Study of inhibition of germination of potato by ethylene.

In Canada, the potato (Solanum tuberosum) is by far the most cultivated vegetable and plays a major nutritional role. However, during storage, the potato can easily undergo germination. In this study we have shown the inhibition potential of ethylene as an anti-germinative agent acting especially on phenols. In both varieties assayed (Yukon Gold and Russet Burbank) in this study, the ethylene treatment led to a decrease in total phenol concentration of about 20%. The analysis of potato extracts showed the decrease of specific phenol concentrations which was dependant on the time and temperature of extraction. Our hypothese that the transformation of phenols into phenolic ethyl ethers via possible radical mechanism were then formulated and confirmed by LC and LC/MS.

Plants of the 'Libellus de Medicinalibus Indorum Herbis' from Mexico, 1552. Zephyranthes fosteri (Amaryllidaceae) Alkaloids.

The Libellus de Medicinalibus Indorum Herbis (Booklet of Indian Medicinal Plants) is the first book of medicinal plants written in the American continent. It was first published in 1939 as 'An Aztec Herbal'. One of the depicted plants is Huetzcanixochitl (laughing flower) interpreted as Zephyranthes fosteri (Amaryllidaceae). No chemical or pharmacological studies are reported for this species; so, we decide to investigate it. The GC/MS of the bulbs and aerial parts extracts indicated that they contain Amaryllidaceae alkaloids, among them: lycorine, 3-O-acetylpowelline, and norlycoramine. An unknown major alkaloid was isolated and identified by 1 H, 13 C-NMR and MS, as 3'-demethoxy-6-epimesembranol (1). The methanolic extract, the alkaloid fraction, and compound 1 inhibited acetylcholinesterase in vitro. Mesembrine alkaloids are found in Sceletium species (Aizoaceae). Several are known as serotonin recapture inhibitors and have been proposed as potential antidepressant drugs. The presence of 1 suggests that Z. fosteri was probably used in pre-Columbian times in Mexico as a 'stimulant and euphoriant', alike Sceletium tortuosum by several ethnic groups in South Africa.

Factors Affecting the Formation of 2:1 Host:Guest Inclusion Complexes of 2-[(R-Phenyl)amine]-1,4-naphthalenediones (PAN) in ß- and γ-Cyclodextrins.

The molecular hosts cyclodextrins form inclusion complexes with a wide variety of guests, resulting in complexes with various host:guest stoichiometries. In the case of a series of 19 1,4-naphthoquinolines as guests with either ß- or γ-cyclodextrin studied using electrospray mass spectroscopy, in most cases only 1:1 complexes were observed, with 2:1 host:guest complexes observed in just 6 out of 38 host:guest combinations. It is shown that these higher-order complexes were observed only in the case of small (or no) electronically withdrawing substituents, and were much less likely in the case of the larger γ-cyclodextrin host. The size and electronic properties of the substituents involved shows that both steric and electronic factors must be taken into account in predicting which cyclodextrin host:guest stoichiometries will be stable enough to form (or once formed, be robust enough to be observed in the ESI-MS experiments). It is clear that the prediction of host-guest stoichiometry for a specific host-guest pair is complicated, and involves a subtle interplay of both electronic and steric factors. However, there are definite trends, which can be used to help predict host:guest stoichiometry for a given host-guest pair.

On the synthesis of cyclodextrin-peptide conjugates by the Huisgen reaction.

A,D-substituted cyclodextrin (CDX) substituted on their primary rim side are ideal scaffolds for the macromolecular assembly and formation of templated structures. Their substitution can be achieved through various reactions. However, the use of the well-known Huisgen reaction in this context is under-reported. We present here results of the synthesis of model conjugates formed between CDX and representative peptides by click chemistry. Notably, bis-conjugation of peptides onto a unique scaffold promotes α-helix formation. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

XAS examination of glutathione-cobalt complexes in solution.

In the present work, we have investigated the coordination modes of cobalt with glutathione (γ-l-glutamyl-l-cysteinyl-glycine, GSH). A systematic study of cobalt-GSH complexes at basic and neutral pH has been undertaken with a multi-spectroscopic approach combined with quantum chemistry calculations. XAS (x-ray absorption spectroscopy) has been performed at the cobalt K edge in order to shed light into the cation coordination sphere and formal oxidation states. XANES (x-ray absorption near edge structure) enabled to show that in basic and neutral media, cobalt oxidation state is equal to +III and +II respectively. EXAFS (extended x-ray absorption fine structure) provided indications on the donor atoms involved in the coordination with cobalt as well as the bond lengths. DFT (density functional theory)-based calculations and NMR experiments have been performed to assess the most stable structure of the cobalt-GSH complex in basic conditions.

