Plant In Vitro Culture Factories for Pentacyclic Triterpenoid Production.

Pentacyclic triterpenoids are a diverse subclass of naturally occurring terpenes with various biological activities and applications. These compounds are broadly distributed in natural plant resources, but their low abundance and the slow growth cycle of plants pose challenges to their extraction and production. The biosynthesis of pentacyclic triterpenoids occurs through two main pathways, the mevalonic acid (MVA) pathway and the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway, which involve several enzymes and modifications. Plant in vitro cultures, including elicited and hairy root cultures, have emerged as an effective and sustainable system for pentacyclic triterpenoid production, circumventing the limitations associated with natural plant resources. Bioreactor systems and controlling key parameters, such as media composition, temperature, light quality, and elicitor treatments, have been optimized to enhance the production and characterization of specific pentacyclic triterpenoids. These systems offer a promising bioprocessing tool for producing pentacyclic triterpenoids characterized by a low carbon footprint and a sustainable source of these compounds for various industrial applications.

Phenolic Profile, Antioxidant and DNA-Protective Capacity, and Microscopic Characters of Ailanthus altissima Aerial Substances.

Invasive species as sources of natural components are of increasing interest for scientific research. This is the case of Ailanthus altissima, which belongs to the top 100 of the most dangerous invasive plant species in Europe, and which is the subject of the present study. The purpose of the research was to analyze the main phenolic compounds in the flowers, leaves, and stem bark of A. altissima and determine the DNA-protective and antioxidant potential of their ethanolic extracts. HPLC profiling revealed the presence of 6 flavonoids and 10 phenolic acids, of which 15 were found in flowers, 14 in leaves, and 11 in the stem bark. Rutin (5.68 mg/g dw in flowers), hesperidin (2.67 mg/g dw in leaves) and (+)-catechin (2.15 mg/g dw in stem bark) were the best-represented flavonoids. Rosmarinic (10.32 mg/g dw in leaves) and salicylic (6.19 mg/g dw in leaves) acids were predominant among phenolic acids. All plant extracts tested showed in vitro antioxidant activity (determined by DPPH, ABTS, FRAP, and CUPRAC assays) and DNA-protection capacity (assay with supercoiled plasmid DNA-pUC19). The highest antioxidant activity was recorded in the flower parts (in the range from 661 to 893 mmol TE/g dw), followed by the leaves. A DNA protective potential for A. altissima leaf and flower extracts has not been established to date. In addition, the main microscopic diagnostic features of studied plant substances were described, with data for the flower parts being reported for the first time. The present study proves that A. altissima could be a natural source of DNA protection and antioxidants.

Antioxidant and DNA-Protective Potentials, Main Phenolic Compounds, and Microscopic Features of Koelreuteria paniculata Aerial Parts.

Interest in plant extracts as a natural source of antioxidants has grown significantly in recent years. The tree species Koelreuteria paniculata deserves attention due to its wide distribution, good adaptability, and growth to the degree of invasiveness in a number of European countries. The purpose of the present study was to analyze flavonoids and phenolic acids of the ethanol extracts from aerial parts of K. paniculata and to screen their antioxidant and DNA-protective activity. HPLC profiling revealed the presence of five flavonoids, with rutin (4.23 mg/g DW), hesperidin (2.97 mg/g DW), and quercetin (2.66 mg/g DW) as the major ones in the leaves, and (-)-epicatechin (2.69 mg/g DW) in the flower buds. Among the nine phenolic acids identified, rosmarinic, p-coumaric, salicylic, vanillic, and gallic acids were the best represented. All the extracts tested showed in vitro antioxidant activity that was determined by DPPH, ABTS, FRAP, and CUPRAC assays. The highest activity was recorded in the flower parts (in the range from 1133 to 4308 mmol TE/g DW). The DNA-protective capacity of the flower and stem bark extracts from the in vitro nicking assay performed, as well as the main diagnostic microscopic features of the plant substances, are given for the first time. According to the results obtained, the aerial parts of K. paniculata could be valuable sources of natural antioxidants.

