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Colloids and Surfaces B: Biointerfaces 94 (2012) 362– 368

Contents lists available at SciVerse ScienceDirect

Colloids and Surfaces B: Biointerfaces

j our na l ho me p age: www.elsev ier .com/ locate /co lsur fb

sotherm kinetics of Cr(III) removal by non-viable cells of Acinetobacteraemolyticus

iti Khairunnisa Yahyaa, Zainul Akmar Zakariab, Jefri Saminc, A.S. Santhana Rajd, Wan Azlina Ahmada,∗

Department of Chemistry, Faculty of Science, Universiti Teknologi Malaysia, 81310 UTM Johor Bahru, Johor, MalaysiaInstitute of Bioproduct Development, Universiti Teknologi Malaysia, 81310 UTM Johor Bahru, Johor, MalaysiaMaterials Science Laboratory, Faculty of Mechanical Engineering, Universiti Teknologi Malaysia, 81310 UTM Johor Bahru, Johor, MalaysiaElectron Microscopy Unit, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Malaysia

r t i c l e i n f o

rticle history:eceived 28 September 2011eceived in revised form 27 January 2012ccepted 14 February 2012vailable online 22 February 2012

eywords:

a b s t r a c t

The potential use of non-viable biomass of a Gram negative bacterium i.e. Acinetobacter haemolyti-cus to remove Cr(III) species from aqueous environment was investigated. Highest Cr(III) removal of198.80 mg g−1 was obtained at pH 5, biomass dosage of 15 mg cell dry weight, initial Cr(III) of 100 mg L−1

and 30 min of contact time. The Langmuir and Freundlich models fit the experimental data (R2 > 0.95)while the kinetic data was best described using the pseudo second-order kinetic model (R2 > 0.99). Cr(III)was successfully recovered from the bacterial biomass using either 1 M of CH3COOH, HNO3 or H2SO4

iosorptionr(III)cinetobacteremovalastewater

TIR

with 90% recovery. TEM and FTIR suggested the involvement of amine, carboxyl, hydroxyl and phosphategroups during the biosorption of Cr(III) onto the cell surface of A. haemolyticus. A. haemolyticus was alsocapable to remove 79.87 mg g−1 Cr(III) (around 22.75%) from raw leather tanning wastewater. This studydemonstrates the potential of using A. haemolyticus as biosorbent to remove Cr(III) from both syntheticand industrial wastewater.

© 2012 Elsevier B.V. All rights reserved.

EM

. Introduction

Chromium is used in various industrial applications such as tan-eries, electroplating, textile dying, wood preservation, petroleum

ndustry as well as metal finishing. Chromium is present in thenvironment in either one of its two most stable forms i.e. theighly soluble and toxic Cr(VI) or the less mobile and less toxicr(III) [1]. Conventional methods to remove heavy metals are gen-rally effective at metal concentrations greater than 100 mg L−1. As

consequence, these techniques may not be suitable to treat thencreasing volumes of wastewater containing low metal concentra-ions [2]. Therefore, it is imperative to develop a cost effective andnvironmental-friendly treatment process. In recent years, muchf the research has been focused on the development of biolog-cal methods for the treatment of industrial effluents, some of

hich are in the process of commercialization. From these meth-ds, biosorption has been demonstrated to possess good potentialor the removal of heavy metals [3]. Amongst its advantages include

he reusability of biomaterial, low operating cost and improvedelectivity for specific metals of interest [4]. Abundant and inex-ensive materials, such as algae, fungi, bacteria, chitosans and

∗ Corresponding author.E-mail address: azlina@kimia.fs.utm.my (W.A. Ahmad).

927-7765/$ – see front matter © 2012 Elsevier B.V. All rights reserved.oi:10.1016/j.colsurfb.2012.02.016

zeolites have proven to have a reasonably high metal sequester-ing capacity which can be used as biosorbents as reported by otherresearchers [5]. Therefore, it is the objective of this study to evaluatethe potential of using non-viable cells of Acinetobacter haemolyticus(A. haemolyticus) as biosorbent for Cr(III) removal.

