Chemical Constituents and Anticancer Activities of the Extracts from Phlomis × commixta Rech. f. (P. cretica × P. lanata).

The present work is the first report on the ingredients of the P. × commixta hybrid, a plant of the genus Phlomis. So far, thirty substances have been isolated by various chromatographic techniques and identified by spectroscopic methods, such as UV/Vis, NMR, GC-MS and LC-MS. The compounds are classified as flavonoids: naringenin, eriodyctiol, eriodyctiol-7-O-ß-D-glucoside, luteolin, luteolin-7-O-ß-D-glucoside, apigenin, apigenin-7-O-ß-D-glucoside, diosmetin-7-O-ß-D-glucoside, quercetin, hesperetin and quercetin-3-O-ß-D-glucoside; phenylpropanoids: martynoside, verbascoside, forsythoside B, echinacoside and allysonoside; chromene: 5,7-dihydroxychromone; phenolic acids: caffeic acid, p-hydroxybenzoic acid, chlorogenic acid, chlorogenic acid methyl ester, gallic acid, p-coumaric acid and vanillic acid; aliphatic hydrocarbon: docos-1-ene; steroids: brassicasterol and stigmasterol; a glucoside of allylic alcohol, 3-O-ß-D-apiofuranosyl-(1→6)-O-ß-D-glucopyranosyl-oct-1-ene-3-ol, was fully characterized as a natural product for the first time. Two tyrosol esters were also isolated: tyrosol lignocerate and tyrosol methyl ether palmitate, the latter one being isolated as a natural product for the first time. Moreover, the biological activities of the extracts from the different polarities of the roots, leaves and flowers were estimated for their cytotoxic potency. All root extracts tested showed a high cytotoxic activity against the Hep2c and RD cell lines.

Biological Activities of Ceratonia siliqua Pod and Seed Extracts: A Comparative Analysis of Two Cretan Cultivars.

Ceratonia siliqua L., commonly known as the carob tree, appears in most Mediterranean countries, often cultivated for the collection of its fruits to be used as food for humans and animals. This study was aimed at the phytochemical characterization of two common Cretan C. siliqua cultivars and the biological evaluation of deseeded pod and seed extracts regarding their putative use in cosmetics. Gas and liquid chromatographic techniques were used to assess their essential oil, fatty acid, and carbohydrate profiles. Cell-free assays, including free-radical scavenging; the inhibition of tyrosinase and collagenase; the blocking of advanced glycation end product (AGE) formation; along with assays in human skin fibroblast cultures, i.e., reactive oxygen species suppression, glutathione stimulation, and protection from oxidative stress and from ultraviolet (UVB) radiation, were also used. Extracts from both cultivars were found to possess antioxidant capacity, tyrosinase- and collagenase-inhibitory activities, an ability to block glucose-induced AGEs, and in certain cases, UVB absorbance and photoprotective activities. Seed extracts were in general more active, while the use of 30% aqueous methanol seemed to be more efficient than n-hexane for extraction. Serial partition of the most active extracts resulted in fractions with enriched biological activities. These properties make Cretan carob extracts and their fractions suitable candidates for use in cosmetics.

The Synthetic Microneurotrophin BNN27 Affects Retinal Function in Rats With Streptozotocin-Induced Diabetes.