HS-SPME-GC-MS Volatile Profile Characterization of Peach (Prunus persica L. Batsch) Varieties Grown in the Eastern Balkan Peninsula.

The volatile compounds of eight peach varieties (Prunus persica L.)-"Filina", "Gergana", "Ufo-4", "July lady", "Laskava", "Flat Queen", "Evmolpiya", and "Morsiani 90"-growing in Bulgaria were analyzed for the first time. Gas chromatography-mass spectrometry (GC-MS) analysis and the HS-SPME technique revealed the presence of 65 volatile compounds; the main identified components were aldehydes, esters, and fatty acids. According to the provided principal component analysis (PCA) and hierarchical cluster analysis (HCA), the relative quantities of the identified volatile compounds depended on the studied peach variety. The results obtained could be successfully applied for the metabolic chemotaxonomy of peaches.

GC-MS Metabolic Profile and α-Glucosidase-, α-Amylase-, Lipase-, and Acetylcholinesterase-Inhibitory Activities of Eight Peach Varieties.

The inhibition of certain digestive enzymes by target food matrices represents a new approach in the treatment of socially significant diseases. Proving the ability of fruits to inhibit such enzymes can support the inclusion of specific varieties in the daily diets of patients with diabetes, obesity, Alzheimer's disease, etc., providing them with much more than just valuable micro- and macromolecules. The current study aimed atidentifying and comparing the GC-MS metabolic profiles of eight peach varieties ("Filina", "Ufo 4, "Gergana", "Laskava", "July Lady", "Flat Queen", "Evmolpiya", and "Morsiani 90") grown in Bulgaria (local and introduced) and to evaluate the inhibitory potential of their extracts towards α-glucosidase, α-amylase, lipase, and acetylcholinesterase. In order to confirm samples' differences or similarities, principal component analysis (PCA) and hierarchical cluster analysis (HCA) were also applied to the identified metabolites. The results provide important insights into the metabolomic profiles of the eight peach varieties and represent a first attempt to characterize the peels of the peach varieties with respect to α-glucosidase-, α-amylase-, lipase-, and acetylcholinesterase-inhibitory activities. All of the studied peach extracts displayed inhibitory activity towards α-glucosidase (IC50: 125-757 mg/mL) and acetylcholinesterase (IC50: 60-739 mg/mL), but none of them affected α-amylase activity. Five of the eight varieties showed inhibitory activity towards porcine pancreatic lipase (IC50: 24-167 mg/mL). The obtained results validate the usefulness of peaches and nectarines as valuable sources of natural agents beneficial for human health, although further detailed investigation should be performed in order to thoroughly identify the enzyme inhibitors responsible for each activity.

Comprehensive Evaluation of Late Season Peach Varieties (Prunus persica L.): Fruit Nutritional Quality and Phytochemicals.

Peaches are one of the most preferred seasonal fruits, and a reliable source of nutrients. They possess biologically active substances that largely differ among varieties. Hence, revealing the potential of several late season peaches is of present interest. Three commonly consumed varieties ("Flat Queen"; "Evmolpiya"; "Morsiani 90") were studied in terms of nutritive and phytochemical content, as well as antioxidant activity with the use of reliable spectrophotometric and High Performance Liquid Chromatographic (HPLC) methods. An analysis of the soil was also made. The phytochemical data were subjected to principal component analysis in order to evaluate their relationship. The "Morsiani 90" variety had the highest minerals concentration (2349.03 mg/kg fw), total carbohydrates (16.21 g/100 g fw), and α-tocopherol (395.75 µg/100 g fresh weight (fw)). Similar amounts of TDF (approx. 3 g/100 g fw) were reported for all three varieties. "Flat Queen's" peel extract was the richest in monomeric anthocyanins (2279.33 µg cyanidin-3-glucoside (C3GE)/100 g fw). The "Morsiani 90" variety extracts had the highest antioxidant potential, defined by 2,2-diphenil-1-picrylhydrazyl (DPPH), ferric-reducing antioxidant power (FRAP) and cupric ion-reducing antioxidant capacity (CUPRAC) assays.

