Evaluation of Bioactive Compounds, Minerals and Antioxidant Activity of Lingonberry (Vaccinium vitis-idaea L.) Fruits.

The extraction efficiency of major classes of phenolics from lingonberries grown in the central region of Poland was evaluated. The ethanol-water solution (60:40, v/v) resulted in the highest extraction yields; however, the results obtained for ethyl acetate were only slightly lower. Total phenolics estimated by Folin-Ciocalteu assay ranged from 468 to 661 mg of GA/100 g fresh weight (fw), while total flavonoids were in the range of 53.2-67.8 µmol/100 g fw. Both solvents exhibited comparable potential for monomeric anthocyanin extraction (26.1-43.0 mg CGE/100 g of fw). The content of several minerals in these fruits and in soil collected from the same places were compared. The essential metal concentrations in all samples increased in the following order: Cr < Cu < Zn < Fe. The levels of toxic elements (Cd, Pb) were acceptable to human consumption for most tested samples. The ethanol-water extracts exhibited the highest scavenging activity against 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radicals, while the highest reducing capacity evaluated by cupric reducing antioxidant capacity (CUPRAC) was obtained for ethyl acetate extracts.

Phytochemical Properties and Antioxidant Activities of Extracts from Wild Blueberries and Lingonberries.

Among Vaccinium species, blueberries (Vaccinium myrtillus L.) and lingonberries (Vaccinium vitis-idaea) are popular in the human diet. In this study, total phenolic, total flavonoid and total monomeric anthocyanin contents in the ethanol-water extracts of blueberry and lingonberry fruits grown wild in the forests in the central region of Poland were assayed. Antioxidant activities of the extracts from each plant were also evaluated for scavenging ability on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals and reducing power by cupric reducing antioxidant capacity (CUPRAC) method. Total phenolics in the blueberry extracts ranged from 4.58 to 5.28 mg GAE CE/g fw. The extracts from lingonberry fruits contained higher total contents of phenolic compounds (5.82-7.60 mg GAE/g fw) as well as total flavonoids (5.22-6.47 µmol CE/g fw) than those from blueberries (3.74-4.18 µmol CE/g fw). For the total monomeric anthocyanin contents, the blueberry extracts presented significantly higher values (3.01-3.93 mg cyanidin-3-glucoside (C3G) equivalent/g fw) in comparison to the lingonberry extracts (0.32-0.47). Blueberry extracts exhibited higher antioxidant activity measured by both assays in comparison to lingonberry extracts. Water extracts from fresh and dried fruits also exhibited significant antioxidant activities for both types of berries. Considering the health benefits that have been associated with polyphenolic consumption, these fruits could appear as a good source of this group of phytochemical compounds for their direct consumption or their use as ingredients for the design of new food products or food supplements.

Changes in Biologically Active Compounds in Pinus sylvestris Needles after Lymantria monacha Outbreaks and Treatment with Foray 76B.

Due to climate warming, the occurrence of Lymantria monacha outbreaks is predicted to become more frequent, causing repeated and severe damage to conifer trees. Currently, the most effective way to control the outbreaks is aerial spraying with the bioinsecticide Foray 76B. The present study aimed to determine the impact of both: (i) L. monacha outbreaks and (ii) treatment with Foray 76B on tree resistance through the synthesis of polyphenols (TPC), flavonoids (TFC), photosynthetic pigments (chlorophyll a and b, carotenoids), lipid peroxidation (MDA), and soluble sugars (TSS) in Pinus sylvestris needles. Samples were collected from visually healthy (control), damaged/untreated, and damaged/Foray 76B-treated plots in 2020 and 2021 (following year after the outbreaks). The results revealed that L. monacha outbreaks contributed to the increase in TPC by 34.1% in 2020 and 26.7% in 2021. TFC negatively correlated with TPC, resulting in 17.6% and 11.1% lower concentrations in L. monacha-damaged plots in 2020 and 2021, respectively. A decrease in MDA was found in the damaged plots in both 2020 and 2021 (10.2% and 23.3%, respectively), which was associated with the increased synthesis of photosynthetic pigments in 2021. The research results also showed that in the following year after the outbreaks, the increase in the synthesis of photosynthetic pigments was also affected by the treatment with Foray 76B. Moreover, the increase in the synthesis of TPC and photosynthetic pigments in the damaged plots in 2021 illustrates the ability of pines to keep an activated defense system to fight biotic stress. Meanwhile, a higher synthesis of photosynthetic pigments in Foray 76B-treated plots indicates a possible effect of the treatment on faster tree growth and forest recovery after L. monacha outbreaks.

