Determination of Trace and Heavy Metals

in Selected Samples of Oregano (Origanum

Vulgare L.) from Bosnia and Herzegovina

Fikreta Behmen1 , Šaćira Mandal2 , Vigor Dundović2 , Mersija Delić1 ,

and Senad Murtić1(B)
1 Faculty of Agriculture and Food Sciences, University of Sarajevo, Sarajevo,
Bosnia and Herzegovina
s.murtic@ppf.unsa.ba
2 Faculty of Pharmacy, University of Sarajevo, Sarajevo, Bosnia and Herzegovina

Abstract. Herbs are commonly used in traditional medicine and as spice around
the world. They consist of a number minerals essential to human health and nutri-
tion. Many of these microelements are crucial for various metabolic processes and
needed for normal physiological functions. The concentration of some essential
metals (Fe, Mn, Cu, Zn, and Cr) and heavy metals (Pb, Ni, and Cd) were analyzed
in selected samples of oregano (Origanum vulgare L.) collected from different
locations in Bosnia and Herzegovina. Preparation of samples was done using
wet digestion with acids, in triplicate and determined by flame atomic absorp-
tion spectrometry. The mean concentration (mg kg−1 ) of Fe, Mn, Cu, Zn, Cr, Pb,
and Ni ranged from 68.79–152.6; 17.48–27.30; 4.63–6.96; 16.81–51.28; 0.023–
0.036; 0.0020–0.0204; and 0.39–2.37, respectively. Cadmium was not detected
in any analyzed samples of oregano. These results were in agreement with other
published data except in the case of Pb which content was lower and within the per-
missible limit; and for Cd which do not detected during analysis. Thus, on the basis
of experimental outcome, it can be concluded that oregano collected from various
locations from BiH are safe and may not produce any harmful effect of metals
toxicity during their applications as spices as well as in different pharmaceutical
formulations.

Keyword: Trace metals · Heavy metals · Oregano · AAS

1 Introduction
Plants in the form of herbs and spices have been used in cuisine and different medicinal
as well as pharmaceutical formulations since ancient times [14, 19]. They contain many
various substances beneficial for human health but at the same time, there are harmful
and toxic elements (e.g. heavy metals) in a wide range of concentrations. Origanum
vulgare L. commonly known as oregano is often used as a spice in cuisines around
the world to improve or increase aroma, taste and smell of food [11, 12]. Also, as a

© The Author(s), under exclusive license to Springer Nature Switzerland AG 2022

M. Brka et al. (Eds.): CE-Food 2020, 10th Central European Congress on Food, pp. 157–162, 2022.
https://doi.org/10.1007/978-3-031-04797-8_14
158 F. Behmen et al.

herb, oregano is used as a medicinal plant in different pharmaceutical formulations of

traditional medicine as well as industrial preparations [2, 10].
A number of studies have demonstrated that O.vulgare L. has important health actions
and medicinal properties such as antioxidant, antimicrobial, antifungal, antigenotoxic,
anti-inflammatory effects, insecticidal and many others activities [3, 4]. Oregano contains
highly chemically and biologically active substances including minerals and trace metals
[16]. Metals have an important role in human health as essential components for a normal
growth of cells, various functions of cells, they catalyze many biochemical reactions in
the organism including the formation of red blood cells, hormones and vitamins, as well
as in the processes of photosynthesis and respiration, oxidation and reduction in plants
[2, 17].
The aim of this study was to determine the trace and heavy metals concentrations
in selected samples of Origanum vulgare L. from different locations in Bosnia and
Herzegovina.

2 Materials and Methods

Samples were collected from three different municipalities in Bosnia and Herzegovina:
Bugojno, Travnik and Sarajevo. Sample 1 as wild oregano were collected in Bugojno,
distanced from roads and industrial pollution. Fresh leaves were cut into small pieces
and air-dried in a dark place for two weeks, then stored in plastic bags until analysis.
Sample 2 as tea samples taken from a local Pharmacy in Travnik, declared as 100%
oregano tea. Sample 3 as a spice purchased at a local market in Sarajevo.

