Review Reports - Sustainable Management Practices to Include Mixtures of Chemicals in Regulatory Approaches Focusing on the Global South

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

This study presents the GLORIES framework, proposing an integrated approach to regulate chemical mixtures in Global South watersheds through chemical, biological, and social pathways. Its core innovation lies in introducing Mixture Assessment Factors (MAF) to adjust regulatory thresholds while advocating for stakeholder collaboration through watershed committees and open data sharing.

While conceptually ambitious, the framework's implementation feasibility appears fundamentally constrained by impractical assumptions about data availability in target regions. The most significant limitation stems from the inherent contradiction between the framework's extensive data requirements and the acknowledged reality of severe data scarcity across Global South nations (Quang et al., 2021). The proposal to build watershed-specific chemical inventories and regionalized SSDs presupposes comprehensive monitoring infrastructure and corporate transparency—conditions explicitly contradicted by the authors' own admission that "monitoring data in the Global South means that using the same models applied to developed countries [...] might lack robustness" and that industries demonstrate "unwillingness to cooperate" with data sharing. This foundational gap renders the database-driven components (AOP modeling, SSD regionalization) theoretically sound but operationally unattainable in the intended contexts.

Technical execution barriers further compound these concerns. The framework's reliance on Adverse Outcome Pathways demands species-specific molecular mechanism data—a requirement fundamentally at odds with the "local species toxicity mechanism research gaps" acknowledged in Section 3.1.5. Similarly, constructing credible SSDs requires toxicity data for ≥10 species (Sala et al., 2012), yet tropical endemic species frequently lack even baseline toxicological studies. The MAF methodology introduces additional uncertainty by failing to address error margins in synergistic indices (e.g., the cited 3.29 factor for diclofenac/sulfamethoxazole), creating risks of regulatory overreach or under-protection without probabilistic uncertainty analysis. Crucially, the exclusion of non-chemical stressors like climate impacts (Section 3.1.3) ignores critical environmental interactions prevalent in Global South watersheds.

To enhance practical viability, the authors should:

Develop phased implementation protocols prioritizing data-rich pilot basins before scaling to resource-limited areas；Integrate proxy methodologies for SSD construction using read-across approaches for taxonomically similar species；Establish clear MAF uncertainty bands based on probabilistic mixture risk modeling；Produce evidence-driven incentives for corporate data sharing (e.g., regulatory flexibility for participating industries)；Include pilot validation studies demonstrating framework applicability in at least one Global South context.

Author Response

Dear reviewer,

We thank you for your thoughtful review and feedback. The alterations are listed below each topic, and the number of lines was added to better indicate where the alterations were performed, as well as edited with “track changes” on.

We hope to have clarified your main issues.

Sincerely,

To enhance practical viability, the authors should:

Develop phased implementation protocols prioritizing data-rich pilot basins before scaling to resource-limited areas;

The text was altered all throughout the paper with track changes on to enhance clarity.

Integrate proxy methodologies for SSD construction using read-across approaches for taxonomically similar species;

Thank you for your suggestions. The text was added on L381.

Establish clear MAF uncertainty bands based on probabilistic mixture risk modeling；

The idea of including bands for the MAF became big enough that it became a different paper. In this other paper a method for the calculation of a MAF was developed and should be published as soon as we finish writing it.

Include pilot validation studies demonstrating framework applicability in at least one Global South context.

Due to the novelty of the paper, the validation of the framework is out of scope for this paper.

Reviewer 2 Report

Comments and Suggestions for Authors

Please see comments in the attached file.

Comments for author File: Comments.pdf

Author Response

Dear reviewer,

We hope to have clarified your main issues.

The alterations in the figure were not performed due to time constraints.

Sincerely,

In the abstract

Clearly state the research gap (lack of mixture-focused regulations in the Global South).
- Added on line 4
Highlight the novelty (integration of civil society and data-sharing principles into regulatory practices).
- Added on line 9
End with a clear statement of expected impact (how the framework can improve regulatory effectiveness and environmental protection).
- Added as the last sentence of the abstract.

