Round 1

Reviewer 1 Report (Previous Reviewer 1)

I have read the corrected version of the manuscript and appreciate the authors' consideration in satisfactorily attending my suggestions made to the original version of the manuscript. I have no further review comments. 

Author Response

Dear Reviewer,

Thank you for taking time to carefully review the paper „The effects of brewer’s spent yeast (BSY) inclusion in dairy sheep’s diets on ruminal fermentation and milk quality parameter”, ID: agriculture-2521086.

Reviewer 2 Report (New Reviewer)

Comments and Suggestions for Authors

Manuscript ID: Agriculture-2521086-peer-review-R1,

entitled " The effects of brewer’s spent yeast (BSY) inclusion in dairy sheep’s diets on ruminal fermentation and milk quality parameter."

General comments.

The manuscript provides an alternative feed additive by the inclusion of brewer’s spent yeast (BSY) in dairy sheep diets.

This work still needs major revision. It could be considered for publication after a major revision. Moreover, the manuscript needs intensive English proofreading.

Abstract

1.      L18 add an article “the” before beer.

2.      L20 add an article “a” before completely.

3.      L23 replace “of propionate and of the acetate/propionate ratio” with “in propionate and in the acetate/propionate ratio. And delete rumen and add ruminal before propionate”.

4.      L26 change to the milk's total polyphenols.

5.      L30 change has to could have. And delete some of.

6.      L31 delete an article “the” before consumers.

Introduction

7.      L35-37 Rephrase to” Nowadays, there is an increasing interest in a healthy and balanced diet, but the explosive population growth makes supplying the human diet with all necessary nutrients become a challenge for the food industry gradually.

8.      L37 correct to it is important.

9.      L37-42 Add references.

10.  L45 you can also add the refrrences of https://doi.org/10.1080/1745039X.2021.1920819; https://doi.org/10.3389/fvets.2020.00181

11.  L59-60 Rephrase it.

12.  L61 Change yet to however.

13.  L65-67 Add references.

14.  L71-76 Rephrase it.

15.  The aim of the study is not clear. Rephrase it.

Material and methods

16.  My greatest perplexity concerns are using two groups and the inclusion of BSY at one level. In addition, dairy sheep were in the late-lactation period. Mostly, in experiments on dairy sheep, the researchers conducted studies on the early-lactation or the mid-lactation periods.  During early-lactation or mid-lactation periods, the lamb is dependent on the milk production of the mother. Therefore, the study of the effect of the inclusion of alternative sources on milk production will be more significant.

17.  Why did you use 5.4% of BSY inclusion in the diet?

18.  L88 change at the end of lactation to the late-lactation period.

19.  What is the duration of milk production of dairy Blackhead sheep?

20.  Mention the number of parities of ewes.

21.  Mention the experimental conditions and when was the experiment conducted.

22.  L83 Insert the subtitle “2.1. Animals, diets, and experimental design” and renumber the rest of the subtitles.  And make a Statistical analysis in subtitle 2.8. Statistical analysis.

23.  L104-109 prefer to mention these data in a separate Table and transfer it into the result section and discuss it also in the discussion section.

24.  L121-122 under Table 2, clarify How Milk Feed Units, Intestinally Digestible Protein allowed by rumen- available Energy and Intestinally Digestible Protein allowed by rumen-available Nitrogen were calculated or determined?

25.  L129 until the completion of determinations convert to until the complete analysis.

26.  19. L137 correct Proximal chemical composition to Proximate chemical composition. And mention it in subtitle 2.3. Proximate chemical composition of diets and milk samples and so on the others.

27.  L171 mention the thiobarbituric acid reactive substances (TBARS).

28.  L180 convert until after the completion of the volatile fatty acid (VFAs) to until measuring the volatile fatty acid (VFAs).

29.  L188 How the statistical analysis of the results was performed by analysis of variance (ANOVA)? There are only two groups, thus the statistical analysis of the results is performed by T-test, not ANOVA. Plz, correct that. 

