{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T22:01:31Z","timestamp":1791324091168,"version":"4.3.1"},"update-to":[{"DOI":"10.12688\/f1000research.165030.1","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T00:00:00Z","timestamp":1769212800000}},{"DOI":"10.12688\/f1000research.165030.2","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T00:00:00Z","timestamp":1769212800000}}],"reference-count":42,"publisher":"F1000 Research Ltd","license":[{"start":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T00:00:00Z","timestamp":1769212800000},"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":"<ns3:p>\n                    Background\n                    <ns3:italic>Plectranthus amboinicus<\/ns3:italic>\n                    is an aromatic herb known for its medicinal properties and is increasingly explored for cosmetic applications. Its bioactive compounds possess antioxidant, antimicrobial, and anti-inflammatory properties, making it a promising candidate for multifunctional skincare formulations. Methods This study investigated the extraction of bioactive compounds from\n                    <ns3:italic>P. amboinicus<\/ns3:italic>\n                    leaves using microwave-assisted ethanol extraction. Two drying techniques\u2014tray drying and freeze-drying\u2014were compared to evaluate their impact on the extraction efficiency. The optimal extract (PF15), prepared using 15-minute microwave extraction and freeze-drying, was selected for further analysis. Bioactive content was assessed through quantification of caffeic acid, total phenolic content, and antioxidant activity via the DPPH assay. The antimicrobial activity of PF15 was tested against\n                    <ns3:italic>Staphylococcus aureus<\/ns3:italic>\n                    ,\n                    <ns3:italic>Staphylococcus epidermidis<\/ns3:italic>\n                    , and\n                    <ns3:italic>Cutibacterium acnes.<\/ns3:italic>\n                    Anti-inflammatory potential of PF15 was evaluated in LPS-stimulated human THP-1 macrophages by measuring cytokine production. Results The PF15 extract yielded the highest levels of bioactive compounds and demonstrated strong antioxidant activity. It exhibited significant antimicrobial effects against all tested skin pathogens. In the anti-inflammatory assay, PF15 significantly decreased pro-inflammatory cytokines (TNF-\u03b1, IL-1\u03b2, and IL-6) while upregulating the anti-inflammatory cytokine IL-10. The extract was formulated into a topical cream, which underwent accelerated stability testing over six heat-cool cycles. The cream remained stable with no signs of phase separation, discoloration, odor change, or microbial contamination, and maintained a pH of 5.5. Conclusions The PF15 extract of\n                    <ns3:italic>P. amboinicus<\/ns3:italic>\n                    demonstrates potent antioxidant, antimicrobial, and anti-inflammatory properties. Its successful incorporation into a stable cream formulation supports its potential as a multifunctional active ingredient in skincare products. These findings highlight\n                    <ns3:italic>P. amboinicus<\/ns3:italic>\n                    as a valuable natural source for the development of cosmetic formulations targeting oxidative stress, microbial infection, and inflammation.\n                  <\/ns3:p>","DOI":"10.12688\/f1000research.165030.3","type":"journal-article","created":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T11:09:09Z","timestamp":1769252949000},"page":"796","update-policy":"https:\/\/doi.org\/10.12688\/f1000research.crossmark-policy","source":"Crossref","is-referenced-by-count":2,"title":["Bioactive Effects of Plectranthus amboinicus Extract Using Microwave Techniques and Its Value Addition in Cosmeceutical Products"],"prefix":"10.12688","volume":"14","author":[{"given":"Chananan","family":"Chinnahong","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Warut","family":"U-Kong","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thiravat","family":"Rattanapot","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-4651-2514","authenticated-orcid":false,"given":"Chetsalit","family":"Hongnueng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3429-1366","authenticated-orcid":false,"given":"Doungporn","family":"Amornlerdpison","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"2560","published-online":{"date-parts":[[2026,1,24]]},"reference":[{"key":"ref1","article-title":"Herbal beauty and personal care products market size, share, and trends analysis report by product (skin care, hair care, fragrance), by gender, by region, and segment forecasts, 2023\u20132032.","year":"2024 cited 2025 Apr 12"},{"key":"ref3","first-page":"65","article-title":"Antioxidant and anti-skin aging potential of selected Thai plants: in vitro evaluation and in silico target prediction.","volume":"12","author":"W Chaiyana","year":"2023","journal-title":"Plants."