Gamma-Ray-Induced Structural Transformation of GQDs towards the Improvement of Their Optical Properties, Monitoring of Selected Toxic Compounds, and Photo-Induced Effects on Bacterial Strains

Structural modification of different carbon-based nanomaterials is often necessary to improve their morphology and optical properties, particularly the incorporation of N-atoms in graphene quantum dots (GQDs). Here, a clean, simple, one-step, and eco-friendly method for N-doping of GQDs using gamma irradiation is reported. GQDs were irradiated in the presence of the different ethylenediamine (EDA) amounts (1 g, 5 g, and 10 g) and the highest % of N was detected in the presence of 10 g. N-doped GQDs emitted strong, blue photoluminescence (PL). Photoluminescence quantum yield was increased from 1.45, as obtained for non-irradiated dots, to 7.24% for those irradiated in the presence of 1 g of EDA. Modified GQDs were investigated as a PL probe for the detection of insecticide Carbofuran (2,2-Dimethyl-2,3-dihydro-1-benzofuran-7-yl methylcarbamate) and herbicide Amitrole (3-amino-1,2,4-triazole). The limit of detection was 5.4 μmol L−1 for Carbofuran. For the first time, Amitrole was detected by GQDs in a turn-off/turn-on mechanism using Pd(II) ions as a quenching agent. First, Pd(II) ions were quenched (turn-off) PL of GQDs, while after Amitrole addition, PL was recovered linearly with Amitrole concentration (turn-on). LOD was 2.03 μmol L−1. These results suggest that modified GQDs can be used as an efficient new material for Carbofuran and Amitrole detection. Furthermore, the phototoxicity of dots was investigated on both Gram-positive and Gram-negative bacterial strains. When bacterial cells were exposed to different GQD concentrations and illuminated with light of 470 nm wavelength, the toxic effects were not observed.


Introduction
Since they were first reported in 2008 by Ponomarenko and Geim [1], graphene quantum dots (GQDs) have been attracting attention in different scientific fields of research [2]. GQDs are 0D fragments of graphene sheets, with a diameter of less than 100 nm and The morphology of GQDs was analyzed using AFM. The concentration of all GQDs dispersions used for deposition was 0.28 mg mL −1 . Samples were prepared by spin-coating deposition (3500 rpm, 1 min) on mica and recorded using Quesant AFM (Agoura Hills, CA, USA) which was operating in the tapping mode, in the air, at room temperature. Standard silicon tip (NanoAndMore Gmbh, Wetzlar, Germany) was used with the constant force of 40 N m −1 . AFM tips that are used in these measurements had the following characteristics: radius minor than 10 nm, 17 µm height, set back of 15 µm, the half cone angle of 20 • ÷ 25 • along the cantilever axis, 25 • ÷ 30 • from the side, and 10 • at the apex. AFM images were analyzed using Gwyddion 2.58 software [36]. Due to the small diameter of GQDs, the accuracy of their lateral size was increased by applying the tip deconvolution. The real GQDs diameters were estimated using the following equation: r c = r (cos θ 0 + (cos 2 θ 0 + (1 + sin θ 0 )(−1 + (tan θ 0 /cos θ 0 ) + tan 2 θ 0 )) 1/2 ) r c is the AFM radius of a particle, r is the particle radius, and θ 0 is the mean half angle of the tip [37]. The histograms of diameter distribution were calculated using 3 different AFM images with the size of 25 × 25 µm 2 for each GQDs sample.

Dynamic Light Scattering and ξ Potential Measurement
DLS and ζ-potential measurements were acquired at an angle of 173 • with a Zetasizer Nano ZS instrument (Malvern, Herrenberg, Germany) equipped with a 633 nm He-Ne laser. Measurements were performed at a fixed position (4.65 mm) with an automatic attenuator and controlled temperature (20 • C). Before measurements, GQD dispersions (0.2 mg/mL) were sonicated with a tip sonicator for 30 min, in ice. For each sample, five independent measurements were averaged, and the number weighted size distribution of the particles was retrieved. Solvent-resistant micro cuvettes(ZEN0040) have been used for experiments G2(τ) = I(t + τ)I(t) / I (1) where τ is the lag time and the brackets represent the ensemble average. The G2(τ) can be related to the field autocorrelation function g 1 (t) through the Siegert relation G2(τ) = 1 + β|g1(t)| 2 (2) where β is an instrumental constant equal to 1 in our setup. ζ-potential of GQDs was calculated from the electrophoretic mobility using the Henry correction to Smoluchowski's equation, implemented in the Z-sizer data-analysis software (Zetaseizer Software 7.13, Malvern Panalytical, 2002 (Worcestershire, UK). For this purpose, 5 independent measurements at pH = 7 were averaged.

X-ray Photoelectron Spectroscopy (XPS)
XPS spectra of GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g were acquired using a PHI5000 VersaProbe II scanning microprobe (ULVAC-PHI, Inc.; Chigasaki, Japan), operating with a monochromatic Al K α source (1486.6 eV). Used X-ray beam had Ø 100 µm (25 W, 15 kV); low-resolution surveys were collected using pass energy (PE) of 117.000 eV, with a resolution of 1.0 eV; high-resolution scans were collected with a PE of 23.500 eV, with a resolution of 0.05 eV. Collected electrons were analyzed in FAT mode. The samples were prepared by drop-casting deposition over aluminum foil; during the data acquisition, the sample surface was kept at an angle of 45 • with respect to the analyzer; charge neutralization was used during all the experiments by means of both electrons and positive ions (Ar + ).

Photoluminescence (PL) Spectroscopy
To examine photoluminescent properties, samples were dispersed in the concentration of 0.075 mg mL −1 in ultrapure water. PL spectra were recorded using the HORIBA Jobin Yvon's Fluoromax-4 spectrometer (Horiba, Kyoto, Japan) at different excitation wavelengths (300, 320, 340, 360, 380, and 400 nm) in the range of 320-580 nm. Excitation and emission slits were 8 and 2 nm, respectively. Integration time was 0.5 s. All spectra were recorded at room temperature and atmospheric pressure. Relative photoluminescence quantum yields (PL QY) were determined by integrating the area under the fluorescence curves using Equation (3) 2 (3) where QY is photoluminescence quantum yield, F is the integrated area under the PL curve, A is the absorbance value, n is the refractive index of the solvent, while REF and GQDs are referred to as the used reference (Rhodamine B, PL QY is 31%) and GQDs samples.