Introduction:

Streamline the narrative to avoid redundancy (similar points about complexity and data scarcity are made in several paragraphs).
- Altered throughout L100 and L113
The citation style has inconsistencies (e.g., “K.ORTENKAMP” typo; inconsistent formatting of dates and initials). Ensure consistent citation formatting and correct typographical errors.
- Corrected throughout the text.
Place greater emphasis on why current models from the Global North are insufficient for developing countries.
- Corrected in L103 and L107

In the methodology

Provide examples of how the evaluation criteria were applied (e.g., “X papers were excluded for focusing on air/soil quality”).

Strengthen the link between the literature review results and the creation of the GlORIES framework.
- Added on L161.

Results and Discussion

Figure 1 is too simplistic and does not illustrate the dynamic interactions between routes.
Expand Figure 1 into a more detailed diagram/flowchart showing how chemical, biological, and social data f low into regulatory decision-making.
Is recommended to separate the discussion from the conclusions.

Suggestions in the references:

Carefully revise for formatting consistency (APA/MDPI style).
Check for duplicate or redundant entries.
Ensure that all in-text citations are properly formatted.
- References revised.

Reviewer 3 Report

Comments and Suggestions for Authors

After careful evaluation of the content, structure, and originality, I recommend rejecting this submission.

Major Concerns:

Suspected AI-Generated Content: The manuscript exhibits several hallmarks commonly associated with text produced by AI tools, such as large language models (e.g., GPT variants). These include:

Repetitive and Generic Phrasing: Sentences often repeat ideas with minor variations, such as emphasizing "integration" across stakeholders without substantial depth or novel insights. For example, the principles section (e.g., GlORIES framework) lists concepts like "Open access" and "Regionalization" in a formulaic manner that lacks nuanced argumentation.
Unnatural Language Flow: Some passages feature awkward or overly formal constructions, such as "It is proposed that watershed management committees be established to integrate all stakeholders and promote workshops organized by academia, industry, regulatory agencies, and civil society, leveraging existing structures to conserve energy in the process." This phrasing prioritizes keyword density over clarity and reads as mechanically assembled.
Inconsistencies in Depth: The abstract and introduction discuss complex topics like Adverse Outcome Pathways (AOPs) and Mixture Assessment Factors (MAFs) at a high level but fail to provide specific, evidence-based examples or critical analysis, which is atypical for human-authored academic work.

Author Response

Dear reviewer,

We thank you for your comments and thoughtful review. However, there is no AI generated text in this paper. The unnatural language flow might come from the use of English as a second language from the authors. We do emphasize that no generative AI was used neither for writing the text or structuring the research.

The number of lines was added to better indicate where the alterations were performed, as well as edited with “track changes” on.

Sincerely,

Reviewer 4 Report

Comments and Suggestions for Authors

Dear Authors,

This work is not a research paper and should be classified as a review. This is also because many interesting observations in this format are worth rejecting.

Sincerely,

Reviewer

Author Response

Dear reviewer,

We understand your point of view and respect it as a valued opinion in the process of revision of this paper. However, we ellaborated a new methodology using existing literature as a basis, and thus we consider that the work performed on this paper constitutes a research article and not a review paper.

In case there needs to be any other alterations, do not hesitate in contacting us.

Sincerely,

Reviewer 5 Report

Comments and Suggestions for Authors

The regulation of chemical mixtures in the Global South is a timely and highly relevant topic that is covered in this manuscript. The suggested GlORIES framework draws attention to the shortcomings of existing procedures, the significance of open-access data, and the integration of social, biological, and chemical dimensions. The manuscript is well-structured, and the interdisciplinary approach is praiseworthy. Some topics, though, call for more thorough explanation, improved methodology, and wider contextualization.