30.  L189 bifactorial (storage time and diet) in the case of TBARS and mono factorial (diet) in the case of the other parameters. This sentence was not clear and should mention the other parameters first and followed by the TBARS parameter, not vice versa.

Results

31.  L205 being close to the “safe zone” from the point of view of the ruminal normal activity (fiber degradation and microbial protein synthesis, etc.). transfer it to the discussion section.

32.  L206-208 rephrase to Ammonia concentration and total VFAs concentration were not influenced by the inclusion of BSY.

33.  L208 replace BSY diet by the inclusion of BSY and in other places in the MS.

34.  L209 delete the main and the whole MS.

35.  L209-211 rephrase it and mention the p-value for statistically significant results while referring to non-significant statistical results as "tended to decrease or increase". And so on the whole manuscript.

36.  L211-212 Rephrase to “Consequently, Acetate: propionate ratio significantly increased (p= 0.006) in the BSY group.

37.  L214-217 rephrase it according to the previous comments.

38.  L217-218 transfer it to the discussion section and no references are mentioned in the results section.

39.  L225 change Ration between omega 6 and omega 3 fatty acids to omega 6: omega 3 ratio.

40.  L233-234 transfer it to the discussion section.

41.  L237 mention the full name of calcium, phosphorus, and zinc in Table 6.

42.  L239-240 rephrase it.

43.  L247 change to the inclusion of BSY led to a significant increase in milk total polyphenol content (p=0.004). However, the milk antioxidant capacity and vitamin E content in milk were not significantly influenced by the inclusion of BSY.

44.  L251-252 delete the thiobarbituric acid reactive substances method and mention it in Table 8 title in L256.

45.  L252 and also in table 8 mention after defrosting time and after 24 h of storage time instead of t0 and t1.

46.  In Table 8 replace diet with treatment.

47.  L258 change it to after 24 h of storage time.

48.  L t0 (fresh sample) This is not a fresh sample, but it is defrosted.

49.  L 263 add increased by increasing storage time.

50.  L278 transfer (p=0.004) to after decreased in L277 and add in milk after the content word.

51.  L278-279 rephrase it to however, the conjugated trienes content in milk tended to decrease by the inclusion of BSY (fig. 1 b).

52.  L298 Unsaturated fatty acids (and, distinctly, omega fractions) did not clear.

53.  L300 add a comma after “primary”.

Discussion

54.   L311 clarify what is meaning this sentence “such as mixing various fractions obtained through the technology flux”

55.  L316-318 rephrase This places BSY between this group and soybean meal, which opens the possibility of replacing the latter in some animal categories, that have intermediate protein requirements.

56.  L322 rephrase this according to the previous comment in the result section.

57.  L336-340 this explanation doesn’t refer to the inclusion of BSY. Delete this explanation.

58.  Avoid writing the P value in the discussion section and mention only the trend of the effect of the inclusion.

59.  L343-352 rephrase this part in the discussion on the effect of BSY inclusion in VFA and add more scientific explanations and add references.

60.  L 353- 364 make these lines as one paragraph.

61.  54. L369 -371 The BSY diet induced a significantly increased in SFA and a significantly decreased in MUFA and PUFA composition in milk. These results are not suitable for consumers' health.

62.  L 404 the antioxidant capacity of the milk was not significantly influenced by the BSY diet, which opposes the increased content of total polyphenols in milk by the BSY inclusion.

63.  L414- 416 add reference.

64.  L420 change but to, however.

65.  L421 change upon inclusion of to when the inclusion of.

66.  L422-424 Transfer to L 416 after “nutritional quality”.  

67.  L424 don’t repeat “ is scarce”. You can use other sentences such as “very few publications can be found”/ very few publications are available in the literature that discusses/ However, studies on ….. are still lacking/ little attention has been paid/given to.

68.  L430 I think, by the BSY, is wrong.

Conclusion 

 

69.  It must be improved and summarized. Avoid redundant sentences and the mention of results.

 

The manuscript needs intensive English proofreading.

