Optimization of Coconut Milk Kefir Beverage by RSM and Screening of Its Metabolites and Peptides

Round 1

Reviewer 1 Report

This manuscript is about the optimization of the coconut milk fermentation with kefir grains to produce antioxidant and antimicrobial peptides.

There is no much information on this topic, then this manuscript could contribute to it.

However, it must be improved previously to accept.

Authors should organize the great quantity of information, decide which figures must be in the manuscript and which ones could be in a supplementary material.

First, the title is confuse, authors should organize the main idea of the job, and based on it, define a clear title.

Line 51-52: the phrase starting with “in the study,…” is confuse. Please reformulate.

Lines 69-71. Rewrite this sentence. A verb is missing.

Line 85. Check equation 1. Apparently, you are dividing Ac by Ac. Maybe a parenthesis is missing.

Line 91. Is this section about antimicrobial or antioxidant?

Line 96. Include information of the spectrophotometer

Lines 104-109. Rewrite this paragraph, the method is not understandable.

Line 120. What does TE mean? This method is for FRAP, the antioxidant activity should be expressed as Fe equivalent, since you use a Fe salt as standard.

Line 127. Add information about the ultrasound equipment used

Lines 142-143. Add more information about the mobile phase, was it a gradient? Or an isocratic flow? HPLC method needs to be described, information about the column used is missing, conditions such as flow, temperature, etc, are missing.

Line 144 how fraction 7 was obtained? Add more information.

Line 167: Table must be in methods section.

Lines 193-207. Please explain this paragraph. First you are sayign that the fitting of your models is above 80% for all but two responses (antioxidant activity), but in the second part of the paragraph, you say “ the results showed a significant 203 (p<0.05) lack of fit for the regression models fitted for ABTS+ scavenging activities, antimi-204 crobial activity against B. subtilis, antimicrobial activity against S. Typhimurium, antimi-205 crobial activity against E. coli, and antimicrobial activity against S. aureus”, then, were de models appropriate or not?

Line 212 it is “case”, not “cause”

Line 223, Include  more information for this table. Even the title does not describe the content of the table. What do RC and PV mean? This meaning should be at the end of the table, instead of the title.

I have more comments on section 3, but it is difficult to identify the exact place to insert the comment, since line numbers are missing.

Figure 2 is not complete.

Both Figure 1 and 2 must be improved, the quality is not good, the size is so small that the axis numbers cannot be seen.

Also, Authors must consider if all this figures are necessary in the main body of the manuscript, or this material could be included in a supplementary section.

There are so many figures grouped in one, and also, authors do not describe each one extensively, that’s why I suggest to uses Supplementary section.

Also, I suggest to combine the regression model with the respective response surface graph, it could more informative than in the present way.

Section 3.3: authors are describing the validation of their models, but they do not mention the optimum values for the independent variables, they should include this values either in the table 4 or in the text.

Figures 3 and 4 must be in a Supplementary section. They do not add valuable information in its present form.

Consider moving Figure 5 or table 7 to supplementary material, both have the same information

In the discussion section of metabolites, review the format of the references, for example “Feng et al. (2014) [29] found” in this case, authors should delete (2014).

There are more errors like this, check all the manuscript.

The conclusion must be expanded.

Author Response

Dear reviewer,

Thank you for the comments and suggestions. Enclosed please find the detailed response for Manuscript ID: fermentation-2283416 as attached.

Author Response File: Author Response.pdf

Reviewer 2 Report

Reviewer comments

The paper has shows good premise and originality in using coconut milk as a functional beverage.

1.      The title is not properly written and might incorrectly or confusingly represent what the authors are reporting. Too much information is included in the title. Reduce the length.

2.      Poor use of English in the manuscript requires a major revision of the writing and preferably review by a native speaker. Some areas that require extensive corrections include:

a.      Page 1, Line 41…and food…correct to …and foods…

b.      Page 1, Line 43…among others…this is not informative. Delete.

c.      Page 2 Lines 48-49…were investigated...do not end the sentence with the verb.

d.      Page 2 Line50-51 – this sentence has no verb. I recommend join this sentence with the following sentence.

e.      Page 2 Line 60…the sample was vortex…correct with …was vortexed…

f.       Page 2 Line 76 poor sentence structure and incorrect use of punctuation mark (:)…rewrite…Four types of bacteria used in this study, Salmonella typhimurium ATCC14028, Escherichia coli 77 ATCC12229, Bacillus subtilis ATCC6633, and Staphylococcus aureus ATCC6538, were…

g.      Page 2 Line 80, correct with…An aliquot (100 mL) of the sample…

h.      Page 8 Line??..it can be explained that….has zero meaning. Authors are recommended to review and improve the language used in the manuscript.

i.       Page 8 Line?? 20 – 35^oC temperature. Delete temperature.

3.      The introduction never mentioned any works by other authors on coconut milk, or its perceived high saturated fat content, which might not be healthy or attractive to vegetarians. On page 2 Line 58, the authors noted that low-fat coconut milk was used without further description of the sample.

4.      Page 2 Section 2.1. Based on Fig. 5 and Table 7, authors failed to describe sample CM or its role in the study in section 2.1. Is CM the control of the study?

5.      Page 4 Section 3.1. Authors are encouraged to cite more of the literature to support their interpretation of the statistical analysis of the product sample optimization.

6.      Texts of Fig. 1 and Fig 2 are unreadable.

7.      Pages 8 onward seem to use a different style of writing and citation, which renders this section inconsistent with the preceding section of the manuscript. Authors are recommended to carefully proofread the manuscript prior to subsequent submissions.

8.      Page 10 Gram harmful bacteria or harmful gram-negative bacteria

9.      GABA appeared in very low concentrations (ppm) in either CMK or CM. Can the authors explain whether GABA had any influence on the bioactivity of CMK?

Author Response

Dear reviewer,

Thank you for the comments and suggestions. Enclosed please find the detailed response for Manuscript ID: fermentation-2283416 as attached.

Author Response File: Author Response.pdf

Reviewer 3 Report

This research  investigated the effects of temperature, time, and inoculum size on coconut milk kefir beverage properties,  Response Surface Methodology (RSM) was used to optimize fermentation conditions.,The optimal conditions for coconut milk kefir drink with high antioxidant and antimicrobial activities predicted from the response surface optimizer were temperature 25 ℃, inoculum size 5.4 g/ 100 mL, and culture time 13.5 h.  The metabolic changes of coconut milk kefir at the beginning and end of fermentation were identified using 1H-NMR based metabolomics. The metabolites that were identified are such as γ-Aminobutyric acid, Biotin, Riboflavin, Butyrate, Lactate, Caprylate, and 11 peptide sequences were identified using LC-MS/MS. Coconut milk  fermentation by kefir have more bioactive metabolites than coconut milk.

I think this manuscript can be published after the authors make some revise. My revision opinions are as follows:

1. It will be better authors can supplement some background about nutritional chemical composition of Coconut milk .

2. Table 1 Inoculum size (g/mL),  should be Inoculum size (g/100mL)

3. It will be better authors can summarize or highlight the changes of chemical nutritional composition before and after kefir fermentation.

4. It will be better authors can refining or compress the manuscript.

Author Response

Dear Reviewer,

Thank you for the comments and suggestions. Enclosed please find the detailed response for Manuscript ID: fermentation-2283416 as attached.

Author Response File: Author Response.pdf