3′-Nitro- and 3′-Aminofluoresceins: Appearance of Previously Missing Dyes
, Iryna V. Omelchenko 2, Svitlana V. Shishkina 2, Andrey O. Doroshenko 3, Kateryna O. Vus 4, Hanna S. Vlasenko 2 and Nikolay O. Mchedlov-Petrossyan 1,*
Round 1
Reviewer 1 Report
Overview and general recommendation:
This manuscript synthesized 3'-nitrofluorescein and 3'-aminofluorescein and explored their spectral properties. In general, there are some obvious typographical errors. Even though the results satisfied the purpose of the experimental designs, the manuscript still needed to be revised carefully to improve the quality, especially the layout, before it could be published. Some comments have been shown blow.
Major Comments:
1. It is not necessary to capitalize all of the letters in the title.
2. The conclusion section is missing in the Abstract section.
3. The last graph of the introduction section should be more exhaustive。
4. The conclusion section is missing.
Minor Comments:
1. In the sentence: “However, to the best of the authors knowledge…” There was a grammatical error on ‘authors’.
2. In the sentence: “the melting point 215 oC of this product was not confirmed by our data.” The writing for the unit of temperature is wrong. Please read through the manuscript and revise it.
3. In the sentence: “…100°C and then for 2 h at 120°C and 60 mmHg, unless otherwise noted.” A blank space is required between the unit of temperature and the number.
4. In the sentence: “…with 40 ml of acetic acid.” The writing for the unit of volume is not correct.
Comments for author File: 
Comments.pdf
It would be better to improve the quality of the English language.
Author Response
please see the attachment
Author Response File: 
Author Response.docx
Reviewer 2 Report
The authors made significant contributions by discovering the synthesis route of 3'-NITRO- and 3'-AMINOFLUORESCEINS and conducting a thorough analysis using NMR, DFT calculation. They aimed to provide readers with in-depth data by comparing the differences between the nitro and amino groups and analyzing the effects of functional group positions. However, the readability is compromised, and there is a lack of discussion on key points. Therefore, it is recommended to make revisions.
1. Why do authors think there is a lack of research or low reliability of references regarding NH2 or NO2 at the 3' position? What are the benefits or expected effects of synthesizing these compounds?
2. Would it be better to place compound 3 above compound 1 in Scheme 2?
3. What does "M" represent in Scheme 2? While 1-M1 and 2-M2 indicate different positions of the NO2 group, what is the relationship for 3-M3?
4. In Scheme 2, is it correct to have one benzene ring missing in route b?
5. Are the pathways from 4 to 5 and from 2 to 5 considered new synthetic methods? Is there a variation in the yields between the two routes? Are there any differences in color, melting point, or other properties?
6. Was Gaussian used for NMR calculations? Please indicate the softwares used in Figure 1, Figure 3, 4, etc.
7. The molar absorptivity difference between 3'-nitrofluorescein and 3'-aminofluorescein is not significant. However, what is the reason for the significant difference in emission?
8. While the positional analysis of Nitro is conducted, is there no analysis regarding Amino?
Author Response
Please see the attachment
Author Response File: Author Response.docx
Reviewer 3 Report
3'-NITRO- AND 3'-AMINOFLUORESCEINS: APPEARANCE OF PREVIOUSLY MISSING DYES
Introduction:
In this manuscript, Shekhovtsov et al. report the preparation of 3'-nitrofluorescein and 3'-aminofluorescein along with their methyl esters derivatives. Experiments required for analyzing these structures were carefully well-done using NMR spectroscopy (1H and 13C) and X-ray analysis for some compounds. Their optical properties were studied by means of the UV-Vis and fluorescence spectroscopies and it was found that the dianionic forms of 3'-nitrofluorescein gave very weak emission as expected from nitroaromatic compound, in contrast, the double-charged anionic form of 3'-aminofluorescein in an organic non-hydrogen donor-solvent, but not in water, exhibits intensive fluorescence, similarly to what was reported for 4'- and 5'-aminofluoresceins. This work will help to extend the library of the nitrofluorescein and aminofluorescein molecules and understand their photophysical properties at the excited state. While I found the subject of the document attractive and rich with a different method of analysis and experiment work along with reassuringly theoretical studies, I must confess that some issues should be addressed before publication.
Major revisions:
Ø Can the authors provide details about the fluorescence quantum yield of 3'-nitrofluorescein and 3'-aminofluorescein compounds.
Ø The spectra of the UV-Visible absorption, excitation (solid line) and fluorescence in Figure 5 should be normalized for more clarity.
Ø For the same figure compound name with the associated spectrum colours could be added inside the graphs, this will offer a better visualization.
Ø In Scheme 1 for the first step, the number 2 next to resorcinol should be deleted
Ø The values of the absorption maxima for the 4'-nitrofluorescein (Table 5) are slightly different from the published value in some other work (J. Phys. Chem. A 2019, 123, 8860−8870) can the authors explain such difference?
Ø A conclusion section should be added at the end of the article.
Ø On page 1 I suggest changing “Scheme 1” to “Figure 1”.
Ø On page 2 the sentence “as result” should be replaced by “as a result”.
Ø On page 2 the sentence “did not exceeded” should be replaced by “did not exceed”.
Ø On page 4 the sentence “induces increase” should be replaced by “induces an increase”.
Ø On page 4 the sentence “thus indicates” should be replaced by “indicating”.
Ø On page 10 the sentence” has low effect to dianion” should be replaced by “has a low effect on dianion”.
Ø Reference 21 should be modified by adding journal title abbreviations.
Ø In reference 27 the journal name should be in italic.
Ø On the supporting information (page 1) the sentence “otained by decomposition” should be replaced by “obtained by decomposition”.
Ø On the supporting information, for some figure’s titles the term “1C” should be replaced by “13C”.
Ø On the supporting information on some pages where 2 figures are present, I suggest adding the terms (top) and (down) to specify the type of the spectra, for example, “Figure S10. 1H (top) and HH COSY (down) NMR spectra of 3'-nitrointermediate, crude (DMSO-d6).”.
Author Response
Please see the attachment
Author Response File: Author Response.docx
Round 2
Reviewer 2 Report
The authors have succeeded in honestly answering and convincing the comments. However, it failed to help readers understand because it was hardly reflected in the text. Therefore, it is requested that the contents of the answer be revised once again so that it is well reflected in the text.
Author Response
According to the recommendation of the reviewer, we added several pieces of text and highlighted them.
Previous improvements are not highlighted now.
Reviewer 3 Report
The manuscript can now be accepted for publication in Colorants.
Author Response
We are thankful to the reviewer for his comments and time.