BNN27, a C17-spiroepoxy derivative of DHEA, was shown to have antiapoptotic properties via mechanisms involving the nerve growth factor receptors (tropomyosin-related kinase A [TrkA]/neurotrophin receptor p75 [p75NTR]). In this study, we examined the effects of BNN27 on neural/glial cell function, apoptosis, and inflammation in the experimental rat streptozotocin (STZ) model of diabetic retinopathy (DR). The ability of BNN27 to activate the TrkA receptor and regulate p75NTR expression was investigated. BNN27 (2,10, and 50 mg/kg i.p. for 7 days) administration 4 weeks post-STZ injection (paradigm A) reversed the diabetes-induced glial activation and loss of function of amacrine cells (brain nitric oxide synthetase/tyrosine hydroxylase expression) and ganglion cell axons via a TrkA receptor (TrkAR)-dependent mechanism. BNN27 activated/phosphorylated the TrkAY490 residue in the absence but not the presence of TrkAR inhibitor and abolished the diabetes-induced increase in p75NTR expression. However, it had no effect on retinal cell death (TUNEL+ cells). A similar result was observed when BNN27 (10 mg/kg i.p.) was administered at the onset of diabetes, every other day for 4 weeks (paradigm B). However, BNN27 decreased the activation of caspase-3 in both paradigms. Finally, BNN27 reduced the proinflammatory (TNFα and IL-1ß) and increased the anti-inflammatory (IL-10 and IL-4) cytokine levels. These findings suggest that BNN27 has the pharmacological profile of a therapeutic for DR, since it targets both the neurodegenerative and inflammatory components of the disease.

Use of costic acid, a natural extract from Dittrichia viscosa, for the control of Varroa destructor, a parasite of the European honey bee.

Costic acid has been isolated from the plant Dittrichia viscosa and its efficacy against Varroa destructor, a parasite of Apis mellifera, the European honey bee, has been studied. Costic acid exhibited potent in vivo acaricidal activity against the parasite. Initial experiments showed that the compound is not toxic for human umbilical vein endothelial cells (HUVEC) at concentrations of up to 230 micromolar (µM), indicating that costic acid could be used as a safe, low-cost and efficient agent for controlling varroosis in honey bee colonies.

Chemical and genetic characterization of Phlomis species and wild hybrids in Crete.

The genus Phlomis is represented in the island of Crete (Greece, Eastern Mediterranean) by three species Phlomis cretica C. Presl., Phlomis fruticosa L., the island endemic Phlomis lanata Willd. and three hybrids Phlomis x cytherea Rech.f. (P. cretica x P. fruticosa), Phlomis x commixta Rech.f. (P. cretica x P. lanata) and Phlomis x sieberi Vierh. (P. fruticosa x P. lanata). This work describes (a) the profile of hybrids and parental species concerning their volatile compounds, (b) the suitability of ribosomal nuclear (ITS region), chloroplast (trnH-psbA), and AFLP markers to identify hybrids and (c) their competence to characterize the different chemotypes of both hybrids and their parental species. The cluster analysis and PCA constructed from chemical data (volatile oils) suggest that there are three groups of taxa. Group IA includes P. cretica and P. fruticosa, group IB includes P. x cytherea, whereas group II consists of P. x commixta, P. x sieberi and P. lanata. Volatile compounds detected only in the hybrids P. x sieberi and P. x commixta correspond to the 3% of the total compounds, value that is much higher in P. x cytherea (21%). Neighbor-joining, statistical parsimony analysis and the observations drawn from ribotypes spectrum of ITS markers divided Phlomis species in two groups, P. lanata and the complex P. cretica/P. fruticosa. In contrast to the ITS region, the plastid DNA marker follows a geographically related pattern. Neighbor-Net, PCA and Bayesian assignment analysis performed for AFLP markers separated the genotypes into three groups corresponding to populations of P. cretica, P. fruticosa, and P. lanata, respectively, while populations of P. x commixta, P. x cytherea, and P. x sieberi presented admixed ancestry. Most of the P. x cytherea samples were identified as F1 hybrids by Bayesian assignment test, while those of P. x commixta and P. x sieberi were identified as F2 hybrids. Overall, high chemical differentiation is revealed in one of the three hybrids, which is likely related with niche variation. Moreover, molecular markers show potential to identify Phlomis taxa.

Fullerene-coumarin dyad as a selective metal receptor: synthesis, photophysical properties, electrochemistry and ion binding studies.

