ARSENIC CONTENT IN SAMPLES TEA PRESENT ON THE REPUBLIC OF SRPSKA MARKET, WITH

ATOMIC ABSORPTION SPECTROPHOTOMETRY METHOD

Mirjana Djermanović1,2 Ljubica .Bojanić1,2, Relja Suručić2

¹Public Health Institute, Banja Luka, Bosnia and Herzegovina

2
Medical Faculty, Department for Pharmacy, University of Banja Luka, Bosnia and Herzegovina

Mirjana Đermanović
Gundulićeva 76, 78 000 Banja Luka
0038765917307
mirjanadjermanovicbl@gmail.com

SUMMARY
1
Introduction:

Arsenic exists in various forms in nature and living organisms. Inorganic compounds are the most toxic arsenic

moieties, unlike the organic ones, which are considered to be harmless, because of the poor absorption by the cell.

Some sources of exposure to arsenic are drinking water, food grown on arsenic contaminated soil, air, smoke from

cigarettes etc. Toxic elements, including arsenic, which are present in some plants, can severely damage

haemopoietic, immune, nervous and reproductive systems. For this reason, a content of heavy metals is one of the

criteria for the use of plant material in the production of traditional medicines and herbal infusions.

This instigates need for constant and organized safety control of plants that are used as raw materials in

pharmaceutical industry.

Objective

The aim of this study is to determine the arsenic content in selected teas which are available on the market of the

Republic of Srpska. As an analytical method for measuring arsenic concentrations, we used atomic absorption

spectrophotometry (AAS).

Methods

All samples were mineralized by dry ashing (AOAC method 999.11 B) and arsenic contents were determined by the

atomic absorption spectrophotometer Agilent Technologies Series 200 with an air-acetylene burner and D2

background correction.

Results

Arsenic content ranged from 0,005mg/kg to 0,155mg/kg.

Conclusion

Arsenic content in samples was in compliance with the current legislation.

Key words: arsenic, tea, toxicity, atomic absorption spectroscopy

INTRODUCTION:

2
 Metals can be classified as one of the oldest poisons known. In the periodic table of the elements, there are

about 80 metal elements among which about 30 elements induce toxic effects in humans. In the past, metal

toxicology mostly followed effects of accute exposure, but today, concentration of metals and other poisons is

intensively monitored in living and working environment. If values are higher than the permitted ones, measures are

undertaken to reduce health risks of persons living or staying for longer periods of time in these areas. [1]. The term

’heavy metal’ refers to any metal chemical complex with a relatively high density, i. e. greater than 5 g/cm³, which

is toxic or poisonous at low concentrations [2].

Heavy metals reach atmosphere in the form of very fine dust particles from where they deposit in water and

soil. Their retention time in atmosphere depends on particle size. The smaller the particles, the longer the retention

times. Larger particles from urban and industrial sources mostly have short retention times, usually about 10 hours.

In case of small particles that can reach stratosphere, retention time may be longer than a year. In waters, particles

quickly settle to the bottom of a body of water as poorly soluble carbonates, sulphates and sulfides. When the

adsorption capacity of the sediment is exhausted, the concentration of metal ions in water increases. The acidic

environment causes metal ions in soil, which are movable and available to plants. Ionic forms are more dangerous

because they can react with other ions and thus form more toxic compounds. This means that there is a possibility of

heavy metals contamination in plants in acidic soils.

Primarily, plants absorb them through the root, from the soil, and, to a lesser extent, through over ground

organs from the atmosphere. Atmospheric heavy metals can be deposited on plant surfaces, especially leaf surfaces,

and mostly on surface soil. In this way, metals enter plants, and upon their consumption, they enter humans and

animals. In such a way, they are introduced to the food chain. Under normal conditions, there are three ways of how

metals can come into the body: through the skin, gastro-intestinal tract and respiratory tract. The International

Agency for Research on Cancer classifies arsenic into the group of compounds for which there is enough evidence

of carcinogenicity in humans. Cases of skin cancer in people exposed to arsenic compounds were described in the

nineteenth century. Carcinogenic effects of metals are particularly important because of their wide distribution,

application and long-term retention in the environment [1,2]

In nature, arsenic can be found in organic and inorganic compounds; and AsH 3 (arsine) is also a significant

compound. Main inorganic arsenic compounds are arsenic trioxide, sodium arsenite, and arsenic trichloride.

