Anal. Method Environ. Chem. J. 4 (1) (2021) 36-45  
Research Article, Issue 1  
Analytical Methods in Environmental Chemistry Journal  
Journal home page: www.amecj.com/ir  
AMECJ  
Speciation and removal of selenium (IV, VI) from water  
and wastewaters based on dried activated sludge before  
determination by flame atomic absorption spectrometry  
Mahdiyeh Ghazizadeha,, Abdollah Abbaslooa and Farzaneh Bivarb  
a Department of Chemistry, Kerman Branch, Islamic Azad University, Kerman, Iran, P. O. Box 167-7635131  
b Department of Chemical engineering, Sirjan Branch, Islamic Azad University, Kerman, Iran, P.O. Box 187-78185  
A R T I C L E I N F O :  
Received 4 Dec 2020  
A B S T R A C T  
In recent decades, large amount of pollutants enters to the environment  
due to development of technology. Therefore, it is necessary to use  
ecofriendly sorbent to eliminate pollutants. In this research, 0.5 g of  
a dried activated sludge (DAS) was used for speciation selenium and  
removal of selenite [Se(IV)] from water and wastewater samples.  
The effect of operating parameters such as solution pH, the amount  
of bio-sorbent, contact time, temperature and initial concentration  
of selenium were studied by flame atomic absorption spectrometry  
(F-AAS). Kinetic data was adjusted to the Langmuir and Freundlich  
kinetic equations. The resulted showed that the Langmuir equation  
with a correlation coefficient of 0.9825 has the best match to  
tetravalent selenium biosorption on DAS. The FT-IR results showed  
that the biosorption mechanism of Se(IV) on DAS is due to functional  
groups on the DAS surface (Se(IV)…. DAS). For reduction of soluble  
selenate [Se(VI), SeO42−] to selenite [Se(IV), SeO32−], the concentrated  
HCl was used at 70oC (30 min). So, the Se(VI) reduced to Se(IV)  
and total selenium (TSe) was determined and the Se (VI) was simply  
calculated by difference of TSe from Se(IV) content. The method was  
validated based on spiking samples in water and wastewater samples  
by F-AAS and using HG-AAS.  
Revised form 9 Feb 2021  
Accepted 30 Feb 2021  
Available online 30 Mar 2021  
------------------------  
Keywords:  
Selenium,  
Water and wastewater,  
Speciation,  
Activated sludge,  
Biosorption,  
Isotherms.  
glass manufacturing, the agriculture and mining  
1. Introduction  
activities increase the selenium concentration in  
the environment matrixes [6-8]. Selenium is also  
used in thermal power stations, the solar panels,  
insecticides, semiconductors and rectifires [9]. Two  
species of this element exist in aqueous systems  
contain Se(IV) and Se(VI) in the form of selenite  
(SeO32-) and selenite (SeO42-), respectively. Se(IV)  
is more toxic than Se(VI) [2, 10]. World Health  
Organization (WHO) proposed the permissible  
limit of selenium concentration in drinking water  
should be below 10 μg L-1 [11-14]. Therefore,  
removal of selenium from wastewaters by an  
Recently, the selenium studies are considered  
strongly because of the direct correlation between  
biologicalfunctionsandtheamountofseleniuminter  
the body [1, 2]. Selenium is an essential bioelement  
and has an important role in the proper biological  
functioning of many organisms [3, 4], although it  
becomes toxic when the concentration is more than  
1.7 μg L-1 [5]. Modern industrial processes such as  
the oil refining, the electrolytic copper refining, the  
*Corresponding Author: Mahdiyeh Ghazizadeh  
Email: ghazizadeh1385@gmail.com  
https://doi.org/10.24200/amecj.v4.i01.119  
Removal of selenium by dried activated sludge  
Mahdiyeh Ghazizadeh et al  
37  
2.2. Apparatus  
economic and effective methods is necessary. The  
most appropriate methods for removing selenium  
from contaminated water include catalytic  
reduction, chemical precipitation, electrochemical  
process, evaporation, floatation, ion exchange,  
membrane processes, biosorption and adsorption  
[2, 15]. Most of these techniques are expensive  
and improper for removal of selenium from  
aqueous samples. However, biosorption can be an  
effective and ecofriendly method for this purpose.  
Low cost and availability are two major factors  
for using biomass to remove the environmental  
pollutant [15, 16]. Biosorption of selenium by  
several sorbents such as seaweed, crustacean shell,  
peanut shell, rice barn, maize, wheat and dry yeast  
biomass is reported [17-20]. Recently the usage  
of DAS for removing selenium is extended [21,  
22]. In addition, the different instrumental analysis  
was used for determination of selenium and other  
metals in different matrixes [23-27]. In this study,  
the removal of selenium by DAS from aqueous  
solutions were studied by F-AAS and validated  
by HG-AAS. The effect of pH, concentration,  
temperature and contact time was investigated.  
