Functionalized C , N-Bipyrazole Receptor Grafted onto Silica Surface for Arsenic Adsorption and Its Antibacterial Activity

The present study is based on the investigation of performance of C,N-bipyrazole receptor grafted onto silica surface (SG2P) of adsorption Arsenic (AS) from aqueous solutions. The effects of operating parameters that include pH, contact time, concentration of As and dosage of adsorbent on adsorption were accomplished. The results clearly showed that the removal efficiency of As was decreased with an increasing of As concentration, pH, and temperature, while it was continuously increasing with time and adsorbent dose. Moreover, the removal efficiency of Cr (VI) adsorption was 75% corresponding to pH; temperature ( ̊C), initial concentration (ppm) and weight of dose (g) were 6, 25, and 0.04 respectively at 24 hours. The adsorption capacity of the synthesized sorbent (SG2P) for arsenic at pH < 7 from pseudo second order in batch experiments was efficient at 20 minutes. Furthermore, the results obtained in this study presented that the antimicrobial activity was limited where the Escherichia coli (ATCC25922) and Staphylococcus aureus (ATCC25932) were as a reference strains, while, the SG2P was able to inhibit growth only at high concentration (MIC = 1.5625 mg/ml).


Introduction
Arsenic (As) is the 20 th most abundant element present in the earth's crust; it is widespread in oxygenated water while As(III) is found in anaerobic conditions like in well water or in groundwater.The concentration of arsenic species is mainly dependent on redox potentials [2] and pH [3].
Recently, the environmental fate and behavior of arsenic (As) are receiving increased attention due to the arsenic (As) pollution in groundwater.It has been a serious health threat to the human beings in the world.Occurrence of arsenic in groundwater above the permissible value (>10 µ g/L WHO) is one of the wide spread problem owing to its toxicity and carcinogenicity [4].In natural waters arsenic normally occurs in the oxidation states +III (arsenite) and +V (arsenate).The removal of As(III) is more difficult than the removal of As(V).Thus, As(III) has to be oxidized to As(V) prior to its removal.In this context, Arsenic is considered as a toxic metal which is toxic to living organisms and harmful to the environment when it is directly discharged into water body.Arsenic can be easily absorbed by fishes and vegetables due to its high solubility in the aquatic environment and may be accumulate in the human body by the means of food chain.So, it is indispensable to treat water and/or wastewater containing toxic metals like arsenic to remove their adverse effects on human and ecology.Recently several methods of treating water for arsenic reduction are being investigated; such as ion-exchange, adsorption, chemical precipitation, membrane filtration, and flocculation and electrochemical methods [5].Adsorption is a fundamental process in the physicochemical treatment of municipal wastewaters, a treatment which can economically meet today's higher effluent standards and water reuse requirements.Also, it considered as a broad spectrum of physical, biological, and chemical processes and operations in the environmental field.Purification of gases by adsorption has played a major role in air pollution control, and adsorption of dissolved impurities from solution has been widely employed for water purification.Adsorption is now viewed as a superior method for wastewater treatment and water reclamation [6] [7].Based on literature reviews, so far, no other studies adsorption on new chemical adsorbent (C,N-bipyrazole with a 3-glycidoxyprpoyl-trimethoxyilane silylant agent) onto a silica surface in short (SG2P) have used.The objectives of this study were to prepare supported SG2P and to test its performance for arsenic removal from water.The effects of various parameters such as adsorbent dosage, pH, contact time, concentration of As and temperature on arsenic removal were assessed.American Journal of Analytical Chemistry

Chemicals and Instruments
The entire chemicals used were of AR grade and were used without further purification.All aqueous solutions were prepared in double distilled water.The distilled water was inspected for arsenic concentration.The concentration was lower than 10 ppb, the detection limit that can be easily accomplished by the analytical methods used in this study [8].The permissible limit of arsenic is 10 ppb.
Stock solutions of arsenic were prepared in reverse deionized (DI) water using arsenic trioxide.Arsenic working solutions were freshly prepared by diluting arsenic stock solutions with DI water.H 2 SO 4 and NaOH (0.1 N) were used for adjusting the pH of the arsenic solution.Then a calibration curve of absorbance versus concentration (Figure 1) was constructed at the time of analysis.Calibration coefficient (R 2 ) was at least 0.983 for all analyses.Appropriately diluted solutions were made from the stock as and when required for calibration purpose.
The instruments used for the experimental work are Electronic Balance, pH meter; Spectrophotometer, Gutzeit apparatus, incubator and Gyro shaker which were used from the college laboratory and all the glass that used were made up of Borosilicate.On the other hand, the amount of metal ion that was sorbed and the percent removal of metal ion by adsorbent were calculated by applying Equation (1) and Equation (2) respectively: where q is the amount of metal ion sorbed by the adsorbent (mg/g); Co is the initial metal ion concentration; (mg/L), Cf is the final ion concentration (mg/L) after the chemo-sorption occurred, V is the volume of aqueous solution (L) in contact with the adsorbent, and m is the mass (g) of adsorbent.

