BIOSORPTION OF PB ( II ) , CR ( III ) AND CD ( II ) FROM SYNTHETIC WASTEWATER USING DRIED RAW WASTE ACTIVATED SLUDGE BY RESPONSE SURFACE METHODOLOGY

In this study the dried raw waste activated sludge (DRWAS) was used as a biosorbent material to remove Pb(II), Cd(II) and Cr(III) from simulated wastewater with different operating conditions to optimize the heavy metal removal process by statically analyzed tool was used to optimized the removal efficiency according to change operating conditions using response surface methodology (RSM). The ranges of these parameters were initial metals concentration 10–50 mg/L, pH 2–5, contact time 10–240 min, temperature 25-40 °C, Agitation speed 50-200 rpm, and biomass loading 0.5-6 g/L. Removal efficiency model was obtaining with high significant and low p-value with predication values of 99%, 85% and 78% for Pb(II), Cr(III) and Cd(II) respectively. The study clearly showed that DRWAS was promising low cost biosorbent to remove heavy metals.


Introduction
Detoxification of water sources is main environmental and human health issue which resulting generally in most development countries [1].Most of heavy metal consider as contaminants to environment and human health.These pollutants have the ability to accumulate in food chain and finally reaching to human tissue [2,3].Heavy metals can change the needed ions in living cells if they pass in the human tissue and can effect in different ways on the normal functions, which cause dangerous carcinogenic effect to human beings and other living organisms.Furthermore, heavy metal ions accumulation has been detected to harm liver, brain, and kidney [4,5,6].Lead, chromium and cadmium ions found in different wastewater of industrial disposal depending on the type of industry.These metals consider important pollutants that should be treated well before releasing to water or soli environments [4].According to the World Health Organization (WHO), the maximum allowable concentration of Cr (III), Cd(II) and Pb(II) in water bodies (lake, river and stream channel) are 0.1, 0.01, and 0.05 mg/L, respectively [2].
Many techniques and conventional treatment methods have been used to remove heavy metal ions from different industries released wastewater.The most common methods are membrane separation, ion exchange, evaporation, precipitation, electroplating [7,8].Nevertheless, these treatment technologies have undesirable effects and require complicated technical setup and high operating cost and need additional treatment to disposal sludge released from these treatment methods [9,10].The specific disadvantage of these method is highly expensive (raw material cost, maintenance and large amount of disposal sludge, large area needed for installed unites) and not work properly when the heavy metal ions were low concentration in high volume quantity of wastewater released from these activities.Therefore, there is needing of a new costeffective and eco-friendly means which can eliminate heavy metal ions from the aqueous solution [11].
Adsorption of heavy metals (biosorption or bioremoval) onto low cost biosorbent takes the advantage in the recent time.The main advantage of this type of treatment is eco-friendly and economy (cheap biosorbent price), excellent efficiency of elimination pollutants, short time required for removal, small area need to install unites [12].More and more consideration has been rewarded to biosorption using low cost biomass, such as by-product waste, agricultural waste and microorganism biomass.Furthermore, it is an economic and desirable process which candidate this method to distinct an extensive range of pollutants from industrial effluents [13].
Bacterial biomass considered easy and effectively low cost biosorbent materials with excellent settling properties and due to its cell wall characterization that contains many extracellular polymeric substances, with many types of functional charged groups such as phosphoric, amine, carboxyl, and hydroxyl groups [2][3][4][5][6].The main mechanisms achieved in biosorption of heavy metal is the attraction between the heavy metal positively charged and cell wall, which is namely physical removal.In the other hand there is complexation mechanism between these metal ions and different charge type of functional groups [6].The biosorption mechanism process is powerfully affected by many factors, including the initial adsorbate concentration, initial pH of the solution, temperature, dosage of the adsorbent, contact time, etc .which effecting on the sensitivity of the process [2][3][4][5][6].
In order to increase the efficiency of the biosorption process at a minimum time and cost, the response surface methodology (RSM) is usually applied as statistical tools for optimization study, increases the accuracy percentage and principal cost decreases experimental time suitable in designing experiments, improving complex processes; predicting and estimating the effect of associated variables with minimum experimental runs.The aim of this paper was to investigate the performance and ability of DRWAS as a biosorbent in the elimination of Pb(II), Cr(III) and Cd(II) from synthetic wastewater.In order to improve results.RSM was applied to achieve a well consideration of the relationship among the operating parameters and also to define the best conditions in Cd(II), Cr(III) and Pb(II) removal.

