Contamination of the waters of the bays (Azito, Banco and Marcory) of the Ebrié lagoon by parabens.

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 Contamination of the waters of the bays
 (Azito, Banco and Marcory) of the Ebrié
 lagoon by parabens.
 KOUAKOU Kouamé1*, ETCHIAN Assoi Olivier3, SEKA Yapoga Jean1, KPIDI Yapo Habib1 and YAPO
 Ossey Benard1, 2.
 1
 Laboratory of Environmental Sciences (LSE), Training and Research Unit in Environmental Sciences and
 Management (UFR-SGE), Nangui Abrogoua University, 02 BP 801 Abidjan 02, Ivory Cost.
 2
 Central Environmental Laboratory of the Ivorian Anti-Pollution Centre (LCE-CIAPOL), 20 BP 650 Abidjan
 20, Ivory Cost.
3
 Laboratory of Animal Biology and Cytology, Natural Sciences Training and Research Unit (UFR-SN), Nangui
 Abrogoua University, 02 BP 801 Abidjan 02, Ivory Cost.

 * Author of correspondence: kkyan24@yahoo.com

Abstract
Emerging pollutants such as parabens are present in the bays of Abidjan. Thus, high concentrations of parabens
were measured in Azito Bay, with average concentrations ranging from 0.553 ±0.058 µg/L of PrP to 0.012
±0.008 µg/L of TCS. At Banco, mean concentrations ranged from 0.334±0.067 µg/L BzP to 0.037±0.011 µg/L
TCS. In Marcory, the values obtained are lower. The highest value is observed for BzP with a content of
0.135±0.055 µg/L. This contamination is marked by the massive entry of wastewater of all kinds through the
bays without prior treatment. Preventive measures for the protection of aquatic species must be taken to prevent
their contamination due to their large consumption by the Abidjan population.

1. Introduction
Over the past decades, the presence of micropollutants in aquatic environments has become a global
environmental issue of increasing concern. Micropollutants, also known as emerging contaminants, concern a
wide variety of substances of natural and anthropogenic origin from industrial products such as pharmaceuticals,
cosmetics and personal care products [1], [2], and [3]. Micropollutants are generally present in aquatic
environments at low concentrations ranging from a few ng/L to several μg/L and can, even in small doses, have
an impact on the environment, and especially on aquatic ecosystems [4].
Parabens, one of many emerging micropollutants, are used as preservatives in drugs, cosmetics and personal care
products to prevent contamination risks [5], [6] and [7]. Thus, their ubiquitous presence in the environment,
especially in aquatic environments is capable of causing toxic effects, in humans and aquatic organisms, even at
very low doses during continuous and chronic exposures due to their estrogenic activity [1], [4], [8], [2] and [9].
Numerous eco-toxicological studies have reported the potential of parabens as endocrine disruptors but they
remain unregulated in developing countries including Côte d'Ivoire.
The Ebrié lagoon system (south of Abidjan) serves several socio-economic needs (aquaculture, fisheries,
dredging, market gardening, transport and tourism) in the development of the city of Abidjan and indeed the
whole of Côte d'Ivoire [10]. Unfortunately, it is subject to several attacks due to the lack of sanitation
infrastructure [11] and [12]. It is thus the receptacle for large quantities of liquid waste (domestic and industrial
wastewater and runoff), often without any primary treatment.
Several studies including those by [1], [5], [4] and [8] have highlighted paraben contamination in aquatic
environments in many countries.
This one aims to assess the state of contamination of water, sediments and some aquatic species in the lagoon
bays (Azito, Banco and Marcory) of the city of Abidjan by parabens (Methylparaben (MeP), Ethylparaben (EtP),
Propylparaben (PrP), Butylparaben (BuP) and Benzylparaben (BzP)). It is part of the reinforcement of the
quality control of the lagoon environment. This study will be carried out in this area in order to shed light on the
contamination of this micropollutant on Ivorian soil.

