Novel nanosensor of cobalt(II) and copper(II) constructed from graphene quantum dots modified with Eriochrome Black T

Detection of heavy transition metals is of importance protecting humans and the environment from their toxicity. Amongst them, copper(II) and cobalt(II) need more attention due to their wide applications in industry, in which exposures with excess amounts of them cause heathy concerns. Hence, fast and efficient distinguishing copper(II) and cobalt(II) could be worthy. While electrochemical approaches could determine these cations, expensive instrumentation and time-consuming analysis are significant drawbacks, leading to concentrations on spectroscopic and colorimetric pathways. Herein, graphene quantum dot was modified with Eriochrome Black T (EBT) to generate a novel sensor with the ability of sensing mentioned cations. This new nanocompound demonstrated new optical properties when complexed with cobalt(II) in natural pH, and copper(II) in acidic media. Both cations successfully were detected by the new nanosensor through revealing distinct spectroscopic signals. Moreover, cobalt(II) was distinguished by this sensor colorimetrical, appearing green solution. Linear ranges of cobalt(II) and copper(II) were obtained as 0.02–0.08 M and 0.015–0.2 M, with the limit of detections 0.014 and 0.018 M, respectively.


Scientific Reports
| (2022) 12:13179 | https://doi.org/10.1038/s41598-022-17616-y www.nature.com/scientificreports/ on the EBT removal from water wastes, and encouraged scientists to come up with the idea of binding EBT on a support 18 . Surprisingly, EBT deposited on various supports demonstrated novel application in detecting various organic and inorganic materials. For example, poly(benzopyrene) films doped with EBT as a Pb 2+ -sensitive sensor 19 , poly(EBT)-modified carbon paste an electrode for Hg 2+ distinguishing 20 , EBT/graphite composite for Theophylline determination 21 , and poly(EBT) on glassy carbon electrode for recognition of Isoniazid are some successful reports in this area 22 . Most of these detections were focused on electrochemical pathways, while we believe that EBT has significant optical potential on account of its extensive π-system. We recently demonstrated that the perching of Alizarine Red S on GQD generates a powerful sensor for detecting metal cations 23 . Easy, rapid, and cost-effective synthesis of GQD in combination with sustainability, and facile modification are some characteristics of this material that encouraged us to its modification with EBT [24][25][26] . Therefore, GQD was modified with EBT (GQD-EBT) to obtain a novel sensor of metal cations via esterification reaction (Scheme 1). This reaction efficiently was progressed by the N,N′-dicyclohexylcarbodiimide (DCC) and 4,4′-dimethylaminopyridine (DMAP) as the esterification agents 27,28 . The sensor successfully detected two significant metal cations, including Co 2+ and Cu 2+ , which their fast detections are not well-stablished till now [29][30][31] .

Experimental section
Materials and methods. All reagents were purchased from Sigma-Aldrich and used without further purification. TEM, and SEM micrographs were obtained by FEI Tecnai G2 F20 SuperTwin TEM, and FESEM TESCAN MIRA3, respectively. FT-IR of AVATAR FTIR instrument of Thermo Company in the range of 600-4000 cm −1 was used to determine functional groups of compounds. XRD of XRD Philips PW1730 was employed with the Cu k α (λ = 1.540598 nm), a voltage of 140 kV, and scanning from 10° to 80°. UV-Vis spectra were obtained by WPA Biowave LifeScience UV-Vis spectrometer using quartz cuvettes with a path length of 10 mm in H 2 O as the solvent.
Synthesis of GQD-EBT. GQD (2.0 g), DCC (0.4 g), and DMAP (0.03 g) were added to a round bottom flask containing 9 ml of dimethyl sulfoxide, and 1 ml of H 2 O. The resulted mixture was stirred magnetically at 90 °C for 10 min. Then, EBT (0.5 g) was added to the reaction mixture, and stirring was continued for 2 h. Next, the precipitate was filtered off, and the residue was washed with H 2 O (3 × 10 ml) to give GQD-EBT as a dark solid after drying at 60 °C.
General procedure for detecting M x+ . To 5 ml of an adjusted M x+ solution (pH = 3 with HCl and pH = 10 with NaOH) in the desired concentration, 0.05 g of GQD-EBT was added. Color changes were recorded as the addition of EBT-GQD. UV-Vis spectroscopies were carried out after calibration of the instrument with the standard solution of media.
Interfere study of cations on Co 2+ . GQD-EBT (0.05 g) was added to 10 ml of 0.02 M solution of Ca 2+ , Cu 2+ , Hg 2+ , Pb 2+ , Fe 3+ , Cd 2+ , Al 3+ , and Ni 2+ . Color changes were recorded as the addition of EBT-GQD. UV-Vis spectroscopy was carried out after calibration of the instrument with the standard media solution.