Adsorbent
The Biomass used in this study was collected from the raw waste activated sludge from Al-Rustomia wastewater treatment plant.This biomass was containing of homegenoues microorganism mainly from bacteria (E-coli , Pseudomonas aeruginosa, Saccharomyces cerevisiae, Clostridium, and Salmonella sp.) some fungi (penicillium sp.) , the collected biomass followed by an oven dried (Hamilton, UK) at 80 ± 3°C until constant weight was obtain from two sequence weights measurement, then the dried raw waste activated sludge (DRWAS) was crashed, sieved, washed three times with deionized water to remove any impurities and kept a desiccator for the exploitation in batch experiments in this study.the physicochemical characterization of DRWAS listed in Table 1.

Adsorbate
All chemicals used in this experimental work were analytically pure and grade.Chromium nitrate, 97% (BHD), Cadmium nitrate tetrahydrate, 98% (merck) and lead nitrate 99% (BHD) were purchased from local market.A stock solution of 1000 mg/L was prepared from dissolving 0.1599, 4.577, and 2.103g of Pb(NO 3 ) 2 , Cr(NO 3 ) 3 and Cd(NO 3 ) 2 in one liter deionized water, then desired concentration of these heavy metal solution was obtained using a flame atomic absorption spectrophotometer Buck, Accusys 211, USA.The characteristics of these pollutants were summarized in Table 2.

Methodology
The pH of heavy metal prepared solution was measured by pH meter (WTW, Germany) and adjusted to desired amount pH using NaOH (0.01 M) and HNO 3 (0.01 M) as buffer solution, the amount of DRWAS need for each run was added into 250 mL conical flask contained 100 mL of Pb(II), Cd(II) or Cr(II) solution then shaking using shaker incubator (Labtech, Koria) with different times according to RSM analysis tool program runs.Samples were then taking at the end of each experiment, filtered by filter paper (Whatman 42 pore, Germany).One drop of HNO 3 (0.01 M) added to separated solution.The concentrations of Pb(II), Cr(III) and Cd(II) before and after adsorption were analyzed according to a standard method [4] using atomic spectrophotometer AA.The removal efficiency (R, %) of Pb(II), Cr(III) and Cd(II) from the solution was obtained using Eq.(1).while the experimental uptake (q e , mg/g) was found using Eq. (2). (1) Where the C 0 , and C e are the initial and final equilibrium concentration (mg/L), respectively, q e is the experimental uptake of heavy metal solution onto DRWAS, mg/g, V is the volume of heavy metal solution, L, and w is the biomass weight in g.

Response surface methodology
There are many methods to optimized the processes such as box behnken design (BBD), face centered design (FCD) and central composite design (CCD).among of these methods the CCD was applied due to its simple structure and good efficiency.RSM consists of a group of mathematical and statistical techniques based on the fit of a polynomial equation to the experimental data in order to predict the behavior of a system and to reduce time and costs.
In this study, the biosorption process of Pb(II), Cd(II) and Cr (III) by DRWAS was optimized with a minimum number of runs.Six of the factors as mentioned in Table 3 affecting the adsorption process were designated to predict their influences on the removal efficiency (Y) of Pb(II), Cd(II) and Cr (III).
The experimental of Pb(II), Cd(II) and Cr (III) removal efficiency and its variation with these six factors are listed in Table 3.A second order polynomial equation Eq. ( 3) was used to fit the data, using the least square method.To evaluate the precision of the model and the significance of the variables, analysis of variance (ANOVA) was performed.
Where, Y is the predicted response, x is the independent variable β 0 , β i , β ii , β ij represent constant, linear, quadratic and interaction coefficients respectively.While ANOVA used to study the model if it would be acceptable using F-value and p-value results.P > F less than 0.05 was regarded as significant.

