Seaweed-Derived Nontoxic Functionalized Graphene Sheets as

Research Article pubs.acs.org/journal/ascecg

Seaweed-Derived Nontoxic Functionalized Graphene Sheets as Sustainable Materials for the Efficient Removal of Fluoride from High Fluoride Containing Drinking Water Mukesh Sharma,†,‡,§,# Dibyendu Mondal,§,# Nripat Singh,†,‡ Kapil Upadhyay,∥ Akhilesh Rawat,⊥ Ranjitsinh V. Devkar,∥ Rosy Alphons Sequeira,†,‡ and Kamalesh Prasad*,†,‡

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†

Natural Products & Green Chemistry Division, CSIR-Central Salt & Marine Chemicals Research Institute, G. B. Marg, Bhavnagar−364002, Gujarat, India ‡ AcSIR−Central Salt & Marine Chemicals Research Institute, G. B. Marg, Bhavnagar−364002, Gujarat, India § Departamento de Química, Universidade de Aveiro, Campus Universitário de Santiago 3810−193 Aveiro, Portugal ∥ Department of Zoology, Faculty of Science, The M.S. University of Baroda, Vadodara−390002, Gujarat, India ⊥ Department of Chemistry, University of Rajasthan, J. L. N. Marg, Jaipur−302004, Rajasthan, India S Supporting Information *

ABSTRACT: Herein we present a sustainable and costeffective approach for the preparation of functionalized graphene nanosheets (GNs) directly from seaweed and deep eutectic solvents (DESs). The seaweed granules remained after the recovery of juice from fresh brown seaweed, Sargassum tenerrimum, was utilized as a raw material and DESs generated by the complexation of choline chloride and metal salts were employed as solvent and catalyst for the large scale and facile production of metal oxide functionalized GNs. Moreover considering the biological application of such GNs, where nontoxic nature of substrates is desirable, we have also evaluated the cytotoxicity of the functionalized GNs (Fe3O4/ Fe, SnO2/SnO/Sn, or ZnO/Zn-functionalized GNs), and most of them were found to be nontoxic against human lung carcinoma cells (A549). Thereafter, efficiency of these GNs was assessed for the removal of F− from fluoride contaminated groundwater (2.72−6.71 mg L−1) used for drinking purposes. After treatment with GNs, the concentration of fluoride was found to reduce to 0.36−1.69 mg L−1 (75−87% removal efficiency). Moreover, after recovery of GNs from the water, no significant contamination of metal ions was found in the remaining water. Thus, seaweed-derived nontoxic GNs can be utilized to produce safe drinking water with permissible fluoride content as per World Health Organization (WHO) norms. KEYWORDS: Biomass, Functionalized graphene, Nontoxic, Fluoride removal

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INTRODUCTION The honeycomb decorated two-dimensional (2D) planar carbon allotrope is termed as “graphene” which is made up of conjugated sp2 carbon.1 The unique properties such as high thermal and electrical conductivity, superior mechanical strength, high specific surface area, etc., make it a potential material for applications in energy storage devices, as membranes in separations and sensors.2−5 There are several methods available for the commercial production of graphene from graphite including Hummer’s method, scotch tape exfoliation of graphite, and chemical vapor deposition (CVD) of volatile organics on metallic surfaces etc.1,6 However, these methods pose some limitations, for instance uses of toxic chemicals, need of metallic surfaces, high temperature, irregular deposition of carbon, and high cost of production.7 Thus, development of cost-effective technology for large scale © 2017 American Chemical Society

production of graphene is important and an ever growing research area. The use of biomass as a resource for the production of graphene has gained considerable attention in past few years.8−14 The production of single layer N-doped graphene film through the pyrolysis of chitosan under an inert atmosphere and porous graphene-like nanosheets from coconut shells has been reported.8,9 Chen et al. reported the production of high-quality graphene from wheat straw without using any catalyst.10 Shams et al. also developed a scalable method for the production of graphene from dead camphor leaves (Cinnamomum camphora).11 Rice husk biomass has been utilized as a biomass resource for the controllable synthesis of graphene Received: January 18, 2017 Revised: February 23, 2017 Published: March 8, 2017 3488

DOI: 10.1021/acssuschemeng.7b00198 ACS Sustainable Chem. Eng. 2017, 5, 3488−3498

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ACS Sustainable Chemistry & Engineering