Inorganic compounds of pentavalent arsenic are arsenic pentoxide, arsenic acid, and lead and calcium arsenate.

3
Organic compounds are considered harmless for their poor cell absorption. Inorganic arsenic compounds are highly

reactive, influence a series of intracellular reactions, and are used as pesticides [1]. Pollution of the environment by

arsenic also occurs as a result of natural phenomena such as volcanic eruptions and fires. The application of

intensive agro-technical measures in modern agricultural production, proximity to industrial plants, mines, roads,

inevitably leads to contamination of soil and plants with pesticides and heavy metals.

Consumption of food grown on arsenic contaminated areas is one of the sources of chronic arsenic

exposure. Arsenic compounds are found not only in food of plant origin such as tea, but also in food of animal origin

such as fish and shells, which contain organic arsenic compounds. This research shows that food can be an important

way of arsenic exposure and that such an exposure might have long-term negative effects on people [2, 3].

Symptoms of arsenic poisoning are fever, vomiting, diarrhea, difficulty breathing and swallowing, enlarged liver,

cyanosis, arrhythmia, coma, oliguria, and tubular necrosis.

Anemia, leukopenia, and especially granulocytopenia, which are reversible, can be noticed. Arsenic in gas form can

interact with hemoglobin causing hemolysis and anemia. Symptoms of chronic arsenic poisoning are changes on

skin and nails (Mees’ lines). Neurotoxic changes in peripheral and central nervous system can be seen after chronic

exposure to arsenic, and they start with sensor changes, paresthesia and muscle weakness. Central nervous system

disorders occur after signs of memory loss and personality changes, which can develop to the stage of obvious

manifestation of encephalopathies [1]. Arsenic belongs to the group A carcinogenic. Most commonly, all types of

skin cancer can appear, i. e. basal cell carcinoma and squamous cell carcinoma, which can develop to local and

distal metastases [1]. A biological indicator for arsenic intoxication is the determination of its content in blood,

urine, and arsenic concentration in hair and nails. Bearing in mind short-term retention in blood (half-retention is

four days), blood determination might be useful only few days after poisoning. Arsenic concentration in urine is

significant in case of chronic poisoning. It remains in hair and nails for years after poisoning [1].

Plants represent direct or indirect sources of minerals in human nutrition. Special significance has plant species for

production of mono component tea or tea blends, which have been applied in traditional medicine. Plantation and

self-seeding plants serve as a source of medicinal plant raw materials [4]. Its large production and consumption

indicate that this drink is very popular, whether black, green or herbal [5].

OBJECTIVE:

4
 Testing samples of various herbal teas, taken from pharmacies, aimed to determine arsenic content in order

to gain some insight into safety of these products, bearing in mind their extensive use.

MATERIAL AND METHODS :

Arsenic content was examined in tea preparations which can be found in the Republic of Srpska and are

available to patients in pharmacies. Tested samples covered 10 different tea-producers. Green tea, chamomile,

mint, and rose hip were tested. The presence and content of arsenic were determined by the atomic absorption

spectroscopy method, on the atomic absorption spectrometer Agilent 200 Series AA Systems, using hydride

technique. The sample is homogenized in accordance with the standard EN 13804. Samples are treated with acid

and ash residues are steamed until dried, and then mineralized at 425 ºC gradually increasing temperature. The

process is repeated until the sample totally burns. An ash has to be white/gray or a little bit coloured. The ash is then

dissolved in hydrochloric acid; and if necessary, the sample is filtered before dissolving to the certain volume.

Solutions for determination of As were pre-reduced (As5+ to As3+) prior to analysis. This was achieved by adding a

reduced solution containing 5% (w/v) KI and 5% (w/v) ascorbic acid. A 10 mL of solution which we used for

determination of mercury was transferred in 50 mL volumetric fl ask and then we added 1 mL of 5% KI and 1 mL

ascorbic acid. Finally, solution was made up to mark with 2M HCl. Arsenic is quantitatively determined at a

wavelength of 193,7nm. For determining a calibration curve, certified standards AccuTraceTM Reference Standard

(USA) concentration 1000 g/l were used. High purity deionized water was used for dilution of standards and

samples. Each sample was measured in duplicate. Arsenic content values measured in herbal tea samples are

expressed in mg/kg. Statistic data processing covers determination of basic parameters in statistical analysis:

arithmetic mean and standard deviation (SD).

Characteristics of tested samples are given in Table 1.

