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In Vitro Evaluation of Mutagenic Potential of

Somed Pesticide Used by the Agricultures in the
Southeast of Algeria

Article · March 2018

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World
Journal of
Life
Sciences
RESEARCH ARTICLE World Scientific Journals Press

In Vitro Evaluation of Mutagenic Potential of Somed Pesticide

Used by the Agricultures in the Southeast of Algeria
I. Ghenabzia*1, T. Lanez2, E. Lanez2,3
1
University of Sciences and Technology Houari Boumediene, Faculty of Natural and Life
Sciences, BP 32 EL Alia 16111 Bab Ezzouar Algiers, Algeria
2
University of El Oued, Faculty of Sciences and Technology, VTRS Laboratory, B.P.789,
39000, El Oued, Algeria
3
University of Ouargla, Chemistry Department, PO Box 511, 30000, Ouargla, Algeria

Abstract
Pesticides are among the most potentially hazardous compounds to human, animals, and the
environment, they can damage the environment and accumulate in ecosystems and cause
serious disturbances. Although they have highly specific actions, some pesticides are capable
of interacting with cellular structures directly or after processing by metabolic enzymes, and
form a covalent bond with DNA nucleosides to produce DNA adducts, The objective of this
study is to evaluate the in vitro mutagenic and/or carcinogenic potential of somed pesticide,
the most used in farmers in the region of El Oued (South Algeria), the evaluation is carried
out using electrochemical techniques based on cyclic voltammetry and chromatography
techniques based on high performance liquid chromatography. Both techniques indicate that
the pesticide somed has a very high in vitro mutagenic and carcinogenic potential.

Keywords: pesticide; cancer; somed; cyclic voltammetry; carcinogen; mutagen.

1. Introduction
Pesticides are chemicals which are synthesized for the purpose of killing living beings; it is a
generic term which includes a variety of categories such as insecticides, acaricides,
fungicides, algaecides, herbicides, weedicides and bactericides [1-2]. These chemicals that are
toxic by nature can contaminate the water we drink, the air we breathe, the food we eat and
the soil on which we walk [3-6].
As shown by numerous epidemiological studies, some types of cancer are increasing
particularly rapidly if the environment (water, air, food, soil ...) is contaminated with
pesticides [7-11]. The development of cancer in this case is due to the penetration of the active
ingredient of a pesticide in the body. This active material can be fixed by covalent bonding to
nucleophilic sites present in the DNA and then form adducts which can be mutagenic in a first
time, then carcinogenic [12,13]. Detection of adducts can therefore be a very useful tool for
evaluating the mutagenic and/or carcinogenic potential of pesticides [14]. In a previous work
we have reported the use of electrochemical techniques as a tool for the evaluation of the in
vitro mutagenic potential of abamectin pesticide [15]. In this paper we present the study of the
reaction of somed pesticide with thymidine using voltammetric and chromatographic
methods.

Received 03rd March 2018, Accepted 24th April 2018

Corresponding author email: imene.gh@gmail.com

World J Life Sci (2018):1(1), 1-6 1

2. Materials and methods
2.1. Chemical
HPLC-grade acetonitrile, dipotassium hydrogen phosphate (K2HPO4) and potassium
dihydrogen phosphate (KH2PO4) were purchased from Fisher Scientific Co. and used as
received. Somed was purchased from local stores in the region of El Oued and used as
received. High purity water was used in all experiments. All other reagents used were of
analytical grade.
2.2. Somed pesticide
Among several commonly used pesticides in the region of El Oued, somed pesticide is chosen
for evaluation of their in vitro mutagenic potential, table 1 resume its characteristics.

Table 1. Characteristics of somed pesticide

Pesticide Active ingredient Empirical formula Concentration dosage
Metalaxyl :8% C15 H21 NO4
Somed 8+65% 0.2-0.3%
Mancozeb: 64% C4 H6 N2 S4 Mn)b – (Zn)Y

2.3. Molecular Structure

The molecular structure of active ingredient of somed pesticide is presented in figure 1.

(A) (B)
Fig.1. Molecular Structure of active ingredient of somed pesticide (A) metalaxyl (B)
mancozeb
2.4. Description
Metalaxyl is a systemic, residual fungicide with curative and preventive properties against
oomycosis and, more specifically, against peronospora (downy mildew). Mancozeb is a
broad-spectrum, preventative fungicide with special activity on leaf diseases caused by
endoparasitic fungi. It has a side effect on mites. The combination of Metalaxyl and
Mancozeb results in a formula with a broader spectrum of activity and important benefits: It
interferes in the synthesis of fungal proteins by preventing the development of promycelium,
mycelium growth and production spores. It prevents the activity of sulfhydrilic enzymes in
general and cysteine in particular forming complexes with enzymes that contain metals such
as those involved in the production of ATP. By acting on multiple processes, controlled by
various genes, it prevents the appearance of resistance. It combines a good shock effect with a
persistence of up to three weeks. Containing Manganese and Zinc, it corrects the deficiencies
of these elements.
2.5. Action spectrum
Somed is authorized as a preventive and curative fungicide in the following crops: - broccoli,
cocombre, cauliflower, poultry, melon, onion, pasteque: Mildew. - Potato: Alternaria,
Anthracnose and Mildew. - tobacco: Blue mold - vineyard: Downy mildew.

