A Hidden Side of the Conformational Mobility of the Quercetin Molecule Caused by the Rotations of the O3H, O5H and O7H Hydroxyl Groups: In Silico Scrupulous Study

Round 1

Reviewer 1 Report

this is an interesting article resulting from a tremendous hard work.  Authors applied MP2 calculations to locate 72 transition states for conformational transitions among 48 Quercetin conformers and concluded that fast thermal equilibrium be reached with 6.5 ns.  It is not easy to grasp the real significance of the HIDDEN side!   However, I recommend this article be accepted after minor typo corrections:  (1) in Reference, Pople, J.A. was misspelled as  People, J.A.  (2)  Debye was misspelled as Debay (3) Conformer 24 in Fig. 2, U =4.07 but 'u=' was missing    and etc

Author Response

Dear Reviewer!

Thank you so much for the careful consideration and analysis of our manuscript!

According the hidden structure – This paper enables to comprehensively understand the conformational mobility of the quercetin molecule and also the potential energy surface of this molecule, which, in particular, is defined by the mobility of its hydroxyl groups. We also hope very much that this paper would make contribution into the better understanding of the physico-chemical properties of the quercetin molecule and its application for the explanation of its activity.  

We have taken into account your remarks, we have corrected the first and second corrections, while we didn’t found mistake 3) and ‘U’ as well.

Reviewer 2 Report

The authors perform a conformational analysis of quercetin using quantum chemical calculations. Specifically, they calculate the relative energy of all possible arrangements of the OH groups of this molecule and the necessary energy to go from one arrangement to another (transition states). It is an interesting work, the topic is somewhat important, and the methodology is adequate. My concern is that I am not 100% sure how their work is related to the general concept of symmetry, and if it is a proper article for this journal. In addition, the used language is rough at many points. I think that the article can be published if the following points are addressed:

The authors use the shorthand notations: O3H, O5H, O7H, rings A, C, HO7C7C8 and other similar ones both in the title and the abstract, which confuses the reader since not everyone is familiar with quercetin. I suggest that the authors keep the title and abstract simpler and more general. Similarly, a scheme with the molecule and numbered atoms is necessary for the reader to follow. The authors prompt the reader to see other previously published work for the notation they use: “In this work standard numeration of the atoms of the quercetin molecule has been used [16, 17].” To me this is not correct strategy. What if someone has no access to these other journals? Lines 94-95: This sentence does not make scientific sense. Free energy calculations needs frequencies and thus geometry optimization and not single point calculations. This has to be explained. Line 168: What does “normal conditions” mean? Do they mean standard conditions of temperature and pressure? Please write the conditions explicitly. Line 109: “default options”. This is not scientifically acceptable. You should either delete this or mention the exact used options. Some language issues are (there are more, but these can help the authors improve their language): Line 33: was -> has been Line 41: Previously, it was found in our previous research … -> In our previous study … Line 45: delete “within the range”. It is redundant. Line 46: Debay -> Debye Line 48: delete “at this” Line 50: “subfamily III” is missing Line 53: delete “however” Line 57: delete “pursued the goal to” Line 61: I don’t get the meaning of this sentence Line 62: “three conformational subfamilies”, but they mentioned four in the paragraph above Lines 66-67: Many, if not the most, of the biological molecules (and quercetin in this case is not the exception) are 67 enantiomers, which effect could be sometimes different on living systems. -> Biological molecules (including quercetin) commonly have various enantiomers with different functionality in living systems. Lines 67-68: I don’t get the meaning of this sentence Line 78: delete “including symmetrical”. It is implied by the preceding word “structural” Lines 80-81: I don’t get the meaning of this sentence Line 82: in details -> in detail Line 87: approved itself in -> benchmarked Line 93: I don’t get the meaning of the sentence Line 113: “OH…HC” are not H-bonds. Please use weak molecular interactions. Line 125: Change the title to “results and discussion” Line 125: Aiming to achieve the goal set in this work and obtain maximally possible … -> Aiming at the maximum possible ... Line 131: What does “non-orthogonal structure” mean? Line 134: “signs +/- correspond to enantioners”. I don’t get it. What enantiomers are they talking about? Why it is + and – in them? Please explain. If they refer to the structures they study, these are not enantiomers, they are isomers. Conclusions: They authors list their conclusions in numbers. I think they either should delete the numbers or add a sentence before the list saying something like “Our main conclusions are listed below” Figure Captions: the authors use the phrase “the initial relatively the terminal” I don’t get it. They mean “relative to”? Table 1, footnote m: please add the scheme of the molecule to make the description of the dihedral angle easier to the reader. Figures 1, 2, and 3 are important but too large. If the journal supports supporting information (SI), the authors may want to add some of them in the SI.

 

Comments for author File: Comments.pdf

Author Response

We are very thankful to Reviewer for the detailed analysis of our manuscript and very useful thoughts and comments. We performed our best in order to improve our paper according to the Reviewer’s comments and also to outline the symmetrical issues.

In our manuscript we used generally accepted designations of the quercetin molecule, which are broadly used in the literature. Also, we provided Scheme 1 for the readers and we also added reference to it after the phrase “In this work standard numeration of the atoms of the quercetin molecule has been used [16, 17].”. Also, numeration of the conformers is provided one-by-one in the order of increasing of its Gibbs free energy. Calculations of the Gibbs free energies have been performed according to the generally accepted protocol. “Normal conditions” was defined as temperature T=298.1 K and pressure 1 atm. We have removed “default options”. “Non-orthogonal structure” means that dihedral angles, which describe the orientation of the OH hydroxyl groups according the rings, are not equal to 90 degree. Sign “+” mean that it is “Right” enantiomer, while “-” – “Left” enantiomer. The phrase “the initial relatively the terminal” means the difference between the initial and terminal. We have also implemented others corrections into the text.

