Production of Hydrogel-Based Curcumin-Loaded O/W Suspoemulsions

Round 1

Reviewer 1 Report

Typographical errors need to be corrected throughout the manuscript [e.g. line No. 298].

Possible mechanism by which sedimentation effect are prevented by suspoemulstions needs to be discussed.

Reason for the expectation of similarity of particle size distribution of the bulk crystals in oil and in water needs to be discussed [Line No. 293].

Author Response

Typographical errors need to be corrected throughout the manuscript [e.g. line No. 298].

The English service proofread the paper.

Possible mechanism by which sedimentation effect are prevented by suspoemulstions needs to be discussed.
The sedimentation in the intermediate products, i.e. the oil, is not decisive from our point of view, since the samples always stood for only a short time and were moved regularly. The sedimentation in the final product is of course central, but we have created such a high viscosity with the hydrogel that the drops cannot sediment (see L 363) and we consider the sedimentation in the drop to be not so decisive. In the final application, e.g. on the skin, the droplets should also eliminate any problems that may arise and thus sedimentation should not play a significant role for the bioavailability of the drug.

Reason for the expectation of similarity of particle size distribution of the bulk crystals in oil and in water needs to be discussed [Line No. 293].

Because the solubility in both environments is not particularly high and the samples were processed quickly, we believe that the particle size did not change significantly. Therefore, the saturation concentration is reached quickly and particles do not change further. Furthermore, we measured it like this. In Fig 6 we have plotted both distributions and it can be seen within the measurement tolerance that the distributions are the same.

Reviewer 2 Report

 

Comments to the Authors:

In the submitted work, the Authors investigate the potential of using so-called suspoemulsions with curcumin-based hydrogels for drug delivery. The manuscript presents the results from optical microscopy and laser diffraction for comparing different routes for forming such systems. The manuscript's text is rather clear and understandable; however, its MAJOR REVISION is required due to several questions as follows. 

 

1. In the Abstract, the Authors used several acronyms (BCS, APIs) that are clear for the expert audience. However, such acronyms are then explained in the Introduction. In the Abstract, the acronyms should also be defined for clarity of the text. 

Besides, there is a question of why the "Curcumin" term used in the Abstract starts with a capital letter, but sometimes it does not (in the Introduction, for example). It should be unified. 

2. The Introduction provides a thorough background for the presented study. However, its ending part must be more accurate and improved for clarity. For instance, in lines 81-84, the Authors very briefly communicated that they used hydrogels, but the motivation needs to be included. Later on, the Authors presented extensively what this manuscript is about. I am unsure if this is necessary for this part of the manuscript. 

3. In the Materials and Methods section, the Authors mentioned TPGS as an additive to water suspensions (see lines 123, 150, etc.). What was the role of using this substance in the experiment? It is not mentioned when introduced. 

4. In line 151, the Authors provided the power (180 W) when describing the sonication of the samples. What was this value? Was it an electric power of the device or ultrasonic power (if so, how was it determined?). 

5.Table 1 may need to be clarified for the readers. The clear division into 2% vs. 10% formulations is suggested to be consistent with the description in the manuscript's text (lines 166-167).

6. In line 187, the Authors explained why they chose a 500-nm wavelength for spectroscopy experiments. How did the Authors know the facts described in lines 187-189? Is it possible to add literature reports or recall their experiments? 

7. Equation (1) might be better edited. Please, check the guidelines for the Authors of the Journal to be consistent with the Journal's style (the use of the word: formula instead of equation, etc.). Besides, the symbols used in the equation could be more explicit and explained in the manuscript's text. The use of sub-indices is suggested. 

8. The main objection to the Materials and Methods section is that it needs to be clarified what happened with hydrogels in the suspoemulsion. The formation of hydrogel (and the definition of which part of the prepared complex system is hydrogel-like) needs to be included. Please, consider the improvement of this section to explain the formation of the hydrogel part of the system straightforwardly. The addition of a scheme that defines the experimental steps is also recommended.

9. In line 262, the Authors referred to Figure 1 IV to discuss the results from the laser diffraction analysis for two methods of formation of curcumin crystals. What is the physical mechanism of encapsulation of crystals into the emulsion (hydrogel?) droplets? For instance, in so-called Pickering emulsions, the submicron or micron particles are attached to the interface (i.e., are from outside the droplets). 