The suggested framework has a solid conceptual foundation, how will the authors confirm that it can be applied to actual case studies?
Could the authors show how the GlORIES framework compares to current regulatory approaches using at least one simulation, pilot study, or comparative example?
How does the framework handle the risk of making decisions based on biassed or incomplete datasets, even though the paper admits that there is a lack of monitoring data in the Global South?
When regionalized SSDs or AOP extrapolations are based on inadequate species data, what protections are suggested to avoid misregulation?
It's creative that local communities and civil society are included. But what safeguards are in place to guarantee that community-reported effects—like fish decline or health symptoms—are methodically verified and not anecdotal?
How will watershed committees handle power disparities (between industry, regulators, and underprivileged communities)?
According to the authors, a limited number of compounds are frequently responsible for toxicity. In complex effluents containing unknown or unmonitored emerging contaminants (such as PFAS and nanoplastics), how will the framework prioritise these compounds?
In the absence of defined regulatory thresholds, how can the framework continue to be flexible enough to accommodate new contaminants?
The methodology makes extensive use of secondary data and literature reviews. How did the authors select and weight the studies to guarantee reproducibility and quality assurance?
Could the authors elaborate on the operationalization of scope, relevance, applicability, and similarity in their assessment?
MAF is emphasised as a promising tool in the manuscript. What are the drawbacks of using a single corrective factor in diverse watersheds with radically different chemical and ecological settings?
Certain sections (such as the explanation of SSD and the biological route) heavily reiterate previously accepted knowledge. To concentrate on new contributions, think about simplifying.
By showing data flows and decision points, the framework's graphical representation (Figure 1) could be made more educational.
To prevent ambiguity, terms like "Energy Saving Principle" should be defined more precisely.
Climate change is mentioned in passing in the framework, but this aspect is not developed. How might mixture toxicity be made worse by extreme events, shifting hydrological cycles, or rising temperatures, and how might GlORIES explain this?

Author Response

Dear reviewer,

We hope to have clarified your main issues.

Sincerely,

The suggested framework has a solid conceptual foundation, how will the authors confirm that it can be applied to actual case studies?

We thank you for your thoughtful review and suggestion, however, the application of the model will be a next step done in a different project. Thus it is out of scope for this paper.

Could the authors show how the GlORIES framework compares to current regulatory approaches using at least one simulation, pilot study, or comparative example?

There is no consensus on the processes due to the novelty of the Most of the processes worldwide occur as stated on the 4^th paragraph of the introduction, with regulators doing literature research on the individual compounds and no country actually applying the mixture toxicity criteria.

How does the framework handle the risk of making decisions based on biassed or incomplete datasets, even though the paper admits that there is a lack of monitoring data in the Global South?

The framework uses the data from toxicological tests done with species as similar as possible from those in the evaluated region. Thus it does not use monitoring data, specifically to avoid the necessity of greater databases. Thus, the bias might come from the mixture toxicity papers chosen for the evaluation, which can be overcome by having several papers considered in the same assessment. The more papers in the process the less bias the regulators will face.

When regionalized SSDs or AOP extrapolations are based on inadequate species data, what protections are suggested to avoid misregulation?

For this step, it is proposed that the updating process should be performed by a group composed of experts to certify the representativeness of the selected organisms. This step would rely on the experts on the committee to provide trustworthy data.

It's creative that local communities and civil society are included. But what safeguards are in place to guarantee that community-reported effects—like fish decline or health symptoms—are methodically verified and not anecdotal?

The committee should listen to all the reports from the community as a part of the process. However, they should also perform connections with the epidemiology and environmental communities from their own regions to investigate whether the proposed address effects are being caused by the chemical pollution and not from other sources.

How will watershed committees handle power disparities (between industry, regulators, and underprivileged communities)?

By attributing the same hierarchical level between organizations during the voting phase. Thus, a set number of votes should be defined and equally divided between all relevant stakeholders.

According to the authors, a limited number of compounds are frequently responsible for toxicity. In complex effluents containing unknown or unmonitored emerging contaminants (such as PFAS and nanoplastics), how will the framework prioritise these compounds?