Scheme 1. Mechanical Expulsion of Sap from Fresh Seaweed Sargassum tenerrimum to Obtain Seaweed Granules and Deep Eutectic Solvent Promoted Preparation of Functionalized Graphene Nanosheets from Seaweed Granules

quantum dots.12 Other carbon sources such as food, insects, and waste have also been used for the production of single layer graphene on the surface of copper through pyrolysis.13 In a recent review article, different methods for the preparation of sustainable carbon materials from biomass have been disclosed.14 All the literature reported methods for the production of graphene nanosheets have some drawbacks/ limitations such as the need for metal surfaces, requirement of further treatment of as obtained graphene nanosheets to obtain functionalized graphene nanosheets, etc. Besides the use of terrestrial biomass which have the “food− fuel” dilemma,15 seaweed biomass/seaweed polysaccharides are considered to be a substitute for nonrenewable fossil resources16 and various synthetic materials used in a number of high end applications.13 A number of seaweeds can be successfully cultivated, which ensures the availability of seaweed biomass for large scale industrial utilization.17 Other than several uses, seaweed biomass can be applicable for the production of graphene from economic, sustainability, and scalability points of view. In this direction, N- and S-doped porous carbon structures were obtained by the pyrolysis of Undaria pinnatifida, a brown seaweed, without using any template.18 Recently, Fe3O4/Fe functionalized graphene nanocomposites (GNs) derived from brown seaweed have been reported as potential electro-catalysts for oxygen reduction reaction (ORR) in an alkaline fuel cell.19,20 The nontoxic graphene nanosheets and graphene nanosheets-based materials at the explored concentration could be applicable for safer biotechnology, electronics, and biomedical applications. Wang et al. demonstrated the biocompatibility of graphene oxide (GO) on human fibroblast cells. The cells were found dead when the dose was higher than 50 μg mL−1.21 Liu et al. have reported the highly biocompatible graphene nanosheets functionalized with gelatin, and these gelatin functionalized graphene nanosheets can be used for drug delivery and cellular imaging.22 Here also a facile method for the production of biocompatible and nontoxic nitrogen doped graphene nanosheets useful as biomimetic electrochemical sensors for the detection of nitric oxide was reported.23 Considering the harmful effects of fluoride due its excessive presence in drinking water, it is mandatory to eliminate excessive concentrations to provide safe drinking water. In view of the above, the present work reports a facile method for the scalable production of metal oxide functionalized GNs (Fe3O4/Fe, SnO2/SnO/Sn, or ZnO/Zn-functionalized GNs) directly from the naturally abundant seaweed biomass, Sargassum tenerrimum (S. tenerrimum), using deep

eutectic solvents (DESs) obtained by the combination of choline chloride (ChCl) as hydrogen bond acceptors and different metal salts (FeCl3, SnCl2, and ZnCl2) as hydrogen bond donors. DESs were utilized because they can simultaneously act as solvent, soft template, and catalyst for the growth of GNs.19,20 Among the GNs synthesized and used, Fe3O4/Fe functionalized GNs have been previously reported by us.19,20 The cytotoxicity and biocompatibility of the functionalized GNs were evaluated against human lung carcinoma cells (A549). The GNs thus produced were also explored for the efficient removal of fluoride from fluoride rich drinking water.

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EXPERIMENTAL SECTION

Materials. The chemicals used for the synthesis of DESs such as choline chloride, ferric chloride (FeCl3), zinc chloride (ZnCl2), stannous chloride (SnCl2), and sodium fluoride (NaF) was purchased from SD. Fine Chemicals, Mumbai, India, and used without any further purification. The brown seaweed S. tenerrimum was collected from Okha (Geographical location: 22° 28′ N, 69° 4′ E), Gujarat (India). Fluoride contaminated drinking water was collected from natural groundwater sources from the village Govind garh, Ajmer, Rajasthan, India (Geographical location: 26° 45′ N, 74° 38′ E) known to have high fluoride content in drinking water. The water samples were collected from three different locations in the village which were free from excessive dust. The collected samples were packed in airtight preautoclaved polypropylene bottles and transported to the laboratory for further analyses. Synthesis of Deep Eutectic Solvents. DESs [ChCl-FeCl3 (1:2), ChCl-ZnCl2 (1:2), and ChCl-SnCl2 (1:2)] were prepared following the method described by Abbott et al.24 (Supporting Information). Method for Extraction of Seaweed Sap and Production of Granules. The freshly harvested S. tenerrimum seaweed was mechanically crushed followed by centrifugation to obtain liquid sap as supernatant which was stored under refrigeration (1 kg fresh S. tenerrimum yielded 600 mL sap). The remaining solid residue (granules) was dried in oven to obtain dried seaweed granules (a 75 g portion of dry granules with 14.2% moisture was obtained from 1 kg of fresh seaweed). Preparation of Graphitic Carbon Nanosheets. GNs from indigenous seaweeds were prepared as per the patented process (Scheme 1).19,20 In a typical experiment, the solid Sargassum tenerrimum granules and DESs were mixed with each other followed by heating at 80 °C for 30 min to obtain a semisolid composite mass. The semisolid composite mass thus obtained was pyrolyzed at three different temperatures in the range of 700−900 °C under inert atmosphere (95% N2 and 5% H2) followed by washing with water for several times to obtain functionalized GNs. The functionalized GNs thus prepared were characterized for its elemental compositions and data are provided in the Supporting Information, Table S1 (for GNs 3489


