thuoc nhuom 1

7/30/2019 thuoc nhuom 1

1/7

Journal of Hazardous Materials 167 (2009) 9951001

Contents lists available at ScienceDirect

Journal of Hazardous Materials

j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / j h a z m a t

Understanding effects of chemical structure on azo dye decolorization

characteristics by Aeromonas hydrophila

Chung-Chuan Hsueh, Bor-Yann Chen, Chia-Yi Yen

Department of Chemical and Materials Engineering, National I-Lan University, I-Lan 260, Taiwan

a r t i c l e i n f o

Article history:

Received 1 December 2008

Received in revised form 20 January 2009Accepted 21 January 2009

Available online 30 January 2009

Keywords:

Biodecolorization

Aeromonas hydrophila

Azo dye

Chemical structure

a b s t r a c t

This novel comparative study tended to disclose how the molecular structures present in seven azo dyes

including two types of azo dyes (i.e., naphthol type azo dyes Reactive Black 5 (RB 5), Reactive Blue 171

(RB 171), ReactiveGreen 19(RG19), ReactiveRed 198 (RR198), ReactiveRed 141 (RR141) andnon-naphthol

type azo dyes Direct Yellow 86 (DY86), Reactive Yellow 84 (RY84)) affected color removal capability of

Aeromonas hydrophila. Generallyspeaking, the decolorization rateof naphthol type azo dye withhydroxyl

group at ortho to azo bond was faster than that of non-naphthol type azo dye without hydroxyl group,

except of RG19. The azo dyes with electron-withdrawing groups (e.g., sulfo group in RR198, RB5 and

RR141) would be easier to be decolorized than the azo dyes with the electron-releasing groups (e.g.,

NH-triazine in RB171and RG19). In addition,the azo dyescontaining moreelectron-withdrawing groups

(e.g.,RR198, RB5and RR141) showed significantly faster rateof decolorization.The azodyes withelectron-

withdrawing groups (e.g., sulfo group) at para and ortho to azo bond (e.g., RR198, RB5 and RR141) could

be more preferred for color removal than those at meta (e.g., DY86 and RY84). The former azo dyes with

para and ortho sulfo group provided more effective resonance effects to withdraw electrons from azo

bond, causing azo dyes to be highlyelectrophilic for faster rates of reductive biodecolorization. However,

since the ortho substituent caused steric hindrance near azo linkage(s), azo dyes with para substituent

could be more favorable (e.g., SO2(CH2)2SO4 in RR198 and RB5) than those with ortho substituent (e.g.,

sulfo group at RR141) for decolorization. Thus, the ranking of the position for the electron-withdrawing

substituent in azo dyes to escalate decolorization was para > ortho > meta. This study suggested that boththepositions of substituentson thearomatic ring andthe electronic characteristicsof substituentsin azo

dyes all significantly affected the performance of biodecolorization ofA. hydrophila.

2009 Elsevier B.V. All rights reserved.

1. Introduction

Azo dyes are the largest chemical class of synthetic dyes, and

widely used as colorants in textile dyeing, leather, plastics, food,

cosmetics, paper printing, with the textile industry as the largest

consumer [13]. Inevitably, the residual azo dyes in wastewa-

ter from the dyestuff or textile industry would be a significant

threat to public and environmental health. Azo dyes are originally

designed to be recalcitrant for long-term use and thus resistantto aerobic wastewater treatment [25]. Moreover, azo dyes are

electron-deficient xenobiotics and thus are capable to be degrad-

able via azo reduction [26]. However, due to diverse structures

present in the synthetic dyes, changes in the chemical structures

(e.g., isomers or the presence of different substituents) would

significantly affect the decolorization capability (e.g., reductive

biodecolorization). Most of researchers have related the capacity

Corresponding author. Fax: +886 39357025.

E-mail addresses: [email protected] , [email protected] (B.-Y. Chen).

of color removal to the dye class rather than the molecular fea-

ture [7,8]. However, the characteristics of substituents and their

relative positions to azo bond all played crucial roles for the per-

formance of azo dye decolorization. Moreover, Zimmermann et al.