10.  What was the reason why Figure 5 cannot be the only figure, as it shows exactly the same data presented before? Please, consider the different presentations of the comparison between methods. 

By the way, the description of the Y axis in Figures 3-5 was not provided in the text. 

11.  The presence of sections 3.2 and 3.3 needs to be clarified. They describe the results not provided in the manuscript (?). The images of suspoemulsions should be added to the manuscript, as discussed in section 3.2. The same should be done regarding the original data and their fitting to obtain the amount of loss curcumin in water. 

 

I understood that this amount evaluated how much curcumin is not inside the emulsion droplet. As stated above, the physical mechanism of such incorporation of crystals inside instead the attachment from outside the droplet should be mentioned in the corresponding literature reports. 

12.  The language of the manuscript is very clear. A few things could be corrected, e.g., lines 291 and 298. These maintain the quality of the scientific effort, but careful proof-reading of the text is suggested. 

 

Author Response

1. In the Abstract, the Authors used several acronyms (BCS, APIs) that are clear for the expert audience. However, such acronyms are then explained in the Introduction. In the Abstract, the acronyms should also be defined for clarity of the text.
   All acronyms are now also explained in the abstract and the text.

Besides, there is a question of why the "Curcumin" term used in the Abstract starts with a capital letter, but sometimes it does not (in the Introduction, for example). It should be unified. 

The writing is now unified for the whole document

2. The Introduction provides a thorough background for the presented study. However, its ending part must be more accurate and improved for clarity. For instance, in lines 81-84, the Authors very briefly communicated that they used hydrogels, but the motivation needs to be included. Later on, the Authors presented extensively what this manuscript is about. I am unsure if this is necessary for this part of the manuscript.
   We have concretized the information in line 86ff
3. In the Materials and Methods section, the Authors mentioned TPGS as an additive to water suspensions (see lines 123, 150, etc.). What was the role of using this substance in the experiment? It is not mentioned when introduced.
   In chapter 2.1 we have revised the tasks of TPGS.
4. In line 151, the Authors provided the power (180 W) when describing the sonication of the samples. What was this value? Was it an electric power of the device or ultrasonic power (if so, how was it determined?).
   The values were not measured, but set on the device. This was worked out more clearly in the text.

5.Table 1 may need to be clarified for the readers. The clear division into 2% vs. 10% formulations is suggested to be consistent with the description in the manuscript's text (lines 166-167).

We have specified the description.

6. In line 187, the Authors explained why they chose a 500-nm wavelength for spectroscopy experiments. How did the Authors know the facts described in lines 187-189? Is it possible to add literature reports or recall their experiments?
   We added the explanation in line 199.
7. Equation (1) might be better edited. Please, check the guidelines for the Authors of the Journal to be consistent with the Journal's style (the use of the word: formulainstead of equation, etc.). Besides, the symbols used in the equation could be more explicit and explained in the manuscript's text. The use of sub-indices is suggested.

 The wording and the equation were edited. The symbols are now explained in the text line 206f

8. The main objection to the Materials and Methods section is that it needs to be clarified what happened with hydrogels in the suspoemulsion. The formation of hydrogel (and the definition of which part of the prepared complex system is hydrogel-like) needs to be included. Please, consider the improvement of this section to explain the formation of the hydrogel part of the system straightforwardly. The addition of a scheme that defines the experimental steps is also recommended.
   This is now explained in more detail in section 2.2.3
   A graphical abstract, as well as a process scheme was added to the paper.
9. In line 262, the Authors referred to Figure 1 IV to discuss the results from the laser diffraction analysis for two methods of formation of curcumin crystals. What is the physical mechanism of encapsulation of crystals into the emulsion (hydrogel?) droplets? For instance, in so-called Pickering emulsions, the submicron or micron particles are attached to the interface (i.e., are from outside the droplets).
   Dear Reviewer, your ideas move us as much as you do, and we would have liked to try to get further clarification here. However, since the particles come from the size to the optical resolution limit of the light microscope, which confirms our laser diffraction, a detection in a multiphase system has not been possible with our means. Thus, we could not determine the EE via the microscope. However, the system is also so complex with the emulsifiers that we cannot make any prediction as to where the particles really are in the end. Our hope is that they remain in the oil phase, but whether a part goes to the interface and how big it is are legitimate questions, which we unfortunately cannot answer at the moment.
10. What was the reason why Figure 5 cannot be the only figure, as it shows exactly the same data presented before? Please, consider the different presentations of the comparison between methods.  By the way, the description of the Y axis in Figures 3-5 was not provided in the text.
    Yes, Fig. 5 has no new data. However, we have deliberately chosen this representation in order to better compare the two processes with each other. The description of the axis was added to the description of the plots.
11. The presence of sections 3.2 and 3.3 needs to be clarified. They describe the results not provided in the manuscript (?). The images of suspoemulsions should be added to the manuscript, as discussed in section 3.2. The same should be done regarding the original data and their fitting to obtain the amount of loss curcumin in water. 

