Carbon Nanoparticles Enhance Drought Tolerance Through the Improvement of Morphological and Physiological Traits in Maize Hybrids

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The authors investigated the effects of carbon nanoparticles (CNPs) on drought stress in maize, assessing physiological traits and photosynthetic performance. The data presented are relatively solid, and the manuscript shows a certain level of logical organization. However, several minor issues need to be addressed:

1. The physical characteristics of CNPs should be provided, such as particle size, size distribution, crystal structure, surface elemental composition, and molecular structure, either through description or experimental characterization.
2. The Results section is too lengthy; it is recommended to divide it into subsections.
3. The Discussion section should specifically address differences between drought-sensitive and drought-tolerant hybrid maize plants.
4. The Discussion should refer to the corresponding figures or tables from your results.
5. It is recommended that cited references be placed at the end of sentences.
6. Line 221: “This reduces leaf production and area.” — the statement is unclear and needs clarification.
7. Lines 226–227: The sentence appears to be missing reference to drought stress response; please check throughout the manuscript.
8. The Discussion section is somewhat list-like; it is recommended to revise and improve the flow of the language.
9. Line 207: There is an extra parenthesis.

Author Response

Response to Reviewer 1 Comments

The manuscript titled "Carbon nanoparticles enhance drought tolerance through the improvement of morphological and physiological traits in maize hybrids" registered under ID: plants-4229926 was reviewed by the authors, and Reviewer 1's comments were answered. The corrections/changes made throughout the manuscript text are highlighted in red. Your comments were important for improving the clarity of our scientific research.

 

Comment 1: The physical characteristics of CNPs should be provided, such as particle size, size distribution, crystal structure, surface elemental composition, and molecular structure, either through description or experimental characterization.

Response 1: I appreciate your comment. A detailed description of the physical characteristics of CNPs has been added to the Material and Methods section.

 

Comment 2: The Results section is too lengthy; it is recommended to divide it into subsections.

Response 2: Thank you very much for your observation. The Results section was divided into three subsections. Subsection 1: Morphophysiological responses compared among maize hybrids. Subsection 2: Effects of foliar CNPs application on maize morphological traits, and Subsection 3: Effects of foliar CNPs application on maize physiological traits.

 

Comment 3: The Discussion section should specifically address differences between drought-sensitive and drought-tolerant hybrid maize plants.

Response 3: A comparative description between drought-tolerant and drought-sensitive maize hybrids was added to the manuscript text. To facilitate the understanding of the morphophysiological responses of different hybrids exposed to drought, an analysis of canonical variables was added to the article.

 

Comment 4: The Discussion should refer to the corresponding figures or tables from your results.

Response 4: Thank you very much for your observation. Throughout the discussion text, the figures and/or tables corresponding to the results of the respective discussion were properly referenced.

 

Comment 5: It is recommended that cited references be placed at the end of sentences.

Response 5: We appreciate your observation. Whenever possible, cited references were moved to the end of the sentence.

 

Comment 6: Line 221: “This reduces leaf production and area.” — the statement is unclear and needs clarification.

Response 6: The sentence has been altered for better clarity and understanding of the discussion text. Sentence was changed to "As result of this effect, there is a reduction in leaf production and leaf area (Figure 1)."

 

Comment 7: Lines 226–227: The sentence appears to be missing reference to drought stress response; please check throughout the manuscript.

Response 7: Thank you for your observation. The sentence has been corrected, and a mention of the drought stress response has been added. This error was verified throughout the manuscript.

 

Comment 8: The Discussion section is somewhat list-like; it is recommended to revise and improve the flow of the language.

Response 8: The Discussion section has been revised, and changes have been made to improve the text's language flow.

 

Comment 9: Line 207: There is an extra parenthesis.

Response 9: The extra parenthesis was removed.

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript entitled “Carbon nanoparticles enhance drought tolerance through the improvement of morphological and physiological traits in maize hybrids” has been submitted to Plants-MDPI Journal. The authors addressed an important issue plant science, particularly in the context of climate change and sustainable agriculture. The manuscript is written well and provides a comprehensive experimental data on the role of carbon nanoparticles (CNPs) in mitigating drought stress in maize. However, there are some issues that need to be addressed.

Major Comments

(1) Why did authors select 1.0 mL L⁻¹ CNP concentration. Explain whether this dose is optimal or based on preliminary trials.

