Structural characterization and evaluation of antimicrobial and cytotoxic activity of six plant phenolic acids

Monika Kalinowska; Renata Świsłocka; Elżbieta Wołejko; Agata Jabłońska-Trypuć; Urszula Wydro; Maciej Kozłowski; Kamila Koronkiewicz; Jolanta Piekut; Włodzimierz Lewandowski

doi:10.1371/journal.pone.0299372

Peer Review History

Original SubmissionNovember 2, 2023
20 Nov 2023 Decision Letter - Jorddy Neves Cruz, Editor PONE-D-23-32977Structural characterization and evaluation of antimicrobial and cytotoxic activity of six plant phenolic acidsPLOS ONE Dear Dr. Kalinowska, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by Jan 04 2024 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. 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The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. We noticed you have some minor occurrence of overlapping text with the following previous publication(s), which needs to be addressed: - Biological Activity of New Cichoric Acid–Metal Complexes in Bacterial Strains, Yeast-Like Fungi, and Human Cell Cultures In Vitro - https://doi.org/10.3390/nu12010154 3. In your revision ensure you cite all your sources (including your own works), and quote or rephrase any duplicated text outside the methods section. Further consideration is dependent on these concerns being addressed. 4. Thank you for stating the following financial disclosure: National Science Centre research project number 2018/29/B/NZ9/01997 Please state what role the funders took in the study. 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We will update your Data Availability statement to reflect the information you provide in your cover letter. 7. We note that you have stated that you will provide repository information for your data at acceptance. Should your manuscript be accepted for publication, we will hold it until you provide the relevant accession numbers or DOIs necessary to access your data. If you wish to make changes to your Data Availability statement, please describe these changes in your cover letter and we will update your Data Availability statement to reflect the information you provide. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Partly Reviewer #3: Yes Reviewer #4: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: I Don't Know Reviewer #3: Yes Reviewer #4: I Don't Know ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes Reviewer #4: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: No Reviewer #3: Yes Reviewer #4: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: (1) Explanation for varying effects of compounds concentrations on cell viability, why lower concentrations of the some cases significantly increased cell viability, while higher concentrations decreased it? (2) The data in Tables 4 and 5 have serious issues in various parts, such as incorrect values. Reviewer #2: The work aims to do some computational calculation characterizing the chemical structures of selected phenolic acids and presents the results of antimicrobial and cytotoxic studies against two microbial strains and two cell lines. General comments: The manuscript is a mix of literature data and experimental results, however the submission is indicated as research work. In the research article the information provided on figure 1 is not suitable. The information provided in tables 4-5 is characteristic for review paper, not for research work. It is not clear what was a purpose of showing it. The introduction section is far too long. It seems like a part from the review manuscript. Before the reader goes to the aim of the work is already overwhelmed by the number of mentioned studies. Also, since so many research was done on the selected phenolic acids, the novelty of this manuscript has to be much more explained and underlined. It is not clear what was a selection criteria to include these phenolic acids. There is some potential in the selection of structures for the research. They differ by the number of factors, not only by the amount of hydroxyl groups. However this potential was not used for the benefit of this research. The chemical calculation performed are not correlated in the discussion with the observed biological activity. The ionization and lypophilicity influences the penetration of the compound into the cells, hence this information could help to explain the activity or lack of it, however this type of analysis of the generated data was not done. The results and discussion is rather a presentation of results and it lacks the attempt to explain the observations. Other remarks: LogP and pKa values of studied phenolic acids are available in the PubChem database, so what was a purpose to calculate them? Table 4 and 5 are shown before tables 1-3. What solvent was used to prepare working concentrations of studied phenolic acids in the antimicrobial test? Table 6 does not show the units of the presented values. Some of the information presented in this table is already available in the literature, eg: https://doi.org/10.1002/bio.2932 Reviewer #3: The article is on the structural characterization and evaluation of the antimicrobial and cytotoxic activity of six plant phenolic acids. It is a well-designed article, fictionally. There are good results in the article. It has high citation potential. This article can be publish with minor revision But introduction is so long, please write short Reviewer #4: The manuscript is well-written and presents a comprehensive analysis. However, I encountered difficulty in interpreting the figures (bar graph). The figures appear unclear, hindering a clear understanding of the presented data and its significance. It is crucial that the figures are enhanced for better visibility and comprehension. ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: No Reviewer #3: Yes: Mustafa Sevindik Reviewer #4: No ******** [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. Attachments Attachment Submitted filename: Kalinowska_manuscript_correct.docx Attachment Submitted filename: PONE-D-23-32977_reviewer (1) 2023.pdf https://doi.org/10.1371/journal.pone.0299372.r001
Revision 1