Photo-Induced Antibacterial Activity
The antimicrobial activity of GQD-EDA samples has been evaluated against reference Gram-positive strain Staphylococcus aureus ATCC 29213, and Gram-negative reference strain Escherichia coli ATCC 35218. Bacterial suspensions were prepared in phosphate buffer (1 × PBS, phosphate saline buffer) to a final concentration of 108 CFU mL −1 .
The microdilution method was used to determine the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of the selected nanoparticles [38]. Two-fold serial dilution of the samples with Luria-Bertani (LB) medium was performed, and each well, except for the sterility control, was inoculated with 20 µL of bacterial suspensions (1 × 108 CFU/mL), reaching a final volume of 200 µL. GQD-EDA samples were tested in the concentration from 0.8-0.025 mg mL −1 . Besides a negative control, a sterility control was prepared. The 96-well microtiter plates were incubated for 16 h at 37 • C under blue light (470 nm, 15 W) at a distance of 20 cm from the light source. A light irradiance in the proximity of the sample was 19 mW cm −2 . After incubation, optical density (OD) was read on a spectrophotometer (Epoch microplate spectrophotometer, Agilent, Santa Clara, CA, USA). 22 µL of resazurin was added and the microtiter plates were incubated for an additional 2 h at 37 • C. According to the resazurin reaction, the lowest concentration without color change was defined as MIC. The lowest concentration that did not show bacterial growth after sub-culturing and incubation was defined as MBC. For CF detection, a series of concentrations from 0 to 100 µmol L −1 was made and mixed with water dispersion of GQDs-EDA/1 g. Samples were incubated for 24 h, to establish interactions between CF and GQDs. For 3-amino-1,2,4-triazole detection, water dispersion of GQDs-EDA/1 g was firstly mixed with 15 µmol L −1 of Pd 2+ ions. These mixtures were incubated for 2 min. After that, 3-amino-1,2,4-triazole in the concentration range of 0-150 µmol L −1 was added and incubated for 5 min.

Photoluminescence Measurement
PL spectra of GQDs-EDA/1 g in the presence of CF and 3-amino-1,2,4-triazole were obtained using HORIBA Jobin Yvon's Fluoromax-4 spectrometer at room temperature at 360 nm excitation wavelength. Excitation and emission slits were 8 and 2 nm. Integration time was 0.5 s.

Morphology Analysis of Gamma-Irradiated GQDs
In Figure 1, AFM images with height histograms of GQD-EDA/1 g (a), GQD-EDA/5 g (b), and GQD-EDA/10 g (c) are presented. All images show spherical, well-dispersed particles (indicated with arrows as dots) with few agglomerates presented in the samples GQDs-EDA/5 g and GQDs-EDA/10 g that are marked with blue arrows, in Figure 1b,c. Height distribution diagrams suggest that the largest number of particles in samples GQDs-EDA/1 g, GQDs-EDA/5 g, GQDs-EDA/10 g are 0.5-1, 2.5-3, and 1-1.5 nm high, respectively. The thickness of the individual GQDs indicates its lamellar structure Nanomaterials 2022, 12, 2714 6 of 23 has consisted of 3-5 graphene layers. Average heights of GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g were 1.25 ± 0.13 nm, 2.57 ± 0.39 nm, and 1.58 ± 0.45 nm, respectively. The histograms of diameter distributions are presented in the lower left corner of each large-scale AFM image (a-c), while the high-magnification images are presented in (g-i). The largest fraction of GQDs in samples GQD-EDA/1 g and GQD-EDA/5 g are those with a diameter of around 35 nm, while in the sample GQD-EDA/10 g, the largest fractions are those with a diameter of around 45 nm. The average values were calculated to be 35.5 ± 3.6, 39.3 ± 1.8, and 41.1 ± 3.8 nm for GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g, respectively.
3.1.1. Morphology Analysis of Gamma-Irradiated GQDs In Figure 1, AFM images with height histograms of GQD-EDA/1 g (a), GQD-EDA/5 g (b), and GQD-EDA/10 g (c) are presented. All images show spherical, well-dispersed particles (indicated with arrows as dots) with few agglomerates presented in the samples GQDs-EDA/5 g and GQDs-EDA/10 g that are marked with blue arrows, in Figure 1b,c. Height distribution diagrams suggest that the largest number of particles in samples GQDs-EDA/1 g, GQDs-EDA/5 g, GQDs-EDA/10 g are 0.5-1, 2.5-3, and 1-1.5 nm high, respectively. The thickness of the individual GQDs indicates its lamellar structure has consisted of 3-5 graphene layers. Average heights of GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g were 1.25 ± 0.13 nm, 2.57 ± 0.39 nm, and 1.58 ± 0.45 nm, respectively. The histograms of diameter distributions are presented in the lower left corner of each large-scale AFM image (a-c), while the high-magnification images are presented in (g-i). The largest fraction of GQDs in samples GQD-EDA/1 g and GQD-EDA/5 g are those with a diameter of around 35 nm, while in the sample GQD-EDA/10 g, the largest fractions are those with a diameter of around 45 nm. The average values were calculated to be 35.5 ± 3.6, 39.3 ± 1.8, and 41.1 ± 3.8 nm for GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g, respectively. The size distribution of GQDs was investigated using DLS and their surface charge was measured by zeta-potential (ζ). The results of DLS and ζ-potential are reported in Figures 2a and 2b, respectively. All the measured GQDs show a monomodal num- The size distribution of GQDs was investigated using DLS and their surface charge was measured by zeta-potential (ζ). The results of DLS and ζ-potential are reported in Figure 2a,b, respectively. All the measured GQDs show a monomodal number-weighted diameter distribution with mode values of 33.4 nm (GQD-EDA/10 g), 42.0 nm (GQDs-EDA/5 g), and 55.7 nm (GQDs-EDA/1 g). The ξ-potential plots are presented in Figure 2b and show that all investigated samples display values in the negative electric region. The following values obtained for GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g were −28.0 ± 1.6 mV, −32.8 ± 1.4, and −25.9 ± 0.9 mV, respectively. An increase of ξ-potential values of irradiated GQDs compared with p-GQDs (−34 eV) [35] was due to a change in the amount of negatively charged functional groups during gamma irradiation in the presence of EDA, probably due to the lower number of carboxyl functional groups.
in Figure 2b and show that all investigated samples display values in the negative electric region. The following values obtained for GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g were −28.0 ± 1.6 mV, −32.8 ± 1.4, and −25.9 ± 0.9 mV, respectively. An increase of ξ-potential values of irradiated GQDs compared with p-GQDs (−34 eV) [35] was due to a change in the amount of negatively charged functional groups during gamma irradiation in the presence of EDA, probably due to the lower number of carboxyl functional groups.