3. Methods
3.1. Reaction of pesticides with nucleosides
The reaction of somed pesticide with thymidine nucleoside was carried out using phosphate
buffer 0.05 M solution at pH equal to 7.2 as a solvent. The general procedure was as follows:
An amount of newly supplied somed pesticide (0.5 mM) was added to 25 ml of phosphate

World J Life Sci (2018):1(1), 1-6 2

buffer containing the thymidine (1 mM). The solutions were shaken until the reaction
mixture become homogenous, and then incubated at 37 °C under agitation for 72 h. The
reaction mixture was then analyzed using cyclic voltammetry and liquid chromatography
techniques.
3.1.1. Cyclic voltammetry
Cyclic voltammetric measurements were carried out using voltalab40 PGZ301
potentiostat/galvanostat (radiometer analytical SAS). Experimentations were made in a
double walled electrochemical cell of 25 mL and conventional three electrode system was
employed. Glassy carbon working electrode (radiometer analytical SAS), having area 0.013
cm2, a platinum wire counter electrode, and an Hg/Hg2Cl reference electrode (3.0 M KCl).
Data acquisitions were accomplished with a Pentium IV (CPU 3.0 GHz and RAM 1 Gb)
microcomputer using VoltaMaster4 software version 7.08 (radiometer analytical SAS).
Graphs plot were carried out using OriginLab software version 2.0 (Integral Software,
France). Cyclic voltammetric measurements were run from 0 to 2.5 V. All measurements
were carried out at (37 ± 1 °C). Figure 1 shows the voltammogram of thymidine nucleoside
before the addition of somed pesticide, the anodic pick current density in this case is equal to
499,32 μA.cm-2, after the addition of somed and incubation at 37 °C for 72 hours, this anodic
pick was almost disappeared, its value become equal to 1.371 μA.cm -2. This decrease in
anodic pick current density is evidence of the formation of thymidine-somed adduct.

dT
1000 dT+ somed

800
Current density [µA/cm²]

600

400

200

0.0 0.5 1.0 1.5 2.0 2.5

Potential [ V]

Fig.1. Cyclic voltammograms of 1 mM solution of thymidine nucleoside in the absence

(green line) and in presence of 0.5 mM of somed pesticide (red line) in 0.05 M phosphate
buffer solution on a glassy-carbon electrode, pH= 7.2 at scan rate 100 mV/s.

The decrease in anodic pick curent density of thymidine nucleoside followed the addition of
somed pesticide can be used to calculate the yield of the formation of the adduct thymidine-
somed obtained from the reaction of thymidine nucleoside and somed besticide at 37 °C for
72 hours. This yield is calculated from electrochemical data using the following equation 1.

 i (t  0) 
Yield %  1 a   100 (1)
 i a ( t  72h ) 

Where i a ( t  0 ) represents the anodic pic current desity of pure thymidine nucleoside and
i a ( t  72h ) represents the anodic pic current desity of thymidine nucleoside in the precence of
somed pesticide after 72 hours of incubition at 37°C. Replacing i a ( t  0 ) and i a ( t  72h ) by
their values from the voltammogram in figure 1,

World J Life Sci (2018):1(1), 1-6 3

  1.371 
Yield %  1   100 99. 72 %
 499.32 

The high value of the yield indicates a quantitative reaction between thymidine nucleoside
and somed pesticide.
3.1.2. Chromatography
This has been done through high performance liquid chromatographic (HPLC) techniques to
detect the formation of thymidine nucleosides adduct using chromatograms. Analyses were
performed utilizing a high performance liquid chromatography (LC 20 AL equipped with
universal injector (Hamilton 25 µl) UV-VIS detector SPD 20A (Shimadzu). After incubation
of the reaction mixture of thymidine nucleoside and somed pesticide for 72 hours at 37 °C, it
was then analyzed under the following conditions: stationary phase, Shim-pack VP-ODS C18
(250 mm L. x 4.6 mm I.D., 5 μm). Flow rate: 1 mL/min. Oven temperature: 25 °C. UV
detector: detection wavelength 260 nm. Injection volume: 20 µL. Run time: 50 min. A linear
gradient was used for mobile phase as follows: linear gradient from 0% B to 12% B, 0-30
min; linear gradient from 12% B to 50% B, 30-50 min.
The obtained chromatogram of thymidine nucleoside shows two picks, the first is at retention
time equal to 2.5 min. and the second is at 30 min. Figure 3.

uV
500000

400000

300000

200000

100000

0
0 10 20 30 40 min

Fig.3. HPLC chromatogram of pure thymidine

After incubation of the mixutre of thymidine nucleoside and somed pesticide for 72 hours at
(37 ± 1 °C), the pick at 30 minutes disappeared and a new pick appeared with the same
intensity at 26.5 minutes, Figure 4.

uV

400000

350000

300000

250000

200000

150000

100000

50000

0
0 10 20 30 40 50 min

Fig.4. HPLC chromatogram of the reaction mixture of somed pesticide and thymidine
nucleoside after 72 hours of incubation under agitation at 37 °C

World J Life Sci (2018):1(1), 1-6 4

4. Conclusion
The in vitro evaluation of the mutagenic potential of somed pesticide, the most used in
agriculture filed in the region of El Oued (South Algeria) was studied using cyclic voltametry
techniques. The obtained results were confirmed by HPLC chromatography techniques. The
study was based on the detection of the thimidine-somed adduct. Both techniques confirmed
the interaction between thymidine nucleosides and somed pesticide. The yield of thymidine-
somed adduct, calculated using the decrease in the anodic pic current density was 99.72 %.

5. Acknowledgements
This research was supported by the Algerian ministry of higher education and was
administrated by VTRS laboratory under faculty of sciences and technology. The authors are
grateful to Dr. Ahmed Chenna for many valuable discussions and insights provided during the
course of this work. We are also indebted to Abdelkirm Rebiai and Ali Tliba for their help.

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