Triterpenoids and Other Non-Polar Compounds in Leaves of Wild and Cultivated Vaccinium Species.

The purpose of the current study was to identify and quantify triterpenoids and other non-polar compounds in the leaves of three high bush blueberry cultivars (Vaccinium corymbosum L. var. Bluegold, var. Bluecrop and var. Elliott) and three natural populations of Vaccinium species (Vaccinium uliginosum L., Vaccinium myrtillus L. and Vaccinium vitis-idaea L.) by means of gas chromatography mass spectrometry (GC-MS) and high-performance liquid chromatography with diode array detector (HPLC-DAD). Metabolite profiles differed significantly among the Vaccinium species analyzed, as well as among the populations of the same species. The populations of V. vitis-idaea predominantly contained relative concentrations of phytosterols (varying between 10.48% of total ion current (TIC) and 22.29% of TIC) and almost twice the content of triterpenes (from 29.84% of TIC to 49.62% of TIC) of the other berry species investigated. The leaves of V. corymbosum varieties biosynthesized the highest relative amount of fatty acids, while the leaves of the populations of V. uliginosum had the highest relative concentrations of fatty alcohols. The results of principal component analysis (PCA) and hierarchical cluster analysis (HCA) showed that the diverse populations of each berry species analyzed differed from each other, most likely due to variations in the climatic and geographical conditions of their localities.

Comparative Study of Early- and Mid-Ripening Peach (Prunus persica L.) Varieties: Biological Activity, Macro-, and Micro- Nutrient Profile.

Exploring the chemical composition and biological activity of different fruit varieties is essential for the valorization of their health claims. The current study focuses on a detailed comparative analysis of three early- and two mid-ripening peach varieties: "Filina" (peach), "July Lady" (peach), "Laskava" (peach), "Gergana" (nectarine), and "Ufo 4" (flat peach). They were characterized in terms of essential nutrients such as carbohydrates (sugars and dietary fibers), amino acid content, and lipids as well as mineral content, fat-soluble vitamins, carotenoids, and chlorophyll. Polyphenolic compounds and the related antioxidant activity were also assessed. The methanolic extract of the peel seems to be richer in the studied biologically active substances compared to the fleshy part of the fruit. Anthocyanins were most abundant in "Gergana" and "July Lady" extracts (6624.8 ± 404.9 and 7133.6 ± 388.8 µg cyanidin-3-glucoside/100 g fw, resp.). The total phenol content of the samples varied from 34.11 ± 0.54 to 157.97 ± 0.67 mg gallic acid equivalents (GAE)/100 g fw. "Filina" and "July Lady" varieties possessed the highest antioxidant activity. Overall, the results of this study confirm that the studied peach varieties have satisfactory nutritional value and are potential sources of biologically active substances. Each variety represents an individual palette of nutrients that should be considered separately from the other.

Metabolite profiling by means of GC-MS combined with principal component analyses of natural populations of Nectaroscordum siculum ssp. bulgaricum (Janka) Stearn.

Nectaroscordum siculum ssp. bulgaricum (Janka) Stearn (Allium siculum subsp. dioscoridis (Sm.) K. Richt.) is a traditional culinary spice from South-East Europe. Studies of N. siculum have focused mainly on the botanical and taxonomic characteristics of this species and there is no data available in the scientific literature about its metabolite profile. Thus, the aim of the current study was metabolite profiling of four wild populations of N. siculum grown in Bulgaria by gas chromatography coupled to mass spectrometry (GC-MS) and subsequent principal component analysis (PCA) of the data obtained. The identified primary metabolites (carbohydrates, amino acids, organic acids and lipids) are initial compounds for the biosynthesis of different plant secondary metabolites, such as polyphenols and flavour compounds with valuable biological activities for humans. The health benefits of the phenolic acids identified in this study have been a prerequisite for the implementation of N. siculum in different food systems in order to increase their quality and biological value.

