How Surface Properties of Silica Nanoparticles Influence Structural, Microstructural and Biological Properties of Polymer Nanocomposites

Round 1

Reviewer 1 Report

Dear Authors,

I carefully reviewed the manuscript materials-1040786, the topic you afford in this work is very interesting and the application of the material you developed are very important.

I would like to know few additional information before the publication:

Did you do some test to measure the eventual residual solvent in the material?

The melting temperature you measured for the materials (all both filled and pristine) is quite high for this kind of polymer (you mentioned in table 3 melting temperature in the range 348 – 357 °C); how do you explain it? Usually at such temperature an important thermal degradation is noticed for this kind of polymers. It is very important that you clarify this point before the publication of the work.

Best Regards

Author Response

Dear Reviewer,

Thank you for a thorough review of our manuscript. I hope that the answers below and additional information that can be found in the reviewed manuscript will provide a better explanation of our research. We also tried to improve the language of the manuscript, sending it for language correction by a specialized scientific translation agency. All corrections in the text to be changed are highlighted in color.

Ad 1. Did you do some test to measure the eventual residual solvent in the material?

The proposed method of nanocomposites preparation by casting method followed by evaporation of the solvent first at room temperature (6h) and then using a dryer (80^oC/24h) was sufficient to obtain clean materials free of DCM solvent fragments. FTIR-ATR for all of materials: pure foil and nanocomposite foils was done. No changes were observed in their FT-IR spectra (Figure 1, Please see the attachment). Additionally, the thermal analysis (TG) was done, which did not revealed any mass changes below 100^oC which confirmed the absence of any residual solvent in the material (Table 1). Please see the attachment

Figure 1. FTIR spectra for the reference material (PLDLA pellets) and the nanocomposite foils PLDLA/hss-SiO₂ and PLDLA/hss-SiO₂

Table 1. Results of thermal analysis of the tested foil materials: PLDLA and nanocomposite films PLDLA/hss-SiO₂ and PLDLA/hss-SiO₂. Mass loss recorded at 100^oC and temperatures with mass drops of 1% and 90% as well as the temperature of thermal decomposition of the material (T_deg)

Ad 2. The melting temperature you measured for the materials (all both filled and pristine) is quite high for this kind of polymer (you mentioned in table 3 melting temperature in the range 348 – 357 °C); how do you explain it? Usually at such temperature an important thermal degradation is noticed for this kind of polymers. It is very important that you clarify this point before the publication of the work.

We thank the Reviewer for reading the manuscript carefully and finding our error in the description of Table 3. The given temperature marked as T_m is, of course, the temperature of thermal degradation of the material T_deg. We corrected the Table 3 description in the manuscript. Indeed, materials based on PLDLA or more popular ones based on PLA degrade in the range of 340-360^oC as evidenced by numerous works on polylactide and its thermal resistance [1-3].

In the results shown in Table 3, glass transition temperature T_g occurred around 50-60°C for all samples. Melting occurred with endothermic peaks at the temperature T_m around 160-170°C for the nanocomposite materials samples. These results show that the crystallinity of the material was higher when the nano-filler was hss-SiO₂.

The probable cause of this phenomenon is the higher specific surface area (SSA) (BET, shown in Table 2 in the manuscript) of the hss-SiO₂ nanoparticles (almost 10 x higher than  SSA of the lss-SiO₂) and stronger interaction between the polymer chain and the particle resulting in the increased crystallinity.

AFM studies of surface of the nanocomposites indicated lower ability of hss-SiO₂ to agglomerate during the material drying. The effect of agglomeration of lss-SiO₂ particles was visible in AFM images, where local spots of faster solvent evaporation (SiO₂ agglomerates) visible as the surface pores on the nanocomposite were created (Figure 4 in the manuscript).

The presence of the nanoparticles slightly reduced (by 4 to 6^oC) the temperature of thermal decomposition (T_deg). The course of the TG curve showed that both nanocomposite foils decomposed in a single-stage degradation and that the addition of SiO₂ reduced the polymer thermal stability.

Table 3. Thermal properties of the nanocomposite foils and the polymer foil

[1] Thermal analysis of polylactic acid －Crystallinity and heat resistance, Application Brief 2007, 81 (9), 11-14

[2] Middleton JC and Tipton AJ. Synthetic biodegradable polymers as orthopedic devices.  Biomaterials.2000; 21(23), 2335-46

[3] Baraúna G., Coraça-Huber D.C., Aparecida de Rezende Duek E., In vitro degradation of Poly-L-co-D, L-lactic acid membranes, Materials Research. 2013; 16(1), 221-226

Author Response File: Author Response.pdf

Reviewer 2 Report

In this manuscript, the authors studied How surface properties of silica nanoparticles influence structural, microstructural and biological properties of polymer nanocomposites in detail. This manuscript needs some major revision before publication. 