2. Materials and methods

2.1. Preparation of biosorbent

A. haemolyticus EF369508 was isolated from the Cr(VI)-containing wastewater from a batek (textile-related) manufactur-ing premise in Kota Bharu, Kelantan, Malaysia [6]. The non-viablebacterial biomass was prepared by harvesting the NB-grown bac-terial cells at early stationery phase via centrifugation at 7000 rpm,4 ◦C for 5 min (B.Braun, SIGMA 4K-15). The supernatant was dis-carded while the pellet was suspended in minimal volume ofdistilled deionized water (DDW) and inactivated by autoclaving at121 ◦C for 15 min.

2.2. Removal of Cr(III) from synthetic solution

The effect of some parameters affecting the biosorption pro-cess such as pH, biomass dosage, initial Cr(III) concentration andcontact time were evaluated as follows: 10 mg dry wt. of the

S.K. Yahya et al. / Colloids and Surfaces B

Table 1Physico-chemical characteristics of raw and treated leather tanning wastewater;four different batches of sampling.

Parameters Unit Raw Treated

pH – 7.8–8.7 6.0–6.3Suspended solid mg L−1 185.5–1067.0 2.0–56.0Colour ADMI 167–8075 17–248Turbidity FAU 53–1100 2–31COD mg L−1 1050–2626 111–482Cr(III) mg L−1 121.5–20888 1.91–6.05Mg mg L−1 16.74–39.40 13.37–39.40K mg L−1 5.46–18.26 10.73–26.39Na mg L−1 439.91–1788.98 889.60–1339.29Zn mg L−1 1.27–4.05 1.05–4.84Pb mg L−1 0.78–0.26 0.19–0.78Cu mg L−1 0.010–0.196 0.251–0.912

b2sebusfwt1vbAd

2

ntstttarvho

2

ctto2bi

2

wfis

Co mg L−1 0.010–0.196 0.005–0.200Mn mg L−1 0.15–5.67 0.15–2.70

acterial biomass suspension was transferred into a series of50 mL Erlenmeyer flasks containing 50 mL of 50 mg L−1 Cr(III)olutions at pH values between 4 and 6. The pH was varied usingither 1.0 M NaOH or 1.0 M HCl. The mixtures were then equili-rated for 24 h at 100 rpm in 25 ◦C. Similar experimental setup wassed for the effect of biomass dosage where 100 mg L−1 of Cr(III)olution was contacted with either 10, 15, 30 or 50 mg cell dry wt.or 30 min. For the isotherm study, 5–200 mg L−1 Cr(III) was mixedith 15 mg cell dry wt. at pH 5 and shaken at 100 rpm for 30 min. In

he kinetic study, 50 and 100 mg L−1 of Cr(III) was contacted with5 mg cell dry wt. followed by periodical sampling. Total workingolume used in this study was 50 mL (mixture of Cr(III) solution andiomass suspension). Cr(III) concentration was determined usingAS. The stock solution for Cr(III), 1000 mg L−1 was prepared byissolving 1.286 g of CrCl3.6H2O in 250 mL of DDW.

.3. Removal of Cr(III) from leather tanning wastewater

Biosorption of Cr(III) from leather tanning wastewater (LTW) byon-viable A. haemolyticus was carried out using optimized condi-ions determined from the synthetic solution study. Two types ofamples i.e. raw and treated LTW, was determined for Cr(VI) andotal Cr concentrations. Since Cr(VI) was not detected in either ofhe samples, total Cr concentration measured would directly relateo the Cr(III) concentration (as total Cr equals to the sum of Cr(VI)nd Cr(III) present), which was determined at 121.15 mg L−1 for theaw LTW and 1.91 mg L−1 for the treated LTW. Based on this obser-ation, raw LTW was chosen for Cr(III) removal study because of theigher Cr(III) concentration. The physico-chemical characteristicsf LTW are listed in Table 1.

.4. Desorption of Cr(III) from non-viable bacterial biomass

The desorption of Cr(III) from Cr(III)-laden bacterial biomass wasarried out using either 0.1 M or 1.0 M H2SO4, HNO3 or CH3COOH ashe desorption agent. Following the biosorption process, the bac-erial cells were first harvested via centrifugation where pelletsbtained were washed using 25 mL DDW followed by contact with5 mL of desorption agents at 200 rpm, 25 ◦C for 2 h. The desorpedacterial biomass (termed as regenerated biomass) was then used

n repeated cycles of adsorption-desorption process.

