A new biocompatible ternary Layered Double Hydroxide Adsorbent for ultrafast removal of anionic organic dyes

It would be of great significance to introduce a new biocompatible Layered Double Hydroxide (LDH) for the efficient remediation of wastewater. Herein, we designed a facile, biocompatible and environmental friendly layered double hydroxide (LDH) of NiFeTi for the very first time by the hydrothermal route. The materialization of NiFeTi LDH was confirmed by FTIR, XRD and Raman studies. BET results revealed the high surface area (106 m2/g) and the morphological studies (FESEM and TEM) portrayed the sheets-like structure of NiFeTi nanoparticles. The material so obtained was employed as an efficient adsorbent for the removal of organic dyes from synthetic waste water. The dye removal study showed >96% efficiency for the removal of methyl orange, congo red, methyl blue and orange G, which revealed the superiority of material for decontamination of waste water. The maximum removal (90%) of dyes was attained within 2 min of initiation of the adsorption process which supported the ultrafast removal efficiency. This ultrafast removal efficiency was attributed to high surface area and large concentration of -OH and CO32− groups present in NiFeTi LDH. In addition, the reusability was also performed up to three cycles with 96, 90 and 88% efficiency for methyl orange. Furthermore, the biocompatibility test on MHS cell lines were also carried which revealed the non-toxic nature of NiFeTi LDH at lower concentration (100% cell viability at 15.6 μg/ml). Overall, we offer a facile surfactant free method for the synthesis of NiFeTi LDH which is efficient for decontamination of anionic dyes from water and also non-toxic.


Synthesis of LDHs.
Ni/Fe/Ti LDHs (NiFeTi1, NiFeTi2, NiFeTi3, NiFeTi4 and NiFeTi5) were synthesized by the hydrothermal route with varied concentrations. The synthesis was carried out by adding Ni(NO 3 ) 2 .6H 2 O, Fe(NO 3 ) 3 .9H 2 O, TiCl 4 and 1.5 g urea in 100 ml decarbonated water. The mixture was vigorously stirred and hydrothermally aged in an autoclave at 160 °C for 2 days. The product so formed was washed with distilled water and dried in an oven at 80 °C. (S2 -Electronic supplementary information) Biocompatibility studies. MHS cell lines were grown in RPMI-1640 media in 5% CO 2 incubator at 37 °C. When the cells reached 70% confluency, they were trypsinized and harvested. Cells were counted on a

Adsorbent
Dye implemented % Removal (approx.) Contact time (approx.) pH Reference MgAl-LDH supported Cu-(BDC) MOF Methyl orange 99 20 min 6 40 Ni4Fe1-CO 3 -LDH Methyl orange 20 120 min 5-6 41 ZnAl-LDH/Al (OH) 3 Methyl Orange 98 50 min 4 42 ZnAl-LDH/Al (OH) 3 Congo Red 99 200 min 6 42 S/NiFe-LDH (1:1), Methyl Orange 82. 6 30 min 3 43 S/NiFe-LDH (2:1) Methyl Orange 63.2 60 min 3 43 Mg−Al−CO 3  Adsorption experiments. All the adsorption experiments were performed in batches by dispersing adsorbent (20 mg) in10 mL dye solution (20 mg/L) for 10 min (pH = actual, temperature = 25 °C). The adsorbent was further separated by centrifugation and the solution was used to evaluate the residual dye concentration by using UV-vis spectrometer. The effect of contact time, pH, amount of the adsorbent and the initial concentrations of the dye on the adsorption process were also evaluated. The adsorption kinetics was studied at time intervals of 1, 2, 4, 6, 8 and 10 min. The adsorption isotherm was studied by varing the initial dye concentrations (ranging from 10-70 mg/L). The dye removal efficiency (DRE) was calculated by the formula: The adsorption capacity at equilibrium, q e (mg/g) was evaluated by the equation: The adsorption capacity at time t, q t (mg/g) was calculated by: where, C 0 is the initial concentration of dyes, C e is the equilibrium concentration of dyes and C t is the concentration of dyes at respective time interval. Reaction mixture volume is denoted by V (L) and adsorbent amount used is represented by W (g).