Structural Characterization of GQDs
In Figure 3, FTIR spectra of p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are shown. The band at 3417 cm −1 located in all spectra stems from the stretching vibrations of O-H bonds in hydroxyl groups attached to carbon atoms. A new band at 3250 cm −1 in GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g is assigned to N-H bonds in amino groups [35]. The bands at 2970, about 2930, and 2870 cm −1 in pristine and irradiated GQDs are attributed to stretching vibrations of C-H bonds in -CH, and -CH2 groups [35]. The band located at 1633 cm −1 in the spectra of all samples steams from stretching vibrations of C=О bonds in amide functional groups [22]. The high-intensity band at 1572 cm −1 detected in the spectrum of the p-GQDs is assigned to stretching vibrations of the C=C bonds in π-conjugated domains of GQDs [35]. In the spectra of irradiated GQDs, this band is slightly shifted at 1567 cm −1 . This change can be attributed to defect formation in the sp 2 region of the GQD structure formed due to irradiation. A new band at 1458 cm −1 appeared in the spectra of GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g arises from C-N bending in N-C=О functional groups [39]. The bands centered at 1330 cm −1 in the spectra of p-GQDs, and GQDs-EDA/5 g, at 1339 cm −1 in the spectra of GQDs-EDA/1 g and GQDs-EDA/10 g, as well as the high-intensity band at 1395 cm −1 found in all spectra, originate from vibrations of the C-OH bonds [40,41]. The bands at 1161 cm −1 observed in all spectra and the one around 1088 cm −1 found in the spectra of p-GQDs, GQD-EDA/1 g, and GQDs-EDA/10 g are assigned to asymmetric [42] and symmetric stretching vibrations of the C-O bonds in epoxy groups [22,43].

Structural Characterization of GQDs
In Figure 3, FTIR spectra of p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are shown. The band at 3417 cm −1 located in all spectra stems from the stretching vibrations of O-H bonds in hydroxyl groups attached to carbon atoms. A new band at 3250 cm −1 in GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g is assigned to N-H bonds in amino groups [35]. The bands at 2970, about 2930, and 2870 cm −1 in pristine and irradiated GQDs are attributed to stretching vibrations of C-H bonds in -CH, and -CH 2 groups [35]. The band located at 1633 cm −1 in the spectra of all samples steams from stretching vibrations of C=O bonds in amide functional groups [22]. The high-intensity band at 1572 cm −1 detected in the spectrum of the p-GQDs is assigned to stretching vibrations of the C=C bonds in π-conjugated domains of GQDs [35]. In the spectra of irradiated GQDs, this band is slightly shifted at 1567 cm −1 . This change can be attributed to defect formation in the sp 2 region of the GQD structure formed due to irradiation. A new band at 1458 cm −1 appeared in the spectra of GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g arises from C-N bending in N-C=O functional groups [39]. The bands centered at 1330 cm −1 in the spectra of p-GQDs, and GQDs-EDA/5 g , at 1339 cm −1 in the spectra of GQDs-EDA/1 g and GQDs-EDA/10 g, as well as the high-intensity band at 1395 cm −1 found in all spectra, originate from vibrations of the C-OH bonds [40,41]. The bands at 1161 cm −1 observed in all spectra and the one around 1088 cm −1 found in the spectra of p-GQDs, GQD-EDA/1 g , and GQDs-EDA/10 g are assigned to asymmetric [42] and symmetric stretching vibrations of the C-O bonds in epoxy groups [22,43].
According to FTIR spectra, all GQDs contain: -OH, -CH 2, -CH, -COOH, C=C, and C-O-C functional groups. In N-doped gamma-irradiated GQDs, N-C=O and -NH functional groups were found. These results indicate the incorporation of N atoms in the structure of GQDs using a gamma irradiation dose of 200 kGy.
Structural properties of irradiated GQDs also were examined using XPS. The analysis of the C 1s region of XPS spectra showed the presence of C-C/C-H (284.80 eV, used as energy reference for all spectra)        Table 1. Atomic % of C, O, and N, and functional groups ratios in the samples p-GQDs, GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g. The analysis of the N 1s region showed the occurrence of (at least) two main components, namely at 399.52 eV and 401.24 eV (Figure 5a-c). Both chemical environments are compatible with nitrogen in an organic matrix (organic carbon-bound nitrogen). Atomic % of C, O, and N in the samples p-GQDs, GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g obtained by XPS analysis are presented in Table 1. The first observed change is an increase of carbon content in all irradiated samples, due to the reduction of the part of the oxygen-containing groups during gamma irradiation. Secondly, the oxygen content is significantly decreased in irradiated samples. This change could be attributed to the transformation of O-C=O to N-C=O groups in the irradiation conditions. In line with that, the content of carboxyl groups decreases, while the content of amide groups became higher with the increase in EDA weight. These results are in agreement with ζ-potentials analysis, which showed that gamma irradiation leads to the formation of lees negatively charged particles.

