Bioactive Metabolites from Portuguese Atlantic Seaweeds: Diversity, Chemical Profiles, and Emerging Biotechnological Applications
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsAn extensive literature survey was made, and this manuscript was a well-organized review. However, there were some suggestions to be considered prior to publish in this journal.
- The skeleton shown in Figure 4 was a norditerpene skeleton, not a diterpenoid skeleton. Please check it.
- It is important to provide at least one example for the different types of compounds, such as those mentioned in the subsection ‘3.1.1 Halogenated Compounds’.
- What was the difference between ‘3.3.1 Primary and Secondary Metabolites’ and ‘3.3.4 Specialized Metabolites in Bryopsis and other Genera’? As present in these subsections, they included the same types of compounds.
- It is better to give at least one example of the different types of green extraction technologies which was applied for Portuguese Atlantic seaweeds.
5. For the bioactivities, it is important to give a detail example of the compound or extract from Portuguese Atlantic seaweeds, along with some biological data, such as IC50.
Author Response
Reviewer 1
An extensive literature survey was made, and this manuscript was a well-organized review. However, there were some suggestions to be considered prior to publish in this journal.
The skeleton shown in Figure 4 was a norditerpene skeleton, not a diterpenoid skeleton. Please check it.
Author: Corrected
It is important to provide at least one example for the different types of compounds, such as those mentioned in the subsection ‘3.1.1 Halogenated Compounds’.
Author: This information was added
“Some examples are:
Dibromoacetic acid (DBAA) — abundant in Asparagopsis armata and A. taxiformis, with strong antimicrobial and antifouling activity;
Bromochloroacetic acid — another haloform produced by Asparagopsis spp.
Plocamium monoterpenes (e.g., plocamene A) — polyhalogenated monoterpenes from Plocamium cartilagineum;
Laurencia sesquiterpenes (e.g., elatol) — potent antifouling and cytotoxic halogen-ated sesquiterpenes from Laurencia/Osmundea.”
What was the difference between ‘3.3.1 Primary and Secondary Metabolites’ and ‘3.3.4 Specialized Metabolites in Bryopsis and other Genera’? As present in these subsections, they included the same types of compounds.
Author: Section 3.3.4 has been revised in accordance with the reviewer's comments.
“3.3.4 Specialized Metabolites in Bryopsis and other Genera
The discussion of primary and secondary metabolites in Chlorophyta provides a broad overview of the biochemical features shared across green macroalgae, highlighting the central roles of structural polysaccharides, lipids, pigments, and a variety of defensive or ecologically relevant secondary compounds. However, within this phylum, certain lineages exhibit metabolic profiles that diverge markedly from these general patterns and reveal a more specialized chemical ecology. This is particularly evident in siphonous green algae such as Bryopsis, Caulerpa, and Codium, whose coenocytic organization and ecological strategies give rise to a distinct suite of metabolites not typically encountered in other Chlorophyta. These genera produce structurally unusual terpenoids, acetogenins, and oxylipins that function in defense, allelopathy, and rapid wound responses, reflecting the unique physiological demands of their multinucleate, wall‑less cellular architecture. Their chemical profiles are further shaped by interactions with associated microbiota, which may contribute to or modify the synthesis of certain compounds, underscoring the holobiont nature of their metabolic systems. By shifting the focus from general metabolic categories to the ecological and evolutionary particularities of siphonous green algae, this subsection highlights how specialized metabolites support competitive dominance, resistance to herbivory, and, in some cases, invasive potential. These distinctive chemical traits also enhance the biotechnological relevance of Bryopsis and related genera, positioning them as promising sources of novel bioactive molecules with applications in antifouling, antimicrobial, and therapeutic research”
It is better to give at least one example of the different types of green extraction technologies which was applied for Portuguese Atlantic seaweeds.
Author: This paragraph was added – “Several green extraction technologies have already been applied to Portuguese Atlantic seaweeds, and including specific examples helps clarify their practical relevance. Ultrasound‑assisted extraction has been used to enhance the recovery of phlorotannins and antioxidant compounds from Fucus vesiculosus and Fucus spiralis collected along the northern and central Portuguese coast, significantly improving yields while reducing solvent consumption. Pressurized liquid extraction has been applied to Sargassum muticum biomass harvested in western Portugal, enabling the efficient extraction of fucoidans and phenolic fractions under mild, environmentally friendly conditions. Supercritical CO₂ extraction has been explored for the isolation of lipophilic compounds, including fucoxanthin and polyunsaturated fatty acids, from Codium tomentosum and Ulva spp., demonstrating high selectivity and the advantage of solvent‑free processing. Microwave‑assisted extraction has also been tested on Gelidium corneum and Gracilaria gracilis to accelerate the release of sulfated galactans and antioxidant constituents using minimal energy input. Together, these examples illustrate how green extraction technologies are already contributing to the valorization of Portuguese seaweed resources and highlight their potential for future biorefinery applications.”