[9] used purified Orange II azoreductase from a Pseudomonas strain

KF46to assessdecolorization efficiency of various Orange dyes. Evi-

dently, the specificity of Orange II azoreductase toward azo linkage

for decolorization is strongly dependent upon the properties (e.g.,

electron-withdrawing capability) of substituents in the proximityof azo linkage(s) and thus strongly determines whether the dye is

susceptible to biodecolorization. On the other hand, the hydroxy

group on the 2-position of the naphthol ring of the azo dye (e.g., 1-

(4-sulfophenylazo)-2-naphthol) provides a beneficial prerequisite

to assist dye decolorization. In contrast, charged groups near azo

bond (e.g., 1-(2-sulfophenylazo)-2-naphthol) significantly hinder

the decolorization efficiency. Zimmermann et al. [9] also quantita-

tively revealed thatthe slopeof the correlation between Hammetts

substituent constant, (an experimental value of the electronega-tivityofasubstituentonaphenylring[10]) of varioussubstrates and

the logarithm of the decolorization rate of these azo dyes was pos-

0304-3894/$ see front matter 2009 Elsevier B.V. All rights reserved.

doi:10.1016/j.jhazmat.2009.01.077
http://www.sciencedirect.com/science/journal/03043894http://www.elsevier.com/locate/jhazmatmailto:[email protected]:[email protected]://localhost/var/www/apps/conversion/tmp/scratch_8/dx.doi.org/10.1016/j.jhazmat.2009.01.077http://localhost/var/www/apps/conversion/tmp/scratch_8/dx.doi.org/10.1016/j.jhazmat.2009.01.077mailto:[email protected]:[email protected]://www.elsevier.com/locate/jhazmathttp://www.sciencedirect.com/science/journal/03043894


2/7

996 C.-C. Hsueh et al. / Journal of Hazardous Materials 167 (2009) 9951001

itive (i.e., the electron-withdrawing groups present on the phenyl

ringescalatethis colorremoval).To have conclusiveremarks, Suzuki

et al. [11] also provided a correlation of aerobic biodegradability of

25 sulfonated azo dyes with their chemical structures. Although

there are many structure-based methods to reveal biodegradation

as well as biodegradability, they still need an abundant supply of

database from experiments to obtain highly adequate model pre-

dictions [12]. Recently, our prior studies presented preliminary

comparisons to propose the plausible reasons for reaction selec-

tivity of azo dye decolorization by Pseudomonas luteola [13,14].

These results disclosed that the sulfo group produced the strongest

electron-withdrawing effect to result in the fastest rate of color

removal compared to the carboxyl and hydroxyl group. In addition,

the sulfo or carboxyl group was ortho to azo bond, the decoloriza-

tion rate significantly decreased compared to the para substituent

to azo bond (e.g., p-MO> o-MO or p-MRo-MR; refer to [14] for

details) likely due to steric hindrance near azo linkage(s). How-

ever, detailed figures of chemical structures related to theirreaction

selectivityin a systematicanalysis were still remained open fordis-

cussion. Although thousands of organic compounds (e.g., azo dyes)

have been developed for myriads of uses, most of the biodegrad-

ability andpersistence of these compoundsin environment arestill

remained unexplored. These compounds in particular recalcitrant

or cytotoxic ones might be invisible threats to the globe [15]. Thus,this study tended to provide a systematized comparison, in general

terms, to uncover these detailed figures.

The model bacterium Aeromonas hydrophila which expressed

a promising capability for color removal was predominantly iso-

lated from fountain springs in Northeast Taiwan [16]. Chen et

al. [16] mentioned that A. hydrophila could exhibit 10-fold color

removal performance of azo dyes as much to P. luteola. However,

the mysteries to have such a high performance of A. hydrophila for

decolorization still remained to be unsolved for possible applica-

tions. In addition, A. hydrophila is known to have high productivity

for biodegradable polymer (e.g., polyhydroxy-alkanoates (PHAs))

[17]. As known, polyhydroxy-alkanoates can be accumulated by

various microorganisms (e.g., A. hydrophila) as carbon and energy

storage materials under unbalanced growth conditions in thepresence of excess carbon source. However, Aeromonas species is

probably more appropriate to be cultivated in closed-vessel biore-

actor due to its possible pathogenic characteristics [18]. Recently,

Chen et al. [19] presented a comparative assessment to quantita-

tively disclose interactive toxicities of some aromatic amines (AAs;

intermediates of decolorization) toA. hydrophila and P. luteola, indi-

cating that A. hydrophila had a higher tolerance to AAs for a more

promising performance of color removal. This follow-up study was

to compare reductive decolorization performance of some model

azo dyes using A. hydrophila, suggesting how the steric hindrance

of substituents at different position (e.g., ortho, meta and para)

to azo bond and electronic effect (e.g., electron-withdrawing or

releasing substituent) in azo dyes affect biodegradability. There-

fore, we could associate higher color removal efficiency of A.hydrophila with higher tolerance to decolorization intermedi-

ates (AAs) and different reactivities of chemical structures in azo

dyes.