},{"key":"ref4","first-page":"45-55","article-title":"Biological activities and essential oil composition of Plectranthus amboinicus.","volume":"49","author":"B Satongrod","year":"2022","journal-title":"Maha. Univ. J. Pharm. Sci."},{"key":"ref5","doi-asserted-by":"publisher","first-page":"1-8","DOI":"10.22270\/ajprd.v11i3.1398","article-title":"Anti-acne emulgel formulation of Torbangun leaf essential oil (Plectranthus amboinicus (Lour.) Spreng) and antibacterial test against Staphylococcus aureus and Propionibacterium acne bacteria.","volume":"12","author":"D Harsep Rosi","year":"2024","journal-title":"Asian J. Pharm. Res. Dev."},{"key":"ref6","doi-asserted-by":"publisher","first-page":"2025336","DOI":"10.31893\/multiscience.2025336","article-title":"Antibacterial activity of Plectranthus amboinicus (Lour.) Spreng extracts against pathogenic bacteria.","volume":"7","author":"W Meesil","year":"2025","journal-title":"Multidiscip. Sci. J."},{"key":"ref7","first-page":"397-402","article-title":"Optimization of Plectranthus amboinicus (Lour.) Spreng extraction process using microwave-assisted technique.","volume":"72","author":"N Samad","year":"2019","journal-title":"Chem. Eng. Trans."},{"key":"ref8","doi-asserted-by":"publisher","first-page":"300-312","DOI":"10.1016\/j.tifs.2005.12.004","article-title":"Recent advances in extraction of nutraceuticals from plants.","volume":"17","author":"L Wang","year":"2006","journal-title":"Trends Food Sci. Technol."},{"key":"ref9","first-page":"1-26","article-title":"Techniques for extraction and isolation of natural products: A comprehensive review.","volume":"16","author":"Q Zhang","year":"2021","journal-title":"Chin. Med."},{"key":"ref10","first-page":"7-18","article-title":"Microwave assisted extraction-an innovative and promising extraction tool for medicinal plant research.","volume":"1","author":"V Mandal","year":"2007","journal-title":"Pharmacogn. Rev."},{"key":"ref11","first-page":"1-7","article-title":"Recent advances in microwave-assisted extraction (MAE) of medicinal plants: A review.","volume":"9","author":"R Aljanabi","year":"2018","journal-title":"Int. Res. J. Pharm."},{"key":"ref12","doi-asserted-by":"publisher","first-page":"369","DOI":"10.3390\/molecules21040369","article-title":"Plectranthus amboinicus (Lour.) Spreng: Botanical, phytochemical, pharmacological and nutritional significance.","volume":"21","author":"G Arumugam","year":"2016","journal-title":"Molecules."},{"key":"ref13","first-page":"373-378","article-title":"Microwave-assisted extraction of Plectranthus amboinicus leaves: Optimization, phytochemical screening and antioxidant activity.","volume":"63","author":"N Abdul Samad","year":"2018","journal-title":"Chem. Eng. Trans."},{"key":"ref14","first-page":"14-22","article-title":"Evaluation of antioxidant activity of aqueous extract of some selected nutraceutical herbs.","volume":"96","author":"J Bor","year":"2006","journal-title":"Food Chem."},{"key":"ref15","first-page":"45-53","article-title":"Evaluation of the antioxidant activity of Plectranthus amboinicus leaf extract using the DPPH method.","volume":"32","author":"J Suntorn","year":"2024","journal-title":"Thai J. Sci. Technol."},{"key":"ref16","doi-asserted-by":"publisher","first-page":"22622-22630","DOI":"10.1039\/C9RA09058D","article-title":"Flaxseed orbitides, linusorbs, inhibit LPS-induced THP-1 macrophage inflammation.","volume":"10","author":"X Zou","year":"2020","journal-title":"RSC Adv."},{"key":"ref17","doi-asserted-by":"publisher","first-page":"75","DOI":"10.3390\/metabo9040075","article-title":"Propolis exerts an anti-inflammatory effect on PMA-differentiated THP-1 cells via inhibition of purine nucleoside phosphorylase.","volume":"9","author":"A Alqarni","year":"2019","journal-title":"Metabolites."