How to Turn a Poisonous Plant into Medicine: Non-Polar Extracts of Rhododendron adamsii (Sagan Dalya) Are Free of Grayanotoxins and Inhibit the SARS-CoV-2 Main Protease

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Overall, this manuscript presents an interesting and valuable exploratory study combining preliminary phytochemical profiling with preliminary antiviral enzyme inhibition assays of Rhododendron adamsii extracts. The sequential extraction strategy, extensive GC-MS characterization, and focus on grayanotoxin exclusion are notable strengths. However, the biological and therapeutic conclusions currently exceed the available experimental evidence.
As mentioned by the authors, the key limitations include lack of cell-based antiviral validation, incomplete compound identification, absence of assay-interference controls, and speculative mechanistic interpretation. Inspite of those limitations, the manuscript could presents an useful priliminary data that add knowledge to natural-product antiviral research. 

Following corrections should be made to the manuscript before the manuscript can be accepted for publication:

1. Title is too long and confused, hence should be rewritten and made catchy and understandable. Pls include common name of the plant.
2. The writting style of the plant name should standardised and corrected throughout the manuscript.
3. Table titles/legends should be refined.
4. Figures of SDS-PAGE may be included. Protease inhibition could have supported by protein docking.
5. Please check the attached manuscript for further comments.   

Comments for author File: Comments.pdf

Author Response

We thank the Reviewer for the positive assessment of the exploratory value of our study and for the constructive critical remarks. We fully accept the identified limitations and have carefully addressed all comments. The manuscript has been revised accordingly as detailed below.

Following corrections should be made to the manuscript before the manuscript can be accepted for publication:

1. Title is too long and confused, hence should be rewritten and made catchy and understandable. Pls include common name of the plant.

We thank the reviewer for this valuable suggestion. We agree that the original title was overly long and potentially confusing. We have therefore rewritten the title to make it more concise, clear, and informative. The common name of the plant, Sagan Dalya, has been added. We also revised the first part of the title to accurately reflect the content of the study — specifically, that we turned a poisonous plant into a source of potential medicine, not the poison (grayanotoxins) themselves. The new title is:

1. The writting style of the plant name should standardised and corrected throughout the manuscript.

We thank the reviewer for this comment. We have standardized the plant name throughout the manuscript as follows: Rhododendron adamsii Rehd. at first mention, R. adamsii thereafter. The common name Sagan Dalya has been added in the title and abstract. All corrections are highlighted in the revised text.

1. Table titles/legends should be refined.

We thank the reviewer for this comment.We have refined all table titles and legends as suggested. Titles were shortened, redundant wording removed, and units were standardized to "mg/100 g dry plant weight". All changes are highlighted in the revised manuscript (see Tables 1–7).

1. Figures of SDS-PAGE may be included. Protease inhibition could have supported by protein docking.

We thank the reviewer for these constructive suggestions.

Regarding SDS-PAGE: We agree that documentation of protein purity is important. Accordingly, we have added Supplementary Figure S1.

Regarding molecular docking: The reviewer raises an interesting point. While we acknowledge that docking studies would provide valuable mechanistic insight, we believe they fall outside the scope of the present work. Our study focuses on (i) the compositional analysis of nonpolar extracts, (ii) demonstration of the absence of grayanotoxins, and (iii) experimental evidence of selective 3CLpro inhibition (including a negative control against HIV-1 protease, ruling out non‑specific effects). The extracts are complex mixtures containing dozens of lipophilic compounds. Reliable docking would require prior isolation of individual active components, which is the subject of our ongoing bioactivity‑guided fractionation study. We have therefore not included docking here.

We hope the reviewer finds this response acceptable.

1. Please check the attached manuscript for further comments.

We have carefully reviewed the annotated version of the manuscript provided by the reviewer. All comments and suggestions marked in the PDF have been addressed. The corresponding corrections have been incorporated into the revised manuscript and are highlighted for clarity. We thank the reviewer for the detailed feedback.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The study constitutes a contribution to an extensive research investigating the pharmaceutical potential of Rhododendron adamsii extracts. The concept is meticulously formulated and the planning is thorough. The authors demonstrated a solid understanding of the subject matter and an awareness of the study's limitations. I have not found any serious factual errors. My comments mainly concern unclear phrasing, minor errors in names, and factual inaccuracies. I have listed them below.

