Thiamethoxam Sensing Using Gelatin Carbon Dots: Influence of Synthesis and Purification Methods

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript presents an interesting study on the synthesis of gelatin-derived carbon dots (CDs) using two different methods (muffle furnace and microwave irradiation) combined with varying purification strategies.

• The SEM images (Figure 4a) and TEM images (Figure 6a,c) lack clearly visible scale bars, or the scale bars are too small and difficult to read. Please revise all microscopy figures to include properly visible scale bars and ensure adequate resolution.
• While the study is well contextualized, some of the key recent works on CDs for pesticide sensing are missing. Consider including additional references on fluorescence quenching-based detection of pesticides using biomass-derived CDs (e.g., from agricultural wastes).
• The manuscript would benefit from a short discussion on the reproducibility of the synthesis and sensing performance.
• Did you assess the long-term stability of CDs (e.g., fluorescence intensity after one month) and whether purification method affects stability?
• Graphene and graphene-based derivatives are well known to suppress fluorescence and simultaneously enhance Raman signals (Graphene-Enhanced Raman Scattering, GERS). Since your study focuses on fluorescence quenching mechanisms, it might be valuable to include a short discussion on whether similar effects could arise when carbon-based nanomaterials such as your gelatin-derived carbon dots interact with analytes. This perspective is relevant because several works have demonstrated how GERS principles can complement fluorescence-based sensing approaches (see for example “The structure of chemical vapor deposited graphene substrates for graphene-enhanced Raman spectroscopy “ Phys.: Condens. Matter36 195303 10.1088/1361-648X/ad238a)

Author Response

The manuscript presents an interesting study on the synthesis of gelatin-derived carbon dots (CDs) using two different methods (muffle furnace and microwave irradiation) combined with varying purification strategies.

 

The Reviewer 2 answers are pink highlight in the text

1-The SEM images (Figure 4a) and TEM images (Figure 6a,c) lack clearly visible scale bars, or the scale bars are too small and difficult to read. Please revise all microscopy figures to include properly visible scale bars and ensure adequate resolution.

Answer: We would like to thank the reviewer for the comments. The scale bars in the Figures have been corrected (Line 361).

 

2-While the study is well contextualized, some of the key recent works on CDs for pesticide sensing are missing. Consider including additional references on fluorescence quenching-based detection of pesticides using biomass-derived CDs (e.g., from agricultural wastes).

Answer: We would like to thank the reviewer for the comments. In the Line 67-72, the references using biomass derived CDs for pesticides detection were inserted.

 

3-The manuscript would benefit from a short discussion on the reproducibility of the synthesis and sensing performance.

Answer: We would like to thank the reviewer for the comments. In lines 293-297 we introduce the text: “The synthesis of CDs-MF was done at least 4 times in to assess the repeatability. The reproducibility was available using the spectroscopic characterization, UV-vis. spectra and fluorescence emission profile. Where band width and height were compared to each sample batch. The CDs-MW obtained from domestic microwave, previously produced by our group (10.1007/s10895-023-03490-y.)”

 

4-Did you assess the long-term stability of CDs (e.g., fluorescence intensity after one month) and whether purification method affects stability?

Answer: We would like to thank the reviewer for the comments. At the time of this study, no stability studies were performed regarding temperature, UV irradiation, or time. The samples were stored in solid state (raw material), and an aqueous suspension was prepared for each characterization technique or analytical application.

