Production of Kojic Acid by Aspergillus niger M4 with Different Concentrations of Yeast Extract as a Nitrogen Source

Round 1

Reviewer 1 Report

Manuscript ID: processes-2412725

The authors reported the optimization of culture medium for the production of kojic acid by the Aspergillus niger M4 strain in a liquid fermentation process. This is an interesting paper, and manuscript is well organized. However, for publication a minor revision is needed. I have just few comments in order to improve the final version of the manuscript before publication, please find them below:

Figures: The superscript letters must be added to show is there a significant difference between the results obtained.

Figure 2: After eight days of incubation, the glucose concentrations begin to increase (which is the explanation for this result).

The interpretation of the results should be expanded (they should be more explained and detailed).

Line 258: 0.5 should be replaced by 0.05 g/L.

Lines 357, 359, and 360: AK should be replaced by KA.

Author Response

Please see the attachment

Author Response File: Author Response.pdf

Reviewer 2 Report

Dear author,

Given the numerous applications of kojic acid, it seems interesting to find different and more economical ways for its production. In the paper, different concentration of yeast extract were tested although it could be interesting to have tested other nitrogen sources more economical. Some consideratation should be considered (listed below), but in general, a deeper discusion of the results is requiered:

-          Line 46. 5-hydroxy- 46 2-hydroxymethyl-γ-pyrone should appers the first time KA is mentioned

-          Line 47. Include the main microorganism that produces KA

-          Line 52-54. Include information about types of substrates uses and references

-          Line 60. For the production of KA but using which microorganism

-          Line 61. Eliminate the space in the formula (NH4)2 NO3

-          Line 63-65. Phrase needs revision

-          Line 64-65. Between 1% and 5%

-          Line 64. Which was the microorganism used to achieve this increment?

-          Line 65-67. Include reference

-          Line 67-69. The phrase is not clear.

-          Line 67-69. What do you mean with remaining?.

-          Line 70-71. Give more information regarding the production of this acid controlling the pH between 3 and 5.

-          Line 70-71. Considering the information included it seems that only A.oryzae can produce this acid. However, in the following phrase you mentioned that A. flavus produces this acid. You must revise the information included in this paragraph.

-          Line 73-75. Do you refer to the composition of the media to grow the microorganisms or to produce the acid. This information is not clear.

-          Line 80. You mention that zinc sulfate is going to be added. However, more information regarding the effect of this compound in the KA production should be included.

-          Line 86-87. English should be revised.

-          Line 87. Proportion of glycerol should be included

-          Line 87-88. Phrase needs revision

-          Line 98. Clarify if the pH was adjusted for each media

-          Line 98. Change to “were conducted in Erlenmeyer..”

-          Line 101. What was monitored at various times?

-          monitoring at various times

-          Line 103. Explain “for a 103 total of 78 reactors at times of up to 552 h”. Considering that you tested 4 different concentration of YE, and you did each experiment in triplicate, you should have 12 reactors. Did you prepare more reactors because you tested several fermentation times?. This information must be clarified.  

-          Line 104. You should explain why you did the study of the pH and why you chose this values of pH.

-          Line 117. Write “Five” in numbers

-          Line 124. Put the formula of Ferric chloride

-          In Analytical determination section indicate if the analysis were performed in triplicate.

-          Line 132. High performance liquid chromatography should be written the first time HPLC appears.

-          Line 140. You should mention to which samples you measure the antioxidant capacity. Why was antioxidant capacity measured?

-          Line 158. After or before the formula you should explain the meaning of Asample, Aref, etc

-          Line 165. What is the “carrying capacity of the system”?

-          Line 172. What do you mean with “with the solution”?

-          Line 176. Phase needs revision.

-          Line 179. I do not undertand why you use the abbreviation SG for new cells. Before, cells were represented with X.

-          Line 233. I do not see an exponential phase in these conditions.

-          Line 233-236. I would change the way to explain the behavior of the biomass. Instead of calculating those percentages, I would recommend to calculate rates and express maximum values to make a better discussion of the figure.

-          Line 245. Remove the word “kinetics”

-          Discussion of the results is missing

-          Line 256. The word “productivity” is not appropriate. You refer to KA production

-          Figure 3. I do not see that the experimental data for the condition of 2.5 g/L yeast extract adjust to the model used. The model used is not appropriate for this condition. I do not know if there is an experimental mistake in sample of 4 days.

-          Line 271-273. The maximum were achieved for 2.5 g/L but for this experiment the model is not appropriate.

-          Line 285 and 286. You do not represent productivity in that figure. The represent the production. Revisa the word in all the text.

-          Figure 3 and 4. I do not understand why you got such a difference in KA production if you are repeating the experiment using 2.5 g/L of yeast extract.

-          Figure 6. The fitting parameter for pH 3.5 was not very high.

-          Figure 7. The confidence limits are very high and the fitting parameter was not very high.

-          Line 324. The caption of figure 7 should be modified. You are not analyzing the product at 120 h. Instead you are measuring the concentration during the whole fermentation.

-          Line 342. Explain EC50

-          Line 340. A better explaination of “Antioxidant capacity by DPPH and ABTS” is needed.

-          Line 355. It is very confusing two different sections giving results of KA. Unification of these section would be more reasonable.

-          Unify the units of KA. In some places are given in g/L and in other mg/L.

-          Line 365. Figure caption of figure 8 should be changed as mentioned in figure 7.

-          It is not clear why the production of KA is different in figure 7 and 8.

-          Line 378. I would change “fermentative kinetics”.

-          Line 384-392. You should make a better discussion. The results obtained should be better related to those published

-          Line 399. I do not really understand. If at day 5 there is more KA than at day 6, there must be a kind of consumption of the acid. Did you find this effect in other published papers?

-          Line 403-405. Could you explain this effect?

-          Line 406. What do you mean with “there is an interaction between YE, glucose, and KH2PO4 in conjunction”?

-          Line 449-459. How could you explain the low concentration of KA obtained in the present work compared to those described in the literature?

 

-          Line 463. Remove “with a retention time 463 of 12 min”.

 

 

Author Response

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Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Dear authors, 

After all the modification, the paper has been significantly improved. Some modifications are recommended:

Line 86  and 493. Change to Zn2+

Line 268. What do you mean with " the growth process is stationary"?

Figure 3. The data do not fit well to the polynomial trend.

Line 491. What do you mean with "with the addition of ZnSO4 there was productivity"?

Line 492-494. Phrase needs revision

Line 534. Change to zinc

Line 550. Remove comma after A. niger

LIne 570. Change "the use of pH"

LIne 568-572. The phrase needs revision.

Line 568-575. The phrase is extremely long

Line 589-592. The phrase needs revision.

LIne 592-598. The phrase is extremely long

Author Response

Please see the attachment

Author Response File: Author Response.pdf