Anti-Influenza A Virus Activity of Rhododendron brachycarpum Extract and Identification of Hyperoside as the Active Constituent
Round 1
Reviewer 1 Report
Comments and Suggestions for Authors
The manuscript “Anti-Influenza A Virus Activity of Rhododendron brachycarpum Extract and Identification of Hyperoside as the Active Constituent” by Park et al., describes the anti-influenza effects of R. brachycarpum and identifies hyperoside as the active compound. The authors tested the antiviral effects against H1N1 and used viral titers, gene and protein expression as readout.
Comment 1:
The arrangement of the results in Figure 1 and the corresponding description do not match.
Comment 2:
Its hard to tell if the IC50 calculation in Figure 1E reliable. Based on the absolute viral titers provided in the manuscript, it looks like effects appeared for 50 and 100 µg/ml. The authors should include additional concentrations in the IC50 calculation, e.g. 25, 30, 35, 40, 45, 50, 55, 60, 70, 80, 90. The authors should also provide the relative viral titers as well as the IC50 calculations.
Comment 3:
As already mentioned in Comment 2, also for Figure 3 D, it is hard to tell if the IC50 value is reliable. The authors should test for additional concentrations and calculate the IC50 of Rb.B again. I furthermore do not understand, why the IC50 evaluation of the Rb.0 in Figure 1 was done in MDCK cells, while the Rb.B evaluation was done in A549 cells. This complicates a direct comparison, as it is done in the discussion section lines 288-291. The IC50 evaluations should both be done in the same cell line.
Comment 4:
The authors should use a HA-pseudotyped system to investigate the effect of Rb.B on the viral internalization. This would make the data shown in Figure 4 even more reliable and would pinpoint Rb.B´s mode of action in the internalization process.
Comment 5:
The discussion generally summarizes the results. A contextualization in the scientific framework is missing. Is hyperoside suitable as an antiviral drug? Is there anything known about antiviral effects against other viruses? Are there similar substances that have been tested in clinical trials?
In addition, the authors did not discuss the limitations of the study: Only one H1N1 strain was tested. Additional subtypes should be tested, such as H5N1, considering the current situation in the US. The study was conducted in cell culture; in vivo or ex vivo data would further substantiate the effects.
Author Response
Please see the attachment.
Author Response File: Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for Authors
Dear authors,
Abstract
IC_50 should be written as IC50.
For the crude extract of Rhododendron brachycarpum and its n-butanol fraction, please provide not only the IC50 values but also CC50 values.
The IC50 and CC50 values of the purified hyperoside should be added to the abstract.
The keyword list should include hemagglutinin.
Introduction
Please include information on other active compounds present in Rhododendron brachycarpum.
In the paragraph discussing quercetin and its activity, please add specific concentration values (IC50, EC50) at which this compound exhibited activity.
When and in which season were the leaves of Rhododendron brachycarpum collected? Does the collection timeaffect the content and yield of hyperoside (quercetin-3-O-galactoside)?
Results
Has the chemical composition of Rhododendron brachycarpum been previously studied, or is this the first reportby the authors? Please provide references.
Can the authors provide molecular docking results of hyperoside with hemagglutinin? I would recommend adding this data.
In line 237, the statement “Flavonoids like quercetin and its glycosides have been reported to exhibit strong anti-influenza activity” should be supported with references.
Throughout the manuscript, IC50 and CC50 should be consistently written as IC50 and CC50.
Apart from the introduction, the reference compound (positive control) is not mentioned anywhere. Its antiviral activity data should be provided and compared to hyperoside in the discussion.
The references 2,3,5,10,16,17, 19, 20,24,25,27 are missing author names and should be formatted according to the journal’s guidelines.
DOIs must be included for all references where available.
The work doesn't contain any new references, the authors should update them where possible
Comments for author File: Comments.pdf
Author Response
Please see the attachment.
Author Response File: Author Response.pdf
Round 2
Reviewer 1 Report
Comments and Suggestions for Authors
I want to thank the authors for addressing my comments. I do not have further comments.
Reviewer 2 Report
Comments and Suggestions for Authors
Thanks to the authors for the revised manuscript. I think that the authors have adequately addressed the comments and most of my concerns. Therefore, I have no further comments. The research paper can be accepted for publication in this journal.