Exploring the Possibility of Enriching Fermented Milks with Young Barley Leaves Powder Preparation
Round 1
Reviewer 1 Report
Although the information is well presented and written, this manuscript should be improved in several aspects to be considered for publication in this prestigious Journal.
Line 159: I don’t think that “fluctuation” be the correct word.
Lines 162 – 167: The characteristics described in this paragraph must be valued, described and discussed in depth, as you did it with the Vmax and pH obtained results.
Scientific names should be italized
Section 3.4 – Discussion should be improved (eg. Lines 330 – 332: why this happen; lines 332 – 336: why this important for this study?, and so on).
Lines: 413 – 417: Why not? Explain.
Author Response
Author's Reply to the Review Report (Reviewer 1)
Reviewer:
Although the information is well presented and written, this manuscript should be improved in several aspects to be considered for publication in this prestigious Journal.
Authors: We wanted to take a moment to express our deepest gratitude for your invaluable effort in reviewing and providing corrections for our manuscript. Your expertise and attention to detail have truly enhanced the quality of the text, and we are incredibly grateful for your time and dedication. Your comments have been carefully analyzed and incorporated into our text. The text has been adapted to all your comments.
Reviewer:
Line 159: I don’t think that “fluctuation” be the correct word.
Authors: We have changed the given sentence as follows. We hope this change is correct:
“The acidification curves are graphical representations of the pH variations that occur during the process of lactic acid fermentation as shown in Figure 1”.
Reviewer:
Lines 162 – 167: The characteristics described in this paragraph must be valued, described and discussed in depth, as you did it with the Vmax and pH obtained results.
Authors: We have supplemented the text with the following description of the data presented in Figure 1:
“The initial pH of the milk averaged 6.67 ±0.2, regardless of the milk samples. During the 14-hour fermentation with mesophilic starter cultures of lactic acid bacteria, the acidity of the milk increased until the final pH measurement averaged 4.50 ±0.3, regardless of whether the samples contained 0.5% YBLP supplementation or not”.
Reviewer:
Scientific names should be italized
Authors: Thank you for this comment. The indicated corrections were made throughout the manuscript.
Reviewer:
Section 3.4 – Discussion should be improved (eg. Lines 330 – 332: why this happen; lines 332 – 336: why this important for this study?, and so on).
Authors: We have improved the discussion of these results as follows:
“Therefore, the discussion of the results was based on other plant-based additives used in fermented milk samples. A study conducted by Mocanu et al. [27] showed that the addition of S. platensis did not lead to significant rheological changes in the fermented product. However, the authors do not explain the reasons for the observations found. In another study [39], samples containing more than 1% roasted barley powder recorded higher values of plastic viscosity, yield stress, consistency coefficient, and apparent viscosity compared with a control sample of milk fermented by a starter culture containing Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus. The aim of the cited research was to investigate the effect of yogurt drink supplementation by different amounts (0, 0.5, 1, 1.5, 2, 2.5, and 3%) of roasted barley powder on the physico-chemical properties, rheological parameters, color attributes, sensory evaluation, and production cost of the formulated yogurt drink. It is worth noting that only dosages greater than 1% of the baked barley powder (i.e., at least double the dosages used in these studies) resulted in higher parameters of the rheological properties. Similarly, Kaur & Riar [40] have found that β-glucan extract (isolated from barley) was particularly effective in set-type products like yogurt, because β-glucan’s ability to entrap water in a three-dimensional product network, helping to maintain the structure of a firm consistency of the yogurt. Researchers cited have observed an increase in the hardness of yogurt samples with the addition of β-glucan and a decrease in the adhesiveness values of yoghurt samples with the addition of β-glucan. Even the addition of 0.5% of the β-glucan preparation led to significant changes in the tested parameters of the yoghurt. However, it should be noted that in this study the YBLP preparation was characterized by a significantly different chemical composition than the β-glucan preparation used in the cited reference. Han et al. [14] reported that young barley leaves contain polysaccharides but in a much lower amount than a purified β-glucan preparation. Studies suggest that barley β-glucan may be involved in hydrophobic interactions between protein molecules, thereby stabilizing the yogurt gel [41,42]. Unfortunately, this effect was not observed in the present study. Although Qu et al. [42] demonstrated that the excessive addition of oat β-glucan (> 0.3%) destroys the three-dimensional network structure of yoghurt. The cited authors hypothesized that the addition of oat β-glucan disrupted the interaction between casein micelles and therefore three-dimensional aggregates cannot form during fermentation. Perhaps some of the results of these studies should be explained by such phenomena”.
