{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T17:00:47Z","timestamp":1786122047867,"version":"build-2736575974"},"update-to":[{"DOI":"10.12688\/f1000research.164618.1","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2025,11,5]],"date-time":"2025-11-05T00:00:00Z","timestamp":1762300800000}},{"DOI":"10.12688\/f1000research.164618.2","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2025,11,5]],"date-time":"2025-11-05T00:00:00Z","timestamp":1762300800000}}],"reference-count":21,"publisher":"F1000 Research Ltd","license":[{"start":{"date-parts":[[2025,11,5]],"date-time":"2025-11-05T00:00:00Z","timestamp":1762300800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["f1000research.com"],"crossmark-restriction":false},"short-container-title":["F1000Res"],"abstract":"<ns5:p>\n                    Background Medicinal plants play a crucial role in pharmacological research, as many pharmaceutical companies rely on them for raw materials. Collard greens, also known as \u201cSukuma wiki,\u201d are a member of the\n                    <ns5:italic>Brassica oleracea<\/ns5:italic>\n                    var. viridis family and are known for their medicinal properties. The aim of this study was to identify phytochemicals in the ethanol extract of collard green leaves that has traditional use in treating male infertility in Uganda. Method The leaves were dried and crushed into fine powder and the powdered leaves (800 g) A was cold macerated in 5 liters of 99% ethanol (1:5 w\/v) for 72 hours in sterile glass container. The ethanol extract (5g) was subjected to Gas Chromatography-Mass Spectrometry (GC-MS) analysis. Results Phytochemical analysis of the Ethanol extract from\n                    <ns5:italic>Brassica oleracea<\/ns5:italic>\n                    var. viridis (Collard green) leaves revealed the presence of various compounds, including flavonoids, alkaloids, phenolic compounds, fatty acids, and terpenoids. Gas Chromatography-Mass Spectrometry (GC-MS) analysis identified 77 bioactive compounds, such as 2-Methoxy-4-vinylphenol, 2, 7-Dimethyl-1, 7-octadien-3-amine, Octadecanoic acid, 9, 12, 15-Octadecatrienoic acid, Pentadecanoic acid, and several phenolic derivatives. Conclusion In conclusion, the phytochemicals identified in\n                    <ns5:italic>Brassica oleracea<\/ns5:italic>\n                    var. viridis (Collard greens), including Phytol, Omega-3 fatty acids, phenols, flavonoids, and sterols, demonstrate potential benefits for enhancing male fertility and reproductive health. These phytochemicals exert their beneficial action through their antioxidant, anti-inflammatory, and possibly neuroprotective properties. There is need for further research to establish the effectiveness of these phytochemicals in clinical settings and provide concrete guidelines for their use in treating male infertility.\n                  <\/ns5:p>","DOI":"10.12688\/f1000research.164618.3","type":"journal-article","created":{"date-parts":[[2025,11,20]],"date-time":"2025-11-20T18:25:13Z","timestamp":1763663113000},"page":"525","update-policy":"https:\/\/doi.org\/10.12688\/f1000research.crossmark-policy","source":"Crossref","is-referenced-by-count":2,"title":["Gas Chromatography-Mass spectrometry (GC-MS) Revealed Bioactive Constituents of Brassica oleracea var. viridis (Collard Greens) Used to Treat Male Infertility in Uganda."],"prefix":"10.12688","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0009-0004-4167-6717","authenticated-orcid":false,"given":"Emmanuel Orire","family":"Ikuomola","sequence":"first","affiliation":[{"name":"Physiology, Faculty of Biomedical Sciences Bushenyi Western Region 20000 Kampala International University - Western Campus"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8264-9131","authenticated-orcid":false,"given":"Daniel Udofia","family":"Owu","sequence":"additional","affiliation":[{"name":"Physiology, Faculty of Biomedical Sciences Bushenyi Western Region 20000 Kampala International