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The presented work is focused on the isotherm study on the Zn2+ ions binding to the individual casein fractions: αS1-, β- and κ-casein (αS1CN, βCN and κCN). The experimental isotherms were evaluated using Freundlich and Langmuir models. Subsequently, the stability of the obtained complexes in the respective Zn2+ ion concentrations (120, 350, 600 mg/L) was determined by carrying out zeta potential measurements. Capillary electrophoresis combined with inductively coupled plasma mass spectrometry (CE-ICP-MS) confirmed the occurring binding process. Additionally, physicochemical characteristics of the obtained metal-protein complexes was performed including scanning electron microscopy (SEM) in two modes (SE and Z-contrast) and the binding sites of caseins to Zn2+ ions were indicated using attenuated total reflectance infrared spectroscopy (FTIR-ATR) and Raman analysis as well as mass spectrometry technique (MALDI-TOF MS). Isothermal studies indicated a heterogeneously complex zinc ion adsorption process, and a stability study showed that the zeta potential is strongly related to the hydrophobicity, size and structure of the casein isoforms studied. Electron microscopy confirmed the modification of casein surfaces due to the addition of Zn2+ ions. Spectroscopic techniques indicated the interaction of zinc ions with polar amino acids of casein, such as glutamic acid (Glu) and aspartic acid (Asp), but also His, Cys. The influence of phosphate groups was also observed. Finally, the study culminated in a molecular docking study of the Zn2+ ion binding process, which confirmed the presence of the listed amino acids responsible for the binding process.

The most critical group of all includes multidrug resistant bacteria that pose a particular threat in hospitals, as they can cause severe and often deadly infections. Modern medicine still faces the difficult task of developing new agents for the effective control of bacterial-based diseases. The targeted administration of nanoparticles can enhance the efficiency of conventional pharmaceutical agents. However, the interpretation of interfaces’ interactions between nanoparticles and biological systems still remains a challenge for researchers. In fact, the current research presents a strategy for using ZnO NPs immobilization with ampicillin and tetracycline. Firstly, the study provides the mechanism of the ampicillin and tetracycline binding on the surface of ZnO NPs. Secondly, it examines the effect of non-immobilized ZnO NPs, immobilized with ampicillin (ZnONPs/AMP) and tetracycline (ZnONPs/TET), on the cells’ metabolism and morphology, based on the protein and lipid profiles. A sorption kinetics study showed that the antibiotics binding on the surface of ZnONPs depend on their structure. The efficiency of the process was definitely higher in the case of ampicillin. In addition, flow cytometry results showed that immobilized nanoparticles present a different mechanism of action. Moreover, according to the MALDI approach, the antibacterial activity mechanism of the investigated ZnO complexes is mainly based on the destruction of cell membrane integrity by lipids and proteins, which is necessary for proper cell function. Additionally, it was noticed that some of the identified changes indicate the activation of defense mechanisms by cells, leading to a decrease in the permeability of a cell’s external barriers or the synthesis of repair proteins.

In this work, the molecular mechanism of Lactobacillus paracasei bio-colloid clumping under divalent metal ions treatment such as zinc, copper and magnesium at constant concentrations was studied. The work involved experimental (electrophoretic - capillary electrophoresis in pseudo-isotachophoresis mode, spectroscopic and spectrometric - FT-IR and MALDI-TOF-MS, microscopic - fluorescent microscopy, and flow cytometry) and theoretical (DFT calculations of model complex systems) characterization. Electrophoretic results have pointed out the formation of aggregates under the Zn2+ and Cu2+ modification, whereas the use of the Mg2+ allowed focusing the zone of L. paracasei biocolloid. According to the FT-IR analysis, the major functional groups involved in the aggregation are deprotonated carboxyl and amide groups derived from the bacterial surface structure. Nature of the divalent metal ions was shown to be one of the key factors influencing the bacterial aggregation process. Proteomic analysis showed that surface modification had a considerable impact on bacteria molecular profiles and protein expression, mainly linked to the activation of carbohydrate and nucleotides metabolism as well with the transcription regulation and membrane transport. Density-functional theory (DFT) calculations of modeled Cu2+, Mg2+ and Zn2+ coordination complexes support the interaction between the divalent metal ions and bacterial proteins. Consequently, the possible mechanism of the aggregation phenomenon was proposed. Therefore, this comprehensive study could be further applied in evaluation of biocolloid aggregation under different types of metal ions.

