Browsing by Author "Brassesco, M. E."
Now showing 1 - 4 of 4
Results Per Page
Sort Options
- Bioactive potential and chromatographic characterization of body mucus from Portugal coastal fish halobatrachus didactylusPublication . Cunha, M. Fernandez; Coscueta, E. R.; Brassesco, M. E.; Almada, F.; Gonçalves, D.; Pintado, M.The body mucus in fish provides a stable physical or chemical barrier against invading pathogens. In mucus, antimicrobial peptides are secreted as a response to immune stimulation. Studies have shown antimicrobial activity against multidrug-resistant pathogens and low toxicity to eukaryotic cells. Previously, body mucus from five captive Halobatrachus didactylus individuals was collected. We aim to explore the mucus studying molecules with bioactive potential. Size exclusion highperformance liquid chromatography (SEHPLC) analyses were performed on the five body mucus samples showing a similar molecular size distribution with a maximum peak of ca. 800 Da. These five mucus samples were pooled to assess the following bioactivities: antioxidant (ABTS and ORAC), antimicrobial (minimal inhibitory concentration), and cytotoxicity (Caco2 and HaCaT human cell lines). The protein content in the mucus, determined by the bicinchoninic acid methodology, was 16836 ± 1020 µg BSA/mL. The antioxidant activity resulted in 268 ± 11 µmol TE/g mucus protein for ABTS and 306 ± 11 µmol TE/g mucus protein for ORAC. The antibacterial activity was assessed against five pathogenic bacteria: Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, within minimal inhibitory concentrations of 421 to 105 µg mucus protein/mL. Moreover, the mucus showed noncytotoxic for Caco2 cells in concentrations between 196 to 25 µg mucus protein/mL, while it showed cytotoxicity for HaCaT cells. In the future, liquid chromatographytandem mass spectrometry (LCMS/MS) analysis will be performed to determine the molecules behind these bioactivities, namely antimicrobial peptides.
- Collagen-based bioactive hydrolysates production from Blue Shark skinPublication . Brassesco, M. E.; Coscueta, E. R.; Pintado, Maria ManuelaPortugal is the country with the highest levels of fish consumption in the world with an average of 59 kg per capita year. As a result, a huge amount of fish residues is produced, of which up to 30% may be skin and bone. Is well-known that fish skin has more than 80% of its total protein content as collagen. This structural protein is present in the extracellular matrices of animals and has widely applications in food, cosmetic, biomedical, and pharmaceutical industries. For decades, the main industrial exploitations of collagen have been bovine and porcine origins. However, due to religious beliefs and infectious diseases, other collagen sources are being debated. In this regard, the use of collagen with marine origin is being considered highly attractive by the industry as an important alternative source. Blue shark skin, which is an abundant by-product of Portugal processing industry rich in collagen, represents a potential source of this protein. Thus, a collagen hydrolysate was obtained from Blue Shark skin with bromelain, a plant enzyme, for the first time. As a result, low molecular weight hydrolysates were obtained with a protein concentration of 0.5%(p/p). The antioxidant activity was analysed in vitro, obtaining a result of 4.077 µmol Trolox Equivalent/mg protein. The ACE inhibition activity was also analysed obtaining an IC50 of 14.3 ug protein/mL at 1h hydrolysis. Therefore, it was possible to obtain an ingredient (hydrolysed collagen) that is currently required by industries such as cosmetic, food and nutraceutical, with high potential antioxidant and antihypertensive activities. For this, it was possible to apply a sustainable process that consumes less time and uses an alternative and abundant raw material.
- Collagen-based bioactive hydrolysates production from blue shark skinPublication . Brassesco, M. E.; Coscueta, E. R.; Pintado, M.Portugal is the country with the highest levels of fish consumption in the world with an average of 59 kg per capita year. As a result, a huge amount of fish residues is produced, of which up to 30% may be skin and bone. Is well-known that fish skin has more than 80% of its total protein content as collagen. This structural protein is present in the extracellular matrices of animals and has widely applications in food, cosmetic, biomedical, and pharmaceutical industries. For decades, the main industrial exploitations of collagen have been bovine and porcine origins. However, due to religious beliefs and infectious diseases, other collagen sources are being debated. In this regard, the use of collagen with marine origin is being considered highly attractive by the industry as an important alternative source. Blue shark skin, which is an abundant by-product of Portugal processing industry rich in collagen, represents a potential source of this protein. Thus, a collagen hydrolysate was obtained from Blue Shark skin with bromelain, a plant enzyme, for the first time. As a result, low molecular weight hydrolysates were obtained with a protein concentration of 0.5%(p/p). The antioxidant activity was analysed in vitro, obtaining a result of 4.077 µmol Trolox Equivalent/mg protein. The ACE inhibition activity was also analysed obtaining an IC50 of 14.3 ug protein/mL at 1h hydrolysis. Therefore, it was possible to obtain an ingredient (hydrolysed collagen) that is currently required by industries such as cosmetic, food and nutraceutical, with high potential antioxidant and antihypertensive activities. For this, it was possible to apply a sustainable process that consumes less time and uses an alternative and abundant raw material.
- P82 Peptide fraction identification by SE-HPLC and LC-MS/MS analysis of the body mucus from Portugal coastal fish Halobatrachus didactylusPublication . Cunha, M.; Coscueta, E. R.; Brassesco, M. E.; Almada, F.; Gonçalves, D.; Pintado, M. ManuelaThe mucus covers the fish's body, working as a protective barrier. Besides physical protection, mucus provides molecules that protect the fish from pathogens damaging 1,2. These include antimicrobial peptides secreted in the mucus, which play an essential role in defense against microbial pathogens since these belong to the innate immune system2,3. In this study, two adult Halobatrachus didactylus individuals were captured from the wild in Sesimbra. Then, mucus collection was performed by scraping the dorsal-lateral body of the fish with a sponge. Our objective was the identification of new peptides with bioactive potential in mucus samples by chromatography analysis. Size exclusion highperformance liquid chromatography (SE-HPLC) analysis performed on mucus samples from the two individuals revealed a similar profile with an intense highlight peak which resulted in a distribution of about 775 Dalton. With interest in that peak, the two mucus samples were pooled for fractionation by SEC. The resulting fraction was analyzed by liquid chromatography-tandem mass spectrometry (LCMS/MS) to identify the most probable peptide sequences. Identification from databases did not provide reliable results, indicating a lack of information on the matrix analyzed. We resorted to de novo sequencing with good results using PEAKS Studio software. Five identified peptides were selected according to their bioactivities predicted in silico. Furthermore, the five identified peptides were synthesized, and the molecular size was validated by SE-HPLC analysis. Overall, this chromatographic approach enabled the identification of promising peptides, which bioactivities will be evaluated in vitro in future work.