Sydnone Imines as a New Class of Promising Plant Growth and Stress Tolerance Modulators—A First Experimental Structure–Activity Overview

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript entitled: "Sydnone imines as a new class of promising plant growth and stress tolerance modulators – a first experimental structure-activity-analysis overview" is a review study.

Climate change and various environmental stress factors (drought, high salinity, heavy metals, low or high temperature, etc.) can strongly affect yield and the quality of plant products. Therefore, in order to satisfy the growing food demand of the human population, the stress tolerance of currently cultivated plants must be improved. Learning about plant defense mechanisms at the molecular level provides a tool for improving the ability to adapt.

Endogenously produced nitric oxide (NO) is one of the key signaling factors that activate stress-resistant mechanisms in plants. Thus, the use of synthetic NO donors as stress-protective phytoeffectors can support the maintenance of plant growth and productivity under stressful conditions. Sydnone-imines (sydnonimines) are heterocyclic NO donors that represent a promising group of candidates in the group of such phytoeffectors.
In the manuscript, the authors provide a sufficient overview of the current situation and progress of the use of sydnone. Imines, as exogenous NO donors in plants, are also analyzed for their herbicidal effect. Their use is also reviewed according to aspects such as e.g. herbicide antidotes, growth promoters and as stress protectors.
Sydnonimines are considered as promising phytoeffector candidates, which are expected to exert a protective effect on plants growing under unfavorable conditions, depending on the exact chemical structure.

The review analyzes a sufficient number of and sufficiently up-to-date sources. Based on their results, he draws correct conclusions regarding the future perspectives of this topic.

After correcting a few minor typing errors, I recommend publishing the manuscript in the form of a scientific article.

Comments on the Quality of English Language

After correcting a few minor typing errors, I recommend publishing the manuscript in the form of a scientific article.

Author Response

We thank the Reviewer for thoughtful review and highly appreciate the valuable comments and suggestions to improve the manuscript. Following these advices, we have corrected statements and supplemented the manuscript with relevant information in corresponding sections.

Remark 1. After correcting a few minor typing errors, I recommend publishing the manuscript in the form of a scientific article.

Answer: In line with your recommendations, we have conducted a comprehensive review of the manuscript, paying close attention to the specific areas:

Chemical Entities and Notations: The notations for “CO₂”, “O₂”, “NAD⁺”, “¹O₂”, “H₂O₂”, “Ca²⁺”, “Ni²⁺”, “Pb²⁺”, “Zn²⁺”, and “Cu²⁺” have been carefully revised. This was to ensure accuracy and consistency with scientific standards. These adjustments have been made across the manuscript, specifically on lines 57, 64, 65, 66, 69, 71 on Page 2; line 104 on Page 3; and lines 666-859 on Pages 18-22 and were marked with yellow color.

Species Names: All species names have been correctly italicized, adhering to the standard scientific format. This includes “Medicago sativa L.”, “Zea mays L.”, and “Triticum aestivum L.” among others. The modifications are found on Line 260 on Page 6; Line 375 on Page10; Lines 563, 564, 578, 593, 596, 598, 600 on Page 18  are marked with yellow color.

Figures and Tables: We have ensured that all figures and tables are cited correctly within the main text and placed near their first citation, as per the journal's guidelines. These revisions are evident on Pages 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, and 17.

Citation Format: The bibliographical sources within the text have been revised to be enclosed in square brackets "[]", with no space between them in parentheses. This comprehensive review and correction process encompassed the entire article, ensuring consistency and alignment with the recommended format.

References Format: The entire References chapter has been meticulously reviewed and updated and all the changes were marked with yellow color. Each citation now adheres to the format “Author 1, A.B.; Author 2, C.D. Title of the article. Abbreviated Journal Name Year, Volume, page range.” Additionally, the Digital Object Identifier (DOI) has been included for each reference where available.

These changes enhance the scientific precision of our manuscript and align it with established conventions in chemical and scientific writing. Ensuring the accuracy of these notations, the proper citation of sources, and the correct presentation of figures and tables is crucial for the clarity and reliability of the research presented.

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

As a Review article, it addressed an interesting topic from a scientific and practical point of view. Numerous scientific articles, in concordance to the topic of the study, were consulted.

Methodology of the study was clearly presented, and appropriate to the proposed objectives.

