Marina Sizonenko’s research while affiliated with North-Caucasus Federal University and other places

Taxifolin (TXL) is a minor bioactive compound from the polyphenol class that may significantly impact human metabolism when included in food products. However, its application is limited by a bitter taste and low bioavailability. We hypothesized that encapsulating TXL in whey proteins using thermomechanical mixing or spray drying could effectively mask its bitterness and enhance bioavailability. Computational simulations indicated that each molecule of β-lactoglobulin (β-Lg) and α-lactalbumin (α-La) can bind at least one TXL molecule. Consequently, experiments used an equimolar ratio of whey proteins to TXL (1:1). Thermomechanical stirring of liquid whey protein concentrate (WPC) at 80 ± 2 °C followed by lyophilisation formed protein aggregates up to 160 μm in size, incorporating large TXL crystals. Encapsulation efficiency was 63 ± 3 %, and the bitter flavour remained unmasked. In contrast, encapsulation via spray drying achieved up to 71 ± 2 % efficiency at an inlet air temperature of 150 °C. The resulting WPC microcapsules, up to 30 μm in size, did not contain large TXL crystals, and bitterness was fully masked. This aligns with the observed reduction in TXL's antioxidant activity. After disrupting WPC microcapsules with ethanol, antioxidant activity of the polyphenol was nearly fully restored. These findings suggest that when such microcapsules are ingested with food, the antioxidant activity of TXL will be expressed in the intestine following proteins breakdown. The results may support the development of novel food products containing whey proteins with encapsulated TXL.

A comparative analysis of mass–volume characteristics of structural and morphological components of hen eggs before incubation and on the fifth (HH25–HH27) and 10th (HH36) days of incubation was carried out. During incubation, egg weight decreased by 9.25% (10 days), mainly due to a decrease in albumen weight (35.8%). The ratio of lipid‐soluble fraction (LSF) and water‐soluble fraction (WSF) in the mixed components and yolk did not change significantly. The total amount of solids in the mixed substances practically does not change during incubation. Antiradical activity of substances increased significantly by more than three times on the fifth day and additionally by 38.6% at the 10th day in relation to the fifth day. Total antioxidant activity increased by 18.9% on the fifth day and by 24.3% on the 10th day, compared to eggs before incubation. Transformation of the main components of WSF and LSF of albumin, yolk, and chicken embryo (CE) was studied using high‐performance liquid chromatography and gas chromatography with mass spectrometry. On the 10th day, an increase in the number of high‐molecular proteins is recorded, which indicates the activation of enzymatic processes of transformation of the main albumen proteins into proteins of organs and tissues of CE. This may cause an increase in the biological activity of substances. It was found that in conditions of in vitro digestion, antiradical activity increases by two times, anti‐inflammatory activity increases by 2.4 times, and an angiotensin‐converting enzyme inhibitory effect occurs in the mixed components of a 10‐day incubation egg.

Assessing the embryotoxicity and teratogenicity of various substances and processes is crucial due to their complexity and resource intensity. The chicken embryo (CE) serves an ideal model for simulating the first months of mammalian embryonic development. This makes the CE a reliable model for testing teratogenic effects, particularly in relation to the nervous system (NS), which experiences developmental abnormalities second in frequency only to cardiovascular teratogenic disorders. Microcomputed tomography (μCT) is a promising method for studying these processes. The advantages of μCT include relatively high research speed, diagnostic accuracy, high resolution and the ability to visualize the entire internal 3D structure of an object while preserving for other types of research. At the same time, there are practically no available databases of normative μCT data, both qualitative and quantitative, which would act as a starting point for screening detection of abnormalities in the development of the NS. In this study, we present a simple method for obtaining very detailed quantitative sets of 2D and 3D μCT data of NS structures of the CE (Gallus Gallus domesticus) at HH22-HH41 embryonic stages with contrasting by 1% phosphotungstic acid. The results of μCT demonstrate the exact boundaries, high general and differentiated contrast of the main and specific structures of the NS of CE, which are quantitatively and qualitatively similar to results of histological analysis. Calculations of the X-ray density and volume of the main structures of the NS at constant exponential growth are presented. In addition to the increase in linear dimensions, significant changes in the structures of various parts of the brain were identified and visualized during the CE development at HH22 to HH41 embryonic stages. The data presented establish the first methodology for obtaining normative data, including subtle localized differences in the NS in CE embryogenesis. The data obtained open up new opportunities for modern embryology, teratology, pharmacology and toxicology.