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Figure 1. FT-IR spectra (a and b), FT-Raman spectra (c and d), and powder X-ray diffraction pattern (d and f) of Sn- and Zn-functionalized graphene nanosheets obtained at varying pyrolysis temperatures. Freundlich adsorption isotherm equation

such as SAR-Fe-700, SAR-Fe-800, SAR-Fe-900, SAR-Sn-700, SAR-Sn800, SAR-Sn-900, SAR-Zn-700, SAR-Zn-800, and SAR-Zn-900). Method for the Removal of Fluoride from Fluoride-Rich Drinking Water. A stock solution (500 ppm) for fluoride was prepared by dissolving sodium fluoride (NaF) in Milli-Q water. This was further diluted to prepare 0.5, 1, 2, 3, 4, 5, 10, 25, 50, and 100 ppm standard solutions. In a typical procedure, 0.025 g of GNs was added in 50 mL fluoride solution in a 150 mL conical flask. The solution was then homogenized by ultrasonication for 5 min. The homogenized solution thus obtained was shaken at room temperature for 30−120 min (for optimization). After that the suspension was filtered through a Whatman filter paper of 0.45 μm. The fluoride concentration in the filtrate was measured on an Orion Versa Star ISE meter fitted with an ionplus sure-flow fluoride electrode (Thermo Scientific, USA). A similar experiment was carried out for real fluoride contaminated drinking water samples which was collected from real water sources (pH 7.7−7.8). The amount of fluoride removed by GNs at equilibrium was calculated using the following eq 1:

qe =

⎛ c0 − ce ⎞ ⎜ ⎟v ⎝ m ⎠

log qe = log KF +

1 log Ce n

(2) −1

Where, Ce is the concentration of fluoride at equilibrium (mg L ), KF is the binding constant, and 1/n is the empirical parameter in the Freundlich adsorption isotherm equation. Langmuir adsorption isotherm equation

Ce C 1 = e + qe qm KLqm

(3) −1

Where, qm is the measurements of the sorption capacity (mg g ) and KL is the binding constant of Langmuir adsorption isotherms. Characterization. Characterization of GNs was carried out using a number of analytical techniques as described in the previous work19 and mentioned briefly in the Supporting Information.

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RESULTS AND DISCUSSION Preparation of Functionalized Graphene Nanosheets. The freshly harvested seaweed biomass was utilized to prepare functionalized GNs. The seaweed biomass was crushed to make a semisolid blend and then centrifuged to separate the juice (60−70% v/w of fresh seaweed biomass) and solid granules as shown in Scheme 1. The solid granules were allowed to dry in an oven to obtain dry seaweed granules (75 g dry granules with ∼14% moisture were obtained from 1 kg of fresh seaweed biomass). The seaweed granules thus obtained were charac-

(1)

Where C0 and Ce are the initial and equilibrium concentrations of fluoride (ppm) respectively, m is the mass of GNs (g), and v is the volume of the solution (L). Determination of Adsorption Isotherms. The fluoride adsorption isotherms were determined using the following equation 3490


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Figure 2. XPS survey spectrum of (a) Sn- and (b) Zn-functionalized graphene nanosheets showing inherent doping of highlighted elements.

cm−1 (2D-band) in a Raman spectrum are the characteristic signature of graphene. The existence of sp2-hybridized carbon atoms is demonstrated by the G-band, whereas the D-band reflects the defects present in graphene and the 2D-band gives an indication of the number of layers in the graphene sample.29 The properties of the graphene materials are also studied by the relative intensity ratio of the G-band (IG) to the D-band (ID); the less the ratio of ID/IG, the better the quality of graphene.28 As can be seen from Figure 1c and d for Sn and Znfunctionalized GNs, D-band and G-band were found to be present in all these samples; however, the 2D-bands were not found in all cases. Besides, the ID/IG ratio was found to be 1.13, 1.25, 1.11, 1.13, 1.27, and 1.12 for SAR-Sn-700, SAR-Sn-800, SAR-Sn-900, SAR-Zn-700, SAR-Zn-800, and SAR-Zn-900 GNs, respectively. The Raman spectra for Fe-functionalized GNs are already reported in a previous article, here also the D-band and G-band was found to be present in all samples; however the 2D band was found only in SAR-Fe-700 and SAR-Fe-800 GNs.19 Thus, the overall results indicates that the quality of graphene was better in the case of SAR-Sn-900, SAR-Zn-900, SAR-Fe700, and SAR-Fe-800. Figure 1e and f shows the powder X-ray diffraction pattern of Sn- and Zn-fuctionalized GNs. As can be seen from Figure 1e and f, SAR-Sn-700 and SAR-Zn-700 GNs show the presence of SnO/SnO2 and ZnO phases when the semisolid composite of seaweed granules and DES pyrolyzed at 700 °C (ref. JCPDS file no.: 04-007-0731, 04-012-0775, and 04-014-0069), respectively. These phases remain similar in SAR-Sn-800 and SAR-Zn-800 GNs when the pyrolysis temperature increased to 800 °C. Upon further increase in the pyrolysis temperature up to 900 °C, that fact that all phases disappeared indicates the absence of metal oxide in SAR-Sn-900 and SAR-Zn-900 GNs. Some other peaks appear in SAR-Sn-900 and SAR-Zn-900 GNs with weak intensity which shows the presence of metal particles indicating that the GNs are functionalized with Sn and Zn metal particles, respectively. The PXRD patterns of Fe-functionalized GNs such as SAR-Fe-700, SAR-Fe-800, and SAR-Fe-900 are already reported in a previous article.19 X-ray photoelectron spectroscopy (XPS) is a technique to investigate the chemical environment of graphene lattices modified by foreign doping.30 The XPS survey of both the Snand Zn-functionalized GNs as shown in Figure 2a and b shows the bands of C, O, N, and Sn/Zn with a trace of sulfur. The appearance of oxygen and the Sn/Zn band in the XPS survey spectrum of Sn- and Zn-functionalized GNs indicates the presence of SnO/SnO2 and ZnO in the GNs, respectively, and these results are consistent with the powder XRD spectra (Figure 1e and f) and elemental analysis data (Table S1). The high resolution C 1s spectrum of Sn-functionalized GNs