Sample Preparation and Analysis

Preparation of sample solutions for metal analysis from plant material was done by wet
digestion using nitric and sulfuric acid by method of Lisjak et al. [15]. Mass of 0.5 g
of dry plant matter was weighed into beakers of 100 ml, 5 ml of HNO3 and 2 ml of
H2 SO4 was added and this solution was left at room temperature for 5–6 h. After that,
the prepared solution was heated on a hot plate until the plant material has completely
decomposed (about 30 min). After cooling, the solution was filtered through filter paper
(blue band) into a 50 ml flask and diluted with redistilled water.
Metal content of collected samples of oregano was determined by FAAS (AAC-
7000 Auto Atomizer Changer, Shimadzu, Japan) as recommended by the method of
ISO 11047:1998. This instrument has a software program that directly reads the results
based on the calibration curve equation obtained for each tested element.
Preparation of working solutions of standards for tested metals.
Working solutions of standards (series of standards) for the element analysis were
prepared from original certified standard solutions (Merck, Darmstadt, Germany) of a
concentration of 1000 ppm. The dilutions were made with redistilled water (Millipure
Q water), and the concentrations in the series of standards for each element are selected
by the expected average values of the content of the test element in the plant material.
All the chemicals and reagents were of analytical grade and purchased from Merck
(Darmstadt, Germany). The sample and blank analyses were performed in triplicate, and
values presented as a mean of analyzed metals mg kg−1 .
 Determination of Trace and Heavy Metals in Selected Samples of Oregano 159

3 Results and Discussion

In this work, the content of trace and heavy metals concentration was determined in the
plant species of O.vulgare L. as a medicinal and aromatic herb, collected from different
locations in Bosnia and Herzegovina. The results of content of trace and heavy metals in
three oregano samples from Bosnia and Herzegovina municipalities (Bugojno, Travnik,
and Sarajevo) labeled as Sample 1, Sample 2, and Sample 3, were presented in Table 1.

Table 1. Trace and heavy metal content in different oregano samples from Bosnia and Herzegov-
ina

Element Sample 1 Sample 2 Sample 3

Cadmium, Cd mg kg−1 n.d.* n.d.* n.d.*
Chromium, Cr mg kg−1 n.d.* 0.036 0.023
Copper, Cu mg kg−1 6.96 4.63 4.87
Iron, Fe mg kg−1 108.7 68.79 152.6
Lead, Pb mg kg−1 n.d.* 0.0204 0.0020
Manganese, Mn mg kg−1 26.28 17.48 27.30
Nickel, Ni mg kg−1 0.39 1.46 2.37
Zinc, Zn mg kg−1 51.28 34.48 16.81
Values presented as mean (mg kg−1 ) of dry matter; *n.d.- not detected.

The concentrations of eight elements (Cd, Cr, Cu, Fe, Pb, Mn, Ni, and Zn) in
the digested samples of oregano were determined by Flame Atomic Absorption Spec-
trometry (FAAS). Among the analyzed metals, lead (Pb) was at the detection limit of
0.002 mg/g and the concentration of the rest of metals are shown as mean of triplicate
(mg kg−1 ) (Table 1). The most abundant metal among the microelements was iron (Fe)
while lead concentration was the lowest in tested samples of oregano. The content of
trace and heavy metals in the selected three samples followed the trend: Fe > Zn >
Mn > Cu > Ni > Cr > Pb, while cadmium (Cd) was not detected in any of the tested
samples of oregano.
The concentration of iron was higher than other trace and microelements. According
to the World Health Organization (WHO) concentration maximum or minimum for Fe
in medicinal herbs has not been established. Our results as well as results from other
authors show wide variations of iron in oregano species depending on various climatic
and geographic conditions [3, 14, 18]. Results of this study are comparable to contents
of iron found in work of Bukva [7] and colleagues which investigated the content of
iron in oregano and different spices from BiH. Also, our findings for Fe are similar
with the results of UAE [8]. Iron has a key function in the body including oxygen
transport, energy production, and immunity. On the other hand, higher concentrations
of iron are associated with toxicity and damage of various organs (e.g., liver damage)
and impairments of many metabolic pathways and the cardiovascular system. Also, its
160 F. Behmen et al.

