Production of rare cycloartane saponins from Astragalus thracicus (Griseb) compared to Astragalus membranaceus (Fisch.) Bunge – native and biotechnological sources

The aim of this study is a comparative metabolomic analysis between the endangered species Astragalus membranaceus and endemic species Astragalus thracicus concerning cycloartane saponins. In addition, in vitro shoots, callus, and suspension cultures of A. thracicus were successfully established to conserve the biodiversity of those endemic species and to increase the amount of produced saponins. The comparison was made according to the quantity of cycloartane saponins astragaloside I (1), astragaloside II (2), and astragaloside IV (4) to the reference standards for the same compounds by UHPLC-HRESI-MS analysis. The in vitro root cultures of A. thracicus reached two folds higher amounts of saponins (1.50 mg/g DW (1), 1.01 mg/g DW (2), and 0.91 mg/g DW (3)) than the native root of A. thracicus (1.14 mg/g DW (1) 0.47 mg/g DW (2), 0.40 mg/g DW (3)), and up to six times higher when compared with roots A. membranaceus (0.23 mg/g DW (1), 0.18 mg/g DW (2) and 0.05 mg/g DW (3))

Arylnaphthalene lignans with a focus Linum species: a review on phytochemical, biotechnological and pharmacological potential

Lignans are a large group of dimeric phenylpropanoids with a long and distinguished history of medicinal use in the ancient cultures of many peoples. The two main groups, -aryltetralin and arylnaphthalene lignans, are leading compounds with important pharmacological properties and a wide range of biological activities. While the first group is well studied mainly for the production of podophyllotoxin, for arylnaphthalene lignans, the data on the availability of a sustainable resource for their production is are still insufficient. The Linum genus, comprising approximately 180 species, is notable for its arylnaphthalene lignans production like justicidin B and isojusticidin B. The pharmacological potential of arylnaphthalene lignans includes cytotoxic, antiviral, anti-inflammatory and antiprotozoal effects. The review highlights the use of biotechnology by in vitro cultures for optimising lignan production. Structural elucidation of novel lignans underscores the ongoing diversity and potential discoveries in this botanical domain, providing an important additional information of arylnaphthalene lignans

Isolation and characterization of 3- O-caffeoyloleanolic acid from Robinia pseudoacacia stem bark

Robinia pseudoacacia, a deciduous tree native to North America, has various medicinal properties, including antioxidant, antitumour, diuretic and antispasmodic effects. The plant contains various bioactive compounds, such as alkaloids, flavonoids, tannins, and phenols. However, caution is advised as all parts of the plant, except the flowers, are poisonous due to the phytotoxin robinin and its glycoside. The bark, on the other hand, shows resistance to rot due to the antifungal compounds dihydrorobinetin and robinetin. This study focuses on the stem bark of R. pseudoacacia from Bulgaria, a widely distributed wild species. Using advanced chromatographic techniques, we isolated and identified 3-O-caffeoyloleanolic acid, a new compound in the genus Robinia and R. pseudoacacia. Structural characterization was performed by state-of-the-art spectroscopic methods, including 1H NMR, 13C NMR and 2D NMR (COSY, HSQC, HMBC), as well as by LC-HRESI-MS analysis

Biotechnological approaches for sustainable production of astragaloside I, II and IV from endemic species of Astracantha aitosensis (Ivan.) and Astragalus membranaceus (fisch.) by in vitro cultures

This study investigates the bioproduction of astragalosides I, II and IV from endemic Astracantha aitosensis (arnacantha) and Astragalus membranaceus species, and the biotechnological methods for increased efficiency. The extracts from established in vitro cultures, including A. aitosensis callus, shoots and roots and A. membranaceus hairy roots, showed higher astragaloside concentrations than native roots. Specifically, in vitro A. aitosensis cultures produced astragaloside I and II at 0.06 and 0.10 mg/g DW, which were absent in native roots. The production of A. membranaceuss hairy roots exceeds 8 to 15 times astragaloside I and II (0.80 and 0.90 mg/g DW) production when compared to native roots (0.10 and 0.05 mg/g DW), and around 3 times high amount related to astragaloside IV. Addressing astragaloside production challenges, this research also reveals biotechnology approaches as an alternative for sustainable production of this rare cycloartane saponins, conserving the natural habitats. A pilot reproducible in vitro cellular platform has been created, and protocol for specific, unconventional induction of the biosynthesis of the desired target compounds, exploiting the enzymatic system of plant cells from the unexplored plant species A. aitosensis has been established. Our findings clearly show the possibility of using in vitro cultures of A. aitosensis and A. membranaceus for biotechnology production of cycloartane type saponins

Biotransformation of quercetin, kaempferol and apigenin to monoglycosylated derivatives by in vitro suspension cultures of Astragalus vesicarius ssp. carniolicus

Biotransformation of exogenous substrates quercetin, kaempferol and apigenin by suspension cultures of Astragalus vesicarius ssp. carniolicus to their monoglycosylated derivatives was performed. The maximal enzymatic potential of cells of A. vesicarius ssp. carniolicus was evaluated by different concentrations of substrate exposure. According to quantitative ultra-high performance liquid chromatography-high resolution electrospray ionization mass spectrometry (UHPLC-HR-ESI-MS) analysis, the highest concentration of kaempferol O-glycoside (14.88 nmol/g dry weight, DW), apigenin O-glycoside (10.55 nmol/g DW) and quercetin O-glycoside (150.83 nmol/g DW) was achieved, when suspension cultures were treated with 4 mg/mL kaempferol, 4 mg/mL apigenin and 3 mg/mL quercetin, respectively. The glycosidic products of biotransformation were not detected in the untreated control