terized for their elemental composition in which concentrations of Na, K, Ca, Mg, C, H, N, and S were found to be significant and characteristic of the seaweed as shown in Table S1. Scheme 1 shows the procedure for the synthesis of functionalized GNs by using solid seaweed granules and DESs. The functionalized GNs prepared were characterized for their elemental compositions, and data are provided in Supporting Information, Table S1 (for GNs such as SAR-Fe-700, SAR-Fe-800, SAR-Fe900, SAR-Sn-700, SAR-Sn-800, SAR-Sn-900, SAR-Zn-700, SAR-Zn-800, and SAR-Zn-900). As can be seen from Table S1, concentrations (%) of N and S were reduced with the increase in pyrolysis temperature, whereas the amount of Fe was found to increase; however, the amounts of Sn and Zn were found to decrease with the increase in pyrolysis temperature. Among the GNs reported herein SAR-Fe-700, SAR-Fe-800, and SAR-Fe-900 were previously reported by us as elctrocatalysts for oxygen reduction reaction;19,20 however, neither the cytotoxicity of these GNs nor their F− removal efficiency was evaluated. Characterization of Functionalized Graphene Nanosheets. All the products obtained after pyrolysis of Sargassum granules and DES [ChCl-FeCl3 (1:2), ChCl-SnCl2 (1:2), and ChoCl-ZnCl2 (1:2)] mixtures were characterized by various spectroscopic and microscopic techniques for the confirmation of synthesis of GNs. Figure 1a and b shows the FT-IR spectra of Sn- and Zn-functionalized GNs obtained after pyrolysis of semisolid composite of seaweed granules and DESs at different pyrolysis temperatures (700−900 °C). As can be seen from the spectra, the peaks appearing at ∼1636 and ∼1576 cm−1 can be assigned to stretching of CC and C−N bonds,25 respectively. The band at ∼1576 cm−1 appears in the SAR-Sn-700 and SARSn-800 GNs which indicates the presence of C−N bonds in both GNs. For SAR-Sn-900, there was no band found for C−N bonds from FT-IR spectra, and the absence of nitrogen in these GNs is also supported by elemental analysis (Table S1). The band appearing at ∼600 cm−1 is ascribed to stretching vibrations of Sn−O or Zn−O bonds.26,27 This indicates the presence of tin oxide (SnO and SnO2) in SAR-Sn-700, SAR-Sn800, and SAR-Sn-900 and zinc oxide (ZnO) in SAR-Zn-700 and SAR-Zn-800, whereas no band appeared at ∼600 cm−1 in the case of SAR-Zn-900 which indicates the absence of ZnO. The FT-IR spectra of Fe-functionalized GNs (SAR-Fe-700, SAR-Fe-800, and SAR-Fe-900 GNs) are already reported in a previous article.19 The presence and absence of metal oxide (SnO, SnO2, and ZnO) in GNs was further confirmed by powder XRD as discussed below in Figure 1e and f. Raman spectroscopy is frequently used for the confirmation of graphene-like structure present in pyrolyzed samples.28 The peaks at 1350 cm−1 (D-band), 1570 cm−1 (G-band), and 2700 3491


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Figure 3. (a−c) SEM images of SAR-Fe-700, SAR-Sn-700, and SAR-Zn-700 graphene nanocomposites, respectively. (d−f) SEM-EDX element mapping on above graphene nanocomposites (red and green color shows oxygen and metal element such as Fe, Sn, and Zn, respectively). (g−i) SEM-EDX profile diagram for above functionalized graphene nanocomposites.