},{"key":"ref18","doi-asserted-by":"publisher","first-page":"159-167","DOI":"10.4103\/0973-1296.197646","article-title":"Technological evaluation of emulsions containing the volatile oil from leaves of Plectranthus amboinicus Lour.","volume":"13","author":"P Lopes","year":"2017","journal-title":"Pharmacogn. Mag."},{"key":"ref19","first-page":"34-42","article-title":"Evaluation of Staphylococcus aureus contamination in acne treatment cream products for consumer safety.","volume":"12","author":"K Phaosan","year":"2021","journal-title":"Thai J. Cosmet. Sci."},{"key":"ref21","first-page":"113-121","article-title":"Effect of extraction time on the yield and composition of phytochemicals from medicinal plants.","volume":"273","author":"S Pradhan","year":"2021","journal-title":"J. Ethnopharmacol."},{"key":"ref22","doi-asserted-by":"publisher","DOI":"10.4172\/2157-7110.1000360","article-title":"Effect of freeze-drying conditions on antioxidant compounds of broccoli.","volume":"05","author":"A Mahn","year":"2014","journal-title":"J. Food Technol."},{"key":"ref23","first-page":"105-115","article-title":"Microwave-assisted extraction of bioactive compounds from medicinal plants.","volume":"25","author":"M Bennett","year":"2020","journal-title":"J. Herb. Med."},{"key":"ref24","first-page":"100262","article-title":"Antioxidant and antimicrobial activities of Plectranthus amboinicus (Lour.) leaves extract prepared by microwave-assisted extraction.","volume":"21","author":"A Abubakar","year":"2020","journal-title":"J. Appl. Res. Med. Aromat. Plants."},{"key":"ref25","first-page":"1097-1105","article-title":"Antimicrobial activity of Plectranthus amboinicus leaf extracts against pathogenic microorganisms.","volume":"107","author":"V Shriram","year":"2009","journal-title":"J. Appl. Microbiol."},{"key":"ref26","first-page":"11-17","article-title":"Antimicrobial and antioxidant properties of Plectranthus amboinicus extracts.","volume":"4","author":"D Bhatt","year":"2012","journal-title":"Int. J. Pharm. Sci."},{"key":"ref27","doi-asserted-by":"publisher","first-page":"4295-4300","DOI":"10.4049\/jimmunol.174.7.4295","article-title":"Requirement for TLR9 in the immunomodulatory activity of Propionibacterium acnes.","volume":"174","author":"C Kalis","year":"2005","journal-title":"J. Immunol."},{"key":"ref28","doi-asserted-by":"publisher","first-page":"381-388","DOI":"10.1038\/jid.2013.309","article-title":"Propionibacterium acnes induces IL-1\u03b2 secretion via the NLRP3 inflammasome in human monocytes.","volume":"134","author":"M Qin","year":"2014","journal-title":"J. Invest. Dermatol."},{"key":"ref29","doi-asserted-by":"publisher","first-page":"1535-1541","DOI":"10.4049\/jimmunol.169.3.1535","article-title":"Activation of Toll-like receptor 2 in acne triggers inflammatory cytokine responses.","volume":"169","author":"J Kim","year":"2002","journal-title":"J. Immunol."},{"key":"ref30","doi-asserted-by":"publisher","first-page":"135-144","DOI":"10.1016\/j.immuni.2007.05.022","article-title":"Toll-like receptor 4 is a sensor for autophagy associated with innate immunity.","volume":"27","author":"Y Xu","year":"2007","journal-title":"Immunity."},{"key":"ref31","doi-asserted-by":"publisher","first-page":"296-303","DOI":"10.1111\/j.1365-2133.2009.09420.x","article-title":"Interleukin-10 secretion from CD14+ peripheral blood mononuclear cells is downregulated in patients with acne vulgaris.","volume":"162","author":"F Caillon","year":"2010","journal-title":"Br. J. Dermatol."},{"key":"ref32","doi-asserted-by":"publisher","first-page":"869-875","DOI":"10.1189\/jlb.0909610","article-title":"TNF-\u03b1, IL-6, and IL-1 expression is inhibited by GAS6 in monocytes\/macrophages.","volume":"87","author":"F Alciato","year":"2010","journal-title":"J. Leukoc. Biol."},{"key":"ref33","doi-asserted-by":"publisher","first-page":"963-969","DOI":"10.4049\/jimmunol.163.2.963","article-title":"Lipopolysaccharide modulates cyclooxygenase-2 transcriptionally and posttranscriptionally in human macrophages independently from endogenous IL-1\u03b2 and TNF-\u03b1.","volume":"163","author":"M Barrios-Rodiles","year":"1999","journal-title":"J. Immunol."},{"key":"ref34","doi-asserted-by":"publisher","first-page":"287-295","DOI":"10.1016\/j.jpba.2004.10.030","article-title":"Assays of physical stability and antioxidant activity of a topical formulation added with different plant extracts.","volume":"37","author":"V Di Mambro","year":"2005","journal-title":"J. Pharm. Biomed. Anal."