Biological Treatment of Reactive Dye in Textile Industry

Introduction
The decolonization of wastewater is still a major environmental concern. Synthetic dyes used in textile industry, are difficult to be removed by conventional waste water treatment systems based on adsorption and aerobic bio degradation. In recent year to solve environmental problems, utilization of microbial flocculants has been anticipated due to biodegradability and harmlessness of their degradative intermediates. A reactive dye is prominent among numerous groups of water-soluble dyes. Pronounced colorization of waste waters is the result of their relatively high solubility in water. Decontamination of these polluted wastewaters requires high effectiveness and low cost removal processes of dyes . Chemical and physical methods including adsorption, coagulation–flocculation, advanced oxidation and electrochemical methods are very efficient in color removal.

These methods are quite expensive and have pose operational problems. Regeneration requirement sand the cost of adsorbents make adsorption an unattractive method for decolonization purpose . However, recent reports indicated the possibility of using some natural or low cost adsorbents such as wood ash, soil, and powdered activated sludge for color removal single biological treatments, the most economical and environmentally friendly ones, are not a suitable alternative when working with toxic and/or non-biodegradable wastewater In fact, most of disposed dyes are of non-biodegradable nature and standard biological treatment of their colored effluents is not effective. The synthetic dyes can be classified as disperse dyes, reactive dyes, sulfur dyes, vat dyes, direct dyes, acid dyes, and basic dye. They are commonly used in the textile industries. Reactive dyes are now important in coloring cellulosic and wool fibers. During the coloration process, reactive dyes combine covalently with the fibers through nucleophilic displacement. The reactive dyes-containing effluents from these industries have caused serious environment pollution because the presence of dyes in water is highly visible and affects their transparency and aesthetics even if the concentration of the dyes is low . Cibacron yellow FN_2R is one of the main dyes that are used in textile industries in Malaysia. Therefore, industrial effluents containing dyes must be treated before their discharge into the environment. Biodegradation using microorganisms is gaining importance as it is cost effective, environmentally friendly, and produces le s sludge. The application of SBR to color removal is rather a new approach compared to anaerobic–aerobic sequential treatment. In the study by Ghoreishi and Haghighi, the COD removal efficiencies were in the range of 76–83 in a combined biological-reduction process. Sequencing batch reactor (SBR) as a modified has been used for many industrial wastewaters such as fiber and dyes wastewater. Main advantages of this system are low build cost, high flexibility and low required space . Disadvantages of this system are high excess sludge production and high SVI index. This paper aims to examine the best performance culture conditions for the production of a bioflocculant by Sphingomonas paucimobilis using dye wastewaters as feeding substrate. Employing reactive dye by spinmongunas bacteria at sequence batch reactor performance is a novel approach for dye removal.

Materials and Methods

Microbial Culture:
One loop, full of bio-flocculants producing bacteria isolated from Closed Drainage System, (CDS) and grown on slant agar at 35°C for 2 days in oven, and . Flasks are under static conditions at its optimum growth temperature, T=35°C.

Bioflocculant-producing Medium and Culture Conditions:
Much type of bioflocculant–producing strains was isolated from Closed Drainage System (CDS) perai wase water samples by using kaolin clay solution as test material. The composition of the basic culture medium for the production of bioflocculant Polyglutamic Acid (PGA) was used as a medium for cultivation of bioflocculant producing bacteria. Compositions of medium were as follows 20(g/L) glucose; L-glutamic acid, 50g/L; yeast extract, 0.5; magnesium sulphate heptahydrate (MgSO4.7H2O), 0.5; bacteriological agar, 15. The initial pH of media was adjusted to 7.0–7.2 using 1.0 M sodium hydroxide (NaOH) and 1.0 Ml hydrochloric acid (HCl).

Determination of the Flocculating Activity:
The flocculating rate was measured using a previous method with a slight modification, in which Kaolin clay was chosen as the suspended solid by Kurane Kaolin clay was suspended in distilled water at the concentration of 5000 mg/L. 4.50 mL of 1% CaCl and 0.5 mL of culture broth were added to 45 mL Kaolin suspended solution in 100 mL beaker in turn. The mixture was vigorously stirred and was allowed to stand for 5 minutes. The optical density (OD) of the clarifying solution (A) was measured with a spectrophotometer at 550 nm. A control experiment was prepared using the same method, but the culture broth was replaced by distilled water (B). The flocculating activity was calculated according to the equation.

Flocculating efficiency (%) = {(B – A)/B} × 100

Where:
A is the optical density of the sample experiment at 550 nm
B is the optical density of control experiment at 550 nm
Effects of pH, flocculant's concentration and various actions on flocculation were investigated.

Media Composition:
The synthetic dye-containing wastewater was used in this study. Seed culture medium: consist of distilled water 1000 mL, the composition of the wastewater was powdered as main carbon source, urea as nitrogen source, K2HPO4 and KH2PO4 as phosphorous sources and the nutrients were also limited. COD: N: P, 500: 25: 5.

Preparation of Dye Waste Water:
Reactive Cibacron FN-2R was used in different concentrations. The dye is one of the main dyes that are used in textile industries in Malaysia. Dye concentration was used to prepare dye solution of concentration 50, 100, 150 and 200 mg/L in reactors R1, R2, R3 and R4, as synthetic dye waste water.