Figure 4. (a−c) TEM images of SAR-Fe-700, SAR-Sn-700, and SAR-Zn-700, respectively. (d−f) HR-TEM images from above graphene nanocomposites. (g−i) HR-TEM images from above graphene nanocomposites showing homogeneous distribution of metal particles on the surface of graphene nanocomposites.

presented in the Figure S1a, shows a high concentration of sp2 carbon with a broad asymmetric tail toward a higher binding energies, a typical signature of graphene samples. The Znfunctionalized GNs also showed a broad asymmetric tail of sp2

carbon toward higher binding energies (Figure S2a). The existense of sp2 carbon in Sn- and Zn-functionalized GNs can be seen at ∼283.1 and ∼294 eV for C 1s which is from the interaction of graphitic carbon in C−OH with O 1s at ∼531 3492


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ACS Sustainable Chemistry & Engineering and ∼541 eV, as shown in the XPS survey spectra, Figure 2a and b. A small band of nitrogen (N 1s peak) also appeared at ∼399 and ∼409 eV for Sn- and Zn-functionalized GNs, respectively, confirms that nitrogen interacts with carbon by attaching to the edge of two graphitic carbon nanosheets (Figure S1b and S2b). From XPS survey spectra, the presence of SnO/SnO2 and ZnO peak edges in Sn- and Zn-functionalized GNs, respectively, is also clear (Figure S1c and S2c). In Sn-functionalized GNs, two peaks appreared for tin at ∼486 eV for Sn 3d5/2 and ∼495 eV for Sn 3d3/2 (Figure S1c). Znfunctionalized GNs also showed two peaks of zinc at ∼1031 eV for Zn 2p3/2 and ∼1055 eV for Zn 2p1/2 (Figure S2c). Thus, the XPS spectra of Sn- and Zn-functionalized GNs indicates the presence of metal oxide in all the GNs except SAR-Zn-900, because no band appeared in the range of 1025−1060 eV which indicates the absence or presence of a very small amount of zinc particles on the GNs surface (Figure S2c); this is also supported by PXRD (Figure 1f) and elemental analysis data as shown in Table S1. The XPS survey spectrum and the high resolution N 1s and Fe 2p spectra for Fe-functionalized GNs such as SAR-Fe-700, SAR-Fe-800, and SAR-Fe-900 are reported in a previous article.19 After confirming the structural properties of functionalized GNs by FT-IR, PXRD, Raman, and XPS spectroscopic techniques, the sheetlike morphology of the GNs were confirmed by SEM and TEM microscopic images. Figure 3a− c shows the SEM images of GNs such as SAR-Fe-700, SAR-Sn700, and SAR-Zn-700, respectively. The sheetlike morphology of graphitic carbon was visible in SEM images, which was consistent with those observed in TEM images. SEM-EDX element mapping was performed for the investigation of the distribution of metal oxide (Fe3O4, SnO/SnO2, and ZnO)/ metal on the surface of GNs (Figure 3d−f). The SEM-EDX element mapping images of GNs showed the homogeneous distribution of metal oxide/metal particles on the surface of GNs (red and green color indicated the presence of oxygen and metal elements, Fe/Sn/Zn, respectively). The EDX profile diagram showed the presence of elements such as C, N, O, and metal particles in appropriate amounts in these GNs (Figure 3g−i). Therefore, the GNs thus prepared directly from the pyrolysis of seaweed granules and DESs in an inert atmosphere are functionalized with metal oxide/metal particles, and the presence of metal oxide/metal particles is also confirmed by FT-IR, PXRD, XPS, and elemental analysis of GNs as discussed above. The SEM images of others GNs such as SAR-Fe-800, SAR-Fe-900, SAR-Sn-800, SAR-Sn-900, SAR-Zn-800, and SARZn-900 showed the sheetlike morphology in all these GNs, and the SEM images of these GNs are provided in Supporting Information Figure S3. Figure 4a−c shows the TEM images of the GNs such as SAR-Fe-700, SAR-Sn-700, and SAR-Zn-700, and it can be observed from the TEM images that the GNs has sheetlike morphology with nanoscale structural distribution. Figure 4d−f provides the high resolution TEM (HR-TEM) images of these GNs. TEM images (Figure 4g−i) also showed homogeneous distribution of metal particles on the surface of GNs indicated the GNs are doped with metal particles. The TEM images of others GNs such as SAR-Fe-800, SAR-Fe-900, SAR-Sn-800, SAR-Sn-900, SAR-Zn-800, and SAR-Zn-900 showed the sheetlike structures in all these GNs and the TEM images of these GNs are provided in Supporting Information Figure S4. After characterization of structural and functional properties of the GNs, their surface properties were also evaluated. Surface

area of the GNs was measured by N2 adsorption−desorption isotherm and was calculated using Brunauer−Emmett−Teller (BET) equation. The N2 adsorption−desorption isotherm and pore volume distributions of functionalized GNs showed in Figures S5 and S6 and the surface area, pore volume, and pore size data are provided in Table 1. In the pore size analysis of the Table 1. BET Surface Area, Pore Volume, and Pore Size of Functionalized Graphene Nanosheets Obtained at Different Pyrolysis Temperature Ranges sample code

BET surface area (m2 g−1)

pore volume (cm3 g−1)

pore diameter (nm)

SAR-Fe-700 SAR-Fe-800 SAR-Fe-900 SAR-Sn-700 SAR-Sn-800 SAR-Sn-900 SAR-Zn-700 SAR-Zn-800 SAR-Zn-900