Sample
%С %О %N p-GQDs [35] 63.00 37.00 0.00 GQDs-EDA/1 g 76. 25 17.36 6.39 GQDs-EDA/5 g 77. 18 16.10 6.71 GQDs-EDA/10 g 76. 60 16.35 7.05 Results obtained by 1 H NMR recording of GQDs-EDA/1 g ( Figure 6, black line), GQDs-EDA/5 g ( Figure 6, red line), GQDs-EDA/10 g ( Figure 6, blue line) show that spectral features are similar for different samples. Signals between 1.00 and 2.50 ppm can be ascribed to the presence of alkyl protons (-CH3, -CH2-). The sharp, intense signal localized between 2.40-2.45 ppm could be attributed to methyl (or methylene) groups connected to electron-withdrawing moieties. Spectral features between 3.00 and 3.50 ppm could be compatible with -CH2-groups bearing an -OH moiety but are less evident in the spectrum recorded for sample GQDs-EDA/10 g. There are low intensity, sharp signals between 6.50-7.50 ppm, which are related to amide groups. However, the presence of amino groups is not excluded, but the typical broad bands at the low field are barely visible due to the limited amount of solubilized material and the weak intensity of the whole spectral profile. Atomic % of C, O, and N in the samples p-GQDs, GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g obtained by XPS analysis are presented in Table 1. The first observed change is an increase of carbon content in all irradiated samples, due to the reduction of the part of the oxygen-containing groups during gamma irradiation. Secondly, the oxygen content is significantly decreased in irradiated samples. This change could be attributed to the transformation of O-C=O to N-C=O groups in the irradiation conditions. In line with that, the content of carboxyl groups decreases, while the content of amide groups became higher with the increase in EDA weight. These results are in agreement with ζ-potentials analysis, which showed that gamma irradiation leads to the formation of lees negatively charged particles.
It was calculated that carboxyl groups were 3.76, 2.94, and 2.53% for GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. The content of amide bonds was similar for all samples, from 6.38 to 6.25, and 7.16%. In the case of C-O, C1s data fitting showed that C-O bonds were 23.54, 20.92, and 21.41% for GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. The bend that was assigned to C-C/C-H showed that samples contained from 66.32 to 69.90%.
Results obtained by 1 H NMR recording of GQDs-EDA/1 g ( Figure 6, black line), GQDs-EDA/5 g ( Figure 6, red line), GQDs-EDA/10 g ( Figure 6, blue line) show that spectral features are similar for different samples. Signals between 1.00 and 2.50 ppm can be ascribed to the presence of alkyl protons (-CH 3 , -CH 2 -). The sharp, intense signal localized between 2.40-2.45 ppm could be attributed to methyl (or methylene) groups connected to electron-withdrawing moieties. Spectral features between 3.00 and 3.50 ppm could be compatible with -CH 2 -groups bearing an -OH moiety but are less evident in the spectrum recorded for sample GQDs-EDA/10 g. There are low intensity, sharp signals between 6.50-7.50 ppm, which are related to amide groups. However, the presence of amino groups is not excluded, but the typical broad bands at the low field are barely visible due to the limited amount of solubilized material and the weak intensity of the whole spectral profile.
Results obtained by FTIR, XPS, and NMR spectroscopies confirmed that gammairradiated GQDs possess C, O, N, and H in their structure. Obtained dots have a larger amount of C in their structure compared to p-GQDs (63% for p-GQDs and around 76% for gamma irradiated), which indicated a chemical reduction of oxygen-containing functional groups. This treatment also induced the incorporation of amino functional groups and the formation of amide bonds. Nanomaterials 2022, 12, x FOR PEER REVIEW 10 of 24 Results obtained by FTIR, XPS, and NMR spectroscopies confirmed that gamma-irradiated GQDs possess C, O, N, and H in their structure. Obtained dots have a larger amount of C in their structure compared to p-GQDs (63% for p-GQDs and around 76% for gamma irradiated), which indicated a chemical reduction of oxygen-containing functional groups. This treatment also induced the incorporation of amino functional groups and the formation of amide bonds.

Optical Properties of GQDs
The optical properties of GQDs were examined using UV-Vis Vis and PL spectroscopies. Absorption spectra of the samples p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are presented in Figure 7. For all samples, the high absorption band at about 240 nm is detected. This band is attributed to the π-π* transition of C-C aromatic domains. A weak shoulder band at 270 nm observed in the spectrum of p-GQDs indicates n-π* transitions in C=O bonds. This band is more pronounced and shifted at 279, and 282 nm in the GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. The shift in this region can be explained by changes in the chemical environment around carbonyl groups in the samples due to functionalization. In Figure 8, 3D emission spectra of p-GQDs (a), GQD-EDA/1 g (b), GQD-EDA/5 g (c), and GQD-EDA/10 g (d) were recorded at different excitation wavelength in the range of 300-400 nm are presented. First, p-and irradiated GQDs demonstrate excitation-depended PL, as observed previously [3,44]. This behavior is more pronounced in

Optical Properties of GQDs
The optical properties of GQDs were examined using UV-Vis Vis and PL spectroscopies. Absorption spectra of the samples p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are presented in Figure 7. For all samples, the high absorption band at about 240 nm is detected. This band is attributed to the π-π* transition of C-C aromatic domains. A weak shoulder band at 270 nm observed in the spectrum of p-GQDs indicates n-π* transitions in C=O bonds. This band is more pronounced and shifted at 279, and 282 nm in the GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. The shift in this region can be explained by changes in the chemical environment around carbonyl groups in the samples due to functionalization. Results obtained by FTIR, XPS, and NMR spectroscopies confirmed that gamma-irradiated GQDs possess C, O, N, and H in their structure. Obtained dots have a larger amount of C in their structure compared to p-GQDs (63% for p-GQDs and around 76% for gamma irradiated), which indicated a chemical reduction of oxygen-containing functional groups. This treatment also induced the incorporation of amino functional groups and the formation of amide bonds.

