Comparison of Gamma-Oryzanol Nanoemulsions Fabricated by Different High Energy Techniques

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Understanding the mechanisms governing the stability of the dispersion system still has important implications for scientific and practical purposes. This study aimed to develop an emulsion with nanometric droplet size and high stability, and then evaluate which technique is the best method to obtain gamma oryzanol nanoemulsions. However, after reading the manuscript, I have the impression that the authors should rephrase Section 3 and first analyze the particle size, then the rheological properties, and finally discuss the physical stability of the nanoemulsion. Because particle size and viscosity significantly affect this parameter (TSI), this will allow for a wider scope for discussion and it will create a more coherent whole. I also have other comments listed below:

L20-21. Insert a comma between each technique.

L24-26. I propose to rephrase this sentence and indicate the best technique for obtaining stable gamma oryzanol nanoemulsion (see L97). TSI cannot be below 0.

L37 and elsewhere. In the text, reference numbers should be placed in square brackets [ ]; for example [1], [1–3] or [1,3].

L59-61. Please add the reference for this fact.

L64-65. 100-300 um? Impossible. Please check! Please add the reference for this fact.

L66. Correct to  ‘microfluidizer’

L65-68. Please add the reference for this fact.

L72. Expand the shortcut UHPH.

L69-75. Please add the reference for this fact.

L81. Please change ‘;’ to dot.

L81084. Please add the reference for this fact.

L85-91. Please expand on this paragraph. What is the current state of knowledge? What effect does it have on the raw material composition of the nanoemulsion? What technique is most often recommended by other researchers? What about the effect of pH?

L101-103. Provide the characteristics and manufacturers of individual nanoemulsion components. Provide the protein content of sodium caseinate. In the case of citrus pectin, provide the degree of methylation.

L103-104. What was the purpose of simulating the raw material composition of the emulsion? Why was it not decided to make all emulsion variants 1-6 at pH 5.3? Didn't increase the share of citrus pectin from 0.3 to 0.7 g cause a decrease in the pH of the emulsion? Was the pH of the emulsion 1-6 adjusted to 7.3?

L115-116. Replace ‘one minute’ to ‘1 minute’.

L117-118. What was used (HCL?) to adjust the pH to 5.3? What decided that the authors made only one variant of the nanoemulsion (5A analog) at pH 5.3?

L127-129. What was the frequency and power of ultrasonic? Was ice cooling of the nanoemulsion used?

L165. Complete the particle size in brackets (Z-average size, D10, D50, D90).

Figure 2. Is it known what happens to nanoemulsions after long storage, e.g. 21 days? It seems that for all variants the TSI index was constantly increasing, which means that they could have been destabilized. Jaki rodzaj niestabilności fizycznej (sedymentacja/kremowanie) wystąpił w przypadku nanoemulsji? Recommended: presentation of delta backscattering profiles of nanoemulsions.

L240. Replace ‘three minutes’ to ‘3 minutes’.

L268. Remove ‘on the other hand’.

L287-291. Style! Please reformulate this sentence.

L300-304. The other authors? Please add references.

L312-313. Please check the statistics for HP2 sample for 0 and 7 days of storage.

L344. Table 100. and UH200)? Correct this sentence.

L348. Correct ‘this same’ to ‘the same’.

L380-382. The emulsion's stability is also significantly influenced by its viscosity. In this case, it seems that viscosity could be a more significant effect than PDI. Recommended: presentation of nanoemulsion viscosity curves.

L402-407. Were the values ​​obtained by other authors similar to this work?

L417. ‘In the studies mentioned before’? Please specify what studies you mean. Add references.

L427-429. Why was this model chosen? Was it the best fit? It is worth providing the matching coefficient (r) in Table 3.

Table 3. Add the results of statistical analysis and standard deviations.

L461. ‘in the oscillatory frequency sweep’? Have these tests been performed? Please explain.

L472. ‘In the three studies’? Please add references in brackets.

L472. It should be ‘flow index’ not ‘consistency index’.

L478. Please add the unit of K.

L477. ‘In the two studies’? Please add references in brackets.

L482-483. morphology analysis on the Figure 8? Please correct this sentence.

L496-501. Style! Unclear. Please rewrite this sentence.

L502. It should be Figure 8G not 8D.

L511. morphology analysis on the Figure 8? Please correct this title.

