Use of Agro-Industrial Waste for Biosurfactant Production: A Comparative Study of Hemicellulosic Liquors from Corncobs and Sunflower Stalks

Round 1

Reviewer 1 Report

The article entitled “Use of agro-industrial waste for biosurfactant production: a comparative study between hemicellulosic liquors from corn-cob and sunflower stalk”.

Current theme, with the premise of the issue of green technologies (use of waste, bioproduct obtained through fermentation), but the solvents used (Hexane, Toluene, Kerosene), not considered ecofriendly.

The theme is current, but already quite explored, becoming clear in the discussion, as results obtained by other researchers are similar, practically only the substrate used changes.

 

Some suggestions:

Only in the material and methods it is verified that it was a submerged fermentation process, it was not mentioned in the abstract or introduction in the form of contexturalization. It would be pertinent to put a paragraph in the methodology describing submerged fermentation, as well as making this clear at the beginning of the work. Even with a schematic, showing all the steps taken.

Line 58. According to (9), you should see the text citation format

Line 68. Here you could clarify that it is a submerged fermentation

Line 108: what is the meaning of MSS

Line 154: “content of hemicellulose (25.3%) and total lignin (34.7%)”, but this methodology is not described in the work. Same thing for “furan derivatives such as hydroxymethylfurfural and furfural” on line 162.

It would be better to join tables 1 and 2, it makes it easier to see

Line 221: ” For this first step, CC and SS liquors were fixed at 20%” what parameters, references were used for this?

 

The results should be better discussed, optimizing the process or even using more “green” processes, ultrasound, microwaves, etc....

Author Response

Many thanks for the suggestions! Changes are in “Track Changes” throughout the manuscript.

The article entitled “Use of agro-industrial waste for biosurfactant production: a comparative study between hemicellulosic liquors from corn-cob and sunflower stalk”.

Current theme, with the premise of the issue of green technologies (use of waste, bioproduct obtained through fermentation), but the solvents used (Hexane, Toluene, Kerosene), not considered ecofriendly.

The theme is current, but already quite explored, becoming clear in the discussion, as results obtained by other researchers are similar, practically only the substrate used changes.

 The authors are grateful for your comments. Indeed, the solvents used for the emulsification tests are not ecofriendly. They are hydrophobic compounds that help us to analyze the biosurfactant’s emulsifying capacity, a valuable property for bioremediation and biodegradation purposes. Bioremediation and biodegradation techniques emerged to treat/remove hazardous contaminants from our ecosystems, including the petroleum-based ones, that is why is so important to evaluate the emulsification index in hydrophobic substances like hexane, toluene, and kerosene. The methodology was proposed by Cooper and Goldberg (1987) and has been widely used ever since in scientific research.

 

Some suggestions:

Only in the material and methods it is verified that it was a submerged fermentation process, it was not mentioned in the abstract or introduction in the form of contextualization. It would be pertinent to put a paragraph in the methodology describing submerged fermentation, as well as making this clear at the beginning of the work. Even with a schematic, showing all the steps taken.

Thank you. Following your suggestion, we made this specification about submerged fermentation in the manuscript’ sections of ‘abstract’, ‘introduction’, and ‘materials and methods’. At the submission, a graphical abstract had been sent to show the process used to produce biosurfactant in the current study. Now, we updated the image to add the information about submerged fermentation, as recommended.

 

Line 58. According to (9), you should see the text citation format

The authors are sorry for this mistake. The citation was corrected as you can check below.

“According to Makkar and Cameotra [25], agro-industrial wastes with high contents of carbohydrates (…)”

 

Line 68. Here you could clarify that it is a submerged fermentation

Thank you. The authors revised and made new additions across the introduction section to attend to all suggestions the reviewers gave. That is why this information regarding the submerged fermentation was indeed added; however, in the subsequent paragraph. Please, check the new version below.

“Therefore, this work compared the use of corncob, sunflower stalk, and a conventional carbon source, such as glucose, as substrates for biosurfactant production by submerged fermentation.”

 

Line 108: what is the meaning of MSS

MSS means ‘mineral salt solution’. The abbreviation first appearance can be found in the lines 302-303.

 

Line 154: “content of hemicellulose (25.3%) and total lignin (34.7%)”, but this methodology is not described in the work. Same thing for “furan derivatives such as hydroxymethylfurfural and furfural” on line 162.

