Journal of Environmental Chemical Engineering 1 (2013) 604–608

Biosorption of Zn(II) from aqueous solutions by Acinetobacter sp. isolated frompetroleum spilled soil

Reza Tabaraki a,*, Salman Ahmady-Asbchin b, Omran Abdi a

a Department of Chemistry, Ilam University, Ilam, Iranb Department of Biology, Ilam University, Ilam, Iran

A R T I C L E I N F O

Article history:

Received 9 April 2013

Accepted 28 June 2013

Keywords:

Biosorption

Acinetobacter sp.

Heavy metals

Zinc removal

A B S T R A C T

In this study, biosorption of Zn(II) from aqueous solutions by Acinetobacter sp. was investigated in batch

experiments. Effects of pH, bacterial dosage, initial Zn(II) concentration, contact time and temperature

were studied. Optimum biosorption conditions were found to be initial pH of 6, bacterial dosage of 0.5 g/l

and initial Zn(II) ion concentration of 100 mg/l at room temperature and contact time of 90 min. The

maximum uptake capacity of Acinetobacter sp. for Zn(II) ions was found to be 36 mg/g at optimum

conditions. The adjusted R squared for the first-order kinetic model was 0.999. The Langmuir and

Freundlich isotherms were also studied and data were better fitted with the Freundlich isotherm. Finally,

Acinetobacter sp. adsorption capacity was compared with other biosorbents. This biomass could be used

for removal of Zn(II) from wastewater.

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Introduction

The increase of industrial activities has intensified environ-mental pollution problems and the deterioration of severalecosystems with the accumulation of many pollutants such astoxic metals. Heavy metals are persistent environmental con-taminants since they cannot be degraded or destroyed. Heavymetal pollution represents an important problem due to its toxiceffect and accumulation throughout the food chain which leads toserious ecological and health problems. Removal and recovery ofheavy metals are very important with respect to environmentaland economic considerations [1,2]. Environmental contaminationby Zn(II) arises as a result of many industrial activities, such as theelectroplating and metal-finishing industries, metallurgical indus-try, tannery operations, chemical manufacturing, and its use inmatches, explosives, photographic materials, fuels and printingprocesses. Environmental quality standards for Zn according to theEuropean Union are 40 mg/l for estuaries and marine waters and45–500 mg/l for freshwater based on its hardness [3].

Some of the treatment methods involve high operating andmaintenance cost. The high cost of the chemical reagents and theproblems of secondary pollution also make the physico-chemicalmethods limited in application. Therefore, there is a need for somealternative techniques, which are efficient and cost-effective [4,5].

* Corresponding author. Tel.: +98 841 2227022; fax: +98 841 2227022.

E-mail addresses: rezatabaraki@yahoo.com, r.tabaraki@mail.ilam.ac.ir

(R. Tabaraki).

2213-3437/$ – see front matter � 2013 Elsevier Ltd All rights reserved.

http://dx.doi.org/10.1016/j.jece.2013.06.024

Biosorption, bioprecipitation and uptake by purified biopolymersderived from microbial cells provide alternative means of cleaningindustrial effluents. Various biomaterials such as bacteria, fungi,algae, yeasts and agricultural by-products have been examined fortheir biosorptive properties [6–8].

Removal of heavy metals by microorganisms is a complexprocess that depends on the chemistry of the metal ions, cell wallcomposition of microorganisms, cell physiology, and physico-chemical factors such as pH, temperature, contact time, ionicstrength, and metal concentration [3]. Various naturally occurringbacteria exhibit high capacity for binding of metals. Intactmicrobial cells (live or dead) and their products can be effectivebioaccumulators of both soluble and particulate forms of metals[6].

A variety of functional groups located on the bacterial cell wallare known to be included in metal biosorption. These includecarboxyl, amine, hydroxyl, phosphate, and sulfhydryl groups. Themechanism of metal biosorption by bacterial biomass occursthrough complexation, coordination, physical adsorption, chela-tion, ion exchange, inorganic precipitation and/or a combination ofthese processes [3].