Reviewer:
Lines: 413 – 417: Why not? Explain.
Authors: You are right, every result and every observation must be justified. The passage indicated has been changed as follows:
“Our results show that the addition of young barley leaves powder preparation at the level of 0.5% does not negatively affect the activity of mesophilic LAB either in model systems (antibacterial activity determined by the agar diffusion method) or under the fermentation conditions of a real dairy product. The positive result can be attributed to several reasons. Some substances contained in young barley leaves powder may promote the growth of beneficial bacteria or may not affect their metabolism, resulting in no adverse effects. In addition, the powder of young barley leaves could contain nutrients and compounds beneficial to the mesophilic LAB. The added nutrients can potentially improve their activity and fermentation processes without hampering their growth. Another explanation is that the young barley leaves powder may not contain substances that inhibit the growth or activity of mesophilic LAB. In addition, the use of 0.5% young barley leaves powder preparation could be in a suitable range that does not overwhelm or disturb the LAB. It is important to note that these are hypotheses only and further research and analysis would be required to determine the exact mechanisms behind the lack of adverse effects”.
Reviewer 2 Report
The manuscript entitled ‘Exploring the Possibility of Enriching Fermented Milks with Young Barley Leaves Powder Preparation” is dealing with the potential effects derived after the addition of YBLP on various LAB cultures. The manuscript is well written and easy to follow the rational of the study, however it lucks solid conclusions. All the analyses presented in this study do not support any specific results regarding a positive effect of this extract in fermented milks. Results on cell viability in some cases show some interest, but I believe that there are not enough to support a full research article. Since the results regarding the effect on LAB are not enough, some organoleptic properties or other types of analyses could be added to enrich the manuscript. For example, the antioxidant activity of the supplement, or the possible monosaccharide content (or oligosaccharide that could actually act some prebiotic).
Some more specific comments:
Line 72: “commercial food supplement”
Line 88: Why did the authors select 27 degrees for the fermentation? Most LAB operate at much higher temperatures (37 degrees mainly)
Line 109: “incubation”
Figure 1: Please remove the legend from inside the figure and describe it in the figure’s description
Why pH=4.5 was a critical value for the study?
Did the protein content of the supplement influenced growth? Some free amino nitrogen analyses could be interesting to be carried out.
Lines 194-195: How do the authors reach this conclusion? It is rather obvious from Table 2 that the addition of YBLP had no effect on acidification regardless the strain employed.
I am a bit skeptical regarding the MANOVA, especially in Table 3. The authors tested the values of the same culture during storage, and also between control and with YBLP. For CHOOZIT MTM 2, there is no statistical difference for 21 days and then, there is a significant difference of pH values 4.38 and 4.33? It is hard for me to believe that these values are truly different. It is easier for me to believe that this statistical difference refers to day 0 when compared to day 28.
Line 272: S. platensis (with italics)
Just some minor spelling errors
Author Response
Author's Reply to the Review Report (Reviewer 2)
Reviewer: The manuscript entitled ‘Exploring the Possibility of Enriching Fermented Milks with Young Barley Leaves Powder Preparation” is dealing with the potential effects derived after the addition of YBLP on various LAB cultures. The manuscript is well written and easy to follow the rational of the study, however it lucks solid conclusions. All the analyses presented in this study do not support any specific results regarding a positive effect of this extract in fermented milks. Results on cell viability in some cases show some interest, but I believe that there are not enough to support a full research article. Since the results regarding the effect on LAB are not enough, some organoleptic properties or other types of analyses could be added to enrich the manuscript. For example, the antioxidant activity of the supplement, or the possible monosaccharide content (or oligosaccharide that could actually act some prebiotic).
Author's Reply: We would like to express our heartfelt appreciation for your meticulous review of our manuscript. Your insightful comments and suggestions have greatly contributed to improving the quality and relevance of our study. We have taken your suggestion to heart and have carefully incorporated these relevant concepts into our revised manuscript. Moreover, we have ensured that the references you suggested have been included in our reference list, thereby strengthening the scientific foundation of our work.