Relevance . The positive aspects of disinfecting ozonation of hatching eggs allow us to expect successful application of the method not only in large but also in small-scale farms, as well as for experimental and industrial purposes in laboratories and bio-enterprises related to the technological process based on the incubation of a small number of eggs. This determines the importance of expanding the range of ozonizers due to numerous portable devices. There are no clear recommendations for these devices and this causes the need to search for the most effective and harmless modes for the embryo, and schemes of disinfecting ozonation. Methods . The study used fertilized chicken eggs “Hysex Brown” and a portable ozonizer “OZON-OviV”. Ozone concentration 2.0 mg / l. Eggs were treated in a specially made chamber. Technology-1: twice for 30 minutes before incubation and on the 3rd day of incubation. Technology-2: three times for 30 minutes before incubation, on days 3 and 5. The range of studies included: assessment of total microbial contamination (densitometry); identification of microorganisms (MALDI-TOF-spectrometry); biological control of incubation (fertility, hatchability, mortality, developmental abnormalities); assessment of the adequacy of the internal organs (MicroCT); embryo morphometry (weight, length, chest circumference) and calculation of development proportionality indices; histological assessment of the liver. Results . The total ozone concentration during treatment by two methods was 240 mg/l and 360 mg/l, respectively. The disinfecting efficiency of ozonation has been proven, providing a decrease in the level of total microbial contamination by 30% and 40% with double and triple treatment. A tendency to maintain a low total microbial contamination, compared to intact eggs, up to 14 days of incubation has been revealed. The dynamics of the microbial landscape indicate the bacteriostatic effect of ozone in the concentrations used on a wide range of microorganisms. Microtomographic and histological methods confirmed the harmlessness of the technologies used. Along with the more pronounced antibacterial effect of technology-2, the presence of a stimulating effect on the body of the developing embryo was revealed, which determines the preference for its choice.

Synthesis of 0.4 ± 0.03 g/L per day of pure and porous bacterial cellulose (BC) scaffolds (scaffBC) and BC scaffolds modified with gelatin (scaffBC/Gel) was carried out using the Medusomyces gisevii Sa-28 bacterial strain. FT-IR spectroscopy and X-ray diffraction analysis showed that the scaffolds largely consist of crystalline cellulose I (Iα, Iß). Heating of BC with gelatin to 60 • C with subsequent lyophilization led to its modification by adsorption and binding of low-molecular fractions of gelatin and the formation of small pores between the fibers, which increased the biocompatibility and solubility of BC. The solubility of scaffBC and scaffBC/Gel was 20.8 % and 44.4 %, respectively, which enhances degradation in vivo. Light microscopy, scanning electron microscopy, and microcomputed tomography showed a uniform distribution of pores with a diameter of 100-500 μm. The chicken chorioallantoic membrane (CAM) model and subcutaneous implantation in rats confirmed low immunogenicity and intense formation of collagen fibers in both scaffolds and active germination of new blood vessels in scaffBC and scaffBC/Gel. The proliferative cellular activity of fibroblasts confirmed the safety of scaffolds. Taken together, the results obtained show that scaffBC/Gel can be used for the engineering of hard and soft tissues, which opens opportunities for further research.