The main objective of this work was to study the maximumbiosorption capacity of Acinetobacter sp. for removal of Zn(II) ions.The most of investigated bacteria in biosorption are Gram-positivebacteria but Acinetobacter sp. is a genus of Gram-negative bacteriabelonging to the Gammaproteobacteria. This bacterium has notbeen used for metal biosorption and particularly was isolated frompetroleum spilled soil. They are important soil organisms becausecontribute to the mineralization of aromatic compounds.

0

5

10

15

20

25

30

35

0 2 4 6 8

q(m

g/g)

pH

Fig. 1. Effect of pH on Zn(II) biosorption on Acinetobacter sp. (T = 25 8C, bacterial

dosage = 1 g/l, Ci = 100 mg/l, contact time = 120 min).

0

5

10

15

20

25

30

35

0 0. 5 1 1. 5 2 2. 5 3

q(m

g/l)

M(g/l)

Fig. 2. Effect of bacterial dosage on Zn(II) biosorption on Acinetobacter sp. (pH = 6, T:

25 8C, Ci = 100 mg/l, contact time = 120 min).

R. Tabaraki et al. / Journal of Environmental Chemical Engineering 1 (2013) 604–608 605

Experimental factors in biosorption were optimized and isothermsand kinetics parameters were determined from biosorptionmeasurements.

Materials and methods

Microorganism and its preparation for biosorption

The microorganism used for this experiment was Acinetobacter

sp. isolated from the soil contaminated with petroleum at the NaftShahr of Kermanshah, Iran. The isolate was purified and identifiedaccording to Bergey’s manual [9]. The bacterial strains werecultured in nutrient broth (pH 7.0) and incubated for 48 h at 30 8C,then harvested by centrifugation for 15 min at 5000 rpm. The cellpellet was washed and rinsed several times with deionized water.The bacterial biomass was used in further experiments for Zn(II)removal from aqueous solutions.

Stock solution of Zn(II)

Stock solution (1000 mg/l) was prepared by dissolving theZn(NO3)2�4H2O salt (Merck) in deionized water. Working solutionswere prepared by diluting the stock solutions to the desiredconcentrations in deionized water. All chemicals used wereanalytical grade.

Biosorption experiments

All biosorption experiments were performed by the batchtechnique and 0.01 M NaNO3 was added as the backgroundelectrolyte to adjust ionic strength. The experiments wereconducted in 250 ml flasks. First, biomass was added to Zn(II)solution and then pH was adjusted to required value using 0.1 MHNO3 and 0.1 M NaOH by a pH-meter. Investigated pH values were2, 3, 4, 5 and 6. In each run, pH was constant. The pH measurementsof test solutions after contact time were shown that pH wasconstant with �0.1 pH unit for each run. Effect of initial Zn(II)concentration from 25 to 150 mg/l and the effect of bacterial dosagefrom 0.5 to 2.5 g/l were studied. The Flasks with mixtures weretransferred on the shaker with 100 rpm for 90 min. The biosorbentwas filtered through a Whatman paper filter and then centrifuged for10 min at 4000 rpm. The final concentration of Zn(II) was determinedby atomic absorption spectrophotometer (Chem Tech Analyticalmodel CTA2000).

Amounts of Zn(II) adsorbed by the biomass were calculatedusing the following equation:

q ¼ VðCi � CeÞM

(1)

where q is the amount of Zn(II) biosorbed by biomass (mg/g); Ci isthe initial concentration of Zn(II) (mg/l); Ce is the concentration ofZn(II) (mg/l) at equilibrium; V is the volume of the metal solution(l); and M is the mass of adsorbent (g) [10,11]. All of theexperiments were carried out in doublets and the experimentalresults were expressed as mean. Statistical analysis was performedby using the Minitab 15.1 (Minitab Inc., State College, PA, USA)software and fitted to a linear regression models. An analysis ofvariance (ANOVA) was then carried out in order to test the modelsignificance and suitability.