The article has an interesting approach, and the results of the scientific review are presented correctly, in relation to the purpose of the study.

The discussions are appropriate, in the context of the results, and was conducted compared to other studies in the field.

The scientific literature, to which the reporting was made, is recent and representative in the field.

Some suggestions and corrections were made in the article.

The following aspects are brought to the attention of the authors.

1.

Correct writing of some substances / compounds / terms / ionic forms

“CO₂/O₂” instead of “CO2/O2”

“CO₂”

“O₂”

“NAD⁺”

“¹[O₂]” or “¹O₂”

“H₂O₂”

“Ca²⁺”

“Ni²⁺”

“Pb²⁺”

“Zn²⁺”

“Cu²⁺”

It is recommended to review and correct the entire article.

2.

Italic Font style for species name

e.g.

“Medicago sativa L.”

“Zea mays L.”

“Triticum aestivum L”

It is recommended to check and correct the entire article, as appropriate.

3.

Presentation of the figures in the content of the article

According to Instructions for Authors, and Microsoft Word template, Stresses journal,

“All figures and tables should be cited in the main text as Figure 1, Table 1, etc.”

“Figures should be placed in the main text near to the first time they are cited.”

It is recommended to check and correct the entire article, as appropriate.

4.

Citation of bibliographical sources in the text

Recommended right bracket "[]"

There is no space between the bibliographic sources in parentheses

It is recommended to check and correct the entire article, as appropriate.

5.

References

According to Instructions for Authors, and Microsoft Word template, Stresses journal

“Author 1, A.B.; Author 2, C.D. Title of the article. Abbreviated Journal Name Year, Volume, page range.”

“Include the digital object identifier (DOI) for all references where available.”

It is recommended to check the entire References chapter, and correct where necessary.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

Revision is necessary for the correct writing of some substances / compounds / terms / ionic forms

Author Response

We thank the Reviewer for thoughtful review and highly appreciate the valuable comments and suggestions to improve the manuscript. Following these advices, we have corrected statements and supplemented the manuscript with relevant information in corresponding sections.

Remark 1. Correct writing of some substances / compounds / terms / ionic forms

“CO₂/O₂” instead of “CO2/O2”

“CO₂”

“O₂”

“NAD⁺”

“¹[O₂]” or “¹O₂”

“H₂O₂”

“Ca²⁺”

“Ni²⁺”

“Pb²⁺”

“Zn²⁺”

“Cu²⁺”

It is recommended to review and correct the entire article.

Answer: In line with your recommendations, we have conducted a comprehensive review of the manuscript and have corrected the writing of the specified chemical entities. The notations for “CO₂”, “O₂”, “NAD⁺”, “1O₂”, “H₂O₂”, “Ca²⁺”, “Ni²⁺”, “Pb²⁺”, “Zn²⁺”, and “Cu²⁺” have been carefully revised to ensure accuracy and consistency with scientific standards.

These corrections have been made across the manuscript and can be found at the following locations:

Lines 57, 64, 65, 66, 69, 71 on Page 2

Line 104 on Page 3

Lines 563, 564, 578, 593, 596, 598, 600 on Page 18

These changes enhance the scientific precision of our manuscript and align it with the established conventions in chemical and scientific writing. Ensuring the accuracy of these notations is crucial for the clarity and reliability of the research presented.

Remark 2.

Italic Font style for species name

e.g.

“Medicago sativa L.”

“Zea mays L.”

“Triticum aestivum L”

It is recommended to check and correct the entire article, as appropriate.

Answer: In compliance with your instructions, we have thoroughly reviewed the entire manuscript and ensured that all species names are now correctly italicized, adhering to the standard scientific format. This includes names such as “Medicago sativa L.”, “Zea mays L.”, and “Triticum aestivum L.”, among others.

These modifications have been comprehensively addressed throughout the manuscript and can be found at the following locations.:

Line 260 on Page 6

Line 375 on Page 10

Lines 564, 578 on Page 18

Line 855 on Page 24

Each of these instances has been meticulously revised to ensure that the nomenclature is in full compliance with the scientific and formatting standards.

Remark 3. Presentation of the figures in the content of the article

According to Instructions for Authors, and Microsoft Word template, Stresses journal,

“All figures and tables should be cited in the main text as Figure 1, Table 1, etc.”

