Optimization of Biosorption Performance of Casuarina Leaf Powder for the Removal of Lead Using Central Composite Design

Water is no alien to all the living beings upon earth. It has no barrier or bars over constituencies or continents, as it leaves only 1/4th of the land of whole ecosystem. The contaminated water may cause anemia, headache, chills, diarrhea and reduction in hemoglobin formation [1]. The influence of pollution on the global environment, through activities such as rapid industrialization, urbanization and mining operations often lead to an increase in the discharge of toxic metals, such as lead, nickel, chromium, cobalt, copper, cadmium etc., into the environment, which results in a contamination of water. These heavy metal contaminants are hazardous to the environment, because they do not naturally degrade [2]. Beyond certain limits, heavy metals are toxic to living organisms and may cause serious hazard to public health [3]. Environmental engineers and scientists are faced with the challenging task to develop appropriate low cost technologies for effluent treatment [4].


Introduction
Water is no alien to all the living beings upon earth. It has no barrier or bars over constituencies or continents, as it leaves only 1/4 th of the land of whole ecosystem. The contaminated water may cause anemia, headache, chills, diarrhea and reduction in hemoglobin formation [1]. The influence of pollution on the global environment, through activities such as rapid industrialization, urbanization and mining operations often lead to an increase in the discharge of toxic metals, such as lead, nickel, chromium, cobalt, copper, cadmium etc., into the environment, which results in a contamination of water. These heavy metal contaminants are hazardous to the environment, because they do not naturally degrade [2]. Beyond certain limits, heavy metals are toxic to living organisms and may cause serious hazard to public health [3]. Environmental engineers and scientists are faced with the challenging task to develop appropriate low cost technologies for effluent treatment [4].
Traditional methods for removal of lead ions from solution include chemical precipitation, ion exchange, electrodialysis and membrane separations. All these methods have various disadvantages, specifically, high capital investment and operating cost, incomplete removal, low selectivity and high energy consumption. Therefore, there is a need for a cost effective treatment method that is capable of removing low concentrations of lead from solution For the last decades, biosorption or sorption of contaminants by sorbents of natural origin has gained important credibility due to the good performance and low cost of these complexing materials [5][6][7][8][9][10]. A multitude of biomass types comprising fungal biomass, bacterial biomass, algae, peat etc., have been studied for their biosorption of metals [11][12][13][14]. Agricultural wastes such as tree bark, peanut skin, hull, tobacco, tomato root tissues and plants waste have been used to remove heavy metals from water [15][16][17].
In view of the above, the authors tried to use a novel biosorbent Casuarina leaf powder to remove lead from aqueous solutions and report the application of Response Surface Methodology using Central Composite Design to develop a mathematical model and predict the response and check the adequacy of the model. The objectives of the present study include identifying the maximum lead uptake capacity of the Casuarina leaves powder with respect to various governing parameters. In addition, biosorption isotherms were described by using Langmuir, Freundlich and Temkin models and the kinetic experimental data were correlated by the pseudo first and second order kinetic models. Thermodynamics for biosorption of lead on Casuarina leaves powder is also studied and fitted in to Van't Hoff equation.

Preparation of metal solutions
Test solutions containing lead ions were prepared from Pb (NO 3 ) 2 at different concentrations ranging from 25, 50, 100, 150, 200, 250 mg/L. The pH of each test solution was adjusted to the appropriate value by using 0.1 N HNO 3 or 0.1 N NaOH solutions. All the chemicals used in preparing the stocks are of analytical grade and the water used is double distilled water prepared from Millipore ELIX-10 unit.

Process optimization
Final experimental runs for optimization were obtained through Response Surface Methodology using Central Composite Design with 2 4 factorial runs, 6-central points and 8-axial points. Design of Experiments (DoE) obtained based on the above optimization technique using STATISTICA software. The extent of biosorption of lead calculated at the preliminary optimum conditions is verified with the final runs for the optimum conditions. For the optimization of biosorption, the regression equation is written in terms of % biosorption of lead (Y) as function of the parameters having greater influence over the response i.e. pH (X 1 ), Co (X 2 ), w (X 3 ), and T (X 4 ). Based on experimental runs and predicted values proposed by CCD design, the following equation represents multiple regression analysis of the experimental data for the biosorption of lead:

Effect of agitation time
Duration of equilibrium biosorption is defined as the time required for heavy metal concentration to reach a constant value during biosorption. The equilibrium agitation time is determined by plotting the % biosorption of lead against agitation time as shown in figure 1 for the interaction time intervals between 1 to 180 min. For 63 μm size of 10 g/L biosorbent dosage mixed in 50 mL of aqueous solution (C o =20 mg/L), 50.4% of lead is biosorbed in the first one minute and reached to 55.7% after 5 minutes of biosorption. The % biosorption is increased briskly up to 60 min reaching 86.5%. Beyond 60 min, the % of biosorption is constant indicating the attainment of equilibrium conditions. The maximum biosorption of 86.5% is attained for 60 min of agitation time. The rate of biosorption is fast in the initial stages because adequate surface area of the biosorbent is available for the biosorption of lead. As time increases, more amount of lead gets biosorbed onto the surface of the biosorbent due to Vanderwaal's forces of attraction and resulted in decrease of available surface area. The biosorbate, normally, forms a thin one molecule thick layer over the surface. When this monomolecular layer covers the surface, the biosorbent capacity is exhausted. Therefore, all other experiments are conducted at this agitation time.