University - Western Campus"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7230-6293","authenticated-orcid":false,"given":"Victor Otu","family":"Oka","sequence":"additional","affiliation":[{"name":"Physiology, Faculty of Biomedical Sciences Bushenyi Western Region 20000 Kampala International University - Western Campus"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sunday Agba","family":"Bisong","sequence":"additional","affiliation":[{"name":"Department of Physiology Calabar Cross River University of Calabar, College of Medical Sciences"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ugwu Okechukwu","family":"Paul-Chima","sequence":"additional","affiliation":[{"name":"Publication and Extension Bushenyi Western Region 20000 Kampala International University - Western Campus"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-2450-9460","authenticated-orcid":false,"given":"Patrick Maduabuchi","family":"Aja","sequence":"additional","affiliation":[{"name":"Biochemistry, Faculty of Biomedical Sciences Bushenyi Western Region 20000 Kampala International University - Western Campus"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"2560","published-online":{"date-parts":[[2025,11,5]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"59","DOI":"10.1186\/s43094-021-00208-4","article-title":"Phytochemical profiling and GC-MS analysis of aqueous methanol fraction of Hibiscus asper leaves.","volume":"7","author":"N Olivia","year":"2021","journal-title":"Futur. J. Pharm. Sci."},{"key":"ref2","doi-asserted-by":"publisher","first-page":"655-673","DOI":"10.1007\/s13596-020-00473-3","article-title":"Ethnobotanical knowledge of medicinal plants used in the treatment of male infertility in southern Benin.","volume":"21","author":"E Agbodjento","year":"2021","journal-title":"Adv. Tradit. Med. (ADTM)."},{"key":"ref3","doi-asserted-by":"publisher","first-page":"14090-14102","DOI":"10.33263\/BRIAC116.1409014102","article-title":"Elucidation of bioactive potential of two commonly grown North Indian Psidium guajava viz., Lalit and Shweta against pathogenic foodborne and MDR bacteria.","volume":"11","author":"A Bano","year":"2021","journal-title":"Biointerface Res. Appl. Chem."},{"key":"ref4","doi-asserted-by":"publisher","first-page":"425-429","DOI":"10.6026\/97320630015425","article-title":"Screening and GC-MS profiling of ethanolic extract of Tylophora pauciflora.","volume":"15","author":"T Starlin","year":"2019","journal-title":"Bioinformation."},{"key":"ref5","doi-asserted-by":"publisher","first-page":"1279","DOI":"10.1186\/s12870-024-06005-4","article-title":"Development of an optimized protocol for protoplast-to-plant regeneration of selected varieties of Brassica oleracea L.","volume":"24","author":"K Stelmach-Wityk","year":"2024","journal-title":"BMC Plant. Biol."},{"key":"ref6","doi-asserted-by":"publisher","first-page":"1043-1052","DOI":"10.2174\/1871530320666200929141140","article-title":"Antihyperglycemic effect of the Moroccan Collard Green (Brassica oleracea var. viridis) in Streptozotocin-induced diabetic rats.","volume":"21","author":"A Amssayef","year":"2021","journal-title":"Endocr. Metab. Immune Disord. Drug Targets."},{"key":"ref7","doi-asserted-by":"publisher","first-page":"437-445","DOI":"10.1016\/j.sajb.2024.09.006","article-title":"italic on fertility in normal mice and rats model.","volume":"174","author":"S Patil","year":"2024","journal-title":"South Afr. J. Bot."},{"key":"ref8","doi-asserted-by":"publisher","first-page":"53-58","DOI":"10.22270\/jddt.v10i2.3914","article-title":"GC-MS analysis of bioactive compounds and phytochemical evaluation of the ethanolic extract of Gomphrena globosa L. flowers.","volume":"10","author":"A Esmat","year":"2020","journal-title":"J. Drug Deliv. Ther."},{"key":"ref9","doi-asserted-by":"publisher","first-page":"419-437","DOI":"10.1080\/10496475.2023.2211288","article-title":"Phytochemical analysis and in vitro assessment of extracts of Rhodobryum roseum for antioxidant, antibacterial and anti-inflammatory activities.","volume":"29","author":"H Sabharwal","year":"2023","journal-title":"J. Herbs Spices Med. Plants."