Astringency Modification of Mandilaria Wines: Vineyard and Winery Strategies

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Abstract

Lines 13, 17-18: The sentences have the same meaning, please reformulate.

Line 14: Please specify the scientific name of the grapevine species, Vitis vinifera, Mandilaria (Vitis vinifera) grape variety.

1. Introduction

Line 35: Please italicize Vitis vinifera.

2. Materials and Methods

Lines 110, 111, 112, 208: Please italicize O for delphinidin-3-O-glucoside, delphinidin-3-O-glucoside,  petunidin-3-O-glucoside,  peonidin-3-O-glucoside, malvidin-3-O-glucoside, and (−)-epicatechin-3-O-gallate.

Line 121: Please add a space between 18.8 ± 1 and °Brix.

Line 146: Did the authors obtain a specific particle size for the powder of the material?

Lines 149, 150: Please replace hours with h.

Line 151: Please add a space between 30 and °C.

Line 173: Please replace minutes with min.

Line 218: Please replace S. cerevisiae with Saccharomyces cerevisiae.

Table 1: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).

Lines 268, 269: Please indicate the units of measurement for (19.0 and 19.4, respectively) and (3.5 and 4.0, respectively).

Table 2: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for CTRL-V, CANE, E-RIF, T-RIF. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).

Table 3: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).

Figure 1: (a) Please replace Brix with °Brix; (b) What Brry weight means? Please indicate the units of measurement in the brackets.

Tables 4, 5: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).

Sections 3.2.1 and 3.2.2: The obtained results should be characterized in contrast with other papers.

Figure 2: Please include the meaning of the superscript letters below the figure. Also, present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30.

Figure 3: Please italicize O for delphinidin-3-O-glucoside, cyanidin-3-O-glucoside, peonidin-3-O-glucoside, petunidin-3-O-glucoside, malvidin-3-O-glucoside, malvidin-3-O-(6-acetyl) glucoside, malvidin 3-O-(6-p-coumaroyl) glucoside. Please italicize p for malvidin 3-O-(6-p-coumaroyl) glucoside. Also, present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, and D-AIR. Please include the meaning of the superscript letters below the figure. What % means? Please indicate vertical axis title and put % in the brackets. Please delete (a) Vineyard treatments and (b) Winery treatments from the figure, because authors had write the meaning in the figure title.

Figure 4: Please include the meaning of the superscript letters below the figure. What % means? Please indicate vertical axis title and put % in the brackets. Please delete (a) Vineyard treatments and (b) Winery treatments from the figure, because authors had write the meaning in the figure title. Please indicate the legend for Dlp, Cyn, Peo, Pet, Mlv, Mlv acet, Mlv coum, CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30.

Table 6: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for MCP, BSA, CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).

Figure 6: Present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Please include the meaning of the superscript letters below the figure.

Conclusions: Please present some relevant numerical data.

References: The references cited should be more recent, with only those published within the last 5 years being considered, especially for the Results and Discussion section.

Author Response

Response to Reviewers
Reviewer 1: 
Abstract:

Lines 13, 17-18: The sentences have the same meaning, please reformulate.
Authors’ response: We have reformulated this part accordingly.

Line 14: Please specify the scientific name of the grapevine species, Vitis vinifera, Mandilaria (Vitis vinifera) grape variety.
Authors’ response: We have specified this part accordingly.

Introduction:

Line 35: Please italicize Vitis vinifera.
Authors’ response: We have changed this part accordingly.

Materials and Methods:

Lines 110, 111, 112, 208: Please italicize O for delphinidin-3-O-glucoside, delphinidin-3-O-glucoside,  petunidin-3-O-glucoside,  peonidin-3-O-glucoside, malvidin-3-O-glucoside, and (−)-epicatechin-3-O-gallate.
Authors’ response: We have changed these parts accordingly.

Line 121: Please add a space between 18.8 ± 1 and °Brix.
Authors’ response: We have changed this part accordingly.

Line 146: Did the authors obtain a specific particle size for the powder of the material?
Authors’ response: No, a specific particle size was not obtained for the powder. The material was ground manually to a fine consistency, but no standardized particle size distribution was measured or controlled during the process.