Phytochemical, nutritional and antioxidant characteristics of whitebeam (Sorbus aria) fruits.

BACKGROUND: Common whitebeam, Sorbus aria (L.) Crantz is an European and Near-Eastern deciduous tree with small red fruits. However, the edible use of this forest fruit is currently not widespread. Also, its nutritional and antioxidative properties remained undiscovered. In this study, the chemical composition and antioxidant properties of common whitebeam fruit were investigated. METHODS: The fruit were collected from Golo Bardo Mountain in Bulgaria. The phytochemical (carotenoids, phenolic compounds, flavonoids), nutritional (moisture, ash, titrable acidity, lipids, proteins, pectin, carbohydrates) and antioxidant activity were evaluated by four assays (DPPH, ABTS, FRAP, and CUPRAC). RESULTS: Sugar analysis demonstrated that only fructose and glucose were detected in the fruit. Pectin content did not exceed 1.30%. Moreover, the fruit had a low lipid content (0.80%). The whitebeam fruit were also characterized as a source of carotenoids (1.69 mg/100 g fresh weight, fw), phenolic compounds (32.42 mg GAE/100 g fw), and flavonoids (20.08 mg QE/100 g fw). Among phenolic acids, only 2,4-dihydrohybenzoic, caffeic, p-coumaric, and sinapic acid were detected. The antioxidant potential of the fruit was from 168.52 mM TE/100 g fw (FRAP) to 244.81 mM TE/100 g fw (CUPRAC assay). CONCLUSIONS: The current research enriched the available information about the nutritional potential and chemical composition of common whitebeam fruit and their low sugar content. The absence of sucrose, together with its high phenolic content, demonstrated the potential of this fruit for future application in food products and supplements.

Analysis of the GC-MS of volatile compounds and the phytochemical profile and antioxidant activities of some Bulgarian medicinal plants.

The study's objective was to investigate the volatile compounds, assess the total phenolic content and phenolic acids profile, determine the antioxidant capacity and evaluate the anthocyanin and flavonoid contents in stinging nettle (Urtica dioica L.), tansy (Tanacetum vulgare L.), bladder campion (Silene vulgaris (Moench) Garcke) and rosehip fruit (Rosa canina L. cv. Plovdiv 1). The total phenolic content ranged from 0.55±0.00 to 47.39±0.41 mg GAE/g dw, total flavonoids ranged from 0.45±0.02 to 17.27±0.45 mg QЕ/g dw and the total anthocyanins ranged from 0.94±0.23 to 58.11±2.85 mg/L. The results demonstrated that the selected research plants show auspiciously useful properties in fields like pharmacy, food preparation, cosmetics, etc. Furthermore, the most promising extraction methods in terms of bioactivity are decoction and tincture (U. dioica; T. vulgare; S. vulgaris) and tincture (R. canina). The current study could be regarded as a first detailed investigation of the biological activity and phytochemical composition of R. canina L. cv. Plovdiv 1.

Carotenoids, tocopherols, organic acids, charbohydrate and mineral content in different medicinal plant extracts.

Nettle (Urtica dioica L.), tansy (Tanacetum vulgare L.), bladder campion (Silene vulgaris (Moench) Garcke, waterpepper (Polygonum hydropiper L.), common centaury (Centaurium erythraea Pers.) and rose hip fruit (Rosa canina L. cv. Plovdiv 1) were used for preparation of different water extracts (infusion, decoction and microwave extract) and ethanol (tincture) extracts. Carotenoids (lutein, lycopene and ß-carotene), tocopherols (α-, γ- and δ-), organic acids (ascorbic, malic, fumaric and citric), five macro- and three microelements, sugars and uronic acids content in the obtained extracts were analyzed. Among the investigated plants, stinging nettle, bladder campion and rose hip fruit were evaluated as most potential with respect to bioactive compounds and microelements. The results showed that the selected six medicinal plants and their extracts can be presented as sources of dietary fibers and micronutrients, which may encourage further application as food supplements and beverages as well as to motivate plant use as a dietary alternative in different foods. The present study is a first detailed analysis with respect to sugar content of decoction, infusion and tincture of S. vulgaris.