2. Materials and method

2.1. Chemicals

Seven azo dyes (Reactive Black 5 (RB5), Reactive Blue 171

(RB171), Reactive Green 19 (RG19), Reactive Red 198 (RR198), Reac-

tive Red 141 (RR141), Direct Yellow 86 (DY86), Reactive Yellow

84 (RY84); Everlight Chemical Ltd., Taipei, Taiwan) were classified

into two categories. Category A dyes, naphthol type azo dyes (e.g.,

RR198 (mono azo dye), RB5, RR141, RB171 and RG19; Fig. 1A), were

synthesizedfrom H acid (8-amino-1-naphthol-3,6-disulfonic acid).

Category B dyes were non-naphthol type dyes (RY84 and DY86;

Fig. 1B).

2.2. Culture conditions

A. hydrophila predominantly isolated fromfountain springs near

Chiao-Hsi in Northeast Taiwan was used as a reporter bacterium

for decolorization performance [16]. To guarantee the consis-tent growth characteristics of cultures for study, a loopful of A.

hydrophila seed taken from an isolated colony on a LB-streak plate

was precultured in 50-mL LB broth, Miller (in g L1: casein pep-

tone 10.0, yeast extract 5.0, sodium chloride 10.0) for 24 h at 30 C,

125rpm using a water bath shaker (SHINKWANG, SKW-12). Then,

precultured broths in appropriate volume ratios were inoculated

into fresh YE medium (in g L1: yeast extract 5.0, sodium chloride

10.0) with ca. 0.3g L1 dye for shake flask cultures. Once cells had

grown to late exponential or early stationary phase (ca. 6 h), shak-

ing was switched off and the culture was kept in static incubation

(i.e., t 0) for reductive decolorization.

2.3. Analytical methods

The model azo-dyes used for decolorization were RB 5

(max = 600nm), RB171 (max = 609 nm), RG19 (max = 631 nm),RR198 (max = 522 nm), RR141 (max = 5 44 nm), DY86(max =393nm), RY84 (max = 411 nm). Dye solutions were ster-ilized by filtration (Millipore Millex-GS 0.22mm filter unit),

since azo dyes may be unstable in moist-heat sterilization. With

appropriate calibrations at specific wavelengths, concentrations

of biomass and dyes were determined using an UVvis spec-

trophotometer (HITACHI Spectrophotometer, model UV-2001).

The concentration of dye was primarily determined by measuring

the optical density (OD) of the supernatant of the sample after cen-

trifugation for5 minat 700g(HSIANGTAI Centrifuge MCD-2000).

In addition, a sterile cell-free medium was chosen as the control

for spectrophotometric measurement. As all samples containedbiomass and dye, concentrations of biomass ((a) and (b)) and dye

(c) were evaluated as follows:

(a) OD600nm of sample mixtureswithoutcentrifugation: ODX+Dye600nm =

ODX600nm +ODDye600nm

,

(b) OD600nm of sample supernatant (sup) after centrifugation:

ODsup600nm = OD

Dye600nm

,

(c) ODmax of sample supernatant after centrifugation: ODX+Dyemax

=

ODXmax +ODDyemax

.

Samples were diluted to an optical density of less than 0.6 when

absorbance was not within the linear arrange (ca. 0.10.7). The

relation ship between the biomass concentration and OD600nm is1.0 ODU=0.373gL1 drycell weight= (1.70.2)108 cfumL1. As

the function of cell density (X) and dye concentration ([Dye]) are

continuous, strictly monotonic and differentiable over time, their

differential terms dX and d[Dye] could be denoted by forward-

difference formulae (e.g., dX= Xt+t

Xt=X, d [Dye] =

[Dye]t+t

[Dye]t= [Dye]) for specific rate determinations. To

ensure the step size tsufficiently small enough for convergence,numerical differentiations were compared with differentiations by

reducing step size ast/2

i.e.,

(dft df

t/2

)