Lines 149, 150: Please replace hours with h.
Authors’ response: We have changed this part accordingly.

Line 151: Please add a space between 30 and °C.
Authors’ response: We have made the necessary adjustments to this section.

Line 173: Please replace minutes with min.
Authors’ response: We have made the necessary adjustments to this section.

Line 218: Please replace S. cerevisiae with Saccharomyces cerevisiae.
Authors’ response: We have made the necessary adjustments to this section.

Results:

Table 1: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).
Authors’ response: This part has been specified as requested.

Lines 268, 269: Please indicate the units of measurement for (19.0 and 19.4, respectively) and (3.5 and 4.0, respectively).
Authors’ response: We have changed this part accordingly.

Table 2: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for CTRL-V, CANE, E-RIF, T-RIF. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).
Authors’ response: Thank you for the comment. We have changed this part accordingly. We would like to clarify that the legend for CTRL-V, CANE, E-RIF, and T-RIF was initially described both in Table 1 and in the main text (Section 2.2 – Experimental Design). However, as requested, we have now also added the explanation directly below Table 2 for clarity.

Table 3: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).
Authors’ response: Thank you for the comment. We have changed this part according to your feedback.

Figure 1: (a) Please replace Brix with °Brix; (b) What Brry weight means? Please indicate the units of measurement in the brackets.
Authors’ response: Thank you for your valuable comment. We have corrected the term Brix to °Brix, as suggested. We also apologize for the typo in “Brry weight” — the correct term is “Berry weight,” referring to the weight of the grape berry, which is now clearly indicated in grams (g) in the figure. We appreciate your attention to detail, which helped us improve the clarity and accuracy of the manuscript.

Tables 4, 5: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).
Authors’ response: Thank you for your observation. We would like to clarify that the acronyms were not redefined in the table legend, as they are already thoroughly explained in Section 2.2 (Experimental Design). However, we understand the value of providing clear and immediate context, so we have now included the corresponding definitions below tables 4 and 5 as well, in line with your suggestion.

Sections 3.2.1 and 3.2.2: The obtained results should be characterized in contrast with other papers.
Authors’ response: Thank you for the suggestion. We have revised the relevant section to include a clearer comparison of our results with those reported in other studies, as recommended.

Figure 2: Please include the meaning of the superscript letters below the figure. Also, present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30.
Authors’ response: Thank you for the comment. We have changed this part according to your feedback.

Figure 3: Please italicize O for delphinidin-3-O-glucoside, cyanidin-3-O-glucoside, peonidin-3-O-glucoside, petunidin-3-O-glucoside, malvidin-3-O-glucoside, malvidin-3-O-(6-acetyl) glucoside, malvidin 3-O-(6-p-coumaroyl) glucoside. Please italicize p for malvidin 3-O-(6-p-coumaroyl) glucoside. Also, present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, and D-AIR. Please include the meaning of the superscript letters below the figure. What % means? Please indicate vertical axis title and put % in the brackets. Please delete (a) Vineyard treatments and (b) Winery treatments from the figure, because authors had write the meaning in the figure title.
Authors’ response: We’ve updated this section to reflect the suggested changes.

Figure 4: Please include the meaning of the superscript letters below the figure. What % means? Please indicate vertical axis title and put % in the brackets. Please delete (a) Vineyard treatments and (b) Winery treatments from the figure, because authors had write the meaning in the figure title. Please indicate the legend for Dlp, Cyn, Peo, Pet, Mlv, Mlv acet, Mlv coum, CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30.
Authors’ response: We have made the necessary adjustments to this section.

Table 6: Please include the meaning of the superscript letters (letters that are presented for each sample in column of the table) below the table. Also, indicate the p-value below the table and present the legend for MCP, BSA, CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Moreover, please specify what the number following ± refers to (e.g., standard deviation, standard error).
Authors’ response: We’ve updated this section to reflect the suggested changes.

Figure 6: Present the legend for CTRL-V, CANE, E-RIF, T-RIF, CTRL-W, D-VIN, D-SUN, D-AIR, SE20, and SE30. Please include the meaning of the superscript letters below the figure.
Authors’ response: We’ve updated this section to reflect the suggested changes.