},{"key":"ref10","first-page":"787-791","article-title":"Effect of aqueous leaf extract of Irvingia gabonensis on gastrointestinal tract in rodents.","volume":"42","author":"F Abdulrahman","year":"2004","journal-title":"Indian J. Exp. Biol."},{"key":"ref11","doi-asserted-by":"publisher","first-page":"1153-1167","DOI":"10.1007\/s10811-023-03168-9","article-title":"Microalgae amino acids: Method validation using gas chromatography coupled to mass spectrometry.","volume":"36","author":"J Furlan","year":"2024","journal-title":"J. Appl. Phycol."},{"key":"ref12","doi-asserted-by":"publisher","first-page":"46","DOI":"10.1007\/s40203-024-00216-1","article-title":"In silico pharmacokinetic and therapeutic evaluation of Musa acuminata peels against aluminium chloride-induced hepatotoxicity in adult BALB\/c mice.","volume":"12","author":"H Onohuean","year":"2024","journal-title":"In Silico Pharmacol."},{"key":"ref13","first-page":"299-304","article-title":"Effects of partially purified bioactive compounds in leaf extract of Jatropha tanjorensis on gonadal hormones of male albino rats.","volume":"6","author":"J Akighir","year":"2018","journal-title":"Int. J. Life Sci. Res."},{"key":"ref14","doi-asserted-by":"publisher","first-page":"836","DOI":"10.3390\/antiox12040836","article-title":"Antioxidant supplementation on male fertility\u2014a systematic review.","volume":"12","author":"F Dimitriadis","year":"2023","journal-title":"Antioxidants."},{"key":"ref15","doi-asserted-by":"publisher","first-page":"552-560.e3","DOI":"10.1016\/j.fertnstert.2019.11.008","article-title":"The effect of antioxidants on male factor infertility: the Males, Antioxidants, and Infertility (MOXI) randomized clinical trial.","volume":"113","author":"A Steiner","year":"2020","journal-title":"Fertil. Steril."},{"key":"ref16","doi-asserted-by":"publisher","first-page":"514-515","DOI":"10.1038\/aja.2012.46","article-title":"The roles of omega-3 and omega-6 fatty acids in idiopathic male infertility.","volume":"14","author":"M Safarinejad","year":"2012","journal-title":"Asian J. Androl."},{"key":"ref17","doi-asserted-by":"publisher","first-page":"245-256","DOI":"10.1080\/19390211.2018.1431753","article-title":"The effect of omega-3 fatty acids, EPA, and\/or DHA on male infertility: A systematic review and meta-analysis.","volume":"16","author":"B Hosseini","year":"2019","journal-title":"J. Diet. Suppl."},{"key":"ref18","doi-asserted-by":"publisher","first-page":"651-715","DOI":"10.1007\/s00204-020-02689-3","article-title":"Antioxidants and antioxidant methods: An updated overview.","volume":"94","author":"I Gulcin","year":"2020","journal-title":"Arch. Toxicol."},{"key":"ref19","doi-asserted-by":"publisher","first-page":"1514-1521","DOI":"10.47750\/pnr.2022.13.S09.185","article-title":"Chemopreventive Anticancer activity of beta sitosterol in human prostate cancer cells (Pc-3) and Emt-mediated signaling regulation: An in vitro study.","volume":"18","author":"C Reddy","year":"2022","journal-title":"J. Pharm. Negat. Results."},{"key":"ref20","doi-asserted-by":"publisher","first-page":"1043-1052","DOI":"10.2174\/1871530320666200929141140","article-title":"Antihyperglycemic Effect of the Moroccan Collard Green (Brassica oleracea var. viridis) in Streptozotocin-Induced Diabetic Rats.","volume":"21","author":"A Amssayef","year":"2021","journal-title":"Endocrine, Metabolic & Immune Disorders-Drug Targets (Formerly Current Drug Targets-Immune, Endocrine & Metabolic Disorders)."