Ján Brindza

Optimization of ultrasound-assisted extraction of bioactive compounds from wild garlic (Allium ursinum L.).

Ultrasound-assisted extraction was used for extraction of bioactive compounds and for production of Allium ursinum liquid extract. The experiments were carried out according to tree level, four variables, face-centered cubic experimental design (FDC) combined with response surface methodology (RSM). Temperature (from 40 to 80 °C), ethanol concentration (from 30% to 70%), extraction time (from 40 to 80 min) and ultrasonic power (from 19.2 to 38.4 W/L) were investigated as independent variables in order to obtain the optimal conditions for extraction and to maximize the yield of total phenols (TP), flavonoids (TF) and antioxidant activity of obtained extracts. Experimental results were fitted to the second order polynomial model where multiple regression and analysis of variance were used to determine the fitness of the model and optimal condition for investigated responses. The predicted values of the TP (1.60 g GAE/100 g DW), TF (0.35 g CE/100 g DW), antioxidant activity, IC50 (0.71 mg/ml) and EY (38.1%) were determined at the optimal conditions for ultrasound assisted extraction: 80 °C temperature, 70% ethanol, 79.8 min and 20.06 W/L ultrasonic power. The predicted results matched well with the experimental results obtained using optimal extraction conditions which validated the RSM model with a good correlation.

Changes in expression of BetV1 allergen of silver birch pollen in urbanized area of Ukraine.

The aim of the study was to determinate the level of expression of silver birch allergen Betv1 in pollen samples from different Ukraine areas by RT-qPCR SYBR Green assay. Protocol for quantifying the expression of Betv1 allergen was developed when testing of three housekeeping genes—cyclophylin, alpha-tubulin and transcription factor CBF1. Samples from urbanized area was analysed by real-time PCR when a sample from forest growth conditions was used as a calibrator. Real-time PCR based quantifying of Betv1 provides a useful method for rapid and sensitive analyses of this silver birch allergen. Our results show higher expression levels in samples from central parts of urbanized area as housing estates when compared to the samples from borders of the urbanized area.

Comparison of culturable Gram-negative bacterial community structures in the rhizosphere of three fruit plants

This research work was oriented to outlining the diversity of Gram-negative culturable portion of the bacterial community in three fruit plants rhizosphere. Rhizosphere samples were taken from European chestnut (Castanea sativa Mill), true service tree (Sorbus domestica L.) and cornelian cherry (Cornus mas L.) plants. Experiments were conducted for three years during the vegetation period, and the bacterial community structure was assessed with cultivation-dependent approach. Many Gram-negative isolates (n = 251) from the rhizosphere survived sub culturing and were identified by biochemical tests. A total of 57 species belonging to 29 genera were identified and assigned to four broad taxonomic groups (Bacteroidetes, Alpha-, Beta- and Gamma-proteobacteria). Several specific bacterial cluster communities were identified inside all the three rhizospheres. Most of the species belonged to the genera Moraxella, Pseudomonas, Pantoea, Enterobacter and Acinetobacter. In addition, while, using the plate count analysis, large discrepancies in numbers among physiological groups of bacteria cultured from three rhizosphere samples have not been revealed, more expressive distinctions among bacterial populations were obtained concerning the relative abundance of different genera, different taxonomic groups as well as different diversity indices. Furthermore, the number of cultured bacteria and their taxonomic distribution in the rhizosphere of all three plants changed not only explicitly during vegetation period but continually during the three years of investigation. It seems that rhizosphere bacterial populations of each plant are under the influence of the specific root-released materials.

Variability of Corylus avellana, L. CorA and profilin pollen allergens expression.

Corylus avellana is the source of inhalant allergies induced by hazel pollen as well as food allergies induced after ingestion of hazelnuts. In this study, real-time PCR approach was used to analyse expression of hazel pollen allergens on the molecular level. Relative quantity of hazelnut allergens Corylus avellana, L. CorA and Corylus avellana, L. pollen profiling in samples from different Ukraine areas were determining and comparing. Differences among the levels of both analysed allergen transcripts were found for hazel CorA and profillin. In both cases, the expression within the urbanized growth conditions was higher when compared to the sample from village area. The average expression for CorA was 0.84 times higher than for profilin and the results are very variable depending on the place of growth. Expression levels here were within the range of 2.957 up to the 52.936. Profilin expression was the highest in the sample from the polluted place of growth—cement plant area with the value of 52 times higher when compared to the sample from the village area. In this study, comparison of expression levels of hazel CorA and profiling pollen allergens was performed for the first time. Real-time PCR assay developed in this study proved the sensitivity for detection of the changes of the hazel pollen allergens expression levels and could benefit labs by fast and reproducible detection method of these allergens.