Optical Properties of GQDs
The optical properties of GQDs were examined using UV-Vis Vis and PL spectroscopies. Absorption spectra of the samples p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are presented in Figure 7. For all samples, the high absorption band at about 240 nm is detected. This band is attributed to the π-π* transition of C-C aromatic domains. A weak shoulder band at 270 nm observed in the spectrum of p-GQDs indicates n-π* transitions in C=O bonds. This band is more pronounced and shifted at 279, and 282 nm in the GQD-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. The shift in this region can be explained by changes in the chemical environment around carbonyl groups in the samples due to functionalization. In Figure 8, 3D emission spectra of p-GQDs (a), GQD-EDA/1 g (b), GQD-EDA/5 g (c), and GQD-EDA/10 g (d) were recorded at different excitation wavelength in the range of 300-400 nm are presented. First, p-and irradiated GQDs demonstrate excitation-depended PL, as observed previously [3,44]. This behavior is more pronounced in Figure 7. UV-Vis spectra of p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g.
In Figure 8, 3D emission spectra of p-GQDs (a), GQD-EDA/1 g (b), GQD-EDA/5 g (c), and GQD-EDA/10 g (d) were recorded at different excitation wavelength in the range of 300-400 nm are presented. First, p-and irradiated GQDs demonstrate excitation-depended PL, as observed previously [3,44]. This behavior is more pronounced in p-GQDs than in N-doped gamma-irradiated samples. As can be seen in Table 2, the emission peak maximum of p-GQDs is moved from 430 to 492 nm, while in GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g, these shifts were from 441 to 465, from 438 to 465, and from 439 to 466 nm, at the excitation wavelength in the range of 300-400 nm. Excitation-depended PL may result from differently sized dots (according to quantum confinement effect with the increase of particle size, band gap decreases), or different surface chemistry. GQDs surface is rich in trap states such as O-functional groups (-COOH,-OH, and C-O-C), the dangling bonds, and sp 2 /sp 3 C atoms [45]. Generally, surface trap states are functional groups, oxygen-related disorder-induced localized states, and surface defects [45]. One study suggested that GQD optical emissions stem most probably from continuous defect states (functional groups), and different defect states are the reason for variation in emissions at different wavelengths [46]. Less dependence on the emission peak position from excited light in irradiated GQDs is attributed to the higher uniformity of size and surface state of these particles [47].
p-GQDs than in N-doped gamma-irradiated samples. As can be seen in Table 2, the emission peak maximum of p-GQDs is moved from 430 to 492 nm, while in GQD-EDA/1 g, GQD-EDA/5 g, and GQD-EDA/10 g, these shifts were from 441 to 465, from 438 to 465, and from 439 to 466 nm, at the excitation wavelength in the range of 300-400 nm. Excitation-depended PL may result from differently sized dots (according to quantum confinement effect with the increase of particle size, band gap decreases), or different surface chemistry. GQDs surface is rich in trap states such as O-functional groups (-COOH,-OH, and C-O-C), the dangling bonds, and sp 2 /sp 3 C atoms [45]. Generally, surface trap states are functional groups, oxygen-related disorder-induced localized states, and surface defects [45]. One study suggested that GQD optical emissions stem most probably from continuous defect states (functional groups), and different defect states are the reason for variation in emissions at different wavelengths [46]. Less dependence on the emission peak position from excited light in irradiated GQDs is attributed to the higher uniformity of size and surface state of these particles [47]. According to the CIE 1931 chromaticity diagrams, the emission of p-GQDs ( Figure  9a) varies from violet-like blue to turquoise part of the spectrum, while all N-doped GQDs exhibit intense blue emission (Figure 9a-c). These diagrams indicate that the position of the emission band center for gamma-irradiated GQDs is less dependent than in the case of p-GQDs. GQDs-EDA/1 g to 10 g still show excitation-dependent PL emission. PL properties of GQDs are controlled by intrinsic and extrinsic factors [48]. It is proposed that the blue emission originated from ordered sp 2 domains in GQDs (intrinsic state emission) [48]. Additionally, intensive blue emission was reported for N-doped GQDs with amide groups [49]. In this case, control factors for strictly blue emission should be both the surface state and graphene core.  According to the CIE 1931 chromaticity diagrams, the emission of p-GQDs (Figure 9a) varies from violet-like blue to turquoise part of the spectrum, while all N-doped GQDs exhibit intense blue emission (Figure 9a-c). These diagrams indicate that the position of the emission band center for gamma-irradiated GQDs is less dependent than in the case of p-GQDs. GQDs-EDA/1 g to 10 g still show excitation-dependent PL emission. PL properties of GQDs are controlled by intrinsic and extrinsic factors [48]. It is proposed that the blue emission originated from ordered sp 2 domains in GQDs (intrinsic state emission) [48]. Additionally, intensive blue emission was reported for N-doped GQDs with amide groups [49]. In this case, control factors for strictly blue emission should be both the surface state and graphene core. In terms of PL intensity, the highest emission band intensity for p-GQDs was recorded when the excitation wavelength was 340 nm, while GQDs-EDA/1 g at 360 nm, and GQDs-EDA/5 g and GQDs-EDA/10 g, were observed when excited with 380 nm. The next observed change in PL spectra of irradiated samples is an increase of intensity at all excitation wavelengths compared to p-GQDs, whereby maximum value was recorded in the PL spectrum of GQDs-EDA/5 g (Table 2).  In terms of PL intensity, the highest emission band intensity for p-GQDs was recorded when the excitation wavelength was 340 nm, while GQDs-EDA/1 g at 360 nm, and GQDs-EDA/5 g and GQDs-EDA/10 g, were observed when excited with 380 nm. The next observed change in PL spectra of irradiated samples is an increase of intensity at all excitation wavelengths compared to p-GQDs, whereby maximum value was recorded in the PL spectrum of GQDs-EDA/5 g (Table 2).
In Table 3, the values of PL QY are presented. These data were calculated using Equation (3) and PL spectra recorded at the excitation wavelength of 360 nm. As a reference, Rhodamine B was used. Relative PL QY of p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are 1.45, 7.24, 6.77, and 5.82%. Obtained results show that the introduction of N-atoms in GQDs structure led to PL QY enhancement, whereby the highest value was found for GQDs-EDA/1 g. Table 3. The values of relative QY at the excitation wavelength of 360 nm.

Sample QY 360 (%)
p-GQDs [35] 1.45 GQDs-EDA/1 g 7.24 GQDs-EDA/5 g 6.77 GQDs-EDA/10 g 5.82 Rhodamine B 31 In Figure 10, the effects of pH and ionic strength on the intensity of emission bands are presented. Only very low (such as 1) and very high (12)  In Table 3, the values of PL QY are presented. These data were calculated using Equation (3) and PL spectra recorded at the excitation wavelength of 360 nm. As a reference, Rhodamine B was used. Relative PL QY of p-GQDs, GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g are 1.45, 7.24, 6.77, and 5.82%. Obtained results show that the introduction of N-atoms in GQDs structure led to PL QY enhancement, whereby the highest value was found for GQDs-EDA/1 g. Table 3. The values of relative QY at the excitation wavelength of 360 nm.