L525. ‘a positive impact on the rheological properties’? What do you mean? Please explain.

Comments on the Quality of English Language

Language proofreading is recommended.

Author Response

Thank you very much for your willingness and time to review this manuscript. We appreciate your corrections and contributions; they will serve a lot to improve the work. I hope our answers resolve your doubts and concerns.

Regarding to your comment with the rephrase of section 3, we agree with the reordering of the results, and the results are now presented in this order: Coarse emulsion physical stability at line 204, particle size and distribution at line 239, rheological properties at line 357, image analysis at line 411 and physical stability of high energy treated emulsions at line 448. Besides that, we put the new line order of your observation in the letter. 

We also changed the title of the work for the suggestion of one of the reviewers, the new title is: “comparison of gamma oryzanol nanoemulsions fabricated by different high energy techniques.”

 

3. Point-by-point response to Comments and Suggestions for Authors

Comments 1: L20-21. Insert a comma between each technique.

 

Response 1: Thank you for pointing this out. We agree with this comment. We changed and appeared in line 19-20 in red color.

 

Comments 2: L24-26. I propose to rephrase this sentence and indicate the best technique for obtaining stable gamma oryzanol nanoemulsion (see L97). TSI cannot be below 0.

 

Response 2: Thank you for your observation. We made changes in the writing in line 22-25 in red color.

 

Comments 3: L37 and elsewhere. In the text, reference numbers should be placed in square brackets [ ]; for example [1], [1–3] or [1,3].

 

Response 3: Thank you, we agreed with your observation, and we changed the format of all references and appeared in red color.

 

Comments 4: L59-61. Please add the reference for this fact.

 

Response 4: Thank you for your point, we put the proper reference in the L63 in red color.

 

Comments 5: L64-65. 100-300 um? Impossible. Please check! Please add the reference for this fact.

 

Response 5: Thank you for your observation, we changed the writing in the manuscript to L60-63 and put the reference in L63. The changes are in red color.

 

Comments 6: L66. Correct to ‘microfluidizer’.

 

Response 6: Thank you for your correction, we made the change in L61 in red color.

 

Comments 7: L65-68. Please add the reference for this fact.

 

Response 7: Thank you for your point, we put the proper reference in L67 with red color.   

 

Comments 8: Expand the shortcut UHPH.

 

Response 8: Thanks for your observation, we put the shortcut earlier in the text at L64 with red color.

 

Comments 9: L69-75. Please add the reference for this fact.

 

Response 9: Thank you for your observation, we put the reference in L67

 

Comments 10: L81. Please change ‘;’ to dot.

 

Response 10: Thank you for your observation, it was made the change in the writing in L73.

 

Comments 11: L81-84. Please add the reference for this fact.

 

Response 11: Thank you for your observation, we put the proper reference in L72 in red color.

 

Comments 12: L85-91. Please expand on this paragraph. What is the current state of knowledge? What effect does it have on the raw material composition of the nanoemulsion? What technique is most often recommended by other researchers? What about the effect of pH?

 

Response 12: Thank you for your observations, we made changes to this paragraph hoping that we provide enough information. The L78-89 present these changes in red color.

 

Comments 13: L101-103. Provide the characteristics and manufacturers of individual nanoemulsion components. Provide the protein content of sodium caseinate. In the case of citrus pectin, provide the degree of methylation.

 

Response 13: Thank you for your observations; we put the information you requested in L98-99 in red.

 

Comments 14: What was the purpose of simulating the raw material composition of the emulsion? Why was it not decided to make all emulsion variants 1-6 at pH 5.3? Didn't increase the share of citrus pectin from 0.3 to 0.7 g cause a decrease in the pH of the emulsion? Was the pH of the emulsion 1-6 adjusted to 7.3?

 

Response 14. The formulation of the coarse emulsions was carried out as follows: The effect of the concentration of citrus pectin was tested, evaluating two levels (0.3 and 0.7 g) and keeping the concentration of sodium caseinate (3g) and oil (5g) constant and low (3 g) at a pH of 7.3. Higher concentrations of pectin were not tested since concentrations higher than 0.7 g formed gels. Once the amount of pectin was estimated, the effect of the concentration of sodium caseinate was tested by evaluating three levels (3, 4 and 7 g), keeping the pectin concentrations constant (0.7 g the one with the best stability) and maintaining a low amount of oil (5 g), at a pH of 7.3. Once the amount of emulsifiers was estimated, the effect of the oil concentration was tested, evaluating three levels (5, 7 and 9 g) and keeping the concentrations of pectin (0.7 g) and caseinate (7 g) (the concentrations with the highest stability) constant at pH 7.3. Finally, the effect of pH was evaluated, modifying it to 5.3, maintaining the concentrations of pectin at 0.7 g, caseinate at 7 g and oil at 7 g (those that had the highest stability tested in the previous formulations). In this way, when we were evaluating the effect of the concentration of each of the components, the concentrations in the formulation with the best stability were chosen for further stability test.