Biological properties of black mulberry-derived food products ( Morus nigra L.)

BACKGROUND: Black mulberry is a good source of bioactive compounds and especially of anthocyanin. OBJECTIVES: The objective of this study was to evaluate the morphological, biochemical, technological and antioxidant properties of black mulberry fruit and fruit-derived food products. MATERIALS AND METHODS: Total anthocyanin content, total sugar content, ascorbic acid content, pH and antioxidant activity was analyzed in black mulberry juice, jam, jelly, syrup, liqueur, compote, wine and cake. All products were produced from selected genotypes of black mulberry grown in Slovakia. RESULTS: Reducing sugar content ranged from 6.50 mg.kg–1 FM in wine to 60.01 mg.kg–1 FM in jam, ascorbic acid content was from 0.27 mg.100 g–1 FM in wine to 1.10 mg.100 g–1 FM in fruits conserved in honey without sterilization. Content of anthocyanin varied from 21.4 in wine to 106.4 mg.dm–3 in fresh juice. Detected pH values ranged from 3.28 in jelly to 3.69 in liqueur. All tested samples had strong DPPH• radical scavenging activity with the best values in black mulberry compote with honey – 74.02% and jam 71.22% FM. Differences between the genotypes were significant and the content of the anthocyanin was significantly higher in genotypes with lower fruit weight (P ≤ 0.05). CONCLUSION: The results showed that black mulberry fruits and derived products are rich in biologically active compounds and can be used in food industry and human nutrition.

Identification of sweet chesnut pollen in bee pollen pellet using using molecular analysis.

Castanea sativa posses many characteristics that are used by human for different purposes, not only as a part of the food. One of them is the utilization of the sweet chesnut pollen for its pharmacological benefits. Actually, no information about the DNA based identification of the sweet chesnut exist. Here, an identification of Castanea sativa based on the specific DNA fragment amplification is described for the first time. Sweet chesnut identification was performed in the very complex sample of bee pollen pellets that were identified as to contain sweet chesnut pollen grains by morphological analysis. First, bioinformatic analysis was performed to find a Castanea sativa conservative part of galactol synthase gene. BLAST alignment of the CDS of GolS1 gene was performed by BLASTtn against plants nucleotide sequences in the NCBI database to ensure for the specifity or existing nucleotide differences. Then, specific primers were subsequently designed and PCR amplification was performed. All the PCRs have run in duplicates for pollen pellet sample and two independent samples of Castanea sativa pure pollen. Restriction cleavage of the PCR amplified fragment was performed to confirm the specifity of the obtained PCR product with the positive confirmation as the predicted three restriction fragments were obtained that fully correspond by the length to those from virtual clevage. Restriction endonuclease Hpy166II was used in restriction cleavage analysis. Castanea sativa pollen grains were confirmed reliable in multifloral pollen pellet by PCR and this approach has the potential to be used effectively for the authentication purposes of sweet chesnut.

SODIUM AZIDE INDUCED MORPHOLOGICAL AND MOLECULAR CHANGES IN PERSIMMON (DIOSPYROS LOTUS L.)

Sodium Azide Induced Morphological and Molecular Changes in Persimmon (Diospyros Lotus L.) Persimmon Diospyros lotus L. is an underutilized species, used as rootstock for Japanese persimmon (Diospyros kaki Thunb.) in many European countries. It has a potential for ornamental uses due to its diploid genome that suggests an easy mutation induction. This study was aimed at unravelling the effects of mutagenic sodium azide (in concentrations of 0.1% and 0.5%) on morphological characters (plant height, maximal leaf blade length, maximal leaf blade width and leaf petiole length) and molecular relations of treated accessions, based on AFLP (amplified fragment length polymorphism). In the M1 generation, the analysed characters of two-year old plants differed among the treatments. Sodium azide influenced persimmon stem and leaves in the opposite way, with increase up to 31.57% in plant height and decrease up to 30.33% in leaf petiole length. Changes in morphological characters were significant, that implies the importance of azide influence on the persimmon. Most genotypes treated with 0.1% azide were without significant differences among them and clustered together. The results suggest that the higher azide concentration of 0.5% is more suitable for persimmon seeds azide treatment to form genotypes, significantly different from the control, at the molecular level.

