Cynara cardunculus L. var. scolymus L. Landrace "Carciofo Ortano" as a Source of Bioactive Compounds.

The preservation of agricultural biodiversity and socioeconomic development are relevant both to enhance domestic production and to support innovation. In the search for new biomolecules, we have focused on the "Carciofo Ortano" landrace, growth in the northern part of the Lazio region. Artichoke cultivation generates substantial by-products, including leaves, stems, and roots, which could serve as valuable sources of biomolecules and prebiotic dietary fiber. To valorize the leaf waste of the "Carciofo Ortano" landrace, a multidisciplinary approach was applied. Chemical analysis using HPLC-DAD identified mono-O- and di-O-caffeoylquinic acids and the sesquiterpene cynaropicrin in all artichoke leaf extracts. SPME-GC/MS analyses detected aliphatic alcohols in the fresh leaf samples. Antiproliferative and cytotoxic studies on cancer (SH-SY5Y, MCF-7, MDA) and normal (MCF-10A) human cell lines revealed that leaf extracts induced a selective dose and time-dependent biological effect. While showing slight activity against environmental bacterial strains, artichoke leaf extracts exhibited significant antifungal activity against the phytopathogenic fungus Alternaria alternata. Overall, the results highlight the potential of "Carciofo Ortano" cultivation by-products as a rich source of biomolecules with versatile applications in humans, animals, and the environment.

Induction of Drug-Resistance and Production of a Culture Medium Able to Induce Drug-Resistance in Vinblastine Untreated Murine Myeloma Cells.

Cancer therapies use different compounds of synthetic and natural origin. However, despite some positive results, relapses are common, as standard chemotherapy regimens are not fully capable of completely eradicating cancer stem cells. While vinblastine is a common chemotherapeutic agent in the treatment of blood cancers, the development of vinblastine resistance is often observed. Here, we performed cell biology and metabolomics studies to investigate the mechanisms of vinblastine resistance in P3X63Ag8.653 murine myeloma cells. Treatment with low doses of vinblastine in cell media led to the selection of vinblastine-resistant cells and the acquisition of such resistance in previously untreated, murine myeloma cells in culture. To determine the mechanistic basis of this observation, we performed metabolomic analyses of resistant cells and resistant drug-induced cells in a steady state, or incubation with stable isotope-labeled tracers, namely, 13C 15N-amino acids. Taken together, these results indicate that altered amino acid uptake and metabolism could contribute to the acquisition of vinblastine resistance in blood cancer cells. These results will be useful for further research on human cell models.

Evaluation of the Local Anesthetic Activity, Acute Toxicity, and Structure-Toxicity Relationship in Series of Synthesized 1-Aryltetrahydroisoquinoline Alkaloid Derivatives In Vivo and In Silico.

Isoquinoline alkaloids constitute one of the most common classes of alkaloids that have shown a pronounced role in curing various diseases. Finding ways to reduce the toxicity of these molecules and to increase their therapeutic margin is an urgent matter. Here, a one-step method for the synthesis of a series of 1-aryl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolines was performed in 85-98% yield by the Pictet-Spengler reaction. This was accomplished using the reaction between 3,4-dimethoxyphenylethylamine and substituted benzaldehydes boiling in trifluoroacetic acid. Furthermore, 1-(3'-amino-, 4'-aminophenyl)-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolines were obtained in 94% and 97% yield by reduction in 1-(3'-nitro-, 4'-nitrophenyl)-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolines with SnCl2 × 2H2O. The structures of the substances obtained were confirmed by infrared (IR) and nuclear magnetic resonance (1H and 13C NMR) spectra. ADMET/TOPKAT in silico study concluded that the synthesized compounds exhibited acceptable pharmacodynamic and pharmacokinetic properties without carcinogenic or mutagenic potential but with variable hepatotoxicity. The acute toxicity and structure-toxicity relationship (STR) in the series of 20 derivatives of 1-aryl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolines (3a-r, 4a, b) was studied via determination of acute toxicity and resorptive action in white mice employing intragastric step-by-step administration. The first compound, 1-phenyl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline hydrochloride (3a), showed the highest toxicity with LD50 of 280 mg/kg in contrast to 1-(3'-bromo -4'-hydroxyphenyl)-6,7-methylenedioxy-1,2,3,4-tetrahydroisoquinoline hydrochloride (3e) which proved to be the safest of the compounds studied. Its toxicity was 13.75 times lower than that of the parent compound 3a. All compounds investigated showed high local anesthetic activity on rabbit eyes in the concentrations studied. Only 3r, 3n, and 4a caused eye irritation and redness. All investigated derivatives (except 4b) in 1% concentration were more active than lidocaine, providing longer duration of complete anesthesia. Therefore, based on the obtained results of in silico tests, local anesthesia, and acute toxicity, a conclusion can be drawn that the experimental compounds need further extensive future investigations and possible modifications so that they can act as promising drug candidates.

