Biosorption of 2,4,6-trichlorophenol from Aqueous Medium Using Agro-waste: Pine (Pinus densiflora Sieb)

Most industrial waste discharges are often contaminated with phenolic compounds, which constitute a major source of water pollution owing to their toxicity and low biodegradability. Development of cost-effective treatment of such industrial wastewater is therefore of paramount importance. Towards this end, we explore the efficacy of Pine bark powder (PBP), which is an agricultural solid waste material, as a low-cost biosorbent without any pre-treatment, for the adsorptive removal of 2,4,6-trichlorophenol (2,4,6-TCP) from aqueous media. The PBP was thoroughly characterized and the effect of important adsorption parameters were examined in the present investigation. The batch equilibrium data were analyzed using well-known isotherm models. Freundlich isotherm model provided the best description of the equilibrium biosorption behavior. At 25 ± 1 °C, the maximum biosorption capacity (qmax) was 289.09 mg/g, which is higher than most biosorbents reported in the literature while the removal as high as 97% was obtained. Moreover, the biosorption process was fast, attaining equilibrium in less than 120 min of contact. The Elovich model accurately described the kinetics data. In view of high biosorption capacity and fast removal rates, PBP can be used for an efficient and cost-effective treatment of 2,4,6-TCP contaminated wastewater.


Introduction
Water pollution is a major global environmental concern that requires immediate attention and sustainable solutions.Its main sources are untreated industrial effluents that release wastewater contaminated with VOCs, metal ions, and phenolic compounds into the environment. 1mong these pollutants, phenolic derivative 2,4,6-Trichlorophenol (TCP), is a known toxin owing to its positioning of the chlorine atoms (at position 2, 4 and 6) in the phenol ring. 2 Often, industrial waste-water from paints, pharmaceuticals, pesticides, wood, pulp and paper industries are heavily contaminated with chlorophenols, 3,4 which require pre -treatment before discharging into the environment.However, the structural stability and persistence of 2,4,6-TCP makes its removal quite challenging. 5The position of chlorine atoms relative to the hydroxyl group and the stable carbon-chlorine bond and are responsible for its toxicity.Exposure of humans to 2,4,6-TCP via inhalation leads to respiratory problems, altered pulmonary function and pulmonary lesions.Besides, even low concentrations (0.1 mg/L) of chlorophenols give undesirable palatability to the potable water. 6Both IARC (International Agency for Research on Cancer) and United States Environmental Protection Agency (USEPA, 1999) classify 2,4,6-TCP as a probable carcinogen to humans under group B2. 7 For the removal of 2,4,6-TCP from the waste-water, several techniques, e.g.][10][11] However, most separation techniques are often expensive, necessitating the development of alternative treatment technology that is both efficient and affordable.Biosorption is a cost-effective, efficient, and environmentally friendly technique that is fast emerging as an alternative to conventional separation techniques. 124][15][16][17][18][19][20][21][22][23][24][25][26] Owing to abundance and ease of availability, Pine bark can prove to be a cost-effective alternative to other adsorbents used for wastewater treatment.In fact, the Pine bark was found to be effective in the removal of phenolic compounds (Phenol, 2-Chlorophenol and 4-Chlorophenol). 27Pine (Pinus densiflora) belongs to the family pinaceae, and is commonly found around the world.
In the current study, we have investigated the efficacy of the low-cost PBP adsorbent for the treatment of wastewater contaminated with 2,4,6-TCP.In the following, we first describe the synthesis of the PBP, followed by its rigorous characterization using BET surface area, FTIR spectroscopy, XRD, SEM, and elemental analysis to determine the carbon, hydrogen, and nitrogen (CHN) and the presence of various functional groups.The effect important adsorption parameters, such as the solution pH, biosorbent dosage, initial adsorbate concentrations and the contact time are examined on the uptake efficiency of the PBP.Both experimental kinetic and equilibrium data were processed with the help of well-known theoretical models.Finally, the removal efficacy of the present biosorbent is compared with other adsorbents reported in the literature.

1. Materials
Chemicals and other materials were obtained from Aldrich (St. Louis, USA).These chemicals were not purified prior to use.The chemical formula of 2,4,6-TCP is C 6 H 3 Cl 3 O.Its molecular weight is 197.46 g/moL.1.0 g of 2,4,6-TCP was dissolved in 1 litre of double distilled water to prepare the stock solution, which was used to prepare different solutions in the concentration range of 100-400 mg/L.We adjusted the solution pH by adding 0.1 M NaOH and 0.1 M HCl solutions.

