Abstract
Marine macroalga are frequently exposed to environmental stresses impairing their overall physiology and growth potential. Among these, Gracilaria cornea (Rhodophyta) is a valuable red seaweed rich in protein and polysaccharides. To investigate its physiological responses under controlled conditions, we cultivated Gracilaria cornea in an indoor culture system at three different salinity levels (30, 40 and 50 ppt), employing continuous aeration, blue and white LED illumination (12:12 light: dark cycle), and exogenous addition of nitrogen and phosphorus. Physiological changes associated with protein content accumulation and amino acid composition were determined using in-situ reflectance spectroscopy (VIS-NIR range 560–674 nm), AI algorithm and GC-MS analysis. We developed novel tools to accurately predict amino acid composition and total protein yield, identified the environmental factors inducing trait accumulation and determined the optimal harvesting day. Hypersaline stress and cultivation day significantly influenced protein content with optimal protein content (> 35% dry weight) achieved on day 14. This peak was not correlated with the specific growth rate (SGR), indicating SGR may not reliably indicate protein yield in this context. The dry weight to fresh weight ratio (DW: FW) was higher under hypersaline conditions, leading to a greater dried biomass and higher protein content, despite a reduced overall growth rate. Protein content was maximal under high ambient pH and high salinity. Day 14 was optimal for the highest yield of essential amino acids (EAA), exceeding 40% of the total amino acids. The algorithmic model accurately predicted specific amino acid proportions.
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Data availability
The spectral and laboratory measurements that support the findings of this study are available from the University of Haifa, but restrictions apply to the availability of these data, which were used under licence for the current study and so are not publicly available. The data are, however, available upon request and with the permission of the University of Haifa.
Abbreviations
- NIR:
-
Near infrared
- ANN:
-
Artificial neural network
- DW:
-
Dry weight
- FW:
-
Fresh weight
- VIS-NIR:
-
Visible-near
- NM:
-
Nano meter
- GC-MS:
-
Gas chromatography mass spectrometry
- AA:
-
Amino acid
- EAA:
-
Essential amino acid
- NEAA:
-
Nonessential amino acid
- Ppt:
-
Parts per thousand
References
Olsson, J., Toth, G. B. & Albers, E. Biochemical composition of red, green and brown seaweeds on the Swedish West Coast. J. Appl. Phycol. 32 (5), 3305–3317 (2020).
Mæhre, H. K., Malde, M. K., Eilertsen, K. E. & Elvevoll, E. O. Characterization of protein, lipid and mineral contents in common Norwegian seaweeds and evaluation of their potential as food and feed. J. Sci. Food. Agric. 94 (15), 3281–3290 (2014).
Holdt, S. L. & Kraan, S. Bioactive compounds in seaweed: functional food applications and legislation. J. Appl. Phycol. 23, 543–597 (2011).
Ashkenazi, D. Y. et al. Enrichment of nutritional compounds in seaweeds via abiotic stressors in integrated aquaculture. Innovative Food Science & Emerging Technologies 80, 103067 (2022).
Machado, M. et al. Amino acid profile and protein quality assessment of macroalgae produced in an integrated multi-trophic aquaculture system. Foods 9(10), 1382. (2020).
Brien, R. O., Hayes, M., Sheldrake, G., Tiwari, B. & Walsh, P. Macroalgal proteins: a review. Foods 11(4), 571 (2022).
de Souza Celente, G., Sui, Y. & Acharya, P. Seaweed as an alternative protein source: Prospective protein extraction technologies. Innovative Food Science & Emerging Technologies 86, 103374 (2023).
Kumar, M., Kumari, P., Reddy, C. R. K. & Jha, B. Salinity and desiccation induced oxidative stress acclimation in seaweeds. In Advances in Botanical Research, Vol. 71 (ed. Bourgougnon, N.) 91–123 (Academic, 2014).
Wen, J. et al. Comparative analysis of proteins involved in energy metabolism and protein processing in Pyropia haitanensis at different salinity levels. Frontiers in Marine Science 7, 415 (2020).
Mensi, F., Ben Ghedifa, A. & Rajhi, H. Effects of seawater sulfur starvation and enrichment on Gracilaria gracilis growth and biochemical composition. Scientific Reports 12(1), 11095 (2022).