In Vitro Anti-Proliferative and Apoptotic Effects of Hydroxytyrosyl Oleate on SH-SY5Y Human Neuroblastoma Cells.

The antitumor activity of polyphenols derived from extra virgin olive oil and, in particular the biological activity of HTyr, has been studied extensively. However, the use of HTyr as a therapeutic agent for clinical applications is limited by its low bioavailability and rapid excretion in humans. To overcome these limitations, several synthetic strategies have been optimized to prepare lipophenols and new compounds derived from HTyr to increase lipophilicity and bioavailability. One very promising ester is hydroxytyrosyl oleate (HTyr-OL) because the chemical structure of HTyr, which is responsible for several biological activities, is linked to the monounsaturated chain of oleic acid (OA), giving the compound high lipophilicity and thus bioavailability in the cellular environment. In this study, the in vitro cytotoxic, anti-proliferative, and apoptotic induction activities of HTyr-OL were evaluated against SH-SY5Y human neuroblastoma cells, and the effects were compared with those of HTyr and OA. The results showed that the biological activity of HTyr was maintained in HTyr-OL treatments at lower dosages. In addition, the shotgun proteomic approach was used to study HTyr-OL-treated and untreated neuroblastoma cells, revealing that the antioxidant, anti-proliferative and anti-inflammatory activities of HTyr-OL were observed in the unique proteins of the two groups of samples.

Hemp (Cannabis sativa L., Kompolti cv.) and Hop (Humulus lupulus L., Chinook cv.) Essential Oil and Hydrolate: HS-GC-MS Chemical Investigation and Apoptotic Activity Evaluation.

In this study, essential oils (EOs) and hydrolates (Hys) from Italian hemp (Cannabis sativa L. Kompolti cv.) and hop (Humulus Lupulus L., Chinook cv.) supply chains were chemically characterized and tested to investigate their apoptotic potential for the first time. Headspace-Gas Chromatography-Mass Spectrometry (HS-GC-MS) techniques were performed to describe their volatile chemical profile, highlighting a composition rich in terpene derivatives such as monoterpenes and sesquiterpenes among which ß-myrcene, limonene, ß-caryophyllene and α-humulene were the main constituents of EOs; in contrast, linalool, cis-p-menth-2,8-dien-1-ol, terpinen-4-ol, α-terpineol, caryophyllene oxide, and τ-cadinol were found in the Hys. The cytotoxicity activity on human leukemia cells (HL60), human neuroblastoma cells (SH-SY5Y), human metastatic adenocarcinoma breast cells (MCF7), human adenocarcinoma breast cells (MDA), and normal breast epithelial cell (MCF10A) for the EOs and Hys was studied by MTT assay and cytofluorimetric analysis and scanning and transmission electron microscopy were performed to define ultrastructural changes and the mechanism of cells death for HL 60 cells. An induction of the apoptotic mechanism was evidenced for hemp and hop EOs after treatment with the corresponding EC50 dose. In addition, TEM and SEM investigations revealed typical characteristics induced by the apoptotic pathway. Therefore, thanks to the integration of the applied methodologies with the used techniques, this work provides an overview on the metabolomic profile and the apoptotic potential of hemp and hop EOs and, for the first time, also of Hys. The findings of this preliminary study confirm that the EOs and Hys from Cannabis and Humulus species are sources of bioactive molecules with multiple biological effects yet to be explored.