“Figures should be placed in the main text near to the first time they are cited.”

It is recommended to check and correct the entire article, as appropriate.

Answer: We have thoroughly reviewed the manuscript and ensured that all figures and tables are cited correctly within the main text. Each is referenced in the format of "Figure 1", "Scheme 1", and ‘Table 1”, in accordance with the journal's guidelines. Moreover, special attention has been paid to the placement of these figures and tables. They have been strategically positioned in the main text close to the point where they are first cited, ensuring a coherent and seamless flow of information for the reader. To facilitate your review, we would like to specifically point out the pages where these figures and tables have been incorporated and revised: Pages 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, and 17.

Remark 4. Citation of bibliographical sources in the text

Recommended right bracket "[]"

There is no space between the bibliographic sources in parentheses

It is recommended to check and correct the entire article, as appropriate.

Answer: In accordance with your instructions, we have diligently revised the citations within the text. Each bibliographical source is now correctly enclosed in square brackets "[]", as recommended. Additionally, we have ensured that there is no space between these bibliographic sources when they appear in parentheses. This comprehensive review and correction process encompassed the entire article. We have checked each citation for consistency and alignment with the recommended format.

Remark 5. References

According to Instructions for Authors, and Microsoft Word template, Stresses journal

“Author 1, A.B.; Author 2, C.D. Title of the article. Abbreviated Journal Name Year, Volume, page range.”

“Include the digital object identifier (DOI) for all references where available.” 

It is recommended to check the entire References chapter, and correct where necessary.

Answer: We are pleased to inform you that all references in the manuscript have been meticulously reviewed and updated to align with the prescribed format. Each citation now adheres to the structured format:

“Author 1, A.B.; Author 2, C.D. Title of the article. Abbreviated Journal Name Year, Volume, page range.”

Furthermore, we have ensured that the Digital Object Identifier (DOI) is included for each reference where available, recognizing its importance for providing a persistent link to the digital content.

The entire References chapter has been thoroughly examined and revised where necessary, ensuring adherence to the recommended standards.

Thank you for highlighting these important aspects of manuscript preparation.

Reviewer 3 Report

Comments and Suggestions for Authors

Review of the paper: The paper addresses intriguing issues related to environmental stresses induced by climate change that may threaten crops, necessitating the enhancement of stress tolerance in agricultural crops. Understanding molecular mechanisms allows for the manipulation of plant metabolism using chemical tools, thereby maintaining crop productivity under unfavorable conditions. The authors present nitric oxide (NO) as a crucial signaling factor activating stress tolerance mechanisms in plants, with synthetic NO donors, such as sydnone imines, proving promising as phytoeffectors protecting against stress. Specifically, derivatives of 4-(α-hydroxybenzyl) sydnone imines exhibit either growth-stimulating or antidote effects, influenced by the nature of the substituent at the N-6 atom.  Sydnone imines, as candidates for phytoeffectors, may offer protective effects or act as herbicides, depending on their structure, contributing to sustainable plant growth in challenging conditions. The paper is worth publishing in its current form, serving as a valuable source of information for interested readers.

Author Response

We thank the Reviewer for thoughtful reading and highly appreciate for his high evaluation and highlighting those important aspects of the manuscript.

Reviewer 4 Report

Comments and Suggestions for Authors

Sydnone imines are used as exogenous NO donors in plants. In this manuscript entitled ‘Sydnone imines as a new class of promising plant growth and stress tolerance modulators – a first experimental structure-activity-analysis overview’, authors have attempted to provide an overview of the regulating effect of Sydnone imines on plant growth (e.g., herbicide, germination, and seed maturation) and stress tolerance (e.g., drought, ROS, and high temperature) and its molecular structure.

There are some comments that required a response from the authors.

[1] Who are the candidate readers for the manuscript? Molecular botanist? This paper is a little out of my professional field.

[2] The manuscript is very similar to the published paper named ‘Sydnone Imines: A Novel Class of Plant Growth Regulators’ (https://doi.org/10.3390/agrochemicals2020015).

[3] To match the scope of the journal ‘Stresses’, the authors should provide an overview of recent studies on the mechanism of Sydnone imines in plant response to abiotic and biotic stresses.

Comments on the Quality of English Language

No comments.