220.73 167.82 132.02 270.72 298.38 331.76 250.65 223.15 216.14

0.063 0.107 0.193 0.145 0.142 0.132 0.077 0.089 0.233

2.80 2.79 2.18 3.07 3.69 3.71 2.68 2.94 2.81

GNs, pore volume was found to be in the range of 0.042−0.233 cm3 g−1 (Table 1). In the same order, average pore diameter were noted to be in the range of 2.68−4.81 nm (Table 1), which is a signature of mesoporous materials. The pore structures of functionalized GNs are complex in nature and tend to be made up of interconnected networks with different size and shape. Thus, the mesoporous nature of functionalized GNs dominates the micropore volume of the materials. The surface areas of the GNs were measured by BET method and found to be high (Table 1). As can be seen from Table 1, the surface area of Fe and Zn-doped GNs decreased with increasing pyrolysis temperature. This may be due to the collapse of pores with the increase of temperature. Whereas, in case of Sn-doped GNs, the surface area increased with increasing pyrolysis temperature. The highest BET surface area, 331.76 m2 g−1, was observed for SAR-Sn-900 (Sn-doped GN). This was because of the flaky properties of GN and high exfoliation during pyrolysis of semisolid composite of seaweed granules and DESs as shown in Scheme 1. The lowest surface area, 132.02 m2 g−1, was observed for SAR-Fe-900 (Fe-doped GN), and this was because of its lower exfoliation during pyrolysis of semisolid composite.19 According to the IUPAC technical report, the N2 adsorption−desorption isotherm plots (hysteresis loop) are classified as various types.31 Thus, the physisorption isotherm of functionalized GNs is type-4, and it comes under hysteresis loop type H2 of all functionalized GNs (Figures S5a and c and S6a). So the GNs with type-4 physisorption isotherm and H2 type loop is indicated the mesoporous adsorbent type materials which may be effective for the adsorption of fluoride ions from fluoride-rich drinking water as discussed here. Cytotoxicity Assay of Functionalized Graphene Nanosheets Tested on Human Lung Carcinoma Cells. Human lung carcinoma cells (A549) were used for the cytotoxicity MTT assay of functionalized GNs (details are described in the Supporting Information). In the present study, GNs did not record cytotoxicity at the highest dose of 200 μg, and hence, their effective dose (ED50) could not be calculated. These results were in agreement with cytological observations wherein cells were adherent and spindle-shaped at the end of treatment 3493


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Figure 5. Cytotoxicity potential of graphene nanosheets such as (a) SAR-Fe-700, (b) SAR-Fe-800, (c) SAR-Fe-900, (d) SAR-Sn-700, (e) SAR-Sn800, (f) SAR-Sn-900, (g) SAR-Zn-700, (h) SAR-Zn-800 and (i) SAR-Zn-900 on human lung carcinoma cells (A549). Results are expressed as mean ± SD for triplicate samples wherein ns = nonsignificant, * p < 0.05, and ** p < 0.01 as compared to control.

The concentration dependent adsorption of fluoride using GNs (SAR-Fe/SAR-Sn/SAR-Zn obtained at different pyrolysis temperature of semisolid composite) from fluoride-rich standard solution (fluoride ion concentration varied from 4 to 100 ppm) was also carried out, and the data are shown in Figure 6b−d. The results indicated that SAR-Fe-700 showed highest adsorption capability in comparison to other two Fe-doped GNs (SAR-Fe-800 and SAR-Fe-900), and it’s adsorption capacity toward fluoride was 79 mg g−1 with the initial concentration of fluoride is 100 mg L−1 (Figure 6b), which indicates that the adsorption of fluoride was decreased with decreasing of BET surface area of Fe-functionalized GNs (Table 1). Thus, BET surface area of GNs can play an important role in the adsorption of fluoride from fluoride rich aqueous solution. As can be seen from Figure 6c and d, SARSn-900 and SAR-Zn-900 showed the highest adsorption capabilities in comparison to other GNs (SAR-Sn-700, SARSn-800, SAR-Zn-700, and SAR-Zn-800), and the adsorption capacity affinity toward fluoride was 78 and 75 mg g−1 when the initial concentration of fluoride was 100 mg g−1, respectively. Here it was evident that, adsorption of fluoride increased with increase of BET surface area, which was opposite to what observed for Fe-doped GNs. Thus, the adsorption of fluoride is not only dependent on the high specific surface area but it can also be affected by the surface functionalization of materials using different metal oxides/metal ions. Iron functionalized materials such as sulfate-doped Fe3O4/Al2O3 nanoparticles, iron−aluminum binary oxides (FeAlOxHy), aluminum modified iron oxides, zirconium−iron oxide, etc., were reported to