Sample
QY360 (%) p-GQDs [35] 1.45 GQDs-EDA/1 g 7.24 GQDs-EDA/5 g 6.77 GQDs-EDA/10 g 5.82 Rhodamine B 31 In Figure 10, the effects of pH and ionic strength on the intensity of emission bands are presented. Only very low (such as 1) and very high (12) pH values caused the small lowering in the intensity of emission bands for all three samples. Thus, it can be concluded that PL emission of gamma irradiated GQDs is stable in the pH range 2-10. PL emission was also stable in the presence of different NaCl concentrations, from 15 to 1275 mM.  Figure 11a shows PL spectra of GQDs-EDA/1 g in the presence of CF concentration from 0 to 100 µmol L −1 . It can be seen that PL intensity increases with increasing CF concentration. The linear relationship between PL intensity was determined using equation A/A 0 = 1 + K SV [Q], where A and A 0 are integrated areas under the emission peak in the presence and absence of CF, respectively, K SV is Stern-Volmer constant and corresponds to the slope obtained by a linear fit of experimental data presented in Figure 10b, while Q is the concentration of the analyte. Linear relationship is described as: A/A 0 = 1.04757 + 0.00173 [CF]. A good linear response (R 2 -coefficient of determination = 0.98) was found within a wide concentration range (15-100 µmol L −1 ). LOD was calculated using the equation LOD = 3S D /K sv , where S D represents the standard deviation of intercept [27]. Obtained LOD was 5.4 µmol L −1 . In the spectra, a significant shift can be seen of emission peak maximum with increasing in CF concentration (from 442 nm at 0 to 455 nm at 100 µmol L −1 ).
Based on these results, we examine the detection of 3-amino-1,2,4-triazole in the concentration range of 0-12 μmol L −1 (Figure 13a). It can be observed that at these concentrations of pesticide, a PL of GQDs/Pd 2+ system was recovered linearly in the whole tested concentration range (Figure 13b). Linearity between probe PL intensity and 3-amino-1,2,4-triazole concentration was presented as: A/A0 = 0.9787 + 0.00848 [triazole]. In this case, R 2 and LOD were 0.96 and 2.03 μmol L −1 , respectively. Presented results showed that tested PL probe GQDs-EDA/1 g can be applied for direct turn-on detection of CF, and turn-off/turn-on detection of 3-amino-1,2,4-triazole using Pd 2+ ions as quenching agents.

Detection Mechanism
The mechanism of interaction between CF and 3-amino-1,2,4-triazole is resolved using FTIR spectroscopy. In Figure 14a, FTIR spectra of CF, GQDs-EDA/1 g, and Presented results showed that tested PL probe GQDs-EDA/1 g can be applied for direct turn-on detection of CF, and turn-off/turn-on detection of 3-amino-1,2,4-triazole using Pd 2+ ions as quenching agents.

