Direct Ethanol Production from Lignocellulosic Materials by Mixed Culture of Wood Rot Fungi Schizophyllum commune, Bjerkandera adusta, and Fomitopsis palustris

Round 1

Reviewer 1 Report

The authors should add information regarding the Nitrogen purity.

LOD for HPLC should be provided

Fig 2 - spell checking anAErobic

Please shortly comment on acids generation and consumption during the fermentation.

Author Response

We thank referees for careful reading our manuscript fermentation-447460 and for giving useful comments. According to the reviewer’s comments and questions, we revised the manuscript. We look forward to hearing from you regarding our submission. We would be glad to respond to any further questions and comments that you may have.

 1. The authors should add information regarding the Nitrogen purity.

- The purity of the N2 gas was more than 99.99 vol.%. We have added this information to the text (P.3, L138).

2. LOD for HPLC should be provided.

- The limit of detection for the HPLC we used is almost 0.05g/L. We provided the information.

P.3, L 117-118,
The limit of detection for the HPLC we used is almost 0.05g/L.

3. Fig 2 - spell checking an Aerobic.

- Thank you for pointing it out. We corrected it.

4. Please shortly comment on acids generation and consumption during the fermentation.

- It‘s known that Fomitopsis palustris produces oxalic acid during wood decay. We tried to detect oxalic acid in this study, however it wasn’t found. Therefore, we didn’t describe about the acid in the manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

The manuscript entitled “Direct ethanol production from lignocellulosic materials by mixed culture of wood rot fungi Schizophyllum commune, Bjerkandera adusta, and Fomitopsis palustris” was much improved compared to the original version, and almost all of my previous suggestions have been included into the text or explained. However, in my opinion, there is still missing the comparison of obtained results with other papers (discussion). Please add a proper discussion to the manuscript.

Also please include the references to the method of determination of biomass composition (method number, year etc).

Author Response

We thank referees for careful reading our manuscript fermentation-447460 and for giving useful comments. According to the reviewer’s comments and questions, we revised the manuscript. We look forward to hearing from you regarding our submission. We would be glad to respond to any further questions and comments that you may have.

1. The comparison of obtained results with other papers

- Thank you for pointing it out. We added the description about the comparison with other papers in the manuscript as follows;

P.6, L232~
In the direct ethanol production from cedar wood in the present study, Schizophyllum commune in the CBP added cellulase produced ethanol at 2.49% of the yield on raw material without any processing. That shows a clue for cost reduction of bioethanol production, however, the yield should be improved for application. Several studies have reported ethanol production from biomass using wood rot fungi. Okamoto et al. reported that the white rot fungus Trametes hirsuta applied to produce ethanol for CBP of crushed rice straw gave the yield of 17% on raw material basis [21]. Maehata et al. reported that Flammulina velutipes in CBP with cellulase performed the ethanol yield of 36% from amorphous cellulose prepared from sugarcane bagasse [33]. Our previous study reported that Schizophyllum commune converted microcrystalline cellulose to ethanol at the yield of 25.9% on raw material basis. These studies indicate the importance of saccharification of biomass. An effective, low-cost technology of saccharification should be established for cellulosic bioethanol production.

2. Measuring methods for biomass composition

- We added the numbers of JIS for measuring methods for biomass composition as follows;

P.2, L 75~
Old;
The contents of holocellulose, lignin, ethanol-benzene extractives, and ash in the cedar wood were determined according to the Japanese Industrial Standard, where the percentage were 52.8%, 31.4%, 3.2%, and 0.6%, respectively.

New;
The contents of holocellulose, lignin, ethanol-benzene extractives, and ash in the cedar wood were determined according to the Japanese Industrial Standard (P 8012, P 8008, P 8010, and P 8251), where the percentage were 52.8%, 31.4%, 3.2%, and 0.6%, respectively.

3. Addition of a reference.

- We added a reference paper for discussion.

P. 8, L343.
33. Maehara, T.; Ichinose, H.; Furukawa, T.; Ogasawara, W.; Takabatake, K; Kaneko, S. Ethanol Production from High Cellulose Concentration by the Basidiomycete Fungus Flammulina velutipes. Fungal Biol. 2013, 117, 220–226. https://doi.org/10.1016/j.funbio.2013.02.002

Author Response File: Author Response.pdf