Michalak, I. & Chojnacka, K. Algae as production systems of bioactive compounds. Eng. Life Sci. 15 (2), 160–176 (2015).
Kazir, M. et al. Extraction of proteins from two marine macroalgae, Ulva sp. and Gracilaria sp., for food application, and evaluating digestibility, amino acid composition and antioxidant properties of the protein concentrates. Food Hydrocoll. 87, 194–203 (2019).
Torres, P., Santos, J. P., Chow, F. & Santos, D. Y. dos A comprehensive review of traditional uses, bioactivity potential, and chemical diversity of the genus Gracilaria (Gracilaria les, Rhodophyta). Algal Research 37, 288–306 (2019).
Sobuj, M. K. A. et al. Floating raft culture of Gracilariopsis longissima for optimum biomass yield performance on the coast of Cox’s Bazar, Bangladesh. Scientific Reports 13(1), 2308 (2023).
Mensi, F., Nasraoui, S., Bouguerra, S., Ben Ghedifa, A. & Chalghaf, M. Effect of lagoon and sea water depth on Gracilaria gracilis growth and biochemical composition in the Northeast of Tunisia. Sci. Rep. 10 (1), 1–12 (2020).
Buschmann, A. H. et al. Seaweed production: overview of the global state of exploitation, farming and emerging research activity. Eur. J. Phycol. 52 (4), 391–406 (2017).
Tadmor-Shalev, N. et al. NIR spectroscopy and artificial neural network for seaweed protein content assessment in-situ. Computers and Electronics in Agriculture 201, 107–304 (2022).
Yu, C. H., Lim, P. E. & Phang, S. M. Effects of irradiance and salinity on the growth of carpospore-derived tetrasporophytes of Gracilaria Edulis and Gracilaria tenuistipitata var Liui (Rhodophyta). J. Appl. Phycol. 25, 787–794 (2013).
Torres, P. et al. Brazilian native species of Gracilaria (Gracilaria les, Rhodophyta) as a source of valuable compounds and as nutritional supplements. J. Appl. Phycol. 31 (5), 3163–3173 (2019).
Dawes, C. J., Orduna-Rojas, J. & Robledo, D. Response of the tropical red seaweed Gracilaria cornea to temperature, salinity and irradiance. J. Appl. Phycol. 10 (5), 419–425 (1998).
Chen, B., Zou, D., Zhu, M. & Yang, Y. Effects of CO 2 levels and light intensities on growth and amino acid contents in red seaweed Gracilaria lemaneiformis. Aquac. Res. 48 (6), 2683–2690 (2017).
Kumar, M., Kumari, P., Gupta, V., Reddy, C. R. K. & Jha, B. Biochemical responses of red Alga Gracilaria Ongissim (Gracilaria les, Rhodophyta) to salinity induced oxidative stress. J. Exp. Mar. Biol. Ecol. 391 (1–2), 27–34 (2010).
Castro, J. Z. & Yokoya, N. S. Growth and biochemical responses of tropical and subtropical strains of Gracilaria domingensis (Gracilaria les, Rhodophyta) to temperature and irradiance variations. J. Appl. Phycol. 31 (1), 607–613 (2019).
Heemboo, M., Thammakhet-Buranachai, C., Makkliang, F. & Buapet, P. Enhalus acoroides seedlings exhibit different high light responses under varying light qualities. Plant Stress 10, 100232 (2023).
Senuma, M., Dobashi, S., Bando, Y., Ko, S. & Shiota, H. Overexpression of eelgrass Rare Cold Inducible 2 (RCI2) maintains chlorophyll content in Arabidopsis subjected to high salinity and dehydration. Plant Stress 6, 100116 (2022).
El-Rafie, H. M., Hammam, H. H. & Ahmed, E. A. E. Nutritional values, antioxidant, and Cytotoxic Activities of Selected Edible Marine Macroalgae: a Comparative Study (Food Measurement and Characterization, 2024).
Donadio, R. et al. Assessing the resilience of the coralline macroalga Ellisolandia elongata in response to a prolonged low tide. Environmental Research 277, 121579 (2025).
Lawton, R. J., de Nys, R., Magnusson, M. E. & Paul, N. A. The effect of salinity on the biomass productivity, protein and lipid composition of a freshwater macroalga. Algal Res. 12, 213–220 (2015).
