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 Analytical Letters

ISSN: 0003-2719 (Print) 1532-236X (Online) Journal homepage: http://www.tandfonline.com/loi/lanl20

Characterization of Heavy Metal Air Pollution in

Romania Using Moss Biomonitoring, Neutron
Activation Analysis, and Atomic Absorption
Spectrometry

Claudia Stihi, Ion V. Popescu, Marina Frontasyeva, Cristiana Radulescu,

Antoaneta Ene, Otilia Culicov, Inga Zinicovscaia, Ioana Daniela Dulama,
Simona Cucu-Man, Radu Todoran, Anca Irina Gheboianu, Alin Bucurica,
Iulian Bancuta & Gabriel Dima

To cite this article: Claudia Stihi, Ion V. Popescu, Marina Frontasyeva, Cristiana Radulescu,
Antoaneta Ene, Otilia Culicov, Inga Zinicovscaia, Ioana Daniela Dulama, Simona Cucu-
Man, Radu Todoran, Anca Irina Gheboianu, Alin Bucurica, Iulian Bancuta & Gabriel Dima
(2017): Characterization of Heavy Metal Air Pollution in Romania Using Moss Biomonitoring,
Neutron Activation Analysis, and Atomic Absorption Spectrometry, Analytical Letters, DOI:
10.1080/00032719.2016.1275661

To link to this article: http://dx.doi.org/10.1080/00032719.2016.1275661

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 ANALYTICAL LETTERS
https://doi.org/10.1080/00032719.2016.1275661

ENVIRONMENTAL ANALYSIS none defined

Characterization of Heavy Metal Air Pollution in Romania

Using Moss Biomonitoring, Neutron Activation Analysis,
and Atomic Absorption Spectrometry
Claudia Stihia,b , Ion V. Popescua,b, Marina Frontasyevac, Cristiana Radulescua,b,
Antoaneta Ened , Otilia Culicovc,e, Inga Zinicovscaiac,f, Ioana Daniela Dulamab ,
Simona Cucu-Mang, Radu Todoranh, Anca Irina Gheboianub, Alin Bucuricab,
Iulian Bancutab, and Gabriel Dimaa,b
a
Faculty of Sciences and Arts, Valahia University of Targoviste, Targoviste, Romania; bInstitute of
Multidisciplinary Research for Science and Technology, Valahia University of Targoviste, Targoviste, Romania;
c
Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Dubna, Russian Federation; dFaculty
of Sciences and Environment, Dunarea de Jos University of Galati, Galati, Romania; eNational Institute for
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Research and Development in Electrical Engineering, Bucharest, Romania; fHoria Hulubei National Institute for
R&D in Physics and Nuclear Engineering, Magurele, Romania; gFaculty of Chemistry, Alexandru Ioan Cuza
University of Iasi, Iasi, Romania; hTechnical University of Cluj-Napoca, North University Center, Baia Mare,
Romania

ABSTRACT ARTICLE HISTORY

Heavy metal air pollution in Romania was investigated by using Received 28 October 2016
passive moss biomonitoring. This study is a component of an Accepted 19 December 2016
international UNECE ICP Vegetation Program – moss survey. A total KEYWORDS
of 330 samples of Hypnum cupressiforme, Hylocomium splendens, Flame atomic absorption
Pleurozium schreberi, and other mosses were collected in Romania in spectrometry (FAAS);
the summer and autumn of 2010. The concentrations of aluminum, graphite furnace atomic
cadmium, chromium, copper, iron, lead, nickel, vanadium, and zinc absorption spectrometry
were determined by graphite furnace/flame atomic absorption (GFAAS); heavy metals; moss
spectrometry and instrumental neutron activation analysis. The biomonitoring; NAA;
results were statistically processed to obtain spatial distribution neutron activation analysis
maps of factor scores based on elemental concentrations in the
moss, together with the spatial distribution maps of heavy metals in
moss. The median concentrations of cadmium, 1.20 mg/kg dry
weight, and lead, 30.8 mg/kg dry weight, were high compared with
other European countries. The results revealed that the atmospheric
deposition of these metals is a problem in north and northwest
Romania.