Results and discussion

Effect of pH

The effect of pH on the biosorptive capacity of Zn(II) byAcinetobacter sp. was shown in Fig. 1. The biosorptive capacity of

Zn(II) by bacteria was very low at low pH values and increased withpH until reaching an optimum at pH 6.0. However, at pH higherthan 6.0, the Zn(II) begins to precipitate due to formation ofZn(OH)2. At low pH values, cell wall ligands are closely associatedwith hydronium ions and restrict the bisorption of Zn2+. As the pHincreases, more ligands such as carboxyl, phosphate, imidazole,and amino groups carry negative charges and attract Zn(II) ontothe cell surface [3].

Effect of bacterial dosage

The effect of initial bacterial dosage was investigated in therange of 0.5–2.5 g/l (Fig. 2). The optimum bacterial dosage was0.5 g/l. The dosage of a biosorbent strongly influences the extent ofbiosorption. In many instances, lower biosorbent dosages yieldhigher uptakes. An increase in the biomass concentration generallyincreases the amount of solute biosorbed, due to the increasedsurface area of the biosorbent, which in turn increases the numberof binding sites. Conversely, the quantity of biosorbed solute perunit weight of biosorbent decreases with increasing biosorbentdosage, which may be due to the complex interaction of severalfactors [12].

Effect of initial Zn(II) concentration

The effect of initial Zn(II) concentration was investigated byvarying initial Zn(II) concentration, ranging from 25 to 150 mg/l atsame pH and room temperature (Fig. 3). The biosorption capacity(q) increased with the increasing Zn(II) concentration at the samepH and temperature. The optimum metal concentration wasobserved 100 mg/l. The higher solute uptake was obtained athigher initial metal concentration. At lower initial soluteconcentrations, the ratio of the initial moles of solute to theavailable surface area is low; subsequently, the biosorption

0

5

10

15

20

25

30

35

0 25 50 75 100 125 150 175

q(m

g/g)

Ci(mg/l)

Fig. 3. Effect of initial concentration of metal on Zn(II) biosorption on Acinetobacter

sp. (pH = 6, T: 25 8C, bacterial dosage = 0.5 g/l, contact time = 120 min).

0

5

10

15

20

25

30

35

40

0 20 40 60 80 100 120 140

q(m

g/g)

t(min)

Fig. 4. Effect of contact time on Zn(II) biosorption on Acinetobacter sp. (pH = 6, T:

25 8C, bacterial dosage = 0.5 g/l, Ci = 100 mg/l).

y = -0.059 7x + 3.7988R² = 0.9994

1

2

3

4

ln(q

e-qt

)

R. Tabaraki et al. / Journal of Environmental Chemical Engineering 1 (2013) 604–608606

becomes independent of the initial concentration. However, athigher concentrations, the sites available for sorption becomefewer compared to the moles of solute present and hence, theremoval of solute is strongly dependent upon the initial soluteconcentration. It is always necessary to identify the maximumsaturation potential of a biosorbent for which experiments shouldbe conducted at the highest possible initial solute concentration[12].

Effect of temperature and contact time

Temperature affected biosorption only to a lesser extent withinthe range from 20 to 35 8C (Table 1). Higher temperatures usuallyenhance sorption due to the increased surface activity and kineticenergy of the solute. However, physical damage to the biosorbentcan be expected at higher temperatures. It is always desirable toconduct/evaluate biosorption at room temperature because thiscondition is easy to replicate [12]. As shown in Table 1, thebiosorption capacity (q) increased with the increasing tempera-ture.

One of the most important factors influencing the biosorptionequilibrium is the contact time. The variation of the biosorbedZn(II) amount with contact time was shown in Fig. 4. The dataobtained from the biosorption showed that a contact time of90 min was sufficient to achieve equilibrium and the biosorbedZn(II) amounts did not change with further increase in contacttime. Therefore, the uptake amount and lead (II) ion concentrationat the end of 90 min were given as the equilibrium values.

Kinetic experiments

Two models were used to determine the data to examine themechanism of biosorption process. The pseudo-first-order kineticmodel assumes that metal ion binds only to one sorption site onthe sorbent surface [13]. In Lagergren model [14], the rate ofoccupation of biosorption sites is proportional to the number ofunoccupied sites:

logqe

qe � qt

� �¼ k1

2:303t (4)

Table 1Effect of temperature (pH: 6, t: 90 min, bacterial dosage = 0.5 g/l, Ci = 100 mg/l).

