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Artificial Manganese Metalloenzymes with Laccase-like Activity: Design, Synthesis, and Characterization
No full textLaccase is an oxidase of great industrial interest due to its ability to catalyze oxidation processes of phenols and persistent organic pollutants. However, it is susceptible to denaturation at high temperatures, sensitive to pH, and unstable in the presence of high concentrations of solvents, which is a issue for industrial use. To solve this problem, this work develops the synthesis in an aqueous medium of a new Mn metalloenzyme with laccase oxidase mimetic catalytic activity. Geobacillus thermocatenulatus lipase (GTL) was used as a scaffold enzyme, mixed with a manganese salt at 50 °C in an aqueous medium. This leads to the in situ formation of manganese(IV) oxide nanowires that interact with the enzyme, yielding a GTL-Mn bionanohybrid. On the other hand, its oxidative activity was evaluated using the ABTS assay, obtaining a catalytic efficiency 300 times higher than that of Trametes versicolor laccase. This new Mn metalloenzyme was 2 times more stable at 40 °C, 3 times more stable in the presence of 10% acetonitrile, and 10 times more stable in 20% acetonitrile than Novozym 51003 laccase. Furthermore, the site-selective immobilized GTL-Mn showed a much higher stability than the soluble form. The oxidase-like activity of this Mn metalloenzyme was successfully demonstrated against other substrates, such as l-DOPA or phloridzin, in oligomerization reactions
Contribution to the knowledge of the temporal and spatial distribution of species from the genus Acartia (copepoda) in the Southern Adriatic
No full textU radu smo prikazali sezonska istraživanja planktonske zajednice Copepoda u Južnom
Jadranu na dve postaje za vrste iz roda Acartia. Podaci o horozontalnom rasprostranjenju
planktona dobijeni su iz materijala koji je sezonski prikupljan u toku jedne godine u
otvorenim vodama Južnog Jadrana. Zaključak je da su sezonske razlike u horizontalnoj
distribuciji povezane s dinamikom hidrografskih uslova Jadranskog mora.In this contribution, we present seasonal research on the plankton community of Copepoda
in the Southern Adriatic at two stations for species from the genus Acartia. Data on the
horizontal distribution of plankton were obtained from material collected seasonally over
one year in the open waters of the South Adriatic. The conclusion is that seasonal variations
in horizontal distribution are linked to the dynamics of the hydrographic conditions of the
South Adriatic Sea
Sr/Mg – doped bioceramic scaffolds for biomedical application
No full textBone is a mineralized connective tissue with remarkable self-healing capability. However, in the
presence of large bone defects (≥ 2.5 cm), bone self-recovery is not efficient, necessitating surgical intervention and the
introduction of a bone substitute. Hydroxyapatite (HAP) is a widely investigated material for bone tissue engineering
(BTE) due to its similarity to the biological apatite found in bones and teeth. Mesoporous bioactive glasses (MBAGs),
quickly bind to surrounding tissues and release ions promoting the formation of new bone [1]. The silica from glass
enhances angiogenesis, which is a pivotal consideration given the high vascularization level of this tissue [2]. Ion-doping
approach of both HAP and MBAG particles has gained great attention due to the ability of therapeutical ions to stimulate
a certain cell response. The project aims to develop and characterize bioceramic scaffolds based on a combination of
Sr/Mg-doped HAP and MBAG, thereby promoting osteogenesis and creating a favourable environment for the
proliferation of endothelial cells.ExcellMater Conference 2024: Innovative Biomaterials for Novel Medical Devices, Belgrade, Serbia, April 10-12, 202
Visualization of Extracellular Vesicles Derived from Dental Mesenchymal Stem/Stromal Cells Using SEM and Immunogold TEM Analyses
No full textExtracellular vesicles (EVs) derived from mesenchymal stem/stromal cells (MSCs)
have emerged as promising therapeutic agents in regenerative medicine. EVs are nanosized,
membrane-limited vesicles containing protein and RNA cargo, released by various cell types
for intercellular communication. It has been shown that EVs derived from MSCs play a significant role in the tissue regeneration process [1]. Dental MSCs such as human stem cells from
the dental pulp of deciduous teeth (SHED cells) are easily accessible MSCs with high proliferative capacity [2]. The objective of this study was to isolate and characterize EVs derived from
SHED, to explore their potential utility in regenerative medicine applications.