Conclusions: Please present some relevant numerical data.
Authors’ response: Thank you for your suggestion. In response, I have added relevant numerical data in the Conclusions section to support and highlight the key findings more clearly.

References: The references cited should be more recent, with only those published within the last 5 years being considered, especially for the Results and Discussion section.
Authors’ response: We’ve updated the references with the most recent published articles to reflect the suggested changes.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript analyses several variables that may influence the astringency of a typical wine from Greece. The scope of the work is interesting and may be the first in a series of studies in the field. However, it has some important deficiencies that need to be addressed before it can be accepted for publication.

Below are the comments for the authors.

Abstract.

Lines 13 to 20. This part of the abstract must be rewritten because it is repetitive in stating the purpose.

Keywords.

Line 31. Do not include abbreviations in Keywords, edit.

Materials and Methods.

Line 120. In food production, especially those highly influenced by environmental and meteorological aspects, one should work on several successive years. This experimental design would make it possible to limit the variability of the data due to a single year. The experimental design implemented by the authors is very limiting.

Line 144. Authors must report on the analysis procedures applied. In particular section 2.4.3. Analysis of anthocyanins does not detail the HPLC procedure.

Line 212. This paragraph, for the purposes of discursive and procedural linearity, should be moved after section 2.2. Experimental design.

Line 228. The authors make no mention in this section of an Ethics Committee opinion and/or informed consent provided to participants. It is suggested that an indication should also be included in this paragraph, as well as at the end of the manuscript.

Line 247. Tables. no footnotes indicating abbreviations, statistical evaluation of the ANOVA. Sometimes the letters of the statistic are inconsistent with the data.

Line 247. Figures. no footnotes indicating abbreviations, measuring units, statistical evaluation of the ANOVA. Sometimes the letters of the statistic are inconsistent with the data.

Line 467. The results of the sensory analysis have been reported very briefly.

Conclusions

Lines 515-518. There is little sensory data reported to demonstrate concrete perceived variability.

Line 509. The conclusions that the authors draw leave several open points on the concrete benefits of the techniques applied.

The authors do not take into consideration that such practices could also affect the volatile composition of wines. It is recommended that the introduction and conclusions be re-evaluated, taking into consideration studies on the changes that practices (such as defoliation) may have on the chemical and sensory characteristics of grapes and wines.

Author Response

Response to Reviewers
Reviewer 2:
The manuscript analyses several variables that may influence the astringency of a typical wine from Greece. The scope of the work is interesting and may be the first in a series of studies in the field. However, it has some important deficiencies that need to be addressed before it can be accepted for publication.
Below are the comments for the authors.
Authors’ response: We thank the Reviewer for their time and effort to improve the quality of our work.
Abstract:
Lines 13 to 20. This part of the abstract must be rewritten because it is repetitive in stating the purpose.
Authors’ response: Thank you very much for your comment. This point had also been noted by a previous reviewer and has already been corrected in the revised version of the manuscript.
Keywords:
Line 31. Do not include abbreviations in Keywords, edit.
Authors’ response: Thank you for your observation. We have renamed it using the full term for clarity.

Materials and methods:
Line 120. In food production, especially those highly influenced by environmental and meteorological aspects, one should work on several successive years. This experimental design would make it possible to limit the variability of the data due to a single year. The experimental design implemented by the authors is very limiting.
Authors’ response: Thank you for this insightful comment. We fully acknowledge the importance of conducting multi-year experiments, particularly in fields such as food production, where environmental and meteorological factors play a significant role in influencing results. In our case, due to the extensive volume and complexity of the measurements required, it was not feasible to repeat the entire experimental design in a second year. However, we did repeat selected measurements in a second growing season, focusing on the most promising practices identified in the initial year. These follow-up results will be presented in detail in a forthcoming publication. We considered it methodologically appropriate not to include these additional data in the current manuscript, as not all treatments were repeated across both years. Including only partial two-year data might have introduced imbalance and confusion in the interpretation. We appreciate your understanding on this point.

Line 144. Authors must report on the analysis procedures applied. In particular section 2.4.3. Analysis of anthocyanins does not detail the HPLC procedure.
Authors’ response: Thank you very much for your valuable and insightful comments. Additional details have been included in section 2.4.3 regarding the analysis of anthocyanins, as well as in section 2.4.4 concerning the analysis of tannins, in order to further clarify the methodology.