},{"key":"ref21","doi-asserted-by":"publisher","DOI":"10.17605\/OSF.IO\/QND7X","article-title":"Gas Chromatography-Mass spectrometric (GC-MS) Revealed Bioactive Constituents of Brassica Oleracea Var. Viridis (Collard Greens) Used as Ethnomedicine to Treat Male Infertility in Uganda.","author":"E Ikuomola","year":"2025, May 14"}],"container-title":["F1000Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/f1000research.com\/articles\/14-525\/v3\/xml","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/f1000research.com\/articles\/14-525\/v3\/pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/f1000research.com\/articles\/14-525\/v3\/iparadigms","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T08:04:07Z","timestamp":1786089847000},"score":1,"resource":{"primary":{"URL":"https:\/\/f1000research.com\/articles\/14-525\/v3"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,11,5]]},"references-count":21,"URL":"https:\/\/doi.org\/10.12688\/f1000research.164618.3","relation":{"has-version":[{"id-type":"doi","id":"10.12688\/f1000research.164618.2","asserted-by":"subject"},{"id-type":"doi","id":"10.12688\/f1000research.164618.1","asserted-by":"subject"}]},"ISSN":["2046-1402"],"issn-type":[{"value":"2046-1402","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,11,5]]},"assertion":[{"value":"Indexed","URL":"https:\/\/f1000research.com\/articles\/14-525\/v3#article-reports","order":0,"name":"referee-status","label":"Referee status","group":{"name":"current-referee-status","label":"Current Referee Status"}},{"value":"10.5256\/f1000research.181158.r388513, Faten Hameed Thamer, Sana'a University, Sana'a, 25 Jun 2025, version 1, 2 approved with reservations","URL":"https:\/\/f1000research.com\/articles\/14-525\/v1#referee-response-388513","order":0,"name":"referee-response-388513","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"<b>Emmanuel Ikuomola<\/b>; \n<i>Posted: 21 Sep 2025<\/i>; \u201cWe sincerely thank the reviewer for their constructive feedback and valuable recommendations. The authors\u2019 detailed responses to the comments are provided below.\u201d &nbsp; \n<b>General reviewer comments<\/b> The abstract is well-structured but can benefit from clearer language, more detail about methods, and precise emphasis on the significance of findings. Clarify the methodology details, especially regarding extraction and GC-MS parameters.    Author\u2019s\n<b> Response<\/b>: The abstract has been modified to contain method of extraction and conclusion emphasized the significance of the phytochemicals identified in the extract. Author\u2019s\n<b> Response:<\/b> The method of extraction has been clarified by including the temperature, volume of solvent used and the storage of the extract obtained in page 4. Highlight the potential health implications of identified compounds more explicitly. Consider adding quantitative data if available, such as relative abundances of key compounds. Ensure consistency in terminology, e.g., &quot;bioactive compounds&quot; vs. &quot;phytochemicals.&quot; Author\u2019s\n<b> Response:<\/b> The quantitative data is not available however table three shows the retention time and area under curve that represents a little bit of abundance of the compounds. The terminology &quot;bioactive compounds&quot; has been replaced with \u201cphytochemicals\u201d as suggested for consistency.   Improve readability with more formal academic language, especially in the conclusion. 1. Introduction to sample collection: Clarify why the specific location was chosen, if relevant. Author\u2019s\n<b> Response<\/b>: This specific location was chosen since it has more farmland where the plant is cultivated. The GPS location has been included in the text for clarity. Correctly cite the method if it is referenced and complete the citation. Author\u2019s\n<b> Response:<\/b> There is no referenced method of picking the plant. It was just by plucking the leaves from the plant.    2. Botanical identification: Specify the authority or botanist who confirmed the identification for scientific rigor (e.g., &quot;identified and authenticated by Dr. XYZ using standard taxonomic keys&quot;). Author\u2019s\n<b> Response:<\/b> The plant was identified and authenticated by Dr. Eunice Olet of the Department of Botany, Faculty of Biological Sciences, Mbarara University of Science and Technology. A voucher specimen was deposited in the university's herbarium under the reference number IOE-24-001.   