Wierzchołki pędów i odcinki węzłowe otoczkowane w alginianie jako alternatywna metoda mikrorozmnażania roślin leczniczych

In recent years, the alginate-encapsulated in vitro-derived shoot tips and nodal segments have been employed as an alternative to somatic embryos method of artificial seed production. Encapsulation of non-embryogenic propagules offers an efficient technique for clonal propagation of elite genotypes e.g. plants of medicinal importance and also enables the establishment of basal collection for gene banks. The artificial seeds technology based on vegetative micropropagules may be useful not only in large-scale propagation but also in short- and long-term conservation and germplasm exchange of desirable species genotypes. This paper presents a brief overview of current status of plants of medicinal value propagated by alginate-encapsulated non-embryogenic plant material.Wierzchołki i odcinki węzłowe pędów pochodzące z kultur in vitro, zamknięte w alginianowych kapsułkach od ponad dwóch dekad wykorzystywane są jako alternatywna – w stosunku do zarodków somatycznych – metoda wytwarzania sztucznych nasion. Kapsułkowanie materiału rozmnożeniowego, innego niż zarodki somatyczne oferuje możliwość rozmnażania klonalnego i krótkoterminowego przechowywania roślin. Technika ta stosowana jest wobec gatunków, dla których nie opracowano metody embriogenezy somatycznej np. elitarnych genotypów roślin o właściwościach leczniczych. Metoda kapsułkowania wierzchołków i odcinków węzłowych pędów umożliwia tworzenie podstawowej kolekcji dla banków genów. Artykuł ten prezentuje przegląd roślin leczniczych rozmnażanych za pomocą wegetatywnych propaguli kapsułkowanych w alginianie

Induction of multi-nucleate microspores in anther culture of salix viminalis L.

The aim of the work was to determine how the induction of androgenesis in selected genotypes of Salix viminalis is affected by thermal factors and medium composition. Anthers isolated from male clones of S. viminalis were pre-cultured at 4, 27 and 32°C for two to eight days in liquid Kyo medium with and without the addition of mannitol and on solid MS or WPM medium. The solid media were supplemented with different concentration of disaccharidesand various combinations of growth regulators including kinetin, 2iP, IAA and IBA. Multi-nucleate microspores indicative of androgenesis were observed in anthers pre-cultured for seven days at 4°C in liquid Kyo medium containing mannitol. The rate of androgenesis was higher when the anthers were transferred to solid modified MS medium containing high concentrations of sucrose and kinetin. In studied genotypes of basket willow early uninucleate microspores underwent sporophytic divisions. Ultrastructural observations showed differences in cellular arrangement of pre-stressed microspores. In Salix viminalis, microspores without starch grains and decreased number of lipid bodies were potentially androgenic cells

Phenylethanoid glycosides in Plantago media L. organs obtained in in vitro cultures

Micropropagation of Plantago media L. and the presence of phenolic compounds in organs of multiplied plants were investigated for the first time. Multiplication of plant material was achieved in shoot-tip cultures and via direct organogenesis on Murashige and Skoog (MS) medium with four variants of plant growth regulators (M1–M4). The best multiplication coefficient – 9.2 was obtained in seedling shoot-tip cultures on MS medium M3 with BA 0.2 mg/L and IAA 1.0 mg/L. Methanol extracts prepared separately from shoots and roots of in vitro- derived plantlets were found to contain typical of the genus Plantago L. phenylethanoid glycosides as the only phenolics. Acteoside and plantamajoside were the major compounds – both known to possess a wide range of promising biological activities applicable for medicinal (therapeutic) and cosmetic uses. Martynoside, as a trace constituent, was also found for the first time in the studied species. The quantitative screening of the extracts by TLC video densitometric method showed a higher content of acteoside in shoots (range 62.43–93.03 mg/g, dry weight) and plantamajoside in roots (range 22.45–44.08 mg/g); the highest recorded values – 93.03 mg/g and 44.08 mg/g, respectively, were found in the organs obtained on MS medium M4 with BA 2.0 mg/L

Przeciwdrobnoustrojowa aktywność trzech gatunków Eryngium L. (Apiaceae)