We are thankful to Reviewer for the valuable comments.

Reviewer 3 Report

This article reports on quantum mechanical geometry computations of transition states (TS) and frequency analyses of the quercetin molecule, a flavoid that has pharmaceutical significance and which, due to a high degree of conformational freedom, may adopt a range of states. The TSs were identified on the potential energy (electronic) surface, calculated with DFT. Gibbs free energy differences between states were subsequently calculated using MP2 theory.

Overall, the work is sound and is a contribution to the field. Nonetheless, the article requires significant refining before being suitable for publication. Below are points that need to be addressed. Some of the grammar/spelling mistakes are pointed out but thorough editing of English language is required. 

--It is not clear to this reviewer how the intrinsic reaction coordinate (IRC) is quantified/determined. The description of the method in "Computational methods" is insufficient. This method needs to be described in more detail and the implementation in the software used should be mentioned.

--As all discussion is based on the structure of the molecule, it would make sense to make Scheme 1 (Figure 1?) of higher quality such that all labels can be read. As it is now, only the rings A, B, and C are clear and the yellow arrows can be seen.

--p2 the authors write that "24 non-planar with Gibbs free energy .." Gibbs free energies are calculated as differences between two thermodynamic states. Do the authors mean that Gibbs free energy differences exist between conformers? This sentence must be rewritten to express that.

--p2 Sentence should read "...information regarding mechanistic details" and not "underground mechanisms"

-- caption of Scheme 1: "designed" should be "designated"

-- p3  replace "in silico scrupulous" with "in-depth in silico"

--p4 please correct the frequency after Eq3 from vi to v_i

--p4 When mentioning standard numeration of atoms please refer to Scheme 1.

--p4 Section should be titled simply "Results and discussion"

--p4 First two sentences of "Results and discussion" should read:

--"The conformational properties of the quercetin molecule associated with the mirror-symmetric turnings ... yield 72 possible TSs. These TSs enable the conformational mobility and give rise to interesting physical-chemical properties."

--p4 Remove "by us" so that sentence reads "Thus all established resulting TSs (C1 point symmetry.."

--The authors refer to "polar" throughout the text but what they mean is "dipole moment", correct? Also, the authors use the phrase "structural, energetic and polar" as "physico-chemical characteristics". More specific would be "geometry, energy and dipole"

--The authors write "Debay" throughout the text and in the tables, but I believe they mean "Debye", the unit of the electric dipole moment that is named after the Dutch scientist.

--Please replace all instances of "analogical" with "analogous"

--Page 6 is virtually unreadable. Please reformat this in a table or Supp Inf.

--p7 Change "We have also revealed" to "The calculations revealed"

--On p7 the authors write "Revealed conformational processes ... 6.5ns" This sentence stands alone and seems not to be related to any subsequent discussion. The next lifetimes are given in "s" but it may make more sense to use the "ps" or "fs" notation. What does "quick" mean in this context? Is 6.5ns quick? "Quick" only makes sense when compared to another time, which the authors do not discuss.

--Top of p7 should read "relative to the A ring"

--On p7 the authors write "quite low" and "quite easily". What is "low" and what is "easily"? These comparative terms are meaningless unless put in comparison. In this paragraph, another figure would be helpful to illustrate the "quasi-planar" structures that are referred to.

--p7 replace "..two pathways of interconversion evidences that.." to "two pathways of interconversion indicate that.."

--p8 replace "row" with "series" when discussing Gibbs free energies

--p8 "Conformational dynamics" implies that sampling under thermal conditions (T not=0K) was carried out. This statement is misleading as no molecular dynamics was carried out. Change "conformational dynamics" to "conformational rearrangement"

--p8 What are "prototropic tautomers"? Could the authors please explain this term?

--p8 Please change "Their characteristic property - exotic values.." to "Their characteristic property is the unusual range of angles.." Why is this range unusual? Are these angles more acute than usual? Perhaps the authors could explain what they mean here.

--p8 Change "We are convinced that.." to "Our data regarding the structurally symmetric mechanisms..are important" (not "is")

Author Response

We are very thankful to Reviewer for the detailed analysis of our work and also for the critical comments and remarks aimed at its improving.

IRC calculations have been provided used the standard procedure, implemented in the Gaussian program. These calculations started from the transition state to the forward and reverse directions. We suggest that more technical details would be excessive for the readers, since it could be easily found in the Gaussian manual. We have improved the resolution of the Scheme 1 as far as it was possible. Yes, it means the difference between the Gibbs free energies between the conformers. We have added explanation “ranging from 0.00 to 25.30”. We have changed to “fundamental mechanisms”. We have corrected caption to “designated”. We have changed to “in-depth in silico”. We have corrected. We have added reference. We have changed to “Results and discussion”. Thank you, we have improved according to your wishes. We have corrected. Yes, we have analyzed dipole moment for defining the polar characteristics of the quercetin molecule. Using the phrases "structural, energetic and polar" we mean a set of different properties, which are provided in the Tables and Figures. We have corrected to Debye. Thank you for the etymology of the origin of this word. We have corrected. We did our best to improve it. We have corrected. We have corrected. We have improved. "Quite low" and "quite easily" means relatively the values, generally accepted in the physical chemistry. We have corrected. We have corrected. We have corrected. We have added “prototropic tautomers - a set of isomeric compounds with the same empirical formula and total charge, but different position of the proton”. We have corrected. We have corrected.

Finally, we thank Reviewer for the great improvements and attention to our paper.