Chemical and Biochemical Properties of Verjuice Obtained from Vitis labrusca Grapes by Using Different Extraction Methods

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

You can find my comments/suggestions in the attached file.

Comments for author File: Comments.pdf

Author Response

Thank you for considering our manuscript. We have carefully taken into consideration all the comments and addressed them accordingly. Please see below our replies in blue. All line numbers below are from all markup view in track changes.

 

REVIEWER #1

- The manuscript addresses an interesting topic and offers valuable insights with the potential to make a meaningful contribution to the field. It is clear that the authors have invested significant effort and engaged thoughtfully with the subject matter. The introduction is well-written, providing a clear overview of the state-of-the-art and effectively highlighting the contribution of the study. The methods are presented in a clear and understandable manner.

Thank you for the comment.

- My main observations are related to grammatical and syntax issues, which, once addressed will further improve the clarity and readability of the manuscript.

Thank you for pointing this out. We have revised the manuscript again, and have made some adjustments in the English. We believe it is suitable for publication as-is now.

Abstract

- L21–22: Replace the hyphen ( - ) with the em dash (—) without spaces for introducing secondary sentences (also in L97–98).

Changes were made, please see in L21-22 and L99-100.

Keywords

- For indexing purposes, it is preferrable to use words that are not already included in the title. For that reason, I suggest replacing the first keyword and adding one or two more.

Thank you for your suggestion. We deleted ‘verjus’ and added the following ones: ‘steam extraction’, ‘centrifuge extraction’ and ‘pressing extraction’. See additions in L36-37.

Introduction

-L48: Are there any economic data regarding verjuice?

We are not sure what the reviewer meant by ‘economic data’. If it is related to the amount of verjuice sales, to the best of our knowledge, current volumes of verjuice production are not available. If it is related to the economic feasibility of verjuice production, there is a recent study that was done in Texas, United States (https://doi.org/10.3390/beverages9030078). However, since this aspect is not the focus of the present research, it was not included in this work.

-L52: Replace “expand the volume” with “expand in volume”.

Thank you. Change was made, please see L54.

-L52: Replace “skin go soften” with “their skin softens”.

Thank you. Change was made, please see L54-55.

- L54: Add a space between values and units (also in L55, L128, L130–131, L138, L144–146, L153–154, L174, L178, L180, L202, L211, L229, L231, L238, L241–245).

Changes were made throughout the manuscript.

- L76: Italicize “in natura” (also in L78). L100: Replace “for its” with “for their”.

Changes were made, please see L78, L80 and L103.

Materials and Methods

- L108: Remove “in” before “November”.

We have edited that sentence, as the 29-31 are not dates in November but the time point/ripening stage of phenological classification scheme proposed by Coombe (1995). Sentence (L111-L112) now reads as: “The grapes were harvested at the ripening stage between 29-31 according to the phenological classification scheme proposed by Coombe (1995), in November 2023 in the Rio Grande do Sul, which is the most Southern state in Brazil”.

- L119: Please indicate which chlorinated compound was used.

We used sodium hypochlorite. The information was added to L123-124 now.

- L166: Replace “titration of” with “titration with”.

Thank you for the correction. Change was made in L173.

- L173: Replace “were mixed” with “was mixed”.

Thank you for the correction. Change was made in L180.

- L173: Replace “and” with a comma.

Thank you for the correction. Change was made in L180.

- L179: Remove the en dash (–).

Thank you for the correction. Change was made in L183.

- L179: Please add the total duration of the run, for replication purposes.

Thank you for the appointment. Total duration was 67 minutes, and the information was added in the text in L187-188.

- L190: Replace “carried out by” with “evaluated using”.

Change was made in L198.

- L191: Replace “capacity” with “assay”.

Change was made in L199.

- L194: Define “TEAC” (also in L200).

We miswrote the abbreviation, apologies. We were referring to TE (Trolox Equivalent). This was amended in L202 and L208 now.

1. If the “AC” stands for “Antioxidant Capacity”, then you should change to “TE” and add the unit (mmol) in the parenthesis.

Thank you for pointing this out. We made the changes in L202 and L208.