The antimicrobial activity of ethanolic extracts from leaves and roots of three Eryngium L. genera (E. planum, E. campestre, E. maritimum) native to Poland was tested by the method of series dilutions against different Gram-positive bacteria (two strains) and fungi (five species). The extracts were analyzed by TLC method to confirm phenolic acids, triterpenoid saponins, flavonoids and acetylenes presence. The antimicrobial activity of extracts compared with the reference substance were expressed by Minimal Inhibitory Concentration (MI C). The results have shown that the ethanolic extracts inhibit the growth of Staphylococcus aureus and all tested fungi.Przeciwdrobnoustrojową aktywność etanolowych ekstraktów z liści i korzeni trzech gatunków Eryngium L. (E. planum, E. campestre, E. maritimum) występujących w Polsce testowano metodą seryjnych rozcieńczeń w bulionie przeciw bakteriom Gram + (dwa szczepy) i grzybom (pięć gatunków). Ekstrakty analizowano metodą TLC w celu potwierdzenia obecności kwasów fenolowych, saponin triterpenowych, flawonoidów i acetylenów. Przeciwdrobnoustrojowa aktywność ekstraktów i związków referencyjnych wyrażona została przez MI C (Minimal Inhibitory Concentration, minimalna wartość hamująca). Wyniki badań wskazały, że ekstrakty etanolowe hamowały wzrost Staphylococcus aureus oraz wszystkich testowanych grzybów

Pentacyclic triterpenoids and polyphenols accumulation in cell suspension culture of Chaenomeles japonica (Thunb.) Lindl. ex Spach

Introduction: Callus and cell suspension cultures are widely applied in investigation of production of highvalue secondary metabolites, which may be used as cosmeceuticals, nutraceuticals and pharmaceuticals. Plant cell cultures are promising alternative to intact plant sources for the production of plant-derived drugs of industrial importance. Objective: The aim of the study was to (i) initiate the cell suspension culture of Chaenomeles japonica from homogenous and uniform callus, (ii) stabilize the selected line and (iii) verify its ability to produce the desired groups of secondary metabolites – pentacyclic triterpenoids and polyphenols. Methods: To establish a cell suspension culture, stabilized and homogeneous callus was selected. Cell cultures were systematically passaged every 2 weeks to fresh liquid medium with the same composition. Biomass from cultures at the growth phase and stationary phase was designated for phytochemical research. UHPLC-DAD-MS analyzes were performed. At the same time, their macroscopic and microscopic observations were carried out. Results: Cells of suspension culture line A2 were characterized by the intense divisions. Cell culture extracts (both from the growth phase and stationary phase) contained pentacyclic triterpenoids. In addition, phenolic compounds (chlorogenic acid and proanthocyanidins type B) and in a small amount also epicatechin are present in the extract of the cells harvested from the growth phase. In the present studies, three pentacyclic triterpenoids were detected and quantified in the extracts of cell suspensions and callus line A2. Ursolic and oleanolic acids were the main triterpenoids in the studied extracts. The cell suspension culture from the growth phase exhibited the highest content of ursolic, oleanolic, and betulinic acid (separately and together). Conclusion: The cell suspension culture of Chaenomeles japonica is a promising source of pentacyclic triterpenoids

Pentacyclic triterpenoids and polyphenols accumulation in cell suspension culture of Chaenomeles japonica

Introduction: Callus and cell suspension cultures are widely applied in investigation of production of highvalue secondary metabolites, which may be used as cosmeceuticals, nutraceuticals and pharmaceuticals. Plant cell cultures are promising alternative to intact plant sources for the production of plant-derived drugs of industrial importance. Objective: The aim of the study was to (i) initiate the cell suspension culture of Chaenomeles japonica from homogenous and uniform callus, (ii) stabilize the selected line and (iii) verify its ability to produce the desired groups of secondary metabolites – pentacyclic triterpenoids and polyphenols. Methods: To establish a cell suspension culture, stabilized and homogeneous callus was selected. Cell cultures were systematically passaged every 2 weeks to fresh liquid medium with the same composition. Biomass from cultures at the growth phase and stationary phase was designated for phytochemical research. UHPLC-DAD-MS analyzes were performed. At the same time, their macroscopic and microscopic observations were carried out. Results: Cells of suspension culture line A2 were characterized by the intense divisions. Cell culture extracts (both from the growth phase and stationary phase) contained pentacyclic triterpenoids. In addition, phenolic compounds (chlorogenic acid and proanthocyanidins type B) and in a small amount also epicatechin are present in the extract of the cells harvested from the growth phase. In the present studies, three pentacyclic triterpenoids were detected and quantified in the extracts of cell suspensions and callus line A2. Ursolic and oleanolic acids were the main triterpenoids in the studied extracts. The cell suspension culture from the growth phase exhibited the highest content of ursolic, oleanolic, and betulinic acid (separately and together). Conclusion: The cell suspension culture of Chaenomeles japonica is a promising source of pentacyclic triterpenoids

