Interaction between Vitamins C and E When Scavenging the Superoxide Radical Shown by Hydrodynamic Voltammetry and DFT

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Intro Paragraph 1 appears to have inconsistency in the font (may just be font size)

line 49.  Not sure if it is necessary to capitalize Vitamin C & E, but capitalization is inconsistent throughout the introduction.  In the first couple paragraph they are always capitalized.  After line 49 it is not.  It feels like this is more correct, but it should be consistent throughout the manuscript.

3.1 As a whole, I am a little concerned about the use of BLYP/DNP.  While I concede these are moderately sized molecules and may not be easily computed with higher accuracy methods/basis set combinations, GGAs such as BLYP are known to underestimate barrier heights.  While it may indeed be a small barrier height, a series of single point energy computations with B3LYP/TZ and wB97xD/TZ may help show these barriers are indeed small and not just an artifact of using an overly simplistic method and small basis set. 

line 208 Font in reaction 1 also looks a little different 

In Figures 2, 5,6,7 (and what appears to be mislabeled as) S.1 (Since this is not in supporting information), 9, 10, 11, S.2 (also mislabeled), 12, 13 labels overlap with each other and with the figure.  These should be moved to make it more legible to the reader.  

Line 243 Remove second period

References capitalization seems a bit inconsistent.  In some article titles, all words are capitalized.  In some article titles only the first word is capitalized.  This should be changed to be consistent with journal format.

Comments on the Quality of English Language

Minor typos, but well written overall

Author Response

Intro Paragraph 1 appears to have inconsistency in the font (may just be font size)

Thanks: We double checked font and font size and now there are no inconsistencies.

line 49.  Not sure if it is necessary to capitalize Vitamin C & E, but capitalization is inconsistent throughout the introduction.  In the first couple paragraph they are always capitalized.  After line 49 it is not.  It feels like this is more correct, but it should be consistent throughout the manuscript.

Thanks for telling us about this inconsistency. It is now updated; only lower case is used.

3.1 As a whole, I am a little concerned about the use of BLYP/DNP.  While I concede these are moderately sized molecules and may not be easily computed with higher accuracy methods/basis set combinations, GGAs such as BLYP are known to underestimate barrier heights.  While it may indeed be a small barrier height, a series of single point energy computations with B3LYP/TZ and wB97xD/TZ may help show these barriers are indeed small and not just an artifact of using an overly simplistic method and small basis set. 

We have used BLYP for our work primarily based on information obtained from this reference: Density Functionals with Broad Applicability in Chemistry. Yan Zhao and Donald G. Truhlar. Accounts of Chemical Research 2008 41 (2), 157-167 DOI: 10.1021/ar700111a). In this paper, the authors state: “it [B3LYP] systematically underestimates reaction barrier heights; it is inaccurate for interactions dominated by medium-range correlation energy, such as van der Waals attraction, aromatic-aromatic stacking”.

We are certainly interested in what the reviewer considers appropriate and, for future studies, are open to suggestions regarding the functionals described by the reviewer.

line 208 Font in reaction 1 also looks a little different 

It is now consistent with the whole manuscript

In Figures 2, 5,6,7 (and what appears to be mislabeled as) S.1 (Since this is not in supporting information), 9, 10, 11, S.2 (also mislabeled), 12, 13 labels overlap with each other and with the figure.  These should be moved to make it more legible to the reader.  

To make the figures more legible to the reader, we consolidated in one figure the pictures of Reagent, TS and Product, whenever a TS is involved in our DFT reactions.  Unfortunately, distances from figures obtained through use of Biovia software cannot be modified. However, to clarify the figures, we specified relevant distances in figure captions. For instance, Figure 2 includes Reagent, TS and Product, and we describe the TS in the figure caption this way:

“The transferred H atom is equidistant from both moieties in this TS: O(superoxide)---H = 1.219 Å, O6---H = 1.209 Å.”

The correct supporting information figure list is now updated at the end of the manuscript.

Line 243 Remove second period

Done

References capitalization seems a bit inconsistent.  In some article titles, all words are capitalized.  In some article titles only the first word is capitalized.  This should be changed to be consistent with journal format.

Done

Reviewer 2 Report

Comments and Suggestions for Authors

In this manuscript, Caruso and co-workers reported a detailed voltametric and computational study interaction between vitamin C and E and efficiency of scavenging the peroxide radical. They reported that the combined form has ~5 times more effective power than individual vitamins. They have also described the mechanism of radical capture and investigated the type of interaction exists between vitamin C and E. Overall, this is a well-executed study which, in my opinion, is important in the biophysical chemistry. The manuscript is nicely written and presented very well. Thus, I support publication of the present manuscript in Biophysica. However, there are certain minor issues which should be addressed prior to publication:

1) Why this peroxide radical scavenging in important? This should be nicely explained in introduction.

2) The authors have used pure DFT functional BLYP for the whole study. I am not sure about the accuracy of this functional in terms of geometry and energy. So, they should describe in manuscript why this method has been used.

3) They made individual figures for each transition states. It is difficult to go through. I would suggest putting similar types of TSs into a single figure and make a nice picture.

4) Did they investigate the explicit solvation model? If yes, then discuss in main manuscript. If no, then I would suggest to investigate that. The reaction involves proton transfer, that might be influenced by explicit solvent molecules.

Author Response

See attached file

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The functional still needs attention.  The reference Yan Zhao and Donald G. Truhlar. Accounts of Chemical Research 2008 41 (2), 157-167 DOI: 10.1021/ar700111a that the authors have added to their response only validates my original concern that BLYP is not an appropriate functional for this study.  While B3LYP may not be a much better choice, the reference by Truhlar shows that Becke's 88 exchange is notoriously bad for medium range correlation.  To fix this, the author may want to consider one of Truhlar's Minnesota functionals (See Zhao and Truhlar, Org. Lett. 2006, 8, 25, 5753–5755 for evidence that M06-2x is superior to both BLYP and B3LYP ) that are in the reference cited in the response.  Or perhaps the authors should look at the B97D3BJ, functional that has been created by Grimme, (See Grimme, S. Angew. Chem., Int. Ed. 2006, 45, 4460. for his original article on medium range correlation) who is often cited as the leading expert on medium range correlation.  This functional is found to be one of the best to treat medium range correlation.  The wb97XD functional from Head-Gordon that I also suggested has been found to be about as good for medium range correlation as Grimme's B97D3BJ.  You may want to use Grimme's GGA, as it is likely more computationally efficient.  My concerns have not yet been addressed and the reference you cite prove your results are not publishable without an update of your Density Functional Approximation.  

Author Response

see attached file

Author Response File: Author Response.pdf

Round 3

Reviewer 1 Report

Comments and Suggestions for Authors

Modifications are acceptable.  I recognize that for medium sized molecules such as this, geometry optimizations can take extensive time.  To clarify, utilizing a GGA to find geometries while using the higher level related hybrid functional at a single point energy computation would verify that barriers are not underestimated  and would be a time efficient way to alleviate concerns in the future.  I might also suggest that open source codes, such as Psi4 or Orca, have an extensive DFT functional library and would not require licensing concerns.