Preparation of Pine Bark Powder Biosorbent
The first step consisted of removing the dust and dirt particles from the pine tree bark by washing with distilled water.It was then dried at 60 °C for 48 h.Next, it was ground using a laboratory mill and sieved to obtain powder with narrow particle size distribution, which varied from 55 to 75 μm.In order to remove the lignin content of the powdered sample, it was soaked in 0.1 M NaOH followed by 0.1 M H 2 SO 4 .It was again washed using distilled water then oven dried at 70° ~ 80 °C for 6 ~ 9 h followed by cooling at ambient conditions.This final product was named 'PBP' (Pine bark powder).

3. Batch Adsorption Studies
Biosorption experiments were conducted using 125 mL conical flasks containing 0.1 g of the biosorbent and 100 mL 2,4,6-TCP solutions with initial concentrations varying from 100 to 400 mg/L.The samples were stirred on a shaking water bath temperature controller (25 ± 1 °C at 220 rpm) till equilibrium was attained.The pH was varied from 2.0-10.0 in the present study.The slurry was filtered using ashless Whatman No.50 filter paper (2.7 µm size particle retention) to obtain the supernatant solution.UV-Vis spectrophotometer (Shimadzu UV-1601 Spectrophotometer, Japan) was used to determine the concentrations of 2,4,6-TCP in the supernatant solutions at the wavelength of 296 nm.The amount of 2,4,6-TCP sorbed onto the PBP, q e (mg/g), was computed using the following material balance relationship, (1)   where, C e and C 0 are equilibrium and initial concentrations of 2,4,6-TCP in the solution (mg/L), respectively.W is the PBP mass in mg and V is the solution volume in L. During the batch kinetics studies, the 2,4,6-TCP uptake at a given time t, q t (mg/g), was computed using, (2)   where C t (mg/L) is the concentration of the adsorbate at time, t (min).The 2,4,6-TCP removal was computed by, (3)

1. Characterization of PBP
The adsorbent was characterized to determine ash content, CHN, moisture content, surface properties, and bulk density.These results are shown in Table 1.

1. 1. FTIR Spectral Analysis
The presence of various functional groups involved in 2,4,6-TCP biosorption were determined using FTIR.Fig. 1(a) and 1(b) shows the FTIR spectral data of the virgin PBP and 2,4,6-TCP loaded PBP in the range of 4000-400 cm -1 .The main broad band peaks observed at 3377 & 3420 cm −1 was assigned to the overlap of -OH stretching vibrations arising from hydroxyl groups and -NH stretching vibration mode of the amine functional groups.The strong absorption band observed at around 2918 cm -1 can be attributed to the C-H stretching vibrations of -CH 3 and -CH 2 functional groups.The peak at 1618 cm -1 indicate the functional group region of C=O, O-H, and C-O groups.The peak at 1518 cm -1 is assigned to a conjugated hydrogen bonded carbonyl group.The presence of car-boxyl groups (-COOH) is evident from the peak at 1440 cm -1 .The peaks at 1059 and 617 cm −1 indicate the presence of alkyl halide (C-N and C-Cl stretch).Fig. 1(a) and (b) show some peaks slightly shifted and/or broadened, in wavenumbers from 2913 to 2918 cm -1 , 1618 to 1621 cm -1 , 1518 to 1510 cm -1 , 1440 to 1432 cm -1 , 1059 to 1063 cm -1 , 878 to 882 cm -1 , 767 to 784 cm -1 , 612 to 617 cm -1 were noticed in the spectra of virgin PBP and 2,4,6-TCP loaded PBP.The functional group(s) involvement is quite prominent as confirmed by the changes observed in FTIR spectrum.The FTIR spectral results further confirmed the fact that PBP is mainly made up of cellulose and lignin, containing functional groups such as hydroxyl, amino and carboxylic groups.

1. 2. Scanning Electron Microscope (SEM) & XRD Analysis
SEM pictures (at different magnifications) revealed dense flaky independent plate like structures with typical agglomeration followed by consistent irregular surface morphology (Fig. 2a, b).The surface of PBP exhibits irregular structure typical of highly porous materials, confirming the possibility of 2,4,6-TCP getting adsorbed on the various parts of the biosorbent (Fig. 2b).X-Ray diffractogram of virgin PBR exhibited broad crested peaks (at 2θ = 29.8°indicating amorphous nature of PBP (figure not shown).