Importance of Collected Flax Germplasm (Linum usitatissimum L.) Characterization

Abstract Our work dealt with detailed characterization of 265 flax genotypes. The collection was evaluated by morphometrical analysis (23 quantitative, 21 qualitative traits); by classification according to an innovated list of descriptors; by image documentation (about 2300 images of leaf, flower, petals, sepals, capsule, and seed); and by mathematical-statistical analyses (descriptive statistics, variability parameters, normality tests, single varietal methods). All obtained data were processed and stored in an information system developed for this purpose. GENOTYP DATA LINUM consists of a completely prepared list of descriptors, catalogue cards of individual genotypes. All parts of this sys are mutually connected and data can be classified, selected, and compared.

Variation of fruits morphometric parameters of Elaeagnus multiflora thunb. germplasm collection

The aim of this study was to determine morphometric parameters of fruits within some genotypes of Elaeagnus multiflora Thunb., which are growing in the Forest-Steppe of Ukraine in M.M. Gryshko National Botanical Garden of NAS of Ukraine (NBG). Their morphometric parameters were following: fruit weight from 0.32 to 1.89 g, fruit length from 7.60 to 19.54 mm, fruit diameter from 4.39 to 10.32 mm, seed weight from 0.10 to 0.41 g, seed length from 7.40 to 13.30 mm, seed diameter from 1.34 to 5.07 mm. The shape indexes of fruits and seeds were found ranging from 1.25 to 1.56 and from 2.90 to 4.04, respectively. The analysis of coefficient of variation showed the difference of variability in morphometric characteristics between Elaeagnus multiflora samples. Data showed that the most variability of important selection characteristics is the average cumulative seeds weight - from 18.72 to 36.61%, seeds diameter - from 10.46 to 24.29%, fruits weight - from 9.15 to 22.24%. The other characteristics are more or less stable. Collected quantitative data were subjected to principal hierarchical cluster analysis. The cluster analysis of morphometric parameters exhibited that the Elaeagnus multiflora genotypes fell into two clusters. These cluster analysis results suggest that broad morphologic diversity was found in Elaeagnus multiflora genotypes examined in this study. The introduction population of the Elaeagnus multiflora , was created in the M.M. Gryshko National Botanical Garden in Kyiv, has a sufficient potential for successful selection work. These preliminary results could open the interest of farmers in this Elaeagnus multiflora and will be precedent for future domestication and introduction of the species in the agroproductive system in Ukraine and the rest of the world.

Alteration of biochemical parameters and microstructure of Fagopyrum esculentum Moench grain in process of germination

Biochemical parameters alteration of Fagopyrum esculentum Moench grain in process of germination was studied. It was found that during germination of Fagopyrum esculentum Moench grain within 24 hours the content of ascorbic acid, thiamine, nicotinic acids, pantothenic acids and routine was increased. The peptide composition of Fagopyrum esculentum Moench grain was studied by gel electrophoresis. The most significant alteration of reserve globulins structure are observed during germination phase from 20 till 24 hours. New low-molecular polypeptides appear during above mentioned period, that indicates embryonic awakening and synthesis of new protein compounds, mainly hydrolases. The process of proteolysis during germination of Fagopyrum esculentum Moench grain promotes a content increase of soluble fractions and sum of albumins and globulins. There is a significant decrease of insoluble protein residue during germination phase change. Chromatographic method was used to determine the change of carbohydrate composition of Fagopyrum esculentum Moench grains during germination. It was established that the content of total carbohydrates amount in grain extracts increases. Electron scanning microscopy revealed that after 12 hours germination of Fagopyrum esculentum Moench grain, swelling of starch grains and minor damage of their packaging in endosperm are observed. After 24 hours, endosperm of germinated grain significantly changed microstructure: starch grains and components of protein matrix had a vague outline, grain disintegration was observed. Evaluation of antioxidant activity of alcohol extract from Fagopyrum esculentum Moench grain germinated during 24 hours showed that percentage of DPPG free radicals inhibition increases with process prolongation. Thus, Fagopyrum esculentum Moench grain germinated within 24 hours is characterized by a high content of biologically active substances and can be used in food technologies for functional products development.