Result and Discussion
Characterization. The XRD pattern of NiFeTi2 LDH can be easily correlated with previously reported data 47 .
The incorporation of CO 3 2− ions and H 2 O molecules in the lattice of synthesized LDH can be confirmed by the 2θ values of 11.12, 22.32 and 34.31° representing (003), (006) and (009) reflections, respectively. The d-spacing of the planes (003) and (110) were found to be 0.796 nm and 0.354 nm, respectively. Due to the similarity in the basal spacing of the synthesized LDH and Ti incorporated LDHs, it could be easily stated that the interlayer CO 3 2− ions and H 2 O molecules have retained a pattern that is similar to the previously reported XRD patterns. Moreover, the presence of (110) and (101) reflections at 25.1° and 36.5° indicates that TiO 2 exists in anatase phase in the synthesized LDH. The XRD pattern of NiFeTi2 LDH is illustrated in Fig. 1(A), XRD patterns of all the synthesized materials is depicted in Fig. S3a(A) and X-ray diffraction parameters are illustrated in Table S1.
During the variation in the prescribed synthesis of LDH, we have performed the PXRD experiment by which we came to observed that the composition of LDH is in its purest form. Simultaneously, we have observed that other material excluding NiFeTi2 LDH contains different oxides other than LDH. Therefore, we have analysed the ICP results of different as-synthesized LDH materials. The pure LDH synthesised as NiFeTi2 LDH (Ni 2.49 Fe 0.2 Ti 1.0 ) and other LDH chemical compositions, with intercorporated CO 3 2− ions and H 2 O molecules, has been provided in Table 2.
FTIR spectra of all the synthesized materials are depicted in Fig. S3a(B) and spectrum of NiFeTi2 LDH is depicted in Fig. 1(B). The broad absorption band located around 3443 cm −1 might be due the stretching vibration occurred because of the presence of layer's hydroxyl group and intercalated water molecules. The shoulders around 3256 cm −1 and 2930 cm −1 could be due to the H-bonding arising between the interlayer H 2 O and CO 3 2− anions. A weaker band at 1632 cm −1 could be assigned to the bending vibrations of hydroxyl group. The observation band at 1387 cm −1 could be asymmetric stretching occurred due to the existence of carbonate ions 48,49 .
The UV-visible spectra of all the synthesized materials are illustrated in Fig. S3a(C). The coordination state and the nature of Ni, Fe and Ti within the layered framework is introduced by the UV-vis of the various LDHs. The hump at ≈230 nm, observed for all the LDHs is attributed to the d-d transitions. The strong adsorption band observed in the range of 250-460 nm can be assigned to typical metal coordinated to the CO 3 2− anion present in the interlayer galleries 47,48,[50][51][52][53][54][55] . A broad shoulder between 385 to 700 nm may be attributed to the existence of Ti 4+ in the brucite-like sheets ( Fig. 1(C)) 47 . Broad valley also signifies supramolecular guest-host interactions (hydrogen bonding, electrostatic attraction and van der Waals forces) or guest-guest interactions (van der Waals forces and hydrogen bonding).
The N 2 adsorption-desorption isotherm of NiFeTi2 is depicted in Fig. 1(D). The isotherm of NiFeTi2 LDH could be easily correlated with H-3 Type hysteresis loop. The hysteresis loop appearing at 0.6-1, suggests the presence of mesoporous material. The calculated BET surface area and Langmuir surface area were 106 m 2 /g and 1702 m 2 /g for NiFeTi2 LDH, respectively. The average pore diameter was also determined by using Barrett-Joyner-Halenda (BJH) method and is obtained as 10.6 nm. The BJH adsorption and desorption pore size distribution volumes were reported as 0.33 and 0.34 cm 3 /g, respectively 49,56 .
In addition, SEM images ( Fig. 2(c,d)) of CO 3 -LDH show platelet structure. The HRTEM images ( Fig. 2(a,b)) clearly indicates the sheets-like morphology of CO 3 -LDH with d-spacing of 0.27 nm.