T (8C) q (mg/g)

20 26

30 30

40 38

where qe, qt and k1 are the amounts of adsorbed Zn(II) ions atequilibrium time, the amounts of adsorbed Zn(II) ions at time t

(mg/g) and the equilibrium rate constant of pseudo-first-orderadsorption (min�1), respectively. The slopes and intercepts of plotof log(qe � qt) versus t were used to obtain k1 and qe. Theadsorption kinetics may also be described by pseudo-second-orderkinetic model. Metal ions are bound to two binding sites on thesorbent surface [13]:

t

qt

¼ 1

k2q2e

þ 1

qe

t (5)

where k2 is the equilibrium rate constant of pseudo-second-orderadsorption [14]. The slope and intercept of plot t/qt versus t wereused to calculate k2 and qe. The straight lines obtained from plot oft/qt versus t showed good fitness of experimental data with thesecond-order kinetic model (Fig. 5). As shown in Table 2, adjusted R

squared for the pseudo-first-order and pseudo-second-ordermodels were found to be 0.999 and 0.964, respectively. Theamounts of adsorbed Zn(II) on the biosorbent at equilibrium (qecal)by first and second order models were 44.70 and 44.84 mg/g,respectively. The predicted residual sums of squares (PRESS)statistic is a form of cross-validation used in regression analysis toprovide a summary measure of the fit of a model to a sample ofobservations that were not themselves used to estimate the model.It is calculated as the sums of squares of the prediction residuals forthose observations. The PRESS statistic can be calculated for anumber of candidate model structures for the same dataset, withthe lowest values of PRESS indicating the best structures. PRESSvalue, squared correlation coefficient of prediction (R2

pred) and F-value were (0.036, 0.998 and 4762.5) and (0.241, 0.889 and 108.93)

-2

-1

00 20 40 60 80 100

�me (min)

Fig. 5. Plot of the pseudo-first-order equation for the biosorption kinetics of Zn(II)

on Acinetobacter sp. (pH = 6, T = 25 8C, bacterial dosage = 0.5 g/l, Ci = 100 mg/l).

Table 2Kinetic models for biosorption of Zn(II).

Parameter Pseudo-first-order model Pseudo-second-order model

qe (mg/g) 44.70 44.84

k 0.060 7.9 � 10�4

R2 0.999 0.973

Table 3Equilibrium models for biosorption of Zn(II).

Freundlich Langmuir

KF n R2 qmax (mg/g) b (mg/g) R2

3.12 1.93 0.977 52.63 0.018 0.974

5

10

15

20

25

30

35

40

q e(m

g/g)

R. Tabaraki et al. / Journal of Environmental Chemical Engineering 1 (2013) 604–608 607

for the pseudo-first-order and pseudo-second-order models,respectively. Hence, it was concluded that this sorption systemwas better described by first-order rate equation.

Another technique used for identifying the mechanism of theadsorption process is fitting the experimental data in anintraparticle diffusion plot. Weber and Morris diffusion modelcan be used to assess this opinion:

q ¼ fDt

r2p

!1=2

¼ kit1=2 (6)

where rp is particle radius, D is the effective diffusivity of soluteswithin the particle, qt (mg/g) is the adsorbed metal ion amount atany time and ki intraparticle rate constant (mg/g min1/2). The slopeof plot q versus t1/2, (ki), was 2.806 (mg/g min1/2) at optimumconditions. The amounts of biosorbed Zn(II) had a multi-linearitywith three steps (Fig. 6). The first and sharper portion is theexternal surface adsorption or instantaneous adsorption stage. Thesecond portion is the gradual adsorption stage, where theintraparticle diffusion is rate-controlled. The third portion is finalequilibrium stage where the intraparticle diffusion starts to slowdown due to extremely low solute concentrations in the solution[15].