1. I am not entirely convinced by the first sentence of the title, “How to turn poison into medicine”. It certainly encourages readers to check out the article, but that phrasing led me to believe that the authors had discovered a medical application for grayanotoxins, which, of course, is not true. I would consider the following sentence: “How to turn a poison plant into medicine”.
2. It would be helpful to provide more context for this study. Rhododendrons have long been studied for their antiviral activity. Furthermore, there is a 2023 study (10.1080/07391102.2021.2021287) that examined the activity of another rhododendron species against the same virus, namely SARS-CoV-2. It would be worthwhile to reference this study in the introduction and/or discussion.
3. Authors should correct the notation of chemical compounds that begin with prefixes. In accordance with the rules, prefixes such as trans- and cis- should be written in italics: https://www.acdlabs.com/iupac/nomenclature/93/r93_74.htm.
4. All prefixes should be attached using a hyphen, with no spaces before or after the hyphen (see the numerous minor errors of this kind in Table 5). The notation of chemical compounds should be consistent throughout the paper.
5. Two entries from the Table 5, i.e. “2-Butanone, 3-phenyl-“ and “Elemenone, beta” should be changed to: “3-Phenyl-2-butanone” and “ß-Elemenone”.
6. In the phrase “(…) of the plant Rhododendron adamsii L.in terms of the weight of the raw material (mg/100g, mg%)” (Title of Tables 2–7) there are three errors.
   a – The author “L.” is incorrect; the accepted taxonomic authority for Rhododendron adamsii is Rehder.
   b – There is no space before “in terms”.
   c – There is a missing space in “mg/100 g”.
7. The lines at the top of the Tables 1–7 are of varying thickness.
8. In line 223, the Latin name, i.e. Rh. adamsii should be italicized.
9. The name E. coli in lines 351, 367 and 370 are not italicized.
10. The phrase “30–650 amu” should be replaced with “m/z 30–650”.
11. The phrase “IC₅₀ values were calculated with standard error” should be modified for its better clarity. I would suggest that the authors consider replacing the following fragment of text:

“For samples where pure compounds or well-defined fractions were tested, IC₅₀ values were calculated with standard error”.

with:

“For samples consisting of pure compounds or well-defined fractions, IC₅₀ uncertainty was expressed as standard errors of the parameter estimates derived from nonlinear regression”.

12. A four-paragraph text is too long for a conclusion. I would skip such a lengthy introduction – which is more typical of a summary – and shorten the text a bit.
13. In the list of abbreviations, some of the explanations are capitalized, while others are not. There is a lack of consistency here.
14. There are editorial errors in the references, e.g., in reference No. 13. Additionally, the Latin names are not in italics. Some names are incorrectly capitalized, e.g., the last reference. The species name (coli) is incorrectly capitalized there.
15. The English name for “Rastitelnye resursy” is “ Vegetation Resources”. I would use the English (international) version of the journal name: https://journals.eco-vector.com/0033-9946/index/index/en_US.

May 18, 2026

 

Author Response

We thank the Reviewer for the positive assessment of the exploratory value of our study and for the constructive critical remarks. We fully accept the identified limitations and have carefully addressed all comments. The manuscript has been revised accordingly as detailed below.

The study constitutes a contribution to an extensive research investigating the pharmaceutical potential of Rhododendron adamsii extracts. The concept is meticulously formulated and the planning is thorough. The authors demonstrated a solid understanding of the subject matter and an awareness of the study's limitations. I have not found any serious factual errors. My comments mainly concern unclear phrasing, minor errors in names, and factual inaccuracies. I have listed them below.

1. I am not entirely convinced by the first sentence of the title, “How to turn poison into medicine”. It certainly encourages readers to check out the article, but that phrasing led me to believe that the authors had discovered a medical application for grayanotoxins, which, of course, is not true. I would consider the following sentence: “How to turn a poison plant into medicine”.

We thank the reviewer for this thoughtful suggestion and completely agree with the concern. The original phrasing could indeed be misinterpreted as implying a medical application of grayanotoxins themselves, which was not our intention. Following the reviewer's advice, we have changed the first part of the title to "How to turn a poisonous plant into medicine". The revised title now accurately reflects the core message of the study: that nonpolar extracts of the poisonous plant Rhododendron adamsii are free of its toxic grayanotoxins and exhibit selective antiviral activity. We believe this new title is both accurate and engaging. The correction has been made throughout the manuscript (title, abstract, and main text where the title is cited).

1. It would be helpful to provide more context for this study. Rhododendrons have long been studied for their antiviral activity. Furthermore, there is a 2023 study (10.1080/07391102.2021.2021287) that examined the activity of another rhododendron species against the same virus, namely SARS-CoV-2. It would be worthwhile to reference this study in the introduction and/or discussion.

We thank the reviewer for providing this important reference. We have now added the suggested study (Lingwan et al., 2023) to the Discussion sections. We compare our findings with those of Lingwan et al., noting that different Rhododendron species and extraction methods (nonpolar R. adamsii extracts vs. aqueous R. arboreum extracts) both target the same viral protease, underscoring the potential of the genus as a source of diverse 3CLpro inhibitors. The reference has been added to the Reference list. All changes are highlighted in the revised manuscript.

1. Authors should correct the notation of chemical compounds that begin with prefixes. In accordance with the rules, prefixes such as trans- and cis- should be written in italics: https://www.acdlabs.com/iupac/nomenclature/93/r93_74.htm.