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2. Method and materials
2.1. Geographical location and sampling stations
The Ebrié lagoon system is located on the southern coast of Côte d'Ivoire where it extends over 125 km between
3°40' and 4°50' West longitude and 5°20' North latitude. It is 4 to 7 km wide and covers an area of 566 km2. It is
made up of the Ebrié lagoon itself and the Aghien and Potou lagoons, which cover 523 km2 and 43 km2
respectively. It is separated from the Atlantic Ocean by a sandy barrier beach 8 km thick. Since 1950, it has been
connected to the sea by the 370 m wide Vridi canal (Figure 1).

 Legend
 Sampling point
 Ebrié lagoon
 Abidjan limit
 Department
 Atlantic ocean

Fig. 1 : Work zone

The Ebrié lagoon system is influenced to the west by the Agnéby River and to the east by the Comoé and Mé
Rivers. In the dry season (December to April), it is fed by freshwater streams, the main one being the Comoé
River during the flood season, from September to November. Rainfall is divided into two rainy seasons, the
major one occurring between late May and August and the minor one centred on October (Figure 2).

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 precipitation temperature

 Month

 S. Average S. Low water S. Flood

Fig. 2 : Precipitations for season lagoon in Ivory Cost (www.tutiempo.org)

This study focused on the typical estuarine area opposite Abidjan (Figure 1), due to the increased pollution in
various enclosed bays in this area. Three bays were selected, all of which are in the Ebrié Lagoon in the highly
urbanised area of Abidjan. These are the bay of Yopougon Azito, the bay of Banco and the bay of Marcory,
which are subject to abusive discharge of domestic and industrial effluents. The major criterion that guided our
choice was the high level of urbanisation around the bays.

2.2 Equipment
2.2.1 Equipment for measuring the physical and chemical parameters of the water
Measurements were made using equipment consisting of a portable multimeter (HACH, type HQ40d) for
measuring pH and water temperature, a WTW type conductivity meter for measuring the conductivity of water
samples, and a GARMIN type GPS (Map 78) for locating the measurement stations.

2.2.2. Equipment for measuring the concentration of parabens in the samples
The water bath, capable of maintaining a temperature of 60 °C±5 °C, was useful for maintaining the temperature
of the samples under favourable conditions. The 50 ml glass vessel with cap was used for the measurement of
the water samples and the UV-visible spectrophotometer for the analysis and reading of the concentrations of the
different samples and an automatic titrator for the measurement of BzP.

2.2.3. Reagents
To determine the concentrations of the preservatives, reagents were used. These were absolute ethanol, methyl 4-
hydroxybenzoate (methyl parahydroxybenzoate), ethyl 4-hydroxybenzoate (ethyl parahydroxybenzoate), n-
propyl 4-hydroxybenzoate (propyl parahydroxybenzoate), n-butyl 4-hydroxybenzoate (butyl
parahydroxybenzoate) methanol, etc. Other solutions such as sulphuric acid solution (H2SO4) = 2 mol/L, an

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ethanol/water mixture and a mixture of 9 volumes of ethanol and 1 volume of water were also used in the
preservative extraction phase.

2.2.4. Methodology
2.2.4.1. Collection of water samples
In six (6) campaigns, two (2) per season, water samples were taken from the bays. Thus, during the rainy season
(June-July 2019), the flood season (October-December 2019) and the low water season (January-February 2020),
water samples were taken at 0.5 m from the surface using a Niskin bottle in 500 mL PIREX glass bottles for
analysis in the laboratory. Immediately after the Niskin bottle was lifted aboard the boat, in situ measurements of
some physico-chemical parameters were made.

2.2.4.2. Principle of paraben analysis
The sample is acidified with sulphuric acid and suspended in a mixture of ethanol and water. After gentle heating
to facilitate quantitative extraction, the mixture is filtered. The preservatives present in the filtrate were
determined by UV-visible spectrophotometry except for BzP of the five parabens.

