Phytochemical and Cytotoxic Aspects of Amaryllidaceae Alkaloids in Galanthus Species: A Review
Round 1
Reviewer 1 Report
Comments and Suggestions for Authors1. The authors describe the chemistry of Amaryllidaceae alkaloids, with a special focus on the Galanthus genus and its associated cytotoxic activities. This is an interesting review; however, why did the author focus on cytotoxicity but not Alzheimer's disease, which is more relevant to Galanthus alkaloids?
2. What are the traditional uses of the Galanthus genus? Please briefly explain in the introduction.
3. What is the objective of this review article? It is unclear in the manuscript.
4. Line 192: Galanthamine type alkaloids; the hyphen is missing.
5. Line 338: Please give a guideline or IC50 ranges to show how the authors define inactive, moderate, or good activity of extracts and alkaloids. Did the authors classify these based on the US National Cancer Institute guidelines?
6. Although the authors emphasise the cytotoxicity of Galanthus alkaloids, the literature data is limited with mainly IC50 values and lacks information on structure-activity relationships and molecular mechanisms. I suggest the title has to be revised. Perhaps as an introductory review.
7. The authors should suggest future studies on the possible mechanism of action of selected potent Galanthus alkaloids (give some examples of pathways that could be explored) in the conclusion section.
Author Response
Thank you to all reviewers for the good evaluation of our work, the insights, questions and keypoints, their solutions will undoubtedly improve the quality of our review paper!
Comment 1: This is an interesting review; however, why did the author focus on cytotoxicity but not Alzheimer's disease, which is more relevant to Galanthus alkaloids?
Response 1: The role of Amaryllidaceae alkaloids (and galanthamine in particular) in the field of neurodegenerative diseases is an exhaustively researched topic where major discoveries have been made. The same could be said about cytotoxicity having in mind all the research on lycorine and narciclasine type alkaloids. However, cytotoxicity is built on several levels like in vitro assays, in vivo assays, clinical trials, etc. which should include both cancer and non-tumor cells. Now, taking into account the diversity in cell lines, alkaloids (incl. structural types) and research methods, the field of cytotoxicity is starting to feel understudied. This is why we decided to focus on cytotoxicity in the biological activities section of our review article.
Comment 2: What are the traditional uses of the Galanthus genus? Please briefly explain in the introduction.
Response 2: Information on this has been added with references and can be found at lines 58-66.
Comment 3: What is the objective of this review article? It is unclear in the manuscript.
Response 3: The main goal of our review article is to offer an update on the alkaloid patterns in Galanthus species. This is very much needed since a similar article was most recently published in 2012. Biological activity is a main feature of Amaryllidaceae alkaloids, so we definitely wanted to include that but we decided to briefly mention their renowned anticholinesterase properties and write more about cytotoxicity which is an interesting and lesser known topic in comparison. Furthermore, many structures occurring in Galanthus spp. have not been a part of cancer research and we would like to attract the attention of specialists to them. To showcase this, the last sentence of the abstract was changed (lines 21-23).
Comment 4: Line 192: Galanthamine type alkaloids; the hyphen is missing.
Response 4: A hyphen was added. It is now located at line 168. The same was added in similar cases, lines 234, 262, 264-265, 276, 284, 313, 369-370, 378, 391, 398, 414, 424, 434, 460, 472, 476, 479, 497, 501, 503.
Comment 5: Line 338: Please give a guideline or IC50 ranges to show how the authors define inactive, moderate, or good activity of extracts and alkaloids. Did the authors classify these based on the US National Cancer Institute guidelines?
Response 5: We did not use NCI guidelines. However, often times we defined the degrees of activity as they are seen in the articles that reported them. An approximate scale for compounds would look like this: 0 < IC50 < 10 µM (potent/active), 11 < IC50 < 30 µM (mild/moderate), 31 < IC50 < 50 µM (weak), IC50 > 50 µM (inactive). For extracts the same scale could be used with µg/ml. This scale is added in the newer version of the manuscript at lines 347-351.
Comment 6: Although the authors emphasise the cytotoxicity of Galanthus alkaloids, the literature data is limited with mainly IC50 values and lacks information on structure-activity relationships and molecular mechanisms. I suggest the title has to be revised. Perhaps as an introductory review.