 

5-Graphene and graphene-based derivatives are well known to suppress fluorescence and simultaneously enhance Raman signals (Graphene-Enhanced Raman Scattering, GERS). Since your study focuses on fluorescence quenching mechanisms, it might be valuable to include a short discussion on whether similar effects could arise when carbon-based nanomaterials such as your gelatin-derived carbon dots interact with analytes. This perspective is relevant because several works have demonstrated how GERS principles can complement fluorescence-based sensing approaches (see for example “The structure of chemical vapor deposited graphene substrates for graphene-enhanced Raman spectroscopy “ Phys.: Condens. Matter36 195303 10.1088/1361-648X/ad238a

Answer: We would like to thank the reviewer for the comments. Over time since the CDs discovery in 2004 (Xu, X.; Ray, R.; Gu, Y.; Ploehn, H.J.; Gearheart, L.; Raker, K.; Scrivens, W.A. Electrophoretic Analysis and Purification of Fluorescent Single-Walled Carbon Nanotube Fragments. J Am Chem Soc 2004, 126, 12736–12737. DOI :10.1021/ja040082h.) a variety of fluorescent carbon nanoparticle has been described in the literature, that include graphene quantum dots (GQDs), carbon quantum dots (CQDs), carbon nanodots (CNDs), polymer carbon dots (PCDs) and carbonized polymer dots (CPDs), which are classified according to the specific carbon core structure and surface groups (functional groups or polymer molecular state) (Xia, C.; Zhu, S.; Feng, T.; Yang, M.; Yang, B. Evolution and Synthesis of Carbon Dots: From Carbon Dots to Carbonized Polymer Dots. Adv. Sci. 2019, 6, 1901316. https://doi.org/10.1002/advs.201901316). The structure of CDs consists of sp²/sp³ carbon skeleton and oxygen/nitrogen-based groups (Zhu, S., Song, Y., Zhao, X. et al. The photoluminescence mechanism in carbon dots (graphene quantum dots, carbon nanodots, and polymer dots): current state and future perspective. Nano Res, 2025, 8, 355–381. https://doi.org/10.1007/s12274-014-0644-3). The use of top-down synthesis, normally produced CDs that have a graphite-type carbon skeleton and are identified as graphene quantum dots (GQDs) or carbon quantum dots (CQDs) (Zeng, Q.; Feng, T.; Tao, S.; Zhu, S.; Yang, B. Precursor-Dependent Structural Diversity in Luminescent Carbonized Polymer Dots (CPDs): The Nomenclature. Light Sci Appl 2021, 10, doi:10.1038/s41377-021-00579-6.). GQDs are small graphene fragments consisting of single or few graphene sheets with obvious graphene lattices and chemical groups on the edge or within the interlayer defect, which contribute the unique properties, such as quantum confinement effect and edge effect (Xia, C.; Zhu, S.; Feng, T.; Yang, M.; Yang, B. Evolution and Synthesis of Carbon Dots: From Carbon Dots to Carbonized Polymer Dots. Adv. Sci. 2019, 6, 1901316. https://doi.org/10.1002/advs.201901316

A variety of techniques have been employed for the detection of pesticides. The outstanding advantage of surface-enhanced Raman spectroscopy (SERS) among these spectroscopic detection techniques is that it is an ultra-sensitive analytical method, in which the Raman signal is boosted by a factor of 10¹⁰–10¹¹ at the single molecule level (Yang, Y.; Kong, L.; Ding, Y.; Xia, L.; Cao, S.; Song, P. High SERS Performance of Functionalized Carbon Dots in the Detection of Dye Contaminants. J Adv Res 2025, 68, 89–98, doi:10.1016/J.JARE.2024.02.004.) Graphene itself provides significant enhancement of the Raman signal and the so-called graphene-enhanced Raman scattering (GERS) (Ilse C. Novoa-De León, Jacob Johny, Sofia Vázquez-Rodríguez, David Avellaneda-Avellaneda, Sadasivan Shaji, and Selene Sepúlveda-Guzmán. Nanocarbon Hybrid Films of Reduced Graphene Oxide and N-Doped Graphene Quantum Dots as a Metal-Free Platform for Graphene-Enhanced Raman Scattering. ACS Applied Materials & Interfaces 2025 17 (11), 17251-17259. DOI: 10.1021/acsami.4c21874) Choopun, et al, the graphene quantum dots (GQDs) are prepared by the electrochemical reaction process (top-down synthesis) of electrolytes in solution and are characterized and investigated SERS signals are observed and analyzed (Panyathip, R.; Sucharitakul, S.; Phaduangdhitidhada, S.; Ngamjarurojana, A.; Kumnorkaew, P.; Choopun, S. Surface Enhanced Raman Scattering in Graphene Quantum Dots Grown via Electrochemical Process. Molecules 2021, 26, 5484. https://doi.org/10.3390/molecules26185484)