 

Author Response

Dear Reviewer,

Thank you for taking time to carefully review the paper „The effects of brewer’s spent yeast (BSY) inclusion in dairy sheep’s diets on ruminal fermentation and milk quality parameter”; ID: agriculture-2521086 and sending us a clear form of your valuable suggestions and observations. After considering all your suggestions we are resubmit the revised form of the paper.

General comments: The manuscript provides an alternative feed additive by the inclusion of brewer’s spent yeast (BSY) in dairy sheep diets. This work still needs major revision. It could be considered for publication after a major revision. Moreover, the manuscript needs intensive English proofreading.

Response: Thank you for your valuable observations and suggestions. We have addressed the issues you have raised. We have used yellow colour to indicate the changes we made to the text. Also, we have updated several lines of the manuscript in order to correct them. (Lines 45-50, 53-54, 55-58, 245-248, 413-415).

Abstract

Point 1: L18 add an article “the” before beer.

Response 1: Thank you for your observation. We have added the article “the” before beer. (Line 18)

Point 2: L20 add an article “a” before completely.

Response 2: Thank you for your observation. We have added an article “a” before completely. (Line 20)

Point 3: L23 replace “of propionate and of the acetate/propionate ratio” with “in propionate and in the acetate/propionate ratio. And delete rumen and add ruminal before propionate”.

Response 3: Thank you for your suggestion. We have added this statement in the text. (Line 23)

Point 4: L26 change to the milk's total polyphenols.

Response 4: Thank you for your observation. We have changed to the milk's total polyphenols. (Line 26)

Point 5: L30 change has to could have. And delete some of.

Response 5: Thank you for your observation. We have made these changes in the text. (Line 29)

Point 6: L31 delete an article “the” before consumers.

Response 6: Thank you for your observation. We have deleted article “the” before consumers. (Line 30)

Introduction

Point 7: L35-37 Rephrase to” Nowadays, there is an increasing interest in a healthy and balanced diet, but the explosive population growth makes supplying the human diet with all necessary nutrients become a challenge for the food industry gradually.

Response 7: Thank you for your suggestion. We have rephrased this phrase. (Lines 35-37)

Point 8: L37 correct to it is important.

Response 8: Thank you for your observation. We have corrected in the text. (Line 37)

Point 9: L37-42 Add references.

Response 9: Thank you for your suggestion. We have added reference in the text. (Line 40)

Point 10:  L45 you can also add the references of https://doi.org/10.1080/1745039X.2021.1920819; https://doi.org/10.3389/fvets.2020.00181

Response 10: Thank you for your suggestion. We have added these references in the test (Line 45)  

Point 11:  L59-60 Rephrase it.

Response 11: Thank you for your suggestion. We have rephrased this phrase. (Lines 60-62)

Point 12:  L61 Change yet to however.

Response 12: Thank you for your suggestion. We have made this change in the text. (Line 59)

Point 13:  L65-67 Add references.

Response 13: Thank you for your suggestion. We have added reference in the text. (Line 67)

Point 14:  L71-76 Rephrase it.

Response 14: Thank you for your suggestion. We have rephrased this phrase. (Line 70-77)

Point 15:  The aim of the study is not clear. Rephrase it.

Response 15: Thank you for your suggestion. We have rephrased the aim of the study. (Lines 78-80)

Material and methods

Point 16:  My greatest perplexity concerns are using two groups and the inclusion of BSY at one level. In addition, dairy sheep were in the late-lactation period. Mostly, in experiments on dairy sheep, the researchers conducted studies on the early-lactation or the mid-lactation periods.  During early-lactation or mid-lactation periods, the lamb is dependent on the milk production of the mother. Therefore, the study of the effect of the inclusion of alternative sources on milk production will be more significant.

Response 16: This experiment follows a previous trial, in which more than one level of inclusion were studied; we have selected the dose that was the most interesting from the point of view of effects on rumen parameters and further sought to detail its effects on fine milk constituents. We chose late lactation as the decrease of milk yield, specific to this stage, causes problems to the farmers, making them interested in new feeding strategies that can lead to premium products (e.g., richer in anti-oxidants).

Point 17:  Why did you use 5.4% of BSY inclusion in the diet?