Author Response

We thank the Reviewer for thoughtful review and highly appreciate the valuable comments and suggestions to improve the manuscript. Following these advices, we have corrected statements and supplemented the manuscript with relevant information in corresponding sections.

Remark 1. Who are the candidate readers for the manuscript? Molecular botanist? This paper is a little out of my professional field.

Answer: The potential readers for this manuscript are interdesciplinary specialists in the field of agriculture science, plant physiology and molecular biology of plant stress tolerance mechanisms and scientists engaged in search of natural and development of synthetic agrochemicals with potential applications in agriculture. The manuscript  describes application of phytoeffector approach with use of synthetic NO donors such as sydnone imines to protect plant against various environment stresses. In this context sydnone imine action might be directed on production NO able triggering signaling cascades leading to activation tolerance mechanisms. One point of their application might be the priming of plant tolerance mechanisms (by a pre-sowing treatment of seeds). Priming is considered as a mechanism which triggers and supports increased “alertness” of plant physiological state without exposure it to stress. This makes plants to respond faster on oncoming drought or another stress.

However, effects of sydnone imines depend on their structure, and they can exert plant protective effects or act as herbicides. Therefore, chemical manipulation of structure of the sydnone imines may provide an opportunity to improve ability of the compounds to increase tolerance of crops to various stress conditions.

In accordance with your question, we decided to sharpen focus on the point that one of the main purposes of sydnone application is to increase stress tolerance of crop plants so we added the following phrase (marked with green color) in the manuscript text (page 4, line 154):

“We focused on this topic as see the potential in sydnone imines as a new class of plant growth modulators and stress protectors.”

Remark 2. The manuscript is very similar to the published paper named ‘Sydnone Imines: A Novel Class of Plant Growth Regulators’ (https://doi.org/10.3390/agrochemicals2020015).

Answer: The current manuscript is a Review, but the published paper named ‘Sydnone Imines: A Novel Class of Plant Growth Regulators’ (https://doi.org/10.3390/agrochemicals2020015) is the most recent experimental study on some of the sydnone imine molecules (namely nine sydnone imines derivatives bearing variable substituents at N(3), C(4), and N6 positions of the heterocycle) that were applied for the priming of plant tolerance mechanisms (by a pre-sowing treatment of winter wheat (Triticum aestivum L., two cultivars) and maize (Zea mays L., two hybrids) seeds) in germinating experiments. This paper was published in Agrochemicals Journal while we were preparing this current manuscript. We state that the text of our manuscript is unique and has no critical similarity to that experimental work. We thank the Reviewer for referring us to the experimental work, we have added the up-to-date information from the paper to our Review on pages 18-19 lines 612-623 and highlighted in green, as follows:

“The most recent studies [93] from 2023 year include the compounds 8, 15, 16, 17, 18, 22, 32, 33 that were utilized for a pre-sowing treatment of winter wheat (Triticum aestivum L., two cultivars) and maize (Zea mays L., two hybrids) seeds in germinating experiments in a wide concentration range (10^-9–10^-4 mol/L). All compounds were found to affect the growth of the axial organs of germinanted seeds, with the growth-stimulating or -inhibitory effect as well as its rate being considerably different for wheat and maize and, in many cases, also for roots and shoots. Compounds 16, 18, 22 and 33 exhibited a growth-promoting effect in germinating experiments and were used at appropriate concentrations for pot experiments. For all compounds, the experiments showed a stimulating effect on the growth of roots (up to 80%), shoots (up to 112%), leaf area (up to 113%), fresh weights of roots (up to 83%), and aerial parts of the plants (up to 87%) or only on some of these parameters.”

Remark 3. To match the scope of the journal ‘Stresses’, the authors should provide an overview of recent studies on the mechanism of Sydnone imines in plant response to abiotic and biotic stresses.