The authors are deeply sorry for this mistake. To characterize the untreated corncob and sunflower stalk, we followed the same bibliography used to characterize their respective liquors. The text has been revised to solve this misunderstanding, as you can check in the following paragraphs.

“The modified Klason method [40] was used to characterize corncob and CC liquor, while the characterization of the sunflower stalk and SS liquor followed the guidelines of the National Renewable Energy Laboratory [41,42].”

 

It would be better to join tables 1 and 2, it makes it easier to see

Thank you. The authors believe that maintaining this information in two separate tables would be less confusing for readers, who would not need to deal simultaneously with a high volume of data.

 

Line 221: “For this first step, CC and SS liquors were fixed at 20%” what parameters, references were used for this?

This choice was made based on the substrates’ characterization and preliminary tests. The experiments were conducted by a multidisciplinary team and the promising results have been further investigated and written for publication. This is the case of the current manuscript, an original work focused on investigating the potential of new carbon/nutrients sources for biosurfactant production, such as corncob and sunflower stalk.

 

The results should be better discussed, optimizing the process or even using more “green” processes, ultrasound, microwaves, etc....

The authors are grateful for your valuable suggestions. They certainly will be taken into consideration for future research. At this moment, we are also grateful for this opportunity as we worked on an investigation regarding the use of hemicellulosic liquors from corncob and sunflower stalks as substrates for biosurfactant production via Bacillus subtilis in submerged fermentation.

Author Response File: Author Response.docx

Reviewer 2 Report

I think that the paper is too technical for a comparative study with potential industrial/everyday application. In my opinion, some sort of economic and social effects discussion (basic economic analysis/comparison) should be included for a more concrete argumentation. I think it is not sufficient to only point out on the work of others to support economic argumentation.

Author Response

I think that the paper is too technical for a comparative study with potential industrial/everyday application. In my opinion, some sort of economic and social effects discussion (basic economic analysis/comparison) should be included for a more concrete argumentation. I think it is not sufficient to only point out on the work of others to support economic argumentation.

The authors appreciate the reviewer’s suggestions and will certainly consider them in future research when we have more time to work on such an in-depth evaluation. However, the present study is focused on qualitative parameters such as surface tension and emulsification index in different hydrophobic compounds.

Reviewer 3 Report

This article reports about the use of agro-industrial waste for biosurfactant production using Bacillus. The manuscript needs the following changes to improve it further.

Comments:

Line 51: Can you mention names of surfactants used for different applications and microbes which produce these? Discuss the recent reference…Microbial biosurfactant: a new frontier for sustainable agriculture and pharmaceutical industries

Mention names of chemical-based surfactants and their effect on humans and the environment.

Line 53: Biosurfactants are secreted extracellularly…..discuss with examples where biomass has been used for biosurfactant production.

Line 63: Cellulose, hemicellulose, and lignin are the main components of natural lignocellulose….mention the various methods used for biomass pretreatments…discuss in support… Recent developments in pretreatment technologies on lignocellulosic biomass: effect of key parameters, technological improvements, and challenges

Discuss results in detail and compare your results with previous reports.

Improve the conclusion part by adding the future scope of the study.

Why the author has not done any effort to extract and identify the biosurfactant produced in this study?

Author Response

Many thanks for the suggestions! Changes are in “Track Changes” throughout the manuscript.

This article reports about the use of agro-industrial waste for biosurfactant production using Bacillus. The manuscript needs the following changes to improve it further.

Comments:

Line 51: Can you mention names of surfactants used for different applications and new frontier for sustainable agriculture and pharmaceutical industries

Mention names of chemical-based surfactants and their effect on humans and the environment.

Thank you. Your suggestions are valuable and welcome to improve the manuscript quality. Therefore, a new paragraph was added discussing the existent microbial surfactants and their applications in different industrial segments. At this point, the reference specified was also properly cited and referenced. We also indicate the most used chemical-based surfactants and their harmful effects on our ecosystems, as you can read below.