},{"key":"ref35","first-page":"113-121","article-title":"Formulation of topical skincare products with plant extracts: an overview.","volume":"31","author":"R Brown","year":"2018","journal-title":"Skin Pharmacol. Physiol."},{"key":"ref36","first-page":"78-85","article-title":"Stability testing of cosmetic formulations under accelerated conditions.","volume":"42","author":"P Kumar","year":"2020","journal-title":"Int. J. Cosmet. Sci."},{"key":"ref37","first-page":"420-427","article-title":"Effect of temperature variations on the stability of topical products.","volume":"108","author":"X Liao","year":"2019","journal-title":"J. Pharm. Sci."},{"key":"ref38","first-page":"950-957","article-title":"Accelerated stability testing of skincare formulations.","volume":"20","author":"J Wang","year":"2021","journal-title":"J. Cosmet. Dermatol."},{"key":"ref39","first-page":"234-242","article-title":"Anti-inflammatory and antimicrobial properties of medicinal plants for acne treatment.","volume":"41","author":"J Choi","year":"2019","journal-title":"Int. J. Cosmet. Sci."},{"key":"ref40","first-page":"153329","article-title":"In vitro antibacterial activities of various plant extracts against skin pathogens.","volume":"79","author":"S Park","year":"2021","journal-title":"Phytomedicine."},{"key":"ref41","article-title":"Guidelines on Good Manufacturing Practices for Cosmetics.","year":"2020","journal-title":"HO Technical Report Series, No. 984."},{"key":"ref42","first-page":"496-502","article-title":"Evaluation of the safety and efficacy of Plectranthus amboinicus leaf extract in cosmetic formulations.","volume":"18","author":"Y Lim","year":"2019","journal-title":"J. Cosmet. Dermatol."},{"key":"ref43","doi-asserted-by":"publisher","DOI":"10.1016\/j.trac.2024.117627","article-title":"Critical analysis of green extraction techniques used for botanicals: Trends, priorities, and optimization strategies-A review.","volume":"173","author":"C Ciro","year":"2024","journal-title":"TrAC Trends Anal. Chem."},{"key":"ref44","doi-asserted-by":"publisher","DOI":"10.5281\/zenodo.15796465","article-title":"Bioactive effects of Plectranthus amboinicus extract using microwave techniques and its value addition in cosmetical products.","author":"C Chananan","journal-title":"Zenodo."}],"container-title":["F1000Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/f1000research.com\/articles\/14-796\/v3\/xml","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/f1000research.com\/articles\/14-796\/v3\/pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/f1000research.com\/articles\/14-796\/v3\/iparadigms","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T11:09:13Z","timestamp":1769252953000},"score":1,"resource":{"primary":{"URL":"https:\/\/f1000research.com\/articles\/14-796\/v3"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,1,24]]},"references-count":42,"URL":"https:\/\/doi.org\/10.12688\/f1000research.165030.3","relation":{"has-version":[{"id-type":"doi","id":"10.12688\/f1000research.165030.2","asserted-by":"subject"},{"id-type":"doi","id":"10.12688\/f1000research.165030.1","asserted-by":"subject"}],"has-review":[{"id-type":"doi","id":"10.5256\/f1000research.195380.r443293","asserted-by":"object"}]},"ISSN":["2046-1402"],"issn-type":[{"value":"2046-1402","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,1,24]]},"assertion":[{"value":"Indexed","URL":"https:\/\/f1000research.com\/articles\/14-796\/v3#article-reports","order":0,"name":"referee-status","label":"Referee status","group":{"name":"current-referee-status","label":"Current Referee Status"}},{"value":"10.5256\/f1000research.181624.r407511, Jukkarin Srivilai, University of Phayao, Phayao, 15 Sep 2025, version 1, 1 approved, 1 not approved","URL":"https:\/\/f1000research.com\/articles\/14-796\/v1#referee-response-407511","order":0,"name":"referee-response-407511","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"<b>Doungporn Amornlerdpison<\/b>; \n<i>Posted: 03 Oct 2025<\/i>; We would like to sincerely thank the reviewers for the valuable and constructive comments. We have carefully revised the manuscript according to all suggestions, and below we provide a point-by-point response. The changes have been made as highlighted in \u201cred font\u201d in revised manuscript. We hope our revision has improved the paper to a satisfactory level.\n<b> <\/b> \n<b>Reviewer 1 (Approved)<\/b> 1. Title: The word \u201cTechniques\u201d should be singular (\u201cTechnique\u201d). &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; Corrected to \u201cTechnique.