DECOLORIZATION OF AZO-REACTIVE DYES BY PSEUDOMONAS PUTIDA Imed Yousfi 1, 2,Néji ladhari 1, Kamel chaieb 2, Tarek Zmantar 2 and Amina Bakhrouf 2 1
Textile Research Unit of ISET Ksar Hellal, B.P 68 Ksar Hellal 5070, Tunisia Tel. (216)73475900
2
Laboratory Analysis and Control of the Chemical Polluents and Microbiological of the Environment, Faculty of pharmacy, Monastir 5000, Tunisia. Email:
[email protected]
Abstract This study was taken up in order to optimize the decolorization azo dyes present in textile wastewater. We are interested in the Pseudomonas putida bacteria because it is know for its high ability to reduce the azo bond and it is not pathogenic. The effect of operational conditions in different combinations (agitation, filamentous fungi, Floating support (Liege of packing) and light) were investigated on microbial decolorization. In optimum conditions, this bacteria was able to decolorize 92 % of reactive Blue 40, 82 % of reactive Yellow 174 and 73 % of reactive Red 220 in 7 days.
Key words Decolorization, Pseudomonas putida, reactive dyes, operational conditions
Introduction The textile dyeing and finishing industry use wide variety of dyestuffs due to rapid changes in the customer‘s demands. The world annual production of the dyestuffs amounts to more than 7×105 tonnes [1]. Azo dyes, containing one or more azo band (-N=N-), account for 60-70 % of all textile dyestuffs used [2]. Despite the attempts of dyestuff manufactures to optimize the exhaustion and fixation characteristic of their dyes from commercial reasons, residual colour in dyehouse effluents continues to cause problems. In fact, estimated that about 10-15 % of the total production of dyes is lost during their synthesis and dyeing processes [3, 4]. Colored industrial effluent is the most obvious indicator of water pollution and discharge of highly colored synthetic dyes effluents is aesthetically displeasing and cause considerable damage to the receiving water bodies by impeding the penetration of light. Many microorganisms like bacteria [5], fungi [6], actinomycetes [7] and algae [8] have been reported for their ability to decolorize azo dyes. The bacteria has many advantages compared to other microorganisms. In fact, the growth of bacteria is very fast and some bacteria are known for their high ability to reduce azo band [6]. The purpose of this work is to optimize the biodegradation of three reactives dyes by the bacteria Pseudomonas putida.
Materials and Methods The azo reactive dyes C.I. Reactive red 220 [Fig. 1], C.I. Reactive Blue 40 [Fig. 2], C.I. Reactive Yellow 174 [Fig. 3] were obtained from manufactory industry, SITEX, Ksar Hellal, Tunisia. O
F
SO3Na
O2S N H
O
SO3Na NH NaO3S
NH2
SO3Na
N
N
N
O
NH
NH
N
SO2
N HO
HN
NaO3S
SO3Na
O
Figure1: C.I. Reactive Red 220 [9]
SO3Na NaO3S
O
Figure 2: C.I. Reactive Blue 40 [10]
1
NaO3S
SO3Na
O HN
NH 2
NH 2 SO2
N
N
NH
CH2
O
SO3Na
NH2
F
Figure 3: C.I. Reactive Yellow 174 [9]
In the preliminary study, adapted microorganisms were isolated from sample collected in the station of treatment of textile wastewater in SITEX. After that, we tested the decolorization ability of 33 bacteria, 2 fungi and 2 yeast. Although these observations cannot give many insights into precise rate of color removals, they do give, at least partially, information on the decolorization ability of the strain tested.15 bacteria strains are selected because they exhibited excellent decolorization from 72 hours. After that, they were identified according to their respective morphological and physiological characteristics. In this study, we are interested only in the bacteria Pseudomonas putida because it is not pathogenic. According to the concentration of dyes in textile wastewater, the concentration of reactive dyes was adjusted to 80 mg/l. The pH was adjusted to 8 [11]. The temperature was maintained to 31° C. Our objective is to optimize the decolorization of reactive dyes by this bacteria. The effect of operational conditions (agitation, filamentous fungi, Floating support and light) were investigated. These factors were used in different combinations to determine the optimum condition of the biodegradation. Dye decolorization was detected by spectrophotomer. It was reported as decolorization (%) = (A0 - At / At) * 100 where A0 and At were the absorbency of the dye solution initially and at cultivation time (t), respectively. Moreover, we detected the removal of pH and DCO by colorimeter.