Response 17: BSY totally replaced the soybean meal; this inclusion level is imprinted by the proportion of soybean meal in the Control group, within a diet that is specific to the animal category (milking sheep)

Point 18:  L88 change at the end of lactation to the late-lactation period.

Response 18: Thank you for your suggestion. We have changed to the late-lactation period in the text. (Line 90)

Point 19:  What is the duration of milk production of dairy Blackhead sheep?

Response 19: This local sheep has a lactation of more than 200 days, provided that appropriate feeding and good management of genetic resources is ensured

Point 20:  Mention the number of parities of ewes.

Response 20: Thank you for your observation. We have mentioned also the parity number, in M&M chapter. (Line 91)

Point 21:  Mention the experimental conditions and when was the experiment conducted.

Response 21: Thank you for your suggestion. We have added this information to the text. (Line 96-101)

Point 22:  L83 Insert the subtitle “2.1. Animals, diets, and experimental design” and renumber the rest of the subtitles.  And make a Statistical analysis in subtitle 2.8. Statistical analysis.

Response 22: Thank you for your suggestion. We have made the changes in the text. (Lines 83, 121, 131, 144, 152, 157, 180)

Point 23:  L104-109 prefer to mention these data in a separate Table and transfer it into the result section and discuss it also in the discussion section.

Response 23: Thank you for your suggestion. We have added these data in one table into the Results chapter and discuss it in the discussion section (Lines 210-211, 338-346)

Point 24:  L121-122 under Table 2, clarify How Milk Feed Units, Intestinally Digestible Protein allowed by rumen- available Energy and Intestinally Digestible Protein allowed by rumen-available Nitrogen were calculated or determined?

Response 24: Line 24: Thank you for your suggestion. We have added the references used to calculate Milk Feed Units, Intestinally Digestible Protein allowed by rumen- available Energy and Intestinally Digestible Protein allowed by rumen-available Nitrogen. (Line 120).

Point 25:  L129 until the completion of determinations convert to until the complete analysis.

Response 25: Thank you for your suggestion. We have made this change in the text. (Line 124)

Point 26: 19. L137 correct Proximal chemical composition to Proximate chemical composition. And mention it in subtitle 2.3. Proximate chemical composition of diets and milk samples and so on the others.

Response 26: Thank you for your suggestion. We have made these changes in the text. (Lines 83, 121, 131, 144, 152, 157, 180)

Point 27:  L171 mention the thiobarbituric acid reactive substances (TBARS).

Response 27: Thank you for your suggestion. We have mentioned the thiobarbituric acid reactive substances (TBARS) in the text. (Line 171)

Point 28:  L180 convert until after the completion of the volatile fatty acid (VFAs) to until measuring the volatile fatty acid (VFAs).

Response 28: Thank you for your suggestion. We have made this change in the text. (Lines 184-185).

Point 29:  L188 How the statistical analysis of the results was performed by analysis of variance (ANOVA)? There are only two groups, thus the statistical analysis of the results is performed by T-test, not ANOVA. Plz, correct that. 

Response 29: Thank you for your observation. We have corrected this phrase. (Lines 194-198)

Point 30:  L189 bifactorial (storage time and diet) in the case of TBARS and mono factorial (diet) in the case of the other parameters. This sentence was not clear and should mention the other parameters first and followed by the TBARS parameter, not vice versa.

Response 30: Thank you for your observation. We have rephrased this statement. (Lines 194-198)

Results

Point 31:  L205 being close to the “safe zone” from the point of view of the ruminal normal activity (fiber degradation and microbial protein synthesis, etc.). transfer it to the discussion section.

Response 31: Thank you for your suggestion. We have transferred this phrase to the discussion section. (Lines 347-349)

Point 32:  L206-208 rephrase to Ammonia concentration and total VFAs concentration were not influenced by the inclusion of BSY.

Response 32: Thank you for your suggestion. We have rephrased this sentence. (Lines 226-228)

Point 33:  L208 replace BSY diet by the inclusion of BSY and in other places in the MS.

Response 33: Thank you for your suggestion. We have replaced BSY diet by the inclusion of BSY in all manuscript.