Introduction
An important worldwide problem is environmental pollution. Heavy metals from
ecological systems (Greger 2004) due to human activity may become pollutants with high
chemical stability and toxicity (Adriano 2001). In Romania, mining, metal smelting, waste

CONTACT Claudia Stihi claudia.stihi@valahia.ro Faculty of Sciences and Arts, Institute for Multidisciplinary Research
for Science and Technology, Valahia University of Targoviste, 2 Carol I Street, 130024 Targoviste, Romania; Antoaneta Ene
aene@ugal.ro Faculty of Sciences and Environment, Department of Chemistry, Physics and Environment, Dunarea de
Jos University of Galati, x 47 Domneasca Street, 800008, Galati, Romania.
Color versions of one or more figures in the article can be found online at www.tandfonline.com/lanl.
This paper is part of a special issue of papers presented at IC-ANMBES 2016 from 29 June to 1 July, 2016 in Brasov, Romania
organized by Monica Florescu.
© 2017 Taylor & Francis
 2 C. STIHI ET AL.

storage and processing, and agriculture are major contributors to the concentrations of
metals in air.
Since the first use of the terrestrial moss as biomonitors (Rühling and Tyler 1973), this
approach has been widely used to estimate atmospheric deposition of heavy metals (Berg
and Steinnes 1997; Harmens, Norris, and the participants of the moss survey 2008;
Harmens et al. 2008). The first systematic study on atmospheric pollution with heavy
metals based on the terrestrial moss analysis in Romania was performed from 1995 to
2001 in north Romania (Lucaciu et al. 2004, 2010; Harmens, Norris, and the participants
of the moss survey 2008). Heavy metals in air in Romania by moss biomonitoring was used
for the first time on the national scale in 2010 as part of the United Nations Economic
Commission for Europe International Cooperative Programme on Effects of Air Pollution
on Natural Vegetation and Crops (UNECE ICP Vegetation). Some results of the 2010
Romanian moss survey were reported in Harmens et al. (2013).
This study reports the spatial distribution of aluminum, cadmium, chromium, copper,
iron, lead, nickel, vanadium, and zinc in terrestrial moss from the 2010 Romanian
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survey by graphite furnace/flame atomic absorption spectrometry (GFAAS/FAAS) and

instrumental neutron activation analysis (INAA). The goal of this study was to identify
the most polluted regions and the sources of these elements.

Experimental
Sampling
A total of 330 moss samples were obtained from the summer and autumn of 2010 using the
experimental protocols of the 2010 and 2011 moss survey (ICP Vegetation 2010). The
species collected included Hypnum cupressiforme, Pleurozium schreberi, Hylocomium
splendens, Polytrichum formosum, and Dicranium scoparium at least 300 m from main
roads, populated areas, and industries, and at least 100 m from smaller roads and houses.
The locations of the sampling sites are provided in Figure 1. The longitude, latitude, and
elevation for each site were established using a global position system. Additional
information for each site included the name; weather conditions; topography described
as plain, slope, or ridge; the species of moss sampled; distance to neighboring shrubs,
the use of surrounding land, distance to the potential emission sources of metals; and
the presence of visible dust particles.

Sample preparation and instrumentation

The collected moss samples were cleaned to remove foreign matter, dried to constant
weight at 40°C for 48 h, and milled to a fine powder.
For atomic absorption, 0.4 g of dried moss was digested using a Speedwave MWS-2
Berghof microwave system. The samples were placed in the Teflon digestion vessels, and
7 mL of concentrated HNO3 and 2 mL of 30% H2O2 were added. The digestion program
included: step 1, 180°C, with a 10 min ramp at 500 W and 20 bar; step 2, 180°C, with a
20 min hold at 500 W and 20 bar. The digested solutions were cooled, filtered, and diluted
to 25 mL.
The concentrations of cadmium, copper, and lead were determined using a GFAAS/
FAAS Avanta GBC spectrometer at the Institute of Multidisciplinary Research for
 ANALYTICAL LETTERS 3
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Figure 1. Distribution of the sampling points in Romanian moss survey conducted from 2010 to 2011.