Our CDs from gelatine are obtained from hydrothermal process (bottom-up synthesis). The majority of the CDs prepared through the bottom-up synthesis method from asymmetrical precursors belong to the category of carbonized polymer dots (CPDs) (Zeng, Q.; Feng, T.; Tao, S.; Zhu, S.; Yang, B. Precursor-Dependent Structural Diversity in Luminescent Carbonized Polymer Dots (CPDs): The Nomenclature. Light Sci Appl 2021, 10, doi:10.1038/s41377-021-00579-6.). The CPDs present special “core-shell”. The core has highly cross-linked rigid polymer network with slight carbonization. While outer shell contains a large number of small polymer chains with large functional groups (Zheng, C.; Tao, S.; Yang, B. The Current Progress and Challenges of Carbonized Polymer Dot-Based Room-Temperature Phosphorescent Materials. CCS Chem. 2023, 6, 1–50. https://doi.org/10.31635/ccschem.023.202303234.).

Reviewer 2 Report

Comments and Suggestions for Authors

The article entitled “Thiamethoxam sensing using gelatin carbon dots: influence of 2 synthesis and purification methods” is in line with the Chemosensors journal. This article is based on original research. The topic is connected with the synthesis of carbon dots. It is up-to-date, interesting and important. The abstract is properly formulated. The article has relevant composition, but it needs slight supplementation before publication, as follows:

 

• Introduction: The novelty aspect should be described more clearly in the last paragraph.
• Chapter 2.1. The additional characteristic of feedstocks is required; please add the basic data for applied materials.
• Chapter 2.1.1 (Line 113): What is the significance of a Teflon coating?
• Chapter 2.1.1 (Line 122): What value of power was used?
• Chapter 2.1.1 (Line 125): What are the conditions in the laboratory (temperature and moisture).
• The discussion could strengthen this article, including comparison with up-to-date literature (properties of CD obtained from other precursors) and add information about the significance of the provided research for further applications.
• Reformulate the conclusion and add there measurable values.

Author Response

The article entitled “Thiamethoxam sensing using gelatin carbon dots: influence of 2 synthesis and purification methods” is in line with the Chemosensors journal. This article is based on original research. The topic is connected with the synthesis of carbon dots. It is up-to-date, interesting and important. The abstract is properly formulated. The article has relevant composition, but it needs slight supplementation before publication, as follows:

We would like to thank the reviewer for the comments and suggestions in our manuscript. We appreciate the opportunity to revise our work, and all the appointments were amended and suggestions applied in the text. We also would like to thank the reviewer for considering our manuscript for publication in Chemosensors.

The Reviewer 1 answers are blue highlight in the text

1-Introduction: The novelty aspect should be described more clearly in the last paragraph.

Answer: We would like to thank the reviewer for the comments In Lines 99-112 the author’s rewrite the paragraph in according the Reviewer suggestion.

 

2-Chapter 2.1 (Line 117). The additional characteristic of feedstocks is required; please add the basic data for applied materials.

Answer: The characteristic of feedstocks was described in detail in the Topic 2.1.

 

3-Chapter 2.1.1 (Line 137-139): What is the significance of a Teflon coating?

Answer: The stainless-steel autoclave can be used with Teflon coating or without it. Teflon-lined autoclaves are often used for their non-stick properties and chemical inertness, particularly when working with corrosive or sensitive materials. In our experiments the suspension solution (ethanol/water) was directly added the stainless-steel container without teflon coating.

 

4-Chapter 2.1.1 (Line 147): What value of power was used?