Ekman, P., Yu, S. & Pedersen, M. Effects of altered salinity, darkness and algal nutrient status on floridoside and starch content, α-galactosidase activity and agar yield of cultivated Gracilaria sordida. Brit. Phycol. J. 26 (2), 123–131 (1991).
Siddiqui, S. A., Agrawal, S., Brahmbhatt, H. & Rathore, M. S. Metabolite expression changes in Kappaphycus alvarezii (a red alga) under hypo-and hyper-saline conditions. Algal Research 63, 102650 (2022).
Contreras-Porcia, L., Thomas, D., Flores, V. & Correa, J. A. Tolerance to oxidative stress induced by desiccation in Porphyra columbina (Bangiales, Rhodophyta). J. Exp. Bot. 62 (6), 1815–1829 (2011).
Kinnby, A. et al. Combining an ecological experiment and a genome scan show idiosyncratic responses to salinity stress in local populations of a seaweed. Frontiers in Marine Science 7, 470 (2020).
Bar-Shai, N., Sharabani-Yosef, O., Zollmann, M., Lesman, A. & Golberg, A. Seaweed cellulose scaffolds derived from green macroalgae for tissue engineering. Scientific reports 11(1), 11843 (2021).
El-Said, G. F. & El-Sikaily, A. Chemical composition of some seaweed from mediterranean sea coast, Egypt. Environ. Monit. Assess. 185 (7), 6089–6099 (2013).
Karsten, U. Seaweed acclimation to salinity and desiccation stress. Seaweed biology: Novel insights into ecophysiology, ecology and utilization, pp.87–107. (2012).
Yin, J. et al. J. and Dynamic changes and transcriptome analyses reveal the microfilament skeleton response to water stress in thalli of Neopyropia yezoensis inhabiting the intertidal zone. Plant Stress 15, 100762 (2025).
Pereira, D. T. et al. Effects of salinity on the physiology of the red macroalga, Acanthophora spicifera (Rhodophyta, Ceramiales). Acta Bot. Brasilica. 31, 555–565 (2017).
Israel, A., Martinez-Goss, M. & Friedlander, M. Effect of salinity and pH on growth and agar yield of Gracilaria tenuistipitata var. Liui in laboratory and outdoor cultivation. J. Appl. Phycol. 11, 543–549 (1999).
Bermejo, A. et al. E. and Influence of irradiance, dissolved nutrients and salinity on the colour and nutritional characteristics of Gracilariopsis longissimi (Rhodophyta). Algal Research 52, 102121 (2020).
Wang, W. et al. Regulatory mechanisms underlying the maintenance of homeostasis in Pyropia haitanensis under hypersaline stress conditions. Sci. Total Environ. 662, 168–179 (2019).
Tadmor-Shalev, N. et al. Light attenuation as a substitute for nutrient supply for maximizing protein content in Gracilaria cornea (Rhodophyta): Modeling nitrogen and phosphorus supplementation using a pharmacokinetic approach. Cleaner Engineering and Technology 26, 100948 (2025).
Krom, M. D., Kress, N., Brenner, S. & Gordon, L. I. Phosphorus limitation of primary productivity in the Eastern mediterranean sea. Limnol. Oceanogr. 36 (3), 424–432 (1991).
Berman-Frank, I. & Rahav, E. Dinitrogen fixation as a source for new production in the Mediterranean Sea: a review. Life in the Mediterranean Sea: A look at habitat changes 199–226. (2012).
Kress, N., Rahav, E., Silverman, J. & Herut, B. Environmental status of israel’s mediterranean coastal waters: setting reference conditions and thresholds for nutrients, chlorophyll-a and suspended particulate matter. Mar. Pollut. Bull. 141, 612–620 (2019).
Hirte, J., Richner, W., Orth, B., Liebisch, F. & Flisch, R. Yield response to soil test phosphorus in Switzerland: Pedoclimatic drivers of critical concentrations for optimal crop yields using multilevel modelling. Science of the Total Environment 755, 143453 (2021).
Ozer, T., Gertman, I., Gildor, H. & Herut, B. Thermohaline Temporal Variability of the SE Mediterranean Coastal Waters (Israel)–Long-Term Trends, Seasonality, and Connectivity. Frontiers in Marine Science 8, 799457 (2022).
