Antifungal and Antioxidant Potential of Methanolic Extracts from Acorus calamus L., Chlorella vulgaris Beijerinck, Lemna minuta Kunth and Scenedesmus dimorphus (Turpin) Kützing

Round 1

Reviewer 1 Report

In this manuscript, the authors reported antifungal and antioxidant activities of several methanolic plant extracts. Total phenolic content and flavonoids content were also tested. While some results are good to know, the whole study lacks enough novelty and significance to warrant publication. 

1. The authors failed to inform the significant scientific and/or social impacts of the study. Why these random chosen plants were chosen? What are the benefits to do the research?
2. The agar well dilution is rather qualitative than quantitative. Although the inhibitory zone diameter and the actual antifungal are positively related, the relationship is usually not linear. There are other better assays that can present inhibitory activity in a more quantitative way, yet they were not chosen. Moreover, why chose 25 ug/mL and only 1 concentration? The word "antimicrobial" is not accurate either; should be "antifungal" all through the paper.
3. What was the purpose to analyze BOTH total phenolic AND total flavonoid? I don't see the necessity. 
4. Figure 1, 2 and 3 can be combined; Figure 4 and 5 did not provide any useful information. The fit model should not use linear model. Well, actually, the data points are too few for any regression analysis.
5. The conclusion was repeating the results. The author should have included some summarized information for the whole study.

As stated reasons above, I am regretfully suggesting that the manuscript should be rejected. 

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments to authors for the article by Toncho Dinev et al.

Introduction

A brilliant and straightforward introduction. I only have one comment here:

Line 72: Maybe the authors should consider rephrasing the term “cardiovascular”. As the sentence reads the syntaxis of “cardiovascular – properties” doesn’t seem right.

And also a general suggestion for this part. It would be great if the authors would consider including a few lines to describe the reasons why the selected strains of Aspergillus flavus, Aspergillus parasiticus, Aspergillus ochraceus, Aspergillus niger, Aspergillus carbonarius, Fusarium graminearum, Fusarium oxysporum, Penicillium chrysogenum and Alternaria alternata have been selected, given that there is a wide range of potential strains to evaluate antifungal activity.

Methodology

The methodology is clear and well written. I only have the following considerations:

Line 93: Seeing that the size and morphology of L. Minuta could lead to misidentification as L. Minor and vice versa (the authors may have already seen the following communication: Morphological study of Lemna minuta Kunth, an alien species often mistaken for the native L. minor L. (Araceae) Simona Ceschin, Ilaria Leacche, Stefano Pascucci, Silverio Abati, Aquatic Botany 131 (2016) 51–56, http://dx.doi.org/10.1016/j.aquabot.2016.01.005) I was wondering how the authors have clarified that the collected plant is in fact L.Minuta.

Line 96: Although the code numbers for C. vulgaris (SKU: 100-CVC00-50) and S. dimorphus (SKU: AC-1002) have been mentioned in previous literature as supplied from the Algae depot – USA, I was unable to visit the site (www.algaedepot.com) and id these products. I was wondering if the authors could provide additional information on these samples, in case the reader (as I have) finds themselves unable to enter the website of the supplier.

Line 98: “Rhizomes of A. calamus were purchased from a local pharmacy”. I wonder if it is possible to have some additional information here like maybe package conditions (example:50g pack of dry rhizomes or fresh rhizomes) and also maybe the origin of the plant might be of value for the reader.

Since further in the results the geographic and climatic conditions are reported to be a potential differentiating factor for the evaluated aspects of these samples it is would be essential to have the basic insights of their origins available in this work.

Results -Discussion

A general suggestion for the presentation of results would be splitting them into two distinct chapters, one for the Antifungal and another for the Antioxidant potential, and also consider pulling all additional discussion in a Discussion segment in between Results and Conclusions.