Food additives and bioactive substances from in vitro systems of edible plants from the Balkan peninsula.

During the last few years there is an increasing demand to the natural biologically active compounds. According to the World Health Organization (WHO) about 11% of the conventional medicines are of plant origin. Nowadays, plant biotechnologies are modern and reliable tool for producing valuable bioactive compounds. Recently, the potential of plant cells as foods also was confirmed. The advantages of plant in vitro systems over the intact plants are well known: growing under controlled and optimized laboratory conditions; independence of climatic and soil differences; preservation of rare and endangered plant species; cultivation in diverse bioreactor systems for increasing production yields of target metabolites. There have been developed many in vitro systems for production of various plant bioactive compounds with potential application in food industries. But potential for industrial implementation of this technology depends on solving problems with the scale-up of bioreactor cultivation, development of additional approaches for improving/modification of bioactivities of the target plant secondary metabolites, and to find way to exclude or replace in the culture media the carcinogenic plant growth regulator 2,4-dichlorophenoxyacetic acid (2,4-D) with its safety analogs, such as α-naphtaleneacetic acid (NAA) and/or indole-3-butyric acid (IBA). The aim of the current mini review is to summarize information about different in vitro systems of edible plants from the Balkan Peninsula with potential for producing food additives and biologically active substances and to describe prospects for successful industrial implementation of this technology.

Ultrasound and Microwave-Assisted Extraction of Elecampane (Inula helenium) Roots.

The aim of the current research was to perform two "green chemistry" extractions (ultrasound and microwave irradiation) with methanol, 70% (v/v) ethanol and water for extraction of biologically active substances from elecampane (Inula heleniun L.) roots and to compare their contents in all extracts. The presence of carbohydrates (sugars, fructooligosaccharides and inulin), total phenols and flavonoids were established. In vitro antioxidant potential was also evaluated by four assays (DPPH, ABTS, FRAP and CUPRAC). Water extracts obtained by ultrasound-assisted extraction (UAE) showed the highest value of inulin (38 g/100 g dry weight plant material). The highest antioxidant activity was possessed by the 70% (v/v) ethanol extracts obtained by UAE: DPPH - 107.2 mM TE/g dw, ABTS - 86.0 mM TE/g dw, FRAP - 67.0 mM TE/g dw and CUPRAC -173.0 mM TE/g dw, respectively. The reason for this probably depended on the highest content of total phenols in the 70% UAE ethanol extract (7.9 mg GAE/g dw, phenolic acids (chlorogenic, caffeic, p-coumaric, sinapic and ferulic acids)}, especially chlorogenic acid (1.84 mg/g) and flavonoids (quercetin, kaempferol and catechin; 26.4 mg QE/g dw). UAE was evaluated as a promising approach for the simultaneous extraction of bioactive compounds (dietary fibers and antioxidants) from elecampane roots in comparison with microwave irradiation.

Alkaloid profiles and acetylcholinesterase inhibitory activities of Fumaria species from Bulgaria.

GC-MS analysis of alkaloid profiles of five Fumaria species, naturally grown in Bulgaria (F. officinalis, F. thuretii, F. kralikii, F. rostellata and F. schrammii) and analysis of acetylcholinesterase inhibitory activity of alkaloid extracts were performed. Fourteen isoquinoline alkaloids were identified, with the principle ones being protopine, cryptopine, sinactine, parfumine, fumariline, fumarophycine, and fumaritine. Protopine contents, defined by HPLC analysis varied between 210.6 ± 8.8 µg/g DW (F. schrammii) and 334.5 ± 7.1 µg/g DW. (F. rostellata). While all of the investigated alkaloid extracts significantly inhibited acetylcholinesterase activity, the F. kralikii demonstrated the highest level of inhibition (IC(50) 0.13 ± 0.01 mg extract/mL).