Seed Treatment with Cold Plasma and Electromagnetic Field: Changes in Antioxidant Capacity of Seedlings in Different Picea abies (L.) H. Karst Half-Sib Families.

In the context of climate change, methods to improve the resistance of coniferous trees to biotic and abiotic stress are in great demand. The common plant response to exposure to vastly different stressors is the generation of reactive oxygen species (ROS) followed by activation of the defensive antioxidant system. We aimed to evaluate whether seed treatment with physical stressors can activate the activity of antioxidant enzymes and radical scavenging activity in young Picea abies (L.) H. Karst seedlings. For this, we applied seed treatment with cold plasma (CP) and electromagnetic field (EMF) and compared the response in ten different half-sib families of Norway spruce. The impact of the treatments with CP (1 min-CP1; 2 min-CP2) and EMF (2 min) on one-year-old and two-year-old P. abies seedlings was determined by the emergence rate, parameters of growth, and spectrophotometric assessment of antioxidant capacity (enzyme activity; DPPH and ABTS scavenging) in needles. The results indicated that the impact of seed treatment is strongly dependent on the genetic family. In the 577 half-sib family, the activity of antioxidant enzymes catalase (CAT), ascorbate peroxidase (APX), peroxidase (POX), and glutathione reductase (GR) increased after EMF-treatment in one-year-old seedlings, while similar effects in 477 half-sib family were induced by CP2 treatment. In two-year-old seedlings, CP1-treatment increased CAT, APX, POX, GR, SOD, DPPH, and ABTS activity in the 457 half-sib family. However, no significant impact of the treatment with CP1 was determined in one-year-old seedlings in this family. The application of novel technologies and the consideration of the combinatory impact of genetic and physical factors could have the potential to improve the accumulation of compounds that play an essential role in the defense mechanisms of P. abies. Nevertheless, for different resistance and responses to stressors of plants, their genetic properties play an essential role. A comprehensive analysis of interactions among the stress factors (CP and EMF), genetic properties, and changes induced in the antioxidant system can be of importance both for the practical application of seed treatment in forestry and for understanding fundamental adaptation mechanisms in conifers.

Genotype-Dependent Jasmonic Acid Effect on Pinus sylvestris L. Growth and Induced Systemic Resistance Indicators.

Due to temperature changes, forests are expected to encounter more stress than before, both in terms of biotic factors, such as increased insect attacks, and abiotic factors, such as more frequent droughts. Priming trees to respond to these changes faster and more effectively would be beneficial. Induced systemic resistance (ISR) is a mechanism that is turned on when plants encounter unfavorable conditions. Certain elicitors, such as jasmonic acid (JA) are known to induce plants' metabolic response. However, even though studies on ISR in herbaceous species are common and varied ISR elicitors can be used in agriculture, the same cannot be said about trees and forestry enterprises. We aimed to investigate whether JA used in different concentrations could induce metabolic changes (total phenol content, total flavonoid content, photosynthesis pigment content, antioxidant enzyme activity) in Pinus sylvestris seedlings and how this varies between different pine half-sib families (genotypes). After six weeks with a single application of JA, pine seedlings in several pine genetic families exhibited increased antioxidant enzyme activity, total phenol content and carotenoid content that correlated positively with JA concentrations used. Results from other genetic families were varied, but in many cases, there was a significant response to JA, with a noticeable increase as compared to the unaffected group. The impact on chlorophyll content and flavonoids was less noticeable overall. A positive effect on seedling growth parameters was not observed in any of the test cases. We conclude that JA can induce systemic resistance after a single application exogenously in P. sylvestris seedlings and recommend that the use of JA needs to be optimized by selecting appropriate concentrations.