Point 34:  L209 delete the main and the whole MS.

Response 34: Thank you for your suggestion. We have deleted the word main.

Point 35:  L209-211 rephrase it and mention the p-value for statistically significant results while referring to non-significant statistical results as "tended to decrease or increase". And so, on the whole manuscript.

Response 35: Thank you for your suggestion. We have mentioned the p-value for statistically significant results and we referred as tendency to non-significant statistical results. (Line 229-231)

Point 36:  L211-212 Rephrase to “Consequently, Acetate: propionate ratio significantly increased (p= 0.006) in the BSY group.

Response 36: Thank you for your suggestion. We have made this change in the text. (Line 231)

Point 37:  L214-217 rephrase it according to the previous comments.

Response 37: Thank you for your suggestion. We have rephrased this sentence. (Lines 235-236)

Point 38:  L217-218 transfer it to the discussion section and no references are mentioned in the results section.

Response 38: Thank you for your suggestion. We have transferred it to the discussion section. (Lines 369-370)

Point 39: L225 change Ration between omega 6 and omega 3 fatty acids to omega 6: omega 3 ratio.

Response 39: Thank you for your suggestion. We have changed this in the text. (Line 243)

Point 40:  L233-234 transfer it to the discussion section.

Response 40: Thank you for your suggestion. We have transferred this to the discussion section. (Lines 405-406).

Point 41:  L237 mention the full name of calcium, phosphorus, and zinc in Table 6.

Response 41: Thank you for your suggestion. We have mentioned the full name of the determined minerals in the table. (Lines 252-253)

Point 42:  L239-240 rephrase it.

Response 42: Thank you for your suggestion. We have rephrased this in the manuscript. (Line 254-256)

Point 43:  L247 change to the inclusion of BSY led to a significant increase in milk total polyphenol content (p=0.004). However, the milk antioxidant capacity and vitamin E content in milk were not significantly influenced by the inclusion of BSY.

Response 43: Thank you for your suggestion. We have rephrased this phrase. (Lines 263-265).

Point 44:  L251-252 delete the thiobarbituric acid reactive substances method and mention it in Table 8 title in L256.

Response 44: Thank you for your suggestion. We have deleted the thiobarbituric acid reactive substances method and mention it in Table 9. (Line 271)

Point 45:  L252 and also in table 8 mention after defrosting time and after 24 h of storage time instead of t0 and t1.

Response 45: Thank you for your suggestion. We have changed t0 and t1 with defrosting time and after 24 h of storage time. (Lines 272-273)

Point 46:  In Table 8 replace diet with treatment.

Response 46: Thank you for your suggestion. We have replaced the diet with treatment (Lines 272-273).

Point 47:  L258 change it to after 24 h of storage time.

Response 47: Thank you for your suggestion. We have mentioned this information in the table and deleted it from the table parameters description.

Point 48:  L t0 (fresh sample) This is not a fresh sample, but it is defrosted.

Response 48: Thank you for your suggestion. We have changed in the text. (Line 276)

Point 49:  L 263 add increased by increasing storage time.

Response 49: Thank you for your suggestion. We have added this in the text. (Line 278)

Point 50:  L278 transfer (p=0.004) to after decreased in L277 and add in milk after the content word.

Response 50: Thank you for your suggestion. We have made this change in the text. (Lines 292-293)

Point 51:  L278-279 rephrase it to however, the conjugated trienes content in milk tended to decrease by the inclusion of BSY (fig. 1 b).

Response 51: Thank you for your suggestion. We have rephrased this statement. (Lines 293-294)

Point 52:  L298 Unsaturated fatty acids (and, distinctly, omega fractions) did not clear.

Response 52: Thank you for your observation. We have changed to unsaturated fatty acids, especially omega 3 and omega 6 fractions. (Line 313).

Point 53:  L300 add a comma after “primary”.

