3D-Printed Conductive Polymers as Alternative for Bioelectrochemical Systems Electrodes: Abiotic Study and Biotic Start-Up

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Summary: The authors present a study in which 3D printed electrodes are prepared and evaluated for use in bioelectrochemical systems. The authors do a reasonable job of evaluating the effects of electrode geometry and enhancement via graphene oxide electrodeposition. The best performing electrodes were then evaluated following biological activation. The results of the work represent a useful addition to the literature in the field of 3D printed electrodes. There are a few items that should be addressed before the manuscript is published, but overall this reviewer recommends that the manuscript be accepted with minor revision. Specific revisions are detailed below.

 1.     The authors should make clear both in the abstract and in the introduction that the electrodes prepared are composite materials. As it currently written, the text seems to imply that PLA and TPU are conductive polymers, which of course is not accurate.

2.     Figure 2A is unnecessary give the information presented in Figure 2B and 2C. The reviewer recommends moving 2A to the supporting information.

3.     Figure 2C should have a y-axis lable of “Peak Separation/ΔE_p” rather than “Voltage/V”

4.     Figure 6 should be replaced by an analysis of the data (similar to what is shown in figure 2B and 2C of the original manuscript).

5.     In the analysis of Figure 7 (text on page 9 of the manuscript), the authors highlight how the results observed without graphene oxide modification are better than those with graphene oxide deposition. The authors should provide a possible explanation for this result.

6.     Figure 8B should have an x-axis that matches the x-axis in 8A.

7.     In the discussion of the TGA data, the authors discuss visible degradation of the electrode. It would be helpful to include images of these electrodes in the supporting information if possible.

8.     In the experimental section the authors state that they measured several parameters of the medium including dissolved oxygen, conductivity, and pH. These results should be included either in the manuscript or the supporting information. Of these, the pH may be especially helpful for understanding the electrode degradation as it is well known that PLA with undergo saponification in the presence of high pH environments.

Comments on the Quality of English Language

A few typographical errors present, but otherwise the manuscript is easy to understand by an English speaker.

Author Response

First of all, the authors wish to thank the reviewer for his/her valuable time and patience, and for his/her comments that helped to improve the manuscript. Below, we indicate point by point how we changed the manuscript consequently:

 

Reviewer #1:

Summary:  The authors present a study in which 3D printed electrodes are prepared and evaluated for use in bioelectrochemical systems. The authors do a reasonable job of evaluating the effects of electrode geometry and enhancement via graphene oxide electrodeposition. The best performing electrodes were then evaluated following biological activation. The results of the work represent a useful addition to the literature in the field of 3D printed electrodes. There are a few items that should be addressed before the manuscript is published, but overall this reviewer recommends that the manuscript be accepted with minor revision.

We appreciate reviewer’s effort in reviewing the manuscript, we understand the minor revision suggestion and we addressed the following comments.

 

Comment#1: The authors should make clear both in the abstract and in the introduction that the electrodes prepared are composite materials. As it currently written, the text seems to imply that PLA and TPU are conductive polymers, which of course is not accurate.

Reply#1: It is indeed appropriate to use the most accurate terms. Therefore, we thank the reviewer for pointing this out and those terms have been changed in the revised version (Abstract and introduction).

C#2: Figure 2A is unnecessary give the information presented in Figure 2B and 2C. The reviewer recommends moving 2A to the supporting information

C#3:   Figure 2C should have a y-axis label of “Peak Separation/ΔEp” rather than “Voltage/V”

R#2 and R#3: We agree with the reviewer about figure 2. Figure 2A provided redundant and less visual information and has therefore been reassigned to figure S1 in supplementary material. In addition, the label for figure 2 has been modified to be in line with what is shown, including also the new title for the y-axis of figure 2C (now renamed 2B) which clarifies what is shown in figure 2C.

C#4:  Figure 6 should be replaced by an analysis of the data (similar to what is shown in figure 2B and 2C of the original manuscript).

R#4: We appreciate the reviewer's advice and we do understand the opinion. However, in this case we consider that these more visual cyclic voltammetries are clearer to represent what we want to show as there are not only differences in peak separation or current generation but also in capacitance (represented by the separation between oxidation and reduction curves) which is a characteristic difference in GO electrodeposition of carbon-based materials. Therefore, we think that we should maintain this figure as it is.