 

Comments 15: L115-116. Replace ‘one minute’ to ‘1 minute’.

 

Response 15: Thank you for your observation, we made the change in the writing in the L114 in red color.

 

Comments 16: L117-118. What was used (HCL?) to adjust the pH to 5.3? What decided that the authors made only one variant of the nanoemulsion (5A analog) at pH 5.3?

 

Response 16: Thank you for your point, the way in which the pH adjustment was carried out is indicated in L117. The reason why we only modified the pH in formulation 7A was because when we were evaluating the effect of the concentrations of the components with a pH of 7.3, the emulsion with 0.7 g of pectin, 7 g of caseinate and 7 g of oil was the most stable in that pH, when we change de pH, the stability increases in a notable way. Further explanation is in response 14.

 

Comments 17: L127-129. What was the frequency and power of ultrasonic? Was ice cooling of the nanoemulsion used?

 

Response 17: Thank you for the observations. We put the information that you requested in L131-132 in red color.

 

Comments 18: L165. Complete the particle size in brackets (Z-average size, D10, D50, D90)

 

Response 18: Thank you for your observation, we made the changes in L164 in red color.

 

Comments 19: Figure 2. Is it known what happens to nanoemulsions after long storage, e.g. 21 days? It seems that for all variants the TSI index was constantly increasing, which means that they could have been destabilized. Jaki rodzaj niestabilności fizycznej (sedymentacja/kremowanie) wystąpił w przypadku nanoemulsji? Recommended: presentation of delta backscattering profiles of nanoemulsions.

 

Response 19: The main destabilization phenomenon that occur in all treated emulsions was creaming. TSI summarizes variations on the backscattering profile, providing in a simpler and condensate form the information contained in the backscatter figures. These figures can be provided in a supplementary material if you want.

 

Comments 20: L240. Replace ‘three minutes’ to ‘3 minutes’.

 

Response 20: Thank you for your observations, we made the change in the L455 in red color.

 

Comments 21: L268. Remove ‘on the other hand’.

 

Response 21: Thank you for your observation we correct the writing in L486.

 

Comments 22: L287-291. Style! Please reformulate this sentence.

 

Response 22: Thank you for your corrections, we made the change in the writing in the L498-502 in red color.

 

Comments 23: L300-304. The other authors? Please add references.

 

Response 23: Thank you for your observations, we put the correct reference in L 513.

 

Comments 24: Please check the statistics for HP2 sample for 0 and 7 days of storage.

 

Response 24: We reviewed the statistics for the sample that you requested, but there appears to be no error. Could you tell us in more detail what you are referring to?

 

Comments 25: L344. Table 100. and UH200)? Correct this sentence.

 

Response 25: Thank you for your observations, we correct the writing at L278 in red color.

 

Comments 26: L348. Correct ‘this same’ to ‘the same.

 

Response 26: Thank you for your observations, we made the correction in L288-289 in red color.

 

Comments 27: L380-382 The emulsion's stability is also significantly influenced by its viscosity. In this case, viscosity could have a more significant effect than PDI. Recommended: presentation of nanoemulsion viscosity curves.

 

Response 27: Thank you for your observations, but we consider that the rheological behavior of the samples is well summarized in Table 3. These figures can be provided in a supplementary material if you want.

 

Comments 28: L402-407. Were the values ​​obtained by other authors similar to this work?

 

Response 27: Thank you for your observations, the particles sizes reported on that work, were larger than our particles sizes values of the emulsion with the ultrasonic treatment.

 

Comments 29: L417. ‘In the studies mentioned before’? Please specify what studies you mean. Add references.

 

Response 29: Thank you for your observations, we put the correct references in L349 in red color.