Response 53: Thank you for your observation. We have added a comma after “primary”. (Line 315)

Discussion

Point 54:   L311 clarify what is meaning this sentence “such as mixing various fractions obtained through the technology flux”

Response 54: Thank you for our observation. Ee have updated the text, indicating the factors of variation of the chemical composition, we also have inserted on reference to support this statement. (Line 325-327)

Point 55:  L316-318 rephrase This places BSY between this group and soybean meal, which opens the possibility of replacing the latter in some animal categories, that have intermediate protein requirements.

Response 55: Thank you for your suggestion. We have rephrased this and added in the text. (Lines 332-334).

Point 56:  L322 rephrase this according to the previous comment in the result section.

Response 56: Thank you for your suggestion. As we were not sure to which of the previous comments to you refer, we have addressed both (Point 23 and Point 55). Therefore, we moved the mentioned text and discussed it (Point 23) and also rephrase it (Point 55). The concerned text is at Lines 338-340.

Point 57:  L336-340 this explanation doesn’t refer to the inclusion of BSY. Delete this explanation.

Response 57: Thank you for your suggestion. We have deleted this explanation.

Point 58:  Avoid writing the P value in the discussion section and mention only the trend of the effect of the inclusion.

Response 58: Thank you for your suggestion. We have deleted the p values from the results section and we have mentioned the trend of the effects observed.

Point 59:  L343-352 rephrase this part in the discussion on the effect of BSY inclusion in VFA and add more scientific explanations and add references.

Response 59:  Thank you for your observation. We have changed updated the discussion on the effect of BSY on VFAs, by using info that are substantiated by references. (Lines 354-367)

Point 60:  L 353- 364 make these lines as one paragraph.

Response 60: Thank you for your suggestion. We have made these lines one paragraph. (Lines 370-379)

Point 61:  54. L369 -371 The BSY diet induced a significantly increased in SFA and a significantly decreased in MUFA and PUFA composition in milk. These results are not suitable for consumers' health.

Response 61: Thank you for your observation. The findings of our study on the main classes of milk fatty acids indicated that the inclusion of the BSY caused a considerable increase in SFA and a significant decrease in MUFA and PUFA composition, which is not suitable for consumers. However, the study also demonstrated that inclusion of the BSY can induce other beneficial effects in milk fatty acids, such as a significant decrease in Ω6 fatty acids, which contributed to a significant decrease in the Ω6/Ω3 ratio, which is important for consumer health considering that a low Ω6/Ω3 ratio may help to prevent some autoimmune, inflammatory, and cardiovascular diseases. Furthermore, the addition of BSY resulted in a considerable rise in short chain fatty acids (SCFA) and some medium chain fatty acids. According to various studies, SCFAs and MCFAs are a better source of energy in human nutrition than long-chain fatty acids due to their smaller molecule and the pathways of their transport through the organism, and they can play a potential role in the protection process against pathogen growth and may exert anti-inflammatory activity.

Point 62:  L 404 the antioxidant capacity of the milk was not significantly influenced by the BSY diet, which opposes the increased content of total polyphenols in milk by the BSY inclusion.

Response 62: Thank you for your valuable observation. The findings of our study demonstrated that the antioxidant capacity of the milk samples did not statistical differ, but the milk total polyphenol content was favourably influenced by the inclusion of the BSY. One probable explanation is that polyphenols are a type of hydrophilic antioxidant, and possibly they are not the primary milk antioxidants. For example, a powerful antioxidant in milk is represented by the casein fractions, whose concentration in milk are substantially higher than that of polyphenols. As can be shown, in our study, the inclusion of the BSY had no effect on casein concentration.

Point 63:  L414- 416 add reference.

Response 63: Thank you for your observation. We have added reference in the text. (Line 432)

Point 64:  L420 change but to, however.

Response 64: Thank you for your suggestion. We have changed this in the text. (Line 433)

Point 65:  L421 change upon inclusion of to when the inclusion of.

Response 65: Thank you for your suggestion. We have made this change in the text (Line 434)

Point 66:  L422-424 Transfer to L 416 after “nutritional quality”.  

Response 66: Thank you for your suggestion. We have made this change in the text. (Line 440-442)

Point 67:  L424 don’t repeat “is scarce”. You can use other sentences such as “very few publications can be found”/ very few publications are available in the literature that discusses/ However, studies on ….. are still lacking/ little attention has been paid/given to.

