DSpace Coleção:

Proteases da biomassa microalgal de Chlorococcum infusionum: obtenção, caracterizações físico-químicas e enzimáticas

2026-07-15T13:18:00Z

Título: Proteases da biomassa microalgal de Chlorococcum infusionum: obtenção, caracterizações físico-químicas e enzimáticas Autor(es): Ferreira, Vanessa Rodrigues Abstract: The biodiversity of microalgae remains a relatively underexplored resource. Its biomass represents a rich source of natural bioactive compounds, such as proteins and lipids, which hold potential applications in human and animal nutrition, as well as in the cosmetic, pharmaceutical, and fine chemical industries. Moreover, they exhibit promise in wastewater treatment and the bioremediation of harmful compounds. Specifically, microalgae belonging to the Chlorococcum genus are considered promising sources of macromolecules; however, their potential as protein sources remains inadequately investigated. In this context, the objective of this study was to extract, isolate, and evaluate the physicochemical and enzymatic characteristics of proteases present in the biomass of Chlorococcum infusionum microalgae produced in a photobioreactor. To achieve this, the microalgae were initially cultivated in Bold's Basal Medium (BBM) in an autotrophic closed-flat plate photobioreactor system. The resulting biomass was characterized and utilized in the preparation of aqueous extracts through a combination of lysis methods (EBCi). The microalgae cultivation yielded an average fresh biomass of 2.35 ± 0.03 g.L-1 after the 12th day of cultivation, with a composition rich in lipids (37.82%) and proteins (27.32%). For the isolation of proteases, EBCi underwent purification steps involving subsequent gel filtration chromatography using a Sephadex G-100 column and ion exchange chromatography using a DEAE-Sepharose™ Fast Flow column. The predominant peak obtained (designated as FPCiT-P3) was subjected to proteolytic activity assays against a nonspecific substrate (azocasein) and through zymograms. Optimal parameter determination assays, stability assays against temperature and pH variations, and structural stability evaluation through circular dichroism spectroscopy were conducted. FPCiT-P3 exhibited an optimum pH for activity around 7.0-9.0 and an optimum temperature of 50°C, being partially inhibited by PMSF and EDTA. The fraction showed low stability at acidic pHs and temperatures above 60°C after a 30-minute incubation. The circular dichroism spectrum of FPCiT-P3 indicated a predominance of β-sheet structure. Tipo: Dissertação

Toxicidade de nanopartículas de prata em modelos aquáticos e a interferência do meio na agregação da nanopartícula

2026-05-12T13:22:50Z

Título: Toxicidade de nanopartículas de prata em modelos aquáticos e a interferência do meio na agregação da nanopartícula Autor(es): Martins, Alexia Riquet Abstract: Silver nanoparticles (AgNP) are of high industrial interest, standing out for their antiseptic and antifungal properties and versatility in drug delivery. However, the expansion of their employability is directly linked to the increase in the disposal of these nanoparticles in the environment. Worldwide, regulations such as REACH and the EPA exist, although not specifically targeted, which cover the use, production, and disposal of nanoparticles. In Brazil, Bill 880/2019 (Legal Framework for Nanotechnology) stands out. Regardless of regulation, risk assessment of a nanomaterial to be used on a large scale is a basic requirement to ensure the safety of use. Aiming to standardize toxicity and experimental parameters, the OECD establishes standards, guidelines, and study models for testing materials. Among the model organisms for studies in aquatic environments and food chains are the microalgae Chlamydomonas reinhardtii, the microcrustacean Artemia salina, and Danio rerio embryos. These organisms were chosen for testing exposure to AgNP concentrations (12.5, 25, and 50 ppm) and testing the trophic level passage of nanoparticles through their synthesis, characterization of AgNP, and analysis of damage caused to the models using microscopy. The AgNP used was synthesized in collaboration and analyzed in UV-Vis, with two syntheses and one standard, and ten (10) times more concentrated; the exposure experiments in the three models were carried out by diluting the synthesized AgNP in the respective model media with analyses every 24 hours. The trophic level experiment consisted of exposing C. reinhardtii previously contaminated with AgNP concentrations to A. salina (nauplii I and II), and at the end of the experiments, the samples were collected and fixed for optical, scanning electron, and confocal microscopy. The results obtained showed that in the A. salinas medium, colloids tend to form, visible in the digestive tract of the specimen without causing morphological changes or mortality. In the homemade TAP medium, AgNP does not show the same tendency to aggregate, in addition to decreasing the replication speed of C. reinhardtii, which form colonies surrounded by a transparent secretion. D. rerio embryos showed 100% mortality, and the trophic level experiment showed no toxicity caused by the ingestion of C. reinhardtii contaminated by A. Salina. It is concluded that the exposure medium directly influences the toxicity of AgNP, which proved to be toxic when kept in its non-aggregated state. Tipo: Dissertação