3. Sample preparation details: Mention how many leaves were collected or the approximate weight of the fresh leaves before drying. Author\u2019s\n<b> Response:<\/b> The quantity of the fresh leaves has been mentioned to be 2 kg in page 4. Specify the drying conditions (e.g., &quot;dried at ambient temperature (~25&deg;C) in the shade&quot; for accuracy). Author\u2019s\n<b> Response:<\/b> The drying condition has been specified as shed dried at ambient temperature of 26\u2103.    4. Extraction process: Complete the citation for the methodology or clarify if it\u2019s a modified version (e.g., &quot;based on the method described by [Author], [Year]&quot;). Author\u2019s\n<b> Response<\/b>: The sentence has been completed to read \u201cThe extraction process was performed using a modified version of the method described by Abdulrahman et al.\n<sup>10<\/sup>\u201d in page 4. Mention the solvent volume in a consistent manner (&quot;5.5 liters of 99% ethanol,&quot; which is clear). Author\u2019s\n<b> Response<\/b>: The solvent volume has been modified to read \u201cA total of 800 g of the powdered leaves was cold macerated in 5 liter of 99% ethanol (1:5 w\/v).\u201d   5. Maceration process: Indicate the container type used and mention if the mixture was shaken periodically during the maceration. Author\u2019s\n<b> Response:<\/b> The container has been specified as \u201csterile glass container\u201d. The mixture was shaken periodically during the maceration process. This has been stated in page 4. Clarify whether the mixture was kept in the dark to prevent light degradation. Author\u2019s\n<b> Response<\/b>: The mixture was kept in the dark as stated in page 4.  6. Filtration and concentration: Specify the conditions of the rotary evaporator, such as temperature (e.g., &quot;at 40&deg;C&quot;) and pressure. Author\u2019s\n<b> Response:<\/b> The temperature is stated as below 40&deg;C and under reduced pressure. This is stated in the relevant section on page 4. Mention how the dryness was confirmed (e.g., no further solvent smell or weight stabilization). Author\u2019s\n<b> Response:<\/b> The filtrate was evaporated to dryness having obtained a constant weight as stated in page 4.  7. Storage: Clarify storage conditions (e.g., temperature, container&nbsp;type). Author\u2019s\n<b> Response:<\/b> The storage condition of the extract has been mentioned as follows in page 4. \u201cThe dried crude extracts were stored at 4&deg;C in an amber glass bottle that was tightly sealed until further analysis\u201d.","URL":"https:\/\/f1000research.com\/articles\/14-525\/v3#referee-comment-14633","order":1,"name":"referee-comment-14633","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"10.5256\/f1000research.188560.r417218, Adeoye-Isijola Morenike, Babcock University, Ilishan-Remo, Ogun State, 13 Oct 2025, version 2, indexed","URL":"https:\/\/f1000research.com\/articles\/14-525\/v2#referee-response-417218","order":2,"name":"referee-response-417218","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"10.5256\/f1000research.188560.r418701, Amanulla Khan, Anjuman Islam Janjira Degree College of Science, Murud Janjira, Raigad, Maharashtra, 01 Oct 2025, version 2, indexed","URL":"https:\/\/f1000research.com\/articles\/14-525\/v2#referee-response-418701","order":3,"name":"referee-response-418701","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"<b>Emmanuel Ikuomola<\/b>; \n<i>Posted: 23 Oct 2025<\/i>; \n<b>Reviewer 3<\/b> Comments:  Grammar \/ Style:Minor corrections:  \u201cThese phytochemicals such as flavonoids, alkaloids\u2026\u201d \u2192&nbsp;\n<i>\u201cThese phytochemicals, including flavonoids, alkaloids\u2026\u201d<\/i> \n<b>Authors\u2019 Response:<\/b> Thank you for the correction. The necessary changes have been made in the introductory section of the manuscript. Replace&nbsp;\n<i>\u201cas shown in Figure 1\u201d<\/i>&nbsp;with a reference to the updated figure. \n<b>Authors response<\/b>: We thank the reviewer for pointing this out. The phrase has been updated as requested to correctly reference the figure.    &nbsp;  Scientific Clarity:  Strengths: Clear ethnomedicinal context. \n<b>Authors response<\/b>: We sincerely thank the reviewer for this positive feedback. We are pleased that the clear presentation of the ethnomedicinal context, which grounds this laboratory investigation in the traditional use of&nbsp;\n<i>Brassica oleracea<\/i>&nbsp;var.&nbsp;\n<i>viridis<\/i>&nbsp;for treating male infertility in Uganda, was recognized as a strength of our work. Improvements: Cite recent studies (last 5\u20137 years) on&nbsp;\n<i>Brassica oleracea<\/i>&nbsp;bioactive compounds and male fertility. \n<b>Authors response: <\/b>We thank the reviewer for this valuable suggestion. We have now integrated recent studies (published within the last 5-7 years) into the&nbsp;\n<b>manuscript <\/b>sections which support the role of&nbsp;\n<i>Brassica oleracea<\/i>&nbsp;bioactive compounds in promoting male reproductive health. This update strengthens the scientific context of our findings.    &nbsp;    Avoid redundancy; streamline phrases like&nbsp;\n<i>\u201cThese constituents support immune function, bone health, digestion, and oxidative balance\u201d<\/i>&nbsp;to maintain focus on reproductive health.&nbsp; &nbsp; \n<b>AUTHORS RESPONSE:<\/b> We thank the reviewer for this valuable suggestion. We have streamlined the text as recommended, removing the broad list of general health benefits and refocusing the narrative specifically on the compounds' relevance to male reproductive health. The changes have been made in the&nbsp;\n<b>Introduction<\/b>&nbsp;section.&nbsp; &nbsp; &nbsp;    &nbsp;  Plant Collection &amp; Identification: Detailed and well-documented. Minor grammar correction:\n<i>\u201cFresh leaves (2 kg) \u2026 were subsequently transported to the herbarium unit at Mbarara University for proper botanical identification and authentication\u201d<\/i>&nbsp;\u2192&nbsp;\n<i>\u201c\u2026were transported to the herbarium at Mbarara University for botanical authentication.\u201d<\/i> \n<b>Authors response<\/b>: We thank the reviewer for this positive feedback and for the suggested improvement. The sentence has been revised as recommended to be more concise.  &nbsp;  Extraction Procedure:\n<b>&nbsp;<\/b>Clearly mention the ratio (w\/v) for ethanol. extraction. Standardize units:&nbsp;\n<i>\u201c26 2&deg;C\u201d<\/i>&nbsp;\u2192&nbsp;\n<i>\u201c26 &plusmn; 2 &deg;C\u201d<\/i>.Use past tense consistently. \n<b>Authors response: <\/b>We thank the reviewer for these precise observations. We have clarified the ethanol ratio, corrected the temperature unit, and ensured the consistent use of past tense in the described procedure. \n<b>Ratio Clarified:<\/b>&nbsp;Changed &quot;(1:5 w\/v)&quot; to &quot;(at a ratio of 1:5 w\/v)&quot; for absolute clarity. \n<b>Units Standardized:<\/b>&nbsp;Corrected &quot;26 2&deg;C&quot; to &quot;26 &plusmn; 2 &deg;C&quot; and added a space before &quot;&deg;C&quot; for &quot;40 &deg;C&quot; to ensure consistency. \n<b>Past Tense &amp; Grammar:<\/b>&nbsp;Corrected &quot;was concentrated to dryness reduced pressure&quot; to &quot;was concentrated to dryness under reduced pressure&quot; and &quot;having a constant weight&quot; to &quot;until a constant weight was achieved&quot; for grammatical correctness and consistent use of the past tense.  &nbsp;  GC-MS Analysis:&nbsp;Clarify split ratio and sample injection mode. Include column specifications (length, diameter, film thickness) for reproducibility. \n<b>Authors response: <\/b>We thank the reviewer for this critical suggestion to enhance the reproducibility of our methods. We have clarified the split ratio and injection mode and have added the standard column specifications (length, internal diameter, and film thickness) to the GC-MS analysis section.  &nbsp;  Tables 2\u20134: Ensure consistent formatting and capitalization for chemical names.