Friedlander, M. & Levy, I. Cultivation of gracilaria in outdoor tanks and ponds. J. Appl. Phycol. 7 (3), 315–324 (1995).
Olmedo-Masat, O. M., Raffo, M. P., Rodríguez-Pérez, D., Arijón, M. & Sánchez-Carnero, N. How far can we classify macroalgae remotely? An example using a new spectral library of species from the southwest Atlantic (Argentine Patagonia). Remote Sensing 12(23), 3870 (2020).
Polinova, M., Salinas, K., Bonfante, A. & Brook, A. Irrigation optimization under a limited water supply by the integration of modern approaches into traditional water management on the cotton fields. Remote Sensing 11(18), 2127 (2019).
Ma, T., Johnston, W. M. & Koran, A. The color accuracy of the Kubelka-Munk theory for various colorants in maxillofacial prosthetic material. J. Dent. Res. 66 (9), 1438–1444 (1987).
Angell, A. R., Mata, L., de Nys, R. & Paul, N. A. The protein content of seaweeds: a universal nitrogen-to-protein conversion factor of five. J. Appl. Phycol. 28, 511–524 (2016).
Lawton, R. J., De Nys, R. & Paul, N. A. Selecting reliable and robust freshwater macroalgae for biomass applications. PloS One. 8 (5), e64168 (2013).
Kashyap, M. et al. Extracting Water-Soluble proteins from the red macroalgae Gracilaria cornea with pulsed electric field in a continuous process. ACS Food Sci. Technol. 3 (4), 562–575 (2022).
Figueroa, F. L. et al. Interactive effects of solar radiation and inorganic nutrients on biofiltration, biomass production, photosynthetic activity and the accumulation of bioactive compounds in Gracilaria cornea (Rhodophyta). Algal Research 68, 102890 (2022).
Martinez, S. et al. Energy sources of the depth-generalist mixotrophic coral Stylophora pistillata. Frontiers in Marine Science 7, 566663 (2020).
McMahon, K. W., Fogel, M. L., Johnson, B. J., Houghton, L. A. & Thorrold, S. R. A new method to reconstruct fish diet and movement patterns from δ13C values in otolith amino acids. Can. J. Fish. Aquat. Sci. 68 (8), 1330–1340 (2011).
Burnham, K. P., Anderson, D. R. & Huyvaert, K. P. AIC model selection and multimodel inference in behavioral ecology: some background, observations, and comparisons. Behav. Ecol. Sociobiol. 65, 23–35 (2011).
Ma, C. et al. Nitrogen enrichment mediates the effects of high temperature on the growth, photosynthesis, and biochemical constituents of Gracilaria blodgettii and Gracilaria lemaneiformis. Environmental Science and Pollution Research 28(17), 21256–21265 (2021).
Choi, T. S., Kang, E. J., Kim, J. H. & Kim, K. Y. Effect of salinity on growth and nutrient uptake of Ulva Pertusa (Chlorophyta) from an eelgrass bed. Algae 25 (1), 17–26 (2010).
(FAO & Rome)., Dietary protein quality evaluation in human nutrition: Report of an FAO Expert Consultation, 31 March-2 April, 2011, Auckland, New Zealand. Food and Agriculture Organization of the United Nations (2013).
Holeček, M. Branched-chain amino acids in health and disease: metabolism, alterations in blood plasma, and as supplements. Nutr. Metabolism. 15, 1–12 (2018).
Chan, P. T. & Matanjun, P. Chemical composition and physicochemical properties of tropical red seaweed, Gracilaria Changii. Food Chem. 221, 302–310 (2017).
Thiviya, P., Gamage, A., Gama-Arachchige, N. S., Merah, O. & Madhujith, T. Seaweeds as a source of functional proteins. Phycology 2 (2), 216–243 (2022).
Brosnan, J. T. & Brosnan, M. E. Glutamate: a truly functional amino acid. Amino Acids. 45, 413–418 (2013).
Li, J. et al. Effects of temperature and salinity on the growth and biochemical composition of the brown Alga Sargassum fusiforme (Fucales, Phaeophyceae). J. Appl. Phycol. 31, 3061–3068 (2019).
Liu, L. & Lin, L. Effect of heat stress on Sargassum fusiforme leaf metabolome. J. Plant. Biology. 63 (3), 229–241 (2020).