Line 212. This paragraph, for the purposes of discursive and procedural linearity, should be moved after section 2.2. Experimental design.
Authors’ response: In accordance with your suggestion, the paragraph has been relocated under the Experimental Design section.

Line 228. The authors make no mention in this section of an Ethics Committee opinion and/or informed consent provided to participants. It is suggested that an indication should also be included in this paragraph, as well as at the end of the manuscript.
Authors’ response: We appreciate your insightful comment. In response, we have included the Ethics Committee approval and participant informed consent in the specified section as well as at the end of the manuscript, as recommended.

Line 247. Tables. no footnotes indicating abbreviations, statistical evaluation of the ANOVA. Sometimes the letters of the statistic are inconsistent with the data.
Authors’ response: Thank you very much for your valuable comment. Initially, footnotes were not included below the tables due to the extensive amount of data, as all abbreviations and relevant explanations are provided in the Materials and Methods section. However, following your suggestion, footnotes have now been added below each table for clarity. In addition, statistical evaluation details from the ANOVA analysis are now also included under each table, and any inconsistencies in lettering have been corrected accordingly.

Line 247. Figures. no footnotes indicating abbreviations, measuring units, statistical evaluation of the ANOVA. Sometimes the letters of the statistic are inconsistent with the data.
Authors’ response: Thank you very much for your insightful comment. At first, footnotes were not placed beneath the figures due to the large volume of information, as all abbreviations and methodological details were described in the Materials and Methods section. Nonetheless, in response to your suggestion, footnotes have now been added below each figure for improved clarity. Furthermore, the statistical evaluation from the ANOVA analysis is now also presented under each figure, and any inconsistencies in lettering have been revised accordingly.
Line 467. The results of the sensory analysis have been reported very briefly.
Authors’ response: We thank the reviewer for this valuable comment. We agree that the initial presentation of the sensory analysis results was rather brief. To address this, we have added Table 7, which summarizes the findings in a more accessible way, allowing for a clearer understanding of the conclusions drawn for each treatment, particularly with respect to the perception of astringency.
Conclusions:
Lines 515-518. There is little sensory data reported to demonstrate concrete perceived variability.
Authors’ response: We thank the reviewer for the thoughtful observation. Indeed, the sensory analysis was conducted with the primary aim of evaluating the overall quality of the produced wines. Although several sensory attributes were assessed, in the present work we chose to report only those directly related to the perception of astringency, in order to maintain focus and avoid potential confusion. We believe this approach better serves the objectives of the study.

Line 509. The conclusions that the authors draw leave several open points on the concrete benefits of the techniques applied.
Authors’ response: Thank you very much for your helpful comment. In response, the Conclusions section has been revised to more clearly outline the concrete benefits of the techniques applied. We have aimed to make the conclusions more focused and specific, directly linking them to the findings presented in the results.

The authors do not take into consideration that such practices could also affect the volatile composition of wines. It is recommended that the introduction and conclusions be re-evaluated, taking into consideration studies on the changes that practices (such as defoliation) may have on the chemical and sensory characteristics of grapes and wines.
Authors’ response: Thank you very much for your thoughtful and constructive comment. In response, I have revised the Introduction section to include references to the potential impact of vineyard practices—such as defoliation—on the volatile composition and overall chemical and sensory characteristics of grapes and wines, as recommended.  However, since the impact on volatile composition was not directly investigated in the present experimental work, this information has been incorporated only in the Introduction as part of the theoretical background. In the Conclusions section, we have chosen to limit the discussion strictly to the parameters that were experimentally assessed, in order to maintain scientific accuracy and avoid overinterpretation of the results.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

This paper is quite interesting and a great number of assays were developed. Even though, some complementary analysis (CIELab and other antioxidant method) could help to understand the results. Discussion is missed in some parts of this manuscript.

So, I encourage the authors to take in mind the comments to improve this paper.

Comments for author File: Comments.docx

Author Response

Response to Reviewers
Reviewer 3: 

Abstract: 
I would report that Mandilaria grapes are typical from Greece (Paros island) although is written in the introduction and please, write that is a red wine.
I also would write that sensory analysis with a trained panel was performed
Authors’ response: We have reformulated this part accordingly.