/ dft

.Only the error less than 1% was defined as achieving of convergent

criteria within the calculated tolerance. Otherwise, the step sizet

was continuously reduced by half for approximations until conver-

gence was reached. Therefore, specific growth rate (SGR), and


3/7

C.-C. Hsueh et al. / Journal of Hazardous Materials 167 (2009) 9951001 997

Fig. 1. (A) Naphthol type azo dyes (Category A dyes RR198, RB5, RR141, RB171 and RG19) used for the comparative study. (B) Non-naphthol type azo dyes (Category B dyes

RY84 and DY86) used for the comparative study.


4/7


specific decolorization rate qp (SDR) could be evaluated as

SDR= qp =

1

X

d [Dye]

dt

=

1

X

[Dye]

t

and

SGR= =dlnX

dt=

1

X

X

t

,

respectively; where X, [Dye], and t denoted for cell concentration,

dye concentration, and time, respectively.

2.4. Construction of phase-plane portraits

According to Perlmutter [20], the (bio)chemical reactor models

could be in terms of the form:

SDR= qp = 1

X

d [Dye]

dt=f1 (X, [Dye] , S) , (1a)

SGR= =1

X

dX

dt=f2 (X, [Dye] , S) , (1b)

whereX, [Dye]and Sdenoted theconcentration of biomass, dye and

limiting nutrient substrate, respectively. Such models lend them-

selves to a graphical analysis in which the dependent variables (or

state variables) are plotted against one another on what is called

a phase-plane. Moreover, for phase-plane analysis it is postulated

that the contribution of other factors (e.g., nutrient substrate, dis-

solved oxygen) was relatively insignificant compared to two major

state variables (i.e., dye and biomass concentration). As the system

evolves continuously over time through a series of states from its

initial to final condition, its mapping on the (X, [Dye]) plane will

trace a trajectory to be a biological fingerprint specific to the cul-

turesystem. Tocombinethis perspective withGadensclassification

scheme of fermentation [21], the phase-plane was modified to dis-

close onthe(, qp) plane since theinstantaneous slope of thecurveinthe(X, [Dye])plane and inthe (, qp) planeare theoreticallyiden-tical. For example, the instantaneous slope m on the (, qp) plane

can be obtained by the formula

qp=1/X

d [Dye]/dt

1/X

dX/dt

=d [Dye]dX

=f1 (X, [Dye] , S)f2 (X, [Dye] , S)

=m. (2)

And the entire trajectory can be drawn as a sequence of small

steps using the computed slope at each step to move on to the next

(i.e., exploitedin the trajectory direction to show time frame on the

phase-plane). This is so-called a set of auxiliary isoclines in the (X,

[Dye]) plane:

f1 =mf2 ,

which is the loci of points where all of their trajectories have slope

m. As Perlmutter [20] indicated, forsystems of the form (1), the tra-

jectory from any point will be unique when the f1 and f2 functions

have continuous first partial derivatives (e.g., the cases of biochem-

ical reactor model shown herein). In light of these, no trajectoriescanever cross,exceptat singular pointswherethe derivativesdX/dt

and d[Dye]/dt are both zero (i.e., lag phases in growth and phe-

nol degradation curves should be excluded) to prevent different

bases for comparison. Thus, it is feasible to apply this phase-plane

analysis to reveal growth-dependent phenomena of fermenta-

tion.

3. Result and discussion

3.1. Naphthol type azo dyes

The ranking of maximal specific decolorization rate

(mgL1 h1 ODU1) of A. hydrophila NIU01, in decreasing

order, was monoazo dye RR198(260.2)> RB5(80.9)> RR141(66.5)

Fig. 2. Comparison on time courses of bacterial growth and decolorization of A.

hydrophila cultureswith variousazo dyes(e.g., naphthol andnon-naphthol typeazo

dyes) at initial concentrations of 300 mg L1 (arrows denoted the maximal specific

decolorization rates (in mg L1 h1 ODU1): RR198(260.2); RB5(80.9); RR141(66.5);

RB171(36.0); RY84(33.7); DY86(30.9); RG19(22.5).