\u201d  2. Abstract: In the Methods section, the last sentence should be revised to: \u201cAnti-inflammatory potential of PF15 \u2026.\u201d Revised: \u201cAnti-inflammatory potential of PF15 was evaluated in LPS-stimulated THP-1 macrophages.\u201d  3.Dosage form: The authors should clearly state the dosage form of the cream throughout the manuscript, or at least specify somewhere that it is an emulsion type. Stated in Part: Product Development and Evaluation, Page 5 \u201cThe extract was incorporated into the cream formulation at a concentration of 0.8%, ensuring antimicrobial efficacy. The aqueous phase containing PF15 was heated to 70 &deg;C, combined with the oil phase, and emulsified for 30 minutes. The cream was then cooled to 35\u201340 &deg;C, mixed until homogeneous, and stored for further analysis. A topical cream was developed by incorporating the PF15 extract into a water-in-oil emulsion system. The detailed composition of the cream is presented in Table 2.\u201d  4.Introduction: In the third paragraph, please verify reference [6] and revise the sentence for better grammatical clarity. Verified and corrected sentence, Page 2. \u201cA recent study demonstrated the antimicrobial efficacy of P. amboinicus leaf extract against pathogens such as Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Candida albicans, and the acne-associated bacterium Cutibacterium acnes, thereby reinforcing its potential application in cosmetic formulations. \n<sup>6<\/sup>  5.Materials and Methods: The description of the 50% ethanol extraction should specify the solvent used (e.g., DI water or another solvent). &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; Clarified as ethanol + deionized water, Page 3. \u201cThe 50% ethanol solution was prepared using a 1:1 mixture of ethanol and deionized water.\u201d  - Chemical analysis (HPLC): The methodology should clarify whether standard compounds were used to construct the calibration curve for determining the relative amount of caffeic acid, or if another approach was applied. Please provide details. Clarified as \u201cThe standard of caffeic acid was used to construct the calibration curve.\u201d&nbsp;  - Table 1: The column \u201cMode\u201d should be labeled \u201cMode of elution.\u201d Corrected. Mode of elution  - Abbreviations: Ensure that abbreviations follow the full name consistently (e.g., Minimal Inhibitory Concentration (MIC)). Please check throughout the manuscript. Checked and corrected; Minimal Inhibitory Concentration (MIC)  - Cream formulation with PF15: Provide more details on the formulation ratio, type of emulsion, and preparation technique. Full formulation ratios, emulsion type, and preparation technique added, Page 5-6.  6.Results and Discussion: - Verify the unit of yield, as it may be incorrect. Corrected to % w\/w dry weight - Figure 3, the retention time of the caffeic acid standard does not match the retention time mentioned in the Results and Discussion section. Please correct this, as it is a critical point for analysis. Corrected to 7.8 min retention time for caffeic acid.  7.Comparative discussion: Expand the discussion to include other extraction techniques or green solvents. You may cite the following articles: [Reference 1] [Reference 2] Expanded with ultrasound-assisted, pressurized liquid, and other green solvent extraction, Page 17-18 References updated: [43]","URL":"https:\/\/f1000research.com\/articles\/14-796\/v3#referee-comment-14709","order":1,"name":"referee-comment-14709","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"10.5256\/f1000research.181624.r413222, Nuntawat Khat-Udomkiri, Mae Fah Luang University, Mueang Chiang Rai, Chiang Rai, 27 Sep 2025, version 1, 1 approved, 1 not approved","URL":"https:\/\/f1000research.com\/articles\/14-796\/v1#referee-response-413222","order":2,"name":"referee-response-413222","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"<b>Doungporn Amornlerdpison<\/b>; \n<i>Posted: 03 Oct 2025<\/i>; \n<b>Reviewer 2 (Not Approved)<\/b>  1. You must cite and discuss prior MAE work on P. amboinicus (e.g., the paper you identified: DOI: 10.3303\/CET1972067). Clearly state how your study differs (new endpoints, different solvent system, biological assays, formulation work, scale-up considerations, etc.). Right now the manuscript reads like a repeat of existing work. \n<u>Response<\/u>: Our study differs by evaluating drying methods (tray vs freeze-drying), biological activities (antimicrobial and anti-inflammatory properties) and formulation into a cream with stability testing.  