A coumarin derivative with a malonate unit has been synthesized and used for the preparation of a fullerene-coumarin dyad through the Bingel cyclopropanation method. The newly synthesized dyad is soluble in organic solvents and has been fully characterized with traditional spectroscopic techniques. Electronic interactions between the two components of the dyad were probed with the aid of UV/Vis spectroscopy, fluorescence emission, and electrochemistry measurements. Our studies clearly show the presence of electronic interactions between C(60) and modified coumarin in the ground state; efficient electron-transfer quenching of the singlet excited state of the coumarin moiety by the appended fullerene sphere was also observed. Time-resolved fluorescence measurements revealed lifetimes for the coumarin-C(60) dyad at a maximum of 50 ps, while the quantum yield was reaching unity. Additionally, the redox potentials of the C(60)-coumarin dyad were determined and the energetics of the electron-transfer processes were evaluated. Finally, after alkaline treatment of C(60)-coumarin, which resulted in the deprotection of carboxylate units, the dyad was tested as a metal receptor for divalent metal cations; ion competition studies and fluorescence experiments showed binding selectivity for lead ions.

A "turn-on" coumarin-based fluorescent sensor with high selectivity for mercury ions in aqueous media.

The synthesis and spectral profile of a "turn-on" fluorescent sensor selective for Hg(2+) ions in aqueous media is described.

Coumarin-based ratiometric fluorescent indicators with high specificity for lead ions.

Total syntheses and spectral properties of fluorescent Pb(2+) indicators are reported.

ICPBCZin: a red emitting ratiometric fluorescent indicator with nanomolar affinity for Zn2+ ions.

A new fluorescent Zn2+ indicator, namely, ICPBCZin was synthesized and the spectral profile of its free and Zn2+ bound forms was studied. The newly synthesized zinc indicator incorporates as chromophore the chromeno [3',2':3,4]pyrido[1,2a] [1,3]benzimidazole moiety and belongs to the dicarboxylate-type of zinc probes. The compound is excited with visible light, exhibits high selectivity for zinc in the presence of calcium and other common biological ions, and its Zn2+ dissociation constant is 4.0 nM. Fluorescence spectra studies of ICPBCZin indicated a clear shift in its emission wavelength maxima upon Zn2+ binding, as it belongs to the class of Photoinduced Charge Transfer (PCT) indicators, along with changes in fluorescence intensity that enable the compound to be used as a ratiometric, visible-excitable Zn2+ probe.

Anti-inflammatory and antioxidant activity of coumarins designed as potential fluorescent zinc sensors.

A series of coumarin analogs, designed and synthesised as potential fluorescent zinc probes were evaluated for their biological activity as anti-inflammatory and antioxidant agents. The effect of the synthesised compounds on inflammation, using the carrageenin-induced rat paw oedema model, was studied. In general, the compounds were found to be potent anti-inflammatory agents (26.5-64%). Compound 5 was found to interact significantly with 1,1-diphenyl-2-picryl-hydrazyl stable free radical (DPPH) whereas the remainder were inactive in this assay. The compounds inhibit in general the soybean lipoxygenase and scavenge superoxide anion radicals. The anti-inflammatory activity seems to be connected with their reducing activity. Their RM values were determined as an expression of their lipophilicity. Theoretical calculations of their lipophilicity as clog P were performed indicating that only a poor relationship exists between their lipophilicity and anti-inflammatory activity.

Quercetin exhibits a specific fluorescence in cellular milieu: a valuable tool for the study of its intracellular distribution.

The elaboration of novel techniques for flavonoid intracellular tracing would elucidate the compounds' absorption and bioavailability and assist molecular and pharmacological approaches, as they are promising candidates for drug development. This study exploited the properties of quercetin (3,3',4',5,7-pentahydroxyflavone), found in high concentrations in the majority of edible plants. Through the use of UV-vis spectroscopy, confocal microscopy, and HPLC-ESI-MS, native quercetin, at physiologically relevant concentrations, was found to exhibit a specific fluorescence (488 nmex/500-540 nmem) upon internalization. This fluorescence shift is due to a non-covalent binding to intracellular targets (probably proteins) and compatible with the settings applied in confocal microscopy. This property provides a valuable, selective alternative technique for quercetin tracing in cellular systems, permitting the quantitative evaluation of its transit at pharmacologically relevant concentrations and the validation of a number of already described molecular functions.