Kinetic models and thermodynamic parameters  
were determined.  
Flame atomic absorption spectrometer (F-AAS,  
Varian spectra 220 model, Australia) with  
wavelength 196.0 nm; slit 1.0 nm; current 10 mA  
was used (10-200 mg L-1). The hydride generation  
atomic absorption spectrometer (HG-AAS, 1-100  
μg L-1) and electro thermal atomic absorption  
spectrometer (ET-AAS, 15- 400 μg L-1) were  
applied as ultra-trace analysis for Se (IV). The  
analytical pH meter (Benchtop meter inoLab pH  
7110 model, WTW company, Germany), analytical  
balance (ALC model, Acculab company, America),  
magnetic hitter stirrer (IKA RH basic 2 model,  
IKA company, Germany), and centrifuge (EBA 20  
model, Hettich company, Germany) were used for  
this study.  
2.3. Preparation of dried activated sludge as a  
biosorbent  
The activated sludge obtained from Zamzam  
company was suspended in a beaker containing 500  
ml of deionized water on a magnetic stirrer for a  
day at 25oC. Let the suspension to precipitate. Then  
the upper liquid was decanted and the remaining  
suspension was centrifuged. The resulted sludge  
was washed with deionized water several times  
to be neutralized. The collected sample was dried  
in oven at 80oC for 36 h. The dried biomass was  
powdered and sieved with mesh No. 25.  
2. Experimental  
2.1. Chemicals  
Sodium selenite (Na2SeO3) with a purity of 98%,  
was purchased from Merck (India) and used as a  
source of Se(IV) ions in the aqueous samples for  
analytical purpose , sodium hydroxide (NaOH)  
with a purity of 98% was purchased from Merck  
(Darmstadt, Germany, http://www.merck.com),  
hydrochloric acid (HCl) with a purity of 37% was  
purchased from Merck (Darmstadt, Germany,  
http://www.merck.com), dried activated sludge was  
obtained from Kerman Zamzam refinery, Iran. The  
different concentration of Selenium was prepared  
by dilution of deionized water (DW) and ultrapure  
water was purchased from Millipore Company. The  
acetate and phosphate buffer was used to adjust the  
pH between 2.6–6.4 and 6.4–8.0, respectively.  
2.4. Preparation of sample and selenium  
solutions  
All glass or PCV tubes were cleaned with a  
2M of HNO3 solution for at least one day and  
then washed for ten times with ultrapure water.  
As low concentrations of Se(IV) and Se(VI) in  
water samples, the ion contamination effected on  
results of analysis, so, we used ultra-trace reagents  
for sampling processes. Sodium selenite was  
used to prepare a selenium stock solution with  
concentration of l000 ppm (mg L-1). The desired  
solutions obtained of diluting stock solution. The  
diluted solutions with concentrations in the range  
2-9.5 ppm were used for calibration.  
Anal. Method Environ. Chem. J. 4 (1) (2021) 36-45  
38  
2.5. SPE procedure and Batch experiments  
from Se(IV) content. The linear range(LR), LOD,  
perconcentration factor (PF) and recovery were  
obtained 0.5-10.2 mg L-1, 0.12 mg L-1 and 19.8, and  
96.5%, respectively  
Biosorption of Se(IV) by DAS was achieved in  
optimized experimental conditions such as pH,  
contact time, amount of biosorbent and temperature.  
The experiments were carried out in 100 ml  
Erlenmeyer flasks. Experiments were achieved with  
pH 2 to 9, contact time 2 to 35 minute, amount of  
biosorbent 0.5 to 3 g, temperature 10 to 40oC and  
selenium concentration 10 to 140 mg L-1. To adjust  
required pH of aqueous solution, HCl 0.2 M and  
NaOH 0.1 M were added. Finally, the kinetic models  
and isotherms were studied. The absorption capacity  
of DAS for Se(IV) was obtained 124.2 mg g-1 by 140  
mg L-1 selenium concentration and 1 g of DAS.  