Experimental Set-up:
In this study, four cylindrical Plexiglas reactors with 14 cm diameter and 46 cm height were used
(Fig.1). The working volume and influent flow rate were 3L and 3.0 L/d, respectively. Four air pumps and four mixers were used for continuous aeration and mixing. The speed of impeller was adjusted at 300 rpm, Do>2 ppm Operation cycle of reactors was 24 hours including 2 min for filling, 22.5 hours for aeration, 1-2 hour for settling, 2 min for discharging and 30min for filling time. After acclimatization period, dye concentration was adjusted 50, 100, 150 and 200 mg/L in reactors R1, R2, R3 and R4, and we add 2 cc of bacteria (Sphingomonas paucimobilis) to each reactors and pH adjustment to pH 7. After magnetic stirring at constant temperature (35°C) for a certain period. 

Fig. 1: Four SBR reactors used in this study
The chemical oxygen demand (COD), dye absorbance ratio, mixed liquor suspended solids (MLSS), mixed liquor volatile solids (MLVSS) of samples were determined using standard methods for examination of water and wastewater.

Color Removal:
The dye solutions' concentrations were measured at a wavelength corresponding the maximum absorbance, ë max by means of a invisible spectrophotometer (Shimadzu UV-160A). The percentage of color removal was calculated by comparing the absorbance value of the supernatant to the standard curve obtained by a known dye concentration. The dye solutions' pH was measured by the pH meter. The dye concentrations were determined through color point (pt/Co) measurement by Hatch DR/2000 Spectrophotometer. The percentage of dye removal was calculated by the following equation:

Dye removal (%) = [(Cr-Ct)/Cr] *100

Where Cr and Ct are the dye concentration in raw and treated solutions.

Result & Discussion

Bioflocculant-producing Properties:
The bacteria was in the highest flocculation of Kaolin suspension attained was 98%. The same phenomenon happens where the flocculating activity paralleled the growth of the microorganism.

Effect of Temperature on Flocculating Activity:
Effect of temperature on flocculating capability of bioflocculant-producing bacteria was determined at different incubation temperatures. Figure 2 shows that flocculation rate of bioflocculant bacteria at different incubation temperatures and various incubation periods. that sample were cultivated at temperatures between 30 C-50 C for 43-77 hours Isolated bacteria from waste water and 0 0 sediment of CDS, Prai showed that the highest flocculation rate at 350C with 98.4% and Fig2 shows that flocculation rate decreased to 55% when incubation temperature raised to 470C. The decreasing in flocculation rate could be due to the thermal effects as reported by (Wuj et al., 2007). 
 
Fig. 2: flocculation rates of bioflocculant –producing bacteria at various incubation temperatures
Effect of Dye Concentration:
Variation of dye removal efficiencies of all reactors are shown in Fig. 3. The minimum dye removal efficiency was obtained in R4 with the dye concentration of 200 mg/L. Maximum dye removal efficiency was obtained in R1 with dye concentration of 50mg/L. 
 
Fig. 3: Variation of dye removal efficiency in the SBR reactors
Effect of HRT on COD and MLVSS:
Determination of COD and MLVSS in different concentration dye (Fig 4) show that during period as time pass, we have decrease in COD and increase in MLVSS (HRT =24h). The results of this study demonstrate that the newly developed treatment technique decrease color, biochemical oxygen demand (BOD), chemical oxygen demand (COD) and increase MLVSS. This shows that bacteria growth during the time. 
 
Fig. 4: Effect of HRT on COD and MLVSS at different concentration
COD Removal Efficiency:
The variation of COD removal efficiencies are shown in Fig 5. The average removal efficiencies were almost in the same range of 97 percent and COD removal efficiencies increased from 95% to 98% during the normal operation period in all reactors. Maximum COD removal efficiency was obtained in the reactor 4 with the dye concentration of 200 mg/L. No significant influence on COD removal efficiency was observed by altering the dye concentration. 
 
Fig. 5: Variation of COD removal efficiencies of SBRs
Conclusion:
A bioflocculant produced by Sphingomonas paucimobilis was effective for the removal of dye, reactive waste water and reactive dyes are more frequently used in dyeing processes due to their good solubility in water. Various culture temperature were tested for 43-77 hours in order to investigate their effect on the bioflocculant production when the culture temperature was 35 C which 0 the flocculating activity of Sphingomonas paucimobilis was up to 98.4% determination flocculating activity, dye removal efficiency COD removal was shown Sphingomonas paucimobilis is biodegradable and increase in number of bacteria during  the time will confirm that. The results show that the optimal flocculant quantity has to be used. In this study, dye removal efficiency was in the range of 31% to 67% and SBR system showed low removal efficiency for the reactive dye, COD removal efficiency of 97.30% was obtained. In our study SBR reactors under similar organic loading rate was used. No significance influence of dye on COD removal was observed. This study was using sequencing batch reactor (SBR) technology different concentration dye waste water between every day (HRT=1day) bacteria growth and produce biopolymer every day and decrease COD, increase MLVSS. 
 
 

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