Production of Human Milk Fat Substitutes by Lipase-Catalyzed Acidolysis: Immobilization, Synthesis, Molecular Docking and Optimization Studies
, Silvana Mattedi 4, Álvaro S. Lima 1,2, Matheus M. Pereira 5, Carla Tecelão 6 and Suzana Ferreira-Dias 7,*
Round 1
Reviewer 1 Report
Usage of the RSM should be highlighted in the introduction section.
Provide the reasoning behind the usage of Central composite design (CCD) for this study in the materials and methods section.
Provide the alpha value used for the CCD as well as the ANNOVA results acquired for the statistical modelling of the performed experiments in results and discussion section.
Add an additional paragraph highlighting the usefulness of different statistical parameters opted here for the selection of most suitable model for the understudy experiment.
Support the morphological results of the biocatalyst via TEM analysis as well.
The unnecessary capitalization of the words such as nature, food industry should be avoided.
Author Response
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Author Response.pdf
Reviewer 2 Report
The manuscript by Cleide M. F. Soares and coworkers describes production of human milk fat substitutes from tripalmitin catalyzed by immobilized lipase in solvent-free medium. The lipase (BCL) was first immobilized in silica by covalent binding and encapsulation, and the immobilized biocatalysts were characterized by FTIR, TGA and SEM. Molecular docking analysis was also conducted with oleic acid and tripalmitin to evaluate the interactions between the BCL and substrate molecules. The highest acidolysis activity was obtained with BCL immobilized by covalent binding in prepared silica with aprotic ionic liquid. Then, the reaction condition was further optimized by a central composite rotatable design, and oleic acid incorporation of 28 mol% was achieved after 48 h. A number of interesting data and results were obtained in this study. The manuscript and the supporting information in general are well prepared and provide some useful information. I think it is an interesting story for the Catalysts and I recommended acceptance after a minor revision (see below).
1. The reaction formula of BCL against tripalmitin and oleic acid molecules should be given in the manuscript.
2. “Results e discussion” should be corrected as “Results and discussion”.
3. More information about calculation of molar incorporation degree (%) should be given. Such as sample preparation, how to detect the product (HMFS) or substrate.
4. The structure of tripalmitin in Figure S2 is wrong.
Minor editing of English language required.
Author Response
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Author Response File: Author Response.pdf
Reviewer 3 Report
The article «Production of Human Milk Fat Substitutes by Lipase-catalyzed Acidolysis: Immobilization, Synthesis, Molecular Docking and Optimization Studies» is written on a relevant topic and has a high applied value.
However, the manuscript is not without significant shortcomings:
1. Figure 2 and description of FTIR. The Figure does not show the bands of amide I and amide II, which are typical for all proteins, so we doubt the presence of immobilized lipase in the samples and/or the correctness of sample preparation.
In addition, in Section 3.3 it is necessary to indicate the method of sample preparation and processing of spectra - whether normalization was done, with respect to which band; what was the amount and/or concentration of the sample; recorded the spectra of wet or dry powder samples, the number of accumulations, etc. Those, the technique must be described in full, and not limited to two lines. The results of spectroscopy strongly depend on the chosen technique of sample preparation and spectra registration.
2. It is necessary to explain why the authors used the TGA method. To increase the number of methods used in the article? TGA is certainly necessary if new (newly synthesized) polymers are used as carriers. In this case, lipase could not withstand temperatures above 100, and even more so 200 or more degrees. What scientific results the authors wanted to take by heating the immobilized enzyme to such high temperatures? This needs to be explained.
In addition, in Section 3.3 it is necessary to indicate the method of sample preparation.
Similar comments apply to the SEM method. It is necessary to add a description of the sample preparation process (for example, whether there was a deposition, what type), the method for registering microphotographs, in particular, the vacuum mode. The authors should not have combined three such methods as FTIR, TGA and SEM into one Section 3.3. Each of them needs a separate detailed description.
3. Formula 3 is a polynomial model. R2 0.73 and 0.60, which is quite small for polynomial models. Interestingly, the authors themselves write that an R2 value of 0.75 is considered satisfactory, and 0.90 is excellent. In addition, this type of models does not have a physical meaning. Authors need to justify the choice of this particular type of model.
Minor remarks:
1. There is no line numbering, so it is inconvenient to write comments on the text.
2. Section 2, probably «Results and discussion».
3. Page 11, last paragraph, Lines 3 and 4 from the bottom, better to write like this: 7.348 × 0.532 × 17.922 Å and 56 × 58 × 58 Å.
Minor editing of English language required
Author Response
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Author Response File: Author Response.pdf
Round 2
Reviewer 3 Report
The authors took into account most of my comments. In the presented form, the article can be published
