Methacryloyl-GlcNAc Derivatives Copolymerized with Dimethacrylamide as a Novel Antibacterial and Biocompatible Coating

Round 1

Reviewer 1 Report

Manuscript ID: Pharmaceutics-1347891

Title: Methacryloyl-GlcNAc derivatives copolymerized with dimethacrylamide as a novel antibacterial and biocompatible coating

Overview and comments:

The authors present the synthesis of functionalized PDMAm hydrogel networks, suitable for polymer surface coating via UV-induced C-H insertion reaction, which results in the formation of coatings via benzophenone crosslinker chemistry. The manuscript presents evaluation of biocompatibility (cytotoxicity & adhesion) and antimicrobial response of the proposed polymeric material.

Comments:

It is a well-written, and organized manuscript; However, the experimental design presents significant flaws. Thus, the data collected is not sufficient (in some cases not appropriate) to support the conclusions presented.

Major comments:

1. The authors claim the novel coating synthesized is a good candidate for implantable medical devices. Can the authors provide additional data that supports the novel coating’s stability (physical, chemical, and mechanical) when treated under standard clinical protocols of sterilization?

2. The manuscript develops its narrative around the idea that the novel coating material presented has the ability to prevent biofilm formation. However, a number of claims are not supported by the microbial physiology as follows:

•  The antibacterial tests presented, speak for the bacteriostatic and bactericidal properties of the material evaluated. Yet, it doesn’t present evidence of its ability to inhibit biofilm formation due to the fact that the tests are run for only 24 hours (Plate testing). To date, there is significant evidence to assume a link between biofilm formation in bacterial cultures and secondary metabolism. The biofilm formation is triggered and regulated by Quorum Sensing (QS), therefore secondary metabolites (produced in the stationary growth phase) must be synthesized prior to triggering the chemical signaling and phenotype modification required, which is commonly reached not before 48 hours of aerated culture.
• Moreover, the antibacterial test (Optical Density), shouldn’t be assumed as a metric to predict biofilm formation. This culture was tested for only 12 hours and the authors clearly stated that they used the exponential phase as the threshold for sampling and further testing which is contradictory to biofilm biology and physiology.

-New data should be collected at 48 & 72 hours.

- Supplementary data is necessary to support these claims (UV-Vis raw data, plate count pictures, microscopic characterization)

• Crystal violet staining is a good approach to preliminary evaluate the biofilm formation. However, it doesn’t differentiate between alive/dead bacteria. Letting alone the previous point on additional growth phases to be evaluated (increased culture time), this test is usually paired with other metabolic batteries & imaging that can provide additional insight

- Could the authors include additional characterization & microscopic analysis (SEM, EPS production [phenol-sulfate method], LIVE/DEAD cell imaging, motility evaluation, etc.)?

Author Response

Dear Editor, Dear Reviewers,

We appreciate the helpful comments of the reviewers which enabled us to further improve the manuscript.
Requested modifications and additions are listed in the attached pdf file, also tagged in the modified version of the manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

• It is not clear for me from the introduction if there are any materials based on  N-acetylglucosamine copolymerized with di- 2 methacrylamide used in this purpose in the literature. Well written introduction but does not provide all the necessary informations, in order to have an overview of the current condition.
• to my understanding the authors intend to use the copolymer as a coating material, and not as a self hydrogel, but no adhesion tests were done;
• what about the morphology of the coating? how does it looks? how do you intend to ensure an uniform coating? what is the average layer size of the coating? did the authors intend to test different concentrations? or different layers, multiple layers? I recommend to have a SEM picture of the tilted and cross-section view in order to observe the morphology.
• did the coating is arranged as brushes? 
• what about the cell culture? how the cells arranged and grow on the coating surface? 

Author Response

Dear Editor, Dear Reviewers,

We appreciate the helpful comments of the reviewers which enabled us to further improve the manuscript.
Requested modifications and additions are listed in the attached pdf file, also tagged in the modified version of the manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The authors addressed all questions from the first review.  No additional comments on the revised version.

Reviewer 2 Report

The authors revised the manuscript properly.