1. In Figure 2, L411, and L415 you use TEC/L. If the “C” stands for “Concentration”, then you should remove the “/L”, as the definition of concentration already contains volume.

The antioxidant activities were expressed as mmol of Trolox equivalent per liter (mmol TE/L). We made the amendments throughout the manuscript and within the figures.

- L194: Add “radical scavenging assay” after “(DPPH)”.

We added this in L203 now.

- L200: Add “assay” after “(RP)”.

We added this in L209 now.

- L214: I suppose you added 800 μL, not 800 mL.

Yes, apologies for the typo. We have amended the unit to µL in L223.

- L235: Replace “analyzes” with “analyses”.

Change was made in L246 now.

- L252: Reference No. 32 is missing here.

Thank you for pointing this out. Addition was made in L266.

Results and Discussion

 L272: Replace “ranged from X and Y” with “ranged from X to Y”.

Thank you for this correction. Changes were made in L298.

- L276: Replace “yield” with “yields”.

Thank you, change was made in L301 now.

- L278: You should add the standard deviation. If you performed the Tukey’s HSD test in these results, you should also add that information.

Since yield was calculated as %, data does not fit the ANOVA presumptions for parametric statistical analysis. Therefore, we would like to keep the presentation of the results as they are.

- L358: Change to “indicating features similar to V. vinifera-based verjuices.”.

Change was made in L384-385.

- L361: Applying SE in CO grapes resulted in a higher verjuice yield when compared with other extraction methods. However, TSS of those samples were not significantly different. How do you explain that?

Steam extraction uses heat to rupture grape skins and pulp, which facilitated more release of juice, resulting in a higher yield. TSS measures the concentration of soluble solids (mostly sugars and acids) in the juice. Although more juice was extracted, the concentration of TSS remained similar, likely because the additional liquid extracted was proportionally similar in composition. However, as discussed in Lopes et al. (2016)’s work, when juices were prepared with steam extraction, the incorporation of water caused a reduction in the concentration of soluble compounds, contributing to the lower TSS values in the juices. This has been now added in L390-393 to cover this aspect.

- L388: The placement of “’s” is inappropriate. You should rephrase as “as demonstrated in the work of …”.

Change was made in L418 now.

- L393: For y axis units of Figure 2c and Figure 2d, see comment for L194.

Thank you for the appointment. We have made the changes in both figures now to reflect the right unit (mmol TE/L).

-L404: You have used lowercase letters to indicate different sections of Figure 2. Please be consistent (also in L406, L411, L416, L426, L430, L431).

Thank you for pointing this out. Changes were made throughout the manuscript.

- L411: See comment for L194 (also in L415).

We made the changes in L441.

-L422: According to Figure 2e, the effect of extraction method is significant (p=0.012). However, you indicate that samples from the same cultivar and different extraction methods belong to the same group (a) according to the Tukey’s HSD test. I suspect that either the p-value or the letters are wrong.

Thank you for the appointment. The statistical data presented in Figure 2e (p_extraction=0.012) is related to the effect of extraction methods. Since the objective of the present work was to evaluate the extraction methods within grape varietals, the statistical data details were not shown in the Figures. Different letters in the Figure indicate significant differences among extraction methods for each grape variety based on ANOVA (p_varietal=0.926). To make it clearer, we rephrased the sentence in L452-455.

-L437: Replace “bounds” with “bonds”.

Amended in L470.

 -L440: Add significance (see comment for L422).

Thank you for the correction. Information was added in L474.

-L452: Restate that alkanes are not included in Figure 3 and Table S1 and why.

We restated the information in L487-488 now and as footnote in Figure 3 and Table S1.

-L493: Change “presented” to “present”.

Corrected in L529.

- L520: Do you mean “thermal”?

Yes. We made this correction in L560.

-L525: Rephrase the last part of the sentece, after the comma.

This sentence has been amended to “Terpenes are often characterized by citric and floral aromas in beverages [44], which have been shown to drive liking of verjuice as a beverage [9].”  See in L566-567.

Conclusions

L542: Since you mention that juice grapes can also be used for verjuice production (L543– 544), you should highlight the fact that V. labrusca is mainly used for juice production.

