Browsing by Author "Horta, Bruno"
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- Anti-biofilm potential of phenolic acids: the influence of environmental pH and intrinsic physico-chemical propertiesPublication . Silva, Sara; Costa, Eduardo M.; Horta, Bruno; Calhau, Conceição; Morais, R. M. S. C.; Pintado, M. E.Phenolic acids are a particular group of small phenolic compounds which have exhibited some anti-biofilm activity, although the link between their activity and their intrinsic pH is not clear. Therefore, the present work examined the anti-biofilm activity (inhibition of biomass and metabolic activity) of phenolic acids in relation to the environmental pH, as well as other physico-chemical properties. The results indicate that, while Escherichia coli was not inhibited by the phenolic acids, both methicillin resistant Staphylococcus aureus and methicillin resistant Staphylococcus epidermidis were susceptible to the action of all phenolic acids, with the pH playing a relevant role in the activity: a neutral pH favored MRSE inhibition, while acidic conditions favored MRSA inhibition. Some links between molecular polarity and size were associated only with their potential as metabolic inhibitors, with the overall interactions hinting at a membrane-based mechanism for MRSA and a cytoplasmic effect for MRSE.
- Antitumor effect of chalcone derivatives against human prostate (LNCaP and PC-3), cervix HPV-Positive (HeLa) and lymphocyte (Jurkat) cell lines and their effect on macrophage functionsPublication . Horta, Bruno; Freitas-Silva, Joana; Silva, Jani; Dias, Francisca; Teixeira, Ana Luísa; Medeiros, Rui; Cidade, Honorina; Pinto, Madalena; Cerqueira, FátimaChalcones are synthetic and naturally occurring compounds that have been widely investigated as anticancer agents. In this work, the effect of chalcones 1–18 against the metabolic viability of cervical (HeLa) and prostate (PC-3 and LNCaP) tumor cell lines was tested, to compare the activity against solid and liquid tumor cells. Their effect was also evaluated on the Jurkat cell line. Chalcone 16 showed the highest inhibitory effect on the metabolic viability of the tested tumor cells and was selected for further studies. Recent antitumor therapies include compounds with the ability to influence immune cells on the tumor microenvironment, with immunotherapy being one actual goal in cancer treatment. Therefore, the effect of chalcone 16 on the expression of mTOR, HIF-1α, IL-1β, TNF-α, IL-10, and TGF-β, after THP-1 macrophage stimulation (none, LPS or IL-4), was evaluated. Chalcone 16 significantly increased the expression of mTORC1, IL-1β, TNF-α, and IL-10 of IL-4 stimulated macrophages (that induces an M2 phenotype). HIF-1α and TGF-β were not significantly affected. Chalcone 16 also decreased nitric oxide production by the RAW 264.7 murine macrophage cell line, this effect probably being due to an inhibition of iNOS expression. These results suggest that chalcone 16 may influence macrophage polarization, inducing the pro-tumoral M2 macrophages (IL-4 stimulated) to adopt a profile closer to the antitumor M1 profile.
- Circular economy: from industrial side streams to nutrient-rich zeolites for agricultural usePublication . Horta, Bruno; Oliveira, Carla; Burgal, João; Oliveira, Catarina
- Effect of 1-carbaldehyde-3,4-dimethoxyxanthone on prostate and HPV-18 positive cervical cancer cell lines and on human THP-1 macrophagesPublication . Medeiros, Rui; Horta, Bruno; Freitas-Silva, Joana; Silva, Jani; Dias, Francisca; Sousa, Emília; Pinto, Madalena; Cerqueira, FátimaXanthone derivatives have shown promising antitumor properties, and 1-carbaldehyde-3,4-dimethoxyxanthone (1) has recently emerged as a potent tumor cell growth inhibitor. In this study, its effect was evaluated (MTT viability assay) against a new panel of cancer cells, namely cervical cancer (HeLa), androgen-sensitive (LNCaP) and androgen-independent (PC-3) prostate cancer, and nonsolid tumor derived cancer (Jurkat) cell lines. The effect of xanthone 1 on macrophage functions was also evaluated. The effect of xanthone 1-conditioned THP-1 human macrophage supernatants on the metabolic viability of cervical and prostate cancer cell lines was determined along with its interference with cytokine expression characteristic of M1 profile (IL-1 ≤ β; TNF-α) or M2 profile (IL-10; TGF-β) (PCR and ELISA). Nitric oxide (NO) production by murine RAW264.7 macrophages was quantified by Griess reaction. Xanthone 1 (20 µM) strongly inhibited the metabolic activity of the cell lines and was significantly more active against prostate cell lines compared to HeLa (p < 0.05). Jurkat was the cell most sensitive to the effect of xanthone 1. Compound 1-conditioned IL-4-stimulated THP-1 macrophage supernatants significantly (p < 0.05) inhibited the metabolic activity of HeLa, LNCaP, and PC-3. Xanthone 1 did not significantly affect the expression of cytokines by THP-1 macrophages. The inhibiting effect of compound 1 observed on the production of NO by RAW 264.7 macrophages was moderate. In conclusion, 1-carbaldehyde-3,4-dimethoxyxanthone (1) decreases the metabolic activity of cancer cells and seems to be able to modulate macrophage functions.
