Rice Bran Biorefinery: A Zero-Waste Approach to Bioactive Oil and Biopolymer Production

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors This manuscript presents a well-structured and comprehensive study on the cascade valorization of rice bran through a zero-waste biorefinery approach. The topic is timely, relevant, and well-suited to Sustainability’s scope, combining agricultural waste valorization, bioprocess engineering, and sustainable development perspectives.

The paper is clearly written, methodologically coherent, and scientifically sound, with data supported by figures and supplementary material. The novelty lies in the integration of multiple valorization pathways (RBO, PHBV, lignin–cellulose–hemicellulose) from a single feedstock, under a cohesive circular framework. Nonetheless, some refinements are required to improve clarity, replicability, and critical discussion depth, particularly regarding statistical robustness, experimental design limitations, and industrial scalability considerations:  

1. Experimental Design and Replicates – Explicitly describe in the Methods section the number of replicates and variability sources.
2. Statistical Treatment – Clarify the statistical tests used, number of replicates, and significance thresholds.
3. In Vitro and In Vivo Correlation – Better articulate the relationship between in vitro inhibition and in vivo efficacy.
4. Environmental Relevance and Toxicity – Include a qualitative or quantitative comparison highlighting environmental benefits.
5. Figures and Supplementary Material – Improve figure resolution and ensure captions are self-contained.
6.  Units and Precision – Standardize units for easier comparison.
7.  References – Verify year alignment and add missing DOIs.

 

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Abstract

The summary lacks a clear logical structure that allows the reader to understand the work performed. It should begin with a general idea, followed by the hypothesis and objectives, then the methodology, results, and finally the main conclusions.

Introduction

The introduction needs substantial improvement to ensure a more fluid and coherent reading, clearly indicating the direction and purpose of the study.

• The first paragraphs reiterate general concepts of the circular economy without progressing toward the specific goal of the work.
• The text does not clearly identify the knowledge gap that the analysis aims to address.
• The narrative mixes technical background (plastics, lignin) with global context, but without adequate transitions between them.

Materials and Methods

• It would be helpful to include a schematic diagram of the methodological process developed.
• The author should explain in greater detail the purpose of Equations 1, 2, and 3.
• Statistical methods are mentioned, but their justification or rationale for use is not provided.
• It would be important to indicate whether negative or positive controls were used, as these enhance the scientific validity of the results.
• The composition and conditions of the culture medium are described with excessive detail but without justification for their selection.
• There is no description of reproducibility analyses or cross-validation of the results obtained.

Results

• The connection between the results and their discussion with respect to other studies is difficult to follow. It is recommended to combine results and discussion in a single section for better readability.
• A mass balance should be included.

Conclusions

• The conclusions are too general, repeating ideas without offering practical applications or scalability perspectives.
• There are no specific future research lines proposed.
• Quantitative conclusions should be highlighted more clearly.
• There is inconsistency between the results and some statements for instance, the text claims “significant effectiveness,” yet only low inhibition levels were achieved for some fungi.
• The section lacks a conceptual closure, as it does not explain how the initial hypothesis was validated nor the novel contribution achieved through this research.

Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript presents a comprehensive and timely study on the valorization of rice bran (RB) within a biorefinery framework. The core concept—a cascading process to extract rice bran oil (RBO) for edible coatings, use the defatted residue (d-RB) for polyhydroxybutyrate-valerate (PHBV) production, and finally recover lignin, cellulose, and hemicellulose from the fermentation residue—is innovative and aligns perfectly with circular economy principles. The research is well-structured, and the ambition to achieve near-zero waste is commendable. The in-vivo evaluation of RBO as an antifungal coating is a particularly strong and application-oriented component. However, the manuscript requires significant revisions, primarily concerning the clarity of data presentation, depth of discussion for certain results, and correction of several inconsistencies, before it can be considered for publication.

 

1. The authors explicitly state in the Conclusions (Lines 425-428) that "the experimental design is inadequate, and the data cannot be considered conclusive" due to limited availability and sample size. This is a critical self-acknowledged weakness that severely undermines the claims made throughout the results and discussion sections. The manuscript must be revised to either remove this disclaimer and provide a robust statistical justification for the sample sizes used, demonstrating that the experiments had sufficient power to support the conclusions drawn; Or tone down all definitive conclusions and frame the findings as preliminary or indicative, requiring future validation with a proper experimental design. The current text makes strong claims (e.g., "considerable industrial applicability") that are directly contradicted by this disclaimer.
2. The yield of PHB is reported as 0.37% of the fermented raw broth, 5.66% of wet biomass, and 30.73% of dry biomass (Lines 289-291). However, the abstract and Figure S5 state a yield of 12.75%. Please clarify this discrepancy. Is 12.75% the yield based on the initial d-RB? A clear calculation pathway from the raw material (RB) to the final PHB product is essential for assessing the process efficiency.
3. The manuscript requires thorough proofreading for English grammar and phrasing. Some sentences are awkwardly constructed (e.g., "Regard rice industry..." in the Introduction should be "Regarding the rice industry..."). Line 10:"The process began with the ethanolic extraction of rice bran oil, yielding 20.58% oil rich in bioactive compounds." -> "The process began with the ethanolic extraction of rice bran oil, which yielded 20.58% (w/w) of oil rich in bioactive compounds." Line 114: "1 g RBO, DMSO (0.5 mL DMSO, 4.5 mL glycerol (density 1.26 g/cm3), and 9.5 mL distilled water." This is grammatically unclear. Rephrase for clarity, e.g., "a solution containing 1 g RBO, 0.5 mL DMSO, 4.5 mL glycerol..., and 9.5 mL distilled water."
4. The pre-inoculum was grown for 7 days. Was the growth (optical density or cell count) monitored to ensure it was in a consistent, optimal growth phase for inoculation?

Author Response

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Author Response File: Author Response.docx

Reviewer 4 Report

Comments and Suggestions for Authors

The authors present a model for a biorefinery using rice bran with a zero-waste approach. The process produces: rice bran oil, the biopolymer PHBV, lignin, cellulose and hemicellulose. 

This article presents a model for biorefinery for agri-food waste, which is a priority of the circular economy.

The innovative of this work involves integrating three stages: extraction, bioconversion and biopolymer separation into a single technological scheme, which is not common in the rice bran literature. 

The methodological part is correct and raises no objections. My only comment is on the profit calculation. What factors were taken into account in material processing? This is not described in any way.

Also, in Table 1, it is not necessary to specify the column with the year of extraction.

Author Response

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Round 2

Reviewer 3 Report

Comments and Suggestions for Authors

I think the authors has revised all comments I mentioned.