Thank you for the appointment. We have highlighted this information in L585.

Abbraviations

L582: There are some abbreviations missing (e.g., TA).

We added the missing information.

 

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

Please consider some recommendations given below:

• Re-arrange the Introduction section as now new reader cannot follow a clear line of presenting the information given in the section;
• The file is uploaded with traces of track changes and comments, please correct
• there are some missing references in section materials and methods
• unify the font used for your figures so that it aligns with the whole manuscript
• try to clear out your conclusions, what is your plot line - to highlight the best extraction or variety studied
• abbreviation list should be updated
• reference list should be updated for minor inconsistencies

Author Response

Thank you for considering our manuscript. We have carefully taken into consideration all the comments and addressed them accordingly. Please see below our replies in blue. All line numbers below are from all markup view in track changes.

REVIEWER #2

Dear Authors,

Please consider some recommendations given below:

- Re-arrange the Introduction section as now new reader cannot follow a clear line of presenting the information given in the section;

Thank you for your suggestion. We appreciate your perspective; however, we believe that the current order of information in the Introduction section aligns with the context and narrative we aim to establish for the readers. The structure was intentionally designed to gradually build the background and lead to the objectives of the study. Therefore, we would like to keep as-is.

- The file is uploaded with traces of track changes and comments, please correct

Apologies for this mistake. The revised version does not contain comments, only the tracked changes indicating the revision in this new version.

- there are some missing references in section materials and methods

Thank you for pointing this out. We added the reference [32] in the VOC section (see L266).

- unify the font used for your figures so that it aligns with the whole manuscript

Thank you for the appointment. We reviewed the figures to keep consistency. All fonts were standardized for Palatino Linotype.

- try to clear out your conclusions, what is your plot line - to highlight the best extraction or variety studied

Thank you for the appointment. We intentionally avoided stating a single "best" extraction method, as verjuice has a range of potential applications, including as a beverage on its own, as an ingredient in cocktails, cooking, etc. As highlighted by Dupas de Matos et al. (2024) [9], the preferred characteristics of verjuice can vary based on the intended use. For example, when consumed as a beverage, higher TSS and lower acidity are likely to be preferred due to sweetness being the main driver of liking. In contrast, for culinary uses such as marinades or salad dressings, higher acidity is often more desirable. Therefore, rather than stating one method superior overall, we clarified in the conclusion that the optimal extraction method depends on the specific application. See additions in L596-601.

 

- abbreviation list should be updated

Thank you, we have now added the abbreviations that were missing.

- reference list should be updated for minor inconsistencies

Thank you for the appointment. We added the additional information.

Reviewer 3 Report

Comments and Suggestions for Authors

Dear Authors

The main objective of this study was to evaluate the effects of three extraction methods (pressing, steam extraction, and centrifugal blender extraction) on the chemical and biochemical properties of verjuice produced from three Vitis labrusca varieties (Bordô, Concord, and White Niagara). However, I´m sending you some observations to improve the document.

Observation 1. Numerical inconsistencies in performance (Table 1 vs. text). In the text (Yield), for example, the WN is reported as 62.12%, CO 57.36%, and SE; in Table 1, the CO is reported as 57.38% and WN 42.12%. Please correct the text and table to ensure they match, and add standard deviations/CI, if available. (This affects interpretation and reproducibility).

Observation 2. I recommend reducing the abstract to ≤200 words in a single paragraph following the recommended order (context–methods–key quantitative results–applied conclusion). Avoid repetition and general phrases.

Observation 3. Statistics: Clarified design and reporting. You indicate "triplicates of two independent extractions" and two-way ANOVA + Tukey test. Specify:

- Actual experimental unit for each variable (bottle/batch/extraction?) and n per cell for ANOVA.

- Assumption checks (normality, homoscedasticity) or transformations.

- Report effects and interactions with F (df1, df2), exact p, and (when applicable) partial η² or ω².

Observation 4. In figures, ensure consistency between asterisks/ “*” and Tukey letters; ideally, use only letters per panel and make n visible.

