude, climatic and other factors.A comparison of the average activity concentrations of 137 Cs in the mosses collected on the territory of the Sokobanja, Banja Jošanica and Gamzigradska banja spas allows us to conclude that the lowest activity concentrations were present in the samples from the Sokobanja municipality.The highest average activity concentrations of 137 Cs in moss collected on the territory of the Sokobanja municipality were in the moss collected on the Lepterija locality (SB2), and they were slightly lower in the moss collected from the Ozren locality (SB4).The average activity concentrations of 137 Cs in moss collected on the territory of the Sokobanja municipality in the observed time period was in agreement with the findings of other authors (Sawidis et al., 2009).
The lowest activity concentrations of 137 Cs were measured in B. rivulare (7) from Ozren (1.12-3.60Bq/kg).As this moss grows besides the Ripaljka Falls, on the Gradašnica River, and that the river water periodically floods this moss, one can assume that the river water has desorbed 137 Cs from the moss and this is the reason why the activity concentrations of 137 Cs are low in this moss.Low activity concentrations of 137 Cs were also noted in the substrate on which B. rivulare moss grows.One can assume that the water also desorbs 137 Cs from the substrate.Investigations of 137 Cs desorption from Cetraria islandica (L.) Ach.lichen by distilled water and solutions simulating the composition of acid rain (Čučulović et al., 2006) favor this assumption.Activity concentrations in mosses growing close to this species, for example H. philippeanum (6), not flooded by the river, are higher (up to 28 Bq/kg, 2010).
In L. sciuroides (11), collected on the territory of Gamzigradska banja, high levels of 137 Cs were noted in 2009 (from 120 to 199 Bq/kg).Due to construction work on the Timok riverbanks since 2009, this moss species is an endangered species and no longer grows on these localities in Gamzigradska banja.Investigations have confirmed that this moss species is a good bioindicator of radioactive pollution in the environment (Sawidis et al., 2009) and it should be used in research.
In Fig. 1 and 2, one can clearly note that changes in the average activity concentrations of 40 K are within measurement error limits, while for 137 Cs, changes in the average activity concentrations are more noticeable, i.e. they decrease with time.
As 40 K is a natural isotope, this similarity in activity indicates a significant even distribution of 40 K on the Earth's surface and in fallouts that is reflected by its content in moss.However, for 137 Cs, the content reduction with time can be the consequence of the radioactive decay of 137 Cs, and washout with water not enriched with 137 Cs.
The ten-year reduction in the average activity concentration of 137 Cs at the SB2 locality is 79.1%, while it is 80.6% at the SB4 locality, as shown in Table 2 for the period 2001-2010.Reduction in activity concentrations of 137 Cs due to radioactive decay was calculated using the radioactive decay equation: where A t is the activity in the sample after t = 10 years, A 0 is the sample activity for t=0, λ is the radioactive decay constant =ln2/T 1/2 =0.023 god -1 , and t is the time of 10 years.
It is 20% for sample SB2, and for sample SB4 21%, which is significantly lower than previously given values.This shows that the reduction in 137 Cs originates from the washout of moss by atmospheric water and its transfer to the environment, i.e. expansion of pollution, while for 40 K this process does not occur as it is naturally distributed over all parts of our planet.
There was no correlation in accumulation between 137 Cs and 40 K in the mosses collected in Sokobanja and Gamzigradska banja.The absence of any correlation between the activity concentrations of these radionuclides in mosses has also been demonstrated in other radioecological studies (Korobova et al., 2007), but it is contrary to the findings of Cevik and Celik (2009).
Activity concentrations of analyzed radionuclides varied with moss species, as well as among samples of the same species collected from different localities.Different activity concentrations of radiocesium in the same moss species from different locations are mostly due to the non-uniform contamination of locations after the Chernobyl accident.While intraspecific variation in radioactivity is due to different deposition and washout rates, interspecific variation of activity concentrations is due to different morphology and anatomy (Sabovljević et al., 2005;Cevik and Celik, 2009), leading to different accumulations of radioactivity from dry and wet deposition.
In this work, mosses have been shown to be good biomonitors for atmospheric radionuclide deposition.Although substantially decreased, 137 Cs is still present in moss tissues.This radioactivity is thought to originate mainly from the Chernobyl accident.Monitoring of 137 Cs activity in mosses thus provides a very sensitive method for the detection of this radionuclide and should be performed as a means of continuous evaluation of environment pollution.

Table 1 .
Continued Fig. 1.Average activity of 137 Cs (Bq/kg) and standard deviation in moss samples collected on localities SB2 and SB4 in the period 2000-2010.

Table 2 .
Average activities of 137 Cs and 40 K (Bq/kg), standard deviation in moss samples collected on localities SB2 and SB4 in the period 2000-2010.