The thermogram (Fig. S3b(A)) of the synthesized materials exhibit two degradation steps. The first degradation occurs from 50-200 °C due to the elimination of surface and interlayer water, followed by second degradation (250-400 °C) due to the decarbonation and dehydroxylation (Fig. 3(A)) 48 . Figures 3(B) and S3b(B) shows the Raman studies of carbonate anions in NiFeTi2 and all LDHs, respectively. In all the LDHs, ν 1 band is observed around 1038 cm −1 . In NiFeTi1, ν 1 is a weak band which becomes stronger in the case of NiFeTi2, which further becomes weak and broad in the case of NiFeTi3 and NiFeTi4 and finally disappears in the case of NiFeTi5. It is also observed that ν 4 band is weakest in NiFeTi1 at 628.28 cm −1 , stronger in NiFeTi2 at 639.1 cm −1 , more stronger for NiFeTi3 (639.14 cm −1 ), broader and stronger in NiFeTi4 at 655.42 cm −1 and broadest and strongest for NiFeTi5. These observed trends in ν 4 bands is due to the overlap of ν 4 band of CO 3 2− and M-OH of hydroxide layer as interlayer anions and this vibration is assigned to the Eg (τ) mode. These two bands overlap result in broadening of ν 4 band.
Adsorbent variation for adsorption study. Initially, MO was used to carry out the adsorption experiment aiming to select the optimum adsorbent from the series, NiFeTi1-NiFeTi5. The dye removal efficiency of each adsorbent was calculated, shown in (Fig. 4(A)). Among all adsorbents (NiFeTi1-NiFeTi5) implemented for adsorption study, the ternary NiFeTi2 which is purely LDH as compared to all other materials showed the highest

Pseudo-first-order
Pseudo-second-order q e (mg/g) k 1 (min −1 ) R 2 q e (mg/g) k 2 (g/mg min) R 2 MO  www.nature.com/scientificreports www.nature.com/scientificreports/ adsorption capacity with 97.44% removal of anionic dye from aqueous solution and attained equilibrium between 4 to 6 min. This might be due to the inherit properties associated with pure LDH, mainly which are layered structure, large surface area and interlayer ion exchange 43 . Along with this, many factors counting electronegativity, electrostatic attractions and hydrogen bonding might be responsible behind such efficient adsorption. A promising mechanism behind such adsorption could be the formation of H-bonds among the dye molecules and -OH groups of NiFeTi2 LDH. Also, the formation of electrostatic attraction between negative and positive charges of dye molecules and NiFeTi2 LDH surface, respectively, results in excellent adsorption of anionic dyes 48 . Dye variation for adsorption study. The adsorption studies for various organic dyes over NiFeTi2 LDH were performed. It was observed from Fig. 5 that the anionic dyes (MO (97.44%), CR (98.63%), MB (96.81%) and OG (98.71%)) are adsorbed more preferentially over the LDH surface than cationic dyes (MeB (22.81%) and RhB (8.5%)). Moreover, the low adsorption value for cationic dyes might be because of the unfavourable resistance of positive charge present in cationic dyes from the positively charged surface of the LDH on contrary to anionic dyes. Therefore, due to the presence of opposite charge, attraction electrostatic forces took place for the adsorption of the anionic dyes and enhanced their adsorption drastically on the LDH surface. Further studies  NiFeTi2 LDH is shown in Fig. 4(B). For all dyes, adsorption was achieved within 10 min from the initial start of the process which might be due to the existence of easily available active sites on the exterior surface of NiFeTi2 LDH [40][41][42][43][44][45][46] . Further, the effect of alteration in the adsorbent amount (5 to 20 mg) on the dye removal efficiency was evaluated ( Fig. 4(C)). It was observed that when the amount of the adsorbent was increased the dye removal (%) also increases. Hence, from the experimental data 20 mg adsorbent amount was chosen for further evalution. Figure 4(D) demonstrates the cleaning efficiency of NiFeTi2 LDH over a wide range of pH = 4 to 12 (Solutions used for adjusting pH = 0.1 M HCl, 0.1 M NaOH). The results showed that the dye removal efficiency of NiFeTi2 LDH was highest at pH 7 (99.73% (MO), 100% (CR), 100% (MB) and 94.51% (OG)). 100% dye removal efficiency is shown by adsorbent for CR and MB at each and every pH. As per the results suggested, the effect of pH change is noteworthy and had given very indicative information for the dye and adsorbent interactions. For lower pH (pH < 7), the adsorbent is prone to the dissolution which results in adsorption and less availability of active adsorbent sites. Henceforth, dissolution of LDH at lower pH has decreased removal efficiency of dyes. According to study by , the negative charge on the adsorbent surface keeps on increasing with increase in value of pH and results in electrostatic repulsion between the anionic organic dye molecules and adsorbent surface. Therefore, in contrast to lower pH, the lower dye removal percentage at higher pH can be attributed the purely electrostatic force of repulsion 1 . Figure 6 depicts the variation in the removal efficiency of NiFeTi2 LDH observed when the initial dye concentration was varied from 10-70 ppm. The results clearly demonstrated a gradual decrease in removal efficiency when dye concentration was increased. The dye removal efficiency decreases from 97. 