Peer review of: Green light triggered [2+2] cycloaddition of halochromic styrylquinoxaline—controlling photoreac  (2020)

REVIEWER COMMENTS 

Reviewer #1 (Remarks to the Author): 

This manuscript describes a pH tunable cycloaddition reaction driven by low energy photonic input 
(green light 550 nm). It presents studies on a model linear polymer and then extends to a dendrimer 
case that leads to cross linked gels. The mechanics of the gels could be tuned by dendrimer load and 
the optical density could be optimized to allow for penetration of the photostimulus at centimeter 
distances through the macroscale material. Several beneficial properties were noted in the context 
of biomaterials applications, and a proof of principle fibroblast live dead assay was performed. 

A major claim meant to justify a Nature level publication is the lowest energy wavelength for 
cycloaddition under catalyst free conditions. This may very well be, and seems to be an important 
advance. I did not mine the literature to corroborate this, and I trust the authors did their due 
diligence on this point. However, what is needed for this report is to assess comparative 
performance with the known art. For example, what are the penetration depts for the prior catalyst 
free photodimerization gel materials? Is there a marked difference in live-dead outcomes with this 
lower energy excitation vs the higher energy precedents? Are the mechanical properties here 
substantial deviations from past work? Although the aspect of lower energy excitation is important 
for the prospective biomedical applications, if the present work provides no significant suggestion 
for dramatic improvement in functional properties, then the work to me would seem more 
incremental and better suited for a specialized macromolecular or materials journal. 

The work claims the first example of a photochemical ligation that can be directly tuned by switching 
the pH. The authors should better defend this claim through analysis of prior art. Many examples of 
carboxylic acid terminated reactants in similar photochemical reactions are known, for example 
where these molecules are templated via H bonds to a template molecule, and in some cases the 
templation interactions are enhanced by the pH of the solution (for example allowing carboxylates 
to interact with the H-bond donor template more strongly than the latent carboxylic acids). The 
work here seems unique but should be prefaced more specifically with prior art. 

Do they know that the nitrogen ring is doubly protonated? This seems like it would require a much 
stronger acid, not simply pH 2. It seems that the same argument could be made for a singly 
protonated quinoxaline ring leading to the same effect. 



Reviewer #2 (Remarks to the Author): 

This manuscript comprises work on a photo-induced [2+2] cycloaddition of two stilbene-type 
molecules (being integrated into a PEG polymer). The key is using a stilbene unit, which contains a 2-
quinoxaline substituent and a PEG-bound benzene ring (SQ). This allows performing photo reactions 
at wavelengths in the visible region. The authors show that the absorption spectrum of the title 
compound is pH dependent and that the cycloaddition is photo-reversible at 360 nm. This can be 
translated into the photo triggered formation of a gel (using PEG-(SQ)8) with additional pH 
dependence. 
Overall, the findings reported here are innovative and suitable for Nature Comm.; however some 
details should be addressed by the authors: 

a) In Figure 1a, the abs spectra between 450 and 550 nm should be shown at an expanded scale (in 
terms of abs) as an insert. It is intriguing that the absorption spectra of PEG-SQ and that ascribed to 
(PEG-SQ)2 are very much alike. Do the authors have an explanation why the compound with a 
smaller chromophore has almost the same UV/Vis spectrum as the conjugated stilbene derivative or 
is the red spectrum just an overlay between the spectrum of PEG-SQ and one band of (PEG-SQ)2, at 
ca. 240 nm with a sh at ca 310 nm? This, then, would also indicate a rather low conversion (quantum 
efficiency) in terms of the cycloaddition. In this view, the authors should also consider the photo-
induced trans/cis conversion of the 'stilbene moiety'. 
b) It would be desirable if the SEC-determined dimerization efficiency could be connected to the 
photo-chemical observations. 
c) The action plots show a clear but not drastic influence of the solvent, accordingly, the statement 
'that solvent interactions play a critical role' is somehow overstated. 
d) In my view, the same holds for the following section: 'SEC data indicate that at acidic levels (pH 2) 
the reactivity decreases significantly, whereas it is not affected at higher pH values (pH >2) (Figure 
3b). As a consequence, the halochromic properties of the styrylquinoxaline unit appear to translate 
directly into its reactivity, marking it as the first example of a photochemical ligation that can be 
directly tuned by switching the pH.' It is straightforward, that the penetration depth at the 
wavelength of 510 nm used for the corresponding experiments is different since it is connected with 
the corresponding absorbance (at 510 nm), which is shifted by a solvatochromic/halochromic effect. 
It should be clearly stated if this is (just) a consequence of the higher absorptivity/lower penetration 
rather than 'other reactivity'. Experiments at lower concentrations of the system should provide 
some evidence here. 



Reviewer #3 (Remarks to the Author): 

The major claims of the paper are the long-wavelength absorption properties of the stilbene-type 
photoactive systems and the pH-dependency of the photodimerization as well as the formation of 
high-molecular weight products from polymeric substrates that also influences the formation of 
supramolecular structures auch as hydrogels. This is of course a step forward that applies research 
on photocycloadditions of styrenes and stilbenes that has been in the focus of photochemisty 
research over the last 4-5 decades and applications in material sciences of these reactions are 
numerous. The novel aspect here is that one part of the chromophore that allows strong red-shift is 
on the other hand also applicable as a proton-induced switch for photoreactivity, i.e. induces a 
halochromic effect. This is a remarkable and obviously useful effect that allows to switch on and off 
the photoreactivity depending on local pH. 
This effect might be important for cellular experiments where pH differences might be mapped by 
the efficiency of photocycloaddition and thus compartimental properties might become sensible by 
this reaction. This would go obviously beyond the idea of this paper but should be shortly 
elaborated. 

Overall, the single ideas behind the specific photochemistry are not completely novel. But the 
combination of halochromic features and substituent effects that allow red-shift together with the 
molecular weight increase that allows supramolecular aggregation effects looks impressive and 
appears to me as an important feature that will influence the thinking in the fields of classic 
photochemistry and supramolecular photochemistry. 

Concerning the data and the experimental information given in the paper, all results are sound and 
appear very well studied and reported. I recommend publication of this important novel 
combination of several features of photochemistry and supra-molecular chemistry that also 
influences the field of modern material chemisry.