2. Do not claim you \u201coptimized\u201d extraction conditions unless a formal design of experiments (e.g., RSM, factorial) or an explicit optimization protocol was applied. If you did not run an optimization design, rephrase to \u201cscreened\u201d, \u201cevaluated\u201d or \u201cinvestigated\u201d. \n<u>Response<\/u>: We agree and have rephrased \u201coptimized\u201d to \u201cevaluated\u201d, Page 2 unless clearly supported by data.  3. Temperature is critical for MAE outcomes and must be reported for every condition. Report temperature profiles for all MAE runs. \n<u>Response:<\/u> We have added the extraction temperature (40 &deg;C under 450 W microwave power) in the Methods section, Page 3  4. Explain the basis for selecting caffeic acid as the principal marker. Was it the most abundant compound in the extract, the most bioactive, or selected for analytical convenience? \n<u>Response<\/u>: We selected caffeic acid as a marker because it was the most abundant phenolic identified in preliminary HPLC profiling. This justification has been added, Page 3.  5. Natural extracts are mixtures \u2014 identify and quantify multiple major phenolics, not only one. Provide HPLC chromatograms and quantification for at least the top 3\u20135 phenolic compounds. \n<u>Response:<\/u> In this study, a representative phenolic compound, caffeic acid was selected as a biomarker for quantification due to its well-documented bioactivity. Furthermore, the use of total phenolic content (TPC) provides a reliable estimation of the overall phenolic load in the extracts. This approach enables holistic evaluation of the extract's functionality while minimizing the complexity, time, and cost associated with full chromatographic profiling. Future work may include expanded HPLC profiling to better understand the full phenolic spectrum and assess potential synergistic effects among individual compounds. 6. Provide a full cream formulation (all ingredients, % w\/w), the exact amount of extract incorporated, and the rationale for the selected concentration(s). \n<u>Response:<\/u> We have revised the section to provide the complete formulation with all ingredients and concentrations (% w\/w) The formulation was based on the minimum inhibitory concentration (MIC) of PF15, determined to be 7.8 mg\/mL against \n<i>Staphylococcus aureus<\/i>. For a 100 g cream formulation (equivalent to approximately 100 mL), the required amount of extract was calculated as 780 mg, corresponding to 0.78% (w\/w) of the total formulation. The 0.8% concentration was selected to ensure antimicrobial efficacy while maintaining formulation stability, Page 5-6.  7. You state extract was included during emulsification: explain and justify this choice. Phenolic compounds is heat-sensitive; if used in a heated phase, show data proving it remains active after processing. \n<u>Response:<\/u> The PF15 extract was incorporated during the emulsification step to ensure uniform distribution of the bioactive compounds within the cream matrix. Although phenolic compounds are generally considered heat-sensitive, our results demonstrate that the antimicrobial activity of the extract was largely retained after processing. The PF15 extract alone showed inhibition zones of 13\u201315 mm (Table 3, Page 12) against selected skin pathogens, while the formulated PF cream exhibited inhibition zones of 11\u201312 mm (Page 16). The slight reduction indicates minor thermal impact; however, the preserved inhibitory activity confirms that the phenolic constituents remained bioactive after emulsification.  8. You cannot assert \u201cno degradation\u201d based on visual inspection alone. Provide objective stability data: HPLC profiles (initial vs after processing and after stability cycles), TPC, and at least one chemical marker quantified before\/after heating and after storage cycles. Replace subjective color assessments with \u0394E measurements and report \u0394E for color change. \n<u>Response:<\/u> Deleted no degradation. We acknowledge the reviewer\u2019s comment regarding the need for objective stability data. In our study, instead of quantifying individual phenolic compounds within the cream formulation, stability was evaluated based on the functional antimicrobial activity of the product. This approach was selected because the cream contains multiple ingredients. Therefore, antimicrobial efficacy against target skin pathogens was used as a practical stability indicator (Page 16), confirming that the bioactive effect of PF15 was retained after processing and throughout the stability cycles.  