We are delighted that you have carefully analyzed our work and appreciate your suggestions for its further expansion. We wholeheartedly agree with your observation regarding the need to enhance the results concerning the effect on LAB. In our subsequent research, we plan to focus on incorporating an analysis of organoleptic properties and other types of investigations, such as the antioxidant activity of the supplement or the analysis of monosaccharide or oligosaccharide content, which may potentially act as prebiotics. Undoubtedly, these additional analyses will enrich our article by providing more comprehensive findings and a better understanding of the potential benefits of the supplement. We are grateful for pointing out these crucial areas for further exploration.
Reviewer: Some more specific comments:
Line 72: “commercial food supplement”
Author's Reply: Thank you for this comment. This is our mistake, which has already been corrected.
Reviewer: Line 88: Why did the authors select 27 degrees for the fermentation? Most LAB operate at much higher temperatures (37 degrees mainly)
Author's Reply: For most of the starter cultures used, the recommended incubation temperature is in the range of 23-30 °C, as they contain mesophilic LAB, only for Chr. Hansen XPL-1 a higher value of the parameter is recommended (30-35 °C). The average temperature for most of the cultures used was assumed to be 27 °C.
Reviewer: Line 109: “incubation”
Author's Reply: Thank you for this comment. This is our mistake, which has already been corrected.
Reviewer: Figure 1: Please remove the legend from inside the figure and describe it in the figure’s description
Author's Reply: Thank you for this comment. Figures 1 – 3 have been modified as recommended and the figure legend has been moved to the figure description.
Reviewer: Why pH=4.5 was a critical value for the study?
Author's Reply: The pH of 4.5 is close to the isoelectric point of casein, an important milk protein. At this pH, the casein molecules tend to interact and coagulate, contributing to yogurt's characteristic thick and creamy texture. For this reason, in many studies, a pH value of 4.5 is assumed to be crucial for the quality of yoghurt.
Reviewer: Did the protein content of the supplement influenced growth? Some free amino nitrogen analyses could be interesting to be carried out.
Author's Reply: Of course, adding young barley leaves powder to milk can affect the growth of LAB due to its protein content. Lactic acid bacteria are known to thrive on nitrogenous compounds, and proteins are a good source of nitrogen for their growth. Young barley leaves powder contains 21.4 grams of protein per 100 grams. This protein content is likely to provide a significant amount of free amino nitrogen, which is the nitrogen present in the form of amino acids and small peptides. To understand the effect of young barley leaves powder on LAB growth, it would indeed be interesting and relevant to perform free amino nitrogen analyses. This analysis would provide valuable information on the availability of nitrogenous compounds in the mixture of milk and young barley leaves powder. It can help determine if there are sufficient nutrients to support the growth of LAB and how the protein content in young barley leaves powder contributes to this process. Keep in mind that other factors, such as the overall composition of the milk and the specific strains of LAB present, can also affect its growth. Nonetheless, the protein content of young barley leaf powder when added to milk is likely to play an important role in supporting the growth of LAB. Thank you for bringing this aspect to our attention. We have added a few sentences to the text (at the end of the paragraph 3.3.):
“YBLP supplement contains 21.4 grams of protein per 100 grams. This protein content likely provides a significant amount of free amino nitrogen. To understand the effect of YBLP on LAB growth, it would be interesting and relevant to perform free amino nitrogen analyzes of YBLP. It can help determine if there are sufficient nutrients to support the growth of LAB and how the protein content in YBLP contributes to this process”.
Reviewer: Lines 194-195: How do the authors reach this conclusion? It is rather obvious from Table 2 that the addition of YBLP had no effect on acidification regardless the strain employed.
Author's Reply: You are right. We have improved what is displayed as follows:
“However, at this point, we demonstrated a lack of negative effect of YBLP supplementation on acidification, albeit depending on the starter culture used.”.
Reviewer: I am a bit skeptical regarding the MANOVA, especially in Table 3. The authors tested the values of the same culture during storage, and also between control and with YBLP. For CHOOZIT MTM 2, there is no statistical difference for 21 days and then, there is a significant difference of pH values 4.38 and 4.33? It is hard for me to believe that these values are truly different. It is easier for me to believe that this statistical difference refers to day 0 when compared to day 28.
Author's Reply: We re-analyzed statistically and found our error in the previous data analysis of Table 3 (the options in the statistics program were set incorrectly). Thank you for your attention. The contents of the table and its interpretation in the text of the manuscript have already been improved. Therefore, we checked the statistical analysis of the other results and found no calculation errors in this case.
Reviewer: Line 272: S. platensis (with italics)
Author's Reply: Thank you for this comment. The indicated corrections were made throughout the manuscript.