> RB171(36.0) > RY84(33.7) > DY86(30.9) > RG19(23.2) (Fig. 2 and

Table 1). As indicated in Fig. 3, the trajectories in phase-plane

plots were all nearly located in +SDR and +SGR axes (except RB5),

indicating that azo dye decolorization was non-growth associated.

That is, the maximal biodecolorization was only taking place when

cell growth was repressed. Although Chen et al. [22] showed the

relative order of initial specific decolorization rates of various dyes

(RR198> RB5> DY86> RY84RR141RB171RG19; Table 1), both

relative rankings were almost in parallel. These consistent results

seemed to support that azo dye decolorization was a genotypically

constitutive characteristics of A. hydrophila regardless of origins

of strain isolation. Note that A. hydrophila NIU01 [16] and DEC

Table 1

Comparison upon decolorization performance of Aeromonas hydrophila isolated

from different sources.

Type of dyes (C.I. number) Decolorization ratea, c Decolorizationb, d

NIU01e DEC2f DEC3f DEC5f

Reactive Red 198 (18221) 260.2 85 2 86 3 82 3

Reactive Black 5 (20505) 80.9 67 3 62 1 66 3

Reactive Red 141 () 66.5 17 2 13 1 10 2

Reactive Blue 171 () 36.0 15 2 13 1 15 2

Reactive Yellow 84 () 33.7 19 2 15 2 14 2

Direct Yellow 86 () 30.9 28 2 20 1 20 2

Reactive Green 19 () 23.2 14 2 10 3 7 2

a Unit: mg L1 h1 ODU1.b Unit: %.c Initial concentration 300 mg L1.d Initial concentration 100 mg L1.e This study.f Data adopted from Chen et al. [22].


5/7


Fig. 3. Phase-plane profiles of SGR versus SDR for the batch cultures with various

azo dyes. The arrows indicated the orientation of all trajectories of profiles as time

went by.

strains [22] were isolated from a unpolluted fountain spring and

dye-laden wastewater treatment sludges, respectively.

In general, the decolorization rates of naphthol type azo dyes

(RR198, RB5, RR141, RB171 and RG19) except RG19 were apparently

higher than those of non-naphthol type azodyes (RY84, DY86). The

reasons to cause such an order in decolorization rates are straight-

forward.All of naphthol typeazo dyes contained a hydroxyl group at

ortho toazo bond, as theywere synthesized fromH acid(Fig. 1A).In

contrast,non-naphthol typeazo dyeswere freeof hydroxyl group(s)

in their chemical structures (Fig. 1B).

Thephenomenaof fast decolorization ratesof naphthol type azo

dyes could also be confirmed in the findings of Zimmermann et al.

[9]. They pointed out that the hydroxyl group on the 2-position

of the naphthol ring in the azo dye (e.g., 1-(4-sulfophenylazo)-2-

naphthol) is very likely prerequisite for dye decolorization. Dueto the hydroxyl group on the 2-position of naphthol ring in azo

dyes, azo-hydrazone tautomerism (i.e., N N NNH) (refer

to Fig.4 foranexampleofRB5; [23,24]) couldtakeplaceandthusthe

hydroxyl group (i.e., an electron-releasing group) would become

carbonyl group (i.e., an electron-withdrawing group). Due to the

presence of an electron-withdrawing group (carbonyl group) dyes

could be reductive to be more favorable for biodecolorization and

could thus stabilize the reduced forms of azo dyes by A. hydrophila

(Fig. 5).

Although the sulfo group at ortho to azo bond in those naph-

thol type azo dyes (e.g., RR198 as shown in Fig. 1A) introduced

steric hindrance to reductive biodegradation of azo dyes, electrons

were still withdrawn by inductive and resonance effect due to high

electronegativity characteristics of the sulfo group. As the redox

potential of azo substrate became more positive, this molecule is

more readily to be reduced. That is, the azo bond became more

electrophilic and was thus more favorable to be reduced, as the

sulfo group stabilized the negative charge generated in reduced

dyes (Fig. 5).

In addition,even the high electronegativity group was present at

para to azo bond, it still could withdraw electron by the resonance

effect as shown in Fig. 5. This led to azo dye more preferred for

reductive biodecolorization compared to the group at ortho, due

to lack of steric hindrance in the proximity of the azo bond [14].