Table 1: The names of medicinal plants and plant parts used in research [4,6]

Fig. 1 shows a calibration curve used to determine arsenic content in tested samples. Correlation coefficient of the

calibration curve is 0.98777, with a linear equation y = 0.03045 + 32.29403x.

Arsenic contents determined by atomic absorption spectrophotometer are shown in a Table 2 and a Table 3.

Table 2. Arsenic content (As) in herbal teas (mg/kg)

5
Table 3. Arsenic content (As) in fruit teas (mg/kg)

DISCUSSION:

Fertile soils supply plants with all chemical elements necessary for growth. Thereby, soil can transmit

toxic elements to plants [7]. Additional sources of contamination of plants with heavy metals are atmospheric

precipitation, protective chemicals, fertilizers, industrial plants, thermal power plants, the proximity of mines and

frequent roads; arsenic is considered to have carcinogenic properties and its very low concentrations are highly toxic

to living organisms [7]. Toxic elements (Hg, As, Pb i Cd) which are present in some plants can severely damage

haemopoietic, immune, nervous and reproductive systems. For this reason, a content of heavy metals is one of the

criteria for the use of plant material in the production of traditional medicines and herbal infusions[8]. This

instigates need for constant and organized safety control of plants that are used as raw materials in pharmaceutical

industry.

The arsenic content in plants is normally lower than in soil and it does not influence plant growth and development.

Some authors appoint stimulant effects of low concentrations of As to root growth. However, a high concentration

of As has a negative effect [9]. A normal concentration of As in developed leaves of various plants is 1-1,7 ppm/dry

matter and the value of 5-20 ppm of As is considered toxic [10]. Karak and Bhagat [11] state the fact that As

concentration in fresh leaves and young tea shoots (China origin) is 0,024– 0,066, i. e. 0,021–0,073 mg/kg.

Maximum value of arsenic content was measured in the sample of elder tea, actually 0,17mg/kg, leading

to a conclusion that herbs were collected in the fields where arsenic from the atmosphere could contaminate elder

flowers used for tea preparation. A minimal value in bearberry leaves was 0,007mg/kg, Bearberry is collected as

a wild plant at elevation above 1200m, where pollution and contamination are certainly less than in urban and

industrial areas; consequently, there is a clear connection between results and herb origin. Low values were also

obtained in samples of: chamomile - flowers, green tea - leaves and mint - leaves.

As content in tea samples is below the value of 1 mg/kg , stipulated by the national legislation [12]. Ražić

and Kuntić examined As content in samples of mint, chamomile and rose hip [5]. Values of As content in mint

samples (original package) of 0,086 mg/kg and of 0,155 mg/kg (bulk) are significantly higher than 0,013±. Also

values of As content in rose hip samples of 0,101 mg/kg (original package) and of 0,088 mg/kg (bulk) are higher

than our results (0,04±0,014 mg/kg).

6
Szentmihályi et al. [13] measured As content of 3,63±1,60 mg/kg in nettle (herba), which is considerably different

from our test results.

CONCLUSION:

Sixteen samples of different types of herbal teas, whose consumption is significant among the population of

the Republic of Srpska, were analyzed for arsenic, a toxic element, in order to gain insight into safety of medicinal

plants used to prepare popular tea beverages. Measured values of As content are low, as expected, and good

regarding safety of teas. Today, there are various products on the market, including the ones that, in some

parameters, are not fit for human consumption and which have negative impacts on health and working ability. For

this reason, constant care and monitoring is required by the competent institutions for all products entering the

human organism. In this regard, the need for faster and more reliable quality control methods for medicinal plants is

imposed, with a special focus on the content of toxic elements.

Conflict of Interest: The authors declare that there is no conflict of interest.

REFERENCES:

7
1. R. Kastori, I. Maksimović, Ishrana biljaka, Vojvođanska akademija nauka i umetnosti, Novi Sad, 2008. (in

Serbian)

2. R. Kastori, Fiziologija biljaka, Feljton, Novi Sad, 1998. (in Serbian)

3. R. Kastori, N. Petrović, I. Maksimović-Arsenijević, Teški metali i biljke, u: R. Kastori (ed.), Teški metali u

životnoj sredini, Naučni institut za ratarstvo i povrtarstvo, Novi Sad, 1997, str. 197–257. (in Serbian)

4. N. Kovačević, Osnovi farmakognozije, Srpska školska knjiga, Farmaceutski fakultet, Beograd, 2002. (in Serbian)

5. S. Ražić, V. Kuntić, Diverse elements in herbal tea products consumed in Serbia using inductively coupled plasma

mass spectrometry, Int. J. Food Prop. 16 (2013) 1–8.

