New synthetic strategies for xanthene-dye-appended cyclodextrins

Xanthene dyes can be appended to cyclodextrins via an ester or amide bridge in order to switch the fluorescence on or off. This is made possible through the formation of nonfluorescent lactones or lactams as the fluorophore can reversibly cyclize. In this context we report a green approach for the synthesis of switchable xanthene-dye-appended cyclodextrins based on the coupling agent 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM). By using 6-monoamino-β-cyclodextrin and commercially available inexpensive dyes, we prepared rhodamine- and fluorescein-appended cyclodextrins. The compounds were characterized by NMR and IR spectroscopy and MS spectrometry, their UV–vis spectra were recorded at various pH, and their purity was determined by capillary electrophoresis. Two potential models for the supramolecular assembly of the xanthene-dye-appended cyclodextrins were developed based on the set of data collected by the extensive NMR characterization

Novel β-cyclodextrin–eosin conjugates

Eosin B (EoB) and eosin Y (EoY), two xanthene dye derivatives with photosensitizing ability were prepared in high purity through an improved synthetic route. The dyes were grafted to a 6-monoamino-β-cyclodextrin scaffold under mild reaction conditions through a stable amide linkage using the coupling agent 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride. The molecular conjugates, well soluble in aqueous medium, were extensively characterized by 1D and 2D NMR spectroscopy and mass spectrometry. Preliminary spectroscopic investigations showed that the β-cyclodextrin–EoY conjugate retains both the fluorescence properties and the capability to photogenerate singlet oxygen of the unbound chromophore. In contrast, the corresponding β-cyclodextrin–EoB conjugate did not show either relevant emission or photosensitizing activity probably due to aggregation in aqueous medium, which precludes any response to light excitation

Enantioseparation and quantitative determination of two homologous beta amino acids found in Fabaceae plants

Non-protein amino acids are important metabolites of the Fabaceae family, possessing valuable biological effects in addition to their toxic properties. We have previously identified two non-protein amino acids homoproline and homopipecolic acid in Ononis species for the first time, and herein the study was extended to investigate further Fabaceae species (O. spinosa, O. arvensis, M. sativa, A. vulneraria) with medicinal, food or cosmetic uses. As the enantiomers of these beta amino acids can carry different activity or toxicity, our aim was to develop a chiral separation method for homoproline and homopipecolic acid enantiomers and apply it to plant samples. For this purpose, dansylated derivatives were prepared and a cyclodextrin-modified capillary electrophoresis in addition to a chiral HPLC method were developed. Although baseline separation was achieved by CE applying mono-(6-N-pyrrolidine-6-deoxy)-β-CD, mono-(6-N-piperidine-6-deoxy)-β-CD or sulfated-gamma-cyclodextrin at pH 6.0, the HPLC method was found to be more suitable for the analysis of the plant samples. Both homoproline and homopipecolic acid were confirmed in plant samples as racemates. The quantitative determination of homoproline and homopipecolic acid in several Fabaceae species were also aimed. Since these molecules can be found in the plants as esters, sample preparation was optimized to liberate the target molecules. Several SPE methods were tested for sample purification and a HPLC-MS/MS method using C8 stationary phase was developed and validated. The presence of homoproline and homopipecolic acid could be confirmed in all species ranging from 1 μg/g through 500 μg/g homopipecolic acid and 6 μg/g to 60 μg/g homoproline and significant changes could be observed between species, geographical origins, and botanical parts. Generally O. spinosa root samples were found to be the richest sources of the two amino acids, but a high variance could be observed between species

Comparative analysis of the full set of methylated β-cyclodextrins as chiral selectors in capillary electrophoresis

