Optimized Preparation of Methyl Salicylate Hydrogel and Its Inhibition Effect on Potato Tuber Sprouting
Round 1
Reviewer 1 Report
(1) I suggest you consider changing the title. The paper devotes much more space to the method of preparation and testing the properties of MeSA hydrogels than to the effect of MeSA on potato tuber sprouting.
(2) It should be clearly written whether pure MeSA or MeSA hydrogels were used for fumigation of potato tubers. How many types of hydrogels were used to fumigate the potato tubers
(3) The preparation of the MeSA hydrogel - is this a method the authors developed for this experiment or is the source of this method described in the literature
(4) Explain the term RSM
(5) Describe in more detail the statistical methods used in the development of the study results
(6) In the captions to Figures 1a and b; 2 a-d; 6; 7 a and b, explain what data they show, e.g. mean±SE
(7) Unit notation: should be mg mL-1 not mg/mL
(8) In the summary and abstract, indicate which of the tested MeSA hydrogel formulations best inhibited potato tuber germination.
Author Response
Response to Reviewer’ Comments
Many thanks to the reviewer for all the very professional and enlightening questions and suggestions. We have modified the manuscript accordingly, and the detailed corrections are listed below point by point.
Response to Reviewer 1:
Comment 1: I suggest you consider changing the title. The paper devotes much more space to the method of preparation and testing the properties of MeSA hydrogels than to the effect of MeSA on potato tuber sprouting.
Response: Thank you for your careful review and professional comments on our manuscript. The title has been modified to “Optimized preparation of Methyl salicylate hydrogel and its inhibition effect on potato tuber sprouting”.
Comment 2: It should be clearly written whether pure MeSA or MeSA hydrogels were used for fumigation of potato tubers. How many types of hydrogels were used to fumigate the potato tubers.
Response: Thanks for your suggestion.
In Line 121, the section of “2.2 Inhibition test of potato sprouting”, pure MeSA was used for fumigation of potato tubers.
In Line 210, the section of “2.6 Inhibition of MeSA hydrogel on potato sprouting”, pure MeSA and MeSA hydrogel were all used for fumigation of potato tubers. Taking pure MeSA as the control, three kinds of hydrogels with highest encapsulation efficiency, lowest encapsulation efficiency and highest predicted value in response surface optimization experiment, were used to treat potato tubers.
All the above modifications were showed in the revised MS in Line 123-124 (Page 3) and in Line 210-228 (Page 5).
Comment 3: The preparation of the MeSA hydrogel - is this a method the authors developed for this experiment or is the source of this method described in the literature.
Response: Thanks for your suggestion. The preparation of hydrogel is from the reference of Mokhtari et al. (2017), and it has been improved and optimized. The references cited are supplemented in the revised MS. (Page 3, Line 138)
Detailed literature information is listed as below:
Mokhtari, S.; Jafari, S.M.; Assadpour, E. Development of a nutraceutical nano-delivery system through emulsification/internal gelation of alginate, Food Chem. 2017, 229, 286-295. https://doi.org/10.1016/j.foodchem.2017.02.071.
Comment 4: Explain the term RSM
Response: Thanks for your suggestion. “RSM” means “Response Surface Methodology”, it has been explained in detail in the revised MS (Page 4, Line 157). Use the full name instead of the abbreviation when the term first appears in the paper, and use the abbreviation for subsequent occurrences.
Comment 5: Describe in more detail the statistical methods used in the development of the study results
Response: Thanks for your suggestion. Details of the data analysis have been added in the revised MS. (Page 6, Line 232-241)
“The experimental design and response surface optimization analysis were performed using Design Expert 12.0 (Stat-Ease Inc., USA, licensed to ICAR-CMFR). Based on the preliminary results of the single factor experiments, determine the variables and their ranges. The response surface factors and the coded levels were shown in Table 1. The independent variables (sodium alginate concentration , CaCl2 concentration, core-wall ratio) and response variable (encapsulation efficiency) were subjected to ANOVA and regression analysis to assess the significance of the constructed model.According to the ANOVA analysis results, the significance of linear terms, interaction terms, and quadratic terms can be obtained. The results of variance analysis are shown in Table 3.”
Comment 6: In the captions to Figures 1a and b; 2 a-d; 6; 7 a and b, explain what data they show, e.g. mean±SE
Thanks for your suggestion. In this paper, each value represents mean ± standard deviation of three replicates. All the data shows are labeled on the Figures 1a and b; 2 a-d; 6; 7 a and b.