Response 6: Cytotoxicity is a secondary topic of our review, the primary one is the phytochemistry of snowdrops. Since the search of novel cytotoxic agents usually starts with in vitro analyses, we provided more info on that. Information on main structure-activity relationships can be seen on lines 424-428, 437-441, 448-452, 455-458, 461-466, 469-471, 488-492, 505-515. Mechanisms of action are briefly provided for the most noteworthy cytotoxic agents amongst Amaryllidaceae alkaloids (lines 384-386, 410-412, 429-432, 445-447, 472-474, 493-496).
Comment 7: The authors should suggest future studies on the possible mechanism of action of selected potent Galanthus alkaloids (give some examples of pathways that could be explored) in the conclusion section.
Response 7: A few suggestions for the research of cellular pathways have been added in the conclusion, they are at lines 525-529.
Reviewer 2 Report
Comments and Suggestions for AuthorsIn the review article prepared by Georgiev and coworkers, alkaloid compounds and their cytotoxic bioactivities have been well introduced. From the view of synthetic chemistry, although the biosynthetic pathways of Amaryllidaceae alkaloids have been described, it is not enough. Since some of them have been developed as acetylchlolinesterase inhibitor for medical use by the FAD. The asymmetric synthesis of these alkaloid molecules is strongly suggested, highlighting the importance of these studies.
After this, the manuscript could be accepted.
Author Response
Thank you to all reviewers for the good evaluation of our work, the insights, questions and keypoints, their solutions will undoubtedly improve the quality of our review paper!
Comment 1: The asymmetric synthesis of these alkaloid molecules is strongly suggested, highlighting the importance of these studies.
Response 1: The topic of synthetic chemistry of Amaryllidaceae alkaloids is quite fascinating, including the fact that there are methods for the synthesis of stereospecific molecules. However, we think it is more suitable for another review article in order to give it the attention it deserves. If we were to add asymmetric synthetic procedures to our manuscript, it would be longer and would deviate from the main topic which is the phytochemistry of snowdrops and their biological activities. Protocols for the synthesis of stereospecific alkaloids are diverse and include distinct chemical reactions, so they cannot be adequately summarized in a short paragraph.
Reviewer 3 Report
Comments and Suggestions for AuthorsThis review research paper highlights the importance of further investigating Galanthus species that have not been studied, as they could be a source of new benefits in the treatment of various diseases. Therefore, it underlines the relevance of alkaloids from a chemical and pharmacological point of view.
It will be very important to highlight in a table the countries where each of the Galanthus species have been documented in the world. As well as the information on their habitat and growth conditions.
It is also suggested to investigate the cultural implications in the world, as this could add an anthropological dimension to the study.
Line 5, has an extra “1” and is not also superscript
Line 141; it is suggested that the article numbers be in parentheses “Turkey (26)”
Line 293; Correct form “Minimum inhibitory concentration (MIC)”
The symbol for “Liters” is “L” and not “l”, it is requested to modify it throughout the manuscript
Author Response
Thank you to all reviewers for the good evaluation of our work, the insights, questions and keypoints, their solutions will undoubtedly improve the quality of our review paper!
Comment 1: It will be very important to highlight in a table the countries where each of the Galanthus species have been documented in the world. As well as the information on their habitat and growth conditions.
Response 1: A table of accepted species with their geographical distribution was added in place of the list of species names in the introduction. It is located at line 52. The information on their habitat and growth conditions has been expanded and can be found at lines 36-43.
Comment 2: It is also suggested to investigate the cultural implications in the world, as this could add an anthropological dimension to the study.
Response 2: Information on this topic has been added and can be found at lines 56-66. The implication of alkaloids in society is also discussed in the beginnings of sections 4.1. and 5. (Alzheimer's disease and cancer, respectively).
Comment 3: Line 5, has an extra “1” and is not also superscript.
Response 3: The unnecessary "1" was deleted.
Comment 4: Line 141; it is suggested that the article numbers be in parentheses “Turkey (26)”.
Response 4: Dashes were removed and parentheses were added (line 127).
Comment 5: Line 293; Correct form “Minimum inhibitory concentration (MIC)”.
Response 5: Thank you for the correction, it has been changed. It is located at line 272.
Comment 6: The symbol for “Liters” is “L” and not “l”, it is requested to modify it throughout the manuscript.
Response 6: Thank you for this observation, l has been changed to L in all instances throughout the text.
