Dilution Method of Menthol Solutions Affects Subsequent Perceptual Thermal Responses during Passive Heat Exposure in Non-Heat Acclimated Participants

Round 1

Reviewer 1 Report

2-3. Write on your title: The effects of two menthol solutions at physiological variables and subsequent thermal responses during passive heat exposure on athletes (it is important to show the level of your participants). 
8-20. Give details about your participants and add some statistical indexes in your results (ex. Some p values only for the variables which show significant differences).
4-5. Add affiliations 
30 & 33. Please, write the full name of the abbreviations TRPM8 and TRP
39-40. Could you give in some rows an example for all these effects? 
43. Do you talk about the menthol’s use as a mouth rinse?
60. Remove comma (,)
Is there in the existing literature any disadvantages from the use of menthol? If yes, it is important to add it. 
71. Write only “Table 1”. 
71. Add the age of your participants in that point of the text and in parenthesis write about Mean and St.Dv. Thus, remove (Mean ± Standard Deviation) from the table’s title. 
The athletic level of your participants was high, medium, or low. It is important to define this detail because probably affects your results. 
81. This information must be added to statistical analysis.
85. Instead of “participants” write “Also they were requested…”
86. CLO. Please, explain the abbreviation
86-87. Please, rephrase the sentence. 
89. Remove (fans)
102-105. Please, rephrase the rows. It is not clear how you have occurred the measurements and how the participants tasted each solution. This explanation will solve some of my questions about statistical analysis.
112-113. I do not understand what do you want to say.
108-112. Please, add the literature that you used for the methodology. 
119-121. I cannot understand why you refer to this in the sentence. In the above sentence, you have defined the scales of TC and TS. 
174; 188; 199. Use an abbreviation for solutions A and B. Also, remove Mean ± 90% 174 CI
182. Write only the p-value in parenthesis. 
208. Change the title on table 2. Put the description about the use of bold under the table as "Bold values". Moreover, I suggest to bold the values which show statistically significant differences.  
222-223. Avoid the statistical indexes in the “Discussion”. Remove them from the other rows too.
234-238. You describe that TS affected in different time points but there is not any statistically significant difference. I suggest referring only statistically significant differences or the lack of it. 
250. Remove “Figure 5” (generally remove the Figures from your “Discussion”) and start the next sentence with “Specifically”. 
252-255. The statistical indexes are related to the “Results” section. You must remove them. Also, you describe that used Bonferroni analysis. You did not describe its use in the “Statistical analysis” neither in your “Results”.
272-282. Do you talk about responders or non-responders in a menthol? How this can be predicted?

Author Response

Thank you for the comments, recommendations and points for clarification. We appreciate the thorough review, and have included a response to each comment below. Where we disagree with the comments made, we have attempted to cite or provide several sources to better convey our stance.

2-3. Write on your title: The effects of two menthol solutions at physiological variables and subsequent thermal responses during passive heat exposure on athletes (it is important to show the level of your participants). 

Whilst we agree that if the participant population were homogenous, defining it narrowly would be appropriate, we feel that although participants were either staff or students within a sport science department, the variety of fitness levels, sport participation time and sports participated in by these individuals would actually increase the variability within the dataset, if it were to be accounted for, and thus further complicate the study design. Therefore, we prefer to leave the title as it is.

8-20. Give details about your participants and add some statistical indexes in your results (ex. Some p values only for the variables which show significant differences).

Thank you for this suggestion, we have amended the abstract to now include the following:

This study compared these two dilution methods, one using a solvent the other using temperature, via a randomized counterbalanced repeated measures design (n = 12; Height: 174.0 ± 8.5 cm Mass: 73.4 ± 13.3 kg Age: 28.7 ± 8.4 y; two exposures to each solution) to assess the effect of solution and heat exposure, upon thermal comfort, thermal sensation and associated physiological parameters in non-heat acclimated participants. 

Thermal comfort was significantly affected by solution (p = 0.041; η2 = 0.017) and time (p < 0.001; η2 = 0.228), whereas thermal sensation was significantly affected by time only (p = 0.012; η2 = 0.133), as was tympanic temperature (p < 0.001; η2 = 0.277).

4-5. Add affiliations 

Amended as requested

30 & 33. Please, write the full name of the abbreviations TRPM8 and TRP

Amended as requested for TRP-M8. We have chosen to hyphenate TRP-M8 to clarify to the reader that the TRP component of both abbreviations is the same.

39-40. Could you give in some rows an example for all these effects?

Menthol has been shown to improve time to exhaustion and time trial performance, but not repeated sprint performance, although it does improve perceptual measures during repeated sprint performance. Whilst we agree that these findings are interesting, given that the focus of the present work is not on exercise performance, we feel that the eight references provided and then directing the reader to the recent consensus statement (which is open access), is likely a better fit for the present paper, so as not to detract from the research question too much.