Detection of Volatiles by HS-SPME-GC/MS and Biological Effect Evaluation of Buddha's Hand Fruit.

The present work aimed to chemically characterize and evaluate the antiradical power and biological effects of Citrus medica var. sarcodactylus essential oil (EO) and hydrolate (Hy) from exocarp as well as methanol extracts, from both exocarp and mesocarp (EEX and MEX). The whole fresh fruit was also investigated by SPME-GC/MS to describe its volatile composition. EO and Hy were analyzed by GC/MS and HS-GC/MS techniques, respectively. Limonene and γ-terpinene were found to be the most abundant compounds both in the fresh parts of the fruit and in the EO, while α-terpineol and terpinen-4-ol were in the Hy. The extracts were also rich in furan and coumarin derivatives. A good antiradical activity of all samples except Hy was detected both against ABTS·+ than DPPH·, removed up to about 50%. The antibacterial activity against Bacillus cereus and Escherichia coli was evaluated by microwell dilution method to determine MIC and MBC values. EEX and MEX showed efficacy at very high concentrations against both tested bacteria. The MIC value of EO against B. cereus was 0.5% v/v, while Hy was not able to inhibit the bacterial growth at the tested concentrations. Cytotoxicity investigated on the HL60 leukemia cell line by MTT assay provided an EC50 of 1.24% v/v for EO. Interesting activity of Hy was also observed.

Headspace/GC-MS Analysis and Investigation of Antibacterial, Antioxidant and Cytotoxic Activity of Essential Oils and Hydrolates from Rosmarinus officinalis L. and Lavandula angustifolia Miller.

In this work, essential oils (EOs) and hydrolates (Hys) of Rosmarinus officinalis L. and Lavandula angustifolia Mill., grown in Tuscany (Italy), were studied to describe their chemical composition and biological activities. The aromatic profile of the EOs liquid phase was carried out by gas chromatography-mass spectrometry (GC-MS), while the volatile composition of vapor phase EOs and Hys was performed by headspace (HS)/GC-MS. The obtained results show that monoterpene hydrocarbons (71.5% and 89.5%) were the main compounds, followed by oxygenated monoterpenes (26.0% and 10.5%) in the liquid and vapor phase of R. officinalis EO, respectively. The oxygenated monoterpenes were the main components of L. angustifolia EO, reaching 86.9% in the liquid phase and 53.7% in the vapor phase. Regarding Hys, they consisted only of oxygenated monoterpenes, and 1,8-cineole (56.2%) and linalool (42.9%), were the main components of R. officinalis and L. officinalis Hys, respectively. Their cytotoxicity was investigated on an SHSY5Y neuroblastoma cell line by thiazolyl blue tetrazolium bromide (MTT) test, showing a notable effect of the EOs with a time-independent manner of activity and half maximal effective concentration (EC50) values quite similar for the two plant species (from 0.05% to 0.06% v/v for the three time points evaluated). A measurable activity of Hys was also obtained although with higher EC50 values. The antibacterial activity against Escherichia coli ATCC® 25922, Pseudomonas fluorescens ATCC® 13525, Acinetobacter bohemicus DSM 102855 as Gram-negative bacteria and Kocuria marina DSM 16420, Bacillus cereus ATCC® 10876 as Gram-positive bacteria, was evaluated by the agar disk-diffusion method and the VPT (vapor phase test) to determinate the MIC (minimal inhibitory concentration) and the MBC (minimal bactericidal concentration) values. Both EOs possessed a high activity against all the bacterial strains with MIC values ranging from 0.19% to 3.13% v/v. Unlike EOs, Hys did not show an inhibition of the bacterial growth at the tested concentrations. Furthermore, antioxidant power was measured by 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt-based (ABTS•+) and the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assays, showing a remarkable ability to reduce radicals by both EOs; Hys were slightly less active. The findings highlighted that R. officinalis and L. angustifolia EOs and Hys have a chemical composition rich in bioactive molecules, which can exert different biological activities.

GC-MS investigation and antiproliferative activities of extracts from male and female flowers of Schinus molle L.