(supporting Figure S7) and were comparable to that of the control cells (Figure S8). But SAR-Sn-700 GN recorded 20% decrement in the cell viability at 10 μg without major alterations in cell morphology (Figure S7d), while the highest dose of 200 μg also elicited similar levels of toxicity response (Figure 5d). Hence, it can be inferred that SAR-Sn-700 shows moderate toxicity even at the highest dose whereas other GNs such as SAR-Fe-700, SAR-Fe-800, SAR-Fe-900, SAR-Sn-800, SAR-Sn-900, SAR-Zn-700, SAR-Zn-800, and SAR-Zn-900 are nontoxic (Figure 5). These observations are vital for safety assessment of functionalized GNs and open new avenues for their use in biomedical applications and removal of fluoride from fluoride-rich drinking water to obtain safe drinking water. Efficient Removal of Excess Fluoride from FluorideRich Drinking Water and Kinetic Study. The study of nontoxic GNs (mesoporous adsorbent) with high specific surface area has opened up a new green, sustainable, and costeffective way to remove excess fluoride from fluoride-rich drinking water. In a typical experimental procedure, 0.025 g of the GNs was added to 50 mL of fluoride-rich drinking water and this was shaken at room temperature for 30−150 min followed by filtration through Whatman filter paper to optimize the time of adsorption. The effect of time on fluoride adsorption by Fe-doped GNs obtained at different pyrolysis temperatures was studied, and the time dependent study is shown in Figure 6a. It can be seen that the adsorption rate of fluoride increased with time and became constant after 120 min, which was considered to be the optimized time duration for maximum fluoride adsorption (Figure 6a). 3494


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Figure 6. (a) Optimization of time for adsorption of fluoride using SAR-Fe graphene nanosheets. (b−d) Isotherms for fluoride removal from standard solution of NaF using SAR-Fe, SAR-Sn, and SAR-Zn graphene nanosheets obtained in different calcining temperature ranges, respectively. (e) Removal of fluoride from drinking water collected from natural sources using SAR-Fe-700. (f) Removal of fluoride from drinking water collected from natural sources using Fe/Sn/Zn-doped graphene nanosheets obtained in different calcining temperature ranges.

Figure 7. (a) Zeta potential for Fe/Sn/Zn-doped functionalized graphene nanosheets at different pH values. (b) pH dependent fluoride removal from standard solution of NaF (fluoride concentration = 25 mg L−1) using SAR-Fe-700, SAR-Sn-900, and SAR-Zn-900. (c−e) SEM-EDX element mapping images with their EDX profile of graphene nanosheets such as SAR-Fe-700 (c), SAR-Sn-900 (d), and SAR-Zn-900 (e) after adsorbing fluoride from fluoride rich water (red color shows fluoride ions, and green color shows metal elements such as Fe, Sn, and Zn).

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ACS Sustainable Chemistry & Engineering Table 2. Adsorption Isotherm Constants for the Removal for Fluoride from Drinking Water Langmuir model

Freundlich model

adsorbent

KL

qm (mg g−1)

R2

KF

1/n

R2

SAR-Fe-700 SAR-Sn-900 SAR-Zn-900

4.7389 × 10−2 1.8513 × 10−2 2.1749 × 10−2

45.5532 75.7576 74.0741

0.9945 0.9784 0.9913

2.4530 1.7531 1.7559

0.7304 0.8154 0.8171

0.9926 0.9993 0.9867

remove fluoride ion from fluoride rich aqueous solution by adsorption technique.32 Thus, the Fe-functionalized magnetic GNs showed highest fluoride adsorption capacity in comparison to Sn- and Zn-functionalized GNs, and the magnetic properties of the Fe-functionalized GNs make them easy to recover from drinking water using an external magnet (Supporting Information, Figure S9). After optimizing the adsorption time with standard fluoride solution, the optimized procedure was applied to remove fluoride (fluoride content was in the range of 2.72−6.71 mg L−1) from naturally occurring drinking water collected from groundwater sources (Govind Garh, Rajasthan, India; geographical location: 26°45′ N, 74°38′ E) regularly used by villagers for drinking. After treating the water sample with SARFe-700 GN, the concentration of fluoride was found to reduce to ∼0.45 mg L−1 from an initial concentration of 3.62 mg L−1 i.e., SAR-Fe-700 showed ∼87% fluoride removal efficiency (Figure 6e). Thus, the final concentration of fluoride in water was below the permissible limit of fluoride (1.5 mg L−1) according to World Health Organization (WHO) and Bureau of Indian Standards (BIS) standards for drinking water. As can be seen from Figure 6f, the Fe/Sn/Zn-doped GNs were tested for their fluoride removal efficiency from fluoride rich drinking water (initial concentration of fluoride is 3.62 mg L−1), and the removal efficiency of fluoride was found to be in the range of 80−87%. The zeta potential at varying pH range and effect of pH on the adsorption of fluoride was also carried out with standard NaF solution (fluoride ion concentration = ∼25 mg L−1) using metal oxide functionalized GNs. It is most significant parameters to define the adsorption of fluoride ion on the GNs due to the GNs’s high specific adsorption capacity. The zeta potential of the all nine metal oxide functionalized GNs showed similar trends at the pH range of 2−12 (Figure 7a). At pH 2.0, GNs showed positively charged surface, however when the pH was increased from 2 to 12, the zeta poteinitial of the GNs also increases and showed negatively charged surface. Figure 7b showed the variation in removal of fluoride ion using Fe-, Sn-, and Zn-functionalized GNs at varying pH range from 2 to 12. The acidic pH showed the highest adsorption of fluoride while the basic pH showed lowest adsorption of fluoride. The trend of removal of fluoride is similar to the trend of zeta potenial. The adsorption of fluoride is constant between pH 4 and 8. The highest adsorption of fluoride in acedic pH was because of the decrease in surface charge on the functionalized GNs and Fefunctionalized GNs showed highest positive charged surface in comparison to Sn- and Zn-functionalized GNs. Due to the more positive charged surface, SAR-Fe-700 showed maximum adsorption of fluoride from fluoride rich drinking water. Thus, the surface charge and high adsorption capacity of GNs is the most significant factor to remove fluoride from fluoride rich drinking water. As discussed in Figure 6a above and by using eqs 2 and 3, the adsorption isotherm models for the fluoride removal was calculated and the various parameters are shown in Table 2.