increase concentrations effects on plant growth which depended on soil properties and
geographic region of growth of plant.
Zinc, after iron is the second trace element in higher concentrations that we detected.
The range of Zn concentrations observed in this study was from 16.81 to 51.28 mg kg−1 .
The content of zinc is the same or similar to the results of other studies [2, 3, 5, 6]. As one
of the more important and essential microelements for normal growth and development
of humans, Zn is responsible for protein and DNA synthesis and many other biochemical
functions. Although our knowledge of zinc toxicity is scarce and not available yet it is
well known that a high content of Zn of above permissible limits causes toxic effects on
the human immune system, blood lipoprotein levels and copper levels.
Manganese is a necessary microelement for plant as well as animal growth. We found
concentrations of Mn in the range of 17.48 to 27.30 mg kg−1 which are comparable with
findings in studies that determined manganese and other trace elements in oregano plant.
As detected concentrations of manganese are well below the max. limit and permissible
levels it does not influence human and plant growth [12, 18].
The concentrations of copper varied from 4.63 to 6.96 mg kg−1 in Sample 1, Sample
2 and Sample 3, which is similar to the amount of copper observed in previous studies
[10, 19]. In line with iron levels, the recommended limits by the WHO have not been
established yet for the Cu in herbal medicine. Therefore, different countries have different
referent and regulatory value limits for copper because this trace metal is an essential
component of many enzymes and plays an important role in a number of physiological
processes including free radical elimination, bone and specific tissues development and
many others.
Nickel (Ni) concentrations in leaves are generally between 0.05 and 5 mg kg−1 in
plants. Our results for concentration of this metal ranged from 0.39 to 2.37 mg kg−1
which is in line with the forementioned values [1, 18]. Nickel is an essential element for
plants as well as humans. In the body it is a structural consistent of the enzyme urease, and
necessary for the regulation of lipid content in tissues. Also, nickel in small quantities
is involved in the formation of red blood cells. The WHO states that the permissible
limit of nickel in medicinal plants is 1.5 mg kg−1 while daily requirement for humans is
1 mg kg−1 . However, Ni toxicity in humans is not a very common occurrence because
its absorption by the body is very low [20]. Higher concentrations of Ni can cause severe
diseases and various pathological effects and its toxicity might be attributed to nickel
interference with the physiological processes of zinc and calcium [21].
Chromium concentrations detected in our study were below 5 mg kg−1 . This finding
was in line with the findings of other investigations [23]. As chromium concentrations
in plant depends of plant parts and it’s decreased presence in the soil.
The content of lead in the analyzed Sample 3 and Sample 2 ranged from 0.0020
to 0.0204 mg kg−1 . In Sample 1 as a wild oregano plant Pb was not detected. The
WHO maximum permissible limit of lead in medicinal herbs is 10 mg kg−1 . The
obtained results showed that the concentrations of lead in Sample 2 and Sample 3
were below the permissible limit which is good having in mind that lead is a highly
toxic heavy metal, especially from environmental pollution/pollutant sources [23]. Lead
 Determination of Trace and Heavy Metals in Selected Samples of Oregano 161

can form complexes with crucial biomolecules and adverse effects or affects their func-
tions. Its exposure may affect blood, immune, nervous, renal, muscular, reproductive
and cardiovascular systems causing severe impairments and seriously defects [21].
Cadmium was not detected in any of the analyzed samples of oregano. In previous
studies, the content of Cd was above the permissible limit set by WHO for medicinal
herbs and plants (0.3 mg kg−1 ). The higher level of cadmium has a serious toxicological
effect on human health, especially for kidneys. Excretion of Cd is very slow and its
accumulation in human kidneys during long periods of time causes irreversible and
damaging effects on the liver, the vascular and immune system, and impairing of the
renal tract [8].

4 Conclusion
Metal contents in oregano (Origanum vulgare L.) were determined by FAAS. In tested
selected samples of this herb i.e. Sample 1, Sample 2, and Sample 3 had the lowest
concentration of lead and cadmium was not detected in any analyzed samples. The results
of this study indicated that selected oregano samples from different location in BiH are
safe for use as tea, spice or medicinal herbs in various pharmaceutical formulations.