Results and discussions In optimum conditions when we have agitation, fungi, floating support and obscurity in the solution, this bacteria was able to decolorize 92 % of reactive Blue 40, 82 % of reactive Yellow 174 and 73 % of reactive Red 220 in 7 days. All of the processing decolorization was achieved in 7 days. The maximum rate of the biodegradation is obtained for the reactive Blue 40. In fact, this dye contains a diazo and an azo group contrary to reactive Red 220 and reactive Yellow 174 that contains only one diazo group. Thus, the bacteria have two possibility to reduce this dye. Morover, the reactive Blue 40 contains only 4 aromatic rings whereas the reactive red 220 contains 7 aromatic rings. We noticed the decolorization decrease if the number of aromatic rings increase. Finally, the number of sulfonate group is more in reactive Red 220 then the reactive Yellow 174. The reactive Red 220 is more soluble . Thus, there is decrease in the surface of contact with the bacteria and the decolorization will to slow down. The influence of any factor was determined by the method of experimental design. We calculed the average of any factor in the level 1 and in the level 2. Effect of operational conditions
80
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(b)
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90 (a)
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70 withouth agitation
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with agitation
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withouth agitation with agitation
10
%Decolorization
60 (%) Decolorization
(b)
80
70
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60
with fungi
50 40
withouth fungi with fungi
30 20
withouth fungi
10
with fungi
0
0 Reactive dyes
Figure 4: Effect of agitation on decolorization of reactive dyes
Reactive dyes
Figure 5: Effect of fungi on decolorization of reactive dyes
2
(a)
(b)
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withouth support
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with s upport
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%Decolorization
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with light
40 withouth light
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0 0
Reactive dyes
Reactive dyes
Figure 6: Effect of support on decolorization of reactive dyes
Figure 7: Effect of light on decolorization of reactive dyes
(a) reactive Yellow 174, (b) reactive Blue 40, (c) reactive Red220.
The figure 4 shows the increase of biodegradation efficiency in the presence of the agitation for reactive Yellow 174 to 10.25 % (53.87 % to 64.12 %), reactive Blue 40 to 10 % (63 % to 73 %) and reactive Red 220 to 8.4 % (47.6 % to 56 %). In fact, the development of bacteria strain was performed in the ventilated medium. The results presented in figure 5 indicate the significant effect of the presence of fungi in optimizing the biodegradation. The decolorization efficiency was increased for reactive Yellow 174 to 19 % (49.25 % to 68.25 %), reactive Blue 40 to 19.5 % (58.25 % to 77.75 %) and reactive Red 220 to 16.8 % (43.36 % to 60.23 %). In fact, Pseudomonas putida was fixed with fungi. The immobilisate bacteria have a high performance to reduce the azo bond in comparison with the free cells. Moreover, the fungi can degrade also the dye but slightly. The results from effect of presence of floating support are shown in figure 6.The percentage of the biodegradation is increased on average to 10.2 % (53.9 % to 64.1%) for reactive Yellow 174, 8.75 % for reactive Blue 40 (63.62 % to 72.37 %) and 10.4 % for reactive Red 220 (46.62% to 56.98%). Just as the fungi, the bacteria are fixed on the support. Thus, their activity increased. If we look at figure 7, we notice that the absence of light slightly increases the decolorization. In fact the influence of light is not significant for Pseudomonas putida and fungi. For the 3 dyes, the best result of the biological degradation is obtained in agitated medium, in absence of the light and in the presence of the support floating and fungi. The rate of the decolorization is high, that proves the good choice of the bacteria and the quantity of biomass that we have met in the solution. Removal of DCO 350
DCO (mg O2/L)
300 250 reactive red
200
reactive yellow
150
reactive blue
100 50 0 1
3
4
5
6
7
8
Time (day)
Figure 8: Evolution of DCO versus time
The figure 8 shows the same shape of curve for the 3 reactive dyes. Versus the 3 first days, the DCO is increased. In fact, the biological agents produced organic substance which influenced the DCO value. After that, the DCO decreased until stabilization after 7 days according to the kinetic of biodegradation. It is the result of the decomposition of reactive dyes.
3
Removal of pH 9 8,8 8,6
pH
8,4
reactive Blue
8,2
reactive Red reactive Yellow
8 7,8 7,6 7,4 1
2
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7
8
9 11
Time (day)
Figure 9: Evolution of pH versus time
During the biodegradation, pH increased for the three dyes. We noticed the appearance of chemicals compound of weak alkalinity.The highest pH is for reactive Blue. Indeed, this one has the greatest rate of decolorization by comparing at the other dyes. Moreover, this dye contains a diazo and an azo group. We have appearance of four aromatics amine (weak alkalinity) for reactive Blue Whereas one has two aromatic amines for reactive Red and reactive Yellow that contains only one diazo group.
Conclusion This work describes the biodegradation of reactive dyes by Pseudomonas putida using various techniques such as Chemical Oxygen Demand (COD), UV-spectroscopy analysis. Furthermore, this work enabled us to show the important effect of agitation, floating support and fungi to optimize the decolorization of the reactive Blue, reactive Red and reactive Yellow.
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textile dyes by newly isolated
[6] Zheng Z, Levin RE, Pinkham JL, Shetty K. Decolorization of polymeric dyes by a novel Penicillium isolate. Proc Biochem 1999;34:31–7. [7] Zhou W, Zimmermann W. Decolorization of industrial e.uents containing reactive dyes by actinomycetes. FEMS Microbiol Lett 1993;107:157–62. [8] Dilek FB, Taplamacioglu HM, Tarlan E. Color and AOX removal from pulping euents by algae. Appl Microbiol Biotechnol 1999;52:585–91. [9] Comminication de la comission au parlement europeen, Elincs 1998 23-45. [10] T. H.Wallace, Biological Reduction of a Synthetic Dye Water and an Industrial Textile Wastewater Containing Azo Dye Compound, june 2001. 7. [11] Chniti N. Contribution a l’optimisation des conditions de traitement des rejets industriels textiles par l’utilisation d’une biomasse marine acclimaté.
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