Keywords:
I would write “grape dehydration” instead grape
“mDp” abbreviation is not understandable, it should be described.
Tannin is increased with treatments. So, I would write Tannin (not tannin reduction).
Authors’ response: I appreciate your helpful suggestion. The keywords have been updated accordingly to better reflect the content and focus of the manuscript.                               
1.    Introduction: 
I would add that a complementary sensory analysis performed by trained assessors was also relevant to
prove the reduction in astringency in order to try to demonstrate the correlation with other parameters.
Authors’ response: Thank you for your suggestion. The statement you recommended has been incorporated into the Introduction section.

2.    Materials and methods:

It is not clear how many different wines were obtained. Why was not winemaking applied to obtain the corresponding wines? How was the tank size to obtain the wines?
Authors’ response: Thank you for your comment and interest in the experimental setup. Each wine produced corresponded to a specific treatment, including the two controls (vineyard and winery-dehydration), 20% and 30% seed removal, long pruning, partial and complete defoliation, as well as the three dehydration treatments. Winemaking was applied individually to each treatment group in order to maintain consistency and allow for accurate comparison across the different practices. We would like also to clarify that all vinifications were carried out at a constant tank volume of 100 liters for each treatment, as is typical in experimental winemaking. This ensured comparability across treatments under standardized conditions.

Water content should be measured during dehydration process of grapes.
Authors’ response: We thank the reviewer for this valuable comment. In response, we have added an additional column in Table 3 reporting the percentage of dehydration during the different treatments. We believe this addition provides a clearer understanding of the dehydration dynamics and its impact on the grapes.

In my opinion, the authors should perform more than 1 method to measure Antioxidant activity. Why only DPPH method was used?
Authors’ response: Thank you for your valuable comment. The DPPH assay was selected for this study due to its wide acceptance, simplicity, and reliability in evaluating antioxidant activity, particularly in wine-related matrices. We acknowledge that incorporating additional methods such as ABTS or FRAP could provide a more comprehensive evaluation, and we will certainly consider including multiple assays in future studies to further validate and expand upon the current findings.

The application of a CIELab colorimeter to obtain more chromatic parameters could add more information related to differences in wine´s color and hue. It is only a recommendation.
Authors’ response: Thank you for your insightful suggestion. We fully agree that the use of a CIELab colorimeter could provide a more detailed and objective assessment of wine color and hue. In the present study, color was evaluated using traditional spectrophotometric methods, but we acknowledge the added value of the CIELab system. We will certainly consider incorporating this approach in future work to enhance the chromatic characterization of wines.

Was the chromatographic column maintained at constant temperature?
Authors’ response: Thank you for your question. Yes, this point has been addressed in the manuscript. The chromatographic column was maintained at a constant temperature during all analyses. 

The questionnaire/ballot for sensory analysis was not showed or reported.    
Authors’ response: We thank the reviewer for the comment. The questionnaire attached as supplementary material provides detailed information regarding the astringency scale. A summary of additional sensory analyses was conducted to assess overall wine quality, including aroma characteristics, taste attributes, mouthfeel, aftertaste, and overall impression. However, these additional parameters were not included, as they were considered beyond the scope of the current study, which focuses specifically on astringency perception.

3.    Results:
A discussion and comparison with other studies, related to some performed measurements are almost 
not made. If there are no studies in Mandilaria you could compare with other Greek grape types or even though with other wines obtained from astringent grapes.
Authors’ response: Thank you very much for your helpful comment. While literature specifically related to Mandilaria is indeed limited, we have expanded the discussion by including comparisons with other studies focusing on similar viticultural and winemaking techniques—such as dehydration and defoliation—rather than specific grape varieties. These comparisons were incorporated for parameters such as color, total phenolics, and antioxidant activity, in order to provide a broader understanding of how such practices influence wine characteristics, regardless of the specific cultivar.

A measurement of water content should be performed, as it affects all parameters as sugar contents and weight evolution of the grapes.
Authors’ response: We thank the reviewer for the insightful comment. In response, the percentage of dehydration has been added in Table 3, providing a clearer presentation of the water loss occurring during the grape dehydration process.