“Microbial surfactants are secreted extracellularly or as a part of the cell membrane. They find promising applications in different industrial sectors, such as cleaning, cosmetics, food, petroleum, environment, agriculture, and pharmaceuticals [11–14]. Evonik Industries, for example, launched a sophorolipid-based biosurfactant produced from Candida bombicola with application in cosmetics, cleaning products, and dishwashing liquids [15]. The French Startup Lipofabrik (Éléphant Vert Group subsidiary) uses Bacillus sp. to produce mycosubtilin and surfactin (lipopeptide biosurfactants) with pharmaceutical applications [16]. Jeneil Biosurfactant Co. also synthesized a rhamnolipid-based biosurfactant, which found a market in enhanced oil recovery and cleaning and oil recovery from storage tanks [17].”                             

Line 53: Biosurfactants are secreted extracellularly… discuss with examples where biomass has been used for biosurfactant production.

The authors followed your suggestions and added five new studies focused on producing biosurfactants from biomass. The following paragraph was inserted in the introduction section.

“Several renewable sources can be used in fermentation processes, such as crop residues, by-products of food processing, and agro-industrial waste [18,19]. This can be observed in the work of Nalini and Parthasarathi [20], who performed a solid-state fermentation using mahua oil cake to produce biosurfactants from Serratia rubidaea SNAU02. Using Pseudomonas aeruginosa M408, Ji et al. [21], produced biosurfactants in media containing olive oil as the sole carbon source. Jadhav et al. [22], used sunflower oil refinery waste as a substrate for biosurfactant production via Starmerella bombicola MTCC1910. Vecino et al. [23], evaluated the utilization of cellulosic sugars extracted from vineyard pruning waste to synthesize biosurfactants with Lactobacillus paracasei A20. Recently, Vieira et al. [24] investigated using pineapple peel juice to produce biosurfactants from a Bacillus subtilis strain. This scenario opens the possibility of overcoming one of the main obstacles in the large-scale production of biosurfactants: high production costs associated with using expensive substrates. According to Makkar and Cameotra [25], agro-industrial wastes with high contents of carbohydrates and lipids are considered very useful in biotechnology processes. These wastes are derived from lignocellulosic biomasses, the most abundant and renewable feedstock available, whose utilization is a promising alternative to solve problems of food shortages, environmental pollution, and energy crises. As the main components of natural lignocellulose, cellulose, hemicellulose, and lignin form a compact structure that requires the application of a pretreatment step to break the crystalline structure – an essential step in bioconversion processes [26,27].”

Line 63: Cellulose, hemicellulose, and lignin are the main components of natural lignocellulose… mention the various methods used for biomass pretreatments…discuss in support… Recent developments in pretreatment technologies on lignocellulosic biomass: effect of key parameters, technological improvements, and challenges

Discuss results in detail and compare your results with previous reports.

Following your suggestions, we added a new paragraph approaching the existent pretreatment technologies and the main factors to be considered to select them. The reference specified was properly cited and referenced. Please, you are welcome to read this new addition.

  “The primary goal of pretreatments is to break the hydrogen bonds and cross-linked hydrophobic interactions between cellulose, hemicellulose, and lignin, disrupting biomass’ crystalline structure [28]. This will increase the accessibility of the polysaccharides to hydrolytic enzymes. The major pretreatment methods are divided into four different approaches, namely: physical (milling, grinding, microwave, ultrasonication, and pyrolysis), physicochemical (steam explosion, carbon dioxide explosion, liquid hot water, and wet oxidation), chemical (alkali, acid, ionic liquids, ozonolysis, and organosolv), and biological (fungi, bacteria, and archaea) [29,30]. According to Wyman [31], there are several criteria for selecting a suitable pretreatment, some of them include the (i) preservation of hemicellulosic fractions; (ii) minimal formation of degradation products, like furfural and hydroxymethylfurfural; (iii) low energy demand; and (iv) low-cost pretreatment catalyst and/or inexpensive catalyst recycle.”

Improve the conclusion part by adding the future scope of the study.

Following your recommendation, we specified that future investigations will focus on evaluating quantitative parameters and characterization techniques to identify and classify the biosurfactants produced from corncob liquor and sunflower stalk liquor (separately and in association with glucose).

      “Future investigations will evaluate quantitative parameters and characterization techniques to identify and classify the obtained bioproducts.”

Why the author has not done any effort to extract and identify the biosurfactant produced in this study?

The authors understand the reviewer query and will certainly take it as suggestion when carrying out future optimization tests focusing on quantitative parameters and characterization analysis.

 

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The authors followed the recommendations and restructured the article for better understanding.