The Optimization of Gel Preparations Using the Active Compounds of Arabica Coffee Ground Nanoparticles
Round 1
Reviewer 1 Report
The paper describes the preparation and optimization of anti-inflammatory gel using Arabica coffee grounds nanoparticles as active compound. The overall design is interesting. But the authors didn't have any anti-inflammatory experiments to validate the results.
Author Response
Dear Reviewer
Thank you very much for your comments and suggestions for our article.
In this article, we limit our discussion to the preparation and optimization of anti-inflammatory gels using Arabica coffee ground nanoparticles as active compounds. For the results of anti-inflammatory experiments we will write in the next article, because this research is advanced and produces a lot of data.
Reviewer 2 Report
The manuscript revealed that the study aims to determine the optimum formulation of anti-inflammatory gel preparations using Arabica coffee grounds nanoparticles as active compounds. Moreover, this manuscript only investigate the optimization of anti-inflammatory gel preparations using active compounds of Arabica Coffee grounds nanoparticles. The preset study did not clear to describe all methods, including how to calculate the percent of solubility, how to confirm the anti-inflammatory effect of Arabica Coffee grounds nanoparticles gel, how to determine the particle size. Authors must be perform. Overall, new experiments must be performed and the results and discussion also rewrite in new manuscript.
Author Response
Dear Reviewer
Thank you very much for your comments and suggestions for our article,
In this article, we limit our discussion to the preparation and optimization of anti-inflammatory gels using Arabica coffee ground nanoparticles as active compounds. For the preparation and characterization of Arabica coffee ground nanoparticles we write in another article as data for research on making nanoparticles from Arabica coffee grounds, PSA results (particle size analysis) and SEM of Arabica coffee ground nanoparticles can be seen in the picture below, but this result is not us show in this article, because we have shown it in other articles. And for the results of anti-inflammatory experiments we will write in the next article, because this research is advanced and generates a lot of data.
Author Response File: 
Author Response.pdf
Reviewer 3 Report
In this work, authors aimed to determine the optimum formulation of anti-inflammatory gel preparations using Arabica coffee grounds nanoparticles as active compounds. In a general way, the work is well designed. Nonetheless, my main concern of the work is related with the characterization of the phenolic profile existing in the nanoparticles. Authors determined the total phenolic contents using spectrophotometric methods, which are subject to several interferences and the results could not correspond to the real ones. In addition, which active compounds are present in the nanoparticles? This is an essential information in this type of work, as well as the concentration of each compound and not the total; since there are some active compounds that depending on the concentration may be toxic. Active compounds should have been characterized and quantified using chromatographic techniques.
Conclusion should highlight the work applicability and relevance.
Minor comments:
- Species name should be presented in italic.
- Page 2, lines 1-5: “Arabica coffee grounds (…) and other active compounds [19], [20]”. Given that flavonoids and chlorogenic acid are phenolics, this sentence needs to be modified.
Author Response
Dear Reviewer
Thank you very much for your comments and suggestions for our article,
Characterization of Arabica coffee grounds using chromatography has been carried out and published in another article entitled "Optimization and characterization of n-hexane extracts from Arabica coffee ground (Coffea arabica L.) from gayo plateau as a source of natural antioxidants" and for GC-MS results it has been we add to the introduction of this article.
In our opinion the conclusion has highlighted the application and relevance of work, namely highlighting the composition of the optimization gel using the response surface methodology.
- We have fixed the name of the species
- Page 2, lines 1-5: we have fixed it “Arabica coffee grounds nanoparticle is one example of the use of nano technology to produce materials used as active substances in pharmaceutical preparations. Arabica coffee grounds contain many bioactive compounds [11] such as alkaloids, flavonoids [12] (phenolics [13] and chlorogenic acid [14]), saponins, karatenoid [15], [16], and other active compounds [17], [18]. Arabica coffee grounds also contain polysaccharides [19] and oil [20]. Nurman et al. [21] GC-MS results from arabica coffee grounds oil showed the presence of compound such as 1,2-benzenedicarboxylic acid, bis (2-etylhexyl) ester (18,09%), methylcyclopentane (14,93%), linoleic acid (9,00%), pentadecylic acid (8,81%), ethyl linoleate (6,36%), 2,3-dimethylbenzofuran (1,61%), cyclohexane (1,36%) and other compounds.”