Figure 1 (Page 6, Line 246); Figure 2 (Page 7, Line 282); Figure 6 (Page 11, Line 384); Figure 7 (Page 12, Line 403).
Comment 7: Unit notation: should be mg mL-1 not mg/mL
Response: Thanks for your suggestion. All unit notations have been checked and revised.
Comment 8: In the summary and abstract, indicate which of the tested MeSA hydrogel formulations best inhibited potato tuber germination.
Response: Thanks for your suggestion. The hydrogel formulation with the best inhibition effect on potato tubers has been added to the summary and abstract.
Abstract (Page 1, Line 21-25): “The optimized formulations of MeSA hydrogel were as follows: 1.9% of sodium alginate, 2.2% of CaCl2, 1.9:1 of core wall ratio and 0.15% of Tween-80. The inhibition effect of MeSA hydrogel was better than that of pure MeSA at 18 days, the sprouitng rate of the MeSA and MeSA hydrogel were 42.50% and 13.33%, and the corresponding sprouitng index of the MeSA and MeSA hydrogel were 8.57% and 2.86%, respectively.”
结论（第12页，第427-434行）：“MeSA水凝胶的优化配方如下：海藻酸钠1.9%，CaCl 2的2.2%，芯壁比的1.9：1和吐温-80的0.15%。储存18 d后，CK组马铃薯块茎的发芽率和发芽指数分别为100.00%和28.57%，用1.0 mL kg-1的纯MesA直接熏蒸后分别为42.50%和8.57%，而0.5 mL kg-1的MesA水凝胶处理效率更高，分别为13.33%和2.86%， 分别（P < 0.05）。结果表明，经过优化制备后，MeSA水凝胶在马铃薯发芽抑制方面比纯MesA更有效，MeSA的使用量可减少一半以上。
Author Response File: 
Author Response.docx
Reviewer 2 Report
The authors have presented a manuscript, which evaluated exploration of Inhibition Effect on potato tuber sprouting. Minor revisions. The manuscript presents interesting data concerning the possibilities of new methods explored for inhibiting potato sprouting, but the document need to be improved. Following, I have included some comments aimed to enhance the paper:
1. I suggest to the authors to add a new section detailing the state of the art. In this section, authors have to describe the relevant related work in which explain.
2. Can the authors include at the end of the introduction, more details of the objectives of their study.
3. This work presents very interesting results and practice to increase the crop of vegetables. I think that the authors can improve the format of results demonstration. The authors can highlight better the importance of the results obtained.
4. Line 367: Conclusions
Consider extending the conclusions and adding a Future works paragraph.
Finally, the topic of this manuscript is interesting, but authors must improve the presentation of their results and discussion.
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Author Response
Response to Reviewer’ Comments
Many thanks to the reviewer for all the very professional and enlightening questions and suggestions. We have modified the manuscript accordingly, and the detailed corrections are listed below point by point.
Comment 1: I suggest to the authors to add a new section detailing the state of the art. In this section, authors have to describe the relevant related work in which explain.
Response: Thank you very much for your advice and comments. They are very helpful to this manuscript and our future works. In the section of Introduction, we add a description of the current applications of hydrogels and potato bud suppressors to help readers increase their understanding of hydrogels and bud suppressors.
The details are as follows:
In Line 77-95 (Page 2), “In recent years, many researchers have used calcium alginate hydrogels to embed essential oils and easily oxidized components. Shin et al. [29] encapsulated volatile and insoluble thyme white essential oil with sulfonated cellulose nanocrystals, and then embedded sodium alginate to form hydrogel beads. Ae. albopictus larvae had the highest mortality rate when treated with SA/PEs hydrogel beads formed by 0.50% CaCl2. In addition, the incorporation of emulsified oils into hydrogels protects sensitive bioactive components, such as ω-3 fatty acids, from chemical degradation [30]. Potiwiput et al. prepared dual-crosslinked Alg/CMC hydrogels using ionic crosslinking and electrostatic interaction for loading drugs tetracycline hydrochloride and silver sulfadiazine [31]. Due to the good anti-bud activity and volatility of essential oils, many researchers have developed anti-bud products in recent years. Ge et al. [32] chosed HP?CD to form inclusion complex with s-(+)-carvone to improve its instability properties and obtain better effect of sprout inhibition, among them, the s-(+)-carvone/HP?CD complex with host-guest ratio was 1:1. The s-(+)-carvone/HP?CD composite treatment can effectively inhibit the potato sprouting, at the storage of 70 d, the sprouting rate is still less than 20%. Arnon-Rips et al. [33] prepared reactive carboxymethyl cellulose films containing coarse emulsions or nanoemulsions of citral were prepared and used as potato packaging. After 28 days of storage, nanoe-mulsified citral carboxymethyl cellulos films inhibited sprouting of 80%, resulting in less weight loss and maintaining the organoleptic properties of potato tubers.”