.5. TEM analysis

TEM analysis of bacterial cell pellets before and after contactingith Cr(III) were carried out as follows; firstly, the cell pellets werexed using 4% (v/v) of glutaraldehyde in 0.1 M phosphate bufferedaline (PBS) for 1 h at room temperature followed by washing using

: Biointerfaces 94 (2012) 362– 368 363

0.1 M PBS and post-fixation with 2% (v/v) osmium tetroxide for 1 h.Then, the cells were dehydrated using increasing concentrations of99.9% (v/v) ethanol (30, 50, 70, 90 and 100%) at 5 min intervals fol-lowed by 2 min washing in 100% (v/v) acetone. The cells were thenembedded in 50 and 100% (v/v) of epoxy resin in acetone for 15 mineach. The cell pellets were again infiltrated with 100% (v/v) epoxyresin and cured at 60 ◦C overnight. Specimens of 90 nm thicknesswere sectioned from the embedded blocks using an ultramicro-tome (Leica-UltraCut UCT) and mounted on 200-mesh copper grids.The specimens were stained with uranyl acetate, post-stained withlead-citrate (5 min each) and viewed using a transmission electronmicroscope (Tecnai G2, Philips).

2.6. Fourier transform infrared (FTIR) analysis

Possible involvement of functional groups from non-viable cellsof A. haemolyticus during the removal of Cr(III) from aqueous solu-tion was elucidated using the Fourier transform infra-red (FTIR)analysis. Overnight dried (60 ◦C) bacterial cell pellets were groundwith KBr (spectroscopic grade) at a ratio of 1:200 in a mortar beforepressed into 10 mm diameter disks under 6 tonnes of pressure. FTIRspectra were obtained on a PerkinElmer FTIR-600 spectrometer.The analysis conditions used were 16 scans at a resolution of 4 cm−1

measured between 400 and 4000 cm−1.

2.7. Data analysis

2.7.1. Adsorption isothermsFour types of adsorption isotherms namely Langmuir, Fre-

undlich, Dubinin–Radushkevich (D–R) and BET were used toelucidate the Cr(III) removal process by non-viable cells of A.haemolyticus. The Langmuir adsorption isotherm model suggeststhe removal of Cr(III) to occur on homogenous surfaces viamonolayer sorption without any interactions between the sorbedanalytes (Eq. (1)).

qe = QmCe

1/b + Ce(1)

where qe represents the equilibrium Cr(III) uptake on the biosor-bent, Qm is the maximum adsorption capacity (mg g−1), Ce is theresidual concentration of Cr(III) at equilibrium (mg L−1) and b isthe Langmuir constant related to the energy of adsorption (L mg−1).The maximum adsorption capacity, Qm and Langmuir constant, bcan be determined from the slope and intercept of linear plot of thelogarithmic equation. The essential features of Langmuir isothermcan be expressed in terms of a dimensionless constant separationfactor, RL (Eq. (2)) where b is the Langmuir constant, C0 is the ini-tial concentration and RL indicates the shape of the isotherm (RL > 1unfavorable, RL = 1 linear; 0 < R < 1 favorable and RL = 0 irreversible)[7].

RL = 11 + bC0

(2)

The Freundlich isotherm assumes a heterogeneous surface witha non-uniform distribution of heat over the surface [8] with generalrelationship as depicted by Eq. (3) where Kc is a constant for relativeadsorptive capacity and n is an affinity constant.

qe = Kf C1/ne (3)

The Dubinin and Radushkevich (D–R) isotherm proposed anequation for the analysis of isotherm to determine whether theadsorption process occurred via a physical or chemical process. The

D–R equation is more general than the Langmuir model becauseit does not assume a homogeneous surface, a constant sorptionpotential nor absence or steric hindrance between adsorbed andincoming particles [8]. The D–R equation is as shown in Equation

364 S.K. Yahya et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 362– 368

Table 2Cr(III) biosorption using various biosorbents at optimum pH.