Reviewer: Comments on the Quality of English Language
Just some minor spelling errors
Author's Reply: We would like to express our gratitude for pointing out the need for minor editing of the English language. We have dedicated considerable effort to improving the language and clarity of our manuscript, addressing any grammatical or stylistic issues that may have affected its readability.
Reviewer 3 Report
I have reviewed the manuscript entitled "Exploring the Possibility of Enriching Fermented Milks with Young Barley Leaves Powder Preparation", submitted to Fermentation Special Issue "New Insights into the Application of Lactic Acid Bacteria Strain in the Fermentation 2.0". The manuscript aimed to investigate the possibility of enriching fermented dairy products (mesophilic-fermented milk) with young barley leaves powder preparation, including different starter cultures of lactic acid bacteria.
I believe that the manuscript is very interesting but needs some minor revisions.
Introduction
The introduction places the study in a broad context and defines the purpose of the work and its significance.
Materials and Methods
L77: Is the chemical composition correct? The manufacturer specifies that there are more sugars than carbohydrates. Fiber content should be listed separately. Please verify.
"carbohydrates 25.1 g (including sugars 35 g and fiber 33.7 g)"
L80: On what criteria was this level (0.5%) of YBLP additive chosen?
L79-83: Please explain how the mixtures were prepared for fermentation. After adding the YBLP to the milk, was the mixture pasteurized? If not, to what temperature was the milk heated with the addition of YBLP? In what amounts and in what form were the starter cultures added? Was it a previously prepared starter or inoculum, under what conditions? Was the level of bacteria in the starters/inoculum known?
122-132: Did the barley descend in the forming milk gel? If so, were the samples mixed before texture measurement?
Results and Discussion
L272, 383: "S. platensis" should be written in italics.
Conclusions
L419-420: The results of the study are very promising. Please, in the future, consider studying the effect of YBLP addition on the organoleptic characteristics of fermented milk.
L444-446: This section should be revised since a specific intake of YBLP may provide this effect. In the study, only 0.5% YBLP was used. Is this amount of YBLP able to provide this effect?
References
Most of the cited literature sources are more recent than the last five years, but their choice is appropriate.
L479: The citation is incorrect (authors' names).
Author Response
Author's Reply to the Review Report (Reviewer 3)
Reviewer: I have reviewed the manuscript entitled "Exploring the Possibility of Enriching Fermented Milks with Young Barley Leaves Powder Preparation", submitted to Fermentation Special Issue "New Insights into the Application of Lactic Acid Bacteria Strain in the Fermentation 2.0". The manuscript aimed to investigate the possibility of enriching fermented dairy products (mesophilic-fermented milk) with young barley leaves powder preparation, including different starter cultures of lactic acid bacteria.
I believe that the manuscript is very interesting but needs some minor revisions.
Authors: We would like to express our sincere appreciation for taking the time to review our paper. We are grateful for pointing out the mistakes we made. Your positive feedback and constructive comments have been immensely valuable to us.
Reviewer: Introduction
The introduction places the study in a broad context and defines the purpose of the work and its significance.
Authors: This is true. The introduction of a scientific study plays a crucial role in providing background information and context for the research. Remember that the introduction may vary depending on the study, its goals, and the guidelines of the target journal.
Reviewer: Materials and Methods
L77: Is the chemical composition correct? The manufacturer specifies that there are more sugars than carbohydrates. Fiber content should be listed separately. Please verify.
"carbohydrates 25.1 g (including sugars 35 g and fiber 33.7 g)"
Authors: We are very sorry; this is our error in transferring the information from the packaging. The correct wording is as follows:
“energy value 1151 kJ/275 kcal, fat 2.5 g (including saturated fatty acids 2.2 g), carbohydrates 25.1 g (including sugars 1.4 g and fiber 23.7 g), protein 21.4 g, and salt 0.17 g”.
Reviewer: L80: On what criteria was this level (0.5%) of YBLP additive chosen?
Authors: The 0.5% YBLP grade is selected based on references [15]. The cited study on the addition of S. platensis in an amount of 0.5 percent showed a positive effect on the survival of bacteria in yoghurt during storage. So far there are no scientific reports describing the influence of young barley preparations on the texture of fermented milk.
Reviewer: L79-83: Please explain how the mixtures were prepared for fermentation. After adding the YBLP to the milk, was the mixture pasteurized? If not, to what temperature was the milk heated with the addition of YBLP? In what amounts and in what form were the starter cultures added? Was it a previously prepared starter or inoculum, under what conditions? Was the level of bacteria in the starters/inoculum known?