Detailed literature information is listed below:
Shin, J.; Seo, S.; Park, I.; Hyun, J. Larvicidal composite alginate hydrogel com-bined with a Pickering emulsion of essential oil. Carbohydr. Polym. 2021, 254, 117381. https://doi.org/10.1016/j.carbpol.2020.117381.
Rahiminezhad, Z.; Gahruie, H.H.; Esteghlal, S.; Mesbahi, G.R.; Golmakani, M.; Hosseini, S.M.H. Oxidative stability of linseed oil nano-emulsions filled in calcium alginate hydrogels. LWT - Food Sci. Technol. 2020, 127, 109392. https://doi.org/10.1016/j.lwt.2020.109392.
Potiwiput, S.; Tan, H.; Yuan, G.; Li, S.; Zhou, T.; Li, J.; Jia, Y.; Xiong, D.; Hu, X.; Ling, Z.; Chen, Y. Dual-crosslinked alginate/carboxymethyl chitosan hydrogel contain-ing in situ synthesized calcium phosphate particles for drug delivery application. Mater. Chem. Phys. 2020, 241, 122354. https://doi.org/10.1016/j.matchemphys.2019.122354.
Ge, X.; Huang, Z.; Tian, J.; Xu, R.; Wu, X.; Tian, S. S-(+)-carvone/HPβCD inclu-sion complex: Preparation, characterization and its application as a new sprout suppressant during potato storage. Food Chem. Advances 2022, 1, 100100. https://doi.org/10.1016/j.focha.2022.100100.
Arnon-Rips, H.; Sabag, A.; Tepper-Bamnolker, P.; Chalupovich, D.; Levi-Kalisman, Y.; Eshel, D.; Porat, R.; Poverenov, E. Effective suppression of potato tuber sprouting using polysaccharide-based emulsified films for prolonged release of citral. Food Hydrocoll. 2020, 103, 105644. https://doi.org/10.1016/j.foodhyd.2020.105644.
Comment 2: Can the authors include at the end of the introduction, more details of the objectives of their study.
Response: Thanks for your suggestion. Details of the objectives of our study were supplemented in the end of the introduction.
In Line 95-110 (Page 3): “In this study, we found that MeSA had the inhibition effect on potato tuber sprouting for the first time. However, MeSA is extremely susceptible to evaporation and decomposition, and has higher price than conventional budding suppressors (such as CIPC), which seriously limits its application. MeSA hydrogel was prepared by ionic crosslinking of sodium alginate and calcium chloride in order to improve the utilization rate of MeSA and save the amount of essential oil. Taking the encapsulation efficiency as the response value, the optimum preparation conditions of MeSA hydrogel were obtained by single-factor test and response surface optimization test, thereafter used for the inhibition test of potato sprouting. According to the release of MeSA in hydrogel, the prepared MeSA hydrogel has the advantage of slow release, which has better inhibition effect on sprouting than using pure MeSA to treat potato tubers, and reduces the amount of MeSA at the same time. We aim to provide a new idea for inhibiting potato sprouting during storage and logistics after dormancy release. MeSA is derived from plants, and its safety has been confirmed, therefore, MeSA hydrogel is expected to be widely used as a green and safe potato bud suppressor.”
Comment 3: This work presents very interesting results and practice to increase the crop of vegetables. I think that the authors can improve the format of results demonstration. The authors can highlight better the importance of the results obtained.