The chiral separation ability of the full library of methylated-β-cyclodextrins towards pharmacologically significant racemic drugs including basic compounds was studied by chiral capillary electrophoresis. The syntheses of all the methylated, single isomer β-cyclodextrins were revised and optimized and the aqueous solubility of the derivatives was unambiguously established. The three most relevant commercially available methylated isomeric mixtures were also included in the screening, so a total of ten various methylated CDs were investigated. The effects of the selector concentration on the enantiorecognition properties at acidic pH was investigated. Among the dimethylated β-cyclodextrins, the heptakis (2,6-di-O-methyl)-β-cyclodextrin isomer (2,6-DIMEB) resulted the most versatile chiral selector. Terbutaline was selected as a model compound for the in-depth investigation of host-guest enantiodiscrimination ability. The association constants between the two terbutaline enantiomers and 2,6-DIMEB were determined in order to support that the enantioseparation is driven by differences is host-guest-binding. The migration order of the enantiomers was confirmed by performing spiking experiments with the pure enantiomers. 1D and 2D NMR spectroscopy was applied to the 2,3-, and 2,6-DIMEB/terbutaline systems to rationalize at molecular level the different enantioseparation ability of the dimethylated β-cyclodextrin selectors. This article is protected by copyright. All rights reserved

The grass root endophytic fungus Flavomyces fulophazii: An abundant source of tetramic acid and chlorinated azaphilone derivatives

Fungal endophytes are remarkable sources of biologically active metabolites of ecological and pharmacological significance. In this study, fungal isolates producing yellow pigments and originating from grass roots, were identified as the recently described grass root colonizing dark septate endophyte (DSE), Flavomyces fulophazii (Periconiaceae, Pleosporales). While analyzing the metabolite composition of 17 isolates of this fungus, 11 previously undescribed compounds, including four tetramic acids (dihydroxyvermelhotin, hydroxyvermelhotin, methoxyvermelhotin, oxovermelhotin), and seven chlorinated azaphilones (flavochlorines A–G), together with the known tetramic acid vermelhotin, were tentatively identified by high performance liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS). Among them, flavochlorine A, flavochlorine G, hydroxyvermelhotin and vermelhotin could be isolated by preparative HPLC, thus their structures were also confirmed by nuclear magnetic resonance (NMR) spectroscopy. Vermelhotin was found to be the main compound, reaching its maximum level of 5.5 mg/g in the in vitro cultures of a selected F. fulophazii isolate. A significant amount of vermelhotin was isolated by preparative HPLC from these cultures (4.8 mg from 1.0 g lyophilized culture), confirming the practical utility of F. fulophazii in high-yield vermelhotin production. The main compounds of this endophyte expressed no activity in standardized plant bioassays (i.e., in the Lactuca sativa seed germination and Lemna minor growth tests). An antiproliferative study of the isolated compounds confirmed moderate activity of vermelhotin against a panel of twelve cancer cell lines, with IC50 ranges of 10.1–37.0 μM, without inhibiting the non-cancer Vero cells, suggesting its selectivity towards cancers

Nine-Dimensional Bioprofiles of Tunisian Sages (Salvia officinalis, S. aegyptiaca and S. verbenaca) by High-Performance Thin-Layer Chromatography – Effect-Directed Analyses

Ethyl acetate extracts of Tunisian Salvia aegyptiaca and S. verbenaca aerial parts and S. officinalis leaves were examined via bioanalytical profiling using high-performance thin-layer chromatography (HPTLC) combined with nine bioactivity assays, namely antibacterial (Aliivibrio fischeri, Bacillus subtilis, and Rhodococcus fascians), antifungal (Bipolaris sorokiniana, and Fusarium avenaceum), radical scavenging (DPPH center dot), and enzyme inhibitory (alpha-glucosidase, acetylcholinesterase, and lipase) ones. The screening, us-ing toluene -ethyl acetate - methanol 6:3:0.5 (V/V/V) as a mobile phase, revealed five bioactive zones (a-e) that were analyzed by HPTLC-electrospray ionization-mass spectrometry (ESI-MS). Zones b and c, observed exclusively in S. officinalis, were active in all assays except alpha-glucosidase, and only c inhib-ited F. avenaceum. Compounds in these zones were identified by HPLC-high resolution tandem MS (LC-HRMS/MS) as rosmanol/epirosmanol and methyl carnosate, respectively. In the bioactive zones a and e, corosolic/maslinic acid and ursolic/oleanolic acid isomer pairs were present, which could be identified in all three Salvia species after their HPTLC separation using pre-chromatographic derivatization with iodine and MS detection. The triterpenes inhibited B. subtilis and R. fascians bacteria and alpha-glucosidase enzyme. Linoleic and linolenic acids were detected in zone d, which showed strong lipase inhibition in all three sage species