Why the sample D-SUN (Table 5) is so low in anthocyanins content? 
Authors’ response: Thank you for your question. The lower anthocyanin content observed in the D-SUN sample is discussed in lines 490–492 of the manuscript. As noted, this finding is consistent with existing literature, which indicates that increased sunlight exposure and higher temperatures can lead to greater skin rupture and subsequently promote oxidative degradation of anthocyanins.
Additionally, the particularly low anthocyanin levels in D-SUN are further supported by Table 5, where the extractability of anthocyanins in this sample is shown to be very low. This reduced extractability is likely related to the extensive dehydration experienced by the berries, which affects anthocyanin stability and availability.

I would like to see a chromatogram for the anthocyanin profile and other for flavanols. 
Authors’ response: We thank the reviewer for the suggestion. However, given the extensive amount of data already presented in this work, we have chosen not to include additional chromatograms at this stage. Moreover, the analytical methods applied for anthocyanin and flavanol profiling are well-established and validated in the literature, and no new methods were developed in the present study. Therefore, we believe that attaching chromatograms would not substantially enhance the clarity or understanding of the results.

I miss a table/Figure showing sensory results.
Authors’ response: Thank you for your observation. In response, Table 7 has been added to present the sensory evaluation results of the wines, providing a clearer overview of the differences among treatments.

Misprint
Line 394: corresponding
Line 410: resulted
Authors’ response: We have reformulated these parts accordingly.

Tables and Figures

In Figure 1b, axis “y” should be typed Berry, not Brry.
Authors’ response: We have reformulated this part accordingly.

In Figure 2, hue variations are almost inappreciable. Please could you show it in other way?
Authors’ response: Thank you for your observation regarding the hue data presentation. To improve clarity and facilitate interpretation, we have now separated color intensity and hue into two distinct graphs—Figure 2a for intensity and Figure 2b for hue. While hue differences are indeed subtle, this separation allows for better visualization and highlights even minor variations between treatments. We believe this revised format more effectively communicates the data and addresses your concern.

The same comment for Figures 3 and 4 where cyanidin and delphinidin contents, respectively, are very low to be clearly distinguished between treatments, despite the statistical notation. 
Authors’ response: We thank the reviewer for this relevant observation. Indeed, in grape berries, malvidin and its derivatives represent the main and most stable anthocyanins, while other anthocyanins such as cyanidin and delphinidin are generally present in much lower concentrations. This explains why differences between treatments for these compounds are not as clearly visible in Figures 3 and 4, despite the statistical annotations. Moreover, in order to maintain a reasonable number of figures and to avoid excessive fragmentation of the data, it was not feasible to present separate graphs for each minor anthocyanin.

In Table 6, tannins in wine are higher in all the treatments compared to control by MCP and BSA methods. So, which is the improvement in tannin content to obtain a lower astringency?
Authors’ response: We appreciate the reviewer’s insightful comment. Indeed, Table 6 shows that tannin content, as measured by both MCP and BSA methods, increased in all treatments compared to the control. Recognizing this, we adjusted the title from "reduction of astringency" to "modification of astringency" to better reflect the nature of our findings. As discussed in the manuscript, not only the quantity but also the source and composition of tannins are crucial factors in modulating astringency. Seed-derived tannins, which are generally associated with harsher, more aggressive astringency sensations, were selectively reduced through seed removal, leading to a softer mouthfeel. Therefore, the observed "improvement" in astringency perception is attributed to the modification of tannin profile rather than a simple decrease in tannin concentration. We have now further clarified this point in the revised manuscript.

In Figure 5, a misprint in “y” axis should be corrected: Astringency. This astringency is the one obtained with sensory analysis. So, the legend should specify it: “Correlation between sensory astringency…”
In this figure, authors should also label every treatment.
Authors’ response: Thank you very much for your detailed and helpful comments. The misprint on the “y” axis of Figure 5 has been corrected to properly indicate “Astringency.” As suggested, the figure legend has also been revised to specify that the astringency values refer to sensory analysis, now reading: “Correlation between sensory astringency and ..” In addition, each treatment has been clearly labeled within the figure to improve clarity and interpretation.