Prioritization, such as described on the paper, should define the toxicity of the compound as the main aspect. Thus, a highly toxic compound should be tackled first with the framework.

In the absence of defined regulatory thresholds, how can the framework continue to be flexible enough to accommodate new contaminants?

For the definition of new regulatory thresholds, the use of the current methodology for single compounds should be the first step. Then the GlORIES framework should be applied

The methodology makes extensive use of secondary data and literature reviews. How did the authors select and weight the studies to guarantee reproducibility and quality assurance?

The quality of the selected papers will be defined by the regulators in charge of the assessment. Thus, it is of great importance that the professionals selected for the framework to be highly qualified. Usually the professionals that perform the water quality thresholds for single compounds have already established methodologies for the definition of such papers

Could the authors elaborate on the operationalization of scope, relevance, applicability, and similarity in their assessment?

Explaining the operationalization on the topics in the order of appearance in the text:

Similarity: The papers were evaluated for their similarity with each other, focusing on those ideas that might be repeated worldwide;

Applicability: Papers with simpler and direct ideas were considered more applicable and thus were given preference when listing the possible reference models for the presented framework;
- When a paper was found, then the applicability was checked by the authors. Methods with more steps were considered less useful for the evaluation.
Relevance: Highly cited papers or guidelines from consolidated organizations were considered highly relevant due to their impact. Organizations such as the World Health Organization (WHO), the United Nations (UN), or the European Commission were deemed “high relevance”.
- Papers of methodologies applied by transnational organizations were prioritized for the application aspect of the framework as they mean more people from different realities were involved in the discussion of the papers, while highly localized papers were used as inspirations for methodology steps.
Scope: Papers concerning methodologies for mixture toxicity evaluation in water were deemed of high importance, while papers that discussed the water quality of single compounds were classified as medium importance.
- As the framework proposes to work with mixture toxicity, greater relevance was attributed to them.

MAF is emphasised as a promising tool in the manuscript. What are the drawbacks of using a single corrective factor in diverse watersheds with radically different chemical and ecological settings?

The main drawbacks for this methodology are firstly homogenizing the different regulatory silos and secondly homogenizing ecological regions. However, this could be corrected by the inclusion of the watershed committees during the definition phase of the MAF.

Certain sections (such as the explanation of SSD and the biological route) heavily reiterate previously accepted knowledge. To concentrate on new contributions, think about simplifying.

Thank you for your contribution.

By showing data flows and decision points, the framework's graphical representation (Figure 1) could be made more educational.

Thank you for your contribution.

To prevent ambiguity, terms like "Energy Saving Principle" should be defined more precisely.

Thank you for your contribution. It was added on L179.

Climate change is mentioned in passing in the framework, but this aspect is not developed. How might mixture toxicity be made worse by extreme events, shifting hydrological cycles, or rising temperatures, and how might GlORIES explain this?

Mixture toxicity could be increased by increased temperatures as studies have shown, since some organisms might be more sensible to pollutants. GlORIES could increase the safety by including studies that have shown this difference in temperature to be significant into the assessment.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Author Response

Dear reviewer,
Thank you for your contribution.

Sincerely,

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

Dear co-authors, thank you very much for your complete responses.