 

Comments 30: Why was this model chosen? Was it the best fit? It is worth providing the matching coefficient (r) in Table 3.

 

Response 30: The average r coefficient in the chosen model was 1, in addition the models with more parameters did not significantly improve the fit to the data. We put the coefficient (r) in table 3.

 

Comments 31: Table 3. Add the results of statistical analysis and standard deviations.

 

Response 31: Thank you for your observations, we put the descriptive statistics in table 3 at line 383.

 

Comments 32: L461. ‘in the oscillatory frequency sweep’? Have these tests been performed? Please explain.

 

Response 32: Thank you for your observation, we made the correction in the writing. The test that was done was an analysis of the flow curve in the viscosity profile adjusted to the previously mentioned model. You can find those changes in L397-400.

 

Comments 33: L472. ‘In the three studies’? Please add references in brackets.

 

Response 33: Thank you for your observation we made the correction, because we only mentioned two studies and we put their proper references. You can find those changes on L403 in red color.

 

Comments 34: L472. It should be ‘flow index’ not ‘consistency index’.

 

Response 34: Thank you for your observation, we made the correction at L403 in red color.

 

Comments 35: L478. Please add the unit of K.

 

Response 35: Thank you for your observation, we put the units in L408 in red color

 

Comments 36: L477. ‘In the two studies? Please add references in brackets.

 

Response 36: Thank you for your observation, we put the proper references in L408 in red color.

 

Comments 37: L482-483. morphology analysis on Figure 8? Please correct this sentence.

 

Response 37: Thank you for your observations, we made the corrections in L411 in red color.

 

Comments 38: L496-501. Style! Unclear. Please rewrite this sentence.

 

Response 38: Thank you for your observation. We corrected the writing at L 428-433 in red, and we hope it is clearer that way.

 

Comments 39: L502. It should be Figure 8G not 8D.

 

Response 39: Thank you for your correction, we made the change in L434 in red color.

 

Comments 40: L511. morphology analysis on Figure 8? Please correct this title.

 

Response 40: Thank you for your observation, we changed the figure title in L442 in red color.

 

Comments 41: L525. ‘a positive impact on the rheological properties? What do you mean? Please explain.

 

Response 41: Thank you for your point, we change the writing in that part hoping that our idea is clearer. The changes are in L457-459 in red color.

 

4. Response to Comments on the Quality of English Language

Point 1: Language proofreading is recommended.

Response 1:   The new version of the manuscript has been proofed

Reviewer 2 Report

Comments and Suggestions for Authors

The authors present a study that compares the resulting nanoemulsions fabricated from different high energy methods (homogenization, high pressure homogenizaiton, ultrahigh pressure homogenization, and ultrasonciation). Although they do present a relevant question (which high energy method should one use and what pros and cons might exist) there are a number of things that should be edited and resolved in the manuscript before it can be suitable for publication.

 

Line 70 - The authors mention here, "rapid heating" of the sample if required. Perhaps it needs to be clarified somewhere why rapid heating would be needed in certain cases as homogenization techniques are not defined by heating.

Lines 119-125 - The authors mention that different formulations were tested (shown in Table 1) and that the one with the highest stability (7A) was used with the high-energy techniques. However, the authors don't clearly indicate how they determined 7A to be the most stable.

 

Lines 164-170 - is there a reason why the authors did not measure zeta potential? Since the authors discuss electrostatic repulsion in lines 263-268, it could be important to have this.

There appears to be no description of the statistical analysis in the methods section. The authors must mention this, since their results and conclusion sections mentions statistical differences (e.g., line 345, 376, Table 3, 518).

In addition to a description of the statistical analysis, it would be helpful for the authors to briefly describe how they reproduced their results. Or, in other words, how many independent replicate coarse emulsions and nanoemulsions were made for their study.

Since the authors are aiming to compare the results of different high energy methods, perhaps they could consider combining Figure 3-7 into one (e.g., Figure 3a-e) so that the readers can better compare the results in the figure.

I'm wondering if the authors can perhaps better clarify in their discussion or somewhere in the manuscript their justification for choosing the specific high-energy conditions that they tested. Are the authors sure that the conditions are optimized for each method, or can they justify that the conditions chosen are fair to compare?

Comments on the Quality of English Language

I would advise that the authors reconsider their title to better reflect their research objective (e.g., perhaps comparison of gamma oryzanol nanoemulsions fabricated by different high energy techniques)

The first two sentences of the introduction are difficult to read and should be revised for clarity.

