Statistical Optimization and Purification of Cellulase Enzyme Production from Trichosporon insectorum

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The paper brings the study of cellulases production by a new isolated strain of Trichosporon insectorum. CMCase and FPase activities were evaluated. Although the subject is interesting and scientifically sound, enzymatic activities are not high when comparing to other studies. In fact, it is preliminary work. The main innovative point is the use of the new isolated strain. So, the work does not bring a significant contribution. However, it could be considered for publication after major revision.

Other comments/suggestions are presented below:

-    - Authors indicated that higher temperatures are significant in terms of CMCase and FPAse activities. Does this condition would be interesting for an industrial process?;

-        - -ANOVA tables could be added to Supplementary Material;

-      - Please, compare the obtained cellulases’ activities with recent reported works. The obtained activities are not very high;

-      -   Revise the titles of some graphics of Figure 4. In this case, the carbon source should be FC (Fiber cellulose)?

- - Revise methodology - FPase activity deteremination must be added.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

The English language needs minor revision. 

Author Response

Thank you for your comments 

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript evaluated the production of cellulases by the yeast Trichosporon insectorum, which is in the scope of the journal and worth of investigation. However, the manuscript is not well written, the introduction is too concise and does not give the background of the state of the art and the novelty of the work; methodology lacks of some important references (such as items 2.5.1; 2.5.2; 2.5.3; 2.5.4) and more detailed protocols. Important points are:

·       A strong English revision is necessary, words such as “amidon” and “Gmnopleurus” are used incorrectly and there are grammatical errors;

·       Cell concentrations must be expressed in optical density or g/L, and the inoculum protocol from the storage culture to the experiments is not clear;

·       CMC and pure cellulose are expensive chemicals, why to optimize the enzyme production using these substrates instead of byproducts and agrowastes?

·       What was considered as UI? This is an important information provided in enzyme manuscripts and allow the comparison of your work with others and it is missing in all enzyme activities methods;

·       Equation 1 is not necessary once it is fully known;

·       What is the difference between protein quantification in items 2.5.3 and 2.5.6? Why to perform different methodologies

·       All enzyme activities need references and the definition of IU. In addition, why there is 2 methodologies for CMCase? (Itens 2.2 and 2.5.5) and most important, why not to use the same protocol for CMCase?

·       The branches of each enzyme reactant (CMC, Birchwood xylan, PNPG and PNPx) are missing;

·       Item 3.1 should be in methodology;

·       Most importantly: Before adjust a model when performing a factorial design of experiments, it is necessary to evaluate if that variable has a significative effect on the response variable. In this sense, equations 3, 4, 5 and 6 should have only the coefficients of the significative factors. In addition, the second order coefficients are only presented when the variables interaction is significant. In this sense, according to table 4 and 5 for CMCase only Temperature and temperature^2 should be in the equation once the others are not significant. The same is valid for FPase;

·       In Model validation, the data observed is the same used for setting the model, in this case a new set of experiments is necessary to validate the model;

·       There is no reason to bring the surface methodology to the manuscript if a variable has not significant effect on the enzyme production. Is not possible to call the performed experiments an optimization of cellulase production once it was not defined a optimal condition with maximum cellulase concentration. To optimize this, a strong curvature should be in the model, with a clear red point of maximum in the center of surfaces.

For all this pointed out, I do not recommend the manuscript publication.

Comments on the Quality of English Language

There are gramatical error. A strong english revision is needed.

Author Response

Thank you for your comments

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

See comments in the attached file

Comments for author File: Comments.pdf

Comments on the Quality of English Language

Author Response

Thank you for your comments

Author Response File: Author Response.pdf