Response 67: Thank you for your suggestion. We have change “is scarce” with several other expressions. (Lines 442, line 432 etc.)

Point 68:  L430 I think, by the BSY, is wrong.

Response 68: Thank you for your suggestion. We have changed to “was significantly increased in the experimental group” (Line 446)

Conclusion 

 Point 69:  It must be improved and summarized. Avoid redundant sentences and the mention of results.

Response 69: Thank you for your suggestion. We have updated the conclusion section according to your observations. (Lines 446-479)

 

Author Response File: Author Response.docx

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

All opinion was emitted with all respect to the efforts of the authors for the preparation of the experiment and its report

General

The purpose of this study was to evaluate The effects of brewer’s spent yeast (BSY) inclusion in dairy sheep’s diets on ruminal metabolism and milk quality parameters. For the above, a milk-performance trial was performed with 30 dairy Blackhead sheep. Even though this reviewer acknowledges the hard work devoted by the authors to the present experience, I also consider that the manuscript,as it is presented, has flaws that limit it consideration for publication in Agriculture Journal. The manuscript is written in a careless way and important information regarding Materials and Method is missing. This study was designed based on replacement techniques. Thus, is essential that physicochemical composition of both SBY tested and the replaced SBM are presented and discussed. The main limitation is that milk yields are not presented. This can affect milk composition parameters. In such a way that if data is not presented and discussed, the inferences become weak.

Specific

Title: More proper “ruminal fermentation” than “ruminal metabolism” please correct it

Abstract: Some information is missing. Needs to be improve

L18: First define before use the acronyms. Represent a byproduct?  Or is a byproduct of beer industry?

L19-21: Need be more descriptive: 30 lactating dairy Blackhead sheep (weight? days on lactation?  Distribution was individually or in groups, was completely randomized? Or block randomized? How the experiment lasted? Sampling form rumen and milk sampling. How? When? Write as (hint): was assed using 30 lactating dairy Blackhead sheep (45± xx kg, 14-d in lactation) distributed under completely randomized design in two groups: control group (with soybean meal and sunflower meal as protein sources) and the BSY group, where soybean meal was totally replaced by BSY (10% inclusion in diet on a dry matter basis). The experiment lasted XX days in which rumen and milk samples were individually taken at xx days of experiment.

L22: “propionate acid” is incorrect! Is propionic acid. Please rewrite as (hint): Inclusion of BSY affected ruminal fermentation, decreasing 25% ruminal propionate increasing 37% the acetate/propionate ratio.

L30: changes in ruminal fermentation parameters and milk..

Introduction:

Is written in a somewhat messy way. Please, rewrite in such a manner that it be more in line with your focus

L40-42:  Statement is confusing. Rewrite

L51-53: Please rewrite as (hint): Due that BSY is rich in α- and β- acids have a potential inhibition to hyper-ammonia bacteria and gram positive bacteria, therefore, BYS inclusion in ruminant’s diets can reduce the ruminal concentration of NH3-N and methane production.

L58: on rumen fermentation parameters

L63: describing the main ruminal fermentation parameters and several..

Material and methods

Poor description! Need improvements

This study was designed based on replacement techniques. Thus, is essential that physicochemical composition of both SBY tested and the replaced SBM are presented and discussed. The main limitation is the DM intake was not proper present, and milk yield are not presented. Those data can affect ruminal fermentation and milk composition parameters! In such a way that if data are not presented and discussed the inferences are weak and not very valid. Sampling method needs to be properly described. 

L70-75: Distribution was individually or in groups, was completely randomized? Or block randomized? How the experiment lasted? Sampling form rumen and milk sampling. How? Diets were offered? Why milk yield was not measured and data was presented?

L73: 10% approximately or adjusted??

L74: Indicate here that, because SBM has greater CP content than SBY, proportion of corn was replace with wheat grain in order to maintaining CP equivalent between diets.

L74: Indicate what was the criterion to determine the restricted quantity of feed offered.