Science and Technology from Valahia University of Targoviste, Romania. The optimal
instrumental parameters and detection limits for the determined elements are provided
in Table 1.
The concentrations of Al, V, Cr, Fe, Ni, and Zn were determined by INAA using an
IBR-2 pulse fast reactor (Frontasyeva and Pavlov 2000), Frank Laboratory of Neutron
Physics from the Joint Institute for Nuclear Research, Dubna, Russian Federation. Two
types of irradiation were performed in this study. Short irradiation was performed for
3 min in a thermal channel for the determination of short-lived isotopes (Al and V) in
0.3 g of moss heat sealed in polyethylene foil. Gamma-ray spectra were recorded
9–10 min following irradiation. The long irradiation was carried out for 72 h in a
cadmium-screened channel to determine the elements associated with long-lived radionu-
clides Cr, Fe, Ni, and Zn in 0.3 g samples of moss in aluminum cups. The spectra of the
induced gamma activity were recorded following 20–23 days of cooling. The gamma
spectra of the moss were measured with an HPGe detector with a resolution of 1.9 keV
for the 60Co 1332 keV line. Data processing was performed using Gennie 2000 and
specialized software described in the literature (Ostrovnaya et al. 1993; Ostrovnaya
2000). The radionuclides used in this study for neutron activation analysis and their
γ-energies are summarized in Table 2.

Table 1. Instrumental conditions and detection limits for Cu, Cd, and Pb determined by AAS analysis of
moss.
Element Wavelength (nm) Lamp current (mA) Detection limit (mg/kg)
Cu 324.7; 327.4 30 0.02; 0.5
Cd 228.8; 326.1 3 0.009; 4
Pb 217.0; 283.3 5 0.16; 2.5
 4 C. STIHI ET AL.

Table 2. Radionuclides used for neutron activation analysis and their γ-energies.
Element Isotope Half life Gamma energy (keV)
28
Al Al 2.2 m 1778.9
51
Cr Cr 27.7 d 320.0
59
Fe Fe 44.5 d 1099.2
58
Ni Co 70.9 d 810.8
52
V V 3.8 m 1434.0
65
Zn Zn 244.0 d 1115.5

Quality control
The quality control for the AAS determinations was provided using the standard reference
material SRM 1571—orchard leaves from the National Institute of Standards and
Technology, NIST. The standard reference was prepared using the same preparation
procedure, and the AAS measurements were performed using the same instrumental
conditions. The measured concentrations of cadmium, copper, and lead were within the
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certified values, and the recoveries were between 98.4 and 103.8%.
The quality control of the INAA measurements was also obtained using standard
reference materials that included SRM 2709 trace elements in soil, 1633b coal fly ash,
and 2711 Montana soil, all obtained from NIST. The SRMS were irradiated using the
same protocol as the moss. The determined concentrations of Al, V, Cr, Fe, Ni, and
Zn were within the recommended values, and the recoveries of the elements were
between 97.4 and 100.8%.

Results and discussion

The results of the statistical analysis of aluminum, cadmium, chromium, copper, iron, lead,
nickel, vanadium, and zinc concentrations in terrestrial mosses collected during the
Romanian moss survey in 2010 are presented in Table 3.
The order of the elements in moss samples from Romania, according to their
abundance, was Cd < Ni < V < Cr < Pb <Cu < Zn < Fe < Al. A high value of the
coefficient of variation (CV) was only found for Cd (121.08%). A moderate variation exists
in the concentrations of V, Cr, and Ni (25% < CV < 50%). A weak variability (CV < 25%)
was obtained for Al, Fe, Zn, Cu, and Pb.
The median values of the elemental concentrations were determined in the moss were
compared with those from other Balkan countries participating in the European survey

Table 3. Descriptive statistics of elements determined in moss in Romania (n ¼ 330) (mg/kg).

Element Mean Median Minimum Maximum Standard deviation Coefficient of variation (%)
Al 5000 3150 220.00 34400 72.91 1.45
V 7.34 4.89 0.38 49.20 2.81 38.33
Cr 8.19 4.98 0.68 62.20 3.00 36.67
Fe 2957 1670 237.00 29500 58.36 1.97
Ni 4.53 3.60 0.39 35.90 2.08 45.95
Zn 56.04 42.30 0.59 1440 9.35 16.69
Cd 2.00 1.20 0.12 101.00 2.42 121.08
Cu 38.52 17.80 0.23 627.38 8.54 22.18
Pb 30.63 30.80 2.20 119.93 3.82 12.49
 ANALYTICAL LETTERS 5

Table 4. Pearson’s correlation coefficients between elemental concentrations in moss in Romania.

Element Al Cd Cr Cu Fe Ni Pb V Zn
Al 1.00
Cd 0.05 1.00
Cr 0.95 0.05 1.00
Cu 0.04 0.52 0.06 1.00
Fe 0.95 0.05 0.98 0.04 1.00
Ni 0.86 0.05 0.87 0.04 0.89 1.00
Pb −0.19 0.59 −0.19 0.44 −0.18 −0.24 1.00
V 0.98 0.05 0.95 0.03 0.96 0.87 −0.17 1.00
Zn 0.49 −0.07 0.49 −0.06 0.46 0.65 −0.35 0.47 1.00
Coefficients in bold are statistically significant at p < 0.05.