This research presents, for the first time, the potential of the Lactobacillus paracasei LC20 isolated from sweet whey as a novel, effective and accessible source for post-cultured ZnO nanocomposites synthesis. The obtained nanocomposites were subjected to comprehensive characterization by a broad spectrum of instrumental techniques. Results of spectroscopic and microscopic analysis confirmed the hexagonal crystalline structure of ZnO in the nanometer size. The dispersion stability of the obtained nanocomposites was determined based on the zeta potential (ZP) measurements&mdash, the average ZP value was found to be &minus, 29.15 &plusmn, 1.05 mV in the 7&ndash, 9 pH range. The ZnO nanocomposites (NCs) demonstrated thermal stability up to 130 &deg, C based on the results of thermogravimetric TGA/DTG) analysis. The organic deposit on the nanoparticle surface was recorded by spectroscopic analysis in the infrared range (FT-IR). Results of the spectrometric study exhibited nanostructure-assisted laser desorption/ionization effects and also pointed out the presence of organic deposits and, what is more, allowed us to identify the specific amino acids and peptides present on the ZnO NCs surfaces. In this context, mass spectrometry (MS) data confirmed the nano-ZnO formation mechanism. Moreover, fluorescence data showed an increase in fluorescence signal in the presence of nanocomposites designed for potential use as, e.g., biosensors. Despite ZnO NCs&rsquo, luminescent properties, they can also act as promising antiseptic agents against clinically relevant pathogens. Therefore, a pilot study on the antibacterial activity of biologically synthesized ZnO NCs was carried out against four strains (Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa) by using MIC (minimal inhibitory concentration). Additionally, the colony forming units (CFU) assay was performed and quantified for all bacterial cells as the percentage of viable cells in comparison to a control sample (untreated culture) The nanocomposites were effective among three pathogens with MIC values in the range of 86.25&ndash, 172.5 &mu, g/mL and showed potential as a new type of, e.g., medical path or ointment formulation.

In this paper, a study of the cytotoxicity of bare and functionalized zinc oxide nanoparticles (ZnO NPs) is presented. The functionalized ZnO NPs were obtained by various types of biological methods including microbiological (intra- and extracellular with Lactobacillus paracasei strain), phytochemical (Medicago sativa plant extract) and biochemical (ovalbumin from egg white protein) synthesis. As a control, the bare ZnO NPs gained by chemical synthesis (commercially available) were tested. The cytotoxicity was measured through the use of (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) dye as well as lactate dehydrogenase (LDH) assays against murine fibroblast L929 and Caco-2 cell lines. As a complementary method, scanning electron microscopy (SEM) was performed to assess the morphology of the tested cells after treatment with ZnO NPs. The microscopic data confirmed the occurrence of apoptotic blebbing and loss of membrane permeability after the administration of all ZnO NPs. The reactive oxygen species (ROS) concentration during the cell lines’ exposure to ZnO NPs was measured fluorometrically. Additionally, the photocatalytic degradation of methylene blue (MB) dye in the different light conditions, as well as the antioxidant activity of bare and functionalized ZnO NPs, is also reported. The addition of all types of tested ZnO NPs to methylene blue resulted in enhanced rates of photo-degradation in the presence of both types of irradiation, but the application of UV light resulted in higher photocatalytic activity of ZnO NPs. Furthermore, bare (chemically synthetized) NPs have been recognized as the strongest photocatalysts. In the context of the obtained results, a mechanism underlying the toxicity of bio-ZnO NPs, including (a) the generation of reactive oxygen species and (b) the induction of apoptosis, is proposed.

The present research is focused on experimental and computational mechanistic study of the nanocomplexes formation through Zn2+(aq)-ovalbumin (OVA) binding. The determination of processes occurring at Zn2+(aq)-OVA interfaces and mechanism of hybrid complexes formation involved the modeling of kinetics and isotherm of Zn2+(aq) binding. Scanning electron microscopy, energy-dispersive X-ray analysis and X-ray diffraction were employed to characterize the nano-crystalline structure and morphology of the Zn2+/ZnO-OVO hybrid complex. Formation of hybrid complex was monitored using Fourier transform infrared spectroscopy. Specific Zn2+(aq) binding sites were pointed using MD simulations. Large-scale MD analysis was carried out to sample conformational space of the formed Zn2+(aq)-OVA complexes. Density functional theory (DFT) calculations were used to investigate the structures of the interaction complexes between zinc(II) and Asp−/Glu− and predict their respective IR spectra, and for mechanistic modeling of oxidation processes involved in the formation and stabilization Zn2+/ZnO-OVO hybrid complex. Negatively charged ovalbumin surface was locally neutralized through rapid Zn2+(aq) adsorption and formation of monolayers via 1:1 Asp−/Glu−/-Zn2+(aq) complexes. The ovalbumin monomer species began to agglomerate, stabilized with Asp−/Glu−-Zn2+(aq)-Glu−/Asp− interface complexes that promoted the formation of nanocomplexes. Finally, antimicrobial assays have shown that Zn2+/ZnO-OVO hybrid complex exhibit an inhibition effect against bacteria (A. baumanii, K. pneumonia) and yeast (C. albicans) strains.