Answer: Answering to this comment we would like to return to the comment [1]:

The manuscript describes application of phytoeffector approach with use of synthetic NO donors such as sydnone imines to protect plant against various environment stresses. In this context sydnone imine action might be directed on production NO able triggering signaling cascades leading to activation stress tolerance mechanisms. This mechanism of sydnone imines in plant response to abiotic and biotic stresses have already been described in lines 143-155 (Pages 3-4)

“Sydnone imines represent a class of mesoionic heterocyclic compounds [31] exhibiting a broad range of biological activities and in this regard they are successfully applicable in medicine. Recently sydnone imines were addressed as the plant phytoeffectors and reported to be potent growth modulators [32]. These compounds are prodrugs which after enzymatic activation and rearrangement can act as potent NO donors, and as NO is involved in the modulation of stress response, this class of compounds can be considered as a source of prospective stress tolerance-enhancers as well. Therefore, here we review NO signaling role in biological systems, and summarized progress in (i) application of sydnone imines in pharma as NO donors, and (ii) in use of different synthetically available sydnone imine derivatives as herbicides or plant growth stimulators. Additionally, we specifically address the structure-activity relationships in the context of the growth modulating activity of the sydnone imines.”

 Also there were done some updates for the English language, everything is highlighted with comments.

Round 2

Reviewer 4 Report

Comments and Suggestions for Authors

Thank the authors for active response and modification. I have no comments on the previous three questions. There are still two problems as follow.

[1] There are a lot of abbreviations in this manuscript, such as ABA (abscisic acid), Ach (acetylcholine), APX (ascorbic peroxidase), ARC (amidoxime reducing component), CAT (catalase), CK (cytokinin), EDRF (endothelial derived relaxing factor), ET (ethylene), ETC (electron-transport chains), GA (gibberellin acid), GMOs (genetic modified organisms), GR (glutathione reductase), IAA (indoleacetic acid), JA (jasmonic acid), MDA (Malondialdehyde), NAD (nicotinamide adenine dinucleotide), NADPH (nicotinamide adenine dinucleotide phosphate), NO (nitric oxide), NOFNiR (nitric oxide-forming nitrite reductase), NR (nitrate reductase), POD (peroxidase), PTM (post translational modification), RNS (reactive nitrogen species), ROS (reactive oxygen species), SA (salicylic acid), SAR (structure-activity relationships), SOD (superoxide dismutase), TBARS (thiobarbituric acid-reactive substances), TBC (thiobarbituric acid), and so on. Thus, it's probably better to add a list for all abbreviations in the manuscript.

[2] The first letter of the genus may be used as an abbreviation in the second and subsequent uses of the name, but the rest of the name is not abbreviated, e.g., Z. mays L. (Line564, Line578, Line606), T. aestivum (Line605).

Comments on the Quality of English Language

No comments.

Author Response

Thank the authors for active response and modification. I have no comments on the previous three questions. There are still two problems as follow.

We thank the Reviewer for the valuable suggestions and did the corresponded corrections and highlighted it with Blue color.

[1] There are a lot of abbreviations in this manuscript, such as ABA (abscisic acid), Ach (acetylcholine), APX (ascorbic peroxidase), ARC (amidoxime reducing component), CAT (catalase), CK (cytokinin), EDRF (endothelial derived relaxing factor), ET (ethylene), ETC (electron-transport chains), GA (gibberellin acid), GMOs (genetic modified organisms), GR (glutathione reductase), IAA (indoleacetic acid), JA (jasmonic acid), MDA (Malondialdehyde), NAD (nicotinamide adenine dinucleotide), NADPH (nicotinamide adenine dinucleotide phosphate), NO (nitric oxide), NOFNiR (nitric oxide-forming nitrite reductase), NR (nitrate reductase), POD (peroxidase), PTM (post translational modification), RNS (reactive nitrogen species), ROS (reactive oxygen species), SA (salicylic acid), SAR (structure-activity relationships), SOD (superoxide dismutase), TBARS (thiobarbituric acid-reactive substances), TBC (thiobarbituric acid), and so on. Thus, it's probably better to add a list for all abbreviations in the manuscript.

Answer: We added the section of abbreviations on the lines 657-676 and highlighted with blue color.

[2] The first letter of the genus may be used as an abbreviation in the second and subsequent uses of the name, but the rest of the name is not abbreviated, e.g., Z. mays L. (Line564, Line578, Line606), T. aestivum (Line605).

Answer: We accepted the suggestion and put the first letters of the genus as abbreviations in the second and subsequent uses in Lines 564, 578, 606 for Z. mays L.; line 605 for T. aestivum and  highlighted with blue color.

Also we checked up and made Scheme 2, Figures 5, 7, 9 and 10 more visually appealing, nothing was changed in the meaning, just visually we made them better and highlighted with grey color.