Preparation of Adsorbent Substance (SG2P)
After converting the hydroxy-bipyrazolic ligand 4 to the alcoolate derivative using sodium metal in tetrahydrofuran, the resulting salt (103 mol) was added to a suspension of 3-glycidoxypropyl-functionalized silica (SiPz) (1.00 g) in 30 ml of dimethylformamide (DMF).The mixture was stirred and refluxed under nitrogen for 24 h.The solid material was filtered, and the residue was washed with DMF, toluene, water (distilled and deionized), methanol, dichloromethane (150 ml of each) and finally dried [9] [10].

Determination of Optimum Dosage of Adsorbent
To determine the optimum dosage of SG2P, it was added to the conical flask in different dosage varying from (0.02 g, 0.04 g, and 0.1 g up to 0.15 g), containing 10 mL concentration of arsenic solution (10 µg/L) at pH 6.The solution in the conical flask was subjected to stirring for optimum contact time, filtered and analyzed for residual arsenic concentration.The dosage which provides minimum residual concentration is chosen as optimum dosage.

Selection of Optimum Contact Time
The adsorption is strongly influenced by the contact time.To study the effect of contact time, 10 mL of 10 µg/L arsenic solution was mixed with 0.02 g of adsorbant (SG2P), stirred at different contact times varying from (5 mins, 15 mins up-to 180 mins).Then filtrate was analyzed for arsenic concentration using uv visible spectrophotometer.

Selection of Optimum pH on Arsenic
It was reported in prior studies that pH is a central factor that affects the performance of adsorption process.The effect of pH on arsenic adsorption was studied by performing equilibrium adsorption tests at different initial pH values.
i.e. from 1.0 to 12.0.The pH of solution was adjusted by using 0.1N H 2 SO 4 or 0.1N NaOH.The pH which gives minimum residual concentration is chosen as optimum pH.

Determination of Optimum Temperature on Arsenic
The effect of temperature was investigated at different degrees, 25˚C, 35˚C, 45˚C and 55˚C.For each temperature, a 0.02 g adsorbent (SG2P) samples was added to 10 mL of As(III) solutions with concentration 10 µg/L at pH around 6. The temperature which gives minimum residual concentration is chosen as optimum temperature.

Selection of Optimum Initial Concentration of Arsenic
Al of these concentrations of 10, 20, 30, 40 and 60 ppb solutions were prepared from the stock solution to find out the optimum concentration, 0.02 g of adsorbent (SG2P) was added to a number of tubes contains 10 ml of the previous concentrations of As(III) solutions under optimized temperature (25˚C) and pH American Journal of Analytical Chemistry ured by Flame Atomic Absorption at 193.7 nm.

Determination of Antibacterial Activity of Adsorbent Substance (SG2P)
Antibacterial activity of SG2P was determined by broth micro-dilution method in which SG2P was serially diluted and examined against the bacteria.Minimum inhibitory concentration (MIC) was considered the lowest concentration of substance that inhibited visible growth of bacteria.

Effect of Contact Time
Effect of contact time was investigated and graph of percentage of arsenic removal versus time in minutes was schemed as shown in Figure 2. From the graph it is clear that the adsorption curves are characterized by sharp rise in removal efficiency of Arsenicat the first 20 minutes.Then it has negligible effect.This is may be due to accumulate of chromium ions at the surface of As(III).In this context, the optimum time for the adsorption of arsenic on SG2P was 20 minutes.

Effect of Adsorbent Dosage
Effect of adsorbent dosage was examined and graph of percentage of arsenic removal versus dosage was plotted as shown in Figure 3. From the graph it is observed that, as the dosage of SG2P increases, amount of arsenic decreases, sharply in the beginning and attains maximum later.This is mainly due to the large contact surface area and adsorption sites available for adsorption.The dosage, at which maximum removal is attained, is taken as maximum dosage.The removal efficiency on arsenic was found to be 77%, 80.5%, and 81.5% with an optimum dosage of 0.05 g, 0.1 g, and 0.15 g respectively.