Park, E. et al. Seaweed metabolomics: A review on its nutrients, bioactive compounds and changes in climate change. Food Research International 163, 112221 (2023).
Gao, G., Clare, A. S., Chatzidimitriou, E., Rose, C. & Caldwell, G. Effects of ocean warming and acidification, combined with nutrient enrichment, on chemical composition and functional properties of Ulva rigida. Food Chem. 258, 71–78 (2018).
Chen, B., Xia, J., Zou, D. & Zhang, X. Responses to ocean acidification and diurnal temperature variation in a commercially farmed seaweed, Pyropia haitanensis (Rhodophyta). Eur. J. Phycol. 54 (2), 184–192 (2019).
Mtolera, M. S. Some properties of glutamate dehydrogenase from the marine red Alga Gracilaria sordida (Harv.) W. Nelson. Western Indian Ocean. J. Mar. Sci. 2 (2), 179–186 (2003).
Bamary, Z. & Einali, A. Nitrogen metabolism and activity of amino acid metabolizing enzymes in the unicellular green Alga Dunaliella sp. under long-term salinity and arginine treatment. J. Appl. Phycol. 35 (6), 2801–2813 (2023).
Bermejo, B., Cara, R., Macías, C. L., Sánchez-García, M. & Hernández, I. J. and Growth rates of Gracilariopsis longissima, Gracilaria bursa-pastoris and Chondracanthus teedei (Rhodophyta) cultured in ropes: implication for N biomitigation in Cadiz Bay (Southern Spain). Journal of Applied Phycology 32, 1879–1891 (2020).
Roleda, M. Y. & Hurd, C. L. Seaweed nutrient physiology: application of concepts to aquaculture and bioremediation. Phycologia 58 (5), 552–562 (2019).
Angell, A. R., Mata, L., de Nys, R. & Paul, N. A. Indirect and direct effects of salinity on the quantity and quality of total amino acids in Ulva Ohnoi (Chlorophyta). J. Phycol. 51 (3), 536–545 (2015).
Norziah, M. H. & Ching, C. Y. Nutritional composition of edible seaweed gracilaria Changgi. Food Chem. 68 (1), 69–76 (2000).
Lourenço, S. O., Barbarino, E., De-Paula, J. C., Pereira, L. O. D. S. & Marquez, M. L. Amino acid composition, protein content and calculation of nitrogen‐to‐protein conversion factors for 19 tropical seaweeds. Phycological Res. 50 (3), 233–241 (2002).
Tabarsa, M., Rezaei, M., Ramezanpour, Z. & Waaland, J. R. Chemical compositions of the marine algae gracilaria salicornia (Rhodophyta) and Ulva lactuca (Chlorophyta) as a potential food source. J. Sci. Food. Agric. 92 (12), 2500–2506 (2012).
WHO/FAO/UNU. Protein and amino acid requirements in human nutrition. Rep. Joint FAO/WHO/UNU Expert Consultation; Tech. Rep. Ser. 935. 935, 1 (2007).
Cebrián-Lloret, V., Martínez-Abad, A., Recio, I., López-Rubio, A. & Martínez-Sanz, M. In vitro digestibility of proteins from red seaweeds: Impact of cell wall structure and processing methods. Food Research International 178, 113990 (2024).
Mabeau, S. & Fleurence, J. Seaweed in food products: biochemical and nutritional aspects. Trends Food Sci. Technol. 4 (4), 103–107 (1993).
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Niva Tadmor-Shalev: Conceptualization, Methodology, Validation, Formal Analysis, Investigation, Data Curation, Writing – Original Draft, Writing – Review & Editing, Visualization, Project Administration. Anna Brook, Software, Formal analysis, Data curation, Writing – Review & Editing. Eli Shemesh: Conceptualization, Methodology, Investigation, Resources, Data curation. Andrea Ghermandi: Writing – Review & Editing, Supervision. Álvaro Israel: Methodology, Writing – Review & Editing. Dan Tchernov: Resources, Supervision.
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Tadmor-Shalev, N., Shemesh, E., Israel, Á. et al. Salinity stress enhances protein content and amino acid profile in Gracilaria cornea (Rhodophyta). Sci Rep (2026). https://doi.org/10.1038/s41598-026-36828-0
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DOI: https://doi.org/10.1038/s41598-026-36828-0