Type of biosorbent Optimum pH Cr(III) removal, mg g−1 (unless stated otherwise) Reference

Streptomyces VITSVK9 spp. 4.0 76% [15]Sorghum straw (acid washed) 4.0 6.96 [16]Oats straw (acid washed) 4.0 12.97 [16]Agave bagasse (acid washed) 4.0 11.44 [16]Aerobic Granules 5.0 37.80 [17]Nymphae sp. 4.5–5.5 6.11 [18]

4cti(

q

lilCsCramta

q

2

tm(aio

l

otbhh

2

nvbassiitaf

Spirogyra spp. 5.0

Eichornia crassipes 4.5–5.5

A. haemolyticus 5.0

[9] where qeq is the biosorption capacity (mol g−1), ̌ is activityoefficient constant (mol2 j−2) related to biosorption energy, qmax ishe maximum biosorption capacity (mol g−1) to form monolayer, εs polanyi potential, T is absolute temperature (K), R is a gas constant8.314 J mol−1 K−1) and Ceq is the equilibrium concentration.

eq = qmax exp[−ˇ(RT ln(1 + 1/Ceq)

]2(4)

The BET model is an extension of the Langmuir model for multi-ayer biosorption. It is based on the assumption that each adsorbaten the first biosorbed layer serves as a biosorption site for the secondayer and beyond. General BET isotherm is given in Eq. (5) whereBET is the constant relating to the energy of interaction with theurface (L mg−1), Cs – saturation concentration of solute (mg L−1),eq – concentration of solute remaining in the solution at equilib-ium (mg L−1), qeq – amount of solute adsorbed per unit weight ofdsorbent and Q◦

max – maximum specific uptake corresponding toonolayer saturation (mg g−1). Linearization of Eq. (5) would yield

he CBET and Qmax values which can be calculated from the slopend the intercept respectively.

eq = Q ◦maxCBETCeq

(CS − Ceq)[1 + (CBET − 1)(Ceq/CS)](5)

.7.2. Pseudo-first order kinetic analysisThe biosorption process may proceed by diffusion of metal ions

hrough the boundary layer at the biosorbent surface and thisay be the rate determining step of the overall process [10]. Eq.

6) shows the first-order rate expression where qt (mg g−1) is themount of adsorbed metal on the biosorbent at time t, k1,ad (min−1)s the rate constant of first-order adsorption and qeq is the amountf metal sorbed at equilibrium (mg g−1).

og (qeq − qt) = log qeq − K1,ad

2.303t (6)

Linear plots of log (qeq − qt) versus t indicate the applicabilityf this model where the qeq and k1,ad can be determined fromhe intercept and slope of the plot, respectively. If the differencesetween the determined qeq value and the experimental values areigh, the reaction cannot be classified as first-order even thoughigh correlation coefficient was obtained [11].

.7.3. Pseudo-second order kinetic analysisFor adsorption that proceeds via a pseudo second-order mecha-

ism, the rate-limiting step may be chemical sorption involvingalency forces through the sharing or exchange of electronsetween sorbent and sorbate as covalent forces. The followingssumptions were made; adsorption via monolayer adsorption, theorption energy for each ion is similar and does not depend onurface coverage, the sorption occurs only on localized sites, nonteractions between the adsorbed ions and the rate of sorption

s almost negligible in comparison with the initial rate of sorp-ion [12]. The pseudo-second order relationship can be expressedccording to Eq. (7) where k2,ad (g mg−1 min−1) is the rate constantor second-order adsorption. A straight line of t/q versus t would

81% [19]6.61 [15]

62.35 This study

indicate the applicability of second-order kinetics where the qeq

and k2,ad values were determined from the slope and intercept ofthe plot, respectively.

t

q= 1

k2,adq2eq

+ t

qeq(7)

3. Results and discussion

3.1. Effect of pH

The non-viable cells of A. haemolyticus showed the highest Cr(III)uptake at pH 5 (62.35 mg g−1) followed by pH 4 (37.38 mg g−1) andpH 6 (35.15 mg g−1). High Cr(III) uptake at pH 5 can be explainedas follows; as the pH increases from 4 to 6, surface of the bacte-ria will be deprotonated, hence exposing more negatively chargedligands which have the ability to bind the cationic Cr(III) speciessuch as Cr(OH)2

+ and Cr(OH)2+ from solution [13]. This study wascarried out between pH 4 and 6 range as it was anticipated thatat pH < 3, predominant species in solution would be the free Cr(III)ions [14]. In this condition, the free Cr(III) ions would compete withH+ ions for the adsorption sites as well as the electrostatic repulsioncondition between the protonated bacterial surface and the Cr(III)ions [13]. At pH > 6, the formation of hydroxylated chromium com-plex (Cr(OH)3) may reduce the availability of Cr(III) species to formcomplex with the negatively charged groups present on the bac-terial cells surface. Table 2 shows the comparison between Cr(III)removal using A. haemolyticus and other biosorbents at optimumpH values.

