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  • Enzymatic soy protein hydrolysis: a tool for biofunctional food ingredient production
    Publication . Coscueta, Ezequiel R.; Campos, Débora A.; Osório, Hugo; Nerli, Bibiana B.; Pintado, Manuela
    This work aimed to evaluate the digestive stability of the peptides previously identified from a Corolase PP soy protein hydrolysate (SPH) and to respond to the uncertainty about the merit of controlled hydrolysis. For this purpose, we applied an empirical and theoretical analysis, determining peptide sequences, oxygen radical scavenging (ORAC) and ACE inhibitory (iACE) activities, and the effect of hydrolysis on solubility. Results showed that during digestion most of SPH peptides were degraded as smaller ones. However, both SPH bioactivities improved significantly after digestion (3.9 ± 0.1 μmol TE/mg protein for ORAC and IC50 = 52 ± 4μg protein/mL for iACE) with similar values for soy protein isolate (SPI). With respect to solubility, the controlled hydrolysis considerably increased this functional property. In conclusion, the results indicated that controlled enzymatic hydrolysis of SPI with Corolase PP produced an ingredient more apt to be incorporated in certain nutritional or nutraceutical formulations.
  • Antihypertensive effect of spent brewer yeast peptide
    Publication . Amorim, M.; Marques, C.; Pereira, J. O.; Guardão, L.; Martins, M. J.; Osório, H.; Moura, D.; Calhau, C.; Pinheiro, H.; Pintado, M.
    Numerous studies have investigated dietary approaches to prevent chronic lifestyle-related diseases, including hypertension. Spent brewer's yeast is the second largest byproduct originated by the brewing industry and it deserves considerable attention because of its high nutritional value, ca. 40% of its dry mass is rich in protein which can be hydrolyzed into biologically active peptides. To upgrade this byproduct, the aim of this study was initially in vitro assessment of biological properties, e.g. ACE inhibition and antioxidant activity, and then, the in vivo effect in short-term oral antihypertensive effect of hydrolyzed yeast fractions on a well characterized model to study hypertension - Spontaneously Hypertensive Rats (SHR). Here, it was demonstrated that the fraction with molecular weight below 3 kDa containing tri and tetra- peptides with hydrophobic amino acid residues - SPQW, PWW and RYW, causes the most noticeable decrease in systolic, diastolic and mean blood pressure of SHR and shows highest antioxidant effect. These properties highlight the potential use of yeast extract as nutraceutical or functional food ingredient for the management and treatment of hypertension with antioxidant effect.
  • Differentiation of Bacillus pumilus and Bacillus safensis Using MALDI-TOF-MS
    Publication . Branquinho, Raquel; Sousa, Clara; Lopes, João; Pintado, Manuela E.; Peixe, Luísa V.; Osório, Hugo
    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) despite being increasingly used as a method for microbial identification, still present limitations in which concerns the differentiation of closely related species. Bacillus pumillus and Bacillus safensis, are species of biotechnological and pharmaceutical significance, difficult to differentiate by conventional methodologies. In this study, using a well-characterized collection of B. pumillus and B. safensis isolates, we demonstrated the suitability of MALDI-TOF-MS combined with chemometrics to accurately and rapidly identify them. Moreover, characteristic species-specific ion masses were tentatively assigned, using UniProtKB/Swiss-Prot and UniProtKB/TrEMBL databases and primary literature. Delineation of B. pumilus (ions at m/z 5271 and 6122) and B. safensis (ions at m/z 5288, 5568 and 6413) species were supported by a congruent characteristic protein pattern. Moreover, using a chemometric approach, the score plot created by partial least square discriminant analysis (PLSDA) of mass spectra demonstrated the presence of two individualized clusters, each one enclosing isolates belonging to a species-specific spectral group. The generated pool of species-specific proteins comprised mostly ribosomal and SASPs proteins. Therefore, in B. pumilus the specific ion at m/z 5271 was associated with a small acid-soluble spore protein (SASP O) or with 50S protein L35, whereas in B. safensis specific ions at m/z 5288 and 5568 were associated with SASP J and P, respectively, and an ion at m/z 6413 with 50S protein L32. Thus, the resulting unique protein profile combined with chemometric analysis, proved to be valuable tools for B. pumilus and B. safensis discrimination, allowing their reliable, reproducible and rapid identification.
  • Impact of thermal treatment and hydrolysis by Alcalase and Cynara cardunculus enzymes on the functional and nutritional value of Okara
    Publication . Voss, Glenise B.; Osorio, Hugo; Valente, Luís M. P.; Pintado, Manuela E.
    Enzymatic hydrolysis of dried okara (autoclaved and non-autoclaved) with Alcalase (AL) and Cynara cardunculus extract (CY) was studied, assessing the impact of heat treatment and hydrolysis on potential antioxidant and antihypertensive activities of final hydrolysates. This study showed that the thermal treatment (sterilization at 121 °C, 1 atm and 15 min) facilitated the enzymatic access to substrate and increased the degree of hydrolysis (DH), especially for AL (37.9%) when compared to CY (3.6%). The antioxidant activity of dried Okara (either autoclaved or not) when hydrolysed with AL was higher (4.2 mg Trolox/mL) than that observed for CY. Additionally, the potential ACE-inhibitory activity was high for samples hydrolysed with both enzymes, however the highest ACE inhibition was also found for AL (IC50 = 9.97 µg/mL). This study allowed the identification of new peptide sequences in dried okara hydrolysed with both enzymes, and some sequences that can explain their bioactivities. The results indicate that dried okara hydrolysates can either be used as functional ingredient or as food supplement for blood pressure lowering or antioxidant applications in the future.