The aim of this study is to define chemical composition and antiproliferative activity of several compounds isolated from Schinus molle male and female flowers on human neuroblastoma (SH-SY5Y) and human leukemia (HL60) cell lines. Three extracts obtained by using solvents with different polarities (petroleum ether, diethyl ether and acetone) were characterized by GC/MS analysis revealing qualitative and quantitative differences in the chemical composition of male and female flowers. Nineteen components were identified and both flower extracts were dominated by the sesquiterpene hydrocarbons fraction, mainly elemol, followed by bicyclic sesquiterpenes and by a smaller amount of monoterpenes. The biological activities of the extracts were evaluated by dose-dependent viability assay on two different cell lines and HL60 cells resulted more sensitive to the treatment. The presence of compounds at low concentration with a well-established cytotoxic activity such as ß-elemene, ß-caryophyllene and γ-eudesmol could exert a synergistic cytotoxic effect.

Chemical Investigation and Screening of Anti-Proliferative Activity on Human Cell Lines of Pure and Nano-Formulated Lavandin Essential Oil.

Lavandin essential oil (LEO), a natural sterile hybrid obtained by crossbreeding L. angustifolia × L. latifolia, is mainly composed by active components belonging to the family of terpenes endowed with relevant anti-proliferative activity, which can be enhanced by proper application of nanotechnology. In particular, this study reports the chemical characterization and the screening of the anti-proliferative activity on different human cell lines of pure and nano-formulated lavandin essential oil (EO). LEO and its formulation (NanoLEO) were analyzed by HS/GC-MS (Headspace/Gas Chromatography-Mass Spectrometry) to describe and compare their chemical volatile composition. The most abundant compounds were linalool and 1,8-cineole (LEO: 28.6%; 27.4%) (NanoLEO: 60.4%; 12.6%) followed by α-pinene (LEO: 9.6%; NanoLEO: 4.5%), camphor (LEO: 6.5%; NanoLEO: 7.0%) and linalyl acetate (LEO: 6.5%; NanoLEO: 3.6%). The cytotoxic effects of LEO and NanoLEO were investigated on human neuroblastoma cells (SHSY5Y), human breast adenocarcinoma cells (MCF-7), human lymphoblastic leukemia cells (CCRF CEM), human colorectal adenocarcinoma cells (Caco-2) and one normal breast epithelial cell (MCF10A) by the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide)-assay. Caco-2, MCF7 and MCF10A normal cells resulted more resistant to the treatment with LEO, while CCRF-CEM and SHSY5Y cells were more sensitive. The antiproliferative effect of LEO resulted amplified when the essential oil was supplied as nanoformulation, mainly in Caco-2 cells. Scanning and transmission electron microscopy investigations were carried out on Caco-2 cells to outline at ultrastructural level possible affections induced by LEO and NanoLEO treatments.

Antiproliferative Properties of Papaver rhoeas Ovule Extracts and Derived Fractions Tested on HL60 Leukemia Human Cells.

Papaver rhoeas plant is common in many regions worldwide and contributes to the landscape with its red flower. In the present study we first carried out morphological investigation by optical and scanning electron microscopy of the ovules within the ovary. After ovules' isolation we prepared extracts to test possible cytotoxic activities on HL60 leukemia human cells and investigated the extracts using thin-layer chromatography (TLC) and gas-chromatography/mass spectrometry (GC-MS). P. rhoeas ovules showed an elongated, round shape and the presence of ordered sculptures on the ovule surface. The ovule extracts showed cytotoxic activity on HL60 human cells mainly found in some TLC-isolated spots. Compounds consisting of active spots were identified by GC-MS investigations. Our findings on the P. rhoeas ovule compounds open perspectives for further investigations of TLC-isolated spots on other human cancer cell lines.

Apoptotic Effects on HL60 Human Leukaemia Cells Induced by Lavandin Essential Oil Treatment.