2. Effect of pH
It is a critical parameter that affects the process of the adsorption by influencing the interaction between the adsorbate and the adsorbent.The determination of the optimum pH is therefore vital for the efficient implementation of the adsorption based separation technology. 28The pH was varied from 2.0 to 10.0 in order to examine its effect  on the uptake capacity of the PBP.There is a strong correlation between the solution pH and the uptake of the chlorophenol as seen in Fig. 3.The profile shows a global maximum at pH 6.The 2,4,6-TCP is in fact a weak acid having pK a value of 6.23.Thus, the acidic pH below the pKa value favours 2,4,6-TCP to remain un-dissociated, and the dispersion interaction predominate. 6When the pH value is greater than 6, the biosorption capacity decreases as a result of electrostatic repulsion between the surface site and the negatively charged chlorophenolate anion or due to the chlorophenolate-chlorophenolate anions interaction in the solution. 29At low pH, the protonated chlorophenols were more amenable to adsorption as compared to their ionized forms.A similar analogy was attributed to the 2,4,6-TCP adsorption on other adsorbents, such as PU@PDA@MSNs sponge, 30 Acacia leucocephala bark, 19 urea-formaldehyde macroporous foams (UFMF), 31 coconut shell-based activated carbons, 32 oil palm empty fruit bunch-based activated carbons and coconut husk-based activated carbon. 33,34Therefore, the optimal pH of 6 was fixed in in the present study.

Influence of Biosorbent Dosage
This is a key parameter that affects the cost-effectiveness of the adsorption technology.The amount of PBP was varied from 0.05 to 0.8 g in the test solution (C 0 = 100 mg/L) to examine its effect on the removal efficacy the 2,4,6-TCP.Since the equilibrium was attained in 2h, the total contact time was kept 3h in all experiments.It is clear from Fig. 4 that the % removal of chlorophenol increases with the increase in the amount of the biosorbent, reaching as high as 97%, when 0.4 g of the biosorbent was used.This can be attributed to greater availability of the solute molecules per unit mass of the sorbent (i.e., higher solute/adsorbent ratio) resulting in greater removal till saturation conditions are reached.Further increase in the amount of PBP however failed to improve the removal efficiency, because increasing the amount of PBP resulted in reduced equilibrium loading capacities, q e (mg/g), which is in fact the amount of the adsorbate removed per unit mass of biosorbent.

4. Effect of the Initial Concentration and the Contact Time
An important aspect of the efficient and cost-effective implementation of the wastewater treatment process is the equilibrium contact time.Shorter equilibrium contact time enhances the efficiency of the separation process.Fig. 5 shows the progress of the biosorption with time for different strength of 2,4,6-TCP solution.The uptake of 2,4,6-TCP (mg/g) increases with the increase in contact time.The uptake rate is initially faster, but tend to gradually decrease with time as the vacant sites available on the biosorbent surface is gradually occupied by the adsorbate.It took 120 min for the equilibrium to be established.Therefore, the 120 min of contact was fixed for the rest of the batch experiments.
For the adsorptive removal of 2,4,6-TCP, a wide range of adsorption rates has been reported in the literature.Using ash derived from wheat (Triticum aestivum L.) straw, Chen et al. reported equilibrium contact time of 7 days, which consisted of initial 3 days of rapid adsorption. 35sing calcined Mg/Al-CO 3 layered double hydroxide (CLDH) obtained equilibrium adsorption time of 180 min for C 0 = 25-400 mg/L. 36Denizli et al. also reported 240 min of equilibrium contact time for C 0 = 500 mg/L. 37In another study, 400 min was required for the removal of 2,4,6-TCP using PU@PDA@MSNs sponge for small initial concentrations (50-100 mg/L). 30Radhika et al. found that the equilibrium time was 60-210 min using coconut shell-based commercial grade activated carbon for the initial concentration of 10-100 mg/L. 32Siva Kumar et al. found that the