3.2. Effect of adsorbent dosage

Cr(III) adsorption increased with increasing amounts of adsor-bent dosage (Fig. 1). This condition can be attributed to the increasein adsorption area and the availability of free adsorption sites.However, further increment in the biosorbent dose would leadto the formation of aggregates that would reduce the availabil-ity of effective adsorption area, hence decreasing the overall metalremoval capacity. The adsorption sites remain unsaturated duringthe biosorption process due to a lower adsorptive capacity uti-lization of biosorbent, which decreases the biosorption efficiency.Highest Cr(III) uptake of 188.58 mg g−1 was obtained at a dosageof 10 mg cells dry wt. while the highest Cr(III) removal of 56.44 at%15 mg cells dry wt.

3.3. Effect of contact time and initial Cr(III) concentration

The non-viable cells of A. haemolyticus showed rapid Cr(III)uptake within the first 0.5–1 h with maximum uptake of51.08 mg g−1 (initial Cr(III) of 50 mg L−1) and 198.80 mg g−1 (initial

Cr(III) of 100 mg L−1). This could be due to the availability of largenumber of vacant adsorption sites on the bacterial cells surface. Thiswas followed by a gradual decrease in Cr(III) uptake until an adsorp-tion equilibrium condition was achieved. It is also worthy to note

S.K. Yahya et al. / Colloids and Surfaces B

Fig. 1. Effect of biomass dosage on the uptake and removal of Cr(III) by A. haemolyti-cut

tucdoocb1tT

woi

TEA

(

us; SDuptake = 0.48–21.35, SDremoval = 16.53, n = 2; Cr uptake (mg g−1) was calculatedsing the normal mass balance equation; Cr removal (%) was derived from dividinghe residual and initial concentration of Cr(III) after equilibrium.

hat higher initial Cr(III) concentration resulted in a higher Cr(III)ptake (difference of 2–3 times as demonstrated in this study). Thisan be explained from the increased adsorption rate and the higherriving force generated at high Cr(III) concentration [21]. Similarbservation was reported by other researcher where the uptakef Cr(III) by aerobic granule increased with increasing initial con-entration [17]. The effect of initial Cr(III) concentration on Cr(III)iosorption capacity of A. haemolyticus was further evaluated using0–200 mg L−1 of Cr(III). The Cr(III) uptake capacity (Quptake) andhe concentration equilibrium (Ceq) of A. haemolyticus are listed inable 3.

The results showed that the Cr(III) uptake capacity increasedith increasing concentration of Cr(III) with maximum uptake

f 330.63 mg g−1 at initial Cr(III) of 175 mg L−1. However, furtherncrease to 200 mg L−1 of Cr(III) cause a slight decrease in the uptake

able 3quilibrium concentration (Ceq) and Cr(III) uptake capacity (Quptake) of non-viable. haemolyticus; SDCeq = 0.36 − 12.51, SDQuptake

= 1.20 − 41.7 and n = 2; Cr uptake

mg g−1) was calculated using the normal mass balance equation.

Initial Cr(III) (mg L−1) Ceq (mg L−1) Quptake (mg g−1)

10 8.17 ± 0.36 6.58 ± 1.2015 10.88 ± 0.47 9.07 ± 1.5525 15.12 ± 1.89 24.56 ± 6.3250 33.18 ± 1.31 46.58 ± 4.3675 46.45 ± 2.19 96.49 ± 7.30

100 39.70 ± 12.51 177.82 ± 41.71125 38.35 ± 10.50 260.72 ± 35.00150 50.33 ± 3.22 282.39 ± 10.72175 65.63 ± 1.63 330.63 ± 5.42200 109.05 ± 3.22 231.06 ± 10.72

Fig. 2. Linearized form of the Lagergren (a) pseudo first-order and (b) pseudo second

: Biointerfaces 94 (2012) 362– 368 365

capacity due to the saturation of binding sites. The increase ofCr(III) uptake capacity with increasing Cr(III) concentration (up to175 mg L−1) could be due to the stronger driving force to overcomeall mass transfer resistances of Cr(III) ions between the aqueoussolution and the biosorbent, which results in higher probability ofcollision between Cr(III) ions and the biosorbent thus leading to agreater biosorption capacity [22].