44     where, q e (mg/g) and q t (mg/g) are the amount of dye adsorbed at equilibrium and at time (t), respectively; k 1 (min −1 ), k 2 (g/mg min) and k i (g/mg min 0.5 ) are the pseudo-first-order, pseudo-second-order and intraparticle diffusion rate constants, respectively and C is the intercept. The pseudo-first-order and the pseudo-second-order kinetic model fittings are illustrated in (Figs 7 and 8), and their parameters q e , k 1 and correlation coefficient (R 2 ) are provided in Table 2.
From the linear form of pseudo-first order kinetic model (Eq. 4), the values of k 1 and q e were estimated from the slope and intercept of linear plots [log(q e -q t ) versus t] (Fig. 7). Similarly, from the linear form of pseudo-second order kinetic model (Eq. 5), the values of k 2 and q e were evaluated (Fig. 8). All the kinetics parameters are summarized in Table 2. In all the cases, it was observed that the adsorption process followed pseudo-second order kinetic model with higher correlation coefficient (R 2 ) value as compared to first order kinetic model.
Using the intraparticle diffusion kinetic model (Eq. 6), the diffusion mechanism of the adsorption process was investigated. The value of k i has been calculated by using the slope and C is the intercept of the plots of q t versus t 0.5 (Fig. 9(A)). Here, the intercept C refers to the boundary layer thickness. For all the cases, it is observed that the adsorption process has been influenced by multiple processes. From the plots, it could be inferred that along with intraparticle diffusion mechanism another mechanism could also be responsible for the process 1 . The initial portion of the plot signifies the adsorption on the outer surface of NiFeTi LDH, the second portion of the graph indicates the pores adsorption (intraparticle diffusion) and the unavailability of the free adsorptive sites after reaching the equilibrium stage is indicated by the third portion of the plot 1,49 . www.nature.com/scientificreports www.nature.com/scientificreports/ Adsorption isotherm. Adsorption isotherms are the route to get qualitative information of the adsorption capacity of adsorbents and the distribution of the adsorbates between liquid and solid phases after reaching equilibrium. Langmuir and Freundlich isotherm models are used to study the adsorption capacity of the adsorbent at different equilibrium concentrations. The linear representation of the Langmuir and Freundlich isotherms are  www.nature.com/scientificreports www.nature.com/scientificreports/ where, C e (mg/L) is the equilibrium concentration of the adsorbate, q e (mg/g) is the equilibrium adsorption capacity of the adsorbent, q m (mg/g) is the maximum adsorption capacity of the adsorbent and b (L/mg) is the Langmuir constant. The linear plot of C e /q e versus C e (Eq. 7) gives the values of q m and b depicted in (Fig. 10) and results are summarized in Table 3. The adsorption results illustrated higher correlation coefficient (R 2 ) values and best fitted the Langmuir adsorption isotherm model for all the cases. The information regarding the favorability of the adsorption is determined by the dimensionless constant separation factor or equilibrium parameter, R L , which is given by the equation, where, b (L/mg) and C o (mg/L) are the Langmuir adsorption constant and the initial dye concentration, respectively. For an adsorption process to be favorable, the R L value must lie between 0 and 1 (0 < R L < 1). For an irreversible process, R L = 0 and for an unfavorable process, R L > 1. In this study, the calculated R L values are in the range of 0 and 1 at different initial dye concentrations of various dyes which indicate that the adsorption of the dyes over NiFeTi2 LDH is a favorable process. The linear form of the Freundlich adsorption isotherm is represented by   www.nature.com/scientificreports www.nature.com/scientificreports/ where, k f and n gives the value of the Freundlich constants. The intercept and the slope of the linear plot of log q e versus log C e (Fig. 11) gives us the value of k f and n, respectively and is shown in Table 3. All the correlation coefficient (R 2 ) values are relatively lower than that of Langmuir isotherm. All the n values lie within 1-10, which indicates that the adsorption process is the favorable one. All the results indicate that the surface of NiFeTi2 LDH is homogeneous and follows monolayer uptake mechanism (Fig. 12).