Results and Discussions
RSM, CCD analyzed method were done by Design Expert 7.0 (trial version), for optimization of significant parameters under the study by twenty-two experiments to perform the locate of maximum removal, with six duplicates experiments in the center point (the lowest and highest values was marked as the center point 0) to obtain the repeatability of the method and to determine the experimental error.
Determination of the effects of main factors and boundary on Pb(II), Cd(II) and Cr(III) onto DRWAS removal efficiency was demonstrated in Table 4. Furthermore, Fig. 1 shows the investigation on the means that clearly identifying that the solution pH, heavy metal concentration and biomass concentration played the major role in biosorption processes.
While minor effects were found due to temperature, agitation speed in bioremoval efficiencies of metal ions.The model equation for actual factors values of the quadratic model fitting the experimental results is illustrated in Eq. (4) to Eq. ( 6).
The quadratic equation was obtained by statistically significance the analysis of variance to find best model represented the experimental bioremoval data of Pb(II), Cd(II) and Cr(III) as shown in Table 5.The values of the coefficient of determination shows that 91%, 93% and 96% of the variability in the response are explained by the model for Cd(II), Cr(III) and Pb(II), respectively, onto DRWAS biomass.
The optimal biosorption conditions for removal of Hg(II) were found to be that temperature 33.5C, pH value 3.5, biomass loading 3.25 g/L, agitation speed 125 rpm, initial metal concentration 50 mg/L and sorption time =125min.Under these optimal conditions, the maximum uptake amount and removal yield were calculated by the quadratic.Table 6 ANOVA shows high F value 20.07, 13.15, and 11.11 for Pb(II), Cr(III) and Cd(II) with a low probability value P < 0.0001 implies high significance of the model.
The most significant parameters effect on Pb(II) and Cr(III) bioremoval onto DRWAS were pH, contact time, biomass loading and agitation speed.While for Cd(II) the pH, contact time and biomass loading these differences in biosorption depended on the attractive between these heavy metal ions associated with different functional groups charge on bacterial surface wall such as -OH, -NH, C-H, C-Cl as proved by M-Ridha [6].From this figure the relationship between these metal ions indicated that the Pb(II) was dominated metals in comparison with Cd(II) and Cr(II), while Cr(III) was the dominated on Cd(II), these results caused by the physicochemical properties of heavy metal ions due to the low solubility of Pb(II) < Cr(III) < Cd(II) with amount of 52, 81 and 136 g/ 100 ml at 20 °C, respectively, and high molecular diffusion of Pb(II) > Cr(III) > Cd(II) with amount of 4.98, 4.22 and 4.2 ×10 -8 m 2 /s, respectively.

Conclusions
This study shows that the DRWAS can be used to remove Pb(II), Cd(II), and Cr(III) from aqueous solution phase of high concentration of 50 mg/L.Based on the statistical analysis using response surface methodology, the optimum conditions for 99%, 85%, and 78% could be obtained for Pb(II), Cr(III) and Cd(II), respectively, when the experimental condition fixed on temperature 40°C, biomass loading 6 g/L, initial metal concentration 10 mg/l and contact time 240 min.the RSM and CCD predicted best model to estimate removal efficiencies of these metals with high significant with low Pvalue less than 0.0001.based on results the DRWAS is promising biomass for removal heavy metal ions with high removal efficiency.

Figure 1 .
Figure 1.Actual and predicted values for Pb(II), Cd(II) and Cr(III) for removal efficiency onto DRWAS

Table 1 .
Main physicochemical characteristic of DRWAS tested

Table 2 .
Main physicochemical characteristic of the Pb(II), Cr(III) and Cd(II) tested

Table 3 .
Factors and levels used in the minimum run resolution v characterization design used to optimized biosorption process using DRWAS.

Table 4 .
Minimum run resolution v characterization design for determination of the most important variables affecting the metal ions removal onto DRWAS.

Table 6 .
Analysis of variance F value and P rob >F for Pb(II), Cd(II) and Cr(III) onto DRWAS removal efficiencies.