Reviewer 4 Report
This manuscript reports Arabica coffee (Coffea arabica L.) grounds nanoparticles contain phenolics compounds. The data is extensive. Appears to be valuable to publish in this journal. However, I have couple of suggestions:
How particle size varies with different loading? What is particle size before and after loading Coffee grounds? any visual evidences for these? like SEM, TEM or DLS data?
Author Response
Dear Reviewer
Thank you very much for your comments and suggestions for our article, In this article, we limit our discussion to the preparation and optimization of anti-inflammatory gels using Arabica coffee ground nanoparticles as active compounds. For the preparation and characterization of Arabica coffee ground nanoparticles we write in another article as data for research on making nanoparticles from Arabica coffee grounds, PSA results (particle size analysis) and SEM of Arabica coffee ground nanoparticles can be seen in the picture below, but this result is not us show in this article, because we have shown it in other articles.
Author Response File: Author Response.pdf
Round 2
Reviewer 2 Report
Dear Authors:
Please added the detail method for 2.4. Characterization of gel preparations
Characterization or response (y) in the design of experiments conducted on gel preparations include acidity degree (pH) test, spreadability test, viscosity test and total phenolic test. " The secondary metabolite content of Arabica coffee grounds nanoparticles are alkaloids,
saponins and phenolics with a total phenolic of 1246,897 mgGAE/g." One gram Arabica coffee grounds nanoparticles contained over one gram galic acid, how to calculate the content of galic acid in Arabica coffee grounds? Authors did not perform any antioxidant and anti-inflammation study for elucidating the bioactivity maintenance. I suggest the manuscript must be added the anti-inflammation study of Optimization of Anti-Inflammatory Gel Preparations. As Table 8 shown, the optimum formulation of Arabica coffee grounds nanoparticles gel formulations are carbopol 940 (0.569%), TEA (0.468%) and nanoparticles (3,000%) that produce a response of acidity degree (5.212), spreadability (5.850 cm), viscosity (3734.244 cps) and total phenolic (669.227 mgGAE/g). However, there are no significant difference among these groups. So, how to confirm the group is optimum formulation?
Author Response
Thank you very much, you have provided input and comments on our article and this is very useful for us to improve our article.
Point 1: Please added the detail method for 2.4. Characterization of gel preparations
Characterization or response (y) in the design of experiments conducted on gel preparations include acidity degree (pH) test, spreadability test, viscosity test and total phenolic test.
Point 2:"The secondary metabolite content of Arabica coffee grounds nanoparticles are alkaloids, saponins and phenolics with a total phenolic of 1246,897 mgGAE/g." One gram Arabica coffee grounds nanoparticles contained over one gram galic acid, how to calculate the content of galic acid in Arabica coffee grounds?
Point 3: Authors did not perform any antioxidant and anti-inflammation study for elucidating the bioactivity maintenance. I suggest the manuscript must be added the anti-inflammation study of Optimization of Anti-Inflammatory Gel Preparations.
Point 4: As Table 8 shown, the optimum formulation of Arabica coffee grounds nanoparticles gel formulations are carbopol 940 (0.569%), TEA (0.468%) and nanoparticles (3,000%) that produce a response of acidity degree (5.212), spreadability (5.850 cm), viscosity (3734.244 cps) and total phenolic (669.227 mgGAE/g). However, there are no significant difference among these groups. So, how to confirm the group is optimum formulation?

Response 1:
Thank you for the input, we have added to the article, like the following
Acidity degree (pH) test
Weighing a sample of 10 g, dissolved in aquadest to 100 mL and stirred evenly. Then the pH of the solution is measured using pH meter.