Answer: In the line we add the brand, power and frequence of domestic microwave (Midea, 700 W, 2450 MHz).

5-Chapter 2.1.1 (Line 151): What are the conditions in the laboratory (temperature and moisture).

Answer: The temperature and humidity conditions in the laboratory were ~30 °C and ~40%, respectively.

 

6-The discussion could strengthen this article, including comparison with up-to-date literature (properties of CD obtained from other precursors) and add information about the significance of the provided research for further applications.

 

Answer: We would like to thank the reviewer for the comments. In line 66-79 and Line 99-112 we introducing a discussion about reviewer suggestions.

 

7-Reformulate the conclusion and add there measurable values.

Answer: We would like to thank the reviewer for the comments. In the Line 663-674, the Conclusion was rewrite.

 

8-The English could be improved to more clearly express the research.

Answer: We would like to thank the reviewer for the comments. the English review was carried out throughout the text.

 

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript provides a comprehensive and systematically organized investigation into the synthesis of gelatin-derived carbon dots (CDs) employing two methodologies (muffle furnace and microwave irradiation), followed by several purifying techniques. The work methodically examines the influence of these parameters on the structural, morphological, and optical characteristics of CDs, as well as their efficacy in thiamethoxam (TMX) detection using fluorescence quenching. The findings largely support the conclusions, and the study provides valuable insights about CD-based sensors for finding pesticides.

Major Revision:

The abstract and introduction could do a better job of highlighting the originality.

First study that compares muffle and microwave-synthesized gelatin CDs for TMX detection? Please explain this.

To do Stern-Volmer analysis or fluorescence anisotropy tests to tell the difference between static quenching and ground-state complex formation, even if IFE is the main thing.

Onion epidermal cells serve as a suitable model; nevertheless, quantified fluorescence data for TMX quenching in the biological matrix are lacking. Adding fluorescent intensity profiles or histograms would strengthen the claim of in situ detection.

The LOD and LOQ numbers are fine, but the publication doesn't provide a table that compares them to other TMX sensors (fluorescence or other types).

The acronyms CDs-MF-C, CDs-MF-F, and CDs-MW are difficult to remember and are presented too soon in the abstract. You might want to shorten the definitions or move them to the methods section and use a table to make them apparent.

There are a lot of grammatical mistakes and unusual phrasing. For example, Line 8: "influence of synthesis and purification methods" might be better as "influence of synthesis and purification methods on their properties and sensing efficiency."

Line 83: "efficiency in pesticide sensing remains relatively underexplored" should be "remains."

Some numbers, like fluorescence spectra, look squished or have labels that overlap.

Make sure that all the graphs are clear and easy to read, especially Figures 7 and 10.
Some of the references aren't formatted correctly (for example, they don't have journal titles or the DOI isn't always in the same location). Follow the Chemosensors style for citations exactly.

Some descriptions are too short and fail to explain the graphic adequately. For example, Figure 6 doesn't explain what the histogram means.

Author Response

The manuscript provides a comprehensive and systematically organized investigation into the synthesis of gelatin-derived carbon dots (CDs) employing two methodologies (muffle furnace and microwave irradiation), followed by several purifying techniques. The work methodically examines the influence of these parameters on the structural, morphological, and optical characteristics of CDs, as well as their efficacy in thiamethoxam (TMX) detection using fluorescence quenching. The findings largely support the conclusions, and the study provides valuable insights about CD-based sensors for finding pesticides.

The Reviewer 4 answers are gray highlight in the text

1-The abstract and introduction could do a better job of highlighting the originality.

Answer: We would like to thank the reviewer for the comments. The abstract was rewrite and insert in the Lines 8-27.

 

2-First study that compares muffle and microwave-synthesized gelatin CDs for TMX detection? Please explain this.