 

C#5:  In the analysis of Figure 7 (text on page 9 of the manuscript), the authors highlight how the results observed without graphene oxide modification are better than those with graphene oxide deposition. The authors should provide a possible explanation for this result.

R#5: While it is true that this only happens in one of the electrodes which subsequently does not affect its biological start-up, we agree with the reviewer on the need to explain the result in the characterisation of the electrodeposited electrodes. A possible explanation for the results obtained has been added in lines 223-227.

C#6:   Figure 8B should have an x-axis that matches the x-axis in 8

R#6: Figure 8 has been modified for x-axis fitting. The title of the figure 8 has been highlighted to emphasise this change.

 

C#7:   In the discussion of the TGA data, the authors discuss visible degradation of the electrode. It would be helpful to include images of these electrodes in the supporting information if possible.

R#7: We understand the recommendation to incorporate some photos of the degradation of the electrodes, unfortunately there are no photos of this experimental period.

 

C#8:  In the experimental section the authors state that they measured several parameters of the medium including dissolved oxygen, conductivity, and pH. These results should be included either in the manuscript or the supporting information. Of these, the pH may be especially helpful for understanding the electrode degradation as it is well known that PLA with undergo saponification in the presence of high pH environments.

R#8: We agree with the reviewer and consider that pH measurements can be an interesting parameter to understand the degradation of electrodes, therefore we have incorporated table S1 in the supplementary material. This table shows the parameters explained in the experimental section. A sentence has also been added to lines 246-247, explaining where these results can be found.

Reviewer 2 Report

Comments and Suggestions for Authors

The review report is in the attached file.

Comments for author File: Comments.pdf

Author Response

First of all, the authors wish to thank the reviewer for his/her valuable time and patience, and for his/her comments that helped to improve the manuscript. Below, we indicate point by point how we addressed the comments accordingly:

 

Reviewer #2:

Summary: The manuscript examines the use of 3D-printed conductive polymers, specifically PLA (polylactic acid) and TPU (thermoplastic polyurethane), as electrodes for bioelectrochemical systems (BES). BES combine electrochemical and biological processes, with applications in energy production and wastewater treatment. The research indicates that TPU electrodes have high charge transfer resistance, rendering them unsuitable for bioelectrodes. In contrast, PLA electrodes, particularly those modified with graphene oxide deposition, exhibited better electrochemical performance and generated higher currents during biological tests, suggesting good development of electroactive biofilms. However, PLA electrodes began to degrade after 56 days of operation, limiting their long-term use.

We appreciate reviewer’s effort in reviewing the manuscript and agree with the main summarized ideas

 

Comment#1: The manuscript presents an intriguing study on bioelectrodes. However, certain parts lack the necessary experiments for calibrating and comparing the bioelectrodes made with PLA. In lines 79 and 80, the authors state, “Subsequently, cyclic voltammetry was used for the electrodeposition (ED) from 0.8 V to -1.5 V, at a rate of 20 mV s-1 with 16 repetitions.” Later, in line 209, they mention, “The graphene electrodeposition led to a significant improvement in the performance of the electrodes for the 3 selected depths (Figure 6), corroborating previous studies with graphene oxide that showed an improvement in the electrochemical performance of the electrode.” This study indicates that electrodeposition with GO is crucial for the conductivity of PLA, so the amount of GO electrodeposited must be standardized and adjusted for optimal electrode performance. However, the authors do not present any experiments demonstrating that the parameters set for electrodeposition by CV ensure the optimal amount of GO. The authors should conduct electrodepositions with varying speeds and cycle counts to determine the optimal electrode performance. Otherwise, it may appear that the variable selection is arbitrary (0.8 V to -1.5 V, at a rate of 20 mV s-1 with 16 repetitions). In citations 24 and 30, the authors justify the importance of electrodeposition with GO, but these studies do not involve PLA, so there is no reference for the optimal amount of GO to deposit. The authors need to adjust this parameter, and if the adjustment experiments have already been conducted, they should include the various CV parameters in the supplementary material to ensure reproducibility for other researchers looking to improve this innovative technique.