Authors: Thank you for this attention. To make the methodological description easier to read, we have added missing information to the information in the manuscript:
“Fermented milk samples were prepared using milk (1.5% fat, SM Łowicz, Łowicz, Poland) previously UHT-treated, then cooled to 27 °C and supplemented with 0.5% by weight of young barley leaves preparation (YBLP) and eight specific mesophilic LAB starter cultures (at the dose of 0.1%), commonly employed in the industrial production of sour milk-type fermented milks. Before addition to the milk, starter cultures were prepared by dissolving them in sterile drinking water. Fermented milk samples without the addition of YBLP were prepared in the same way (control tests).”.
Reviewer: 122-132: Did the barley descend in the forming milk gel? If so, were the samples mixed before texture measurement?
Authors: No, YBLP at 0.5% concentration did not precipitate in milk or yoghurt samples. The YBLP formulation was mealy. Therefore, the samples were not mixed prior to texture measurement. Regardless, the samples were mixed with sterile metal spoons before the other analyzes were performed.
Reviewer: Results and Discussion
L272, 383: "S. platensis" should be written in italics.
Authors: Thank you for this comment. The indicated corrections were made throughout the manuscript.
Reviewer: Conclusions
L419-420: The results of the study are very promising. Please, in the future, consider studying the effect of YBLP addition on the organoleptic characteristics of fermented milk.
Authors: The suggestion to study the effect of YBLP addition on the organoleptic characteristics of fermented milk is valuable. By conducting further research on the organoleptic characteristics, scientists and food manufacturers can ensure that adding YBLP to fermented milk does not negatively impact its taste or texture. If the sensory properties are maintained or even improved, it could enhance the appeal and consumption of such products among consumers. The conclusion paragraph was modified as follows:
“The results of this study unequivocally highlight the need for further exploration of the use of the additive YBLP in fermented milk. In the future, consider studying the effect of YBLP addition on the organoleptic properties of fermented milk. By further studying the organoleptic properties, scientists and food manufacturers can ensure that adding YBLP to fermented milk does not adversely affect its taste or texture. Understanding how YBLP affects the flavor and overall sensory experience of fermented milk is critical to consumer acceptance and marketability. Research and development in this area could lead to the formulation of YBLP-enriched fermented milk products with potential health benefits and improved consumer acceptance.”
Reviewer: L444-446: This section should be revised since a specific intake of YBLP may provide this effect. In the study, only 0.5% YBLP was used. Is this amount of YBLP able to provide this effect?
Authors: Thank you for this suggestion. In fact, we now see that the conclusions were wrongly formulated. The last paragraph of the conclusion has been changed as follows:
“It is important to note that in addition to the beneficial effects of LAB, the YBLP formula itself contributes essential nutrients for the human body. It is rich in vitamins, minerals, and essential amino acids, supporting the immune system and showing proven anti-cancer effects [65]. The effectiveness of any substance, including YBLP, depends on various factors, such as the intended effect, the target group, the method of administration, and the desired result. It is worth investigating in the future whether a portion of 0.5% YBLP can have a sufficient health-promoting effect. In this study, it was only shown that 0.5% YBLP failed to inhibit LAB activity. As for the effectiveness of 0.5% YBLP, it is difficult to determine its effect without the specific context of the study and its objectives.”
Reviewer: References
Most of the cited literature sources are more recent than the last five years, but their choice is appropriate.
Authors: We want to assure you that we have taken your suggestions to heart and have diligently addressed the issues you raised. We have carefully corrected the mistakes and made the necessary revisions to ensure the accuracy and professionalism of our work. Thank you once again for your contribution to our manuscript.
Reviewer: L479: The citation is incorrect (authors' names).
Authors: Thank you for this comment. This is our mistake and we have already corrected it:
“Osawa, T.; Katsuzaki, H.; Hagiwara, Y.; Hagiwara, H.; Shibamoto, T. A Novel Antioxidant Isolated from Young Green Barley Leaves. J. Agric. Food Chem. 1992, 40, 1135–1138, doi:10.1021/jf00019a009”.
Round 2
Reviewer 1 Report
All my suggestion were well addressed by the authors. Thus, this manuscript can be considered for publication in this Journal.
Reviewer 2 Report
The authors have carried out all the corrections suggested by the reviewers and adequately responded to all comments.