Author Response

Dear reviewer,

Thank you for your contributions,

Sincerely,

Author Response File: Author Response.docx

Reviewer 3 Report

Comments and Suggestions for Authors

Major Comments

Lack of Empirical Validation: The framework is proposed based on a literature review, but there is no testing or validation (e.g., no case study from a Global South watershed like the Amazon or Mekong). This makes the paper speculative. Suggest adding a section with a hypothetical or real-world application (e.g., applying GlORIES to a Brazilian watershed using existing data from LABTOX or similar). How would MAF be calculated in practice? Provide a worked example or simulation using tools like AOP or SSD.
Methodological Clarity: The methodology (Section 2) describes a literature review but lacks details on search results (e.g., number of papers screened, PRISMA flow diagram). How were the 20-year period and keywords selected? The evaluation criteria (similarity, applicability, etc.) are good but could be quantified (e.g., using a scoring system). Expand on how the framework was derived—did it involve stakeholder consultations or modeling?
Depth on Key Concepts: Sections on AOPs, SSDs, and MAFs are introductory and assume reader familiarity. Elaborate on limitations (e.g., AOPs are data-intensive; SSDs may overestimate protection if species data is sparse). Discuss uncertainties in mixture interactions (synergism/antagonism) more critically—reference Liu & Sayes (2024) for modeling challenges. The social route is underdeveloped; how exactly would community reporting integrate with regulatory decisions? Address potential biases in self-reported health alterations.
Global South Focus: While centered on the Global South, comparisons with Global North practices (e.g., EU Water Framework Directive) are brief. Strengthen by discussing adaptations for specific contexts (e.g., informal industries in Africa or Asia). The "energy saving principle" is vague—provide examples of existing structures (e.g., Brazil's watershed committees) and how they save resources.
Limitations and Future Work: Section 3.3 acknowledges exclusions (e.g., plastics, groundwater), but expand on feasibility barriers (e.g., data privacy laws conflicting with open access, costs of workshops). Suggest next steps, like pilot testing or integration with AI tools for AOP modeling.
Sustainability Linkages: Tie more explicitly to SDGs—e.g., how GlORIES supports Goal 12 (responsible production) via industry integration. Discuss climate-chemical interactions briefly, as hinted in the introduction.

Minor Comments

Abstract and Highlights: The abstract is clear but could specify key tools (e.g., AOPs, SSDs, MAFs) earlier. Highlights are concise but repetitive of the abstract; make them more punchy (e.g., "Regionalized SSDs reduce dependency on expensive quantification in Global South").
Figure 1: The schematic is useful but could be improved with labels for routes (chemical, biological, social) and arrows showing feedback loops. Ensure high resolution in the final version.
Repetition: Phrases like "bridge the gap between academia and regulatory practices" appear multiple times (e.g., Intro, Section 3.1.4). Consolidate for conciseness.
Terminology Consistency: Use consistent terms (e.g., "Global South" vs. "developing countries"). Define acronyms on first use (e.g., NOAEL in Intro).
Length and Tables: The paper is concise (13 pages), but consider adding a table comparing GlORIES to existing frameworks (e.g., EU SOLUTIONS project) for clarity.

Grammar, Typos, and Style

General: Language is mostly clear but has awkward phrasing (e.g., "polinizers" in Section 3.1.1.5 should be "pollinators"; "autochthones" in Section 3.1.4 should be "autochthonous"). Proofread for consistency in tense (mix of present/past).
Specific Typos:
- Page 1, Line 4: "Maria Elisa Magri 1." → Add affiliation superscript if needed.
- Page 2, Line 52: "Bench-mark dose" → "Benchmark dose".
- Page 3, Line 92: "Mode of Action (MoA)" → Consistent capitalization.
- Page 5, Line 223: "Daphnia magna" → Italicize species names throughout.
- Page 7, Line 296: "whole effluent" → "whole-effluent" for consistency.
- Page 8, Line 325: "compounds approach" → "component-based approach" (standard term).
- Page 10, Line 427: "synergistic effect" → Specify it's greater than additive.

Author Response

Dear reviewer,

Thank you for you in depth analysis of our work and your valuable contribution. As for your comments here are the responses for each topic.

Major Comments

Lack of Empirical Validation: The framework is proposed based on a literature review, but there is no testing or validation (e.g., no case study from a Global South watershed like the Amazon or Mekong). This makes the paper speculative. Suggest adding a section with a hypothetical or real-world application (e.g., applying GlORIES to a Brazilian watershed using existing data from LABTOX or similar). How would MAF be calculated in practice? Provide a worked example or simulation using tools like AOP or SSD.