- Fractionating of synthetic biology waste stream into nutrient rich extract for cosmetic industryPublication . Horta, Bruno; Cálix, Carla; Burgal, João; Oliveira, CatarinaAmyris is a world-leading manufacturer of sustainable ingredients producing environmentally friendly products. Biofene® (or trans-β-farnesene) is a synthetic biology derived product using yeast fermentation of sugarcane. This production originates spent broth, an aqueous waste stream typically applied to land by fertirrigation. On one hand, spent broth contains recoverable added-value compounds; on the other hand, an improper disposal of this waste stream could result in negative effects and give rise to environmental concerns. Hence the importance of implementing the bio-waste refinery and circular bio-economy concepts for recovering high-value bio-products. Cosmetic industries are committed to find natural, sustainable, and functional ingredients meeting consumer’s and entities rigorous demands. Spent broth may be a source of such ingredients. Thus, the objective of this work was to assess the fractionation of the spent broth stream from Biofene® production to obtain a concentrated nutrient rich extract with potential application on cosmetics. The spent broth’s was pretreated for mineral recovery, using a chemical free process developed within the Alchemy Project. The pre-treated spent broth was then fractionated in a cascade membrane filtration process, in an attempt to recover a rich peptide and minerals fractions. This process comprised three steps: Microfiltration (MF), ultrafiltration (UF), and nanofiltration (NF). The composition of the obtained fractions was monitored in terms of total proteins and minerals contents, amino acids profile, and antioxidant capacity. All experiments were done in a crossflow filtration unit (Sepa CF, Sterlitech), coupled to a pump (G13-X, Hidra-Cell®) applying a feed recirculation of ca. 250 L/h. Microfiltration aimed to the clarification of the pre-treated spent broth, removing suspended mineral and organic particles; a Toray’s sanitary TMF membrane presented a total solids rejection factor of 51.1%, and rejection factors towards total proteins and minerals of 60.5% and 26.4%, respectively. The antioxidant capacity was higher in the obtained permeate than in the pre-treated spent broth (560 ±59 and 317 ±56 μmol TE/g by ORAC, respectively). The subsequent ultrafiltration of the MF permeate with a 5 kDa MWCO membrane (Toray Sanitary TUF) had rejection factors towards total proteins of 5.9% and of 20.0% towards minerals. The retained fraction had an interesting antioxidant capacity (685 ±64 μmol TE/g by ORAC), amino acids content (61.9 ±0.9 mg/g-dw), comprising mainly glutamic acid and aspartic acid (27 and 26 wt%, respectively), and minerals (103.6 ±5.6 mg/g-dw), mainly calcium (ca. 65 wt%). The UF permeate was further filtrated using a Toray’s sanitary NF membrane, which retained 61% of total solids, thus being a concentration step. The retained fraction was similar to that retained on the UF membrane, presenting higher antioxidant capacity (738 ±64 μmol TE/g by ORAC) and minerals content (232.4 ±17.7 mg/g-dw, mainly calcium ca. 65 wt%). The fractions resultant from the cascade filtration process, namely the UF and the NF retentates, presented interesting characteristics for cosmetics applications, particularly significant amino acids and minerals contents, and high antioxidant capacity. Indeed, amino acids are highly desired in cosmetics for their capacity to improve skin and hair quality [1]; mineral based salts, including calcium and magnesium act as absorbents, abrasives, and astringent agents, promoting skin softening and moisture [2]. Powerful antioxidant and free radical scavenging properties are also extremely valuable in anti-aging and skin protection products [3]. This work demonstrates a potential practical valorization for developing increased value bio-derived products from the Biofene® fermentation spent broth stream. The product preliminary characterization reveals interesting nutrients and bioactivities, suggesting a potential application in cosmetic industry.