Seed Treatment with Electromagnetic Field Induces Different Effects on Emergence, Growth and Profiles of Biochemical Compounds in Seven Half-Sib Families of Silver Birch.

In the context of climate change, strategies aimed at enhancing trees' resistance to biotic and abiotic stress are particularly relevant. We applied an electromagnetic field (EMF) seed treatment to observe changes in the establishment and content of biochemical compounds in silver birch seedlings induced by a short (1 min) seed exposure to a physical stressor. The impact of EMF treatment was evaluated on seedling emergence and growth of one-year-old and two-year-old seedlings from seven half-sib families of silver birch. The effects on numerous biochemical parameters in seedling leaves, such as total phenolic content (TPC), total flavonoid content (TFC), amounts of photosynthetic pigments, total soluble sugars (TSS), level of lipid peroxidation level, antioxidant activity and activity of antioxidant enzymes, were compared using spectrophotometric methods. The results indicated that, in one-year-old seedlings, two of seven (60th and 73rd) half-sib families exhibited a positive response to seed treatment with EMFs in nearly all analyzed parameters. For example, in the 60th family, seed treatment with EMFs increased the percentage of emergence by 3 times, one-year-old seedling height by 71%, leaf TPC by 47%, antioxidant activity by 2 times and amount of chlorophyll a by 4.6 times. Meanwhile, the other two (86th and 179th) families exhibited a more obvious positive response to EMF in two-year-old seedlings as compared to one-year-old seedling controls. The results revealed that short-term EMF treatment of silver birch seeds can potentially be used to improve seedling emergence and growth and increase the content of secondary metabolites, antioxidant capacity and photosynthetic pigments. Understanding of the impact of EMFs as well as the influence of genetic differences on tree responses can be significant for practical applications in forestry. Genetic selection of plant genotypes that exhibit positive response trends can open the way to improve the quality of forest stands.

Phenolic and Antioxidant Compound Accumulation of Quercus robur Bark Diverges Based on Tree Genotype, Phenology and Extraction Method.

Oak bark is a rich niche for beneficial bioactive compounds. It is known that the amount of the compounds found in plant tissues can depend on species, genotype, growth site, etc., but it is unclear whether oak phenology, i.e., late or early bud burst, can also influence the amount of phenols and antioxidants that can be extracted. We tested two Quercus robur populations expressing different phenology and five half-sib families in each population to see how phenology, genotype, as well as extrahent differences (75% methanol or water) can determine the total phenol, total flavonoid content, as well as antioxidant activity. Significant statistical differences were found between half-sib families of the same population, between populations representing different oak phenology and different extrahents used. We determined that the extraction of flavonoids was more favorable when using water. So was antioxidant activity using one of the indicators, when significant differences between extrahents were observed. Furthermore, in families where there was a significant difference, phenols showed better results when using methanol. Overall, late bud burst families exhibited higher levels in all parameters tested. Thus, we recommend that for further bioactive compound extraction, all these factors be noted.

Changes in Content of Bioactive Compounds and Antioxidant Activity Induced in Needles of Different Half-Sib Families of Norway Spruce (Picea abies (L.) H. Karst) by Seed Treatment with Cold Plasma.