ICPBC and C12-ICPBC: two new red emitting, fluorescent Ca2+ indicators excited with visible light.

Two new, visible-excited and red-emitting fluorescent Ca(2+) indicators were synthesized and the spectral profiles of their free and Ca(2+) bound forms were studied. The fluorescent properties of these probes are due to the extended conjugation of the chromeno[3',2':3,4]pyrido[1,2a][1,3]benzimidazole chromophore incorporated in their BAPTA-type, Ca(2+) chelating structure. The compounds, namely ICPBC and its N-dodecyl analog C12-ICPBC exhibit Ca(2+) dissociation constants of 7.7 and 18.0 microM, respectively. The fluorescence spectra of the probes showed a clear shift in excitation wavelength maxima upon Ca(2+) binding along with a large Stokes shift and changes in fluorescence intensity, indicating their potential use as Ca(2+) indicators. The ability of ICPBC to trace high calcium spikes was tested in the human HepG2 cell line with positive results.

Composition and insect attracting activity of the essential oil of Rosmarinus officinalis.

The essential oil and a number of extracts of Rosmarinus officinalis L. in solvents of increasing polarity were isolated, and their components identified and tested as pest control agents. Ethanol and acetone extracts attract grape berry moth Lobesia botrana. However, none of the extracts had a significant effect on western flower thrips Frankliniella occidentalis, which is attracted by 1,8-cineole, a major essential oil component.

The coumarin moiety as chromophore of fluorescent ion indicators in biological systems.

Fluorescent probes have evolved into an extremely useful tool for the detection of ions in biological systems. The design of ion indicators is based in the proper choice of the ion chelating group as well as the chromophore moiety. The chromophores of choice should fulfill a number of requirements concerning the photostability of the group, the range of the excitation and emission wavelengths of the indicators, the Stokes shift, the fluorescence quantum yield, the excitation and/or emission wavelength shift upon coordination of the probe with its target ion, the lipophilicity of the indicators, and their possible cell toxicity. Coumarin and its analogues have been extensively used in ion detection by incorporation of the coumarin chromophore in the larger indicator framework. Coumarins fulfill all the aforementioned requirements since they are relatively photostable and their excitation and emission maxima, in many cases, are long enough to minimise "background" fluorescence of cellular components, tissues and biological fluids. They exhibit Stokes shifts large enough to avoid significant overlap of the excitation and emission spectra, their fluorescence quantum yields allow for ion detection at low indicator concentrations, and they can be introduced to cells either by microinjection or as membrane permeable derivatives without causing cell death. Synthetic approaches, aiming at the optimisation of indicator properties, have extended the conjugated coumarin system either by introduction of substituents or by expansion of the heterocyclic system. In this review, the basic rationale for the selection of the particular coumarin analogues is analysed, synthetic pathways leading to the desired structures are presented, and properties and relative advantages in the use of these probes are described.

Correction of the structure of a new sesquiterpene from Cistus creticus ssp. creticus.

In an attempt to identify the structure of a sesquiterpene from Cistus creticus ssp. creticus proposed in the literature as 1,1,4a,6-tetramethyl-5-methylene-1,2,3,4,4alpha,5,8,8alpha-octahydronaphthalene, the synthesis of its cis isomer 2 was carried out in 11 steps and 9.5% yield. Comparison of the spectra of 2 and those reported earlier for the synthetic trans isomer 1 with the spectral profile of the isolated natural product indicated that the latter was not compatible with either 1 or 2. The correct structure was assigned, by detailed spectroscopic analysis of the natural product, as 6-isopropenyl-4,4a-dimethyl-1,2,3,4,4a,5,6,7-octahydronaphthalene (3).