Determination of equilibrium models

Langmuir and Freundlich isotherms were used to describe theequilibrium state for metal ion adsorption. The Freundlichisotherm is a nonlinear sorption model. This model proposes amonolayer sorption with a heterogeneous energetic distribution ofactive sites, accompanied by interactions between adsorbedmolecules [14,16]. The logarithmic form of this model is:

log qe ¼ log KF þ 1=n log Ce (7)

where KF (mg/g) and n are the Freundlich constants.The Langmuir model represents one of the first theoretical

treatments of nonlinear sorption and suggests that uptake occurson a homogeneous surface by monolayer sorption withoutinteraction between adsorbed molecules. In addition, the modelassumes uniform energies of adsorption onto the surface and notransmigration of the adsorbate [17]. The general Langmuir

Fig. 6. Plot of the intraparticle diffusion (Weber–Morris model) for the biosorption

kinetics of Zn(II) on Acinetobacter sp. (pH = 6, T = 25 8C, bacterial dosage = 0.5 g/l,

Ci = 100 mg/l).

equation is commonly presented as the equation may be linearizedas follow:

Ce

qe

¼ 1

qmaxbþ Ce

qmax(8)

where qe is the amount of metal ion removed (mg/g), Ce is theequilibrium concentration (mg/l), b is the Langmuir constantrelated to affinity, and qmax is the maximum metal uptake underthe given conditions.

The Freundlich and Langmuir constants were calculated fromthe corresponding plots. The Freundlich constants (KF and n) areshown in Table 3. The n value greater than 1.0 represents favorablebiosorption conditions [18]. The maximum metal uptake (qmax)under the optimal conditions and adsorption binding constant (b)in Langmuir model are shown in Table 3. Adjusted R squared,PRESS value, squared correlation coefficient of prediction (R2

pred)and F-value were (0.972, 0.034, 0.955 and 172.06) and (0.968,0.227, 0.934 and 150.90) for the Freundlich and Langmuiradsorption isotherm, respectively. The values for two modelswere same but predictability of Freudlich model was slightlybetter. The Freundlich type adsorption isotherm is an indication ofsurface heterogeneity of the adsorbent while Langmuir modelhints surface homogeneity of the adsorbent. The experimental datafor the biosorption of Zn(II) by Acinetobacter sp. was given in Fig. 7.

Comparison with other bacteria

Table 4 compares the maximum adsorption capacities obtainedin this study with some other results reported in the literature. The

00 50 10 0 15 0

Ce(mg/l)

Fig. 7. Isotherm of Zn(II) biosorption on Acinetobacter sp. (pH = 6, T = 25 8C, bacterial

dosage = 0.5 g/l, contact time = 90 min).

Table 4Comparison of maximum biosorption capacity of Zn(II) on different bacteria.

Bacteria pH T (8C) qmax (mg/g) Reference

Pseudomonas aeruginosa 6 30 66.6 [2]

Bacillus cereus 6 30 83.3 [2]

Geobacillus toebii sub.sp. decanicus 5 80 21.1 [1]

Geobacillus thermoleovorans 4 70 29 [1]

Streptomyces rimosus 7.5 20 30 [19]

Delftia tsuruhatensis 6 25 14 [20]

Streptomyces rimosus 5 30 30 [19]

Acenitobacter sp. 6 25 36 This study

R. Tabaraki et al. / Journal of Environmental Chemical Engineering 1 (2013) 604–608608

value of Zn(II) uptake by Acinetobacter sp. found in this work iscomparable with other bacteria.

Conclusion

Biosorption properties of Acinetobacter sp. were studied as afunction of pH, initial Zn(II) concentration, bacterial dosage,contact time and temperature. The biosorption capacity of Zn(II)increased with the increasing temperature and decreasing with theincreasing bacterial dosage at the same initial concentration ofZn(II). The optimum biosorption conditions were determined asinitial pH 6, temperature 25 8C, biosorbent concentration 0.5 g/land initial Zn(II) concentration 100 mg/l. It was found that theZn(II) biosorption attained to equilibrium after 90 min and thiscontact time was taken as the equilibrium. The maximum uptakecapacity of Acinetobacter sp. for Zn(II) ions was found to be 36 mg/gat optimum conditions. The data were better fitted with theFreundlich isotherm model. The first-order kinetic model wasbetter described the system.

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