SHED cells were isolated from deciduous teeth from three donors, and their morphology and osteogenic potential were assessed through optical microscopy and Alizarin Red
staining of cells cultured in osteogenic cell media for 21 days. SHED cells were then cultured in EVs depleted cell culture media and EVs were isolated from conditioned cell media
by differential ultracentrifugation. The isolated EVs pellets underwent further characterization, including Western Blot for the detection of CD63 surface marker, Nanoparticle Tracking
Analysis (NTA) for the size distribution, Bicinchoninic acid assay (BCA) to determine protein
concentration and electron microscopy analyses for visualization. For the scanning electron
microscopy (SEM) analysis EVs were fixed with glutaraldehyde, placed on aluminum stubs,
and gold-sputterd for analysis. For immuno-gold transmission electron microscopy (TEM)
analysis, samples were fixed on carbon-coated copper TEM grids, followed by immunoreactive
binding of streptavidin-gold nanoparticles (10 nm) to the CD63-positive population of EVs.
Optical microscopy unveiled typical MSC morphology, exhibiting elongated spindle,
cuboidal, and polygonal shapes consistent with 2D cell culture isolates. Alizarin Red staining,
observed through optical microscopy, confirmed mineral matrix deposition by cells treated
with osteogenic supplementation, indicating their differentiation capacity. Characterization of
isolated EVs revealed the presence of the transmembrane protein CD63 by Western blot analysis. Additionally, the median size of the isolated EVs was 156 nm measured via NTA, while
BCA analysis determined a total protein concentration of 200 µg/ml. The SEM analysis confirmed the presence of EVs in the isolates (Figure 1), capturing individual particles with spherical morphology and sizes ranging from 70 nm to 250 nm. Immunogold TEM analysis revealed
cup-shaped and spheroid morphologies, characteristic of EVs’ appearance when using TEM, as well as positive immunogold-labeling with transmembrane extracellular protein CD63. The
presence of gold nanoparticles bound to the majority of EVs indicated a significant number of
CD63-positive vesicles.
In conclusion, EVs derived from SHED cells were successfully isolated and characterized. The EVs exhibited characteristics typical for MSC EVs, including CD63 positivity [3],
appropriate size distribution, protein content, and morphology [4], [5]. The potential utility of
these EVs in regenerative medicine will need to be confirmed by assessing their function [6]
Physicochemical Characterization of Polysaccharide–Protein Carriers with Immobilized Yeast Cells Obtained Using the Freeze-Drying Technique
No full textNew techniques for the immobilization of yeast cells have the potential for enhancement of the beer production process. Alongside conventional materials for cell immobilization, there is a rising trend toward polysaccharide–protein systems. This study focused on the immobilization of yeast cells (Saccharomyces pastorianus) via a freeze-drying process. The whey protein isolate, sodium alginate, maltodextrin, inulin, and their blends were used for carrier preparation. The effect of a 1.0% inulin solution as a cryoprotectant on the viability of the yeast cells after the freeze-drying process was also analyzed. The powders were assessed for cell viability, moisture content, water activity, solubility, particle size, and surface charge. According to the results, the addition of whey proteins reduced the moisture content, while solubility did not significantly decrease. Samples containing whey protein showed slight diameter variations. The negative surface charge observed in all samples, especially the control, indicates a cell’s tendency to aggregate, demonstrated by optical microscopy. SEM micrographs showed successful cell immobilization in polysaccharide–protein carriers. Furthermore, inulin and whey protein addition enhanced cell protection during the immobilization of cells. The freeze-drying technique demonstrates efficacy in immobilization of yeast cells, indicating its potential for applications in the food and beverage industry
Pro-Osteogenic Properties of Titanium-Based Scaffold Combined with SHED-Derived EVs Embedded in Collagen