By solid phase extraction procedure (SPE), 0.5 g  
of biosorbent of DAS added to 100 mL of water  
and wastewater solution and shaked for 15 min at  
pH=5. After adsorption, based on chemical bonding  
between DAS with Se(IV) [−NH+:−NH2+----SeO32−]  
the solid phase separated/collected in bottom of  
tube and removed upper liquid phase of water/  
wastewater. Finally, the Se(IV) determined with  
F-AAS after desorption Se(IV) from DAS by adding  
of HNO3 (0.5 M, 5 mL). The concentration Se(IV)  
validated by HG-AAS after dilution with DW. For  
reduction of Se(VI) to Se(IV) the concentrated HCl  
(50%) was used at 70oC for 30 min. After reduction,  
the total selenium (TSe) was determined and the Se  
(VI) was simply calculated by difference of TSe  
3. Results and discussion  
3.1. FT-IR analysis  
Fourier Transform Infrared (FT-IR) spectrum of  
DAS was recorded (Fig. 1) to gain the information  
about surface functional group. As seen in this  
spectrum, the stretching vibrations of hydroxyl  
group (−OH) on DAS surface gives the broad and  
strong band at 3443 cm-1. The weak peaks at about  
2300 cm-1 show the stretching vibrations of –NH,  
−NH+, −NH2+ functional groups of DAS. The band  
peak at 1646 cm-1 refers to stretching vibrations of  
−C═O group. The stretching vibrations of −C−O  
group appears at 1088 cm-1. The band peak at 876  
cm-1 is concerned to carbonate group.  
3.2. Effect of pH  
The effect of pH on the biosorption of Se (IV)  
by DAS were studied at pH in the range of 2 to  
11 for SPE. First, 100 mL selenium solution with  
concentration 2-9.5 ppm (mg L-1) and 0.5 g DAS  
at temperature of 25oC (15 min) was used. The  
results were shown in Figure 2. Three species  
of selenium in these aqueous solutions include  
selenite (SeO32-), biselenite (HSeO3-) and selenious  
Fig.1. Fourier Transform Infrared (FT-IR) spectrum of DAS  
Removal of selenium by dried activated sludge  
Mahdiyeh Ghazizadeh et al  
39  
3.3. Effect of contact time  
acid (H2SeO3) [28, 29]. The selenious acid prevails  
when pH decreases below 3.5, biselenite prevails  
when pH is in the range of 3.5 to 9 [2]. The lowest  
selenium biosorption at pH less than 3.5 is because  
of inability of neutral selenious acid to interact  
electrostatically with the DAS. In this work, the  
highest chemical biosorption of Se(IV) based on  
DAS was achieved with high recovery more than  
95% for batch system and SPE procedure at pH=5.  
The effect of contact time, as the next parameter  
was investigated in the range of 2 to 35 minute at  
pH=5. As observed in Figure 3, the most proper  
contact time for selenium biosorption was obtained  
15 minutes for SPE. After this contact time,  
equilibrium occurred. The best time for batch  
system was obtained 30 min (2 g) for selenium  
concentration 10 to 140 mg L-1.  
Fig.2. The effect of pH on Se(IV) removal from water and wastewater by DAS  
Fig.3. The effect of contact time for removal of Se(IV) from water and wastewater by DAS  
Anal. Method Environ. Chem. J. 4 (1) (2021) 36-45  
40  
3.4. Effect of amount of biosorbent  
optimized amount of DAS. Therefore, a number of  
Se(IV) ions remain in solution and biosorption yield  
decreased. At higher amount of DAS, biosorption  
yield is almost unchanged. Because most of Se(IV)  
ions interact with DAS surface. For SPE, the 0.5 g  
of DAS is favorite mass for removal of Se (IV) in  
water samples with high recovery more than 95%.  
The effect of amount of biosorbent was investigated  
under optimized conditions (pH=5 and contact  
time: 30 min.). As shown in Figure 4, the selenium  
biosorption increased slowly with the DAS amount  
up to 2 g for batch system. The DAS surface  
becomes saturated with the extra Se(IV) ions in  
120  
100  
80  
60  
40  
20  
0
0.2  
0.5  
1
1.5  
2
2.5  
3
3.5  
Amount of DAS( g)  
Fig. 4. The effect of biosorbent amount on Se(IV) removal in batch system (green)  
and SPE procedure(blue) in water and wastewater by DAS  
10  
15  
20  
25  
30  
35  
40  
45  
Temperature (oC)  
Fig. 5. The effect of temperature on Se(IV) removal in batch system (blue) and SPE  
procedure(green)from water and wastewater samples by DAS  
Removal of selenium by dried activated sludge  
Mahdiyeh Ghazizadeh et al  
41  
3.5. Effect of temperature  
3.7. Kenetic isotherms for Se(IV) and Se(VI)  
The effect of temperature on selenium biosorption  
was investigated between 10-45oC in optimized  
condition. The results showed us, the optimum  
temperature was achieved 30oC in optimized  
condition (pH, contact time and amount of  
biosorbent DAS were 5, 30 minute and 2 g,  
respectively). Due to Figure 5, by increasing  
temperature, the selenium biosorption decreased. It  
indicated the biosorption by DAS is an exothermic  
reaction. The best temperature for SPE procedure  
for DAS was 25-30oC.  