 
43. Do you talk about the menthol’s use as a mouth rinse?

The consensus paper does distinguish between rinsing and ingestion of menthol, however given we are outlining possible safety concerns here, we feel it best to cautious and group both rinsing and ingestion as oral application of menthol. We have included in parentheses (e.g. as a mouth rinse or ingested)

60. Remove comma (,)

Amended as requested

Is there in the existing literature any disadvantages from the use of menthol? If yes, it is important to add it. 

We refer the reviewer to the following on lines 46/48 'but also highlighted important safety considerations, concluding that if menthol is to be applied orally, that solutions are prepared according to scientific literature and in a well-ventilated area [20]. ' This is the consensus across papers which have outlined menthol's safety, with most of the evidence of harm resulting from supraphysiological doses and toxic exposure due to inappropriate working conditions. We feel the above statement suggests that avoidance of such conditions is to be strongly encouraged, but will include further comment in the discussion.

71. Write only “Table 1”. 
71. Add the age of your participants in that point of the text and in parenthesis write about Mean and St.Dv. Thus, remove (Mean ± Standard Deviation) from the table’s title. 

Thank you for the above comments. We feel that this counters journal guidelines regarding information to provide in table titles, and also believe it is more economical to note in the title that all data presented are Mean ± Standard Deviation, than to write Mean ± SD repeatedly as column headings, alongside the presented sample sizes.

The athletic level of your participants was high, medium, or low. It is important to define this detail because probably affects your results. 

Given that there is no exercise component to the trial, and we are investigating non-heat acclimated individuals for a period of ~1 hour of heat exposure, we are unaware of evidence to suggest that this would be the case. We acknowledge that physiological factors such as adiposity may increase the thermoregulatory strain experienced by individuals in the heat, but unfortunately we did not take any further anthropometric assessment beyond height and weight to confirm this.

81. This information must be added to statistical analysis.

Amended as requested.

85. Instead of “participants” write “Also they were requested…”

Amended as requested.

86. CLO. Please, explain the abbreviation

CLO is not an abbreviation but an international unit of measurement for insulation provided by clothing, hence our writing it without providing an explanation. We have provided supporting references which have employed similar protocols and quantification of insulation.

86-87. Please, rephrase the sentence. 

We are unclear as to which sentence the reviewer wishes us to revise. We have revised the previous sentence regarding the use of participants, but feel that the sentences discussing clothing worn by participants and the environmental conditions for the test are clear, and do not warrant revision.

89. Remove (fans)

Amended as requested.

102-105. Please, rephrase the rows. It is not clear how you have occurred the measurements and how the participants tasted each solution. This explanation will solve some of my questions about statistical analysis.

Thank you for this suggestion, we understand that this was not well-written and have amended it to read as follows:

'During testing participants were required to sit in the chamber for 10 minutes prior to any measures being recorded. During their first visit, participants would swill either solution A [28] or solution B [13] following 10 minutes of passive heat exposure (time point 1) and then swill the alternate solution 15 minutes later (time point 5). In their second visit, participants would swill the solutions in the opposing order to their first visit.' 

112-113. I do not understand what do you want to say.

We have amended this section to read as follows, we hope this clarifies the reviewer's comment 'The participants swilled each menthol mouth rinse solution, for a duration of ~ 5 seconds, before expectorating the solution into a cup. Thermal comfort, thermal sensation, heart rate, tympanic temperature was recorded ~ 30 seconds prior, ~ 30 seconds post, ~ 5 minutes post, ~ 10 minutes post and ~ 15 minutes post swilling of the solutions (Time points 1-5 (first swill) and 5-9 (second swill)). In the latter half of the exposure, the 15 minute post swill time point also acted as the 30 seconds prior time point (time point 5). ' 

108-112. Please, add the literature that you used for the methodology. 

Thank you, we have added the following, we hope this change satisfies the reviewer:

'We have previously used a similar design to assess differing solutions applied during passive heat exposure [31], and the time-period between swills was devised following pilot data gathered during heat acclimation studies, which are yet to be published.'

119-121. I cannot understand why you refer to this in the sentence. In the above sentence, you have defined the scales of TC and TS. 

We presume the reviewer is referring to the use of -0.5 and +0.5 for statistical purposes here. This is an important point that would allow replication of statistical analysis or for researchers to use this scale and analyse their data in a similar fashion to ours. This point is not made in the original work that is referenced, but as we go on to say fits with the direction of comfort experienced by the participant and ensures statistical tests can be carried out properly, by not treating both outcomes as 0 i.e. having no numerical value that agrees with the direction of perception.