Answer: We would like to thank the reviewer for the comments. In our research group, we previously obtained carbon dots via domestic microwaves (10.1007/s10895-023-03490-y.; 10.1021/acsomega.4c05011.) Regarding gelatin carbon dots, most of them are prepared in the literature via hydrothermal processes in muffle furnaces (10.1016/j.carbon.2013.04.055). In the lines 265-275, we proposed a discussing about reaction mechanism for gelatin carbon dots based in muffle and microwave.

 

3-To do Stern-Volmer analysis or fluorescence anisotropy tests to tell the difference between static quenching and ground-state complex formation, even if IFE is the main thing.

Answer: We would like to thank the reviewer for the comments. A linear relationship in the I/I₀ vs [Q] graphic is most associated with dynamic quenching, where the quenching is mainly due to collisions between the emitter and quencher molecules. However, the linear relationship on the Stern-Volmer plot can also be associated with static quenching system. In this case, the best technique to verify whether the quenching of fluorescence emission occurs via a dynamic or static process is to determine the lifetime in the excited singlet state. Since static quenching does not show changes in the emission lifetime as a function of the quencher concentration lifetime measurements can be used to distinguish between the two (Lakowicz, J. R. Principles of Fluorescence Spectroscopy; Springer Science & Business Media, 2006).

               The fluorescence lifetime obtained to CD-MF-C, CDMF-F and CD-MW were reported in the Table 1, Line 451, where the average lifetime: 8.30, 7.73 and 3.63 ns, respectively. After TMX addition (Table S1, Supplementary Material) reveal that the fluorescence lifetime remains nearly constant. We conclude that, in static quenching, the presence of TMX leads to forming a non-fluorescent ground-state complex with CD-MF, which reduces fluorescence without altering the excited state dynamics. (10.1016/J.JFCA.2024.107109). The text was modified in the lines 622-626.

4-Onion epidermal cells serve as a suitable model; nevertheless, quantified fluorescence data for TMX quenching in the biological matrix are lacking. Adding fluorescent intensity profiles or histograms would strengthen the claim of in situ detection.

Answer: We would like to thank the reviewer for the comments. The representative histogram from each image were obtained from ImageJ (Fiji) software.

 

5-The LOD and LOQ numbers are fine, but the publication doesn't provide a table that compares them to other TMX sensors (fluorescence or other types).

Answer: We would like to thank the reviewer for the comments. The compare TMX fluorescence detection based in carbon dots was insert as Table S1 in Supplementary Material.

 

6-The acronyms CDs-MF-C, CDs-MF-F, and CDs-MW are difficult to remember and are presented too soon in the abstract. You might want to shorten the definitions or move them to the methods section and use a table to make them apparent.

Answer: We would like to thank the reviewer for the comments. Below topic 2.2 (Line 127). Synthesis of CDs, was insert the Table with each acronyms definition.

 

7-There are a lot of grammatical mistakes and unusual phrasing. For example, Line 8: "influence of synthesis and purification methods" might be better as "influence of synthesis and purification methods on their properties and sensing efficiency."

Answer: We would like to thank the reviewer for the comments. the English review was carried out throughout the text.

 

8-Line 83: "efficiency in pesticide sensing remains relatively underexplored" should be "remains." Some numbers, like fluorescence spectra, look squished or have labels that overlap.

Answer: We would like to thank the reviewer for the comments. the English review was carried out throughout the text. All figures in the manuscript were redone to provide better visualization of the information.

 

9-Make sure that all the graphs are clear and easy to read, especially Figures 7 and 10. Some of the references aren't formatted correctly (for example, they don't have journal titles or the DOI isn't always in the same location). Follow the Chemosensors style for citations exactly.

Answer: We would like to thank the reviewer for the comments.

 

10-Some descriptions are too short and fail to explain the graphic adequately. For example, Figure 6 doesn't explain what the histogram means.

Answer: We would like to thank the reviewer for the comments. In the lines 351-354, the histogram is a graphical representation of the distribution of particle sizes when used the microscopy image. The horizontal axis of the histogram represents particle size (e.g., diameter), and the vertical axis represents the number (or frequency) of particles of that size.