2.2.4.3. Preparation of the stock solution
This phase consists of mixing 36 volumes of water and 64 volumes of methanol. To do this, 0.025 g of methyl
parahydroxybenzoate, 0.025 g of ethyl parahydroxybenzoate, 0.025 g of propyl parahydroxybenzoate and 0.025
g of butyl parahydroxybenzoate were weighed out and placed in 4 volumetric flasks of 100 mL each. Each of the
powders was dissolved and made up with the ethanol/water mixture. The solutions remain stable for one week in
the refrigerator. From this solution, 5 mL of each preservative standard (parabens) is put into four 25 mL
volumetric flasks. Using a graduated pipette, add 1 mL of sulphuric acid solution to each vial, make up to
volume with the ethanol/water mixture and then take a spectrophotometer reading.

2.2.5. How it works
2.2.5.1. Preparation of extracts
The extracts are made by accurately measuring approximately 0.025 g of sample into a 50 mL glass tube with
stopper. Using a graduated pipette, 5 mL of the sulphuric acid solution and 50 mL of the ethanol/water mixture
are introduced into the tube (close the flask and shake vigorously until a homogeneous suspension is obtained (at
least one minute). Place the tube for 5 minutes in a water bath at 60 °C ± 5 °C to facilitate extraction of the
preservatives in the ethanolic phase. Immediately cool the tube in a stream of cold air and store the extract in the
refrigerator for one hour. This extract is filtered on filter paper (porosity 0.45 µm), pour the filtrate into a 100
mL volumetric flask by rinsing the walls of the extraction flask with the ethanol/water mixture. The volume is
made up with the ethanol/water mixture, of which about 5 mL is transferred from the solution into a 25 mL
volumetric flask. Finally, 1 mL of sulphuric acid solution is added to each flask using a graduated pipette and the
ethanol/water mixture is made up to the mark with a volumetric flask.
The analysis is carried out by UV-visible spectrophotometer under spectrophotometric conditions.
The extracts can be stored in the refrigerator and analyzed by UV-visible spectrophotometer within 24 hours.

2.2.5.2. Spectrophotometric conditions are as follows:
- solution: water/methanol mixture;
- quartz cell;
- wavelength: MeP: 254 nm, EtP: 277 nm, PrP: 280 nm, and BuP: 272 nm;
- read off each of the reference solutions and the test solution (3 times) from the optical density thus obtained.
The quantities of preservatives determined by this method are expressed as %/g or µg/g of the samples weighed
exactly.
 A ech x C temoin
 ℎ =
 A temoin
 With :
- Cech: Concentration of the sample;
- Ccontrol: Control concentration;

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- Aech: Absorbance of the sample;
- Atemoin: Control absorbance.

2.2.5.3. Determination of benzylparaben by automatic titrator
The determination of BzP by automatic titrator consisted of adding 2 grams of benzyl benzoate to each water
sample. To this mixture 50 mL of 0.5 M alcoholic potassium hydroxide was added. The whole mixture was
boiled under reflux gently for 1 hour. After this step, the resulting solution was titrated hot with 0.5 M
hydrochloric acid in the presence of phenolphthalein. This titration was carried out in the blank.
1mL of 0.5 M alcoholic potassium hydroxide corresponds to 106.1 mg of paraben benzyl (C12H12O2).