3.4. Biosorption isotherm study

The Langmuir and Freundlich adsorption isotherm models fit-ted well with the Cr(III) removal data by non-viable A. haemolyticuswith R2 values of 0.9515 and 0.9742 respectively. For the Lang-muir isotherm, maximum uptake capacity, Qe, of 20.03 mg g−1 islower than the predicted maximum uptake value (Qm) of 44 mg g−1.This indicate that the Cr(III) removal by non-viable A. haemolyti-cus would proceed at higher Cr(III) concentrations. The constantb value of 0.0166 indicates significant binding strength affinityof Cr(III) towards non-viable A. haemolyticus. The RL (dimension-less separation factor) values of between 0 and 1 also suggestedthe favourable homogenous monolayer mode of adsorption usingnon-viable A. haemolyticus as biosorbent. Moreover, the Freundlichisotherm model also gave the best fit to the experimental data withR2 values of 0.9742. The n value of 2.0792 (which is within then of 1–10 interval) indicates that the biosorption of Cr(III) usingnon-viable A. haemolyticus is favourable at the experimental con-dition used. On the other hand, the D–R and BET isotherms didnot fit well with the experimental data due to the low R2 values of0.7877 and 0.096 respectively suggesting the unlikely occurrence ofmultilayer adsorption, hence limiting the overall biosorption pro-cess to monolayer adsorption only. The mean free energy (Ea) of<8 kJ mol−1 suggests that physical sorption may also take placeduring the biosorption process along with the primary chemicalbinding. The adsorption isotherm parameters are summarized inTable 4.

3.5. Pseudo-first order and second-order kinetic analysis

Analysis on the kinetic parameters i.e. theoretical uptake (qcal),experimental uptake (qexp), correlation coefficient (R2) and thekinetic rate constants (K1 and K2) are as summarized in Table 5while the linearized form of pseudo-first order and pseudo-secondorder is illustrated in Fig. 2.

It can be clearly seen that the pseudo second-order model fit-ted very well (R2 of 0.9814 and 0.9966) with the experimental datacompared to pseudo first-order kinetic model (R2 of 0.0071 and

0.0583). The qe(cal) values increased with increasing initial Cr(III)concentration while the opposite trend was observed for the K2values. The suitability of using the pseudo second-order Lagergrenkinetic model to describe the biosorption of metal ions by various

-order kinetic model for Cr(III) removal by non-viable cells of A. haemolyticus.

366 S.K. Yahya et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 362– 368

Table 4Isotherm constants of Langmuir, Freundlich, D–R and BET models for biosorption ofCr(III) using A. haemolyticus.

Isotherm Parameter A. haemolyticus

Langmuir Qe,max (mg g−1) 20.03Qmax (mg g−1) 44b (L mg−1) 0.0166R2 0.9515RL 0.2525–0.8559

Freundlich Kf 0.0811N 2.0792R2 0.9742

Dubinin–Radushkevich(D–R)

ˇ (mol2 J−2) 5 × 10−5

E (kJ mol−1) 0.1R2 0.7877

tCtcPtptib

3

aHltpH1arp

Fig. 3. Desorption of Cr(III) from Cr(III)-loaded A. haemolyticus using various con-centrations of CH3COOH, HNO3, H2SO4; initial Cr(III) of 100 mg L−1.

TP

BET CBET (L mg−1) 0.562R2 0.096

ypes of biosorbent has also been reported for the biosorption ofu(II) using the chemically modified Uncaria gambir [23], biosorp-ion of Zn(II) using living and non-living strain of Streptomycesiscaucasicus CCNWHX 72-14 [24] and biosorption of Cd(II) usingleurotus platypus [25]. Therefore, it is suggested that the biosorp-ion of Cr(III) by A. haemolyticus can best be described using theseudo second-order kinetic model based on the assumption thathe rate limiting step may be chemical sorption or chemisorptionnvolving valency forces through sharing or exchange of electronsetween biosorbent and Cr(III) ions.