Micropropagation of Eryngium campestre L. via shoot culture provides valuable uniform plant material with enhanced content of phenolic acids and antimicrobial activity

An efficient micropropagation protocol for production of genetically uniform clones of Eryngium campestre L. was developed. To determine the effect of nutritional and hormonal factors on shoot and root development and bioactive compounds production, three variants of media differing in the content of macro- and micronutrients, as well as plant growth regulators of various types and concentrations were tested. The highest regeneration (100%), with over 13 shoots per explant, was induced on Murashige and Skoog (MS) medium with 1.0 mg lˉ¹ benzyladenine (BA) and 0.1 mg lˉ¹ indole-3-acetic acid (IAA). The in vitro derived shoots multiplied through axillary bud formation were rooted and transferred to an experimental plot with 78% frequency of survival. Flow cytometry showed no variation in nuclear DNA between the seedlings and micropropagated plants. Preliminary thin layer chromatography (TLC) analysis indicated that phenolic acids, saponins, flavonoids and acetylenes were present in plant biomass. Ultra high performance liquid chromatography (UHPLC) analysis revealed that shoots and roots from in vitro derived plants and root cultures maintained the ability to produce rosmarinic acid (RA), rosmarinic acid hexoside (RA-HEX) and chlorogenic acid (CGA). The highest phenolic acid content was detected in roots of in vitro regenerated plants. The extract from those roots expressed the highest inhibitory effect against bacteria Staphylococcus aureus, as well as dermatophytes Trichophyton mentagrophytes and T. rubrum

Właściwości przeciwbakteryjne ekstraktów i ich frakcji trzech gatunków Eryngium L.

Introduction: Due to increasing resistance against antibiotics and antifungal agents, crude plant extracts, fractions, and isolated pure compounds became a new interest as antimicrobial agents. Objectives: The antimicrobial activity of methanolic extracts and fractions of Eryngium planum L., E. campestre L., and E. maritimum L. was evaluated against selected bacteria, yeast and mould, and compared in tested Eryngium species and in their organs. Methods: The antimicrobial activity was studied with use of broth microdilution method. The antibacterial (Staphylococcus aureus, Pseudomonas aeruginosa) and antifungal (Candida albicans, Aspergillus niger) activity of selected extracts and fractions compared with the reference substance was expressed by Minimal Inhibitory Concentration (MIC) and Minimal Bactericidal/Fungicidal Concentration (MBC/MFC).The extract and fraction compounds were identified on the basis of TLC examination. Results: The saponin-phenolic acid fractions of E. maritimum and E. planum and a saponin fraction of E. planum showed the highest activity against S. aureus (MIC = 1–2.5 mg·ml-1). The growth of C. albicans was inhibited by methanolic extract of E. planum cell suspension culture (MIC = 7.8 mg·ml-1). Conclusion: The antimicrobial activity depends on the Eryngium species, tested biomass, and microorganism.Wstęp: Z powodu rosnącej oporności na antybiotyki i środki przeciwgrzybicze rośnie zainteresowanie ekstraktami roślinnymi, frakcjami i wyizolowanymi czystymi związkami jako środkami przeciwdrobnoustrojowymi. Cel: Badano aktywność przeciwdrobnoustrojową ekstraktów metanolowych i frakcji Eryngium planum L., E. campestre L. i E. maritimum L. w stosunku do wybranych bakterii, drożdżaka i grzyba pleśniowego oraz porównywano badane gatunki Eryngium i ich organy. Metody: Aktywność przeciwdrobnoustrojową badano z zastosowaniem metody seryjnych rozcieńczeń w bulionie. Aktywność przeciwbakteryjna (Staphylococcus aureus, Pseudomonas aeruginosa) i przeciwgrzybicza (Candida albicans, Aspergillus niger) wybranych ekstraktów i frakcji, w porównaniu z substancją referencyjną, wyrażona została za pomocą minimalnego stężenia hamującego (MIC) i minimalnego stężenia bakterio- lub grzybobójczego (MBC/MFC). Wyniki: Frakcje saponinowo-fenolokwasowe E. maritimum i E. planum oraz frakcja saponinowa E. planum wykazały najwyższą aktywność wobec S. aureus (MIC = 1–2,5 mg∙ml -1). Wzrost C. albicans był hamowany przez metanolowy ekstrakt zawiesiny komórkowej E. planum (MIC = 7,8 mg∙ml -1). Wnioski: Aktywność przeciwdrobnoustrojowa zależy od gatunku Eryngium, testowanej biomasy i mikroorganizmu