Reply#1: We appreciate the comment on the electrodeposition of PLA electrodes and agree with the need of deposition optimization. The protocol used for electrodeposition is well implemented in our group and we have seen its usefulness for different carbon surfaces (some are experimental studies without published results), so we decided to see if this same standardised protocol was useful in the case of PLA. However, we agree that an optimisation of the process would be required to ensure the optimum amount and correct distribution of the graphene oxide over the entire surface of the material. In our opinion this study is a starting point and we consider the optimization experiment as a very interesting future work that will hopefully be published in the future.

 

C#2: In lines 85 and 86, the authors state, “The electrode dimensions were 3 cm long x 2 cm wide x 3 mm thick. It was decided to use 2 mm isosceles triangles (Figure 1) as the pattern as a compromise between optimization of the electroactive surface and fluid shear response.” However, the authors do not explain why these specific dimensions were chosen for the electrode or why 2 mm triangles were selected as the pattern instead of another potentially more efficient geometric shape. This clarification is important because one of the advantages of this technique is its versatility in creating custom 3D-printed electrodes. As mentioned in the previous paragraph, the authors should optimize both the size and shape of the patterns to achieve the optimal performance of the electrode. If these experiments have already been conducted, the authors should include them in the supplementary material to ensure reproducibility for other researchers.

R#2:   We are grateful to receive your comment and to be able to specify the choice of dimensions. The dimensions selected were the smallest dimensions allowed by the precision of the printer used. We found them sufficient for a representative micro-scale pattern experiment and then we used electrodeposition for the modification of the surface roughness. As stated in the previous comment, we consider this study a starting point for the evaluation of 3D printed materials in BES, and we find very interesting the possibility of optimizing the pattern and its size. We deeply consider that option as a future work to continue our investigations.

 

C#3: In lines 157 and 158, the authors state, "In any case, the mean value of ΔEp is approximately 1.3 V, which is similar to the values reported for traditional carbon felt electrodes." While the authors compare the peak separation results of the PLA-GO electrode with a carbon felt electrode, they do not include a bioelectrode as a benchmark in the rest of the experiments, such as CV, thermogravimetry, and electrochemical impedance. The traditional carbon felt electrode should be used as a comparison standard to determine if the PLA-GO electrodes offer any comparative advantages under the same conditions and parameters. A similar issue arises in the biological characterization. The authors should conduct the same experiments under the same conditions and variables using a comparison electrode (either carbon felt or another with better performance). If such comparisons were made, the results should be included in the manuscript's graphs to clearly show the differences between the comparison electrode and the PLA-GO electrode. While the manuscript demonstrates an innovative technique in the creation of a PLA-GO bioelectrode, the absence of the experiments mentioned in this review report makes it impossible to determine the advantages and disadvantages of the PLA-GO bioelectrode compared to existing bioelectrodes under the same experimental conditions. Therefore, the authors must provide the requested data.

R#3:   We agree with the reviewer on the importance of comparison with other more widely used materials in order to fully understand the extent of the advantages and disadvantages of the technical-scientific innovation presented in this paper. For this reason, the quoted sentence appears referenced to a paper by this same group in which work was carried out with different carbon-based electrodes. The purpose of this paper is to make a comparison of different conductive composites among themselves, as well as to check the viability of the pattern design, so we focused on the use of different depths of the pattern, which together with the need to use a relatively small volume (500 mL) to have a controlled experimental situation, made it unfeasible to add one more electrode. We understand and share the desire to obtain comparative results between other types of electrodes and the electrodes proposed here, and once we are aware of the opportunities and weaknesses offered by the latter, as well as their optimal performance range (both in design and operation), we believe that future work should be carried out in the future. Moreover, ΔEp is a parameter that can be easily compared between electrodes as it is for example independent of the size, which is a difficult characteristic to standardize among different 3D materials such as carbon felt and our 3D printed electrodes.

 

Comment#4: In various sections of the manuscript, the authors use abbreviations without first providing their full meanings. For instance, the abbreviations "PLA" and "TPU" are included in the abstract, but their meanings are only explained in lines 51 and 59, respectively. Additionally, "graphene oxide" is mentioned in line 76 without introducing its abbreviation, "GO," which is subsequently used from line 115 onwards to refer to the compound. It is recommended that the authors review all abbreviations and their corresponding meanings to ensure they are presented in the correct order, making it easier for the reader to follow.

Reply#4: We thank you for your comment on the abbreviations and they have been revised along the whole manuscript.

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

No comments