1. Abstract should be more quantitative.
2. The introduction section should be a comparative study with recently PLA-based articles. Please mention the novelty of this work.
3. In line 113 authors need to check carefully. Change from 80oC to 80 ^oC maintain consistency.
4. In line 72, Hap was used for hydroxyapatite, whereas in line 55 HA was used, check it.
5. 3 caption, change from SiO2 to SiO₂. Please maintain consistency.
6. In Figure 3, the author should indicate the peak for clear comparison. Why one new peak appeared at (3700-3750 cm^-1) in fig d, please explain it?
7. Please provide the DSC colling and heating curves. Why the authors mention a decrease in melting temperature was due to the particle size. It could affect the crystal size, need more explanation.
8. The author should provide TGA data of the prepared samples.
9. Please provide contact angle images of the samples.
10. Regarding morphology, indicate the location of SiO₂ by using an arrow mark. Provide improved SEM image quality.
11. Regarding references, please follow the same format according to the journal requirement (Refs 17, 24, 29, 33 48, 49, and 56). Some refs have abbreviations and some have full form, maintain consistency.

Comments for author File: Comments.pdf

Author Response

Dear Reviewer,

Thank you for a thorough review of our manuscript. I hope that the answers below and additional information that can be found in the reviewed manuscript will provide a better explanation of our research. We also tried to improve the language of the manuscript, sending it for language correction by a specialized scientific translation agency. All corrections in the text to be changed are highlighted in color.

Ad 1-2. Abstract should be more quantitative. The introduction section should be a comparative study with recently PLA-based articles. Please mention the novelty of this work.

Thank you for your commnts. The changes were made to the abstract as well as to the introduction section. The changes were highlighted in red in the manuscript.

Ad 3-5. In line 113 authors need to check carefully. Change from 80oC to 80 ^oC maintain consistency. In line 72, Hap was used for hydroxyapatite, whereas in line 55 HA was used, check it. 3 caption, change from SiO2 to SiO₂. Please maintain consistency.

Thank the Reviewer for reading the text carefully and pointing out the editing errors, they were corrected and marked red in the text.

Ad 6. In Figure 3, the author should indicate the peak for clear comparison. Why one new peak appeared at (3700-3750 cm^-1) in fig d, please explain it?

As already mentioned in the manuscript; bands (not peaks) in the 3680-3750 cm^-1 range are corresponding to weak hydrogen bond vibrations. In the literature such vibrations can be observed in the case of inorganic oxides or zeolites and come in our case from OH groups on SiO₂ particles. The 3700–3750 cm^-1 spectral region represents weakly hydrogen-bonded inorganic oxide hydroxyl groups [1-2]. Weak OH bands in the 3680-3750 cm^-1 range can only be captured when using a special powder spectroscopic examination technique such as DRIFTs, other spectroscopic methods are unable to capture such nuances. In this case, it means that the large surface area of hss-SiO₂ (BET, shown in Table 2 in manuscript) adsorbs water from the air very quickly which causes the appearance of chemically active OH groups. Probably, the hydroxyl groups are able to generate the second-row interactions between the polymer chain and the hss-SiO₂ nanoparticle, hence its bioactivity (the zeta potential indicating bioactivity of the hss-SiO₂ after 48 h of the incubation in SBF - Figure 7 in the manuscript) and higher viability and metabolic activity of mesenchymal cells (Figure 8A and 8B). Such properties were not present in the lss-SiO₂ nanoparticle, which results in lower crystallinity comparable to the pristine polymer (PLDLA), as well as lack of bioactivity (no nucleation of apatite on the surface. Figure 6B in the manuscript) and lower biological activity is visible in the vitality and metabolic activity of the mesenchymal cells (Figure 8A and 8B in the manuscript).

[1] Sitarz M.  Handke M. Mozgawa W., Identification of silico-oxygen rings in SiO₂ based on IR spectra. Spectrochimca Acta, Part A, 2000, Volume 56, pp 1819–1823,

[2] Audrey L. Ingram, Tara M. Nickels, Dalia K. Maraoulaite Robert L. White, VT-DRIFTS Investigations of Interactions Between Benzoic Acid and Montmorillonite Clay, Spectroscopy, 2015, Volume 30, Issue 10, pp 32–42

Ad 7. Please provide the DSC colling and heating curves. Why the authors mention a decrease in melting temperature was due to the particle size. It could affect the crystal size, need more explanation.

Thank you for that comment. More detailed results of thermal analysis with TG and DSC (with heating and cooling in different condition i.e. temperature rate) combined with FT Raman and the grazing incidence X-ray diffraction which allow to calculated the size of spherulites and shown the influence of the presence of the nanoparticle on polymer chain reorganization is the subject of the manuscript which is being prepared and we would not like to describe it in more detail here.

In practice, in order to detect differences between the samples, only DSC measurements were performed at the standard conditions - like those in our study. The DSC tests were performed at the same heating rate of 10^o/min in protective atmosphere (helium) (Figure 1, please see the attachment). The results obtained in combination with the spectroscopic analysis suggest, that hydroxyl groups present on the developed surface of hss-SiO₂ nanoparticles improve the interaction with the polymer chain and result in an increase in crystallinity of the PLDLA/hss-SiO₂ nanocomposite. Growing crystallinity (about 40%) and increase of melting temperature of this nanocomposite (up to 168^oC) indicate higher share of the crystalline phase in the matrix - the system must get more energy to be destabilized. This behavior is different from that observed for PLDLA/lss-SiO₂, where lower crystallinity translates into lower melting temperature of the material. Our conclusions about the size of crystallites (spherulites) is based on the knowledge from other studies: observations in the contrast-phase microscope and SEM of the bottom side of the film - the solvent evaporation is difficult to achieve in the long run for chain reorganization and the spherulites formation (Figure 2, please see the attachment).