In addition, Pearcea et al. [24] also mentioned that the substi-

tution of electron-withdrawing groups (SO3H, SO3NH2) in the

para position of the phenyl ring, relative to the azo bond, causes an

increase in the reduction. In contrast, compared to ortho and para

in naphthol type azo dyes, a sulfo group at meta toazo bond in non-

naphthol type azo dyes resulted in the least electron-withdrawing

capability through the weak inductive effect. Apparently, the rel-

ative position (e.g., ortho, meta, para) of the electron-withdrawing

substituent to azo bond considerably influenced the capability ofbiodecolorizationofA. hydrophila. In contrast,azo dyes without any

electron-withdrawing substituent on aromatic ring would decrease

the efficiency of azo dye decolorization [14].

Among five naphthol type azo dyes, the decolorization rate of

RR198 was ranked the top one due to the sulfo group at ortho and

SO2(CH2)2SO4 at para to azo bond. Both high electronegativ-

ity groups would withdraw electron from azo bond via resonance,

leading to the azo bond more electrophilic and reductive for color

removal. Plus, they could also stabilize the negative charge on

reductive azo dye (Fig. 5). Moreover, a monoazo dye (i.e., RR198)

could be more favorable to be decolorized than polyazo dyes. That

is, the more the azo bonds, the less readily the molecule is reduced

[13,22,25].

RB5 was ranked the second fast for biodecolorization in thenaphthol type azo dyes. RB5 was synthesized from H acid to be

a symmetric diazo dye (Fig. 1A). Two group (SO2(CH2)2SO3Na)

at para to azo bond on aromatic ring withdrew electrons from

azo bond via resonance, causing azo dye more electrophilic for

reductive decolorization. Thus, the negative charge was stabilized

when azo dye was reduced as indicated previously in RR198. How-

ever, the biodecolorization rate of RB5 was less than that of RR198

since diazo dye was less reductive for color removal than monoazo

dye.

Fig. 4. Example demonstration of azo-hydrazone tautomerism of RB5.


6/7


Fig.5. Examplepresentation ofresonanceeffectof RR198.The sulfo group atortho andSO2(CH2)2SO4 atpara toazo bondin RR198are bothhighelectronegativitysubstituents.

These substituents may stabilize the negative charge present in the reduced intermediates, since azo dyes can be reduced through the inductive effect and resonance effect.

Regarding RR141, it was a symmetric diazo dye (i.e., symmetric

centerat thebenzene ring as shown in Fig.1A)andhadasulfogroup

at ortho to azo bond on each of four naphthalene rings. Although

these introduced more considerable steric hindrance for azo bond

breakage,theystillcouldwithdraw electron from azobondby reso-

nance,allowing azo dyemore electrophilic tobe reduced. Moreover,

the sulfo group could stabilize the negative charge of reduced azo

dye as mentioned before(RR198,RB5). Plus, dueto steric hindranceeffect from sulfo group at ortho to azo bond, the biodecolorization

rate of RR141 was thus less than that of RB5.

RB171 and RG19 are isomers of a symmetric diazo dye and both

were synthesized from H acid with an electron-releasing group

(NH-triazine). This electron-releasing group was present at meta

to azobondin RB171, butatpara toazobond inRG19.In the effectof

resonance,para position was more influencedto azobond breakage

than meta position. That is, the azo bond of RG19 was less elec-

trophilic because of an electron-releasing group (NH-triazine) at

para to azo bond. This appreciably changed the azo bond less fea-

sible to be reduced, causing the negative charge to be less stable

at reduced azo dye. Thus, its biodecolorization rate was ranked the

last in seven azo dyes.

In comparison with RB171 and RR141, the decolorization rateof RB171 was slower than that of RR141 because of an electron-

releasing group (NH-triazine) existing in RB171. As RR141 was

more electrophilic than RB171, RR141 was more feasible to be

reduced for color removal [24].

In general, the less electron density in the proximity of azobond

induced this azo bond more electrophilic for reductive decoloriza-

tion. That is, substituent(s) with high electronegativity near azo

bond could stabilize the negative charge formed at reduced azo

dye, considerably escalating the decolorization rate of azo dye.

3.2. Non-naphthol type azo dyes

Although there existed a methyl group at ortho to azo bond in

non-naphthol type azo dyes RY84 and DY86 (Fig. 1B), the decol-

orizationrates of bothdyeswereslowerthan those ofnaphtholtype

azo dyes (except RG19) with hydroxyl group at ortho to azo bond.