Recent scientific investigations have reported a number of essential oils to interfere with intracellular signalling pathways and to induce apoptosis in different cancer cell types. In this paper, Lavandin Essential Oil (LEO), a natural sterile hybrid obtained by cross-breeding L. angustifolia × L. latifolia, was tested on human leukaemia cells (HL60). Based on the MTT results, the reduced cell viability of HL60 cells was further investigated to determine whether cell death was related to the apoptotic process. HL60 cells treated for 24 h with LEO were processed by flow cytometry, and the presence of Annexin V was measured. The activation of caspases-3 was evaluated by western blot and immunofluorescence techniques. Treated cells were also examined by scanning and transmission electron microscopy to establish the possible occurrence of morphological alterations during the apoptotic process. LEO main compounds, such as linalool, linalyl acetate, 1,8-cineole, and terpinen-4-ol, were also investigated by MTT and flow cytometry analysis. The set of obtained results showed that LEO treatments induced apoptosis in a dose-dependent, but not time-dependent, manner on HL60 cells, while among LEO main compounds, both terpinen-4-ol and linalyl acetate were able to induce apoptosis.

Chemical Investigation of a Biologically Active Schinus molle L. Leaf Extract.

The pepper tree Schinus molle L. is an evergreen ornamental plant belonging to the Anacardiaceae family, native to South America and widespread throughout the world. It has biological activities and is used in folk medicine. This paper aims to contribute to a deeper knowledge of its chemical composition and biological properties. S. molle leaf extracts were obtained by sequential extraction with solvents of different polarities and subsequently tested on the HL-60 human leukaemia cell line to define a possible cytotoxic activity. Among the investigated extracts, the petroleum ether extract revealed a high cytotoxic activity, and its chemical composition was further investigated. By a silica column chromatography, eight fractions were obtained, and their compositions were determined by GC-MS analysis. Compounds and relative abundance differed widely among the fractions; sesquiterpenes resulted the main component and alcoholic sesquiterpenes the most abundant.

Liquid and Vapour Phase of Lavandin (Lavandula × intermedia) Essential Oil: Chemical Composition and Antimicrobial Activity.

Essential oils from Lavandula genus and the obtained hybrids are widely used for different purposes such as perfume production in the cosmetic field and for its biological properties. This is the first study on the liquid and vapour phase of Lavandula × intermedia "Grosso" essential oil grown in the Lazio Region, Italy, investigated using headspace coupled to gas chromatography and mass spectrometry (HS-GC/MS). The results showed the most abundant components were linalool and linalyl acetate, followed by 1,8-cineole and terpinen-4-ol, while lavandulyl acetate and borneol were identified as minor compounds, maintaining the same proportion in both the liquid and vapour phase. Furthermore, we tested lavandin liquid and vapour phase essential oil on gram-negative bacteria (Escherichia coli, Acinetobacter bohemicus, and Pseudomonas fluorescens) and gram-positive bacteria (Bacillus cereus and Kocuria marina).

GLC-MS profiling of non-polar extracts from Phlomis bucharica and P. salicifolia and their cytotoxicity / Perfiles GLC-MS de extractos no polares de Phlomis bucharica y P. salicifolia y su citotoxicidad

Phlomis species (Phlomis bucharica Regel and P. salicifolia Regel) have been traditionally used by Uzbek people as stimulant, tonic, diuretic, and in the treatment of ulcers, hemorrhoids, wounds and gynecological problems. In the present study, we characterized the chemical composition of non-polar extracts from P. bucharica and P. salicifolia by high resolution GLC-MS and evaluated their cytotoxicity. Concentrations of hexadecanoic acid in hexane and chloroform extracts were higher in P. bucharica than in P. salicifolia. 1,8- Cineol, camphor, borneol, alfa-terpinol, thymol, and isobornyl acetate were detected in P. bucharica but not in P. salicifolia. About 45 components were identified in P. bucharica and 40 in P. salicifolia. The chloroform extract from P. bucharica showed cytotoxicity in HeLa and HL-60 cells, with IC50 values of 26.07 and 29.42 ug/ml, respectively.