This step is relevant for the application of the model and this paper is a part of a bigger project. The MAF calculation was so relevant and took such a big part of this paper that it became its own paper. Therefore, the MAF calculation is in a different paper that is being finished for future submission.

Methodological Clarity: The methodology (Section 2) describes a literature review but lacks details on search results (e.g., number of papers screened, PRISMA flow diagram). How were the 20-year period and keywords selected? The evaluation criteria (similarity, applicability, etc.) are good but could be quantified (e.g., using a scoring system). Expand on how the framework was derived—did it involve stakeholder consultations or modeling?

The methodology involved literature review on the topic using the preestablished criteria. Due to time constraints it was not possible to generate a PRISM flow diagram.

Depth on Key Concepts: Sections on AOPs, SSDs, and MAFs are introductory and assume reader familiarity. Elaborate on limitations (e.g., AOPs are data-intensive; SSDs may overestimate protection if species data is sparse). Discuss uncertainties in mixture interactions (synergism/antagonism) more critically—reference Liu & Sayes (2024) for modeling challenges. The social route is underdeveloped; how exactly would community reporting integrate with regulatory decisions? Address potential biases in self-reported health alterations.

Altered. The main topics are at L386, L411.

Global South Focus: While centered on the Global South, comparisons with Global North practices (e.g., EU Water Framework Directive) are brief. Strengthen by discussing adaptations for specific contexts (e.g., informal industries in Africa or Asia). The "energy saving principle" is vague—provide examples of existing structures (e.g., Brazil's watershed committees) and how they save resources.

The adaptations were done starting on line 266.

Limitations and Future Work: Section 3.3 acknowledges exclusions (e.g., plastics, groundwater), but expand on feasibility barriers (e.g., data privacy laws conflicting with open access, costs of workshops). Suggest next steps, like pilot testing or integration with AI tools for AOP modeling.

The inclusions start at L490.

Sustainability Linkages: Tie more explicitly to SDGs—e.g., how GlORIES supports Goal 12 (responsible production) via industry integration. Discuss climate-chemical interactions briefly, as hinted in the introduction.

Altered. Included on L353 and L388.

Minor Comments

Abstract and Highlights: The abstract is clear but could specify key tools (e.g., AOPs, SSDs, MAFs) earlier. Highlights are concise but repetitive of the abstract; make them more punchy (e.g., "Regionalized SSDs reduce dependency on expensive quantification in Global South").

Altered. Alterations start at L50.

Figure 1: The schematic is useful but could be improved with labels for routes (chemical, biological, social) and arrows showing feedback loops. Ensure high resolution in the final version.

Figure design altered for clarity at L292

Repetition: Phrases like "bridge the gap between academia and regulatory practices" appear multiple times (e.g., Intro, Section 3.1.4). Consolidate for conciseness.

Altered. Mainly on L149.

Terminology Consistency: Use consistent terms (e.g., "Global South" vs. "developing countries"). Define acronyms on first use (e.g., NOAEL in Intro).

Altered throughout the text.

Length and Tables: The paper is concise (13 pages), but consider adding a table comparing GlORIES to existing frameworks (e.g., EU SOLUTIONS project) for clarity.

Even though the table would be a great addition to the paper, due to time constraints it was not possible to elaborate one.

Grammar, Typos, and Style

General: Language is mostly clear but has awkward phrasing (e.g., "polinizers" in Section 3.1.1.5 should be "pollinators"; "autochthones" in Section 3.1.4 should be "autochthonous"). Proofread for consistency in tense (mix of present/past).

Revised throughout the paper and altered when needed.

Specific Typos:

All specific typos were corrected as demanded.

Reviewer 4 Report

Comments and Suggestions for Authors

Dear Authors and Editors,

After applying the changes in the M&M section, the publication is ready for publication.

Sincerely,

Ł. Sikorski

Author Response

Dear reviewer,

Thank you for your contributions,

Sincerely,

Author Response File: Author Response.docx

Round 3

Reviewer 3 Report

Comments and Suggestions for Authors

The authors respond to previous comments and did the suggested changes.