Line 42 - what do authors mean by "biodisponibility"?

Lines 289-291, could the authors rephrase this for clarity?

 

Author Response

Thank you very much for your willingness and time to review this manuscript. We appreciate your corrections and contributions; they will serve a lot to improve the work. I hope our answers resolve your doubts and concerns.

We inform you that as one of the reviewers commented, we reordered the results, placing the analysis of the physical stability of the emulsions with the high energy treatments at the end, in the sense that the results were first correlated with the particles size and distribution, the rheological properties, the image analysis and finally his impact on stability. In that sense, the results are now presented in this order: Coarse emulsion physical stability at line 204, particle size and distribution at line 239, rheological properties at line 357, image analysis at line 411 and physical stability of high energy treated emulsions at line 448. Besides that, we put the new line order of your observation in the letter. 

We also changed the title of the work as you recommended, the new title is: “comparison of gamma oryzanol nanoemulsions fabricated by different high energy techniques”.

 

 

3. Point-by-point response to Comments and Suggestions for Authors

Comments 1: Line 70 - The authors mention here, "rapid heating" of the sample if required. Perhaps it needs to be clarified somewhere why rapid heating would be needed in certain cases as homogenization techniques are not defined by heating.

 

Response 1: Thank you very much for your observation, and we agree with your comment regarding that rapid heating has no relevance in the homogenization technique for the reduction of the droplet size of the emulsion, however the homogenization in the milk industry cannot, so the milk in preheating before the homogenization. The changes in the writing are presented in red on line 63-67.

 

Comments 2: Lines 119-125 - The authors mention that different formulations were tested (shown in Table 1) and that the one with the highest stability (7A) was used with the high-energy techniques. However, the authors don't clearly indicate how they determined 7A to be the most stable.

Response 2: Thank you for your observation, we agreed, and we made changes in the writing in that section of the methods in line 123-124 in red color.

 

Comments 3: Lines 164-170 - is there a reason why the authors did not measure zeta potential? Since the authors discuss electrostatic repulsion in lines 263-268, it could be important to have this.

Response 3: thank you for your observations but we do not measure zeta potential. The specific reason to that, are that we think that we have enough information with the particle size determinations (z-average size, distribution curves, D10, D50, D90), rheological properties and the physical stability. Besides that, the emulsions were not presenting aggregation problems among particles.

 

Comments 4: There appears to be no description of the statistical analysis in the methods section. The authors must mention this, since their results and conclusion sections mention statistical differences (e.g., line 345, 376, Table 3, 518).

In addition to a description of the statistical analysis, it would be helpful for the authors to briefly describe how they reproduced their results. Or, in other words, how many independent replicate coarse emulsions and nanoemulsions were made for their study.

Response 4: Thank you for your observation. We agree with your point. We add in the materials and methods section the statistical analysis and the reproduction of the results in line 204-208

Comments 5: Since the authors are aiming to compare the results of different high energy methods, perhaps they could consider combining Figure 3-7 into one (e.g., Figure 3a-e) so that the readers can better compare the results in the figure.

 

Response 5: Thank you for your point, however we tried to combine all the figure in one or presented in a separate way, but the differences in range of all individual distribution causes that the distribution curves cannot fully appreciate and the tables of D10, D50 and D90 disappeared. Besides that, we consider that the main idea is to present the distributions in separate ways for in that manner we can see the effect of the high energy techniques on the particle distribution. We gave an example, however, if you think that is the best way to present those results, we can try again a new way to reorder these figures.

 

 

Comments 6: I'm wondering if the authors can perhaps better clarify in their discussion or somewhere in the manuscript their justification for choosing the specific high-energy conditions that they tested. Are the authors sure that the conditions are optimized for each method, or can they justify that the conditions chosen are fair to compare?

 

Response 6: Thanks for your observation. In all the techniques we use, we test more conditions in preliminary studies not reported: higher pressures in the HP and UH treatments and a greater or lesser amplitude and time in the US treatment. But in the HP and UH treatments it resulted in a separate emulsion with a paste-like texture, so they were no longer continued. In the US treatment, the other amplitudes and times tested resulted in emulsions with extremely low stability, less than one hour. Thats why we use those conditions and report the results on those treatments.