Results and discussion
Characterization of GQD-EBT. Organic transformations could be quickly approved by FT-IR spectroscopy, which highlights the addition, elimination, and transformation of functional groups with active vibrational modes. FT-IR spectra of GQD before and after modification indicated remarkable changes. Three prominent peaks of GQD appeared in both spectra, including stretching modes of OH, C=O, and C=C. The peak of C=O indicated a blueshift when GQD was modified with EBT due to the esterification reaction ( Fig. 1) 24 . Raman spectrum of GQD-EBT was also employed to characterize the structure with the ability to provide worthy information about the graphene-based structures. The spectrum of GQD-EBT demonstrated peaks of D-band at 1313 cm −1 attributed to the bending vibrations of C-H at the defects/edges and G-band at 1598  www.nature.com/scientificreports/ ascribed to in-plane C=C vibrations, respectively. The high intensity of the D-band compared to G-band is a reliable sign for the presence of abundant functional groups on GQD (Fig. 2). Also, a strong peak appeared at 1313 cm −1 for EBT relating to C=C bonds 32 . 1 H NMR spectroscopy was also employed to affirm the modification reaction, where H-aromatic of EBT in the GQD-EBT spectrum could confirm loading of the indicator on GQD. Spectrum of GQD-EBT indicated peaks for H-aromatic at about 7.0 and 8.2 ppm (Fig. 3), which did not observe in the GQD spectrum ( Figure S1). These peaks clearly confirm the loading of EBT on GQD.
Elemental analysis was conducted by Energy Dispersive X-ray Spectroscopy (EDX) on GQD-EBT to determine various elements in the structure (Fig. 4). X-ray detector of SEM instrument revealed C and O atoms of GQD with high intensities. The analysis also indicated atoms ascribed to EBT, including S, Na, and N. To disclose the EBT distribution on GQD surface, elemental mapping was carried out, in which homogeneous distribution of S, N, and Na atoms observed (Fig. 3). This homogeneity in the elements' dispersal implies success of the homogeneous sensor synthesis.
TEM micrographs of GQD were prepared before and after modification to see the structure and changes made during EBT loading on GQD (Fig. 5). TEM image of GQD showed the formation of nanoparticles in the range of 4-8 nm. The reaction time of citric acid transformation to graphene derivatives determines the dimensions of the synthesized particles with the fine particles in a shorter time 33   Detecting heavy metals by GQD-EBT. The optical behavior of GQD-EBT was studied by a spectrophotometer in the presence and absence of metal cations. GQD-EBT showed an absorption at 550 nm without any cation in a natural pH (Fig. 6). While the addition of GQD-EBT to most of the cations solutions (0.02 M) such as Ca 2+ , Cu 2+ , Hg 2+ , Pb 2+ , Fe 3+ , Cd 2+ , Al 3+ , and Ni 2+ did not create impressive changes in the absorption wavelength in the natural pH, charging to Co 2+ solution (0.02 M) led to the redshift of absorption to 514 nm (Fig. 6). Surprisingly, GQD-EBT could not spectroscopically or colorimetrical detect Ca 2+ opposite to EBT. Intriguingly, the new sensor distinguished Co 2+ colorimetrical by changing the brown GQD-EBT solution to green. The amount of GQD-EBT was optimized as 0.05 g, in which the best color detection of Co 2+ was obtained at the lowest amounts of the sensor. A standard addition technique for Co 2+ detection by GQD-EBT revealed a calibration curve of y = 1.80x + 0.12 with a linear range lying between 0.02-0.08 M. Limit of detection (LOD) was calculated as 0.014 M.
To elucidate the effect of solution pH on the detection ability of GQD-EBT, recognitions of the mentioned cations were carried out in two pHs of 10 and 3. As the first result, the detection of Co 2+ was lost in both acidic and basic conditions. However, the result of this experiment was fascinating for Cu 2+ in pH = 3, in which a new peak appeared at about 800 nm in addition to a peak of 550 nm (Fig. 7). A standard addition technique for Cu 2+ detection by GQD-EBT demonstrated a calibration curve of y = 3.0653x + 0.1314 with a linear range lying between 0.015-0.2 M and an LOD of 0.018 M. The colorimetric distinguishing of Cu 2+ was impossible due to the no detectable color change with the addition of the cation to the sensor solution.
Selectivity of the sensor was examined in a solution containing various cations at pH = 7, leading to the successful determination of Co 2+ by spectroscopic and colorimetric methods. The peak of Co 2+ /GQD-EBT was observed in the spectrum of the solution which gained deep green color, indicating noninterference of other cations. Investigation of the selectivity at pH = 3 was also showed the possibility of the Cu 2+ detection spectroscopically by observing the related peak at about 800 nm. Real samples were studied by GQD-EBT, with the various concentrations of Co 2+ and Cu 2+ in the tape and river water, in which both cations were successfully distinguished by the sensor after adjusting pH. Table 1 describes the results of detecting various concentrations of Co 2+ and Cu 2+ .
A remarkable number of sensors has been reported for the Co 2+ and Cu 2+ distinguishing, in which some of them indicated superior sensitivity and selectivity than GQD-EBT 23,30,31 . However, introducing new approach to the functionalizing GQD with an indicator such as EBT could be a promising pathway to afford a lot of new sensors via this protocol. Easy synthesis of GQD from inexpensive citric acid, high optical activity of GQD, facile GQD modification through carboxylic acid groups, useful changing in the GQD's optical properties due to modification with an indicator are some of significant features highlighted in this study. Therefore, synthesis -0. 0.5 1.5 2.5 3.5 4.5 5.5 6.5 7.5 f1 (ppm) NMR L1

Conclusion
In summary, a novel sensor was synthesized from GQD and EBT by the facile and efficient pathway of esterification reaction. GQD modified EBT demonstrated different performance in the detection of cations compared to EBT, such as inability in Ca 2+ detection. The new sensor successfully detected Co 2+ , and Cu 2+ in natural, and acidic solutions, respectively. Moreover, colorimetric detection of Co 2+ was also affordable with a green color induced by the sensor. This study offers that modifying GQD with optically active compounds could generate