.6. Desorption of Cr(III)

All acid solutions used showed good potential as desorptiongent for Cr(III) with more than 90% of recovery for 0.1 and 1.0 M2SO4, 1.0 M CH3COOH and 1.0 M HNO3 (Fig. 3). This situation is

ikely to occur via the exchange of protons with bound Cr(III) onhe bacterial cells surface, which can be supported from the lowH values of each acids evaluated; 1.0 M H2SO4 (pH 1.32), 0.1 M2SO4 (pH 1.47), 1.0 M HNO3 (pH 1.26), 0.1 M HNO3 (pH 1.66),

.0 M CH3COOH (pH 1.86) and 0.1 M CH3COOH (pH 2.03). Mineralcids can be used as a potent desorbing agent based on its ability toeplace the adsorped metal ions on the biosorbent’s surface withrotons. In view of the weak nature of the metal-binding forces,

Fig. 5. FESEM micrographs of non-viable cells of A. haemolyticus (a) be

able 5seudo first-order and pseudo second-order kinetic rate constants for the biosorption of

Initial Cr(III) conc. (mg L−1) qe(exp) (mg g−1) Pseudo first-order rate cons

qe(cal) (mg g−1) K1 (m

50 38.43 5.499 1.15100 169.43 1.683 2.99

Fig. 4. Biosorption–desorption cycles of Cr(III) by cells of A. haemolyticus; initialCr(III) of 100 mg L−1; SDsorp = 0.89–8.31, SDdesorp = 0.33–5.68 and n = 2.

it is also possible to desorp the bound metal from the biosorbent

using desorbing solution containing other cations such as H+ andCa2+. The effectiveness of desorption process strongly depends onthe binding strength of the metal ion to the active site. Mineral

fore and (b) after desorption using 1.0 M CH3COOH; bar = 1 �m.

Cr(III) by non-viable cells of A. haemolyticus.

tant Pseudo second-order rate constant

in−1) R2 qe(cal) (mg g−1) K2 (min−1) R2

× 10−3 0.0583 36.63 2.4 × 10−3 0.9814 × 10−3 0.0071 120.48 8.05 × 10−4 0.9966

aces B: Biointerfaces 94 (2012) 362– 368 367

af

3

br4acHgd(tfot(aiiraafitCS[

Fb

S.K. Yahya et al. / Colloids and Surf

cids have also been reported to desorp 90% of Pb(II) and Cd(II)rom macrofungus [26].

.7. Biosorption–desorption cycles

During the first biosorption–desorption cycle, non-viableiomass of A. haemolyticus showed high percentage of Cr(III)emoval from initial Cr(III) of 100 mg L−1 with values ranging from5 to 72% of Cr(III) removal. Upon desorption using HNO3, H2SO4nd CH3COOH, only 5–15% of Cr(III) remained on the bacterialells surface (indicating Cr(III) recovery percentage of 85–95%).owever, the percentage of Cr(III) removal and recovery wasreatly reduced in the second cycle where the percentage removalropped to 8–18% while percentage recovery was 5–20% onlyFig. 4). The decrease in the loading capacities following desorp-ion could be due to various factors such as loss of biosorbentrom the reactor, structural damage of biosorbent and blockagef binding sites by metal complex. This can be supported fromhe FESEM micrographs of biosorbent before and after desorptionFig. 5). It can be observed that the biosorbent cells were deformednd clumped after exposure to CH3COOH (1.0 M) thus, resultingn a decrease in surface area/volume ratio hence, less availabil-ty for binding. Also, the blockage of active binding sites by Cr(III)esidue hindered the interaction with newly adsorped Cr(III). Thedsorption of Mn(II) by Pseudomonas sp., Staphylococcus xylosusnd Blakeslea trispora cells were reported to decrease after therst cycle of sorption-desorption when 0.1 M HNO3 was used as

he desorption agent [28] as well as a 50% reduction in terms ofr(III), Cd(II) and Cu(II) uptake capacities for the blue–green algaeprulina sp. when 1.0 M HNO3 was used as the desorption agent29].

ig. 7. TEM micrographs of non-viable cells of A. haemolyticus (a) cells only (b) in 25 mgar = 200 nm; red arrows indicates Cr deposition as electron-dense microparticles.