No full textBiomaterials for the restoration of large bone defects, such as 3D Ti-based scaffolds, have
good mechanical properties; however, they do not induce bone regeneration. The aim of this
study was to encapsulate extracellular vesicles (EVs) derived from stem cells from human
exfoliated deciduous teeth (SHED cells) in a 3D-printed Ti6Al4V scaffold and analyze their
combined osteogenic potential. SHED cells were isolated from deciduous teeth, and their MSC
properties were characterized using fluorescence-activated cell sorting (FACS). EVs were
isolated from SHED-conditioned media using differential ultracentrifugation, and
characterized with SEM, TEM, NTA, and ONI. The size distribution, morphology, and
presence of tetraspanin markers CD9, CD63, and CD81 confirmed successful isolation of
SHED-EVs. EVs were further encapsulated in collagen hydrogel, and their release rate was
analyzed using fluorescence-based NTA. In the first three weeks, around 50% of EVs were
released from collagen in PBS. Ti6Al4V scaffolds were coated with graded silicate glasses
and HAP powder doped with silver. Their cytocompatibility was tested on SHED cells,
showing a higher cell metabolic rate in coated samples compared to bare Ti. The osteogenic
induction of EVs + Ti scaffold was tested on SHED cells, and gene expression was analyzed
using qRT-PCR. Both RUNX-2 and BMP-2 genes were upregulated after 10 days of treatment,
indicating pro-osteogenic properties of material filled with SHED-EVs. EV-loaded Ti6Al4V
scaffold shows potential for bone regeneration applications in large defects
Harnessing the power of green and rooibos tea aqueous extracts for obtaining colored bioactive cotton and cotton/flax fabrics intended for disposable and reusable medical textiles
No full textThis study harnesses the potential of green and rooibos tea (GT and RT) aqueous extracts for obtaining colored bioactive cotton and cotton/flax fabrics with intended applications in medical textiles. The chemical characterization of the tea aqueous extracts was conducted using LC–HRMS/MS analysis, resulting in the detection of 129 bioactive compounds. GT demonstrates 2.2 times higher total phenolic content, a 14.7% lower total flavonoid content, and 3 times higher reducing power than RT. Both extracts exhibit excellent antioxidant activity (> 99.8%) and antibacterial activity (99.99%) against both tested bacteria, E. coli and S. aureus. Cotton and cotton/flax fabrics functionalized with GT or RT display outstanding antioxidant (99.63–100%) and antibacterial activity against S. aureus (90.95–99.33%), and high color strength values (5.48–11.08). The cytotoxicity assay confirmed the non-cytotoxic nature of 100% cotton fabric functionalized with GT. This sample additionally demonstrated an antibacterial reduction against E. coli and S. aureus higher than 99% and the highest release of bioactive compounds rendering it highly suitable for disposable medical textiles-wound dressings. To address the shortcomings of functionalized fabrics observed after washing, including decreased antioxidant activity (55.8–81.0%), diminished bacterial reduction, and reduced color strength values (0.80–1.36), copper-based nanoparticles (CuNPs) were biosynthesized in situ on their surfaces utilizing GT and RT aqueous extracts as reducing agents. The successful fabric decoration with CuNPs was proven by quantifying Cu2+ uptake, and characterization of the surface chemical composition and morphology of CuNPs. Colored CuNPs-decorated cotton and cotton/flax fabrics exhibited excellent antioxidant (> 98.28%) and antibacterial (99.99%) activity that remained almost unchanged after washing (94.44–98.90% and 99.99%, respectively). These fabrics are non-cytotoxic and characterized by small quantities of released bioactive compounds and Cu2+ ions into the physiological saline solution and hold promise as protective, reusable medical textiles suitable for producing gowns and drapes
Концентровани сок брескве обогаћен олигосахаридима изолованим из споредних токова индустрије соје са пребиотском и антиоксидативном активношћу
No full textКонцентровани сок брескве садржи
влакна и олигосахариде са пребиотском и антиоксидативном активношћу.