Nigam et al. [26] mentioned that azo compounds with a hydroxyl

group or an amino group are more likely to be degraded than those

with a methyl, methoxy, sulfo or nitro group. This was due to the

azo-hydrazone tautomerism about naphthol type azo dyes with

hydroxyl group at ortho to azo bond as shown previously.

Even RY84 andDY86 were both synthesized from sulfonaphtha-lene, the decolorization rate of RY84 was relatively faster than that

of DY86. This was because there was four sulfo groups (electron-

withdrawing group) conjugate with azo bond in RY84. However,

there is an electron-releasing group (i.e., NH(CH2)2OH) conjugate

with azo bond in DY86. The sulfo group could withdraw electron

from azo bond by resonance, and the environment in the prox-

imity of azo bond became more electrophilic for higher reductive

decolorization.

4. Conclusion

Considering chemical structures and decolorization rates of

seven model azo dyes by A. hydrophila, we could reach a con-

clusive remark. For similar structures of azo dyes, monoazo dyeswere easily biodecolorized than polyazo dyes [13,24]. Azo dyes

with hydroxyl group at ortho to azo bond were beneficial to biode-

colorization by A. hydrophila [9]. The azo bond was thus in a less

electron density to become more electrophilic for reductive biode-

colorization. Literature [9,14] also confirmed that azo dyes with

electron-withdrawing groups would be more feasible for reduc-

tive decolorizationthan thosewith electron-releasing groups. Thus,

azo dyes with more electron-withdrawing groups near azo bond

would more feasible to be reduced for decolorization. Plus, as the

position of electron-withdrawing groups to azo bond significantly

influenced the reductive decolorization, their ranking of the decol-

orization rate waspara > ortho > meta. Sincepara and ortho position

could more efficiently withdraw electron from azo bond through

resonance, azo bond became more electrophilic to be reduced for


7/7


color removal [6,14]. However, compared to substituents at para

the steric hindrance at ortho would become more significantly to

result in a slowerdecolorization. This study explored more system-

atized details of chemical structure effect in azo dye decolorization

byA. hydrophila. This findingwas almostconsistentwith thefinding

for P. luteola (i.e., membrane-enclosed biocatalyst) [14] and puri-

fied azo reductase (i.e., naked biocatalyst) [9]. Follow-up studies

would focus on effects of other chemical structures in azo dyes

using different biodecolorizers to grasp a general perspective for

biodecolorization.

Acknowledgements

Financial supports (NSC 95-2221-E-197-005, NSC 96-2221-E-

197-012, NSC 97-2221-E-197-019) from National Science Council,

Taiwan, R.O.C. for this research and seeding grants for Biochemical

Engineering Laboratorysdg of National I-Lan University (NIU) from

the Ministry of Education, Taiwan, R.O.C. are very much appreci-

ated. The authors also extend sincere appreciation to Distinguished

Professor Jo-Shu Chang (Department of Chemical Engineering,

National Cheng Kung University) for valuable suggestions.

References

[1] S.V. Kulkarni, C.D. Blackwell, A.L. Blackard, C.W. Stackhocese, M.W. Alexander,Textile Dyesand DyeingEquipment, Classification, Propertiesand Environmen-tal Aspects,U.S. Environmental Protection Agency, Research TrianglePark, N.C.,1985, EPA-600/2-85/010.

[2] A. Stolz, Basic and applied aspects in the microbial degradation of azo dyes,Appl. Microbiol. Biotechnol. 56 (2001) 6980.

[3] A. Pandey, P. Singh, L. Iyengar, Bacterial decolorization and degradation of azodyes, Int. Biodeterior. Biodegrad. 59 (2007) 7384.

[4] K.T. Chung, S.E. Stevens Jr., Degradation of azodyes by environmental microor-ganisms and helminths, Environ. Toxicol. Chem. 12 (1993) 21212132.

[5] P.G. Rieger, H.M. Meier, M. Gerle, U. Vogt, T. Groth, H.J. Knackmuss, Xenobioticsin the environment: present and future strategies to obviate the problem ofbiological persistence, J. Biotechnol. 94 (2002) 101123.

[6] H.J. Knackmuss, Basic knowledge and perspectives of bioelimination of xeno-biotic compounds, J. Biotechnol. 51 (1996) 287295.