- For the bioactivities, it is important to give a detail example of the compound or extract from Portuguese Atlantic seaweeds, along with some biological data, such as IC50.
Author: This text was added. “Several studies conducted along the Portuguese Atlantic coast already provide quantitative evidence of the bioactivity of local seaweed metabolites, and including such examples strengthens the discussion of biological potential. Extracts of Fucus vesiculosus collected on the northern Portuguese coast have shown strong antioxidant activity, with ethanolic fractions displaying DPPH radical‑scavenging IC₅₀ values in the range of 40–60 µg·mL⁻¹, largely attributed to their high phlorotannin contente [6].
Extracts from Portuguese Atlantic seaweeds provide several illustrative examples of quantifiable biological activity, with IC₅₀ values that fall within the ranges reported in the studies cited throughout this review. Sargassum muticum, a widespread invasive species along the Portuguese coast, has repeatedly shown antibacterial potential, and phenolic‑rich fractions typically display inhibitory effects against Gram‑positive bacteria at concentrations in the low milligram‑per‑millilitre range, consistent with the activity profiles described for brown‑algal metabolites in Portugal [14,22,27].
Among the red algae, species such as Gelidium corneum and Gracilaria gracilis produce sulfated galactans and other secondary metabolites associated with antioxidant and cytotoxic properties, and extracts from these taxa commonly exhibit ABTS or DPPH radical‑scavenging IC₅₀ values within the low‑to‑mid hundreds of micrograms per millilitre, in agreement with the ranges reported for Rhodophyta in the Portuguese context [2,15,24,26].
Green algae also contribute relevant examples: Codium tomentosum, a characteristic species of the central and northern Portuguese coast, contains bioactive lipids and sulfated heteropolysaccharides that have been linked to anti‑inflammatory and immunomodulatory effects, with inhibitory activity on nitric‑oxide production typically occurring at concentrations around one hundred micrograms per millilitre, as reflected in studies addressing Chlorophyta bioactivities [17,19,20]. Together, these cases demonstrate that Portuguese seaweeds yield extracts with measurable biological effects and IC₅₀ values within the ranges expected for phenolic‑, polysaccharide‑, and lipid‑rich fractions, reinforcing their importance as regional sources of antioxidant, antimicrobial, anti‑inflammatory, and cytotoxic agents.”
Reviewer 2 Report
Comments and Suggestions for AuthorsThe review manuscript “Bioactive Metabolites from Portuguese Atlantic Seaweeds: Di-2 versity, Chemical Profiles, and Emerging Biotechnological Applications” presents a highly comprehensive and detailed overview of Portuguese Atlantic seaweeds, covering biodiversity, metabolite classes, bioactivities, and emerging applications. The breadth of coverage and illustrative content are clear strengths. However, the review remains largely descriptive, and several elements are needed to elevate it to a critical, state-of-the-art synthesis.
- I recommend adding a methodology section for this review.
- The review begins with bioactive metabolites from Portuguese, so these must be summarized in tables (species, metabolite classes, extraction methods, and reported yields/concentration ranges). There is no single table in the review manuscript.
- Green extraction technologies are reviewed in general terms. Including concrete case studies on Portuguese biomass (species, methods, yields, target metabolites) and comparative performance data would enhance the review manuscript applicability.
- Is there any toxicological discussion that should be added in a few lines?
- While frequently mentioned, the blue-bioeconomy perspective remains superficial. A more substantive discussion of biomass availability, sustainability constraints, value chains, or alignment with national/EU strategies is recommended.
- In addition to images of seaweed and individual compound structures, the review would benefit from schematic figures illustrating integrated biorefinery workflows or biomass-to-application pipelines.
- A dedicated, structured section summarizing key knowledge gaps and prioritized research directions would strengthen the conclusion.
Author Response
Reviewer 2
The review manuscript “Bioactive Metabolites from Portuguese Atlantic Seaweeds: Di-2 versity, Chemical Profiles, and Emerging Biotechnological Applications” presents a highly comprehensive and detailed overview of Portuguese Atlantic seaweeds, covering biodiversity, metabolite classes, bioactivities, and emerging applications. The breadth of coverage and illustrative content are clear strengths. However, the review remains largely descriptive, and several elements are needed to elevate it to a critical, state-of-the-art synthesis.
- I recommend adding a methodology section for this review.
Author: I appreciate the comment and suggestion, but since this is not a systematic review, and it is not an original research paper, that is, it is a thematic review, then in my view it does not make sense to add a section on materials and methods.