Detection Mechanism
The mechanism of interaction between CF and 3-amino-1,2,4-triazole is resolved using FTIR spectroscopy. In Figure 14a, FTIR spectra of CF, GQDs-EDA/1 g, and GQDs-EDA/1 g-CF are presented. In the FTIR spectrum of CF, bands at 1231 and 1264 cm −1 are attributed to stretching vibrations of C-O-C, where one C atom is attached to the aromatic ring, while the other is bonded to the aliphatic structure [51]. A high-intensity band at 1717 cm −1 is assigned to the bending vibration of C=O groups [51]. The bands in the range 2882-3000 cm −1 correspond to symmetric and asymmetric stretching of -CH 2 and -CH 3 moieties [51]. At 1334 and 3363 cm −1 , bands are assigned to stretching vibrations of C-N, and N-H groups, respectively [51,52].
FTIR spectra of GQDs-EDA/1 g + CF and GQDs-EDA/1 g are compared and presented in Figure 14b. First, the band at 3417 cm −1 in the untreated GQDs spectrum from O-H bonds is shifted to 3422 cm −1 after CF addition. A small shift in the hydroxyl group band is probably due to hydrogen bond formation between CF and the functional groups at the GQDs surface [52]. A very weak band at around 1630 cm −1 observed in the GQDs-EDA/1 g is assigned to the carboxyl functional group. After treatment with CF, this band became defined and more pronounced. That change can be attributed to O-H bending in hydrogen bonds established between CF molecules and carboxyl groups of GQDs [53]. High-intensity bands at 1088 cm −1 and 1395 cm −1 found in the GQDs-EDA/1 g spectrum from C-O bonds in epoxy groups and C-OH became weaker and they almost disappeared from the spectrum when CF was added to the system. Their lowering may indicate the dissociation of such bonds in the binding mechanism of carbofuran to GQDs [54]. this band became defined and more pronounced. That change can be attributed to O-H bending in hydrogen bonds established between CF molecules and carboxyl groups of GQDs [53]. High-intensity bands at 1088 cm −1 and 1395 cm −1 found in the GQDs-EDA/1 g spectrum from C-O bonds in epoxy groups and C-OH became weaker and they almost disappeared from the spectrum when CF was added to the system. Their lowering may indicate the dissociation of such bonds in the binding mechanism of carbofuran to GQDs [54]. Carbon materials such as sugarcane bagasse biochar [52], and others [53][54][55], adsorb CF from the water via hydrogen bonds. CF at pH > 3.78 acts as an H-acceptor, while -OH groups of biochar play the role of an H-donor. Additionally, it was reported that the N-H group of CF are donated protons and created H-bond with oxygen functional groups of the biochar [52]. According to the FTIR spectrum of the GQDs-EDA/1 g-CF, shifts in the bands assigned to O-H, -COOH, -C-O, and -C-OH groups can be assigned to the formation of hydrogen bonds between CF molecules and O-functional groups of GQDs. Considering a ξ-potential result, carboxyl functional groups on GQDs surface are deprotonated at pH 7, so CF probably served as an H-bond donor in complex with GQDs (Scheme 1). Carbon materials such as sugarcane bagasse biochar [52], and others [53][54][55], adsorb CF from the water via hydrogen bonds. CF at pH > 3.78 acts as an H-acceptor, while -OH groups of biochar play the role of an H-donor. Additionally, it was reported that the N-H group of CF are donated protons and created H-bond with oxygen functional groups of the biochar [52]. According to the FTIR spectrum of the GQDs-EDA/1 g-CF, shifts in the bands assigned to O-H, -COOH, -C-O, and -C-OH groups can be assigned to the formation of hydrogen bonds between CF molecules and O-functional groups of GQDs. Considering a ξ-potential result, carboxyl functional groups on GQDs surface are deprotonated at pH 7, so CF probably served as an H-bond donor in complex with GQDs (Scheme 1). This binding mechanism is explaining the increase of GQDs PL intensity. In several previous studies, it was found that hydrogen bonds formed between carbon dots and pesticide atrazine [56], B,N,S-doped GQDs and glucosamine [57], and B-doped GQDs and glucose [58] lead to the increase in PL intensity due to structural rigidification. In a rigid structure, intermolecular motions are restricted which caused the blocking of non-radiative channels. Relaxation of excitons was directed through radiative channels increasing GQDs PL intensity. Interaction of GQDs functional groups with CF caused the shift of the emission band.
This binding mechanism is explaining the increase of GQDs PL intensity. In several previous studies, it was found that hydrogen bonds formed between carbon dots and pesticide atrazine [56], B,N,S-doped GQDs and glucosamine [57], and B-doped GQDs and glucose [58] lead to the increase in PL intensity due to structural rigidification. In a rigid structure, intermolecular motions are restricted which caused the blocking of non-radiative channels. Relaxation of excitons was directed through radiative channels increasing GQDs PL intensity. Interaction of GQDs functional groups with CF caused the shift of the emission band.
In Figure 15a, an FTIR spectrum of 3-amino-1,2,4-triazole is presented. The bands at 3083 cm −1 and 3054 cm −1 are assigned to C-H aromatic vibrations [59]. A strong band at 3211 cm −1 is attributed to stretching vibrations of N-H bonds in amino groups [59], while those at 1531 and 1472 cm −1 originated from C=C stretching vibrations of aromatic domains in 3-amino-1,2,4-triazole molecules [59]. High-intensity bands located at 1595 and 1045 cm −1 indicate the presence of endocyclic N=N and C-N-C, respectively [60]. After the addition of 3-amino-1,2,4 triazole, the band at 3417 cm −1 is fully recovered, due to releasing of -OH groups from the complex with Pd 2+ . The band at 3250 cm −1 is also recovered and moved at 3150 cm −1 . The bands at around 2970 and 2930 cm −1 are also partially rehabilitated. The one at 1630 cm −1 is completely released from the complex and moved at 1691 cm −1 . The band assigned to the C=C bond is not fully recovered as well as not returned to the previous position, which proved incomplete regeneration of the π-conjugated system of GQDs. After introducing a 3-amino-1,2,4 triazole in the GQDs-EDA/1 g-Pd 2+ system, bands at 1458, 1395, and 1088 cm −1 are not located. These changes indicate that there is no re-establishment of the C-N and C-O bonds. Results obtained by FTIR analysis confirmed a proposed mechanism of 3-amino-1,2,4 triazole detection whereas PL of GQDs was quenched in the presence of Pd 2+ ion, after the addition of pesticide was restored (Scheme 2).

Photo-Induced Antibacterial Activity
In order to explore a possible application of amide functionalized GQDs in antibacterial photodynamic therapy, we treated two representative Gram-positive and Gram-negative bacterial lines, Streptococcus aureus (S. aureus) and Escherichia coli (E. coli), respectively, with different concentrations of irradiated GQDs and exposed them to blue lights for 16 h. Our previous study showed that p-GQDs induced bacterial cell death at a concentration of 125.00 mg mL −1 for S. aureus, and 62.5 mg mL −1 for E. coli [68]. In the same study, it was found that carbon quantum dots with amino functional groups (CQDNH) caused the death of S. aureus at 62.5 mg mL −1 and E. coli at 31.25 mg mL −1 . In the tested range of concentrations (0.8 to 0.025 mg mL −1 ), GQD-EDA/1 g and GQD-EDA/10 g did not show antimicrobial activity (Table 4). In Figure 15b, FTIR spectra of GQDs-EDA/1 g, GQDs-EDA/1 g-Pd 2+ , and GQDs-EDA/1 g-Pd 2+ -3-amino-1,2,4-triazole are presented. To explore the mechanism of 3-amino-1,2,4-triazole detection using Pd 2+ ions, firstly FTIR spectrum of GQDs-EDA/1 g in the presence of ions is analyzed. By comparing the spectrum of GQDs and the one recorded in the presence of Pd 2+ ions, several changes are observed. First, the high-intensity band at 3417 cm −1 is slightly reduced and shifted to 3442 cm −1 due to complex formation [61]. Bands at 2970, 2930, and 2870 cm −1 in irradiated GQDs attributed to stretching vibrations of C-H bonds in -CH and -CH 2 groups completely disappeared after the addition of Pd 2+ ions. Band located at 3250 cm −1 attributed to stretching vibration of N-H group is also reduced in the presence of Pd 2+ ions as a consequence of their coordination with amino groups [62]. The band at 1633 is moved to 1712 cm −1 and became more pronounced due to the metal-ligand complex formed between the carboxyl group and Pd 2+ [61]. The band at 1567 assigned to the C=C bond is shifted at 1634 cm −1 which confirmed cation-π interactions [63,64]. The band at 1458 cm −1 attributed to C-N vibrations of amide groups in GQDs is reduced and moved at 1496 cm −1 due to the building of a complex, where N atoms play the role of electron donor [65]. Another change is observed: the band at 1395 associated with C-O is reduced and moved at 1404 cm −1 due to surface complexation [66,67]. Additionally, the band at 1088 cm −1 attributed to C-OH vibrations completely vanished.
After the addition of 3-amino-1,2,4 triazole, the band at 3417 cm −1 is fully recovered, due to releasing of -OH groups from the complex with Pd 2+ . The band at 3250 cm −1 is also recovered and moved at 3150 cm −1 . The bands at around 2970 and 2930 cm −1 are also partially rehabilitated. The one at 1630 cm −1 is completely released from the complex and moved at 1691 cm −1 . The band assigned to the C=C bond is not fully recovered as well as not returned to the previous position, which proved incomplete regeneration of the π-conjugated system of GQDs. After introducing a 3-amino-1,2,4 triazole in the GQDs-EDA/1 g-Pd 2+ system, bands at 1458, 1395, and 1088 cm −1 are not located. These changes indicate that there is no re-establishment of the C-N and C-O bonds. Results obtained by FTIR analysis confirmed a proposed mechanism of 3-amino-1,2,4 triazole detection whereas PL of GQDs was quenched in the presence of Pd 2+ ion, after the addition of pesticide was restored (Scheme 2). not returned to the previous position, which proved incomplete regeneration of the π-conjugated system of GQDs. After introducing a 3-amino-1,2,4 triazole in the GQDs-EDA/1 g-Pd 2+ system, bands at 1458, 1395, and 1088 cm −1 are not located. These changes indicate that there is no re-establishment of the C-N and C-O bonds. Results obtained by FTIR analysis confirmed a proposed mechanism of 3-amino-1,2,4 triazole detection whereas PL of GQDs was quenched in the presence of Pd 2+ ion, after the addition of pesticide was restored (Scheme 2).