In order to ensure sufficient food resources for a constantly growing human population, new technologies (e.g., cold plasma technologies) are being developed for increasing the germination and seedling growth without negative effects on the environment. Pinaceae species are considered a natural source of antioxidant compounds and are valued for their pharmaceutical and nutraceutical properties. In this study, the seeds of seven different Norway spruce half-sib families were processed for one or two minutes with cold plasma (CP) using dielectric barrier discharge (DBD) plasma equipment. At the end of the second vegetation season, the total flavonoid content (TFC), DPPH (2,2- diphenyl-1-picryl-hydrazyl-hydrate), and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) antioxidant activity, and the amounts of six organic acids (folic, malic, citric, oxalic, succinic, and ascorbic) were determined in the needles of different half-sib families of Norway spruce seedlings. The results show that the TFC, antioxidant activity, and amounts of organic acids in the seedling needles depended on both the treatment duration and the genetic family. The strongest positive effect on the TFC was determined in the seedlings of the 477, 599, and 541 half-sib families after seed treatment with CP for 1 min (CP1). The TFC in these families increased from 118.06 mg g-1 to 312.6 mg g-1 compared to the control. Moreover, seed treatment with CP1 resulted in the strongest increase in the antioxidant activity of the needles of the 541 half-sib family seedlings; the antioxidant activity, determined by DPPH and ABTS tests, increased by 30 and 23%, respectively, compared to the control. The obtained results indicate that the CP effect on the amount of organic acids in the needles was dependent on the half-sib family. It was determined that treatment with CP1 increased the amount of five organic acids in the needles of the 541 half-sib family seedlings. The presented results show future possibilities for using cold plasma seed treatment in the food and pharmacy industries.

Effect of Extracts from Dominant Forest Floor Species of Clear-Cuts on the Regeneration and Initial Growth of Pinus sylvestris L. with Respect to Climate Change.

Climate change influences the ecological environment and affects the recruitment of plants, in addition to population dynamics, including Scots pine regeneration processes. Therefore, the impact of cover-dominant species extracts on the germination of pine seeds and morpho-physiological traits of seedling under different environmental conditions was evaluated. Increasing temperature reinforces the plant-donor allelochemical effect, reduces Scots pine seed germination, and inhibits seedling morpho-physiological parameters. Conditions unfavourable for the seed germination rate were observed in response to the effect of aqueous extracts of 2-year-old Vaccinium vitis-ideae and 1-year-old Calluna vulgaris under changing environmental conditions. The lowest radicle length and hypocotyl growth were observed in response to the effect of 1-year-old C. vulgaris and 2-year-old Rumex acetosella under increasing temperature (+4 °C) conditions. The chlorophyll a + b concentration in control seedlings strongly decreased from 0.76 to 0.66 mg g-1 (due to current environmental and changing environmental conditions). These factors may reduce the resistance of Scots pine to the effects of dominant species and affect the migration of Scots pine habitats to more favourable environmental conditions.

Priming of Resistance-Related Phenolics: A Study of Plant-Associated Bacteria and Hymenoscyphus fraxineus.

European ash (Fraxinus excelsior) is highly affected by the pathogenic fungus Hymenoscyphus fraxineus in all of Europe. Increases in plant's secondary metabolite (SM) production is often linked tol enhanced resistance to stress, both biotic and abiotic. Moreover, plant-associated bacteria have been shown to enhance SM production in inoculated plants. Thus, our hypothesis is that bacteria may boost ash SM production, hence priming the tree's metabolism and facilitating higher levels of resilience to H. fraxineus. We tested three different ash genotypes and used Paenibacillus sp. and Pseudomonas sp. for inoculation in vitro. Total phenol (TPC), total flavonoid (TFC) and carotenoid contents were measured, as well as the chlorophyll a/b ratio and morphometric growth parameters, in a two-stage trial, whereby seedlings were inoculated with the bacteria during the first stage and with H. fraxineus during the second stage. While the tested bacteria did not positively affect the morphometric growth parameters of ash seedlings, they had a statistically significant effect on TPC, TFC, the chlorophyll a/b ratio and carotenoid content in both stages, thus confirming our hypothesis. Specifically, in ash genotype 64, both bacteria elicited an increase in carotenoid content, TPC and TFC during both stages. Additionally, Pseudomonas sp. inoculated seedlings demonstrated an increase in phenolics after infection with the fungus in both genotypes 64 and 87. Our results indicate that next to genetic selection of the most resilient planting material for ash reforestation, plant-associated bacteria could also be used to boost ash SM production.