Table 1.  How the MFL, PDIE, PDIN were calculated? Include the reference.

L89: Described as: rumen fermentation parameters

L91: First ruminal sample was discarding? Or not?

L92: ruminal pH was determined immediately. Ruminal fluid was filtered?  No acid was added to controlling fermentation? Describe!!!

L98 How the milk samples were taken?? The same day? When was that? 3 days after the start of the experiment? 50 days after the start of the experiment? Need to describe more specifically!

L102: What type of minerals? Be specific!

L127: The statistical analyses of data was… What type of experimental design was used? Completely random design? Complete randomized block design?  All data present normality distribution? Data were tested for normality using the Shapiro-Wilk test? Please, specify.

Results

Again, milk production data is missing

L138-146: Composition data of BSY and SBM must be presented in a Table. Mineral composition, Polyphenols, vitamin E, etc., were determined in BSY? I think not (not described in M&M section). Then should not be presented as a result. Correct this.

L147: on rumen fermentation

L149:  and after inclusion of BSY? Then, you had one group that evaluated before and after that feed BSY? Or have two groups?  Please describe it properly!

L151: Please, present data of Table 2 and Table 3 in a single Table.

L154:  pH level? This is not proper description to pH. Rewrite as: Ruminal pH value registered at 4-h post-feeding was not affected by treatments, averaging 6.20. Remove sheep’s diet here and through document, is leftover.

L158-160: Please, present data of Table 2 and Table 3 in a single Table.

L209: According to the data presented in Table 8, authors used a factorial analysis. Why this is not described in statistical analyses in M&M section? Please, include the analyses.

L271: include mineral determination for SBY in M&M section

L272: What do you attribute its higher iron concentration? Explain?

L283-289: Be careful with this statement!! Your results were exactly the opposite! (see Tables 2 and 3). Inclusion of BSY tended to increase (23%, P=0.06) NH3-N, and significantly decreased ruminal propionate (25%, P=0.019). Therefore, the statement exposed not sustain your results. It is necessary to deepen this in your discussion. Not all effects on ruminal fermentation parameters can be explained by nutraceutical properties of BYS. I allow myself to give some clues about the observed changes in ruminal fermentation. Regarding NH3-N, it is necessary to consider the changes made in diet formulation. Even when the total grain in ration is the same (73%) in both treatments, wheat replaced corn (34% of total corn was replaced by wheat in BSY diet). Protein in wheat is mostly degraded in rumen than corn protein (64 vs 43%). This is the main cause of increased on ruminal NH3-N, because BSY has lower ruminal CP degradability than SBM (~35% vs ~58%), but due its low inclusion in diet (10%), was not possible reflex it property on ruminal fermentation. Regarding acetate, BSY has both higher NDF than SBM, in addition its NDF is very ruminal soluble, therefore, increased soluble NFD could promote greater quantities of acetate.

L327: Remove sheep’s diet here and through document, is leftover.

L328: If Milk casein was not affected. How do you explain the Ca increases in milk? Please, explain this.

L336-337: Redundant.  Remove sheep’s diet here and through document, is leftover.

Conclusions

L389: can replace soybean meal up to 10% in diet. Production performance?? Where is the milk yield data??? Again, it is essential that include milk yields data in the document.

L391: ruminal fermentation parameters. What about of ruminal NH3-N?

L398: Your experiment has only two groups. Remove “compared to control group” is leftover

L405. Can affect ruminal fermentation.

Reviewer 2 Report

The manuscript submitted for review treats a popular topic of using food industry by-products (such as BSY) as an animal feed source. However, it needs to be better written, and the experiment methods need extensive change. After completing the necessary changes, please re-submit the manuscript.

Please find attached the review report.

Comments for author File: Comments.pdf

The English language needs to be edited.

Reviewer 3 Report

I stopped reviewing the work because the authors did not specify the rearing and feeding conditions of the animals (sigola and group), nor the number of samples taken, nor the level of ingestion of the animals. Without this information it is impossible to review the work and all the affirmations in the d iscussion are meaningless.

Comments for author File: Comments.pdf

Engish must be improved