Observation 5. Supplement and citation (“Table S1”). The link “https://www.mdpi.com/article/doi/s1” is a placeholder. Cite the supplement within the text (“Table S1: …”) and upload the file as Supplementary Materials on the platform; the final link will be added by the production team. Maintain consistency of compound codes between Figure 3 and Table S1 according to the MDPI templates.

Observation 6. Technical English and consistency of the manuscript. Correct typos: “thermased” → “thermal,” “Niagara/Niagara” (consistency), “varietal(s)” vs. “variety/varieties.” Units and spaces should be unified (e.g., g/L, ºBrix), decimal separator period, and p style (lowercase, p = 0.0035). I recommend a thorough review of the entire document.

Observation 7. Methods—replicable details. I recommend indicating the make/model and parameters of all instruments already listed (ok), but add many critical reagents when they matter, and exclusion criteria (if applicable). HS-SPME/GC-MS: add the number of blanks, detection limits/quantities (if available), or state that it is semi-quantitative by IS (mention it; highlight it in the Limitations section).

Observation 8. Author Contributions (CRediT model). The proposed paragraph is appropriate, but MDPI prefers to use CRediT roles (Conceptualization, Methodology, Validation, Formal Analysis, Investigation, Writing—Original Draft, Writing—Review & Editing, Supervision, Funding). These roles align exactly with the journal template.

Observation 9. I recommend carefully reviewing the format of the references section.

Observation 10. Discussion: Compared with V. vinifera verjuices; please strengthen.

-Thermal dilution effect on SE (TA, TSS, sugars) with quantitative data and reference.

-Phenolic–ACE activity link: If there are no correlations, a Pearson/Spearman correlation between TPC/ABTS/DPPH and E-I (even in supplements) could be added.

-Sensory driver VOCs: Map families (esters, terpenes, aldehydes) with key aroma descriptors and connect with reported drivers of liking (ref. verjuice consumers).

Author Response

Thank you for considering our manuscript. We have carefully taken into consideration all the comments and addressed them accordingly. Please see below our replies in blue. All line numbers below are from all markup view in track changes.

REVIEWER #3

Dear Authors

- The main objective of this study was to evaluate the effects of three extraction methods (pressing, steam extraction, and centrifugal blender extraction) on the chemical and biochemical properties of verjuice produced from three Vitis labrusca varieties (Bordô, Concord, and White Niagara). However, I´m sending you some observations to improve the document.

Thank you for the comment.

- Observation 1. Numerical inconsistencies in performance (Table 1 vs. text). In the text (Yield), for example, the WN is reported as 62.12%, CO 57.36%, and SE; in Table 1, the CO is reported as 57.38% and WN 42.12%. Please correct the text and table to ensure they match, and add standard deviations/CI, if available. (This affects interpretation and reproducibility).

Thank you for noticing this, and apologies for the typo. Data in Table 1 is correct and we have now amended the text in L298, 301-302.

- Observation 2. I recommend reducing the abstract to ≤200 words in a single paragraph following the recommended order (context–methods–key quantitative results–applied conclusion). Avoid repetition and general phrases.

Thank you for your suggestion. We have revised the abstract to reduce words (196 words currently), while following the appropriate structure.

- Observation 3. Statistics: Clarified design and reporting. You indicate "triplicates of two independent extractions" and two-way ANOVA + Tukey test. Specify:

Thank you for your comment. We have clarified now in 2.2 Extraction procedures that each extraction method was performed in duplicate (independent experiments). The bottles of each extraction were analyzed, with a total of three bottles (triplicate) per extraction method. We added this information in L159-161.

- Actual experimental unit for each variable (bottle/batch/extraction?) and n per cell for ANOVA.

We added detailed information between L159-161 (“Each extraction method for each grape varietal was performed twice (independent experiments) and three bottles of each experiment were used to perform the instrumental analysis (triplicates).”), and we believe our experimental and statistical procedures are clearer now.

- Assumption checks (normality, homoscedasticity) or transformations.

We have checked all data and added the information in the data analysis section that was missing, please see information in L273-276.

- Report effects and interactions with F (df1, df2), exact p, and (when applicable) partial η² or ω².