 

 

4. Response to Comments on the Quality of English Language

Point 1: I would advise that the authors reconsider their title to better reflect their research objective (e.g., perhaps comparison of gamma oryzanol nanoemulsions fabricated by different high energy techniques)

Response 1: Thank you for your point, we changed the title of the work as you recommended.

 

Point 2: The first two sentences of the introduction are difficult to read and should be revised for clarity.

 

Response 2: thank you for your observations, we made a change to the writing in that part of the introduction. You can find those changes in line 30-32 in red color.

 

Point 3: Line 42 - what do authors mean by "biodisponibility"?

 

Response 3: Thank you for your correction, we made the correction of that phrase. You can find the change on line 40 in red color.

 

Point 4: Lines 289-291, could the authors rephrase this for clarity?

 

Response 4: thank you for your observations, we made a change in that part in the writing on lines 498-502 in red color.

Reviewer 3 Report

Comments and Suggestions for Authors

This article provides a detailed study of the effects of component ratios and pH on the stability of crude emulsions; Then the most stable formula is used to load gamma oryzanol, and four kinds of high-energy treatment are carried out to explore the differences in the stability and internal structure of the obtained lotion, so as to evaluate the effect of the four high-energy treatment methods. The results provide important theoretical guidance for the preparation of nanoemulsion loaded with bioactive substances, stabilizing treatment and improving the transfer effect of bioactive substances, but there are some problems to be raised.

Abstract: A more detailed summary of the research results should be made, including the stability effects of the components, microstructure, etc.

Keywords: Should be more refined and accurate, such as stability and viscosity.

Line 129: The homogenization time should be explained, and the following treatment is the same.

Line 215: The stability evaluation of sample 4C seems to deviate from the figure.

Line 330: The horizontal axis of the particle size distribution map should be labeled in more detail within the particle size range, so that the particle size can be clearly seen.

Line 490: The image positioning for the processing method is incorrect.

Conclusion: The experimental results should be more completely summarized, such as the difference between the effects of different treatment methods, and the lotion formula with the highest stability.

Author Response

Thank you very much for your willingness and time to review this manuscript. We appreciate your corrections and contributions; they will serve a lot to improve the work. I hope our answers resolve your doubts and concerns.

We inform you that as one of the reviewers commented, we reordered the results, placing the analysis of the physical stability of the emulsions with the high energy treatments at the end, in the sense that the results were first correlated with the particles size and distribution, the rheological properties, the image analysis and finally his impact on stability. In that sense, the results are now presented in this order: Coarse emulsion physical stability at line 204, particle size and distribution at line 239, rheological properties at line 357, image analysis at line 411 and physical stability of high energy treated emulsions at line 448. Besides that, we put the new line order of your observation in the letter. 

We also changed the title of the work at the suggestion of one of the reviewers, the new title is: “comparison of gamma oryzanol nanoemulsions fabricated by different high energy techniques.”

 

3. Point-by-point response to Comments and Suggestions for Authors

Comments 1: Abstract: A more detailed summary of the research results should be made, including the stability effects of the components, microstructure, etc.

 

Response 1: Thank you for your point; we changed some of the results you requested in the abstract section. The changes are presented on lines 21-25 in red color

 

Comments 2: Keywords: Should be more refined and accurate, such as stability and viscosity.

 

Response 2: Thank you for your observations, we made the change in the keywords as you recommended. You can find those changes on line 27 in red color.

 

Comments 3: Line 129: The homogenization time should be explained, and the following treatment is the same.

 

Response 3: Thank you for your observation. Time was not a changeable variable within the conventional, high and ultra-high pressure treatment conditions, since the emulsion was only passed through the equipment without measuring the time it took to pass through this equipment.

 

Comments 4: Line 215: The stability evaluation of sample 4C seems to deviate from the figure

 

Response 4: Thank you for your observation, we made changes in figure 1 to make the results clearer to the 4C formulation. You can find those changes in figure 1 on line 225.

 

Comments 5: Line 330: The horizontal axis of the particle size distribution map should be labelled in more detail within the particle size range, so that the particle size can be clearly seen

 

Response 5: Thank you for your observation, but the distribution curve values are presented in logarithmic scale, showing constant values in the horizontal axis.

 

Comments 6: Line 490: The image positioning for the processing method is incorrect.