Fig. 6. Biosorption of Cr(III) from leather tanning wastewater using non-viable cellsof A. haemolyticus; SDremoval = 0.20–2.28, SDuptake = 5.85–17.99 and n = 2.

3.8. Biosorption of Cr(III) from LTW

Around 79.87 mg g−1 of Cr(III) was removed from LTW afterthree consecutive adsorption–desorption cycles (Fig. 6), which wascarried out in similar optimized condition as the simulated solu-tion. However, this value is significantly lower compared to thesimulated solution (198.80 mg g−1) which can be attributed to thepresence of cations such as Na+, K+ and Mg2+ (13–1340 mg L−1) that

interferes with Cr(III) binding to the functional groups present onthe bacterial cells surface. This condition can be supported fromone report [30] where the Cr(III) uptake capacity of Spirogyra con-densata and Rhizoclonium hieroglyphicum were lower compared to

L−1 Cr(III) (c) in 100 mg L−1 of Cr(III), bar = 500 nm and (d) in 100 mg L−1 of Cr(III),

3 aces B

sM

3

cwdcfmbbdt

3

ct[[iNid[

sira1Cttc

4

boec

A

TeYiT

R

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[anions (F− ,H2PO4

−) by viable and autoclaved cells of the Gram-negative bac-

68 S.K. Yahya et al. / Colloids and Surf

imulated Cr(III) solution due to interference from cations such asg2+.

.9. TEM analysis

The bacterial cell structure remained intact even after auto-laving at 121 ◦C. However, the disappearance of the bacterial cellall outer layer was noticeable. Uneven distribution of electron-ense microparticles on the outer region of the bacterial cellslearly indicates the deposition of Cr(III) onto the bacterial cellsunctional groups without further translocation into the cytoplas-

ic region (Fig. 7). The absence of electron-dense structure at theacterial cytoplasmic region strongly suggest that Cr(III) removaly non-viable A. haemolyticus proceeds via the non-metabolicallyependent biosorption followed by microprecipitation on the bac-erial cells outer region.

.10. FTIR analysis

The band assignments for native non-viable cells of A. haemolyti-us showed strong absorption at 3414.3 cm−1 that can be attributedo the overlapping stretching vibrations of NH and OH groups31], 2924.38 cm−1 due to CH2 asymmetric stretching vibration32], 1651.81 cm−1 for the typical amide band i.e. C O stretch-ng, 1544.45 cm−1 which corresponded to amide II (combination ofH bending and CN stretching modes), 1401.81 cm−1 for CO bend-

ng from carboxylate group and 1083.61–1240.13 cm−1 which wereue to the vibrational motion of the carboxyl and phosphate groups33].

Upon contacting with Cr(III), several peaks were significantlyhifted notably 3414.34 cm−1 to 3431.91 cm−1 indicating thenvolvement of hydroxyl and amine group during the Cr(III)emoval process, the amide and amide II peaks at 1651 cm−1

nd 1544.45 cm−1 that were slightly shifted to 1649 cm−1 and540.56 cm−1 respectively which suggested the complexation ofr(III) ions with functional groups from protein [17], 1401.81 cm−1

o 1388.88 cm−1, 1240.13 cm−1 to 1234.84 cm−1 and 1083.61 cm−1

o 1071.49 cm−1 which may be due to the binding of Cr(III) witharboxyl, phosphate and carbonyl groups respectively [34].

. Conclusion

The non-viable cells of A. haemolyticus showed great potential toe used as an alternative biosorbent to remove Cr(III) from aque-us solution. Its superior metal uptake ability compared to otherxisting biosorbents warrants further investigation on its uptakeapacity in view of accelerating its industrial application.

cknowledgments

The authors acknowledge the support from Ministry of Science,echnology and Innovation (MOSTI), Malaysia for the National Sci-nce Fellowship (NSF) scholarship awarded to Siti Khairunnisaahya, the Ministry of Higher Education (MOHE), Malaysia for fund-

ng of the project (FRGS Vote 78465 and 78532) and Universitieknologi Malaysia for the GUP grant Q.J13000.7125.00H52.

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