References
1. Akram, S., Najam, R., Rizwani, G.H., Abbas, S.A.: Determination of heavy metal con-
tents by atomic absorption spectroscopy (AAS) in some medicinal plants from Pakistani
and Malaysian origin. Pak. J. Pharm. Sci. 28(5), 1781–1787 (2015)
2. Annan, K., Kojo, A.I., Cindy, A., Samuel, A.-N., Tunkumgnen, B.M.: Profile of heavy metals
in some medicinal plants from Ghana commonly used as components of herbal formulations.
Pharmacognosy Res. 2(1), 41–44 (2010)
3. Arceusz, A., Wesolowski, M.: Essential metals and phenolic acids in commercial herbs and
spices. Multivariate analysis of correlations among them. Open Chem. 13, 1196–1208 (2015)
4. Arora, D.S., Kaur, J.: Antimicrobial activity of spices. Int. J. Antimicrob. Agents 12(3),
257–262 (1999)
5. Başgel, S., Erdemoğlu, S.B.: Determination of mineral and trace elements in some medicinal
herbs and their infusions consumed in Turkey. Sci. Total Environ. 359, 82–89 (2006)
6. Bielicka-Giełdoń, A., Ryłko, E.: Estimation of metallic elements in herbs and spices available
on the Polish Market. Pol. J. Environ. Stud. 22(4), 1251–1256 (2013)
7. Bukva, M., Kapo, D., Huseinbašić, N., Gojak-Salimović, S., Huremović, J.: Iron content in
fruits, vegetables, herbs and spices samples marketed in Sarajevo Bosnia and Herzegovina.
Kem. Ind. 68(7–8), 281–287 (2019)
8. Dghaim, R., Khatib, S.A., Rasool, H., Khan, M.A.: Determination of heavy metals concentra-
tion in traditional herbs commonly consumed in the United Arab Emirates. J. Environ. Public
Health, 2015, Article ID 973878, 1–6 (2015)
9. Fischer, A., Brodziak-Dopierała, B., Loska, K., Stojko, J.: The assessment of toxic metals in
plants used in cosmetics and cosmetology. Int. J. Environ. Res. Public Health 14(1280), 1–12
(2017)
10. García-Galdeano, J.M., Villalón-Mir, M., et al.: Zn, Cu, and Fe concentrations in dehydrated
herbs (Thyme, Rosemary, Cloves, Oregano, and Basil) and the correlation with the microbial
counts of listeria monocytogenes and other foodborne pathogens. Foods 9(1658), 1–18 (2020)
162 F. Behmen et al.

11. Gilling, D.H., Kitajima, M., Torrey, J.R., Bright, K.R.: Antiviral efficacy and mechanisms of
action of oregano essential oil and its primary component carvacrol against murine norovirus.
J. Appl. Microbiol. 116, 1149–1163 (2014)
12. Huremović, J., Badema, B., Muhić-Šarac, T., Selović, A., Memić, M.: Heavy metal contents
in spices from markets in Sarajevo Bosnia and Herzegovina. Kem. Ind. 63(3–4), 77–81 (2014)
13. ISO 11047:1998 Soil quality — Determination of cadmium, chromium, cobalt, copper, lead,
manganese, nickel and zinc — Flame and electrothermal atomic absorption spectrometric
methods
14. Kostić, D., Mitić, S., Zarubica, A., Mitić, M., Veličković, J., Randjelović, S.: Content of trace
metals in medicinal plants and their extracts. Hem. Ind. 65(2), 165–170 (2011)
15. Lisjak, M., Špoljarević, M., Agić, D., Andrić, D.: Practicum in Plant Physiology. Faculty of
Agriculture, Osijek, p. 25 (2009)
16. Morshedloo, M.R., Salami, S.A., Nazeri, V., Craker, L.E.: Prolonged water stress on growth
and constituency of Iranian of oregano (Origanum vulgare L.). J. Medicinally Active Plants
5(2), 7−19 (2016)
17. Odoh, R., Ajiboye, O.E.: Quality assessment of some selected herbal medicinal products
consumed in Wukari, Taraba State. Acta Sci. Microbiol. 2(9), 28–36 (2019)
-
18. Randelović, S.S., Kostić, D.A., Zarubica, A.R., Mitić, S.S., Mitić, M.N.: The correlation of
metal content in medicinal plants and their water extracts. Hem. Ind. 67(4), 585–591 (2013)
19. Ražić, S., Onjia, A., Ðogo, S., Slavković, L., Popović, A.: Determination of metal content in
some herbal drugs – empirical and chemometric approach. Talanta 67(1), 233–239 (2005)
20. Rodak, B.W., de Moraes, M.F., Lopes Pascoalino, J.A., de Oliveira Junior, A., de Castro, C.,
Pauletti, V.: Methods to quantify nickel in soils and plant tissues. R. Bras. Ci. Solo, 2015. 39
788−793 (2015)
21. Sabudaka, T., Kaykioglua, G., Ongena, A., Dokmecia, H., Celika, S.O., Dokmeci, I.: Deter-
mination of nickel and lead contents in soil and plant in Corlu, Turkey. J. Environ. Prot. Ecol.
9(3), 557–565 (2008)
22. WHO guidelines for assessing quality of herbal medicines with reference to contaminants
and residues
23. URL:apps.who.int/medicinedocs/documents/s14878e/s14878e.pdf (15.3. 2020.)
24. Zaguła, Z., Fabisiak, A., Bajcar, M., Czernicka, M., Saletnik, B., Puchalski, C.: Mineral
components analysis of selected dried herbs. Econtechmod. Int. Quart. J. 5(2), 121–124
(2016)