Regarding the antioxidant potential of the examined samples, it is clear that the authors have made a considerable effort in collecting and comparing previous data for each sample to their findings. However, the way they are presented is causing me great uncertainty to draw conclusions as to where these findings stand in the comparison. It is clear that the extraction method, the extraction solvent, and the preparation of the sample have a significant effect on the overall outcome and in this regard, the comparisons of the methanol extracts of this study ought to be initially made with methanol extracts of previous research (which maybe have also employed similar sample preparations) and then with extracts of hexane or ethanol or any other extraction solvent (which could and would be great to be included in the discussion). To this end, I suggest that the authors consider reorganizing their findings into levels of comparisons which may make it more clear for the reader to understand if similar samples are in fact compared or not and also maybe lead to a better view of which solvent or extraction method or sample preparation may be optimum to work with.

In a similar context, the role of geographic and climatic conditions for antioxidant potential of different samples of the same species is presented as a potential reason for differences in this work's findings as compared to others (and in between others). I was wondering if the authors could elaborate on that. Were there any similarities observed that can relate their samples to others in terms of environmental conditions? (apart from working on the same species) or maybe what might have caused -in terms of environmental factors involved – the samples of this study to respond better/worse than others. This information might be an addition the authors should consider for the discussion of the results and a very helpful one for further research to be able to compare their results with this work in the future.

Line 230-231 and lines 232-233: “C. vulgaris 025”and “another strain of this species – C. vulgaris 030”

I see that these are codes of different strains of the species as described in previous research which is referenced by the authors. However, I wonder if a short explaining of the codes should be included so that it would be clear for the reader - since at the moment these numbers don’t seem to have any particular point other than saying that these are different strains.

Another point would be that a third strain (C. vulgaris 012), as reported in the provided reference (Ghasemi et al.) appears to also have inhibited the growth of A. niger (inhibition zone diameter: 14mm). This point is not mentioned in the manuscript. Perhaps it should be included and combined with the stain mentioned in the manuscript (C. vulgaris 025 – inhibition zone diameter: 11mm) in order to support the overall result of a strain-specific tendency that is already highlighted by the authors.

Line 246: Kindly consider elaborating on the code of the strain.

Line 260: Maybe “mm” is missing after the numbers.

Lines 282-283: Kindly consider altering the order of the references from “Effiong and Sanni [33] and Abbassy et al. [32]” to “Abbassy et al. [32] and Effiong and Sanni [33]”

Lines 311-313: Kindly consider properly altering the order of the references in the following text “Abdel-Karim et al. [44] – 3.17 g GAE/kg, Bhuvana et al. [45] – 2.13 g GAE/kg, El-Chaghaby et al. [43] – 0.71 g GAE/kg, and much lower compared to the findings of Gürlek et al. [46]”

Author Response

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Author Response File: Author Response.doc

Reviewer 3 Report

The paper “Antifungal and Antioxidant Potential of Methanolic Extracts from Acorus calamus, Chlorella vulgaris, Lemna minuta and Scenedesmus dimorphus” is interesting but, in my opinion, several points should be improved before the publication.

1. All plant species, the first time they are named in the text, must be named as genus, species and botanical classifier. Even in the title.
2. Paragraph 2.3. Indicate the range of concentrations used during the reaction, please
3. Paragraph 2.7. I would suggest adding anova analysis or explain better the results please

Author Response

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Author Response File: Author Response.doc

Round 2

Reviewer 1 Report

The authors replied comments from all reviewers and revised the manuscript in a timely manner. I agree with some of the responses and changes, and highly appreciate the authors' great effort. However, I would like to keep my recommendation of "Reject" on this manuscript, as the study and findings are not significant enough to warrant publication. Basically, 1. the authors found that methanolic extracts of some plants have more or less antifungal activity but the assay was rather qualitative; 3. the higher total phenolic content comes with higher flavonoids content, which is nothing interesting or new; 3. the higher GAC and CE come with higher radical scavenging which is nothing new either.

There are too many things left to be desired. Overall, I don't see any novelty or any important findings that are worthy of being published. 

Reviewer 2 Report

Thank you to the authors that have fully addressed all suggestions and the quality of the manuscript has been substantially improved. I consider the current version of the manuscript to be suitable for publication.