Conclusions
Tannin levels found with treatments applied are higher compared to control samples. So, the wines should be more astringent. The authors explain this effect in Results and Discussion (lines 462-465). But there is not a consistent conclusion for this effect.
Authors’ response: We thank the reviewer for this important observation. As correctly pointed out, although the differences in tannin levels and their relationship to perceived astringency were discussed in the Results and Discussion section, a clear and consistent conclusion was missing.
To address this, we have now added a specific paragraph at the end of the Conclusions section that summarizes and clarifies the impact of the applied treatments on wine astringency, emphasizing the role of tannin quality and source, not only quantity, in shaping the sensory outcome. We believe that this addition improves the overall coherence and completeness of our findings, and we sincerely appreciate the reviewer’s suggestion.

References:
The cited references are appropriate.
In References 7, 10, 26, page numbers are skipped.
Authors’ response: Thank you for your observation. Regarding Reference 10, the article is identified by its article number, 112656, as the journal uses article numbers instead of traditional page ranges. Similarly, Reference 26 is identified by article number 1583, but we also found the page number. For Reference 7, we reviewed and include the appropriate page numbers to maintain consistency and accuracy in the reference list.

In Reference 23, the journal abbreviation is not the official one: J. Int. Sci. Vigne Vin. 
Authors’ response: We thank the reviewer for pointing this out. The journal abbreviation in Reference 23 has been corrected to the official format: J. Int. Sci. Vigne du Vin

A dot is missed in Reference 33: Y Glories. Il Colore…
Authors’ response: We have reformulated this part accordingly.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The authors have addressed the necessary revisions, and the manuscript is now suitable and of interest for publication.

Author Response

Response to Reviewers 
Reviewer 1:
The authors have addressed the necessary revisions, and the manuscript is now suitable and of interest for publication.
Authors’ response: We sincerely thank the reviewer for the positive evaluation and kind comments. We are grateful for the constructive feedback provided throughout the review process, which has helped us to improve the quality and clarity of the manuscript. We are pleased that the revised version is now considered suitable for publication.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The authors have well adjusted the manuscript on the basis of the comments received. However, the experimental project cannot be based on one year alone. For this reason, I believe that the manuscript cannot be published before a reconsideration of this aspect, perhaps even reconsidering the data to be included.

Author Response

Response to Reviewers
Reviewer 2: The authors have well adjusted the manuscript on the basis of the comments received. However, the experimental project cannot be based on one year alone. For this reason, I believe that the manuscript cannot be published before a reconsideration of this aspect, perhaps even reconsidering the data to be included. 
Authors’ response: Dear Reviewer,
Thank you once again for your insightful feedback and your careful consideration of our manuscript.
We fully understand your concern regarding the limitation of a single year's dataset. We would like to clarify that the experimental work was conducted in triplicate, both in the vineyard and in the winery, ensuring a robust and representative data set. Furthermore, the study continued beyond the year presented, and the data collected from the subsequent year confirm and reinforce the current findings.
However, due to the extensive volume of data generated, and in order to maintain clarity and avoid confusion in the interpretation of the results, we decided to present the two experimental years as separate studies. Specifically, in the second year, only the most promising treatments from the first year were repeated, focusing the design while still maintaining the required number of replications for both viticultural and oenological practices.
For these reasons, and to ensure that each study remains coherent and methodologically sound, we believe it is appropriate to present the current dataset independently, with the follow-up data to be developed in a subsequent publication.
We sincerely hope this explanation addresses your concerns and are, of course, open to any further recommendations you may have.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

In legend for Table 7: "Descriptive sensory evaluation of wines" please, change it by: Sensory evaluation of wines astringency.

Author Response

Response to Reviewers
Reviewer 3: In legend for Table 7: "Descriptive sensory evaluation of wines" please, change it by: Sensory evaluation of wines astringency.
Authors’ response: We thank the reviewer for the helpful suggestion. As advised, we have modified the legend of Table 7 accordingly. It now reads: " Sensory evaluation of wines astringency from different treatments based on intensity levels". We appreciate your attention to detail, which has contributed to improving the clarity of the manuscript.

Author Response File: Author Response.pdf