Влакна и олигосахариди су изоловани
иновативним поступцима из сојиних
љуспица применом комбинованог
поступка који се заснива на физичком
предтретману и ензимској екстракцији
применом сопствено произведених
ензима, ксиланаза. Ксиланаза је
произведена техником ферментације
на чврстој подлози из властитих сојева
(Penicillium rubens) и као таква показала
је потенцијал за хидролизу отпадне
биомасе и добијање олигосахарида
дефинисаног хемијског састава и
пребиотске активности
Al/Cu joints with AlxCuy interlayers produced under static and dynamic conditions
No full textThe thickness and phase composition of the AlxCuy intermetallic compound (IMC) interlayersincontinuous drive friction welded (CDFW) Al/Cu joints are crucial to their mechanical integrity. Most previousstudies have focused on the formation of these IMC interlayers under static conditions (heat treatment) toevaluate their impact on Al/Cu joint properties. In contrast, the present study produced two sets of Al/Cujoints using CDFW with similar welding parameters. Both sets of as-welded joints featured discontinuousAl2Cu IMC interlayers with similar thicknesses below 0.5 µm. In the first set, IMC interlayers weregrownviaheat treaetment at 325°C for 1 and 12 h (static conditions). In the second set, IMC layers were developedbyextending the friction time of the CDFW process to 2 and 4 s while maintaining other welding parametersconstant (dynamic conditions). This approach aimed to compare the interfacial microstructure andtensileproperties of Al/Cu joints containing AlxCuy IMC interlayers formed under different conditions. Thejointswere evaluated using light microscopy (LM), scanning electron microscopy (SEM), and uniaxial tensiletesting.Joints heat treated at 325°C for 1 h contained an IMC interlayer comrising continuous Al2CuandAl4Cu9sublayers with a total thickness of approximatelly 1 µm. These joints exhibited higher strength andductilitycompared to joints produced with a friction time of 2 s, which had an IMC interlayer cconsistingofacontinuous Al2Cu sublayer and a discontinuous Al4Cu9 sublayer with a total thickness of less than0.8µm. Furthermore, joints heat treated at 325°C for 12 h, with an IMC interlayer measuring approximatelly7.3µmin thickness and composed of continuous Al2Cu, AlCu, and Al4Cu9 sublayers, demonstrated higher strengthand comparable ductility to those produced with a friction time of 4 s. The latter joints containedanIMCinterlayer less than 1.6 µm thick, consisting of a continuous Al2Cu sublayer and a discontinuous Al4Cu9sublayer. Overall, heat-treated Al/Cu joints exhibited higher tensile strength than those formed withvaryingfriction times, despite more substantial IMC interlayer development in terms of thickness andphasecomposition. This outcome does not align well with the current understanding of CDFWjoining of Al andCu, which typically associates thicker and more complex IMC layers with reduced mechanical performancedueto increased brittleness.Book of Abstract
Application of model fitting methods to study thermal degradation of waterborne polyurethanes at late stages
No full textThe thermal degradation of the waterborne polyurethanes (WBPUs) based on
poly(dimethylsiloxane) (PDMS), isophorone diisocyanate (IPDI) and 2,2-bis-
(hydroxymethyl)propionic acid (DMPA) is a complex, three-step process. Study of the kinetics of
the second and third degradation steps of these WBPUs has been presented here. During the second
step, PDMS chains decomposed, while in the third step degradation of residual components
occurred. A comprehensive analysis of the obtained results revealed that the Zhuravlev-Lesokin-
Tempelman and Avrami-Erofeev equations represent the best kinetic models that describe
degradation mechanism of the examined decomposition steps of WBPUs