 Chapters 3.2 and 3.3 have been revised. The results have been presented in more detail to improve readability. A picture and the original data have been added in the annex.

 

I understood that this amount evaluated how much curcumin is not inside the emulsion droplet. As stated above, the physical mechanism of such incorporation of crystals inside instead the attachment from outside the droplet should be mentioned in the corresponding literature reports. 

See Remark on 9.

 

12. The language of the manuscript is very clear. A few things could be corrected, e.g., lines 291 and 298. These maintain the quality of the scientific effort, but careful proof-reading of the text is suggested. 

The English service proofread the paper.

Reviewer 3 Report

The study performed by authors “Production of hydrogel-based Curcumin-loaded o/w 2 suspoemulsions”, there are some minor points to be further addressed.

 

 Comments:

1.      Please explain why these specific processes were chosen and how they compare to other potential methods.

2.      It is recommended to compare your study with similar studies and thoroughly explain its strengths and weaknesses.

 

3. How was the particle size of curcumin samples characterized?
4. What are the differences in particle size distribution of curcumin samples produced in water using different processes?
5. How does sonofragmentation affect the particle size distribution of curcumin in Miglyol?
6. What is the preferred process for producing curcumin particles smaller than the emulsion's droplet diameter?
7. What was the best alternative method for determining encapsulation efficiency in suspoemulsions?
8. The results and conclusion section of the manuscript needs to be more concise and presented in a fluent manner.
9. There are a few minor grammatical errors.

Author Response

1. Please explain why these specific processes were chosen and how they compare to other potential methods.
   Added in line 89 based on the work of Schilde et al.
2. It is recommended to compare your study with similar studies and thoroughly explain its strengths and weaknesses.

We compared our studies to similar studies, where they were available:

Production of nanoparticle: L 269 ff

Strengths of the new method: L327

Gel-based Suspoemulsion: However, to produce suspoemulsions in general, no similar papers are available.

1. How was the particle size of curcumin samples characterized?
   We describe the characterization of the particle size distributions in water and oil by laser diffraction in section 2.2.1. This is a standard method. In the final product, we were unable to measure the size so far.
2. What are the differences in the particle size distribution of curcumin samples produced in water using different processes?

Since we have discussed the differences in particle sizes in detail in the paper, we suspect that the reviewer here means the different mechanisms of disruption and stabilization. See Line 222ff and L274ff.

1. How does sonofragmentation affect the particle size distribution of curcumin in Miglyol?
   The ultrasonic process is mainly based on cavitation. Although this cavitation effect is reduced in oil due to the higher viscosity, it can be positively influenced by the solubility of gases in the oil. The mechanism of the particle breakage is consistent with a direct interaction between the shockwaves created by the ultrasound (through acoustic cavitation) and the solid particles in the slurry. 10.1002/chem.201605857
   However, we think that this is going too far in the paper to elaborate. Or do you think this is necessary for the thread?
2. What is the preferred process for producing curcumin particles smaller than the emulsion's droplet diameter?
   The bead milling produces higher amounts of nanoparticles, resulting in a left curve shift. (e.g. line 313)
3. What was the best alternative method for determining encapsulation efficiency in suspoemulsions?
   We proposed different alternative methods to determine the encapsulation efficiency. However, at this point no results were produced with those methods. (see L390) The methods we see today are NMR or Raman (see L457).