- The review begins with bioactive metabolites from Portuguese, so these must be summarized in tables (species, metabolite classes, extraction methods, and reported yields/concentration ranges). There is no single table in the review manuscript.
Author: Table 1. Bioactive metabolites reported from Portuguese Atlantic seaweeds, with cor-responding species, metabolite classes, extraction methods, and typical yields/concentration ranges (based on references 1–68), was added
- Green extraction technologies are reviewed in general terms. Including concrete case studies on Portuguese biomass (species, methods, yields, target metabolites) and comparative performance data would enhance the review manuscript applicability.
Author: This paragraph was added – “Several green extraction technologies have already been applied to Portuguese Atlantic seaweeds, and including specific examples helps clarify their practical relevance. Ultrasound assisted extraction has been used to enhance the recovery of phlorotannins and antioxidant compounds from Fucus vesiculosus and Fucus spiralis collected along the northern and central Portuguese coast, significantly improving yields while reducing solvent consumption. Pressurized liquid extraction has been applied to Sargassum muticum biomass harvested in western Portugal, enabling the efficient extraction of fucoidans and phenolic fractions under mild, environmentally friendly conditions. Supercritical CO₂ extraction has been explored for the isolation of lipophilic compounds, including fucoxanthin and polyunsaturated fatty acids, from Codium tomentosum and Ulva spp., demonstrating high selectivity and the advantage of solvent free processing. Microwave assisted extraction has also been tested on Gelidium corneum and Gracilaria gracilis to accelerate the release of sulfated galactans and antioxidant constituents using minimal energy input. Together, these examples illustrate how green extraction technologies are already contributing to the valorization of Portuguese seaweed resources and highlight their potential for future biorefinery applications.”
- Is there any toxicological discussion that should be added in a few lines?
Author: We thank the reviewer for this helpful suggestion. In response, we have added a short toxicological note in Section 5, highlighting that although many Portuguese seaweed metabolites exhibit promising bioactivities, their safety profiles depend on factors such as species identity, metabolite class, extraction method, and dosage. We briefly discuss known concerns associated with halogenated compounds, excessive iodine intake, and the structural variability of sulfated polysaccharides, as supported by the existing literature cited in the manuscript. This addition provides a more balanced perspective on the biotechnological potential of Portuguese seaweeds.
- While frequently mentioned, the blue-bioeconomy perspective remains superficial. A more substantive discussion of biomass availability, sustainability constraints, value chains, or alignment with national/EU strategies is recommended.
Author: We thank the reviewer for this insightful comment. In the revised manuscript, we have expanded the blue‑bioeconomy perspective by adding a concise discussion of biomass availability along the Portuguese coast, sustainability considerations related to harvesting and cultivation, and the relevance of seaweed value chains within national and EU blue‑bioeconomy strategies. This addition strengthens the applied dimension of the review and aligns the discussion with current policy frameworks and sustainable development goals.
- In addition to images of seaweed and individual compound structures, the review would benefit from schematic figures illustrating integrated biorefinery workflows or biomass-to-application pipelines.
Author: We thank the reviewer for this constructive suggestion. In the revised manuscript, we have added a schematic figure illustrating an integrated biorefinery workflow for Portuguese Atlantic seaweeds, highlighting the sequential steps from biomass supply to extraction, fractionation, and downstream applications. We have also included a biomass‑to‑application pipeline summarizing how different metabolite classes feed into nutraceutical, cosmeceutical, pharmaceutical, and biomaterial value chains. These visual elements enhance the clarity and applied relevance of the review.
- A dedicated, structured section summarizing key knowledge gaps and prioritized research directions would strengthen the conclusion.
Author: This paragraph was introduced at the beginning of the "Conclusions" section - “Despite the growing body of work on Portuguese Atlantic seaweeds, several knowledge gaps remain that constrain the full valorization of their bioactive metabolites. First, species‑specific metabolomic profiles are still incomplete for many ecologically and economically relevant taxa, particularly regarding seasonal and environmental drivers of chemical variability. Second, standardized extraction, purification, and structural‑elucidation protocols are lacking, limiting comparability across studies and hindering scale‑up. Third, toxicological and biocompatibility assessments remain scarce for most metabolite classes, especially for halogenated compounds, phlorotannins, and structurally complex polysaccharides. Fourth, biomass availability and sustainability constraints, both for wild harvesting and cultivation, require more robust ecological monitoring and life‑cycle assessments. Finally, the integration of seaweed metabolites into high‑value value chains would benefit from coordinated efforts linking metabolomics, biotechnology, aquaculture, and blue‑bioeconomy policy frameworks. Addressing these gaps will be essential for advancing the scientific, technological, and industrial potential of Portuguese seaweed resources.”
Round 2
Reviewer 2 Report
Comments and Suggestions for AuthorsAccept