9. DPPH must include a positive control (e.g., Trolox or ascorbic acid) and report results as IC\u2085\u2080 or mg Trolox equivalent\/g for comparability. \n<u>Response<\/u>: We used gallic acid as the positive control, consistent with its use in the phenolic content determination. Figure 7 has been updated to include gallic acid as the control.  10. Antimicrobial claims cannot use \u201cbroad-spectrum\u201d without testing a representative panel. \n<u>Response<\/u>: Deleted broad-spectrum from conclusion.  11. Key MAE factors were omitted (type of solvent, solvent concentration, power, temperature, time, solid:liquid ratio). If you want to claim optimization, perform a DOE (factorial or RSM). Otherwise clearly present the work as a screening study. \n<u>Response<\/u>: We have now reported solvent type and concentration (50% ethanol), power (450 W), temperature (40 &deg;C), extraction time (10\u201320 min), and solid\u2013liquid ratio (1:10 v\/v) in the Methods section, Page 3.  12. Rephrase or remove any absolute claims (e.g., \u201cbroad-spectrum\u201d, \u201cno degradation\u201d) unless supported by data. \n<u>Response<\/u>: Statements such as \u201cno degradation\u201d and \u201cbroad-spectrum\u201d have been rephrased to reflect the data accurately.  13. Reword the heat-cycling sentence: heating\u2013cooling is a preliminary shelf-stress test, not a full simulation of long-term stability. Use careful language. \n<u>Response<\/u>: We have reworded this as a \u201cpreliminary stress test to evaluate its stability under accelerated conditions\u201d, Page 16  14. Update and expand references: several cited works are dated \u2014 include recent MAE and cosmeceutical formulation literature. \n<u>Response<\/u>: We have updated the reference with recent MAE\n<b> <\/b>and other green extraction, Page 17-18 References updated: [43]   15. Drying methods, whether heat-based (e.g., tray drying) or non-thermal (e.g., freeze-drying), are well-documented for their influence on phenolic compound stability and retention. Please clarify the rationale for selecting this factor as the focus of investigation rather than prioritizing the optimization of parameters directly affecting the extraction process. \n<u>Response<\/u>: Our unique focus on drying and extraction methods relevance for industrial-scale processing and use and effective cost for commercial.  We believe these revisions strengthen the manuscript significantly by clarifying novelty, improving methodological transparency, and ensuring accurate claims. We hope that the revised version now satisfactorily addresses all reviewer concerns and will be considered suitable for approval","URL":"https:\/\/f1000research.com\/articles\/14-796\/v3#referee-comment-14710","order":3,"name":"referee-comment-14710","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"10.5256\/f1000research.189257.r443296, Nguyen Thanh Triet, University of Medicine and Pharmacy at Ho Chi Minh City, Ho Chi Minh City, 14 Jan 2026, version 2, indexed","URL":"https:\/\/f1000research.com\/articles\/14-796\/v2#referee-response-443296","order":4,"name":"referee-response-443296","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"<b>Doungporn Amornlerdpison<\/b>; \n<i>Posted: 16 Jan 2026<\/i>; \n<b>Response to Reviewer\u2019s Comments<\/b> We sincerely thank the reviewer for the constructive and insightful comments. Our responses and corresponding revisions are provided below. The changes have been made as highlighted in \u201cred font\u201d in revised manuscript (Version 3).  1.\n<b> <\/b>The authors should mention who identified the plant sample? When was the sample collected? What is the name of plant voucher specimen? The plant material (\n<i>Plectranthus amboinicus) <\/i>was taxonomically identified and authenticated by Assoc. Prof. Dr. Yuwalee Unpaprom, a botanist at the Center of Excellence in Agricultural Innovation for Graduate Entrepreneur (Agri Inno), Maejo University, Chiang Mai, Thailand. Fresh leaves were collected in April 2025 from the Maejo area, San Sai District, Chiang Mai Province, Thailand. A voucher specimen has been prepared and deposited at Agri Inno under voucher number Agri Inno- 2574. This information has now been added to the Materials and Methods section of the revised manuscript.  