(2) In several sections, the discussion is descriptive rather than analytical. Authors should avoid repeating results and instead explain underlying physiological mechanisms and provide deeper insights into, why CNPs improve water use efficiency? How they influence photosynthesis and stomatal regulation?

(3) In conclusion add practical implications (e.g., field application potential), Address limitations of the study and recommend future research directions (e.g., molecular-level validation)

(4) There are some grammatical and typographical errors throughout the manuscript.

(5) Add more recent references (2023–2025) to strengthen relevance.

(6) Some abbreviations are used before definition (e.g., WUEi, A/Ci). Ensure all abbreviations are defined at first mention.

Minor comments:

(7) Line 28-29: “However, foliar CNP application showed strong potential to alleviate drought's adverse effects…”

 “Strong potential” is not clear.

Example: “However, foliar CNP application significantly alleviated the adverse effects of drought…”

(8) Line 34-35 (Keywords): “Keywords: C-Dots; nanofertilizer; nutritional management; sustainable agriculture; Zea mays.”

“nutritional management” and “C-Dots” may not match the terminology used in the text. Instead add Keywords: Example: carbon nanoparticles; nanofertilizer; drought stress; water use efficiency; Zea mays.”

(9) Line 39-40: “Maize (Zea mays L.) ranks among the world's primary cereal crops, owing largely to its extensive applications…”

 “primary cereal crops” is not clear (better: major/global staple crops)

Example: “Maize (Zea mays L.) is one of the world’s major cereal crops due to its extensive use in food, feed, biofuel, and industrial applications.”

(10) Line 43-44: “The Brazilian Cerrado region produces about two-thirds of the national maize production.”

Repetition: “produces” and “production” Revise the sentence as

Example: "The Brazilian Cerrado region accounts for nearly two-thirds of national maize production.”

(11) Line 56-57:  Nanofertilizers represent an innovative and sustainable approach to enhancing plant growths, physiology, and metabolism under drought conditions (Rivera-Solís et al., 2024). Add recent reference here.

Reference: Rivera-Solís, L. L., Medrano-Macías, J., Morelos-Moreno, Á., Sahito, Z. A., & Benavides-Mendoza, A. (2024). Biostimulation of plants with nanocomposites: a new perspective to improve crop production. In Nanocomposites for Environmental, Energy, and Agricultural Applications (pp. 217-276). Woodhead Publishing.

(12) Line 65-66: Carbon nanoparticle (CNP)-based nanofertilizers offer a sustainable option for         alleviating drought stress in agricultural systems, as their molecules are biodegradable and environmentally benign. CNPs also possess unique structural and mechanical properties, with their use in drought management expanding recently.

The logical flow of the paragraph feels disconnected. It needs smoother transition. Add a sentence like this to connect the story. It also discusses research gap on CNPs in maize drought tolerance.)

“Among nanomaterials, carbon nanoparticles (CNPs) have emerged as promising tools in sustainable agriculture due to their high surface reactivity and multifunctional roles in soil remediation and plant stress tolerance (Sol-Magdaleno et al., 2025).” Carbon nanoparticle-based nanofertilizers offer a sustainable approach for alleviating drought stress in agricultural systems, as these materials are biodegradable and environmentally benign. Moreover, CNPs possess unique structural and mechanical properties, and their application in drought management has expanded in recent years.” Add a recent reference.

Reference: Sol-Magdaleno, M., Aguilar-Aguilar, J. I., Beltrán-Naturi, E., Valencia-Ordóñez, L. D., Díaz-González, A., Trejo-Flores, P., ... & González-Moscoso, M. (2025). Carbon nanomaterials as an environmental technology in the remediation of agricultural soils contaminated with heavy metals: a review. Discover Soil, 2(1), 26.

(13) Line 73-74: “…cowpea (Vigna radiata L.) Wilczek] plants.”

Typographical error: extra bracket “]”

Example: “…cowpea (Vigna radiata L. Wilczek) plants.”

(14) Line 95-96: “Plants were grown in pots containing soil.”

Add sufficient detail (soil type, composition, pot size)

Example:“Plants were grown in plastic pots (X L capacity) filled with a soil mixture (sand:soil:organic matter, ratio X:X:X).”

(15) Line 97-99 (Experimental Design): “The experiment followed a factorial design with replications.”

Incomplete description? Add number of factors and replicates

Example: "The experiment followed a 2 × 2 × 3 factorial design (hybrid × CNP treatment × drought level) with four replicates.”