Impact of Plant-Associated Bacteria on the In Vitro Growth and Pathogenic Resistance against Phellinus tremulae of Different Aspen (Populus) Genotypes.

Aspens (Populus tremula and its hybrids), economically and ecologically important fast-growing trees, are often damaged by Phellinus tremulae, a rot-causing fungus. Plant-associated bacteria can be used to increase plant growth and resistance; however, no systematic studies relating the activity of symbiotic bacteria to aspen resistance against Phellinus tremulae have been conducted so far. The present pioneer study investigated the responses of two Populus tremula and two P. tremula × P. tremuloides genotypes to in vitro inoculations with, first, either Pseudomonas sp. or Paenibacillus sp. bacteria (isolated originally from hybrid aspen tissue cultures and being most closely related to Pseudomonas oryzihabitans and Paenibacillus tundrae, respectively) and, in the subsequent stage, with Phellinus tremulae. Both morphological parameters of in vitro-grown plants and biochemical content of their leaves, including photosynthesis pigments and secondary metabolites, were analyzed. It was found that both Populus tremula × P. tremuloides genotypes, whose development in vitro was significantly damaged by Phellinus tremulae, were characterized by certain responses to the studied bacteria: decreased shoot development by both Paenibacillus sp. and Pseudomonas sp. and increased phenol content by Pseudomonas sp. In turn, these responses were lacking in both Populus tremula genotypes that showed in vitro resistance to the fungus. Moreover, these genotypes showed positive long-term growth responses to bacterial inoculation, even synergistic with the subsequent fungal inoculation. Hence, the studied bacteria were demonstrated as a potential tool for the improved in vitro propagation of fungus-resistant aspen genotypes.

The Impact of Stress Caused By Light Penetration and Agrotechnological Tools on Photosynthetic Behavior of Apple Trees.

The aim was to find out the impact of stress, caused by agrotechnological tools on photosynthetic behaviour of apple trees. The apple tree (Malus domestica Borkh.) cultivar Rubin was grafted on dwarfing rootstocks P60, planted in single rows spaced 1.25 m apart with 3.5 m between rows. In contrast to plant senescing reflectance index and nitrogen balance index, the photochemical reflectance index was significantly higher in 2018 compared with 2017. Such differences might be caused by drought stress on the summer and fast recovery before harvest time when measurements were made. The movement of nutrients and water disrupted by trunk incision had significantly negative effect on reflectance indices regardless on the year. Mechanical pruning with trunk incision and calcium-prohexadione lead to decreased dry to fresh weight ratio by 10-12% in first year of treatment. Mechanical pruning had significantly negative impact on photosynthetic rate, compared to pruning by super spindle it decreased 47%. Strong positive correlation between PRI and NBI 0,89-0,94, and strong negative correlations between PRI, NBI and PSRI -0.88 - (-0.91) were determined.

Mineral Composition of Wild and Cultivated Blueberries.

The concentrations of 13 elements (Al, Ca, Cd, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, Pb, and Zn) were determined in several samples of native (wild) naturally growing and cultivated blueberry fruits. The total metal contents after mineralization were analyzed by inductively coupled plasma optical emission spectrometry. Reliability of the procedure was checked by the analysis of the certified reference materials Mixed Polish Herbs (INGT-MPH-2) and Leaves of Poplar (NCS DC 73350). In the fruits collected in the forest (wild blueberries), higher contents of Ca, Na, and Mg as well as Mn and Zn were observed. Similar levels of Cu, Cr, Fe, and Ni were detected in both wild-growing and cultivated plants. The significantly higher content of Fe and Cd in cultivated blueberries was connected with the content of these metals in soil samples collected from the same places. The metal extraction efficiency by hot water varied widely for the different blueberries (wild or cultivated) as well as their form (fresh or dried).