Reviewer 4 Report

Comments and Suggestions for Authors

The paper reported the effect of the influence of synthesis and purification methods on the thiamethoxam sensing using gelatin carbon dots. There are two distinct methodologies and two purification methods. Three methods were ultracentrifugation (CDs-MF-C), syringe filtration (CDs-MF-F), and CDs-MW.  

The experimental method is very detailed and reasonable. However, the data analysis needs to be strengthened.

1. In Table 1. “Fluorescence Lifetime for CDs-MF and CDs-MW in aqueous solution at 298 K.”, how many replicates of each run? If they have replicates, please show the means and standard deviations.
2. In Table 2 “ LOD and LOQ values of the CDs studied, determined at two distinct TMX con- centration ranges”, how many replicates of each run? If they have replicates, please show the means and standard deviations.
3. In Figure 9. “Fluorescence emission versus pH plots for (a) λexc = 250 nm and (b) λexci = 320 nm. Both with excitation/emission window 5/10.” and lines 449-453, “

 As observed, upon excitation at 250 nm, no significant differences in fluorescence intensity were detected between CDs-MF and CDs-MW. Therefore, pH 7 was selected as the working condition for subsequent TMX sensing experiments, considering that represents a neutral environment and avoids potential hydrolysis of the TMX molecule under more acidic or basic conditions [64,65].”

 Please use the quantitative method to find the solution, for example, at pH =7, the differences between the three methods ( CDs-MF-C, CDs-MF-F, and CDs-MW) are maximum.  

4. It is very important to find an adequate calibration. The calculation of the LOD and the LOQ is affected by σ and S. Please use the Piecewise regression technique to find adequate calibration equations and use these equations to calculate the LOD and the LOQ.

Author Response

The paper reported the effect of the influence of synthesis and purification methods on the thiamethoxam sensing using gelatin carbon dots. There are two distinct methodologies and two purification methods. Three methods were ultracentrifugation (CDs-MF-C), syringe filtration (CDs-MF-F), and CDs-MW. The experimental method is very detailed and reasonable. However, the data analysis needs to be strengthened.

We would like to thank the reviewer for the comments and suggestions in our manuscript. We appreciate the opportunity to revise our work, and all the appointments were amended and suggestions applied in the text. We also would like to thank the reviewer for considering our manuscript for publication in Chemosensors.

The Reviewer 2 answers are green highlight in the text

1. In Table 1. “Fluorescence Lifetime for CDs-MF and CDs-MW in aqueous solution at 298 K.”, how many replicates of each run? If they have replicates, please show the means and standard deviations.

Answer: We would like to thank the reviewer for the comments. Fluorescence lifetime measurements were not performed in replicates. Time-correlated photon counting (TSPC) technique was used for lifetime analysis. The Time-resolved spectroscopy present, Target Std. Deviation (± ns) ± 0.01 – 0.1 ns and Relative Std. Dev. (± %) ~0.5–2%. The (Chi-squared test) χ² value is the parameter for good measurement quality. χ² values should be between 0.9 and 1.2 (Bevington, P.R.; Robinson, D.K. Data Reduction and Error Analysis for the Physical Sciences, 3rd ed.; McGraw-Hill: New York, NY, USA, 2003)

 

2. In Table 2 “ LOD and LOQ values of the CDs studied, determined at two distinct TMX con- centration ranges”, how many replicates of each run? If they have replicates, please show the means and standard deviations.

Answer: We would like to thank the reviewer for the comments. The Table 2 (Line 450) were completed with means and standard deviations. The measurements were done in duplicated.

 

3. In Figure 9. “Fluorescence emission versus pH plots for (a) λexc = 250 nm and (b) λexci = 320 nm. Both with excitation/emission window 5/10.” and lines 449-453, “As observed, upon excitation at 250 nm, no significant differences in fluorescence intensity were detected between CDs-MF and CDs-MW. Therefore, pH 7 was selected as the working condition for subsequent TMX sensing experiments, considering that represents a neutral environment and avoids potential hydrolysis of the TMX molecule under more acidic or basic conditions [64,65].” Please use the quantitative method to find the solution, for example, at pH =7, the differences between the three methods (CDs-MF-C, CDs-MF-F, and CDs-MW) are maximum.