(16) Line 100-102 (CNP Treatment): “Carbon nanoparticles were applied as foliar spray.”

Add details about the concentration, timing, and application method.

Example: “Carbon nanoparticles (CNPs) were applied as a foliar spray at a concentration of 1.0 mL L⁻¹ using a hand sprayer until runoff, at the V3 growth stage.???”

(17) Line 106-108 (Drought Treatment): “Drought stress was induced using PEG solution.”

Specify PEG type and concentration for reproducibility

Example: “Drought stress was induced using polyethylene glycol (PEG-6000) solutions to achieve osmotic potentials of 0, −0.4, and −0.8 MPa.”…………..

(18) Line 115-118 (Physiological Measurements)

“Photosynthesis and related traits were recorded.”

Add instrument details and conditions

Example: “Photosynthetic rate (A), stomatal conductance (gs), and transpiration rate (E) were measured using a portable infrared gas analyzer (IRGA, model, manufacturer) under standardized light and CO₂ conditions.”

(19) Line 119-120: “Water use efficiency was calculated.”

Mention Formula

(20) Line 136-138: “Application of CNP improved plant growth under stress conditions.”

 “improved” how much? Add statistical data to support.

Example: “CNP application significantly improved plant growth under drought stress, increasing biomass by ??-??% compared to untreated stressed plants (p ≤ 0.05).”

(21) Line 151-155: “There was a significant interaction between treatments.”

Not clear, which interaction? (H × D, D × CNP, or H × D × CNP?)

Example: “A significant interaction between hybrid, drought level, and CNP treatment (H × D × CNP) was observed for most measured traits (p ≤ 0.05).”

Author Response

Response to Reviewer 2 Comments

The manuscript titled "Carbon nanoparticles enhance drought tolerance through the improvement of morphological and physiological traits in maize hybrids" registered under ID: plants-4229926 was reviewed by the authors, and Reviewer 1's comments were answered. The corrections/changes made throughout the manuscript text are highlighted in Blue. Your comments were important for improving the clarity of our scientific research.

 

Comment 1: Why did authors select 1.0 mL L⁻¹ CNP concentration. Explain whether this dose is optimal or based on preliminary trials.

Response 1: The concentration of 1 mL/L of Arbolina applied to maize leaves was based on the nanofertilizer manufacturer's recommendation. This is essential to ensure the effectiveness and safety of the product. This information has been added to the manuscript text.

 

Comment 2: In several sections, the discussion is descriptive rather than analytical. Authors should avoid repeating results and instead explain underlying physiological mechanisms and provide deeper insights into, why CNPs improve water use efficiency? How they influence photosynthesis and stomatal regulation?

Response 2: The discussion section has been improved to address your comments. Beneficial effects of CNPs on water use efficiency, photosynthesis, and stomatal regulation have been added to the manuscript text.

 

Comment 3: In conclusion add practical implications (e.g., field application potential), Address limitations of the study and recommend future research directions (e.g., molecular-level validation)

Response 3: Thank you for your suggestion. Information on practical applications and recommendations for future research has been added to the Conclusions.

 

Comment 4: There are some grammatical and typographical errors throughout the manuscript.

Response 4: Thank you for your observation. A careful grammatical review has been performed throughout the manuscript.

 

Comment 5: Add more recent references (2023–2025) to strengthen relevance.

Response 5: Thank you for your suggestion. Three recent citations have been added to the manuscript.

1. Thabet, S.G., Alqudah, A.M. Unraveling the role of nanoparticles in improving plant resilience under environmental stress conditions. Plant Soil, 503, 313–330 (2024). https://doi.org/10.1007/s11104-024-06581-2
2. Inoue, S.; Tedla, B.; Sobze, J.-M.; Thomas, R. Multi-Walled Carbon Nanotube Application Alters Stomatal Behavior in Boreal Shrubs Under Drought Conditions. Nano 2025, 6, 20. https://doi.org/10.3390/applnano6040020
3. Alabdallah, N.M.; Alluqmani, S.M. Carbon nanotechnology-based sustainable solutions for mitigating drought stress in coffee plants. Coffee Science, 20, e202387, 2025. DOI: 10.25186/.v20i.2387.

 

Comment 6: Some abbreviations are used before definition (e.g., WUEi, A/Ci). Ensure all abbreviations are defined at first mention.

Response 6: Thank you for your observation. All abbreviations in the manuscript have been revised.