6. N. Kovačević, R. Jančić, 100 Lekovitih biljaka, kroz tradiciju i savremeni život srpskog naroda, Srpska školska

knjiga, Beograd, 2003. (in Serbian)

7. L. Kolomejceva-Jovanović, Hemija i zaštita životne sredine, Ekološka hemija, Savez inženjera i tehničara Srbije,

Beograd, 2010. (in Serbian)

8. D. Kostić, S. Mitić, A. Zarubica, M. Mitić, J. Veličković, S. Ranđelović, Content of trace metals in medicinal

plants and their extracts, Hem. Ind. 65 (2011) 165–170.

9. R. Kastori, N. Petrović, Uticaj teških metala na biljke. Teški metali i pesticidi u zemljištu, Zbornik, Poljoprivredni

fakultet, Institut za ratarstvo i povrtarstvo, Novi Sad, 1993, str. 55–72. (in Serbian)

10. A. Kabata-Pendias, H. Pendias, Trace elements in soils and plants. Third Edition, CRC PRESS, Boca Raton, FL,

2001.

11. T. Karak, M. R. Bhagat, Trace elements in tea leaves, made tea and tea infusion: A review, Food Res. Int. 43

(2010) 2234–2252.

12. Pravilnik o maksimalno dozvoljenim količinama za određene kontaminante u hrani Službeni glasnik RS, br.

68/2014. (in Serbian)

8
13. K. Szentmihályi, G. Marczal, M. Then, Medicinal plants in view of trace elements, Thaiszia- J. Bot. 16 (2006)

99 –107.

9
Table 1: The names of medicinal plants and plant parts used in research [4,6]

Material Part of the plant

Sample number Latin name
Commercial name
1 ZeleniR Tea siensis folium
2 Crni R
Tea siensis folium
3 KamilicaR Matricaria chamomilla flos
4 Nana R
Mentha piperita folium
5 Majčina dušica R
Thymus serpyllum folium, flos and stems
6 AnisR Anis fructus
7 ZovaR Sambuci flos flos
8 UvaR Uvae ursi folium
9 SljezR Althaea officinalis radix
10 KoprivaR Urtica dioica folium
KantarionR folium, flos and stems
11 Hypericum perforatum
ŠipakR fructus
12 Rosa canina
Hajdučka travaR folium, flos and stems
13 Achillea millefolium

10
Table 2. Arsenic content (As) in herbal teas (mg/kg)

Material First Second Standard

Sample number Average
Commercial name measurement measurement deviation
1 ZeleniR 0.012 0.014 0.013 0.001
2 Crni R 0.017 0.020 0.014 0.003
3 Kamilica R 0.011 0.007 0.009 0.003
4 Nana R 0.014 0.012 0.013 0.001
5 Majčina dušicaR 0.021 0.022 0.0215 0.001
6 AnisR 0.02 0.025 0.0225 0.003
7 ZovaR 0.17 0.12 0.145 0.035
8 UvaR 0.007 0.011 0.009 0.003
9 SljezR 0.09 0.07 0.08 0.014
10 KoprivaR 0.03 0.04 0.035 0.007
KantarionR 0.02 0.04 0.014
11 0.03
ŠipakR 0.03 0.05 0.014
12 0.04
Hajdučka travaR 0.026 0.027 0.001
13 0.0265

11
Table 3. Arsenic content (As) in fruit teas (mg/kg)

Material First Second Standard

Sample number Average
Commercial name measurement measurement deviation
1 Fruit mix 0.02 0.035 0,0275 0.010
2 Aronia 0.017 0.021 0.019 0.002
3 Cranberry 0.012 0.016 0.014 0.003

12
Fig. 1 shows a calibration curve used to determine arsenic content in tested samples. Correlation coefficient of the

calibration curve is 0.98777, with a linear equation y = 0.03045 + 32.29403x.

0,5

0,4

0,3
A

0,2
Equation y = a + b*x
Adj. R-Square 0,98777
Value Standard Error
B Intercept 0,03048 0,01464
0,1 B Slope 32,29403 1,79391

0,0

0,000 0,002 0,004 0,006 0,008 0,010 0,012 0,014 0,016

k o n ce n tra c ija , m g /L

13