Table 5. Factor analysis of the correlation matrix for moss from Romania.
Variable Factor 1 Factor 2 Factor 3
Al 0.80 0.21 0.14
V 0.79 0.19 0.15
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Cr 0.85 0.26 0.18

Fe 0.90 0.21 0.14
Ni 0.85 0.16 0.10
Zn 0.09 0.82 0.05
Cd 0.09 0.06 0.66
Cu 0.10 −0.04 0.64
Pb 0.11 0.24 0.63
%Variance 37 21 16
Eigenvalues 16.16 2.57 2.71
Coefficient loadings >0.60 are provided in bold.

(Harmens et al. 2013; Harmens et al. 2015). The median values of Al, V, Cr, Fe, and Ni
were comparable with those determined in Bulgaria, Albania, and Macedonia. Higher
median values were obtained in this work for Zn, Cu, Cd, and Pb.
Pearson’s correlation analysis was carried out to characterize the origin of the elements
in moss. The results are shown in Table 4. Strong positive correlations were
found between Ni and Al; Cr and Fe; Fe and Al, Cr; V and Al, Fe, Ni, Cr; and Cr and
Al (p < 0.05). Together with the correlation analysis, factor analysis (FA) was used. Three
factors were extracted from the results and interpreted as source categories contributing
to the concentrations of the determined elements (Table 5). Factor 1 is a mixture of
heavy crust components: Al (0.80), V (0.79), Cr (0.85), Fe (0.9), and Ni (0.85). Factor
2 is of anthropogenic origin: Zn (0.82), the main contributors to this factor being
mining and industrial sites from the northwestern of the country. Factor 3 includes
Pb (0.63), Cd (0.66), and Cu (0.64) which may be from gasoline, lead production, or
copper mining.
The spatial distribution maps of factor scores based on elemental concentrations
together with the spatial distribution maps of Fe, Zn, and Pb moss collected in the 2010
Romanian survey are shown in Figures 2, 3, and 4, respectively. The maps in Figures 3
(Factor 2) and 4 (Factor 3) suggest high metal concentrations, especially for the related
elements of anthropogenic origin, Zn, Cu, Cd, and Pb, in north and northwest Romania
due to mining operations associated with the flotation and smelting. The metallurgical
industry in the Baia Mare, Copsa Mica regions has been conducted for approximately
100 years. Previous studies from north Romania demonstrated high concentration of
 6 C. STIHI ET AL.

Figure 2. Geographical distribution of factor 1 based on elemental concentrations and iron in moss
collected in the Romanian in the 2010 survey.
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Figure 3. Geographical distribution of factor 2 based on elemental concentrations and zinc in moss
collected in 2010 Romanian survey.

Figure 4. Geographical distribution of factor 3 based on elemental concentrations and lead in moss
collected in the 2010 Romanian survey.

metals in the atmosphere (Damian et al. 2008; Lucaciu et al. 2004, 2010; Big, Lăcătuşu,
and Damian 2012; Mihali et al. 2013). Hence, it may be concluded that metal pollution
is a historical problem in this region.
 ANALYTICAL LETTERS 7

Conclusion
This study confirms that moss biomonitoring is valuable to characterize metal pollution in
air and to identify the regions with a high atmospheric pollution. The results from the 2010
Romanian moss survey show that high concentrations of metals were present in north and
northwest Romania due to historical pollution. Romanian moss monitoring may be used in
the future to supplement air programs because of its ability to characterize concentrations
at many sites and provide long-term trends. This study may provide the national human
health risk assessment with epidemiological measurements.

Acknowledgement
We thank Professor Octavian Duliu from the University of Bucharest for the help with data
interpretation.
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Funding
The research leading to these results has received funding from the research project “Nuclear and
related analytical techniques for Environmental and Life Sciences”, based on scientific protocols
nos. 3869-4-08/10 and 4080-4-11/13 between Valahia University of Targoviste (UVT), Romania
and the Joint Institute for Nuclear Research, Dubna, Russia.

ORCID
Claudia Stihi http://orcid.org/0000-0002-2058-1856
Antoaneta Ene http://orcid.org/0000-0002-6976-0767
Ioana Daniela Dulama http://orcid.org/0000-0003-0200-0867

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