 

Comment 7: Line 28-29: “However, foliar CNP application showed strong potential to alleviate drought's adverse effects…” “Strong potential” is not clear.

Example: “However, foliar CNP application significantly alleviated the adverse effects of drought…”

Response 7: Thank you very much for your suggestion. The sentence has been changed to improve understanding.

 

Comment 8: Line 34-35 (Keywords): “Keywords: C-Dots; nanofertilizer; nutritional management; sustainable agriculture; Zea mays.”

“nutritional management” and “C-Dots” may not match the terminology used in the text. Instead add Keywords: Example: carbon nanoparticles; nanofertilizer; drought stress; water use efficiency; Zea mays.”

Response 8: I appreciate your suggestion. The keywords were changed to enhance search indexing terms.

 

Comment 9: Line 39-40: “Maize (Zea mays L.) ranks among the world's primary cereal crops, owing largely to its extensive applications…”

“primary cereal crops” is not clear (better: major/global staple crops)

Example: “Maize (Zea mays L.) is one of the world’s major cereal crops due to its extensive use in food, feed, biofuel, and industrial applications.”

Response 9: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 10: Line 43-44: “The Brazilian Cerrado region produces about two-thirds of the national maize production.”

Repetition: “products” and “production” Revise the sentence as

Example: "The Brazilian Cerrado region accounts for nearly two-thirds of national maize production.”

Response 10: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 11: Line 56-57:  Nanofertilizers represent an innovative and sustainable approach to enhancing plant growths, physiology, and metabolism under drought conditions (Rivera-Solís et al., 2024). Add recent reference here.

Reference: Rivera-Solís, L. L., Medrano-Macías, J., Morelos-Moreno, Á., Sahito, Z. A., & Benavides-Mendoza, A. (2024). Biostimulation of plants with nanocomposites: a new perspective to improve crop production. In Nanocomposites for Environmental, Energy, and Agricultural Applications (pp. 217-276). Woodhead Publishing.

Response 11: The reference has been added to the text.

 

Comment 12: Line 65-66: Carbon nanoparticle (CNP)-based nanofertilizers offer a sustainable option for alleviating drought stress in agricultural systems, as their molecules are biodegradable and environmentally benign. CNPs also possess unique structural and mechanical properties, with their use in drought management expanding recently.

The logical flow of the paragraph feels disconnected. It needs smoother transition. Add a sentence like this to connect the story. It also discusses research gap on CNPs in maize drought tolerance.)

“Among nanomaterials, carbon nanoparticles (CNPs) have emerged as promising tools in sustainable agriculture due to their high surface reactivity and multifunctional roles in soil remediation and plant stress tolerance (Sol-Magdaleno et al., 2025).” Carbon nanoparticle-based nanofertilizers offer a sustainable approach for alleviating drought stress in agricultural systems, as these materials are biodegradable and environmentally benign. Moreover, CNPs possess unique structural and mechanical properties, and their application in drought management has expanded in recent years.” Add a recent reference.

Reference: Sol-Magdaleno, M., Aguilar-Aguilar, J. I., Beltrán-Naturi, E., Valencia-Ordóñez, L. D., Díaz-González, A., Trejo-Flores, P., ... & González-Moscoso, M. (2025). Carbon nanomaterials as an environmental technology in the remediation of agricultural soils contaminated with heavy metals: a review. Discover Soil, 2(1), 26.

Response 12: I appreciate your suggestion. The sentence has been modified to improve understanding of the text. The reference has been added to the text.

 

Comment 13: Line 73-74: “…cowpea (Vigna radiata L.) Wilczek] plants.”

Typographical error: extra bracket “]”

Example: “…cowpea (Vigna radiata L. Wilczek) plants.”

Response 13: The extra bracket was removed.

 

Comment 14: Line 95-96: “Plants were grown in pots containing soil.”

Add sufficient detail (soil type, composition, pot size)

Example:“Plants were grown in plastic pots (X L capacity) filled with a soil mixture (sand:soil:organic matter, ratio X:X:X).”

Response 14: The details of the plastic container size and the type/composition of substrate used for growing the corn plants are described in subsection 2.1 (Plant Growth Condition) of the materials and methods section. This can be viewed on lines 360-362.

 

Comment 15: Line 97-99 (Experimental Design): “The experiment followed a factorial design with replications.”

Incomplete description? Add number of factors and replicates

Example: "The experiment followed a 2 × 2 × 3 factorial design (hybrid × CNP treatment × drought level) with four replicates.”