Effect of biosorbent size
The variations in % biosorption of lead from the aqueous solution with biosorbent size are drawn in figure 2 with percentage biosorption of lead as a function of biosorbent size. The percentage of biosorption is decreased from 86.9% to 80.2% as the biosorbent size increases from 63 to 105 μm. This phenomenon is expected, as the size of the particle decreases, surface area of the biosorbent increases; thereby the number of active sites on the biosorbent also increases.

Effect of temperature
The effect of temperature on the equilibrium metal uptake was significant. The effect of changes in the temperature on the lead uptake is shown in figure 6. Lead uptake marginally increased from 92.2 to 94.6% with increasing temperature from 283 K to 323 K indicating that the biosorption of lead on to Casuarina leaf powder is endothermic process. Adsorption processes are normally exothermic and as the temperature increases the % adsorption decreases in accordance with Le Chatelier principle. The reverse phenomena could be activation of non living biomass under moderate temperatures and Increasing the temperature is known to increase the rate of diffusion of the adsorbate molecules across the external boundary layer and in the internal pores of the adsorbent particles, owing to decrease in the viscosity of the solution.

Effect of pH
Biosorption is controlled by pH by the influence of surface charge of the biosorbent, the degree of ionization and the species of biosorbate. In the present investigation, effect of pH on lead biosorption is obtained in the pH range of 2 to 8 of the aqueous solution (C o =20 mg/L) using 10 g/L of 63 μm size biosorbent. The effect of pH of aqueous solution on % biosorption of lead is shown in figure 3. The % biosorption of lead is increased from 64.6% to 91.4% as pH is increased from 2 to 5 and decreased beyond the pH value of 6. Percentage of biosorption is decreased to 62.1% from 84.3% for pH variation of 6 to 8. Since the pH of aqueous solution influences the solution chemistry of the heavy metals, the binding of metal ions by surface functional groups is strongly pH dependent. The increase in % removal when pH increases from 2 to 5 could be due to decrease in competition between hydrogen ions and metal species for appropriate sites on the biosorbent surface and also by the decrease in positive surface charge on the adsorbent. However, with increasing pH above 5 lead tends to hydrolyze and precipitate instead of adsorption and adsorbent was deteriorated with accumulation of metal ions, making the true adsorption studies unpredictable.

Effect of initial concentration of lead
The effect of % biosorption on initial concentration of lead in the aqueous solution is shown in figure 4. The percentage biosorption of lead is increased from 73.3% to 90.1% with decrease in C 0 from 150 mg/L to 20 mg/L. Such behavior can be attributed to the increase in the amount of biosorbate to the unchanging number of available active sites on the biosorbent.

Effect of biosorbent dosage
The percentage biosorption of lead is drawn against biosorbent dosage for 63 μm size biosorbent in figure 5. The biosorption of lead increased from 87.2% to 93.4% with an increase in biosorbent dosage from 5 to 35 g/L. Such behavior is obvious because with an increase in biosorbent dosage, the number of active sites available for lead biosorption would be more. The change in percentage biosorption of lead is marginal from 93.4% to 93.8% when 'w' is increased from 35 to 50 g/L. Hence all other experiments are conducted at 35 g/L dosage.

Kinetics
The order of biosorbate -biosorbent interactions have been described using kinetic model. Traditionally, the first order model of Lagergren finds wide application. In the case of biosorption preceded by diffusion through a boundary, the kinetics in most cases follows the first order rate equation of Lagergren: (dq t /dt)= K ad (q e -q t ) Where, q e and q t are the amounts adsorbed at t, min and equilibrium time and K ad is the rate constant of the lagergren first order biosorption.
The above equation can be presented as ∫ (dq t / (q e -q t )) = ∫ K ad dt Applying the initial condition q t = 0 at t = 0, we get log (q e -q t ) = log q e -(K ad /2.303) t Plot of log (q e -q t ) versus't' gives a straight line for first order kinetics, facilitating the computation of adsorption rate constant (K ad ).
The pseudo second order kinetic equation given below: (dq t /dt) = K (q e -q t )2 Where, 'K' is the second order rate constant.
Substituting these values in above equation, we obtain: 1/ (q e -q t ) = Kt + (1/q e ) Rearranging the terms, we get the linear form as: (t/q t ) = (1/ Kq e 2 ) + (1/q e ) t (12) The pseudo second order model based on above equation, considers the rate -limiting step as the formation of chemisorptive bond involving sharing or exchange of electrons between the biosorbate and biosorbent. If the pseudo second order kinetics is applicable, the plot of (t/q t ) versus't' gives a linear relationship that allows computation of q e and K. The first order and the second order kinetics plots are given in figures 10 and 11 respectively. The rate equations obtained and comparative lead uptake capacities are given in table 2 and table 3 respectively. The correlation coefficients indicate that the system under consideration is more appropriately described by pseudo-second order model. The regression coefficient of 0.9897 shows that that the model can be applied for the entire adsorption process. The confirmation of pseudo second order kinetics indicates that in the adsorption process, concentrations of both adsorbent and adsorbate are involved in rate determining step. In the range of studied parameters, the metal uptake is very good for Casuarina leaf powder.