 

Response 7: Thank you for your observations but we cannot fully understand your point, can you clarify that part please? That figure now appeared on line 442.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The authors followed most of my comments. Sometimes I would expect more comprehensive answers, but I appreciate the effort.

Other comments:

L24-25. ‘…and high-pressure treatment the greatest stability (TSI<0.3).’ For what storage time of nanoemulsions is the TSI value given? Complete.

 

L35. Put dot after brackets.

L66. Put dot in the end of line 66.

L67. Remove dot before brackets.

L115. Replace ‘one minute’ to ‘1 minute’.

L117. Put dot in the end of line 117.

L124. Put dot in the end of line 124.

L181, L360, Table 3 and elsewhere. Consistency index. K or k? Unify the abbreviation throughout the article.

L186 and elsewhere. Parameter n (flow behavior, flow behavior index or flow index). The authors use different terms of parameter n. Unify.

L244-245 and Table 2. I asked about the HP2 sample to check the statistics to see if there was an error due to the fact that these values for 0 and 7 days are different and have low standard deviations. But if it has been checked again and it is ok, I have no comments.

Table 3. Add statistical analysis (comparison of two factors (processing and storage) with K and n values). Use different letters to mark important differences.

L547-549.’ …. facilitating the direct incorporation into food products without’ Without what? Part of the text is missing. Complete missing text.

Comments on the Quality of English Language

The text has been corrected. The last sentence at the end of the conclusion section requires completion.

Author Response

1. Summary
Thank you very much for your willingness and time to review this manuscript once again. We appreciate your time in giving us your feedback on our work. We hope that our answers resolve all your doubts. The modifications requested by Reviewer 1 and the editor are highlighted in red. Acknowledgements have been added and some minor typing and grammatical mistakes amended.
3. Point-by-point response to Comments and Suggestions for Authors
Comments 1: L24-25. ‘…and high-pressure treatment the greatest stability (TSI<0.3).’ For what storage time of nanoemulsions is the TSI value given? Complete
Response 1: Thank you for your point. We added the information that you requested at lines 23-24.
Comments 2: L35. Put dot after brackets.
Response 2: Thank you for your correction, we made put the dot at line 35. Comments 3: L66. Put dot in the end of line 66. Response 3: Thank you for your correction, we made put the dot at line 66. Comments 4: L67. Remove dot before brackets. Response 4: Thank you for your corrections, we made the change on line 67. Comments 5: L115. Replace ‘one minute’ to ‘1 minute’. Response 5: Thank you for your observation, we made the change at line 115. Comments 6: L117. Put dot in the end of line 117 Response 6: Thank you for your correction, we put the dot at line 117. Comments 7: L124. Put dot in the end of line 124 Response 7: Thank you for your correction, we put the dot at line 124. Comments 8: L181, L360, Table 3 and elsewhere. Consistency index. K or k? Unify the abbreviation throughout the article. Response 8: Thanks for your observation, we changed and unified the abbreviation (k) on the formula in materials at methods at line 181 Comments 9: L186 and elsewhere. Parameter n (flow behavior, flow behavior index or flow index). The authors use different terms of parameter n. Unify. Response 9: Thank you for your observation, we unified to flow behavior index and made changes to all the text at line 361, 400, 401 and 403 Comments 10: L244-245 and Table 2. I asked about the HP2 sample to check the statistics to see if there was an error due to the fact that these values for 0 and 7 days are different and have low standard deviations. But if it has been checked again and it is ok, I have no comments Response 10: Thank you for your observation and we understand what you meant. Apparently, there was a small reduction in the particle size of the emulsion with the HP2 treatment, however we attribute this to the increase in the PDI and the possible increase in the distribution and average size. And no statistical differences were observed in that sample.
Comments 11: table 3 Add statistical analysis (comparison of two factors (processing and storage) with K and n values). Use different letters to mark important differences. Response 11: Thanks for your point, we did the statistical analysis by two-way ANOVA with treatment and storage time as factors and added the output to Table 3 Comments 12: L547-549.’ …. facilitating the direct incorporation into food products without’ Without what? Part of the text is missing. Complete missing text. Response 12: Thank you for your observation, we made the correction in the text at line 552.

Reviewer 2 Report

Comments and Suggestions for Authors

The authors resolved the concerns I mentioned.

Author Response

Thanks to Reviewer 2 for his/her positive evaluation of the revised version of the manuscript. The comments received have been useful to improve the final version.