2. The authors should mention in detail how they established the optimization of extraction method? We acknowledge that a formal optimization design, such as Response Surface Methodology or factorial design, was not conducted in this study. Accordingly, the manuscript has been revised to avoid the use of the term \u201coptimized.\u201d Instead, the extraction conditions were \n<b>systematically evaluated and screened<\/b> based on microwave-assisted extraction times (5, 10, and 15 minutes) in combination with two drying methods (tray drying and freeze-drying). The selection of the PF15 condition was based on comparative outcomes including extraction yield, total phenolic content, caffeic acid content, and antioxidant activity. The revised manuscript now clearly reflects this screening-based approach.  3.\n<b> <\/b>The authors need to explain why they used caffeic acid as a marker to evaluate the quality of the extract and optimize the extraction method? Caffeic acid was selected as a marker compound because it is a well-documented major phenolic constituent of \n<i>P. amboinicus<\/i> and is closely associated with its biological activities. Preliminary HPLC screening showed that caffeic acid was the most prominent and consistently detectable peak. Screening with standards of other phenolics, including chlorogenic acid and quercetin, did not show detectable peaks under the applied conditions. Therefore, caffeic acid was used as the representative marker for extract evaluation.  4.\n<b> <\/b>Bioactivity assays need to be described in detail, especially which indicators were used to evaluate the results and the calculation formulas. Bioactivity assays are now described in greater detail in the revised manuscript, including the indicators used and calculation approaches. The concentration of PF15 incorporated into the cream formulation was selected based on antimicrobial efficacy and formulation feasibility. The minimum inhibitory concentration (MIC) of PF15 against \n<i>Staphylococcus aureus<\/i> was determined to be \n<b>7.8 mg\/mL<\/b>. To ensure effective antimicrobial performance in a topical dosage form, PF15 was incorporated at \n<b>0.8% w\/w<\/b>, corresponding to approximately \n<b>8 mg\/g of formulation<\/b>, consistent with the MIC obtained from in vitro testing.  This concentration was selected to ensure sufficient antimicrobial activity while maintaining formulation stability, acceptable sensory properties, and suitability for topical application. Antimicrobial testing of the finished PF15 cream demonstrated inhibition zones of 11\u201313 mm against selected skin pathogens, confirming that the bioactivity of PF15 was retained after emulsification and thermal processing. Although a slight reduction in inhibition zone diameter compared with the crude extract (13\u201315 mm) was observed, the extract remained functionally active within the cream matrix.  5. Did the authors see other peaks after 15 minute-run in HPLC chromatogram? As shown in Figures 3 and 4, no additional significant peaks were observed within the 15-minute HPLC run. The chromatographic analysis was designed as a targeted method focusing on the retention time of caffeic acid, which was clearly detected and consistently resolved under the applied analytical conditions. Therefore, the analytical window was limited to 15 minutes to enable reliable and reproducible comparison of extraction conditions using caffeic acid as the representative marker compound.","URL":"https:\/\/f1000research.com\/articles\/14-796\/v3#referee-comment-15285","order":5,"name":"referee-comment-15285","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"10.5256\/f1000research.189257.r445681, Supenya Chittapun, Thammasat University, Khlong Nueng, 22 Jan 2026, version 2, indexed","URL":"https:\/\/f1000research.com\/articles\/14-796\/v2#referee-response-445681","order":6,"name":"referee-response-445681","label":"Referee Report","group":{"name":"article-reports","label":"Article Reports"}},{"value":"The author(s) declared that no grants were involved in supporting this work","order":7,"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"}]}}