3. Results and discussion
3.1 Results
The results presented in Tables 1, 2 and 3 are the average concentrations per season.
The mean values in Table 1 obtained in Azito Bay are significant in the rainy season (SP) with the exception of
BzP. The concentrations of MeP, EtP, PrP and BuP vary from the rainy season to the flood season (SC) through
the low water season (SE) (SP˃SE˃SC). As for the BzP, they vary from the low water season to the rainy season
(SE˃SC˃SP). Statistical analysis reveals significances between seasons for parameters such as EtP (p= 0.0199)
and BuP (p= 0.0474).
In Banco Bay, the average paraben concentrations vary from season to season. As in Azito, the levels obtained
during the wet season are high and then from the flood season to the low water season. The highest values during
the wet season are 0.663±0.139 µg/L MeP, 0.623±0.058 µg/L BzP. Contrary to the variation of the other
parabens, EtP is more abundant in low water (0.348±0.190 µg/L). The variation in values between the low and
high water seasons is low for some parameters such as MeP, BuP and BzP. Highly significant values are noted
between seasons for the parameters MeP (p= 0.0016) and BzP (p= 0.0002).
The evolution of paraben concentrations between seasons in Marcory Bay is variable from one parameter to
another. MeP varies from the low water season to the rainy season. EtP and BuP change from low water to flood
season. PrP concentrations vary from flood season to low water. In Marcory, the highest concentration is
obtained during the rainy period in BzP (0.331±0.115 µg/L) and decreases from the low water season to the
flood season.

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Table 1: Seasonal variation of emerging pollutants in Azito bay (µg/L).

Sampling seasons MeP EtP PrP BuP BzP TCS MeTCS TCC

 Rainfall 0.171±0.077 0.232±0.006 0.667±0.153 0.040±0.017 0.00±0.00 0.00±0.00 0.040±0.00 0.00±0.00

 Flood 0.020±0.009 0.473±0.015 0.472±0.015 0.072±0.019 0.036 ±0.020 0.00±0.00 0.099±0.045 0.00±0.00

 Low water 0.210±0.032 0.489±0.098 0.519±0.082 0.140±0.033 0.054±0.018 0.036 ±0.020 0.081±0.057 0.070±0.029

 Average 0.196±0.026 0.398±0.046 0.553 ±0.058 0.084±0.017 0.030±0.010 0.012 ±0.008 0.073±0.023 0.023 ±0.013

 Probability 0.8241 0.0199 0.4020 0.0474 0.1003 0.1003 0.6151 0.0245

 Significance NS S NS S NS NS NS S

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 In each column, the means followed by the same letter are not significantly different at the 5% level (Newman-Keuls test). VHS: very highly significant, HS: highly significant,
 S: significant, NS: not significant.

 Table 2: Seasonal variation of emerging pollutants in Banco bay (µg/L).

 Sampling MeP EtP PrP BuP BzP TCS MeTCS TCC
 seasons

 Rainfall 0.663±0.139 0.179±0.010 0.276±0.159 0.059±0.029 0.623±0.058 0.036±0.020 0.290±0.010 0.298±0.015

 Flood 0.104±0.039 0.104±0.064 0.273±0.193 0.028±0.007 0.199±0.061 0.127±0.080 0.193±0.026 0.232±0.024

Low water 0.103±0.033 0.348±0.190 0.188±0.087 0.026±0.017 0.181±0.020 0.038±0.020 0.153±0.017 0.137±0.052

 Average 0.290 ±0.091 0.210 ±0.068 0.245±0.080 0.037±0.011 0.334±0.067 0.066±0.028 0.212±0.020 0.222±0.026

Probability 0.0016 0.3561 0.8999 0.4476 0.0002 0.3717 0.0020 0.0276

Significance HS NS NS NS THS NS HS S

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In each column, the means followed by the same letter are not significantly different at the 5% level (Newman-Keuls test). VHS: very highly significant, HS: highly significant,
S: significant, NS: not significant.

Table 2: Seasonal variation of emerging pollutants in Marcory bay (µg/L).