 

1. The results and conclusion section of the manuscript needs to be more concise and presented in a fluent manner.

We reworked the results and conclusion chapters. Hopefully it is now more concise.

1. There are a few minor grammatical errors.

The English service proofread the paper.

Reviewer 4 Report

The manuscript develops curcumin suspoemulsion. However, the manuscript is written poorly and cannot be read smoothly. The experiments are not described well. The results cannot demonstrate the formation of curcumin suspoemulsion. The manuscript should not be published, even after revision. More experiments are needed. 

1. How will curcumin suspoemulsion be administered?  by oral or by transdermal? At the beginning, authors talk about bioavailability, but later talk about transdermal. 

2. The experiments about measuring entrapment efficiency failed. There are no other experiments demonstrating the formation of suspoemulsion in which nanocrystals are entrapped inside of oil drops. Particle size data and microscope data cannot demonstrate the formation of suspoemulsion.

3. Hydrogel and emulsion are completely different formulations. How could o/w suspoemulsion be hydrogel-based? 

4. Very poor English writing:  for example, "Besides lipid-based and polymer-based delivery systems, different emulsions and 47 molecular complexes (Anand et al., 2010; Araiza-Calahorra et al., 2018; IM et al., 2012; 48 Nayak et al., 2016); " is an incomplete sentence. 

Author Response

The manuscript develops curcumin suspoemulsion. However, the manuscript is written poorly and cannot be read smoothly. The experiments are not described well. The results cannot demonstrate the formation of curcumin suspoemulsion. The manuscript should not be published, even after revision. More experiments are needed.
The paper is reworked and additional data are added.

1. How will curcumin suspoemulsion be administered?  by oral or by transdermal? At the beginning, authors talk about bioavailability, but later talk about transdermal.
   The main idea behind the suspoemulsion is a (trans)dermal application. However, suspoemulsions may also be administered by other routes. In our paper, we focus on the fundamental production of such suspoemulsions regardless of the route and curcumin is also only a model active ingredient. The administration route, and, therefore detailed formulation is in our opinion, at this point of research not significant. Also, bioavailability is not limited only to oral drug systems since the bioavailability reflects the amount of drug which reaches the targeted tissue.

We address this topic in the introduction in Line 87ff.

2. The experiments about measuring entrapment efficiency failed. There are no other experiments demonstrating the formation of suspoemulsion in which nanocrystals are entrapped inside of oil drops. Particle size data and microscope data cannot demonstrate the formation of suspoemulsion.
   Yes, the paper shows which ways of determining encapsulation efficiencies do not work and why not. Therefore, we consider this to be worthy of publication.

We have clearly shown that we have supersaturated the oil solution, and thus particles exist in the oil. We have further demonstrated that we could significantly change the curcumin particle in particle size. If this oil phase is emulsified into the water phase and the solubility of curcumin in water is close to zero (BSV IV), then the particles can only be in the oil phase or at the interface. However, we could not definitively confirm the final position.

3. Hydrogel and emulsion are completely different formulations. How could o/w suspoemulsion be hydrogel-based?
   The emulsion is hydrogel-based by forming a hydrogel in the continuous aqueous phase. The oil phase is therefore embedded in a gel. This enables us to stabilize the emulsion without the need for another emulsifier. We have included this in the introduction.
4. Very poor English writing:  for example, "Besides lipid-based and polymer-based delivery systems, different emulsions and 47 molecular complexes (Anand et al., 2010; Araiza-Calahorra et al., 2018; IM et al., 2012; 48 Nayak et al., 2016); " is an incomplete sentence.
   Line 47; was corrected. The English service proofread the paper.

Round 2

Reviewer 2 Report

All my concerns have been adressed. When it comes to Q9 and Q11, it would be worth investigating in the future as the presented suspoemulsions are going to be used in future medicine. 

Author Response

Thank you very much!

Yes, we will continue to work on the pharmaceutical formulas and see in which applications it proves useful.

Reviewer 4 Report

I cannot read the revised manuscript. The manuscript went through the major revision in English. The Word track in the revised manuscript really bothers me for a normal reading. The authors should clean-up changes and provide a clean manuscript for reading. Only when they go to the minor revision stage, use Work track. 

Author Response

We have revised the paper and deleted the major revisions. Now only have minor revisions in it.