Thank you for the appointment. The p-values from ANOVA were presented in the Figures. Regarding F-values, thus, we do not believe this is necessary or add much value to this manuscript, following as most of previous literature on the topic. Regarding η² or ω².values, ANOVA from XLSAT does not calculate these parameters. Thus, we respectfully would like to keep the presentation of results as-is.

- Observation 4. In figures, ensure consistency between asterisks/ “*” and Tukey letters; ideally, use only letters per panel and make n visible.

Thanks for the comment. The a,b,c letters indicate differences among extraction methods (CJE: centrifuge juicer extraction; PE: pressing extraction; SE: steam extraction) for each grape variety (□ in black: Bordô, ○ in red: Concord, ◊ in blue: White Niagara), which was our main focus. Since we have varietal and extraction, as fixed factors, we also looked at their interaction. When the interaction was significant, this was indicated with * next to each method in x axis. This information has been already shown in the figure’s notes.

-Observation 5. Supplement and citation (“Table S1”). The link “https://www.mdpi.com/article/doi/s1” is a placeholder. Cite the supplement within the text (“Table S1: …”) and upload the file as Supplementary Materials on the platform; the final link will be added by the production team.

We have uploaded the supplementary material into the submission platform previously. Same file has been uploaded again. Table S1 was previously cited in the manuscript, see in L478-479, 486, 521, 523, 530, 532, 557, 559, and 562 now.

Maintain consistency of compound codes between Figure 3 and Table S1 according to the MDPI templates.

Thank you for the comment. We reviewed the items.

Observation 6. Technical English and consistency of the manuscript. Correct typos: “thermased” → “thermal,” “Niagara/Niagara” (consistency), “varietal(s)” vs. “variety/varieties.” Units and spaces should be unified (e.g., g/L, ºBrix), decimal separator period, and p style (lowercase, p = 0.0035). I recommend a thorough review of the entire document.

We have revised the entire manuscript and corrected these inconsistencies.

Observation 7. Methods—replicable details. I recommend indicating the make/model and parameters of all instruments already listed (ok), but add many critical reagents when they matter, and exclusion criteria (if applicable). HS-SPME/GC-MS: add the number of blanks, detection limits/quantities (if available), or state that it is semi-quantitative by IS (mention it; highlight it in the Limitations section).

VOC determination procedure was updated. The method used is semiquantitative, and the signal-to-noise method used considered a 3:1 ratio. See new information in L234-235, 239, 257-260, 267.

-Observation 8. Author Contributions (CRediT model). The proposed paragraph is appropriate, but MDPI prefers to use CRediT roles (Conceptualization, Methodology, Validation, Formal Analysis, Investigation, Writing—Original Draft, Writing—Review & Editing, Supervision, Funding). These roles align exactly with the journal template.

We have now changed the information to match the CRediT model.

-Observation 9. I recommend carefully reviewing the format of the references section.

We reviewed the references again, thank you for mentioning this.

Observation 10. Discussion: Compared with V. vinifera verjuices; please strengthen.

-Thermal dilution effect on SE (TA, TSS, sugars) with quantitative data and reference.

Thank you for the appointment. Reference from Bresolin et al. (reference number 34) was added. We do not have the information about the exact amount of exogenous water added into the juice, so unfortunately we cannot comment on that.

- Phenolic–ACE activity link: If there are no correlations, a Pearson/Spearman correlation between TPC/ABTS/DPPH and E-I (even in supplements) could be added.

Thank you for the appointment. We have now performed the Person correlation analysis although it was not the focus of the present work (see table below). Since the manuscript is already close to 9000 words, we believe that this information will not add much value in the discussion. Thus, we prefer to keep the information reported as-is.

r-values of Pearson correlation below.

	ABTS	DPPH	Reducing powder	Anti-hypertensive	Total polyphenolics
ABTS	-	0.864934	0.924986	-0.271247981	0.337481289
DPPH		-	0.860547	-0.314140325	0.035008871
Reducing Power			-	-0.23258133	0.422875679
Anti-hypertensive				-	0.060565098
Total polyphenolics					-