Effect of pH
The adsorption of arsenic from aqueous solution using GS2P in this research was found to be a lower pH (pH 3) dependent process.The graph of percentage of arsenic removal versus pH was plotted as shown in Figure 4. From the graph it is observed that, arsenic is removed more effectively in slight acidic range.This is because the solution contains protons that bind with electrons from oxygen, so the negative charge on the surface attract the positive ions of As, where at high pH in alkaline conditions the result showed is not favorable for arsenic sorption because carboxyl, hydroxyl, and amide groups of the adsorbent become negatively charged and a high density of OH − at alkaline conditions would compete with all anionic species of As(III) [11].

Effect of Temperature
Effect of temperature was investigated and graph of percentage of arsenic removal versus temperature was plotted as shown in Figure 5. From the graph it is observed that, arsenic is removed more effectively in high temperature range.As expected, increasing temperature will lead to increase in temperature results in increase in mobility of large metal ions, which indicates that the adsorption process is endothermic, leads to swelling effect within the internal structure of adsorbent that in turn, encourages the metal ions to penetrate further and increase in the kinetic energy of adsorbent particles that increase the collision between adsorbent and adsorbate molecules results in enhancing the removal of metals from adsorbent surface [12].

Effect of Arsenic Concentration
The effect of arsenic concentration on the adsorption was studied under optimized pH found from our previous results.Concentrations of Arsenic varied from 10 -40 ppb. 10 ml of each concentration of arsenic was treated with the adsorbent at pH value of 3.0.The results are illustrated in Figure 6, which show GS2P is more efficient adsorbent at low concentrations of arsenic.Because at the beginning the removal of As(III) ions is high due to large surface area of adsorbent and when the saturation point is reached, at equilibrium the capacity of the (SG2P) to adsorb As(III) ions is controlled and the ions transported from the exterior to the interior sites of the (SG2P) particles [13].

Adsorption Isotherm of Arsenic
Adsorption isotherms are used to explain the equilibrium of metal ions that occurs between the solid phase of adsorbent and the aqueous solution.Freundlich and Langmuir isotherms are the most models that widely used to determine some kinetic and thermodynamic parameters that can give a clearer image about the binding mechanism.

Freundlich Adsorption Isotherm
The Freundlich isotherm is an empirical equation employed to describe heterogeneous systems.This model is specified with the following equation [14]: The linear form of this equation can be written as: adsorption.If n lies between one and ten, this indicates a favorable sorption process [15].To determine the constants K F and n, the linear form of the equation may be used to produce a graph of ln(Q e ) against ln(C e ) as shown in Figure 7.
From Table 1 the values of n = 1.6949 and 1/n = 0.59, as Freundlich isotherm model, these values indicate that the sorption of As(III) on SG2P is favorable and the R 2 value is 0.911.

Langmuir Adsorption Isotherm
This describes quantitatively the formation of a monolayer adsorbate on the outer surface of the adsorbent, and after that no further adsorption takes place.
Thereby, the Langmuir represents the equilibrium distribution of MB dye between the solid and liquid phases.The Langmuir isotherm is valid for monolayer adsorption onto a surface containing a finite number of identical sites.The model assumes uniform energies of adsorption onto the surface and no transmigration of adsorbate in the plane of the surface.Based upon these assumptions, Langmuir represented the following equation [14] [16]: where: C e = the equilibrium concentration of adsorbate (mg/L); Q e = the amount of As(III) adsorbed per gram of the adsorbent (mg/g); Q m = maximum monolayer coverage capacity (mg/g); K L = Langmuir isotherm constant (L/mg).The values of Q m and K L were computed from the slope and intercept of the Langmuir plot of C e /Q e versus C e .From Langmuir plots which is shown in Fig-     C 0 (mg/l) American Journal of Analytical Chemistry terms of a dimensionless constant separation factor R L that is given by the following equation [15]: ( ) where, C o is the highest initial concentration of adsorbate (mg/L).The value of R L indicates the shape of the isotherm to be either unfavorable (R L > 1), linear (R L = 1), favorable (0 < R L < 1), or irreversible (R L = 0).The R L values between 0 and 1 indicate favorable adsorption.The value of R L in the present investigation was found to be 0.166 at 25˚C indicating that the adsorption of As(III) dye on (SG2P) is favorable [17].