The most popular isotherms are Langmuir [21,30]  
and Freundlich [21, 31] models. The Langmuir  
model describes monolayer adsorption, however  
Freundlich model show heterogeneous surface.  
The linear form of Langmuir model is given by  
following equation I:  
(Eq. I)  
where Ce (mg L-1) is the equilibrium concentration of the  
solution, qe (mg g-1) is the amount of metal adsorbed per  
specificamountofadsorbent,qm(mgg-1)isthemaximum  
amount of metal ions required to form monolayer, K (L  
mg-1) is the adsorption equilibrium constant.  
The linear form of Freundlich model is given by  
following equation II:  
3.6. Effect of initial concentration of selenium  
Effect of initial concentration of selenium in the  
rangeof20-250ppmwasinvestigatedforabsorption  
capacity. The results indicated that increasing  
selenium concentration caused to more absorb of  
the selenium on DAS and decreased the selenium  
concentration in the solution. The number of sites  
on DAS were interacted with Se(IV) ions and can  
be saturated at high concentrations of selenium  
ions. According to obtained results, the adsorption  
capacity of Se(IV) ions on DAS increased up to  
124.2 mg g-1 [AC; mg per gram]. The results have  
presented in Figure 6.  
(Eq. II)  
where n is the adsorption intensity and KF is the  
adsorption capacity.  
TheamountofSe(VI)adsorbedonDASatequilibrium  
(qe, mg g-1) was calculated by Equation (III):  
qe= (C0-Ce) × V/m  
(Eq. III)  
140  
120  
100  
80  
60  
40  
20  
0
20  
70  
90  
110  
140  
150  
160  
250  
Conc. Se(mg L-1)  
Fig. 6. The effect of initial concentration of Se(IV) ions on absorption capacity  
of DAS in water and wastewater samples  
Anal. Method Environ. Chem. J. 4 (1) (2021) 36-45  
42  
where C0 and Ce (mg L-1) are the initial and  
equilibrium Se(VI) concentrations, respectively, V  
(L) is the volume of the solution and m (g) is the  
mass of the adsorbent. (C0 = 10-250 mg L-1, C0=250  
mg L-1 and Ce=190 mg L-1, V=0.1 L, m=0.05 g).  
So the qe, qmax and Ce/qe was obtained as 120 mg  
g-1, 120 mg g-1 and 2.08, respectively. As different  
concentrations, the Ce/qe were calculated based on  
Langmuir model between 0.02-2.08.  
isotherm (Fig. 7). The Se(VI) in solutions with  
different initial concentrations (C0 = 10-250 mg  
L-1) were used. Langmuir constants, KL and qm  
were calculated from the slope and intercept of the  
plot Ce/qe versus Ce.  
As Figure 8, the linear Freundlich isotherm of Se(IV)  
and Se(VI) is a another kenetic model for DAS.  
Freundlich isotherm parameters, KF and 1/n were  
calculated from the slope and intercept of linear plot.  