5. Biosorption Kinetics
Adsorption kinetics critically affects the performance of an adsorbent.This aspect was therefore investigated for different initial 2,4,6-TCP concentrations in the present study.The experimental data were analyzed using Pseudo-first-order, pseudo-second-order, Elovich model and intra-particle diffusion kinetic models, which are listed in the following.The pseudo-first-order kinetic model is usually represented as: 38 (4) where k 1 (min -1 ) is the first order rate constant, which can be evaluated from the slope of the log (q e -q t ) versus t (figure not shown).Table 2 lists k 1 values for C 0 = 100-400 mg/L along with correlation coefficients (R 2 ), which is a statistical measure of the goodness of fit.The actual experimental q e values are also presented in the table.There is a substantial difference between predicted and experimental values of q e , which tends to increase with the increase in the solution concentration.For low initial concentrations, the difference is almost 30%, which increase several folds at higher solution concentrations.Clearly, the first order kinetics fails to provide an adequate description of the biosorption of 2,4,6-TCP onto PBP.
The pseudo-second-order kinetic model is generally represented as: 39 (5) The plot of t/q t versus t (figure not shown) in the above equation yields the q e as the slope and k 2 as the intercept.These values are listed in Table 2 along with corresponding values of R 2 .The agreement between predicted and experimental values of q e is excellent.The difference between the two is low for C 0 = 100 mg/L, which unlike the first order kinetic fit, tends to decrease as C 0 increases.For the highest solution concentration considered here, the difference is hardly 2.5%.This confirms the superiority of the pseudo-second-order kinetics as compare to the first-order kinetics.
The biosorption kinetics of 2,4,6-TCP onto PBP was also examined using Elovich equation: 40 (6)   where a (mg/g/min) is the initial biosorption rate and b (g/ mg) is the desorption constant related to the chemisorption activation energy and the extent of the surface coverage of the adsorbent.The plot of lnt versus q t yields parameters (1/b) and (1/b)ln(ab) as its slope and the intercept, respectively.From the Table 2, it can be seen that the computed q e values from Elovich model show good agreement with experimental data.

6. Intraparticle Diffusion Model
The transport of the solute molecules from the bulk liquid to the interiors of the porous structures of the adsorbent involves two mass transport resistances.One is due to the film or the boundary layer surrounding the adsorbent.The greater the thickness of this layer, the higher will be the resistance to the mass transport.Another resistance to the mass transport is due to the movement of the external species inside the pores of the adsorbent, which is also often termed as the intraparticle diffusion.In the literature, the dependence of the concentration of the adsorbate on the time is mathematically expressed as: 41 (7)   where k id (mg/g.min 1/2 ) is the slope of the square root of time versus q t plot while C is the intercept, which is a measure of the boundary layer thickness (figure not shown).The computed k id values are shown in Table 2.If equation ( 7) yields a linear trend passing through the origin, then the intraparticle diffusion is the lone rate limiting step.From the table, it is clear that the intercept C is not zero.Therefore, the presence of other mass transport mechanism, besides the intraparticle diffusion, cannot be ruled out in the biosorption of 2,4,6-TCP.
Figure 6a-d shows the comparison of the experimental data and model predictions for the 2,4,6-TCP concentrations ranging from 100-400 mg/L.The pseudo-first order, pseudo-second order, intraparticle diffusion and Elo-

7. Validity of Kinetic Models
The validity of three kinetic models in accurately describing the biosorption process was carried out by computing the normalized standard deviation Δq t (%).It is defined as: (8)   where 'exp' , 'cal' and 'N' denote experimental, calculated values and the number of data points, respectively.Lower values of normalized standard deviation Δq t (%) indicate better data fit.The computed parameters of the models, their corresponding correlation coefficients (R 2 ), and Δq t (%) values are listed in Table 2.The pseudo-first-order kinetic model gives Δq t (%) as high as 83% while the pseudosecond order and Elovich models give Δq t (%) values ranging from 1.4-3.5% and 0.50-1.2%,respectively.Based on the correlation coefficients (R 2 ) and normalized standard deviation Δq t (%) values, the 2,4,6-TCP adsorption on PBP is best described by the Elovich model model.