174; 188; 199. Use an abbreviation for solutions A and B. Also, remove Mean ± 90% 174 CI

Solutions A and B are already effectively abbreviations for the methods of dilution undertaken. We do not feel that these solutions can or should be abbreviated beyond a single letter. The word solution is required. We also feel that the inclusion of Mean ± 90% CI is required, as both Mean ± SD and Mean ± 90% CI are presented at varying points throughout this paper. This distinction is both statistically and ecologically/practically important.

182. Write only the p-value in parenthesis. 

We respectfully disagree with this recommendation for revision. It is our understanding that when reporting F values as the result of conducting an ANOVA of any sort that the degrees of freedom for the effect and the error should be reported, alongside the resultant p value, as they convey differing statistical information. Likewise we have included eta squared as a measure of effect size that commonly accompanies ANOVA calculations on programmes such as SPSS and JASP, and is used to describe the variance explained by the variable or interaction of interest. Given its similarity in this instance to r², we feel it is of importance and of interest to readers.

208. Change the title on table 2. Put the description about the use of bold under the table as "Bold values". Moreover, I suggest to bold the values which show statistically significant differences.  

We have partially made this amendment. We have included the bold values as a footnote as suggested, and have double checked present values and had indeed missed some statistically significant values, which also presented clear effects. These have now been included.

222-223. Avoid the statistical indexes in the “Discussion”. Remove them from the other rows too.

We respectfully disagree with the reviewer here. The statistical inferences presented here and throughout the discussion are to ensure that the data presented and discussed are done so transparently, in a manner that allows the reader to make an informed decision regarding the precision and uncertainty of the estimate of an effect(s). This is important when one considers that people may wish to use this intervention in a non-research setting, so they may have to weigh the potential effect against other interventions. Likewise, in a research setting inclusion of the presented statistical indices allows for those wishing to undertake meta-analyses to do so more easily. 

We refer the reviewer to the following papers on the matter:

Lakens, D. (2013). Calculating and reporting effect sizes to facilitate cumulative science: a practical primer for t-tests and ANOVAs. Frontiers in psychology, 4, 863.

Bakker, A., Cai, J., English, L., Kaiser, G., Mesa, V., & Van Dooren, W. (2019). Beyond small, medium, or large: points of consideration when interpreting effect sizes. Educational Studies in Mathematics, 102(1), 1-8.

234-238. You describe that TS affected in different time points but there is not any statistically significant difference. I suggest referring only statistically significant differences or the lack of it. 

Based on the articles referenced above, and an increasingly common approach across a range of scientific disciplines, we decline to take this position. We feel that presented results as either statistically significant or not, presents a false dichotomy and prefer to incorporate the observed size of an effect and a degree of its uncertainty, as opposed to just an assessment of whether or not the observed data or a more extreme datum could have occurred by chance. To reiterate, this practice is widely adopted and we encourage the reviewer to consider the points raised in the Bakker paper above, and the work of Daniel Lakens on this issue, as well as the papers below:

Kraemer, H. C., Neri, E., & Spiegel, D. (2020). Wrangling with p‐values versus effect sizes to improve medical decision‐making: A tutorial. International Journal of Eating Disorders, 53(2), 302-308.

Lakens, D., & Etz, A. J. (2017). Too true to be bad: When sets of studies with significant and nonsignificant findings are probably true. Social Psychological and Personality Science, 8(8), 875-881.

Sullivan, G. M., & Feinn, R. (2012). Using effect size—or why the P value is not enough. Journal of graduate medical education, 4(3), 279-282.

Greenwald, A., Gonzalez, R., Harris, R. J., & Guthrie, D. (1996). Effect sizes and p values: what should be reported and what should be replicated?. Psychophysiology, 33(2), 175-183.

250. Remove “Figure 5” (generally remove the Figures from your “Discussion”) and start the next sentence with “Specifically”. 

Thank you for this suggestion. We have amended this to now read Figures 3 and 4, and begun the next sentence with specifically. We have chosen to include the reference to figures so as to better direct readers attention to the appropriate figure.

252-255. The statistical indexes are related to the “Results” section. You must remove them. Also, you describe that used Bonferroni analysis. You did not describe its use in the “Statistical analysis” neither in your “Results”.

We disagree with the reviewer regarding including results that have already been reported in the discussion. This is common practice in many fields and arguably most papers would make little sense without including some data in their discussion, especially when discussing multiple measures across different times and or treatments. We acknowledge that we do not report a Bonferroni correction being performed in the methods; this has been added. We do however feel that it is important to acknowledge the range of p-values and the fact that significant results were obtained despite the conservative nature of the Bonferroni correction, given our relatively small sample size. This is important given the magnitude of some findings, in light of the robustness of the statistical correction(s) that have been performed.

272-282. Do you talk about responders or non-responders in a menthol? How this can be predicted?

