Effects of Methyl Jasmonate and Nano-Methyl Jasmonate Treatments on Monastrell Wine Volatile Composition

Reviewer 1

List of changes in the current main document / or a rebuttal

The results and discussion should come after the materials and methods section

How was de pH meter verified

GC-MS verification

There has been usage of multivariate techniques done in the past to separate out wines and spirits, the authors should cite the following paper: Burns, Rachel L., et al. “ A fast, straightforward and inexpensive method for the authenticacion of Baiju Spirit Samples by Fluorescence Spectrospocopy

Our sincere thanks for the review of the article, and for the opinions generated by the reviewer. This journal has a template to present the paper and we have followed the instructions given by the journal to the authors. This is the reason why we have written the different sections of the article in this order.

We have not detailed the calibration mode of the pH meter due to the fact of it is a basic operation that we perform every day in our lab before any series of measurements, and we considered that was not necessary to specify , either way we calibrate it following the instrument instructions and using two certified standards at pH 4 and pH 7

The equipment is verified daily in our laboratory before analyzing each batch of samples by performing an e-tune. This is an operation to verify that all the parameters of the equipment are correct. Once this is done, we begin to analyze our samples.

We appreciate the suggested article, we have reviewed it and it is interesting the use of emission spectra to differentiate different types of liquor samples. But we do not see a direct relationship with the determination of the aromatic profile of the wines. The used of discriminant analysis to separate groups is very common and because it is a statistical tool we didn´t consider necessary to add any reference.

Reviewer 2

Please carefully reread the manuscript, there are many tipo mistakes.

Please write the numbers of units in superscript an d the numbers of chemical formula in subscript

L425: replace ppb and ppm by µg/L and mg/L

Have the wines been analyzed over 3 years (meaning wines made in 2019 analyzed in 2019, wines from 2020 analyzed in 2020…) or have they been analyzed at the same time (all three wines analyzed in 2021)?

If the wines were analyzed at the same time this could also explain why the lowest volatile content was observed in the first year wine.

Same question for sensory analysis

Concerning volatile compound analyses by HS-SPME-GC-MS, why did the authors choose san mode instead of SIM mode? SIM mode is much more appropriate for quantitation than SCAN mode.

Moreover, the authors prepared calibration solutions in model wine. Did the authors investigate a potential matrix effect? Is it possible that the quantification is overestimated with a calibration curve prepared in model wine?

Concerning sensory analysis, please provide results of statistical analysis of triangular test. The preference test require a large number of tasters, which is not the case in this study since there are only 12 tasters. The statistical results on taster preference do not really make sense here with only 12 persons, and therefore I suggest removing the results and only discussing the results of the triangular test.

Statistical analyses: the authors used one-way ANOVA to analyze the data. Did they verify if the data was normally distributed? Please provide results of normality tests.

The errors mentioned in addition to others have been fixed through the manuscript in the following lines:

L15, L39,L52,L59,L63,L100,L113,L116,L119,L177,L183-185,L191,L207,L212,L216,L-220-221, L223,L234,L248,L265,L267,L294,L297,L311-314,L319,L335,L337,L352-356,L368-369,L376,L392,L397,L399,L433,L437,L441,L456

We have changed it in the manuscript.

The different wines were analyzed over 3 years for them to prevent evolution processes that would mask the effect of the treatments. So that question can be better  understood we have rewrite L402/403 :” All samples wine were analyzed by triplicate at final alcoholic fermentation and in the same season that them were produced”

If this were the case it would be probable but because every wine was analyzed at its production year and close to time of having finished the alcoholic fermentation, the probable reason on this difference was the season, as we said, 2019 register an important rainfalls before harvest time.

The sensory analysis was performed over the three years. The judges were the same, the workers of Oenological Station of Jumilla trained previously.

As the reviewer suggests, the SIM method is more appropriate to SCAN once the compounds of interest have been identified, but our methodology is that used by Moreno-Olivares et al. 2020, which in turn is based on that of Gomez-Plaza et al. 2012, which used the SCAN mode. Since our analyses are already done, we keep it in mind for the future.

As the reviewer suggests, this overestimation could occur, but the interest of our work was to compare the aromatic composition in wines in which different treatments had been applied. Therefore, this overestimation, if it were to occur, would not influence us in the comparison, since in all cases we are using the same matrix.

Regarding statistical analysis of triangular test, Roessler et al. ¹ provides tables which indicate the number of tasters and the number of correct answers  to each significance level. The minimum number of tasters is 5.

As for preference test we admit that 12 tasters would be not enough to considerer statistical results. We have removed from the text these statistical consideration.

¹Roessler, EB;Pangborn, RM;Sidel, JL; Stone H. Expanded statistical tables for estimating significance in paired-preference, paired-difference, duo-trio and triangle tests. J Food Sci 43:940–943 (1978).

We are verified the normality distribution of the data using Shapiro wilks test because our parameters has less of 50 data. We show some examples of it, because showing all the results would be too long.

Reviewer 3

1. Check the words, symbols and spaces in the text and results.

2.The text words presented in fig. 2-3 are not clearly visible. At least, it needs to be modified to a similar size so that it can look like fig. 1

3. The amount of total esters in the grape wine treated with Nano- MeJ was rather decreased compared to the control in 2019 and 2021 Even considering the error value of the control, the average value of the Nano-MeJ treatment group was low. What do you think is the reason?

4. Do you think the treatment technology using these nanoparticles will have an effect on the quality improvement of other fruit fermented alcoholic beverages?

5. Although calcium phosphate nanoparticles have various advantages, there are articles reporting that this induces apoptosis. Can you comment on this?

We have done it and corrected them in the following lines in the manuscript:

L15, L39,L52,L59,L63,L100,L113,L116,L119,L177,L183-185,L191,L207,L212,L216,L-220-221, L223,L234,L248,L265,L267,L294,L297,L311-314,L319,L335,L337,L352-356,L368-369,L376,L392,L397,L399,L433,L437,L441,L456

We have replace for another figures with a bigger typography  in the manuscript.

The decrease in total esters is due to the fact that the ethyl acetate decrease, if we consider de fruity esters (total amount- ethyl acetate contribution) we observed and increased all the treatments vs. control and all the seasons.

We think that because MeJ induce the secondary metabolism in plants, its effects would be found in others fruits and would be transferred to other fermented beverages made with them, in addition as can be observed in our study season is an important factor to be considerered.

We have read the comment article and we think that is an important factor to considerer but in our work we have had no notice about apoptosis signals in the treated vines so we have no considered this fact.