Thermodynamics of biosorption
Biosorption is temperature dependant. In general, the temperature dependence is associated with three thermodynamic parameters namely change in enthalpy of biosorption (∆H), change in entropy of biosorption (∆S) and change in Gibbs free energy (∆G). Enthalpy is the most commonly used thermodynamic function due to its practical significance. The negative value of ∆H will indicate the exothermic/ endothermic nature of biosorption and the physical/chemical in nature of sorption. It can be easily reversed by supplying the heat equal to calculated ∆H.
The ∆H is related to ∆G and ∆S as log (q e /C e ) = -0.40566 (1 / T) + 1.029489 (15) Where, (q e /C e ) is called the biosorption affinity.
If the value of ∆S is less than zero, it indicates that the process is highly reversible. If ∆S is more than or equal to zero, it indicates the reversibility of process. The negative value for ∆G indicates the spontaneity of biosorption, whereas the positive value indicates non spontaneity of sorption. Experiments are conducted to understand the biosorption behavior varying the temperature from 283 to 323 K. The Van't Hoff's plot for the biosorption data obtained is shown in figure  12. From the plot the equation obtained is log (q e /C e ) = -0.40566 (1 / T) + 1.029489. The values obtained from the equation are ∆G = -5964.899, ∆H = 7.7672 and ∆S = 19.71177. ∆H is positive indicating that the biosorption is endothermic. The negative value of ∆G indicates the spontaneity of biosorption. As ∆S is more than zero, it indicates the irreversibility of biosorption.

Optimization using Response Surface Methodology (RSM)
In order to determine an optimum condition for Pb ions removal, the parameters having greater influence over the response is to be identified. In the present study, the relationship between four independent variables and percent of Pb ions biosorption fitted well with the quadratic model. Levels of different process variables in coded and un-coded form (Table 4), Results from CCD ( Table 5), Analysis of variance (ANOVA) ( Table 6), Estimated regression coefficients ( Table 7) and Comparison of optimum values obtained from CCD and experimentation (Table 8)   First order log (q e -q t )=-0.019698 t-0.073348 0.04513 0.9507 Second order t/q t =0.5935 t + 1.85514 0.18987 0.9897   -0.008 X 1 X 2 + 0.022 X 1 X 3 + 0.000 X 1 X 4 + 0.037 X 2 X 3 + 0.001 X 2 X 4 + 0.000 From the Central Composite Design, the Pareto Chart (Figure 13), Observed Vs Predicted values plot ( Figure 14) and Surface contour plot (Figures 15 a-f) for biosorption of lead on to Casuarina leaf powder are presented below. The optimal set of conditions for maximum percentage biosorption of lead is pH= 4.9884 biosorption dosage (w) =35.3702 g/L, initial lead concentration (Co) =18.0555 mg/L and temperature=306.4727 K and the % of biosorption calculated at these values found to be 95.73%.

Conclusions
The analysis of the experimental and theoretical data resulted that the equilibrium agitation time for lead biosorption is 60 minutes. The % removal of lead from an aqueous solution increases with a decrease in the particle size of the biosorbent and increases with increase in biosorbent dosage. With an increase in the initial concentration of             lead in the aqueous solution, the percentage removal of lead from the aqueous solution is decreased. Percentage removal of lead from the aqueous solution increases significantly with increase in pH from 2 to 5, thereafter percentage removal decreases for further increase in pH. In the range of variables studied, percentage removal is increased from 50.44% to 93.9%. The maximum uptake capacity of 15.0797 mg/g is obtained at temperature 303 K. The kinetic studies show that the biosorption of lead is better described by pseudo second order kinetics It is also found that the data are well represented by Langmuir isotherm with higher correlation coefficient of 0.9944, followed by Freundlich and Temkin isotherms. The biosorption of lead on to Casuarina leaf powder is irreversible, spontaneous and endothermic in nature. The present study involves the use of statistical experimental design to optimize process conditions for maximal biosorption of lead from aqueous solution using CCD involving RSM. The maximum biosorption of lead (95.73%) onto Casuarina leaf powder is observed when the processing parameters are set as follows: pH = 4.988, w = 35.37 g/L, C o =18.0555 mg/L and T=306.47 K. Therefore the above said Casuarina leaf powder is effective and efficient biosorbent and is capable of removing lead.