Response 15: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 16: Line 100-102 (CNP Treatment): “Carbon nanoparticles were applied as foliar spray.”

Add details about the concentration, timing, and application method.

Example: “Carbon nanoparticles (CNPs) were applied as a foliar spray at a concentration of 1.0 mL L⁻¹ using a hand sprayer until runoff, at the V3 growth stage.???”

Response 16: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 17: Line 106-108 (Drought Treatment): “Drought stress was induced using PEG solution.”

Specify PEG type and concentration for reproducibility

Example: “Drought stress was induced using polyethylene glycol (PEG-6000) solutions to achieve osmotic potentials of 0, −0.4, and −0.8 MPa.”…………..

Response 17: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 18: Line 115-118 (Physiological Measurements)

“Photosynthesis and related traits were recorded.”

Add instrument details and conditions

Example: “Photosynthetic rate (A), stomatal conductance (gs), and transpiration rate (E) were measured using a portable infrared gas analyzer (IRGA, model, manufacturer) under standardized light and CO₂ conditions.”

Response 18: The sentence has been modified to improve understanding of the text.

 

Comment 19: Line 119-120: “Water use efficiency was calculated.”

Mention Formula

Response 19: I appreciate your suggestion. The equation for calculating water use efficiency (WUE) has been added. WUE was calculated by the ratio between carbon assimilation (photosynthesis) and water loss through transpiration (WUE = A/E).

 

Comment 20: Line 136-138: “Application of CNP improved plant growth under stress conditions.”

 “improved” how much? Add statistical data to support.

Example: “CNP application significantly improved plant growth under drought stress, increasing biomass by ??-??% compared to untreated stressed plants (p ≤ 0.05).”

Response 20: I appreciate your suggestion. The sentence has been modified to improve understanding of the text.

 

Comment 21: Line 151-155: “There was a significant interaction between treatments.”

Not clear, which interaction? (H × D, D × CNP, or H × D × CNP?)

Example: “A significant interaction between hybrid, drought level, and CNP treatment (H × D × CNP) was observed for most measured traits (p ≤ 0.05).”

Response 21: Thank you for your suggestion. The sentence has been modified to improve understanding of statistical effects.

Reviewer 3 Report

Comments and Suggestions for Authors

Please read the attached file.

Comments for author File: Comments.pdf

Author Response

Response to Reviewer 3 Comments

The manuscript titled "Carbon nanoparticles enhance drought tolerance through the improvement of morphological and physiological traits in maize hybrids" registered under ID: plants-4229926 was reviewed by the authors, and Reviewer 3's comments were answered. The corrections/changes made throughout the manuscript text are highlighted in Green. Your comments were important for improving the clarity of our scientific research.

 

Comment 1: *In the Abstract section, please clarify. Did you spray with nanoparticles just once? At what stage? Or at some intervals throughout the 25-day experiment.

Response 1: Thank you for your observation. Foliar application of CNPs was performed only at the V₂ growth stage. This information has been added to the Abstract.

 

Comment 2: What do you consider early growth to mean? Line 31.

Response 2: Thank you for your observation. The technical term has been changed to plant initial growth.

 

Comment 3: Lines 46-55. There are repetitions in meaning. I suggest we shorten it.

Response 3: Thank you for your observation. The sentence has been modified/reduced to improve text comprehension.

 

Comment 4: Lines 104-106. Please clarify information.

Response 4: The sentence has been modified/reduced to improve text comprehension.

 

Comment 5: The authors of lines 207-243 describe the harmful effects of drought in great detail. These things are generally obvious. I propose to shorten this part.

Response 5: Thank you for your observation. The sentence has been modified to improve text comprehension.

 

Comment 6: Why is the Discussion section not discussing the impact on the two drought-resistant varieties of corn? Why would you do that if you're not discussing it? After all, there are also many discussions and separate conclusions about how varieties respond to nanoparticle treatment.

Response 6: A comparative description between drought-tolerant and drought-sensitive maize hybrids was added to the manuscript text. To facilitate the understanding of the morphophysiological responses of different hybrids exposed to drought, an analysis of canonical variables was added to the article.

 

Comment 7: Part of the manuscript Discussion requires correction.

Response 7: Thank you for your observation. The Discussion section has been improved to enhance comprehension and address the suggestions of the three ad hoc reviewers.

Round 2

Reviewer 3 Report

Comments and Suggestions for Authors

The authors took all my comments and remarks into account.