&nbsp; \n<b>Authors response:<\/b> &nbsp;&nbsp;We thank the reviewer for highlighting this lack of consistency. We have carefully revised all three tables {2,3 and 4} to ensure chemical names follow standard IUPAC capitalization rules (sentence case for common names, proper capitalization for systematic names) and that formatting is uniform throughout.  &nbsp;  &nbsp;\n<b>&nbsp;<\/b>Figures:&nbsp;Figure 1: Replace with a high-resolution, original photograph of collard greens collected from the field.&nbsp;Figure 2\n<b>&nbsp;<\/b>(GC-MS chromatogram): Replace with a high-resolution chromatogram; label peaks and retention times clearly.&nbsp;Figure 3\n<b>&nbsp;<\/b>(biosynthetic pathway): Replace with original\/redrawn diagrams relevant to the identified compounds; avoid screenshots from other sources. \n<b>Authors response: <\/b>We thank the reviewer for these critical recommendations to enhance the quality and originality of our figures. We have taken the following actions: \n<b>Figure 1:<\/b>&nbsp;The original diagram has been replaced with a new, high-resolution, original photograph of&nbsp;\n<i>Brassica oleracea<\/i>&nbsp;var.&nbsp;\n<i>viridis<\/i>&nbsp;(collard greens) collected from the field in Bwejuragye, Bushenyi, Uganda, as specified in the methodology. \n<b>Figure 2:<\/b>&nbsp;The GC-MS chromatogram has been replaced with a high-resolution version. Key peaks corresponding to the major bioactive compounds discussed in the manuscript are now clearly labeled with their respective retention times. \n<b>Figure 3:<\/b>&nbsp;The previous pathway diagram has been removed. It has been replaced with an original, self-created schematic that illustrates the proposed mechanistic pathways.  &nbsp;  Strengths:&nbsp;Detailed plant authentication and voucher specimen deposition. Comprehensive GC-MS analysis. Effective linkage of phytochemicals to male fertility mechanisms. \n<b>Authors response:<\/b> We sincerely thank the reviewer for their positive assessment and for recognizing the strengths of our work, particularly the rigorous plant authentication, comprehensive phytochemical profiling, and the functional interpretation of the results in the context of male fertility.  &nbsp;  Limitations \/ Recommendations:&nbsp;Include quantitative analysis (concentration of compounds). Discuss potential toxicity or adverse effects. Provide experimental reproducibility details (replicates, GC-MS calibration). Ensure figures and tables are high-quality and original. \n<b>Authors response: <\/b>We thank the reviewer for these critical and constructive points. We have addressed them by adding a dedicated &quot;Limitations and Future Perspectives&quot; section to the manuscript. The key additions are summarized below.  \n<b>Key additions <\/b>  \n<b>Added a New Section:<\/b>&nbsp;A &quot;Limitations and Future Perspectives&quot; section has been inserted before the conclusion. \n<b>Addressed Quantification:<\/b>&nbsp;Explicitly stated the qualitative\/semi-quantitative nature of the study and recommended quantitative analysis for future work. \n<b>Addressed Toxicity:<\/b>&nbsp;Acknowledged the lack of toxicity data and recommended future toxicological studies. \n<b>Addressed Reproducibility:<\/b>&nbsp;Specified that the analysis was done in triplicate and mentioned instrument calibration. \n<b>Confirmed Figure Quality:<\/b>&nbsp;Reiterated that all figures have been replaced with high-quality, original versions as per the reviewer's previous comment.","URL":"https:\/\/f1000research.com\/articles\/14-525\/v3#referee-comment-14831","order":4,"name":"referee-comment-14831","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"The author(s) declared that no grants were involved in supporting this work","order":5,"name":"grant-information","label":"Grant Information"},{"value":"This is an open access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.","order":0,"name":"copyright-info","label":"Copyright"}]}}