Production of Curcuminoids in different in vitro organs of Curcuma longa.

Curcuma longa L. (turmeric) is one of the most important spice and safe food additives. Its main constituents, curcuminoids, showed anti-inflammatory, antitumor and antioxidant effects. In the present work, an in vitro propagation method was developed to achieve selected plant organs with quantified curcuminoid content. I n vitro plants were obtained from sprouting buds as primary explants. The major curcuminoid constituents, such as curcumin (CUR), demethoxycurcumin (DEM), and bis-demethoxycurcumin (bis-DEM) were examined in different organs by LC-DAD-ESI-MS. A significant production of curcumin (more than 260 μg g−1fresh weight) was obtained from in vitro microrhizomes, especially grown in a Murashige and Skoog medium (MS) supplemented with kinetin (0.1 mg L−1), α-naphthaleneacetic acid (NAA, 1 mg L−1), sucrose (6%), agar (5%) and activated charcoal (0.1%). The analyzed microrhizomes showed reduced amounts of DEM and bis-DEM in comparison with CUR levels. In addition a shoot culture line was suitable to biosynthesize curcuminoids, in a ratio very similar to that identified in the fresh rhizomes of parent plants. This study represents the first direct quantification of curcuminoids in turmeric in vitro shoots and microrhizomes to be used in dietary supplements

Daidzein Production and HeLa Cytotoxicity of Bituminaria bituminosa hairy root Cultures

Bituminaria bituminosa (L.) C.H. Stirt is a perennial species widely distributed in the Mediterranean basin and the Canary Islands. This species is used in folk medicine and currently has considerable pharmaceutical interest for its content in phenylpropanoids, furanocoumarins and pterocarpans. In vitro cultures (shoots and hairy roots) have been performed to obtain plant material useful for the production of these metabolites. Hairy root cultures were successfully established after inoculation of hypocotyls with the LBA 9402 A. rhizogenes strain. The HRPB3 line was selected for further analysis and elicited with chitosan and salicylic acid. All the HRPB3 cultures showed higher polyphenol content and greater DPPH-antioxidant activity than shoots cultured in vitro. The presence of isoflavone daidzein was detected in the hairy root extracts. The cytotoxic effect of HR extracts has been further tested on HeLa cells: the salicylic acid elicited HR exhibited good antiproliferative effects

Activity of Salvia dolomitica and Salvia somalensis Essential Oils against Bacteria, Molds and Yeasts

Essential oils (EOs) fromSalvia dolomiticaandSalvia somalensis, widely employed in the cosmetic and perfume industry, were analyzed for composition and tested against bacterial and fungal pathogens isolated from clinical and environmental specimens. The analyses were carried out againstStaphylococcus aureus,Staphylococcus pseudointermedius,Pseudomonas aeruginosa,Escherichia coli,Streptococcus canis,Streptococcus pyogenes,Klebsiella pneumoniae,Proteus mirabilis,Microsporum canis,Microsporum gypseum,Trichophyton mentagrophytes,Aspergillus niger,Aspergillus flavus,Candida albicans,Candida krusei,Mucorsp. andTrichothecium roseum. Both EOs showed similar percentages of total monoterpenes and sesquiterpene hydrocarbons. The main constituents were 1,8-cineole andβ-caryophyllene inS.dolomiticaand bornyl acetate and camphor inS.somalensis. The selected EOs have no relevant antifungal or antibacterial activities if compared to conventional drugs

1,3-di(benzo[d]oxazol-5-yl)urea acts as either adventitious rooting adjuvant or xylogenesis enhancer in carob and pine microcuttings depending on the presence/absence of exogenous indole-3-butyric acid

Asexual propagation in Ceratonia siliqua L. (carob), species of economic value, is difficult because of adventitious rooting recalcitrance. In Pinus radiata adventitious rooting of hypocotyl cuttings is enhanced by two urea-derivatives, 1,3-di(benzo[d]oxazol-5-yl)urea (5-BDPU) and 1,3-di(benzo[d]oxazol-6-yl)urea (6-BDPU), combined with exogenous indole-3-butyric acid (IBA). The research was aimed to define the role of these urea-derivatives in adventitious root (AR) formation of carob, and to identify morphogenic roles induced in carob, but also in pine, a distantly-related forest species. In carob, 5-BDPU (10 μM) highly promoted AR formation in combination with IBA (1 μM) when applied for 3 days, followed by a transfer onto hormone free medium (HF) up to culture end (4 weeks). IBA alone (1 μM) was more effective than IBA + kinetin (Kin, 10 nM), whereas Kin alone and 5-BDPU alone were not AR-inductive. The histological analysis showed that the cambial cells initiated the ARs, and similar numbers of AR-primordia were visible at day 12, independently of the AR-inductive treatment (i.e., IBA, IBA + 5-BDPU, IBA + Kin). No cutting treated with Kin alone, and rare HF (±5-BDPU)-treated ones, showed AR-primordia at day 12. The number of AR-forming explants increased under IBA + 5-BDPU. By contrast, the cambial cells were stimulated to initiate deuteroxylem instead of ARs under 5-BDPU alone. The histological analysis in pine microcuttings treated with IBA and/or 5-BDPU at the same concentrations confirmed that 5-BDPU applied alone enhanced xylogenesis, highlighting that this urea-derivative exhibits a dual morphogenic role being involved in the switching between adventitious rooting and xylogenesis depending on the presence of exogenous auxin in both species

