Introduction
Matrix metalloproteinases (MMPs) are enzymes that play pivotal roles in various biological processes, including tissue remodeling and inflammatory responses across multiple organs . Certain bioactive compounds found in macroalgae—such as sulfated polysaccharides and phlorotannins—have attracted increasing attention due to their capacity to act as matrix metalloproteinase inhibitors (MMPIs), thereby modulating extracellular matrix (ECM) turnover and attenuating inflammation (Jayawardhana et al ., 2023). In several teleost fish species, intestinal inflammation—sharing pathophysiological similarities with human inflammatory bowel disease—can be induced by diets containing plant-based ingredients (Krogdahl et al ., 2015). However, to date, no studies have explored the potential of algal-derived MMPIs to mitigate such diet-induced enteropathies. This preliminary study aimed to: (a) determine the presence of MMPI activity in a concentrated extract obtained from the green macroalga Ulva spp ; (b) evaluate the compound’s stability under in vitro conditions simulating gastrointestinal digestion in Sparus aurata (gilthead seabream); and (c) assess its effects on intestinal function in this species when included in experimental diets.
Materials and methods
A protein-enriched concentrated extract was obtained from the green macroalga Ulva spp. through a multi-step extraction process conducted at ANFACO-CECOPESCA (Vigo, Spain). The extract’s potential to inhibit matrix metalloproteinases (MMPs) from Clostridium histolyticum was evaluated using the EnzChek™ Gelatinase/Collagenase Assay Kit (Thermo Fisher Scientific). Upon confirmation of inhibitory activity, the extract was incorporated at 12% (w/w) into an experimental diet containing 50% plant-derived ingredients, including 10% soybean-based products. This formulation was subjected to an in vitro digestion assay simulating seabream (Sparus aurata ) gastrointestinal conditions, following the methodology described by Martínez-Antequera et al. (2023). A control diet, identical in composition but lacking the algal concentrate, was used for comparison. Both diets were subsequently administered to juvenile seabream (68.7 ± 0.1 g) over a 6-week feeding trial. At the end of the trial, fish were sampled and intestinal functionality was assessed by measuring epithelial electrical resistance and paracellular permeability using Ussing chamber techniques, following the procedure outlined by Gregorio and Fuentes (2018).
Results and discussion
The concentrated U. lactuca extract exhibited strong inhibitory activity against C. histolyticum MMPs, achieving nearly complete inhibition (∼ 100%) at a concentration of 50 mg/mL. This result is consistent with previously reported values for other algal products (Martínez-Antequera et al., 2025). The simulated digestion of the diets demonstrated that a substantial MMP inhibitory effect was retained in the digestate following sequential exposure to gastric and intestinal enzymes (Table 1). These observations support the hypothesis that, under in vivo conditions, the extract may exert modulatory effects on intestinal MMP activity, potentially reflected in altered intestinal barrier functionality.
The inclusion of the algal extract in the diet significantly increased t issue resistance (Rt) in the anterior intestine of gilthead seabream from values of 241 in the control diet to 330 Ω·cm² , indicating enhanced epithelial barrier integrity. Although apparent permeability to Lucifer Yellow showed a trend toward lower values in fish fed the algal-supplemented diet, the differences were not statistically significant (Fig. 1).
The significant increase in epithelial resistance strongly suggests that bioactive compounds delivered through the algal extract played a pivotal role in enhancing intestinal tissue integrity compared to fish fed the control diet. This positive response may be attributed to either an alteration in tissue repair mechanisms or a reduction in inflammatory activity, potentially mediated by the MMP-inhibitory properties of the algal compounds. This finding may provide additional support for the incorporation of algal-derived bioactives as functional ingredients in aquaculture nutrition strategies.
References
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Acknowledgements
This work has been financed by project ALEHOOP (H2020-BBI-JTI-2019, proposal nº: 887259, UE) funded by the European Union under the Horizon Europe Programme.