 Sampling seasons MeP EtP PrP BuP BzP MeTCS TCC

 Rainfall 0,043±0,002 0,074±0,036 0,031±0,019 0,012±0,001 0,331±0,115 0,013±0,010 0,00±0,00

 Flood 0.053±0.021 0.056±0.015 0.136±0.078 0.008±0.004 0.036±0.020 0.063±0.017 0.00±0.00

 Low water 0.132±0.076 0.081±0.021 0.039±0.030 0.033±0.027 0.038±0.021 0.045±0.022 0.008±0.006

 Average 0.076±0.026 0.070±0.013 0.068±0.029 0.017±0.009 0.135±0.055 0.079±0.014 0.002±0.002

 Probability 0.3616 0.7861 0.3005 0.5333 0.0212 0.0171 0.2483

 Significance NS NS NS NS S S NS

In each column, the means followed by the same letter are not significantly different at the 5% level (Newman-Keuls test). VHS: very highly significant, HS: highly significant,
S: significant, NS: not significant.

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3.2. Discussion
 Parabens are actually present in Ivorian lagoon estuaries with high concentrations. This presence is due to the
wide use of parabens in pharmaceutical, food, cosmetic and personal care product formulations [8]. Frequent
consumption and use of products preserved with parabens participate in the contamination of the aquatic
environment. For [1], WWTPs are the main source of contamination of the natural environment by these
micropollutants that are detected at the level of continental waters. For studies conducted in Côte d'Ivoire, [13]
estimated that daily domestic wastewater discharges are the major source of parabens from cosmetic and
personal care products (make-up remover, shampoo, toothpaste, etc.) that are applied daily.
This claim is supported by several authors [14], [15] and [16] who have measured the concentration of parabens
in grey water entering WWTPs. According to them, the usual toilet hours of households correspond to an
increase in paraben concentrations. Similarly, paraben concentrations in WWTP influents are higher and more
regular on weekdays than at weekends. Thus, these domestic waters are discharged into the Ivorian aquatic
environment without treatment [17]. Also, from medicines and food, ingested parabens are rapidly metabolised
and eliminated in the urine, mainly in the form of metabolites, which in turn enter domestic wastewater [4] and
the immediate environment. In addition, direct industrial discharges are at the origin of the release of parabens
into the environment. Thus, Ivorian lagoon bays such as Azito, Banco and Marcory are contaminated by
parabens.
This would have resulted in very high concentrations (Table 1, 2 and 3) in Ivorian estuaries. These substances,
which are known to be endocrine disruptors, could pose a threat to the ecosystem of Côte d'Ivoire. As far as the
Abidjan bays are concerned, the entry of wastewater into the Ebrié lagoon could have effects on aquatic species
due to the high concentrations of parabens obtained. According to several authors, acute and chronic effects exist
on certain aquatic species, notably fish and crustaceans, which are the most vulnerable. In order to determine the
acute toxicity of parabens, [18] tested the LD50 of the Medaka fish and the EC50 of the crustacean D. magna. This
study allowed them to obtain the most toxic BzP of the parabens in fish. In crustaceans, BuP and BzP were the
most toxic parabens. In chronic tests, they determined the 21-day and 14-day NOEC of the crustacean D. magna
and the fish O. latipes respectively. The predicted values allowed [18] to state that BzP is more toxic to the more
sensitive fish.
Indeed, some authors have shown the advantages of wastewater treatment plants in the treatment of emerging
pollutants. In a study, [19] explained the efficiency of treatment plants above 90% in general. In particular, [20]
and [21] claimed that this device applied to parabens could reach 99%. This treatment has resulted in
concentrations in five (5) outfalls in France of which the most abundant emerging pollutant is MeP with an
average concentration between 11,672 and 21,483 ng/L [22].

4. Conclusion
Emerging pollutants including parabens are indeed present in the raw water of the Ebrié lagoon bays. The levels
in the different bays are higher in the rainy season. In contrast to this lagoon season, the flood and low water
seasons record lower concentrations. In general, the concentrations measured at the edges of the economic
capital of Côte d'Ivoire are high due to the absence of a wastewater treatment plant at the outlet of all kinds of
wastewater. As a result, wastewater containing parabens is discharged into the bays without any prior treatment.
Thus, these substances, described as endocrine disruptors, are at the origin of several harmful effects on aquatic
species present in the Ivorian aquatic system. This is why it is important to take an interest in monitoring them.

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