Resilience of Stevia rebaudiana (Bertoni) Bertoni in the Underwater Biospheres of Nemo's Garden®: Adaptation to New Cultivation Systems

The Nemo's Garden® project is an alternative production system for areas with scarce cultivable land but significant presence of water; thus, it is an interesting intervention to address the climate crisis. This work aimed to evaluate the micromorphological, biochemical, and phytochemical characteristics of Stevia rebaudiana (Bertoni) Bertoni grown underwater compared to the terrestrial specimens. The micromorphological analyses, performed on the leaves using light microscopy, fluorescence microscopy, and scanning electron microscopy, evidenced a general uniformity of the trichome morphotype and distribution pattern. The histochemical investigation indicated the simultaneous presence of terpenes and polyphenols in the trichome secreted material from the underwater samples and a prevailing polyphenolic content in the terrestrial specimens; this was also confirmed by biochemical analyses (26.6 mg GAE/g DW). The characterization of non-volatile components, performed using HPLC-MS, showed similar chemical profiles in all the samples, which were characterized by phenolic compounds and steviol glycosides. The volatile compounds, evaluated using HS-SPME coupled with GC-MS, showed sesquiterpene hydrocarbons as the main class in all the analyzed samples (80.1-93.9%). However, the control plants were characterized by a higher content of monoterpene hydrocarbons (12.1%). The underwater biosphere environment did not alter S. rebaudiana micro-morphological characters, although slight qualitative changes were evidenced for the compounds produced as a response to the growth conditions

Analytical methods for the extraction and identification of secondary metabolite production in 'in vitro' plant cell cultures

The production of plant secondary metabolites by in vitro culture is one of the most challenging and thrilling field of recent scientific researches. In the few last years, pharmaceutical and food industry demand in phytochemicals has increased steadily. Therefore, the establishment of in vitro plant protocols has to be monitored by phytochemical investigation of their selected extracts in order to supply standardized raw material. In this chapter, the advantages and disadvantages of some modern techniques have been described for the sampling, extraction and analysis of the in vitro plants and derivatives. Depending on the volatile or nonvolatile substances produced by in vitro plant raw material, different kinds of laboratory facilities are needed for the extraction and quali‑quantitative analysis. Recent extraction technology such as Accelerated Solvent Extraction or Microwave Assisted Extraction in combination with hyphenated techniques such as Gas Chromathography‑Mass Spectrometry (GC‑MS) and Liquid Chromatography‑Mass Spectrometry (LC‑MS) represent a modern approach to perform fast and reproducible analytical methods for the quality control of secondary metabolite production in ‘in vitro’ plant materia

Volatilomic Analysis of Four Edible Flowers from Agastache Genus

Volatilomes emitted from edible flowers of two species of Agastache (A. aurantiaca (A.Gray) Lint & Epling, and A. mexicana (Kunth) Lint & Epling) and from two hybrids (Agastache ‘Arcado Pink’ and Agastache ‘Blue Boa’) were investigated using a solid-phase microextraction technique as well as the extraction of its essential oils. Oxygenated monoterpenes were almost always the predominant class (>85%) of volatile organic compounds (VOCs) in each sample of A. aurantiaca, A. ‘Blue Boa’ and A. mexicana, with the exception of A. ‘Arcado Pink’ (38.6%). Pulegone was the main compound in A. aurantiaca (76.7%) and A. ‘Blue Boa’ (82.4%), while geranyl acetate (37.5%) followed by geraniol (16%) and geranial (17%) were the principal ones in A. mexicana. The essential oil composition showed the same behavior as the VOCs both for the main class as well as the major constituent (pulegone) with the same exception for A. mexicana. Total soluble sugars, secondary metabolites (polyphenols, flavonoids and anthocyanins) and antioxidant activity were also investigated to emphasize the nutraceutical properties of these edible flowers