Thank you for this question. Response to menthol, and how this can be accurately assessed, is potentially problematic if we were wanting to define someone as a responder or non-responder. We say this because even individuals without taste or smell can still experience menthol related effects, although they may lack perceptual awareness of menthol's most common factors. We have included commentary regarding genotyping, but have now also included a general comment regarding response heterogeneity, and how this may be incorporated in future assessments of menthol work.

Reviewer 2 Report

This is a very well-written manuscript reporting the effects of two methol dilution methods while exposed to heat in an environmental heat chamber. The authors found small differences and potential benefits from each solution, suggesting that personalized approaches are required for athletes/users. 

Please include some of the statistical results in the abstract

Please define abbreviations in the first instance for readers who are not familiar with them

Introduction flows well and includes appropriate background literature

Strong methods, even number of female and male participants

The limitations are well stated. Please consider combining into their own paragraph, with future directions afterwards

In the conclusion, please briefly state what Solution A and B are

Author Response

Thank you for taking the time to review our manuscript, please find below comments, amendments and changes made.

This is a very well-written manuscript reporting the effects of two methol dilution methods while exposed to heat in an environmental heat chamber. The authors found small differences and potential benefits from each solution, suggesting that personalized approaches are required for athletes/users. 

Thank you for these comments

Please include some of the statistical results in the abstract

Amended as requested, the abstract now includes the following:

'Thermal comfort was significantly affected by solution (p = 0.041; η2 = 0.017) and time (p < 0.001; η2 = 0.228), whereas thermal sensation was significantly affected by time only (p = 0.012; η2 = 0.133), as was tympanic temperature (p < 0.001; η2 = 0.277).' 

We have chosen not to include p values or effect sizes for the other stated effects, as this would clutter the presentation of the abstract and make it harder for the reader to interpret.

Please define abbreviations in the first instance for readers who are not familiar with them

Amended as requested

Introduction flows well and includes appropriate background literature

Strong methods, even number of female and male participants

Thank you for the above comments

The limitations are well stated. Please consider combining into their own paragraph, with future directions afterwards

Thank you for this suggestion, we have done so and include further recommendations (lines 294 - 322)

In the conclusion, please briefly state what Solution A and B are

Thank you for this suggestion, the conclusion now includes 'solution A (0.1% menthol; diluted with ethanol and water) is recommended; whereas if lesser but more sustained effects are sought, choose solution B (0.1% menthol; heated to 40 ºC following dilution in water)'

Round 2

Reviewer 1 Report

2-3 It is important on the title to identify your participants. If the term “athletes” does not match use another term.

74. Write only “Table 1“ in parenthesis.

176, 189, 199. I still believe that the information “Mean ± 90% CI” is better to described only in the text of results. Not on the title of the figures.

183. Remove the “Figure” inside parenthesis and write it at a separate spot in the text. Of course, P or F – values must be included in your results.

222-223. In your methodology, you write that conducted measurements on the 10th and 15th minutes. Nowhere in your “Results”, these timepoints exist. I respect the reason that you add the statistical indexes in “Discussion” but it is important firstly to show all of your results in the “Results” section and then to use them in “Discussion” where is necessary.

Author Response

Thank you for providing a quick review upon the revised version, we really appreciate it, and have made the recommended revisions as outlined below:

2-3 It is important on the title to identify your participants. If the term “athletes” does not match use another term.

We have provided the amendment 'in non-heat acclimated individuals'

74. Write only “Table 1“ in parenthesis.

Amended as requested

176, 189, 199. I still believe that the information “Mean ± 90% CI” is better to described only in the text of results. Not on the title of the figures.

We have amended as requested, and feel that the wording on line 151, is sufficient as an explanation.

183. Remove the “Figure” inside parenthesis and write it at a separate spot in the text. Of course, P or F – values must be included in your results.

Thank you for this recommendation, we have removed Figure from the parentheses and in doing so have realised it needn't be mentioned here or elsewhere in this section, in relation to this point.

222-223. In your methodology, you write that conducted measurements on the 10th and 15th minutes. Nowhere in your “Results”, these timepoints exist. I respect the reason that you add the statistical indexes in “Discussion” but it is important firstly to show all of your results in the “Results” section and then to use them in “Discussion” where is necessary.

Apologies, this is due to a lack of clarification on our part. In terms of the 10th and 15th minutes post-swilling, this generally relates to time points 4 and 5, and 8 and 9, depending on the swill being discussed (see lines 118-119 for initial explanation). We believe the clause following the section you have highlighted int he discussion, support the above. 'Solution B was rated higher for thermal comfort by a clear small – moderate extent at 5 (ES: 0.74; 90% CI: 0.18 to 1.26; p = 0.027) and 10 minutes (0.49; 90% CI: 0.03 to 0.98; p = 0.121) post-swilling, early in the exposure, corresponding to time points 3 and 4.'  (Lines 222-225). We hope this satisfies the reviewer.