Response: Thank you very much for your affirmation of our results. We found that MeSA had an inhibition effect on potato tuber sprouting for the first time, and obtained a product with good effect and low dosage through the optimized preparation of hydrogel dosage form. We added these results to the abstract and conclusion, so that readers could understand our experimental results more quickly and clearly. The focus of our study is to improve the inhibition efficiency on sprouting and save the cost by preparing hydrogels, relevant data have been added to the revised MS:
In Line 21-25 (Page 1): “The optimized formulations of MeSA hydrogel were as follows: 1.9% of sodium alginate, 2.2% of CaCl2, 1.9:1 of core wall ratio and 0.15% of Tween-80. The inhibition effect of MeSA hydrogel was better than that of pure MeSA at 18 days, the sprouitng rate of the MeSA and MeSA hydrogel were 42.50% and 13.33%, and the corresponding sprouitng index were 8.57% and 2.86%, respectively.”
In Line 426-433 (Page 13): “The optimized formulations of MeSA hydrogel were as follows: 1.9% of sodium alginate, 2.2% of CaCl2, 1.9:1 of core wall ratio and 0.15% of Tween-80. After 18 days of storage, the sprouting rate and sprouting index of potato tubers in CK group was 100.00% and 28.57%, and it was 42.50% and 8.57% after direct fumigation with pure MeSA of 1.0 mL kg-1, whereas, MeSA hydrogel treatment of 0.5 mL kg-1 showed the more efficient which was 13.33% and 2.86%, respectively (P < 0.05). The results showed that after the optimized preparation, MeSA hydrogel was more effective than that of pure MeSA in potato sprouting inhibition, and the usage of MeSA can be reduced by more than halved.”
In Line 412-417: “It should be emphasized that the effective dose of hydrogel is 0.5 mL kg-1, which is only half of the previous effective dose of pure MeSA (1.0 mL kg-1). Under the same storage conditions, the inhibition effect on sprouting of hydrogel is significantly better than that of pure MeSA fumigation. After direct fumigation with pure MeSA (1.0 mL kg-1), the sprouting rate and sprouting index of potato tubers at 18 days were 42.50% and 8.57%, respectively.”
Comment 4: Line 367: Conclusions Consider extending the conclusions and adding a Future works paragraph.
Response: Thanks for your suggestion. The conclusion has been extended and new section of “Future works” has been added to the end of the paper.
Conclusion (Line 420-438, Page 12): “In this paper, we first found that MeSA had an inhibitory effect on potato tuber sprouting. MeSA hydrogel prepared by the RSM had an encapsulation efficiency of 83.25% and it released slowly and steadily, the release of MeSA was 0.0085 mg mL-1 on the 7th day, whereas MeSA direct fumigation released sharply on the first day, and MeSA content decreased to 0.0083 mg mL-1 on 4th day and 0.0011 mg mL-1 on 7th day, respectively. The optimized formulations of MeSA hydrogel were as follows: 1.9% of sodium alginate, 2.2% of CaCl2, 1.9:1 of core wall ratio and 0.15% of Tween-80. After 18 days of storage, the sprouting rate and sprouting index of potato tubers in CK group was 100.00% and 28.57%, and it was 42.50% and 8.57% after direct fumigation with pure MeSA of 1.0 mL kg-1, whereas, MeSA hydrogel treatment of 0.5 mL kg-1 showed the more efficient which was 13.33% and 2.86%, respectively (P < 0.05). The results showed that after the optimized preparation, MeSA hydrogel was more effective than that of pure MeSA in potato sprouting inhibition, and the usage of MeSA can be reduced by more than halved. The result suggests a promising technology for essential oils in potato tuber sprouting inhibition.”
Future Work (Line 439-452, Page 13): “MeSA, as a plant-derived essential oil, is non-toxic and harmless to human body, which can be used as a new green and safe potato sprout suppressant for promotion. However, there are still some problems to be solved in the future in theory and applications, such as (1) the specific physiological, biochemical and molecular mechanisms of how hydrogels inhibit potato sprouting need to be further studied; (2) how to reduce or mask the odor of MeSA after use to make consumers accept them; (3) the preparation process of MeSA hydrogel is not easy to operate, it is necessary to improve the operation process on the basis of optimizing the formula for the promotion of MeSA hydrogel sprout suppressant; (4) at present, the application effect of MeSA hydrogel is only carried out in the laboratory. In the next step, appropriate optimization and validation of the test conditions will be carried out, and relevant experiments will be conducted under actual transportation or storage conditions, thus paving the way for the production of green and effective potato sprout suppressant.”
Author Response File: Author Response.docx
Round 2
Reviewer 1 Report
After reviewing the amendments made, I believe that the article can be published.