Acetylcholinesterase-inhibiting alkaloids from Zephyranthes concolor.

The bulbs and aerial parts of Zephyranthes concolor (Lindl.) Benth. & Hook. f. (Amaryllidaceae), an endemic species to Mexico, were found to contain the alkaloids chlidanthine, galanthamine, galanthamine N-oxide, lycorine, galwesine, and epinorgalanthamine. Since currently only partial and low resolution (1)H-NMR data for chlidanthine acetate are available, and none for chlidanthine, its 1D and 2D high resolution (1)H- and (13)C-NMR spectra were recorded. Unambiguous assignations were achieved with HMBC, and HSQC experiments, and its structure was corroborated by X-ray diffraction. Minimum energy conformation for structures of chlidanthine, and its positional isomer galanthamine, were calculated by molecular modelling. Galanthamine is a well known acetylcholinesterase inhibitor; therefore, the isolated alkaloids were tested for this activity. Chlidanthine and galanthamine N-oxide inhibited electric eel acetylcholinesterase (2.4 and 2.6 × 10(-5) M, respectively), indicating they are about five times less potent than galanthamine, while galwesine was inactive at 10(-3) M. Inhibitory activity of HIV-1 replication, and cytotoxicity of the isolated alkaloids were evaluated in human MT-4 cells; however, the alkaloids showed poor activity as compared with standard anti-HIV drugs, but most of them were not cytotoxic.

Development of normal phase-high performance liquid chromatography-atmospherical pressure chemical ionization-mass spectrometry method for the study of 6,6'-bis-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-benzo[1,2,4]-triazin-3-yl)-[2,2']-bipyridine hydrolytic degradation.

In the field of nuclear waste management, the 6,6'-bis-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-benzo[1,2,4]-triazin-3-yl)-[2,2']-bipyridine (CyMe(4)BTBP) is a polycyclic N-based molecule eligible to remove actinides from spent nuclear fuel by liquid-liquid extraction processes. In such processes, the organic phase containing the extracting molecules will undergo hydrolysis and radiolysis, involving degradation products. The purpose of this work was to develop a normal phase chromatography (NP-HPLC) coupled to atmospherical pressure chemical ionisation-mass spectrometry (APCI-MS) method to separate and identify degradation products of CyMe(4)BTBP dissolved in octanol, submitted to HNO(3) hydrolysis. 1 mol L(-1) HNO(3) hydrolysis conditions were used regarding the selective actinides extraction (SANEX) process, while 3 mol L(-1) HNO(3) conditions were applied for the group actinide extraction (GANEX) process. The first step consisted in optimizing the chromatographic separation conditions using a diode array detection (DAD). Retention behavior of a non hydrolyzed mixture of N,N'-dimethyl-N,N'-dioctyl-hexyloxyethyl-malonamide (DMDOHEMA), a malonamide used in the SANEX process to increase the kinetic of extraction, and CyMe(4)BTBP were investigated on diol-, cyano-, and amino-bonded stationary phases using different mobile phase compositions. These latter were hexane with different polar modifiers, i.e. dioxane, isopropanol, ethanol and methylene chloride/methanol. The different retention processes in NP-HPLC were highlighted when using various stationary and mobile phases. The second step was the setting-up of the NP-HPLC-APCI-MS coupling and the use of the low-energy collision dissociation tandem mass spectrometry (CID-MS/MS) of the precursor protonated molecules that allowed the separation and the characterization of the main hydrolytic CyMe(4)BTBP degradation product under a 3 mol L(-1) HNO(3) concentration. Investigation of the CID-MS/MS fragmentation pattern was used to suggest the potential ways leading to this hydrolysis degradation product. This NP-HPLC-APCI-MS method development is described for the first time for the CyMe(4)BTBP and should provide separation methods regarding the analysis of polycyclic N-based extracting molecules and more generally for the investigation of the organic phase coming from liquid-liquid extraction processes used in nuclear fuel reprocessing.

A new approach for the purification and characterisation of PA49.5, the main prebiotic of Lactococcus lactis subsp. cremoris.