Rate Order of the Adsorption Process
Uptake of each metal ion from different initial concentrations of the metal can be used to study the dependency of the rate of adsorption on the concentration of metal ion left in solution.Thus, the reaction order of the adsorption process can be determined.Pseudo first-order equation was applied for evaluation the adsorption kinetics for As(III) onto SG2P, the rate constant for the adsorption K 1 was evaluated.The pseudo first-order equation expressed as Equation ( 4): ( ) where Q e is the adsorption capacity of the SG2P at equilibrium (mg/g), Q t is the amount of As(III) adsorbed at time t (mg/g) and K 1 is the pseudo first order rate constant (min −1 ).
A linear plot of log(Q e − Q t ) against time allows obtaining the rate constant (Figure 9).If the plot was found to be linear with good correlation coefficient, indicating that Lagergren's equation is appropriate to As(III) sorption on SG2P so the adsorption process is a pseudo first order process.The Lagergren's first order rate constant (K 1 ) and Q e determined from the model are presented in Table 3 along with the corresponding correlation coefficients.It was observed that the pseudo first order model did not fit well.It was found that the calculated Q e values do not agree with the experimental Q e value This suggests that the adsorption of As(III) does not follow first-order kinetics.
From Figure 9, the R 2 value shows that the adsorption As(III) onto SG2P does not obey the pseudo first-order kinetics, and the pseudo first-order model is not appropriate for the adsorption.
The Lagergren pseudo first-order rate constant (K 1 ) and (Q e ) values are represented in Table 3.
The pseudo second order kinetics may be expressed in a linear form as integrated second order rate law [13]: where K 2 is the pseudo second order rate constant (g•mg −1 •min −1 ).American Journal of Analytical Chemistry  Pseudo second-order adsorption model for As(III) adsorption onto SG2P was applied and the rate constant for the adsorption K 2 was evaluated as shown in Figure 10.
The results obtained shows that the value of linear regression coefficient R 2 is 0.999, the values of Q e experimental, K 2 , R 2 and Q e calculated were listed in Table 4.
The results shows that the Pseudo second-order kinetic model perfect fit with experimental data and the value of R 2 = 0.9998, by comparing (Q e ) experimental and (Q e ) calculated values from Table 4, there is agreement between experimental and calculated values for pseudo second order model, and K 2 value greater than K 1 value, so the pseudo second order model is favorable model for As(III) adsorption onto SG2P.

Adsorption Thermodynamics
Adsorption thermodynamics were determined using the thermodynamic equilibrium coefficients obtained at different temperatures and concentrations to verify possible adsorption mechanisms.The adsorption characteristics of a material can be expressed in terms of thermodynamic parameters such as ΔG (Gibbs free energy change), which can be calculated by the following equation [17]: where K d is the thermodynamic equilibrium constant (L•g −1 ).
According to thermodynamics, the Gibbs free energy is the difference between  the adsorption enthalpy (ΔH) and adsorption entropy (ΔS) multiplied by the temperature.In this manner, by applying this concept to Equation ( 9), the thermo chemical parameters ΔH and ΔS can be determined using Van't Hoff's plot (Figure 11), according to Equation ( 12) [18]: H ∆  and S ∆  were calculated from the slope and intercept of the linear plot of lnK d versus 1/T respectively.The results show that the enthalpy of adsorption H ∆  was −3.7886 kJ•mol −1 and S ∆  was 9.9 J•mol −1 •K −1 .G ∆  was calculated at different temperatures from the following equation: The obtained thermodynamic values are given in Table 5.
The negative G ∆  values indicate that the adsorption is spontaneous at these temperatures.The negative value of H ∆  reflects an exothermic adsorption and indicates that the adsorption is favored at low temperature.In the other hand, the positive value of S ∆  suggests that some structural changes occur on the adsorbent and the randomness at the solid/liquid interface in the adsorption system increases during the adsorption process.

Antibacterial Activity of SG2P
The results of the minimum inhibitory concentration of micro dilution tray of the examined microorganism are shown in Figure 12 and in Table 6.

Figure 1 .
Figure 1.Linear calibration curve of absorbance vs. concentration for As (III) concentrations in the range 10 -60 µg/L.

Figure 2 .
Figure 2. Effect of contact time.

ure 8 .
amount adsorbed for monolayer formation (Q m ), Langmuir adsorption-desorption equilibrium constant (K L ) and regression constant (R 2 ) were determined and values are shown in Table2.The essential characteristics of the Langmuir isotherm can be expressed in
American Journal of Analytical Chemistry

Figure 12 .
Figure 12.The inhibitory activity of SG2P against E. coli and S. aureus.

Table 1 .
Parameters and correlation coefficient of Freundlich isotherm model for adsorption of As(III) onto SG2P.

Table 2 .
Parameters and correlation coefficient of Langmuir isotherm model for adsorption of As(III) onto (SG2P).

Table 3 .
Pseudo first-order parameters for As(III) adsorption onto SG2P at 25˚C.

Table 4 .
Pseudo second-order parameters for As(III) adsorption onto SG2P at 25˚C.

Table 5 .
The values of the thermodynamic of As(III) adsorption at various temperatures.

Table 6 .
Demonstration of bacterial growth and inhibition of growth by SG2P in each well.