To study the adsorption mechanism, FTIR spectra of pure NiFeTi2 LDH, MO and adsorbed MO over NiFeTi2 LDH were compared and are shown in Fig. 13(A). Creation of new bands in the case of MO adsorbed LDH in the wavenumber range of 1000-1300 cm −1 , evidences the adsorption of MO over the surface of NiFeTi2 LDH. The bands appearing around 1022 and 1120 cm −1 could be assigned to the vibration of the -SO 3 − group and 1,4-substituent of the benzene ring of MO dye, respectively 49 . The bands observed at 1176 cm −1 and at 1610 cm −1 could be attributed to the stretching vibration of C-N and C=C of the benzene ring of MO respectively. Hence, we can conclude that MO is adsorbed to the surface of NiFeTi2 LDH.
The reusability test of NiFeTi2 LDH was carried out and is illustrated in Fig. 13(B). Before carrying out the reusability test, desorption study was performed first, by suspending the used NiFeTi2 LDH in the solution of  www.nature.com/scientificreports www.nature.com/scientificreports/ Na 2 CO 3 and stirring it for next 24 h. After 24 h, the adsorbent was collected by centrifugation process, washed with decarbonated water and dried at 80 °C for 12 hours. The recovered adsorbent was again subjected for multi-cycles of adsorption for MO under similar reaction conditions. The recovered NiFeTi2 LDH showed dye removal efficiency of 96, 90 and 88% for three successive cycles. Thus, it can be concluded that NiFeTi2 LDH could be considered as a potential adsorbent for complete elimination of organic dyes from contaminated wastewater.
Biocompatibility. The result shows that NiFeTi LDH are biocompatible in nature. At a concentration of 15 μg/ml 100% cells are viable while on increasing the concentration the viability of cells decreases, but at a higher concentration of 125 μg/ml more than 50% cells remain viable (Fig. 14). So we can conclude that NiFeTi LDH is biocompatible in nature 56 .

conclusion
In summary, we have successfully synthesized NiFeTi LDH using the hydrothermal method and the resulting material was successfully employed in the adsorptive removal of MO, CR, MB and OG from aqueous solutions. Anionic dyes such as MO (97.44%), CR (98.63%), MB (96.81%) and OG (98.71%) showed comparatively greater adsorption than cationic dyes, MeB (22.81%) and RhB (8.5%) over NiFeTi2 LDH at pH = 7.35. NiFeTi2 gave the best dye removal efficiency (approx. 100% removal in 6 min) with greater adsorption capacity and higher kinetic rate constant. Adsorption equilibrium was attained only within 6 min at T = 25 °C. Pseudo-second order kinetic equation and Langmuir equation perfectly describes the adsorption equilibrium data. The dye adsorption over the LDH surface involved a physisorption process through hydrogen bonding. The reusability of the LDH for multicyclic adsorption process was also explored leading to the conclusion that NiFeTi LDH can be considered as a fast and efficient adsorbent for the removal of anionic dyes from contaminated water. The biocompatibility studies carried on MHS cell lines concluded the non-toxic behaviour of NiFeTi LDH.