Spreadability test
The gel is weighed as much as 0.5 g then placed on graph paper coated with glass. Then put another glass above the gel mass. The gel diameter was calculated by measuring the diameter length of several sides, then added an additional load of 150g, allowed to stand for 1 minute then measured again the diameter of the gel as before.
Viscosity test
Gel as much as 100 mL is put into a container, then placed on a viscometer and installed spindle no. 64. Then the spindle is lowered into the gel to the specified limit. Next set the speed of 0.6 rpm and see the viscosity value shown on the tool.
Total phenolic test
Gallic acid was weighed as much as 125 mg then 96% ethanol was added up to 25 mL to obtain a 5,000 mg/mL concentrated mother solution. From the main solution 10 mL pipette and then diluted with ethanol 96% to 50 mL volume so that the second mother solution was obtained to concentrate 1,000 mg/mL. The second mother solution was then pipetted 3, 4, 5, and 7 mL, then diluted with 96% ethanol to a volume of 10 mL. The resulting solution has concentrations of 300, 400, 500, and 700 mg/L. Each gallic acid solution of various concentrations pipetted 0.2 mL then added 15.8 mL of distilled water and then added 1 mL of Folin-Ciocalteu reagent and shaken until homogeneous. The solution was allowed to stand for 8 minutes then added 3 mL of 20% Na2CO3 solution and shaken homogeneously. The solution was then incubated for 2 hours at room temperature. Uptake was measured by a UV-Vis spectrophotometer at a wavelength of 725 nm.
A total of 0.3 grams of gel was weighed and then dissolved to 10 mL with 96% ethanol. The solution was pipetted 0.2 mL and added 15.8 mL of distilled water and 1 mL of the Folin-Ciocalteu reagent; homogeneous shaken solution. The solution is allowed to stand for 8 minutes then 3 mL of Na2CO3 20% of the solution is allowed to stand for 2 hours at room temperature. Uptake was measured by a UV-Vis spectrophotometer at a wavelength of 725 nm.
Response 2:
We are Sorry, we were wrong in writing the unit, the correct one is 1246,897 µgGAE/g (in microgram units not in milligrams) and we have corrected it in the article.
To determine the content of gallic acid in Arabica coffee grounds, we used a regression formula produced from the calibration curve Y = 0.0005X + 0.1024 with a value of R2 = 0.9996.
The concentration of gallic acid produced is:
The total phenolic in the sample is read as gallic acid
To determine the total phenolic in Arabica coffee grounds using the following formula:
Response 3:
True, in this article we do not do antioxidants and anti-inflammatory, because in this article we only focus on determining the physicochemical optimization of gel preparations from coffee grouds nanoparticles using the surface response method. For the antioxidant and anti-inflammatory test an optimal gel preparation will be used from the results of this study (this article) also uses the surface response method and we will discuss it in the next article. To emphasize this, can we change the title of this article from "Optimization of Anti-inflammatory Gel Preparations Using Active Compounds of Arabica Coffee Grounds Nanoparticles" to "Optimization of Gel Preparations Using Active Compounds of Arabica Coffee Grounds Nanoparticles"?
Response 4:
True, Table 8 does not show a significant difference for each group, but we chose Solution number 8 was chosen as the prediction of optimum conditions because it has a high w / o interval value (0.994) and high desirability (0.981). Other solutions, although they have the same w / o intervals and desirability values, but produce a lower total phenolic.
Author Response File: Author Response.pdf
Reviewer 3 Report
I read the revised version of the article and I noticed that the authors did not answer to most of the comments. I understand that the authors may have a different opinion; however, they should clarify their point of view and comment the questions.
Total phenolic content was determined using spectrophotometric methods, which are subject to several interferences.
Authors concluded that "Arabica coffee (Coffea arabica L.) grounds nanoparticles can be used as a source of active compounds (...)". Which compounds exist in this nanoparticles? This is a relevant issue in this type of work; since there are some active compounds that depending on the concentration may be toxics.