8. Biosorption Isotherm Models
In order to mathematically describe the equilibrium distribution of the adsorbate in the liquid solution and the adsorbent, adsorption isotherm models are used.These models accounts for the type of coverage, the nature of the interaction between the adsorbate and the adsorbent surface, and its homogeneity/heterogeneity. In the present study, we focus our attention on three different isotherms models, i.e.Langmuir, Freundlich and Dubinin-Radushkevich to analyze the equilibrium experimental data.
The generalized form of Langmuir isotherm is represented by: 42 Siva Kumar et al.: Biosorption of 2,4,6-trichlorophenol from Aqueous ... (9)   where q e (mg/g) is the amount of the solute adsorbed per unit mass of adsorbent, C e (mg/L) is its equilibrium concentration in solution, q m is the monolayer biosorption capacity of the adsorbent (mg/g) and K L (mg/L) is the Langmuir equilibrium constant, respectively.The graph of 1/q e versus 1/C e has been plotted to determine both q m and K L .The suitability of Langmuir type adsorption can be analysed using the following dimensionless parameter (R L ): 43 (10)   where C 0 is the initial adsorbate concentration in solution (mg/L).For values of R L less than unity, the biosorption is favored.R L values greater than unity indicate that the biosorption process is unfavorable.In the current study, R L values were found varying from 0 to 1, indicating that Langmuir adsorption is favorable.
Another widely used relationship, Freundlich isotherm model is given as: 44 (11)   where K F is a constant related to the adsorption capacity and (1/n) is related to the adsorption intensity.For a favourable sorption, n >1.Taking the log of both sides, the above equation can be linearized as follows: (12)   In order to gain a greater insight into the biosorption process, one can also use the Dubinin-Radushkevich isotherm model to fit the experimental data.The linearized form of this isotherm model can be mathematically expressed as: 45 (13)   where q m (mg/g) is the maximum adsorption capacity, B is a the activity coefficient related to mean adsorption energy, and ε is the Polanyi potential defined as : (14)   where R is the gas constant (8.314J/mol K) and T is the absolute temperature.The mean free energy 'E' of adsorption per molecule of the adsorbate is determined using the constant 'B' using the following relationship: A plot of ε 2 versus ln(q e ) enables determination of constants E and q m .The nature of the biosorption process is characterized by the parameter E. Smaller values of E (< 8 kJ/ mol) indicate that biosorption is purely physical.On the other hand, higher E values (8 < E < 16 kJ/mol) indicate that the sorption process is chemical ion-exchange.We present the model parameters for all the three isotherms, i.e.Freundlich, Langmuir and Dubinin-Radushkevich in Table 3 for comparison along with values of the correlation coefficient (R 2 ).The predictions of all the isotherm models along with the experimental data are presented in Figure 7. Clearly, the Freundlich isotherm fits the experimental sorption data better than both Langmuir and Dubinin-Radushkevich models.cases, R 2 values exceeded 0.997 and low Δq e (%) were obtained in the present study (Table 3).It is however clear from the table that the Freundlich isotherm described the experimental data better than the other two models.This suggests that surface or pore heterogeneity plays a role in 2,4,6-TCP adsorption.The present results also agree with earlier works, which reported that the Freundlich model provided better representation of the 2,4,6-TCP adsorption using oil palm empty fruit bunch and activated clay. 4,33

8. 2. Comparison of PBP with Other Adsorbents
Fig. 8 compares the adsorption capacities of various adsorbents reported in the literature..Its uptake capacity is found to be 289.09mg/g, which is higher than most other agro-waste based adsorbents.This clearly establishes the efficacy of the PBP for the cost-effective removal of 2,4,6-Trichlorophenol from an aqueous media.

8. 1. Validity of Isotherm Models
The capability of the present isotherm models in accurately describing the biosorption process was evaluated by computing the normalized standard deviation Δq e (%) as follows: (16)   where 'q e , i ' , 'q mod , i ' and 'N' denote experimental, model values and the number of data points, respectively.In all

Conclusion
In this study, the potential of PBP (agro-waste material) for the 2,4,6-TCP removal from wastewater over a Siva Kumar et al.: Biosorption of 2,4,6-trichlorophenol from Aqueous ... wide range of concentrations has been examined.The FTIR study confirms the presence of hydroxyl, amine and carboxyl functional groups on the surface of the adsorbent, while SEM study reveals irregular porous surface morphology.The biosorbent performed better under acidic conditions (optimal pH of 6).The biosorption process was fast since equilibrium was achieved within 120 minutes of contact.The sorption behaviour was accurately represented by the pseudo-second order kinetics as compared to other kinetic models.With an increase in the PBP dosage, the equilibrium adsorption capacity (mg/g) gradually decreased.On the other hand, the removal capacity as high as 97% was obtained for 0.4 g/L of adsorbent.However, further increase in its dosage failed to improve the removal efficiency.Three different isotherm models (Langmuir, Freundlich and Dubinin-Radushkevich) were used to represent the experimental equilibrium data.The Langmuir isotherm model predicted the maximum biosorption capacity of 289.09 mg/g at 25 ± 1 °C.These findings conclude that PBP, owing to its ease of availability and environment friendly nature, can potentially replace existing sorbents for the removal of 2,4,6-TCP from aqueous media.

Acknowledgments
Authors appreciate the support of the Deanship of Scientific Research at the King Saud University for the Research Group RGP-1437-003.

Figure 6 .
Figure 6.Comparison of experimental and calculated values obtained from the pseudo-first order, pseudo-second order, intraparticle diffusion and Elovich kinetic models (a-d).

Figure 7 .
Figure 7.Comparison of experimental data with predictions of isotherm models.

Table 1 .
Elemental analysis, physical and surface characterization of PBP