12 Jan 2024 Author Response Please find enclosed the answers to reviewers’ comments. Thank you for the comments, suggestions that improved the manuscript. Reviewer #1: (1) Explanation for varying effects of compounds concentrations on cell viability, why lower concentrations of the some cases significantly increased cell viability, while higher concentrations decreased it? Answer: Thank you for the valuable comment. In both tested lines, decreases in cell viability were observed at higher concentrations of the analyzed compounds. However, in fact, a different response was observed for both tested lines at the lower concentrations analyzed. An increase in proliferation was observed in the Caco-2 line. This may be due to the fact that, although these are lines of colorectal adenocarcinoma, they are different from each other. The DLD-1 line is a cell line that is more tumorigenic and exhibited great chances of resistance and recurrence. However, the Caco-2 line is less tumorigenic. The different response to the applied compounds may result from differences in the structure and functioning of both cell lines. It may also be related to the chemical structure of the analyzed compounds. To explain why low concentrations of some of the tested compounds have a stimulating effect on the Caco-2 line, more thorough and detailed research should be carried out on the mechanisms of action of the tested compounds in both cell lines, which is in our scientific plans for the next experiment. The explanation was added to the manuscript. (2) The data in Tables 4 and 5 have serious issues in various parts, such as incorrect values. Answer: Table 4 and 5 have been corrected. Reviewer #2: The work aims to do some computational calculation characterizing the chemical structures of selected phenolic acids and presents the results of antimicrobial and cytotoxic studies against two microbial strains and two cell lines. General comments: The manuscript is a mix of literature data and experimental results, however the submission is indicated as research work. In the research article the information provided on figure 1 is not suitable. The information provided in tables 4-5 is characteristic for review paper, not for research work. It is not clear what was a purpose of showing it. Answer: Thank you for you valuable comments. Figure 1 as well as the tables 4 and 5 have been moved to the supplementary data. The Introduction has been significantly shortened. The introduction section is far too long. It seems like a part from the review manuscript. Before the reader goes to the aim of the work is already overwhelmed by the number of mentioned studies. Answer: The Introduction has been significantly shortened. Also, since so many research was done on the selected phenolic acids, the novelty of this manuscript has to be much more explained and underlined. It is not clear what was a selection criteria to include these phenolic acids. Answer: Thank you for the comment, it helps us to improve the manuscript. The motivation for undertaking the research and the explanation of the selection of compounds are additionally included in the Introduction. “In this study, we will evaluate the antimicrobial activity of these six natural phenolic acids against a bacterial strains living in the digestive system, i.e. Escherichia coli and Lactobacillus rhamnosus based on 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide colorimetric assay. Studies of antimicrobial activity aim to determine the suitability of the tested compounds as potential preservatives with antimicrobial properties against selected food pathogens (E.coli). Moreover, the studies on the effect of the analysed compounds on L. rhamnosus were conducted, which are part of the human intestinal microflora to exclude any negative effects on beneficial probiotic bacteria We will also investigate those phenolic acids cytotoxic effects of on two colorectal adenocarcinoma cell lines:Caco-2 and DLD-1 cell lines in order to analyze potential anticancer activity of studied compounds. Additionally, some structural parameters of the phenolic acids such as lipophilicity coefficient (logP) and reactivity of molecules on the basis of parameters calculated in Gaussian 09W program such aslike ionization potential, electron affinity, electronegativity, chemical hardness, chemical softness, dipole moment and electrophilicity index. The six natural phenolic compounds were chosen on purpose. They have proven biological properties such as antioxidant, antimicrobial or chemopreventive evaluated in different model systems of bacteria or cell lines.The chosen ligands are derivatives of two important aromatic acids, i.e. benzoic and cinnamic acids which differ in the presence of vinyl moiety between the aromatic ring and carboxylic group. Moreover they differ in the number of hydroxyl substituents in the ring (i.e. p-coumaric, caffeic and gallic acid), some of them are caffeic acid esters (i.e. rosmarinic and chlorogenic acids) or gallic acid ester (i.e. tannic acid). All this structural difference influence on the solubility, lipophilicity and reactivity of molecules. Therefore the following questions may arise: (1) does the number of -OH substituents in the aromatic ring is the main factor that affect the antimicrobial and cytotoxic activity of compounds (toward selected bacteria and cell lines), (2) does the conjugation of caffeic and gallic acid with other molecules influence on the physico-chemical and biological properties of molecules, (c) do the selected experimental (logP) and theoretical (e.g. energy of HOMO or LUMO orbitals) correlates with the biological activity of molecules?” There is some potential in the selection of structures for the research. They differ by the number of factors, not only by the amount of hydroxyl groups. However this potential was not used for the benefit of this research. The chemical calculation performed are not correlated in the discussion with the observed biological activity. The ionization and lypophilicity influences the penetration of the compound into the cells, hence this information could help to explain the activity or lack of it, however this type of analysis of the generated data was not done. The results and discussion is rather a presentation of results and it lacks the attempt to explain the observations. Answer: Thank you for the comment. The discussion was improved