[7] A.J. Greaves, D.A.S. Phillips, J.A. Taylor, Correlation between the bioelimina-tion of anionic dyes by an activated sewage with molecular structure. Part 1.

Literature review, J. Soc. Dyers Colorists 115 (1999) 363365.

[8] H.R. Hitz, W. Huber, R.H. Reed, The absorption of dyes on activated sludge, J.Soc. Dyers Colorists 94 (2) (1978) 7176.

[9] T. Zimmermann, H.G. Kulla, T. Leisinger, Properties of purified orange II azore-ductase, the enzyme initiating azo dye degradation by Pseudomonas KF46, Eur.

J. Biochem. 129 (1982) 197203.[10] L.P. Hammett,The effect of structure uponthe reactions of organic compounds

benzene derivatives, J. Am. Chem. Soc. 59 (1937) 96103.[11] T. Suzuki, S. Timofei, L. Kurunczi, U. Dietze, G. Schuurmann, Correlation of

aerobic biodegradability of sulfonated azo dyes with the chemical structure,Chemosphere 45 (2001) 19.

[12] J.W. Raymond, T.N. Rogers, D.R. Shonnard, A.A. Kline, A review of structure-

based biodegradation estimation methods, J. Hazard. Mater. B84 (2001)189215.

[13] C.C. Hsueh, B.Y. Chen, Comparative study on reaction selectivity of azo dyedecolorization by Pseudomonas luteola, J. Hazard. Mater. 141 (2007) 842849.

[14] C.C.Hsueh, B.Y. Chen, Exploring effects of chemical structure on azo dye decol-orization characteristics by Pseudomonas luteola, J. Hazard. Mater. 154 (2008)703710.

[15] L.P. Wackett, L.B.M. Ellis, Predicting biodegradation, Environ. Microbiol. 1 (2)(1999) 119124.

[16] B.Y. Chen, W.M. Chen, F.L. Wu, P.K. Chen, C.Y. Yen, Revealing azo-dye decol-orizationof indigenousAeromonashydrophila fromfountainspringin NortheastTaiwan, J. Chin. Inst. Chem. Eng. 39 (2008) 495501.

[17] S.Y. Lee, Bacterial polyhydroxyalkanoates, Biotechnol. Bioeng. Rev. 49 (1996)114.

[18] B.Y. Chen, W.M. Chen, H.Y. Kuo, C.C. Hsueh, Comparative assessment upon dyeremovalcapability of indigenous bacterial strains fromLanyang Plain in north-east Taiwan, J. Hazard. Mater. 161 (1) (2009) 526533.

[19] B.Y. Chen, K.W. Lin, Y.M. Wang, C.Y. Yen, Revealing interactive toxicity of aro-matic amines to azo-dye decolorizer Aeromonas hydrophila. J. Hazard. Mater.

(2009), in press.[20] D.D. Perlmutter, Geometry of dynamics, in: Stability of Chemical Reactors,

Prentice-Hall, U.S.A., 1972, p. 51, Chapter 3.[21] E.L. Gaden Jr., Fermentation process kinetics, J. Biochem. Microbiol. Tech-

nol. 1 (4) (1959) 413429 (Reprinted Biotechnol. Bioeng. 67(6) (2000)629635).

[22] K.C. Chen, J.Y. Wu, D.J. Liou, S.C.J. Hwang, Decolorization of the textile dyes bynewly isolated bacterial strains, J. Biotechnol. 101 (2003) 5768.

[23] J.A. Libra, M. Borchert, L. Vigelahn, T. Storm, Two stage biological treatment ofa diazo reactive textile dye and the fate of the dye metabolites, Chemosphere56 (2004) 167180.

[24] C.I.Pearcea,J.R. Lloydb,J.T.Guthriea,The removalof colour fromtextile wastew-ater using whole bacterial cells: a review, Dyes Pigments 58 (2003) 179196.

[25] T.L. Hu, Kinetics of azoreductase and assessment of toxicity of metabolic prod-ucts from azo dyes by Pseudomonas luteola, Water Sci. Technol. 43 (2) (2001)261269.

[26] P. Nigam, I.M. Banat, D. Singh, R. Marchant, Microbial process for the de-colorization of textile effluent containing azo, diazo and reactive dyes, Process

Biochem. 31 (5) (1996) 435442.

Documents

thuoc nhuom 1