We thank the reviewer for this important correction. We have revised the manuscript according to IUPAC nomenclature rules (R-0.1.6.3), which require italicization of stereodescriptors such as cis- and trans-. These corrections are highlighted in the revised manuscript. We appreciate the reviewer's attention to detail.

1. All prefixes should be attached using a hyphen, with no spaces before or after the hyphen (see the numerous minor errors of this kind in Table 5). The notation of chemical compounds should be consistent throughout the paper.

We thank the reviewer for this careful observation. We have thoroughly revised the manuscript to ensure consistent and correct notation of chemical compound names, with particular attention to Table 5 as suggested. The following corrections have been made throughout the paper (including all tables and the main text).

1. Two entries from the Table 5, i.e. “2-Butanone, 3-phenyl-“ and “Elemenone, beta” should be changed to: “3-Phenyl-2-butanone” and “ß-Elemenone”.

We thank the reviewer for these precise corrections. The two entries in Table 5 have been corrected as suggested:

2-Butanone, 3-phenyl- → 3-Phenyl-2-butanone

Elemenone, beta → β-Elemenone

These changes are highlighted in the revised Table 5. We have also checked the rest of the table for similar issues and made appropriate corrections where necessary.

1. In the phrase “(…) of the plant Rhododendron adamsii L.in terms of the weight of the raw material (mg/100g, mg%)” (Title of Tables 2–7) there are three errors.
   a – The author “L.” is incorrect; the accepted taxonomic authority for Rhododendron adamsii is Rehder.
   b – There is no space before “in terms”.
   c – There is a missing space in “mg/100 g”.

We thank the reviewer for these corrections. The following changes have been made in the titles of Tables 2–7:

• (a) The incorrect authority L. has been replaced with the correct Rehd. (Rehder).
• (b) A space has been added before in terms.
• (c) A space has been inserted in the unit: mg/100g → mg/100 g.

The corrected table titles now read:"of the plant Rhododendron adamsii Rehd. in terms of the weight of the raw material (mg/100 g, mg%)"

All changes are highlighted in the revised manuscript.

1. The lines at the top of the Tables 1–7 are of varying thickness.

We thank the reviewer for this careful observation. We have standardized the horizontal lines across all tables (Tables 1–7). All top and bottom borders are now set to the same thickness (1 pt), and the header separator lines are also uniform. The corrected tables are presented in the revised manuscript.

1. In line 223, the Latin name, i.e. Rh. adamsii should be italicized.

Corrected. The Latin name in line 223 is now in italics.

1. The name E. coli in lines 351, 367 and 370 are not italicized.

Corrected. E. coli is now in italics in all three locations.

1. The phrase “30–650 amu” should be replaced with “m/z 30–650”.

We agree with the reviewer. The notation has been corrected in the GC‑MS analysis section (4.4). The phrase "30–650 amu" has been replaced with "m/z 30–650" to conform to standard mass spectrometry terminology. The change is highlighted in the revised manuscript.

1. The phrase “IC₅₀ values were calculated with standard error” should be modified for its better clarity. I would suggest that the authors consider replacing the following fragment of text:

“For samples where pure compounds or well-defined fractions were tested, IC₅₀ values were calculated with standard error”.

with:

“For samples consisting of pure compounds or well-defined fractions, IC₅₀ uncertainty was expressed as standard errors of the parameter estimates derived from nonlinear regression”.

We thank the reviewer for this helpful suggestion. The proposed wording is indeed more precise and clearly describes how the uncertainty was estimated. We have replaced the sentence in section 4.6 (Statistical Analysis) exactly as recommended. The revised text now reads:

"For samples consisting of pure compounds or well-defined fractions, IC₅₀ uncertainty was expressed as standard errors of the parameter estimates derived from nonlinear regression."

The change is highlighted in the revised manuscript.

1. A four-paragraph text is too long for a conclusion. I would skip such a lengthy introduction – which is more typical of a summary – and shorten the text a bit.

We thank the reviewer for this constructive suggestion. The Conclusions section has been substantially shortened and now consists of two concise paragraphs. 

1. In the list of abbreviations, some of the explanations are capitalized, while others are not. There is a lack of consistency here.

We thank the reviewer for this observation. The list of abbreviations has been standardized. All explanations now begin with a capital letter (capitalized style), as is common for the journal.

1. There are editorial errors in the references, e.g., in reference No. 13. Additionally, the Latin names are not in italics. Some names are incorrectly capitalized, e.g., the last reference. The species name (coli) is incorrectly capitalized there.

We thank the reviewer for carefully checking the references. All references have been thoroughly revised.

1. The English name for “Rastitelnye resursy” is “ Vegetation Resources”. I would use the English (international) version of the journal name: https://journals.eco-vector.com/0033-9946/index/index/en_US.

We thank the reviewer for this correction. The journal name in reference 2 has been changed from Rastitelnye resursy to the official English name Vegetation Resources, as provided in the link. The corrected reference is highlighted in the revised manuscript.

Author Response File: Author Response.pdf