and the structure-activity discussion was in-depth, taking into account theoretical parameters and lipophilicity. The discussion is in the section Results and discussion, Antimicrobial activity, Cytotoxic activity and final remarks in Conclusions. Other remarks: LogP and pKa values of studied phenolic acids are available in the PubChem database, so what was a purpose to calculate them? Answer: The table was removed to supplementary. Collecting data in one place shortens the time needed to search it. Table 4 and 5 are shown before tables 1-3. Answer: It was corrected. What solvent was used to prepare working concentrations of studied phenolic acids in the antimicrobial test? Answer: The stock solution was prepared by dissolving it in TrisHCl buffer prior analysis. Table 6 does not show the units of the presented values. Some of the information presented in this table is already available in the literature, eg: https://doi.org/10.1002/bio.2932 Answer: We agree. The table was removed to supplementary. Reviewer #3: The article is on the structural characterization and evaluation of the antimicrobial and cytotoxic activity of six plant phenolic acids. It is a well-designed article, fictionally. There are good results in the article. It has high citation potential. This article can be publish with minor revision. But introduction is so long, please write short Answer: The Introduction has been significantly shortened. Reviewer #4: The manuscript is well-written and presents a comprehensive analysis. However, I encountered difficulty in interpreting the figures (bar graph). The figures appear unclear, hindering a clear understanding of the presented data and its significance. It is crucial that the figures are enhanced for better visibility and comprehension. Answer: Thank you for the comments. The figures were corrected. Journal Requirements: 2. We noticed you have some minor occurrence of overlapping text with the following previous publication(s), which needs to be addressed: - Biological Activity of New Cichoric Acid–Metal Complexes in Bacterial Strains, Yeast-Like Fungi, and Human Cell Cultures In Vitro - https://doi.org/10.3390/nu12010154 Answer: Thank you for the comments. The experimental part, which is similar in both publications, has been corrected the to get rid of repetitions in the description of the methodology. 3. In your revision ensure you cite all your sources (including your own works), and quote or rephrase any duplicated text outside the methods section. Further consideration is dependent on these concerns being addressed. Answer: Thank you. It was checked. 4. Thank you for stating the following financial disclosure: National Science Centre research project number 2018/29/B/NZ9/01997 Please state what role the funders took in the study. If the funders had no role, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript." If this statement is not correct you must amend it as needed. Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf. Answer: It is correct, the funder (National Science Centre) had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. 5. Thank you for stating the following in the Acknowledgments Section of your manuscript: This research was funded by National Science Centre (NCN), Poland, under the research project number 2018/29/B/NZ9/01997. We note that you have provided funding information that is not currently declared in your Funding Statement. However, funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form. Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows: National Science Centre research project number 2018/29/B/NZ9/01997 Please include your amended statements within your cover letter; we will change the online submission form on your behalf. Answer: I understand. The information about funding was deleted from the Acknowledgement and removed to the Funding Statement. 6. In your Data Availability statement, you have not specified where the minimal data set underlying the results described in your manuscript can be found. PLOS defines a study's minimal data set as the underlying data used to reach the conclusions drawn in the manuscript and any additional data required to replicate the reported study findings in their entirety. All PLOS journals require that the minimal data set be made fully available. For more information about our data policy, please see http://journals.plos.org/plosone/s/data-availability. Answer: The data that support the findings of this study will be openly available in a repository. Unfortunately, we are currently still waiting to obtain a doi number. It will be added as soon as possible. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0299372.r002
9 Feb 2024 Decision Letter - Jorddy Neves Cruz, Editor Structural characterization and evaluation of antimicrobial and cytotoxic activity of six plant phenolic acids PONE-D-23-32977R1 Dear Dr. Kalinowska, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Jorddy Neves Cruz Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0299372.r003
Formally Accepted
29 Apr 2024 Acceptance Letter - Jorddy Neves Cruz, Editor PONE-D-23-32977R1 PLOS ONE Dear Dr. Kalinowska, I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team. At this stage, our production department will prepare your paper for publication. This includes ensuring the following: * All references, tables, and figures are properly cited * All relevant supporting information is included in the manuscript submission, * There are no issues that prevent the paper from being properly typeset If revisions are needed, the production department will contact you directly to resolve them. If no revisions are needed, you will receive an email when the publication date has been set. At this time, we do not offer pre-publication proofs to authors during production of the accepted work. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few weeks to review your paper and let you know the next and final steps. Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. If we can help with anything else, please email us at customercare@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Jorddy Neves Cruz Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0299372.r004

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