Photo-Induced Antibacterial Activity
In order to explore a possible application of amide functionalized GQDs in antibacterial photodynamic therapy, we treated two representative Gram-positive and Gram-negative bacterial lines, Streptococcus aureus (S. aureus) and Escherichia coli (E. coli), respectively, with different concentrations of irradiated GQDs and exposed them to blue lights for 16 h. Our previous study showed that p-GQDs induced bacterial cell death at a concentration of 125.00 mg mL −1 for S. aureus, and 62.5 mg mL −1 for E. coli [68]. In the same study, it was found that carbon quantum dots with amino functional groups (CQDNH) caused the death of S. aureus at 62.5 mg mL −1 and E. coli at 31.25 mg mL −1 . In the tested range of concentrations (0.8 to 0.025 mg mL −1 ), GQD-EDA/1 g and GQD-EDA/10 g did not show antimicrobial activity (Table 4).

Photo-Induced Antibacterial Activity
In order to explore a possible application of amide functionalized GQDs in antibacterial photodynamic therapy, we treated two representative Gram-positive and Gram-negative bacterial lines, Streptococcus aureus (S. aureus) and Escherichia coli (E. coli), respectively, with different concentrations of irradiated GQDs and exposed them to blue lights for 16 h. Our previous study showed that p-GQDs induced bacterial cell death at a concentration of 125.00 mg mL −1 for S. aureus, and 62.5 mg mL −1 for E. coli [68]. In the same study, it was found that carbon quantum dots with amino functional groups (CQDNH) caused the death of S. aureus at 62.5 mg mL −1 and E. coli at 31.25 mg mL −1 . In the tested range of concentrations (0.8 to 0.025 mg mL −1 ), GQD-EDA/1 g and GQD-EDA/10 g did not show antimicrobial activity (Table 4). Table 4. Minimum inhibitory concentrations of GQD-EDA/1 g and GQDs-EDA/10 g (in mg mL −1 ). In parallel, for the sample GQD-EDA/1 g, OD values were obtained to determine whether there was a reduction of OD values in the negative control (Table 5). It was observed that a higher optical density was observed in the treatment compared to the control. Therefore, this method did not show to be reliant on assessing antimicrobial activity. Therefore, the results of antimicrobial activity were read exclusively by resazurin reaction and reinoculation to the appropriate nutrient medium. Although our previous studies showed the ability of GQDs to induce bacterial cell death when they were exposed to blue light [68], these experiments revealed that GQDs with both amide and amino functional groups lost their phototoxic activity. Our previous studies revealed that GQDs are very potent singlet oxygen produced [8,[68][69][70]. Thanks to this ability, together with small size and fast internalization, dots induced oxidative stress in bacterial cells. Here, the introduction of amide and amino groups probably resulted in the inability of dots to transfer the energy toward molecular oxygen. Studies presented in Table 6 showed that amino functionalized and N-doped GQDs are able to induce death of various bacterial strains under conditions presented in the table. Table 6. Studies regarding GQDs with N in structure and its antibacterial photodynamic efficiency.

Material Bacterial Strain Illumination Wavelength (WL) Effects
nitrogen-doped GQDs and amino-functionalized N-GQDs (amino-N-GQDs) [ As Table 6 shows, there is a potential for studying these dots under different illumination conditions.

Conclusions
In summary, we successfully introduced N atoms in GQDs structure in the form of amide groups in a simple, easy, and eco-friendly process using gamma irradiation. At a dose of 200 kGy and 1, 5, and 10 g weight of EDA in the samples, the percentages of introduced nitrogen varied from 6.38-7.16%. FTIR, XPS, and NMR spectroscopies confirmed the presence of N in GQDs structure in the form of amide and amino moieties. After irradiation in the presence of EDA, PL QY was enhanced from 1.45 to 7.24, 6.77, and 5.82% in the GQDs-EDA/1 g, GQDs-EDA/5 g, and GQDs-EDA/10 g, respectively. GQDs-EDA/1 g were investigated as a potential platform for PL detection of carbofuran, and 3-amino-1,2,4triazole. Emission spectra of GQDs-EDA/1 g recorded in the presence of CF show a linear turn-on response (R 2 = 0.98) with LOD 5.4 µmol L −1 . Pesticide 3-amino-1,2,4-triazole was detected through a turn-off/turn-on mechanism using Pd(II) ions as PL quencher. With concentration increase (0-12 µmol L −1 ), PL was resolved linearly (R 2 = 0.96). LOD was 2.03 µmol L −1 . This is the first time that PL detection of 3-amino-1,2,4-triazole using GQDs as a potential probe has been reported. Using FTIR spectroscopy showed the formation of hydrogen bonds between GQDs-EDA/1 g and carbofuran. Produced dots were unable to induce an antibacterial response on E. coli and S. aureus. Although illumination with blue light was applied for 16 h, bacterial cells survived, indicating than modified dots are not phototoxic towards selected bacterial lines under these conditions.