Answer: We would like to thank the reviewer for the comments. Many synthetic methods have been developed to prepare carbon dots. Interestingly, although the synthesis methods, raw materials, structure, and morphology of carbon dots are quite different, most CDs possess pH-sensitive fluorescence emission properties, despite their pH response behaviors (Liu, C.; Zhang, F.; Hu, J.; Gao, W.; Zhang, M. A Mini Review on PH-Sensitive Photoluminescence in Carbon Nanodots. Front Chem 2021, 8, 626663. DOI: 10.3389/fchem.2020.626663.). The changes in pH might change either the molecular structure or the electronic structure of the pH-responsive CDs. Electronic structure changes can lead to a change in wavelength or intensity of light emission (Zhang, W.; Li, W.; Wang, Q.; Zhang, J.; Li, L.; Zhou, Z.; Lyu, B.; Ma, J. Synthesis of PH-Responsive Fluorescent Carbon Dots Using Waste Leather Scrap for Anti-Counterfeiting. Dyes and Pigments 2024, 231, 11243. DOI: 10.1016/J.DYEPIG.2024.112431.). In general, the literature discussed the pH sensitivity of CDs originating from the protonation and deprotonation of oxygen-containing groups on the surface, which could change the energy level of CDs. There are also many researchers who believed that the pH sensitivity of CDs was caused by the pH induced aggregation of carbon dots. In addition to changing the energy level, protonation, and deprotonation of functional groups could also cause electron transfer between functional group and emission site of CDs as pH changes, thereby resulting in the fluorescence emission change. Yang et al. obtained gelatin carbon dots from stainless steel autoclave with a Teflon liner under heating at 200 °C for 3 h. In this study many H⁺ or OH^- ions induce the significant change of functional groups, and then the electronic transition of some defects would be disrupted or even prohibited (Liang, Q.; Ma, W.; Shi, Y.; Li, Z.; Yang, X. Easy Synthesis of Highly Fluorescent Carbon Quantum Dots from Gelatin and Their Luminescent Properties and Applications. Carbon N Y 2013, 60, 421–428. DOI:10.1016/j.carbon.2013.04.055.). Recently, Shanker et al., synthesized N-CQDs from Citrus sinensis discussed about the intensity of fluorescence emission decreased with a variation in acidity and basicity values. Both protonation and deprotonation characteristics of N-CQDs are indirectly affected by the existence of H⁺ or OH^- ions interacting with the functional groups of these particles and causing such changes (Rishabh, I.; Rani, M.; Shanker, U. Fluorescence-Based Detection of Thiamethoxam in Agricultural and Fruit Samples by Green Synthesized N-CQDs from Citrus Sinensis. Journal of Food Composition and Analysis 2025, 139, 107109. DOI:10.1016/J.JFCA.2024.107109).

               A new discussion was insert in Line 456-503.

 

4. It is very important to find an adequate calibration. The calculation of the LOD and the LOQ is affected by σ and S. Please use the Piecewise regression technique to find adequate calibration equations and use these equations to calculate the LOD and the LOQ.

Answer: We would like to thank the reviewer for the comments. The calibration equation based in Piecewise regression were plotted and insert in the Figure S6 (Supplementary Material). The discussion inserts in the Line 538-545.

 

5-Figures and tables must be improved

Answer: We would like to thank the reviewer for the comments. All figures in the manuscript were redone to provide better visualization of the information.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

I recommend the publication of this article

Reviewer 3 Report

Comments and Suggestions for Authors

Author responded to all comments satisfactorily

Reviewer 4 Report

Comments and Suggestions for Authors

All problems have been addressed adequately.