Chlorogenic acid is a phenolic compound but not a flavonoid. Thus, this sentence "(...) such as alkaloids, flavonoids [12] (phenolics [13] and chlorogenic acid [14]) (...)" is not correct.
Author Response
Thank you very much, you have provided input and comments on our article and this is very useful for us to improve our article.
Point 1: I read the revised version of the article and I noticed that the authors did not answer to most of the comments. I understand that the authors may have a different opinion; however, they should clarify their point of view and comment the questions.
Point 2:Total phenolic content was determined using spectrophotometric methods, which are subject to several interferences.
Point 3:Authors concluded that "Arabica coffee (Coffea arabica L.) grounds nanoparticles can be used as a source of active compounds (...)". Which compounds exist in this nanoparticles? This is a relevant issue in this type of work; since there are some active compounds that depending on the concentration may be toxics.
Point 4:Chlorogenic acid is a phenolic compound but not a flavonoid. Thus, this sentence "(...) such as alkaloids, flavonoids [12] (phenolics [13] and chlorogenic acid [14]) (...)" is not correct.
Response 1:
We're sorry if there are comments or questions that aren't answered
Response 2:
We agree that the spectrophotometric method is subject to several disorders. But in this study we used the spectrophotometric method for the determination of total phenolics, because we wanted to see the total phenolic compounds in gels from arabica coffee grounds nanoparticles and the resulting data was easy for us to insert into the softer Design Expert Version 10.0.3.0 to determine the optimum composition of gel preparations.
Response 3:
Thank you very much for your comment, this comment is very useful for us for the perfection of this research, the active compound here we mean is a phenolic compound. But we will conduct further analysis to see the active compounds and compounds that may be toxic contained in the preparation of arabica coffee nanoparticles gel using chromatographic techniques as suggested in the previous comments and we will discuss in another article.
Response 4:
We fixed the paragraph, it became “(...)such as alkaloids, phenolics [12] (flavonoids [13] and chlorogenic acid [14]) (...)”
Reviewer 4 Report
Authors responded well to my comments.
Author Response
Dear reviewer
Thank you very much for your response to our article
Round 3
Reviewer 2 Report
1. Page 4, Line 13-14. A nano-sized active drug has the advantage of being able to pass through cell walls and membranes to reach the target cell. In addition, a nano-size also ensures high solubility and absorption efficiency. This results can provide antioxidant and anti-inflammatory effects. Author must be performed cellular uptake test for confirming the absorption efficiency.
2. Figure 1 showed particle size of arabica coffee ground nanoparticles is 396 nm, but Figure 2 observed bigger than 10 µm particle size, why?
Author Response
Thank you very much for your comments and input for our article
Point 1: Page 4, Line 13-14. A nano-sized active drug has the advantage of being able to pass through cell walls and membranes to reach the target cell. In addition, a nano-size also ensures high solubility and absorption efficiency. This results can provide antioxidant and anti-inflammatory effects. Author must be performed cellular uptake test for confirming the absorption efficiency.
Response 1: “A nano-sized active drug has the advantage of being able to pass through cell walls and membranes to reach the target cell. In addition, a nano-size also ensures high solubility and absorption efficiency. This results can provide antioxidant and anti-inflammatory effects”. Is a quote from the literature. But we thank you very much, this comment was very useful for our article. Cellular uptake test to ensure the efficiency of absorption of nanoparticles gel arabica coffee grounds we will do and we will report in the next article.
Point 2: Figure 1 showed particle size of arabica coffee ground nanoparticles is 396 nm, but Figure 2 observed bigger than 10 µm particle size, why?
Response 2: Figure 2 shows a larger particle size for nanoparticle samples arabica coffee grounds are easy to agglomerate, so that coagulation makes the particle size becomes larger.
Reviewer 3 Report
I read with interest the second revised version of this manuscript. Authors detailed the experimental section and improved the results section. Thus, in my opinion the work can be accepted for publication.
Author Response
Thank you very much for your response to our article
Round 4
Reviewer 2 Report
No other comments.