Linear Langmuir equation was considered to gain  
2.5  
2
y = 0.0108x  
R2=0.9825  
1.5  
1
0.5  
0
0
50  
100  
Ce(mg L-1)  
150  
200  
Fig. 7. Linear Langmuir equation for selenium removal by DAS biosorbents  
from water and wastewater samples  
0.2  
0
y = 0.4205x - 0.84  
R² = 0.9547  
-0.2  
-0.4  
-0.6  
-0.8  
-1  
Se (VI)  
y = 0.3827x - 0.9139  
R² = 0.9621  
-1.2  
-0.7  
-0.2  
0.3  
0.8  
1.3  
1.8  
logCe  
Fig. 8. Linear Freundlich isotherm for selenium removal by DAS biosorbents  
from water and wastewater samples  
Removal of selenium by dried activated sludge  
Mahdiyeh Ghazizadeh et al  
43  
3.8. Validation of SPE procedure  
a satisfactorily result for determination of Se(IV)  
and Se(VI) in water samples (Table 1). Moreover,  
the real water samples were analyzed with HG-  
AAS/ET-AAS and used for validation of results  
of SPE/F-AAS procedure. The results showed,  
the favorite efficiency and reliability of proposed  
method for determination selenium in water and  
wastewater sample which was compared to ET-  
AAS and HG-AAS (Table 2)  
The selenium was removed and determined in  
100 mL of water and wastewater samples based  
on DAS with SPE procedure at pH=5. The mean  
concentration of Se(IV) more than Se(VI) in water  
samples. The spiked water was used to demonstrate  
the reliability of the method for determination  
of Se(IV) and Se(VI) in water samples by SPE  
procedure. The recovery of spiked samples showed  
Table 1. Validation of SPE procedure based on DAS for speciation selenium (VI, IV) in water  
and wastewater samples (mg L−1; n=8)  
Added  
Se(IV)  
Added  
Se(VI)  
*Found  
Se(IV)  
*Found  
Se(VI)  
Total  
TSe  
Recovery Recovery  
Se(IV) (%) Se(VI) (%)  
Sample  
Wastewater 1  
-----  
4.0  
-----  
0.5  
4.45± 0.19  
1.23± 0.05 5.68 ± 0.24  
-----  
96.0  
-----  
98.8  
-----  
104  
8.29 ± 0.37 1.75 ± 0.07 10..04± 0.45  
3.86± 0.18 0.56 ± 0.03 4.42 ± 0.22  
Wastewater 2  
Wastewater 3  
River  
-----  
4.0  
-----  
0.5  
-----  
94.7  
-----  
98.6  
-----  
103.5  
7.81 ± 0.35  
1.95 ± 0.14  
3.98 ± 0.19  
ND  
1.03 ± 0.05  
1.47± 0.08  
2.95 ± 0.14  
ND  
8.84 ± 0.42  
3.42 ± 0.15  
6.93 ± 0.34  
ND  
-----  
2.0  
-----  
1.5  
-----  
101.5  
-----  
94.9  
-----  
2.0  
-----  
2.0  
1.97 ± 0.11  
2.07 ± 4.4 4.04 ± 0.21  
*x ± ts /√n at 95% confidence (n=8)  
Well water prepared from Varamin agricultural  
Wastewater 1 prepared from drug company  
Wastewater 2 prepared from petrochemical factory  
Wastewater 3 prepared from paint factory  
River water prepared from Karaj  
Table 2. Comparing of proposed procedure for selenium determination by F-AAS/DAS with HG-AAS and ET-AAS  
Sample  
F-AAS/DAS(mg L−1)  
ET-AAS(μg L-1)*  
*HG-AAS(μg L-1)  
Wastewater*  
Water  
0.55 ± 0.25  
ND  
ND  
5.36 ± 0.52  
40.9± 13.81  
41.3 ± 13.81  
x ± ts /√n at 95% confidence (n=5)  
*Wastewater 1 prepared from drug company, 1 mL of sample diluted with DW up to 100 (1:100)  
Anal. Method Environ. Chem. J. 4 (1) (2021) 36-45  
44  
biosorption of selenium (IV) ions onto  
4. Conclusions  
Ganoderma Lucidum Biomass, Sep. Sci.  
Tech., 48 (2013) 2293-2301.  
In this study, the results showed tetravalent  
selenium ions (Se IV) biosorption were successfully  
achieved by DAS from contaminated aqueous  
solutions. The maximum removal of Se(IV) ions  
was 96% at optimized experimental conditions  
by SPE/F-AAS. The interaction between Se(IV)  
ions and functional groups of DAS surface was  
exothermic. The experimental data were fitted to  
Freundlich isotherm. Also the speciation Se(IV)  
and Se (VI) ions determined based on DAS by SPE  
procedure for 0.5 g of DAS at pH=5. The method  
was validated by ET-AAS and HG-AAS. The  
absorption capacities for Se(IV) and Se (VI) ions  
with DAS were achieved 124.2 mg g-1 and 121.8  
mg g-1, respectively.  
[6] H. Robberecht, R.V. Grieken, Selenium  
in environmental water: determination,  
speciation and concentration levels, Talanta,  
29 (1982) 823-844.  
[7] J. Lessa, A. Araujo, G. Silva, L. Guilherme,  
G. Lopes, Adsorption-desorption reactions  
of selenium (VI) in tropical cultivated and  
uncultivated soils under Cerrado biome,  
Chemosphere, 164 (2016) 271-277.  
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Botelho, Selenium contaminated waters: an  
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5. Acknowledgments  
[9] E.I. El-Shafey, Sorption of Cd(II) and Se(IV)  
from aqueous solution using modified rice  
husk, J. Hazard. Matter., 147 (2007) 546- 555.  
[10] F.Sahin, M. Volkan, A.G. Howard, O.Y.  
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The authors would like to thank from Department  
of Chemistry, Kerman Branch, Islamic Azad  
University, Kerman, Iran.  
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