Biotechnological approaches for sustainable production of astragaloside I, II and IV from endemic species of Astracantha aitosensis (Ivan.) and Astragalus membranaceus (fisch.) by in vitro cultures

This study investigates the bioproduction of astragalosides I, II and IV from endemic Astracantha aitosensis (arnacantha) and Astragalus membranaceus species, and the biotechnological methods for increased efficiency. The extracts from established in vitro cultures, including A. aitosensis callus, shoots and roots and A. membranaceus hairy roots, showed higher astragaloside concentrations than native roots. Specifically, in vitro A. aitosensis cultures produced astragaloside I and II at 0.06 and 0.10 mg/g DW, which were absent in native roots. The production of A. membranaceuss hairy roots exceeds 8 to 15 times astragaloside I and II (0.80 and 0.90 mg/g DW) production when compared to native roots (0.10 and 0.05 mg/g DW), and around 3 times high amount related to astragaloside IV. Addressing astragaloside production challenges, this research also reveals biotechnology approaches as an alternative for sustainable production of this rare cycloartane saponins, conserving the natural habitats. A pilot reproducible in vitro cellular platform has been created, and protocol for specific, unconventional induction of the biosynthesis of the desired target compounds, exploiting the enzymatic system of plant cells from the unexplored plant species A. aitosensis has been established. Our findings clearly show the possibility of using in vitro cultures of A. aitosensis and A. membranaceus for biotechnology production of cycloartane type saponins

In vitro production of flavonoids in cultures of Gypsophila glomerata

Effects of increased concentration of calcium chloride on growth and production of flavonoids in newly established shoot and callus Gypsophila glomerata cultures were studied. The highest impact of CaCl2 on the growth index was determined in callus cultures (GI = 0.92), while in shoot cultures calcium treatment reduced the amount of biomass (GI = 0.38). Total flavonoids in shoot cultures grown on MS medium and MS medium supplemented with double amount of CaCl2 were 0.36 mg/g d. w. In both callus cultures, 2 mg/g d. w. total flavonoids were quantified. Shoots and callus grown on non-modified media accumulated 0.02 mg/g d. w. quercetin derivatives. Unlike these, both shoots and callus grown on calcium-enriched media accumulated 0.03 and 0.05 mg/g d. w. of isorhamnetin derivatives. In vitro shoot cultures grown on MS medium enriched in twice the amount of CaCl2 accumulated the highest amount of saponarin (0.138 mg/mg d. w.)

Antiproliferative activity of extract from in vitro callus cultures of Astragalus vesicarius ssp. carniolicus (A. Kern.) Chater

Five isoflavonoids, i.e. 5-hydroxy-7-methoxy-2’, 5’-dihydroxyisoflavone (AV4), 5, 7-dihydroxy-4’-methoxyisoflavone (AV6), 7-methoxy-5-hydroxy-4’-methoxy-2’-hydroxyisoflavone (AV7), 8-pregnyl genistein (AV9), 5,7-dihydroxy-8-pregnyl-4’-methoxy-2’-hydroxyisoflavone (AV10) and one coumarochromone – sophorophenolone (AV8) were isolated from EtOAc of in vitro callus cultures of Astragalus vesicarius ssp. carniolicus, after enzymatic hydrolysis with β-glucosidase. Their structures were tentatively elucidated by spectroscopic mean (1H NMR and HR-ESI-MS spectra). Antiproliferative activity of EtOAc extract and isolated aglycones against chemosensitive human promyelocyte cell line HL-60 and its multidrug-resistant variant HL-60/Dox was assessed in vitro. Despite the strong activity of EtOAc (IC50 8.8 µg/mL (HL-6, 72 h) to 11.8 µg/mL (HL-60/Dox, 72 h)), prenylated compound AV9 showed also antiproliferative activity – 36.1 µg/mL (HL-60 and HL-60/Dox, 72 h)

Induction of flavonoid biosynthesis by in vitro cultivation of Astragalus glycyphyllos L.

Establishment of in vitro cultures from Astragalus glycyphyllos, determination of biomass and analysis of total flavonoids, rutin and camelliaside A were performed. To increase flavonoid production various combinations of plant hormones and light/dark regimen were investigated. Suspension cultures with exogenous quercetin were evaluated for possible increase in flavonoid production. Shoots, calli and suspensions were successfully established. Rutin and camelliaside A were proved in highest amount in shoots. Calli, cultivated on modified G48 medium, with double amount of Ca2+ and Mg2+, achieved higher total flavonoid content (2.37 and 2.03 mg/g DW). Suspensions cultures, cultivated on modified G48 medium with 10, 20 and 30 mg/mL quercetin achieved higher total flavonoid content (0.09, 0.10 and 0.13 mg/mg DW). Biotransformation of quercetin to isoquercitrin was achieved. The highest concentration of isoquercitrin (56.73 ng/mg DW) was observed on suspensions cultures cultivated on modified G48 medium, with 20 mg/mL quercetin