Desenvolvimento de um protocolo de simulação por dinâmica molecular para avaliação estrutural e energética de variantes da l-asparaginase humana

2026-03-27T16:25:50Z

Título: Desenvolvimento de um protocolo de simulação por dinâmica molecular para avaliação estrutural e energética de variantes da l-asparaginase humana Autor(es): Silva, Juliana Meneses de Sena Abstract: L-asparaginase is an enzyme widely employed in the treatment of acute lymphoblastic leukemia (ALL) due to its ability to degrade L-asparagine (Asn), which is essential for the survival of leukemic cells. However, currently available formulations are derived from bacterial sources and are often associated with severe immune reactions. Although the human body possesses a homologous enzyme, hASNase1, its low catalytic activity precludes its direct clinical application. To overcome this limitation, mutations have been proposed to enhance its catalytic efficiency. This study aimed to develop a computational protocol, based on molecular dynamics (MD) simulations and structural analyses, to evaluate the structural and energetic effects of hASNase1 variants and to identify molecular features associated with increased catalytic activity. The systems were built based on the native structure of the enzyme and simulated under physiological conditions. Structural stability was assessed through root mean square deviation (RMSD), while interatomic interaction potentials (PII) and binding free energy calculations were used to analyze the interactions between hASNase1 variants and Asn. MM/PBSA calculations revealed increasing binding affinities in the order D87S < D84N < A186V < D87N, consistent with substrate residence times in the catalytic site. Experimentally, catalytic activity increases of 17× (D87S), 19× (D84N), 50× (A186V), and 52× (D87N) were observed compared to the wild-type enzyme, confirming the effectiveness of the DM + MM/PBSA protocol in identifying high-performance variants. In conclusion, the DM and MM/PBSA protocol elucidated the structural determinants underlying the experimentally observed activity gains in hASNase1 variants, identifying D87N and A186V as promising candidates for an optimized human L-asparaginase. Understanding the relationship between structure and catalytic activity in hASNase1 variants reinforces the use of in silico approaches in the rational development of safer and more effective biopharmaceuticals for the treatment of ALL. Tipo: Dissertação

Avaliação in vitro da atividade antimicrobiana e antibiofilme da quitosana de Ucides cordatus contra bacterias gram-negativas resistentes aos antimicrobianos

2026-03-04T15:46:06Z

Título: Avaliação in vitro da atividade antimicrobiana e antibiofilme da quitosana de Ucides cordatus contra bacterias gram-negativas resistentes aos antimicrobianos Autor(es): Carvalho, Francisco Sérvulo de Oliveira Abstract: Chitosan, derived from the deacetylation of chitin, has gained prominence due to its broad potential for applications in biomedical fields and the development of technologies in the food industry, particularly because of its antimicrobial activity. Traditionally extracted from the exoskeleton of crustaceans, the growing demand for this biopolymer has driven the exploration of the crab Ucides cordatus, found in mangrove ecosystems along the Atlantic coast. Thus, the present study aimed to evaluate the antimicrobial and antibiofilm activities of Ucides cordatus chitosan extracts against antimicrobial-resistant Gram-negative bactéria and to formulate an appropriate coating composition for application on coalho cheese. Antibacterial activity was determined by assessing the minimum inhibitory concentration and minimum bactericidal concentration at concentrations ranging from 5 to 0.009 mg/mL. Antibiofilm activity was evaluated using inhibition and eradication assays, quantified by the crystal violet staining method. To determine the appropriate coating formulation for coalho cheese, the wettability technique was employed through contact angle measurements on the surface. Toxicity assessments were conducted using the extracts and coating solutions in cellular models. With a degree of deacetylation of 92%, Ucides cordatus chitosan demonstrated good antimicrobial properties, showing inhibition of Gram-negative strains at bactericidal concentrations of 0.625 mg/mL, as well as inhibition of biofilm formation by approximately 98% in Pseudomonas aeruginosa and 86% in Escherichia coli resistant strains isolated from coalho cheese. These concentrations were found to be non-toxic. Regarding coating potential, based on the wettability technique on coalho cheese surfaces, the composition containing 2.0% Ucides cordatus chitosan with 0.2% glycerol was identified as the most suitable for coating application. The results obtained were comparable to those of commercially available chitosan, thus highlighting the biotechnological potential of Ucides cordatus chitosan as an antimicrobial agent in the biomedical field and the food industry. Tipo: Dissertação