- In vivo study of the effect of sugarcane bagasse lignin supplementation on broiler chicken diet as a step to validate the established chicken gastrointestinal tract in vitro modelPublication . Carvalho, Nelson Mota de; Souza, Carla Giselly de; Costa, Célia Maria; Castro, Cláudia; Fangueiro, Joana F.; Horta, Bruno; Outor-Monteiro, Divanildo; Teixeira, José; Mourão, José Luís; Pinheiro, Victor; Amaro, Ana L.; Costa, Patrícia Santos; Oliveira, Catarina S. S.; Pintado, Manuela Estevez; Oliveira, Diana Luazi; Madureira, Ana RaquelSince the global restrictions on antibiotics in poultry systems, there has been a growing demand for natural and sustainable feed additives for disease prevention and poultry nutrition. This study evaluated the effects of incorporating sugarcane bagasse (SCB) lignin into broiler chicken diets. The performance of the chickens, including body weight, feed intake, and mortality, as well as intestinal histomorphometry, and cecum content pH, microbiota, and volatile fatty acids were assessed. In addition, we also aimed to validate an in vitro gastrointestinal tract (GIT) model developed by Carvalho et al. (2023). One hundred and eight 1-day-old Ross 308 chicks were randomly and equally divided into two groups. The first group was fed a basal diet (BD group), while the second group was fed a basal diet supplemented with 1% (w/w) SCB lignin (BD + SCB lignin group) for 36 days. The in vivo conditions of the chicken GIT were replicated in an in vitro model. In the in vivo study, SCB lignin increased cecum acetate and butyrate levels while reducing Bifidobacterium and Enterobacteriaceae, without affecting productivity (body weight, feed intake, and mortality). The in vitro assessment reflected microbiota trends observed in vivo, although without statistical significance. The divergence in organic acid production between the in vivo and in vitro conditions likely resulted from issues with inoculum preparation. This study demonstrates that SCB lignin incorporation positively influences cecal microbiota composition without impacting the animals’ productivity and physiology, suggesting its potential as a functional feed additive. For a more reliable in vitro model, adjustments in inoculum preparation are necessary.
- Omega-3- and resveratrol-loaded lipid nanosystems for potential use as topical formulations in autoimmune, inflammatory, and cancerous skin diseasesPublication . Caldas, Ana R.; Catita, José; Machado, Raul; Ribeiro, Artur; Cerqueira, Fátima; Horta, Bruno; Medeiros, Rui; Lúcio, Marlene; Lopes, Carla M.Resveratrol (RSV) and omega 3 (!3), because of their biological favorable properties, have become subjects of interest for researchers in dermocosmetic and pharmaceutical industries; however, these bioactives present technological limitations that hinder their effective delivery to the target skin layer. To overcome the stability and skin permeation limitations of free bioactives, this work proposes a combined strategy involving two different lipid nanosystems (liposomes and lipid nanoparticles) that include ω3 in their lipid matrix. Additionaly, RSV is only encapsulated in liposomes that provid an adequate amphiphilic environment. Each formulation is thoroughly characterized regarding their physical–chemical properties. Subsequently, the therapeutic performance of the lipid nanosystems is evaluated based on their protective roles against lipid peroxidation, as well as inhibition of cicloxygenase (COX) and nitric oxid (NO) production in the RWA264.7 cell line. Finally, the lipid nanosystems are incorporated in hydrogel to allow their topical administration, then rheology, occlusion, and RSV release–diffusion assays are performed. Lipid nanoparticles provide occlusive effects at the skin surface. Liposomes provide sustained RSV release and their flexibility conferred by edge activator components enhances RSV diffusion, which is required to reach NO production cells and COX cell membrane enzymes. Overall, the inclusion of both lipid nanosystems in the same semisolid base constitutes a promising strategy for autoimmune, inflammatory, and cancerous skin diseases.
- Valorization of spent sugarcane fermentation broth as a source of phenolic compoundsPublication . Oliveira, Carla Maria; Horta, Bruno; Leal, Tânia; Pintado, Manuela; Oliveira, Catarina S. S.A methodology based on a solid phase extraction (SPE) was optimized for the recovery of phenolic compounds from the spent fermentation broth generated from Biofene® (trans-β-farnesene) production. For this purpose, two resins (XAD-2 and HP-20) and three desorption solutions (water, 50/50 ethanol/water, and ethanol) were tested. The most efficient resin revealed to be the HP-20, using ethanol as desorption solution, reaching an overall total phenolic compound recovery of ca. 80% when 6 BV (bed volume) of both feed and ethanol were applied. The optimization of the resin’s process cycle pointed to 15 BV feed to be treated per cycle and using the same volume of ethanol in the desorption step, with no need for an extra resin regeneration step, stably yielding 48% total phenolic compound recovery from the spent broth for at least 4 cycles, translating into 60 BV of feed being treated per BV of resin, and with the resin being still perfectly active. The extract was characterized using LC−ESI−UHR−QqTOF−MS, and a total of 82 and 15 compounds were identified, in negative and positive ionization modes, respectively. Organic acids were the main class of compounds identified in the phenolic-rich extract, followed by phenolic compounds, saccharides, peptides or amino acids and vitamins. Additionally, the extract revealed a significant antioxidant capacity (914.1 ± 51.6 and 2764.5 ± 142.8 µmol Trolox equivalents/g-dw, respectively, with ABTS and ORAC methodologies), which might be interesting for a wide variety of applications.