Las especies Phlomis (Phlomis bucharica Regel y P. salicifolia Regel) se han utilizado tradicionalmente por la gente de Uzbekistán como estimulante, tónico, diurético, y en el tratamiento de las úlceras, hemorroides, heridas y problemas ginecológicos. En el presente estudio, hemos caracterizado la composición química de los extractos no polares de P. bucharica y P. salicifolia por GLC-MS de alta resolución y se evaluó su citotoxicidad. Las concentraciones de ácido hexadecanoico en extractos de hexano y cloroformo fueron mayores en P. bucharica que en P. salicifolia. 1,8-cineol, alcanfor, borneol, se detectaron alfa-terpinol, timol, y acetato de isobornilo en P. bucharica pero no en P. salicifolia. Cerca de 45 componentes fueron identificados en P. bucharica y 40 en P. salicifolia. El extracto de cloroformo a partir de P. bucharica mostró citotoxicidad en células HL-60 y HeLa, con valores de CI 50 de 26,07 y 29,42 ug/ml, respectivamente.

Antiproliferative activity of yatein isolated from Austrocedrus chilensis against murine myeloma cells: cytological studies and chemical investigations.

CONTEXT: Fitzroya cupressoides (Molina) I. M. Johnst. and Austrocedrus chilensis (D. Don) Pic.Serm. & Bizzarri are two Chilean Cupressaceae that are naturally resistant to biodegradation. Secondary metabolites from these species display a variety of biological activities. OBJECTIVE: To evaluate the antiproliferative activity of two lignans, a diterpene and a flavonol isolated from A. chilensis and F. cupressoides, to elucidate their cytological effects on P3X murine myeloma cells. MATERIALS AND METHODS: The antiproliferative activity of yatein, isotaxiresinol, ferruginol, and isorhamnetin was evaluated in vitro using the MTT assay. The effect of yatein at the cellular level, due to its high antiproliferative activity was evaluated. P3X cells treated for 24 h with 12.5 and 25 µg/mL of yatein were also examined at the cytological level using immunofluorescence and scanning and transmission electron microscopy. RESULTS: Yatein, a lignan isolated from A. chilensis, potentially inhibited P3X murine myeloma cell proliferation, resulting in approximately 75% cell death in response to a 25 µg/mL treatment with the lignan. P3X cells lost membrane integrity at the nuclear and cytoplasmic levels, including organelles, in response to yatein treatment (12.5 µg/mL), and we observed changes in the cytoplasmic organization and distribution of microtubules. The other compounds tested had low activity. DISCUSSION AND CONCLUSIONS: Yatein is a lignan precursor of podophyllotoxin, a key agent in anticancer drugs. Due to its structural similarities to podophyllotoxin, yatein could have similar cytoplasmic target(s), such as the microtubular apparatus. These findings suggest that yatein may be of potential pharmacological interest and warrants further investigation in human cell lines.

Identification and characterization of plasma membrane proteins that bind to microtubules in pollen tubes and generative cells of tobacco.

The organization and function of microtubules in plant cells are important in many developmental stages. Connections between microtubules and the endomembrane system of plant cells have been discovered by microscopy, but the molecular characteristics of these relationships are mostly unknown except for a few cases. Using two antibodies raised against microtubule-associated proteins (MAPs) from maize, we have identified two polypeptides that share properties of the MAP family in the pollen tube of Nicotiana tabacum. The two polypeptides (with an apparent Mr of 161 and 90 kDa) bind efficiently to animal and plant microtubules and are found in association with the cellular membranes of the pollen tube, from which they can be solubilized with a zwitterionic detergent. One of these proteins has been purified and shown to promote the assembly of tubulin and, to a lesser extent, the bundling of microtubules. Subcellular fractionation indicated that the two proteins are associated with the plasma membrane compartment. The two proteins are found to co-localize in situ with cortical microtubules in the vegetative cytoplasm of tobacco pollen tubes; co-localization is also evident in the generative cell. According to these data, both the 161 and 90 kDa polypeptides are likely to mediate the interactions between the plasma membrane and microtubules in pollen tubes. In addition, functional data indicate that these MAP-like proteins take part in the process of microtubule assembly and reorganization occurring during cell growth. The evidence that both proteins associate with different cellular compartments also suggests a broad-spectrum role in mediating the dynamic relationships between microtubules and plant cell membranes.