Relevance . The chicken embryo model is ideal for studying the development of the heart, which is the first functioning organ in embryogenesis. Particular attention is drawn to the embryonic period, when the embryonic heart is both functioning and forming, and small deviations in this process can lead to developmental anomalies. The current lack of results of accurate measurements of the morphology of the heart of a chicken embryo requires high-quality visualization, the complexity of which is determined by dynamic changes in the morphology of the heart, small size and complex spatial details of the organ. Threedimensional imaging methods are indispensable tools in this aspect, among which X-ray microtomography has special advantages. Methods . Embryo scanning (4–8 days, embryonic stages HH22–HH34) was performed using a highresolution X-ray microtomograph Skyscan 1176. A 1% solution of phosphoric-tungstic acid was used as a contrast. Post-processing, reconstruction, visualization of 3D-images, morphometry and X-ray density assessment were carried out using software packages DataViewer (1.5.6.2), CTvox (3.3.0r1403), CT-analyser (1.18.4.0). Results . The results obtained consist of three components: 1) general three-dimensional X-ray microtomographic visualization of chicken embryos in the early stages of embryogenesis with the designation of internal organs, allowing to determine the correct location of the heart; 2) microtomography of the heart with the designation of the main structures; 3) morphometric parameters of the embryo, heart and their X-ray density in units of the Hounsfield scale (HU). The methodological approach used has shown high efficiency. With an increase in the linear size of the chicken embryo and internal organs, including the heart, a high level of both general X-ray contrast and diffraction remains. At all the studied stages of the embryonic period of embryogenesis (4–8 days; HH22–HH34), the main visual and quantitative characteristics of cardiac morphogenesis were revealed.

X-ray contrast techniques were tested on the chick embryos in early periods of embryogenesis. For contrast stain, reagents with radiopacity in various concentrations were used: silver proteinate, eosin, Lugol’s solution (I2KI), phosphomolybdic acid and phosphotungstic acid under heating at 25 °C and 40 °C and exposure for 24 and 48 h. The use of silver proteinate, eosin and I2KI in various concentrations in the contrast of the chick embryo in the early period of embryogenesis did not make it possible to obtain microtomographic results that provide reliable microstructural analysis. The most optimal and effective method of embryo staining at the HH22–HH34 embryonic stages reliably determined the staining of 1% phosphotungstic acid at 40 °C heating and exposure for 24 h. Taking into account the size of the chick embryos and their structures at the HH22–HH34 embryonic stages, the features of the development, location of organs, and the minimum permissible parameters of microtomography for obtaining high-quality and reliable results were determined by the isometric spatial resolution of 8.87 μm, X-ray voltage 50 kV, X-ray current 500 μA, and the use of filters started from Al 0.5 mm. Microtomographic results were obtained, characterized by the appearance of the chick embryo at the HH22–HH34 embryonic stages, and they visualized the locations and structures of the chick embryo organs and provided calculation of their volume and X-ray density. The results of the work open up significant prospects for using the chick embryo at the early embryonic period of embryogenesis as an alternative model for screening teratogenicity.

Unlabelled: The aim of the investigation was to study the level of amylolytic activity and microtomographic index of synovial fluid density as well as to substantiate their clinical and pathogenetic significance by identifying correlations with the known informative indicators reflecting characteristic features of the pathological process in various joint diseases. Materials and methods: Samples of synovial fluid from 95 patients with various joint pathologies at the stage of the disease progression characterized by copious effusion into articular cavities have been examined. Synovial fluid samples obtained by knee arthrocentesis served as a material for the investigation. Conventional methods were used to determine the concentration of uric acid, inorganic phosphorus, total protein, and amylolytic activity level in the selected samples while X-ray density was identified by computed microtomography. Results: All samples of pathological joint fluid have shown a high level of amylolytic activity as compared to the synovial fluid from healthy joints. The relationship between the level of amylolytic activity in synovia and specific joint pathology has been identified. It has also been found that uric acid values, inorganic phosphorus concentrations, and total protein in various types of joint damage may influence X-ray density of the synovial fluid. Correlations between the studied indices have been established. Conclusion: New data on the level of synovia amylolytic activity has been obtained in one non-inflammatory and six different inflammatory diseases. Pathogenically determined correlation between the microtomographic index of synovial fluid density and concentrations of uric acid, inorganic phosphorus, total protein has been confirmed. Specific indicators of X-ray density of synovia in various joint pathologies as well as unidirectional and multidirectional data in comparison with the norm allow us to consider X-ray microtomography as a method that reveals additional details during investigation of synovial fluid density and brings new surrogate markers for the study of pathogenetic mechanisms of the development, differentiation, and treatment of various joint pathologies.