Figure 1. DSC curves measurement at standard conditions for the PLDLA pellet, PLDLA foils and the nanocomposite foils: PLDLA/hss-SiO₂ and PLDLA/lss-SiO₂

Figure 2. Microscopic images of the bottom side of nanocomposites based on PLDLA with lss-SiO₂ (a) and hss-SiO₂ (b)

Ad 8. The author should provide TGA data of the prepared samples.

We carried out thermal analysis (TGA) tests of the nanocomposites (PLDLA/hss-SiO₂ and PLDLA/lss-SiO₂) and the reference material (pristine polymer film). The aim of the study was to determine the possible presence of residual solvent - DCM. Thermal analysis was done, and we did not observe any mass changes under 100^oC which confirmed, that no residual solvent in the material still exist (Table 1). The only change that was noticed was the faster thermal decomposition of the PLDLA/hss-SiO₂ nanocomposite, probably due to faster thermal degradation of crystalline areas. The lack of significant changes in the position of characteristic temperatures resulted that such results were not included in the manuscript.

Table 1. Results of thermal analysis of tested foil materials: PLDLA and nanocomposite films PLDLA/hss-SiO₂ and PLDLA/hss-SiO₂. Mass loss recorded at 100^oC and temperatures with mass drops of 1% and 90% as well as the temperature of thermal decomposition of the material (T_deg)

Ad 9. Please provide contact angle images of the samples.

Wetting contact angle images were entered into the manuscript in Figure 5

Ad 10. Regarding morphology, indicate the location of SiO₂ by using an arrow mark. Provide improved SEM image quality.

In the Figure 10 in manuscript were shows SEM observations indicating characteristic crystalline forms for the apatite (A) and pure surface without the apatite (B). On the Figure 10A you can see cauliflower forms, whose composition obtained by an average analysis from the EDS micro-area shows that it is apatite with characteristic elements such as P and Ca, while C and O comes from a carbon layer allowing SEM observation (a conductive layer). In the case of Figure 10B only C and O derived from the conductive layer applied on the PLDLA/lss-SiO₂ nanocomposite film can be seen. It is not possible to determine of Si from the surface by EDS average analysis. Probably if we have the agglomerate of this powder and their size will be big enough to be seen without destroying the polymer surface with electron beam – this analysis will be sufficient. Here, a better solution would be to make TEM analysis with a suitable ionic dimming preparation (FIB), which would show the distribution of Si in the matrix. Unfortunately, we are not able to perform such measurements at the present moment.

Ad 11. Regarding references, please follow the same format according to the journal requirement (Refs 17, 24, 29, 33 48, 49, and 56). Some refs have abbreviations and some have full form, maintain consistency.

Thank you for your attention. The changes were made in references, they were corrected and marked red in the text.

Author Response File: Author Response.pdf

Reviewer 3 Report

1. Paragraph: The first line of some natural paragraphs is not indented, such as lines 114, 124, 158, 240, 251, 267, 290, 303, 323 and 439;
2. Pictures: The font of the text in each picture is not uniform, and some text is not clear enough;
3. Conclusion: It is recommended to write the conclusion separately, corresponding to the content of each part of this article;
4. Simulation: There are few introductions to the setting method of the simulation model, please add and perfect it if possible. Besides, if all relevant formulas are not included in the article, please complete them.

Author Response

Dear Reviewer,

Thank you for a thorough review of our manuscript. I hope that the answers below and additional information that can be found in the reviewed manuscript will provide a better explanation of our research. We also tried to improve the language of the manuscript, sending it for language correction by a specialized scientific translation agency. All corrections in the text to be changed are highlighted in color.

Ad 1. Paragraph: The first line of some natural paragraphs is not indented, such as lines 114, 124, 158, 240, 251, 267, 290, 303, 323 and 439;

Thank you for your comment, we corrected the introduction taking into account the suggestions for ticking the listed paragraphs.

Ad 2. Pictures: The font of the text in each picture is not uniform, and some text is not clear enough;

Thank you for your comment, we corrected the font on the figures and their description.

Ad 3. Conclusion: It is recommended to write the conclusion separately, corresponding to the content of each part of this article;

Thank you for your comment, we took into account the suggestions for a separate conclusion for each part of the study, but we also included a general conclusion that was already in the manuscript text.

Ad 4. Simulation: There are few introductions to the setting method of the simulation model, please add and perfect it if possible. Besides, if all relevant formulas are not included in the article, please complete them.

There are no simulation models or simulations methods in our manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Tha authors answered to alla issues outlined in previous review.

The work is now fine

Reviewer 2 Report

The authors have addressed all the major concerns. So I recommended it for publication.