Considering the correlation coefficient, the equilibrium data for the F adsorption process on various GNs, it was observed that the adsorption phenomenon by SAR-Fe-700 and SAR-Zn-900 best fit the type 2 Langmuir isotherm with R2 = 0.9945 and R2 = 0.9947, respectively. However, the adsorption on SAR-Sn900 best fit to the Freundlich isotherm. Moreover, the R2 for all of the isotherms were more than 0.9 which indicates that the adsorbents follow the isotherms models well. The removal of fluoride using GNs was also detected in SEM-EDX element mapping images to determine the adsorption of fluoride on the surface of GNs (Figure 7c−e). The SEM-EDX images of recovered GNs after fluoride adsorption from fluoride rich aqueous solution after shaking for 2 h at room temperature showed homogeneous distribution of fluoride (red color dots) with metal oxide particles (green color dots) on the surface of functionalized GNs. There were no major contaminants from the GNs added into the water during adsorption process was observed indicating the water obtained after fluoride removal is safe for drinking purposes. The toxic heavy elements such as mercury and manganese were found more than permissible limits in naturally collected drinking water (Table S2). These elements were also adsorbed significantly by functionalized GNs such as SAR-Fe-700 and SAR-Sn-900 (Table S2), and finally both elements were found in permissible limits in drinking water after treatment with GNs. Thus, the GNs can also adsorb toxic heavy elements apart from the fluoride from natural drinking water, and finally, the nontoxic GN-treated water rich with essential elements can be safe for human consumption.

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CONCLUSION

In conclusion, a scalable, sustainable, and cost-effective approach was developed for the synthesis of metal oxide functionalized graphene nanosheets directly from the pyrolysis of seaweed granules and deep eutectic solvents. The seaweed granules remaining after the recovery of sap from fresh brown seaweed, Sargassum tenerrimum, are utilized as a raw material, and deep eutectic solvents generated by the complexation of choline chloride and metal salts such as FeCl3, SnCl2, ZnCl2 are employed as solvents as well as catalysts for the large scale production of metal oxide functionalized graphene nanosheets. We also evaluated the cytotoxicity and biocompatibility of the metal oxide functionalized graphene nanosheets, and most of them found to be nontoxic against human lung carcinoma cells (A549). Afterward, these graphene nanosheets are tested for the efficient removal of fluoride from fluoride-rich natural drinking water. After treatment of water with graphene nanosheets, the concentration of fluoride is reduced to its permissible limit and no significant contamination of metal ions was found in the water. Thus, seaweed derived nontoxic graphene nanosheets can be utilized to produce fluoride safe (below WHO and BIS standards) drinking water. 3496


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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acssuschemeng.7b00198. Tables describing elemental composition of different constituents present in Sargassum granules, functionalized graphene nanocomposites obtained after pyrolysis of granules and DES mixtures at different pyrolysis temperatures, and naturally collected water before and after treatment with functionalized graphene nanosheets and comparison with WHO and BIS standards for drinkable water. High-resolution XPS graphs, SEM, and TEM images of functionalized graphene sheets (PDF)

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]; [email protected]. Phone no.: +91-278 2567760. Fax no. +91-278-256756 (K.P.). ORCID

Mukesh Sharma: 0000-0002-3438-3367 Dibyendu Mondal: 0000-0002-1715-5514 Kamalesh Prasad: 0000-0003-2366-1664 Author Contributions #

M.S. and D.M. contributed similarly to this work.

Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS K.P. thanks CSIR, New Delhi, for the grant of the CSIR− Young Scientist Awardees Project and overall financial support. M.S., N.S., and D.M. thank UGC and CSIR for research fellowships and AcSIR for Ph.D. registration. Mr. Arka Saha is acknowledged for useful discussions and help. The authors are also thankful to “Analytical Discipline and Centralized Instrumental Facilities” for providing instrumentation facilities. The work reported in this paper is also presented in a patent.20

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REFERENCES

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