The main autolysin PA49.5, an enzyme that hydrolyzes or destroys the components of a biological endogenous cell or a tissue, was purified 3045 times from the homogenate of a whole cell extract of Lactococcus lactis subsp. cremoris ATCC 9596 (Mc5), with a recovery yield of 52%. The purification of the protein was carried out through a micro-purification technique using SDS-BigCHAP polyacrylamide gel electrophoresis and concentrated with a Microcon-10 filtration system. SDS-polyacrylamide gel electrophoresis of the purified enzyme confirmed the presence of only one band having a molecular weight of 49.5 kDa. In view of its insolubility, PA49.5 contained in the cell extract precipitate was solubilized in the presence of 0.1% (w/v) of BigCHAP, a non-ionic detergent. Higher concentrations of this detergent completely inhibited the activity of solubilized PA49.5 or prevented its solubilization. The optimal pH and temperature for PA49.5 enzymatic activity are 7.5 and 45 degrees C respectively. In addition 0.1% or less of PA49.5 significantly increased Mc5 lysis. We observed 55% more lysis with 0.25 mug of purified PA49.5 compared to the control. Gas chromatography analysis of the components of the crude cell extract, of the precipitate and of the supernatant indicates the presence of at least 6 fatty acids. The long-chained fatty acids (e.g. C18:0 and C18:3) detected represent 81.65% of the precipitate from which PA49.5 was purified. Of these two acids, the C18:0 (stearic acid) alone represents 47.40% of the precipitate. Mc5 releases proteins at the beginning (major peak) and at the end (moderate peak) of the exponential stage of growth. Analysis by denaturing polyacrylamide gel electrophoresis with Mc5 cell walls incorporated as the autolysin's substrate identified a band corresponding to PA49.5 in the second peak of protein secretion.

A structural study of model peptides derived from HIV-1 integrase central domain.

The HIV-1 integrase (IN) catalyzes the integration of viral DNA in the human genome. In vitro the enzyme displays an equilibrium of monomers, dimers, tetramers and larger oligomers. However, its functional oligomeric form in vivo is not known. We report a study of the auto-associative properties of three peptides denoted K156, E156 and E159. These derive from the alpha4 helix of the IN catalytic core. The alpha4 helix is an amphipatic helix exposed at the surface of the protein and could be involved in the oligomerization process through its hydrophobic face. The peptides were obtained from the replacement of several amino acid residues by more helicogenic ones in the alpha4 helix peptide. K156 carries the basic residues Lys156 and Lys159, which have been shown important for the binding of IN to viral DNA. In E156 and E159 they are replaced with the acidic residue Glu. A fourth peptide K(E)156 obtained from the replacement of hydrophobic residues with Glu in K156 in order to abolish the auto-associative properties is used as a negative control. The capacity shown by peptides for alpha-helical formation is demonstrated by circular dichroism (CD) analysis performed in aqueous solution and in aqueous trifluoroethanol (TFE) mixtures. Both electrospray ionization mass spectrometry (ESI-MS) and glutaraldehyde chemical cross-linking show that peptides adopt different solvent-dependent equilibriums of monomers, dimers, trimers and tetramers. Oligomerization of peptides in aqueous solution is related to their ability to form helical structures. Addition of a small amount of TFE (<10%) stimulates helix stabilization and the interhelical hydrophobic contacts. Higher amounts of TFE alter the hydrophobic contacts and disrupt the oligomeric species. In addition to hydrophobic interactions, the patterns indicate that the biologically important Lys156 and Lys159 residues also participate in helix association. K(E)156 despite its ability to adopt a helical structure is unable to associate into oligomers, demonstrating the importance of hydrophobic contacts for oligomerization. Thus, the designed peptides provide us information on the functional properties of the alpha4 IN that seems to hold a dual role in DNA recognition and protein oligomerization.

Cytotoxic effects of mammea type coumarins from Calophyllum brasiliense.

Calophyllum brasiliense (Clusiaceae) is a big tree from the Tropical Rain Forests of the American continent. The organic extracts from the leaves yielded coumarins of the mammea type: mammea A/BA, A/BB, B/BA, B/BB, C/OA, C/OB, B/BA cyclo F, B/BB cyclo F, and isomammeigin. The triterpenoids friedelin and canophyllol, as well as the biflavonoid amentoflavone, protocatechuic and shikimic acids, were also obtained. Most of the isolated compounds were tested in vitro against K562, U251, and PC3 human tumor cell lines. The coumarins were cytotoxic against the three cell lines, the highest activity was shown by mammea A/BA (IC50 = 0.04 to 0.59 microM). The mixtures of mammea A/BA + A/BB, mammea B/BA + B/BB and mammea C/OA + C/OB were also highly active (IC50 < 4.05 microM). Friedelin was cytotoxic only against PC3, and U251 lines. Inhibition of HIV-1 reverse transcriptase was also assayed in vitro; however, none of the tested compounds (250 microM) prevented the activity of this enzyme. Most of the isolated compounds were also inactive against fourteen bacterial strains; however mammea A/BA + A/BB, and mammea C/OA + C/OB inhibited the growth of Staphylococcus aureus, S. epidermidis and Bacillus subtilis.