Conformationally Restricted 2-Substituted Wyosine Derivatives. 1H, 13C,and 15N NMR Study

Abstract

It was found by ¹H, ¹³C and ¹⁵N NMR study that substitution of 4,9-dihydro-4, 6-dimethyl-9-oxo-3-(2′,3′,5′-tri-O-acetyl-β-D-ribofuranosyl) imidazo [1,2-a]purine (wyosine triacetate, 1) at C2 position with electronegative groups CH₃O and C₆H₅CH₂O results in a noticeable electron distribution disturbance in the “left-hand” imidazole ring and a significant increase in the North conformer population of the sugar moiety.     

Conformational evaluation of labeled C3′‐O‐P‐13CH2‐O‐C4″ phosphonate internucleotide linkage,a phosphodiester isostere

Modified internucleotide linkage featuring the C3′‐O‐P‐CH₂‐O‐C4″ phosphonate grouping as an isosteric alternative to the phosphodiester C3′‐O‐P‐O‐CH₂‐C4″ bond was studied in order to learn more on its stereochemical arrangement, which we showed earlier to be of prime importance for the properties of the respective oligonucleotide analogues. Two approaches were pursued: First, the attempt to prepare the model dinucleoside phosphonate with ¹³C‐labeled CH₂ group present in the modified internucleotide linkage that would allow for a more detailed evaluation of the linkage conformation by NMR spectroscopy. Second, the use of ab initio calculations along with molecular dynamics (MD) simulations in order to observe the most populated conformations and specify main structural elements governing the conformational preferences. To deal with the former aim, a novel synthesis of key labeled reagent (CH₃O)₂P(O)¹³CH₂OH for dimer preparation had to be elaborated using aqueous ¹³C‐formaldehyde. The results from both approaches were compared and found consistent. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 514–529, 2009. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

Emission of Methane and Nitrous Oxide by Australian Mangrove Ecosystems

Abstract: The fluxes of the greenhouse gases methane (CH₄) and nitrous oxide (N₂O) were measured in mangrove wetlands in Queensland, Australia, using the closed chamber technique. Large differences in the fluxes of both gases from different study sites were observed, which presumably depended on differences in substrate availability. CH₄ emission rates were in the range of 20 to 350 μg m^‐2 h^‐1, whereas N₂O fluxes were lower, amounting to ‐ 2 to 14 μg m^‐2 h^‐1. In general, the field sites with high substrate availability showed higher emissions than sites with poor nutrient supply. This assumption is supported by the observation of dramatically increased N₂O emissions (150 ‐ 400 μg m^‐2 h^‐1) if study sites were artificially fertilised with additional N. As expected, N fertilisation did not alter CH₄ fluxes during the period of investigation. In the present study, it was confirmed that the mangrove vegetation may play a role as a transport path for CH₄ and N₂O by facilitating diffusion out of the soil. Prop roots from Rhizophora stylosa emitted CH₄ and N₂O at rates of 2.6 and 3.3 μg m^‐2 root surface h^‐1, respectively, whereas the soil of this stand acted as a sink for CH₄. As a consequence, the ecosystem as a whole could constitute a CH₄ source despite CH₄ uptake by the soil. In contrast to prop roots, the presence of pneumatophores in Avicennia marina led to a significant increase in CH₄ emissions from mangrove soils, but did not enhance N₂O emissions. These findings indicate that mangrove ecosystems may be considered a significant source of N₂O and that anthropogenic nutrient input into these ecosystems will lead to enhanced source strengths. For an up‐scaling of greenhouse gas emissions from mangrove forests to a global scale, more information is needed, particularly on the significance of vegetation.     

VARIATIONS IN THE SUBSTITUTED 3‐LINKED MANNANS CLOSELY ASSOCIATED WITH THE SILICIFIED WALLS OF DIATOMS1

The polysaccharides from cleaned frustules of the diatoms Pinnularia viridis (Nitzsch) Ehrenberg, Craspedostauros australis Cox, Thalassiosira pseudonana Hasle et Heimdal, and Nitzschia navis‐varingica Lundholm et Moestrup were extracted with hot alkali that dissolved the silica and were characterized by constituent sugar and linkage analyses. The polysaccharides from P. viridis were investigated further by permethylation, partitioning according to solubility, desulfation, and CD₃I‐methylation. Yields of carbohydrate in the hot alkali extracts ranged from 0.9% to 1.8% w/w based on the dry weight of the silica. Mannose was the dominant sugar in the polysaccharides from all four species (54–69 mol% of constituent sugars), although 14 other monosaccharides, including neutral sugars (glucose, galactose, xylose, arabinose, rhamnose, fucose), acidic sugars (glucuronic acid, galacturonic acid, 2‐O‐methylglucuronic acid), and O‐methylated neutral sugars (2‐O‐methylrhamnose, 3‐O‐methylrhamnose, 2,3‐di‐O‐methylrhamnose, 3‐O‐methylxylose, 4‐O‐methylxylose) were also detected in varying proportions among the four samples. The polysaccharides were predominantly composed of a 3‐linked mannopyranose backbone with a prevalence of linkage and/or substitution at O‐2 of the 3‐linked mannopyranosyl residues, and they were polyanionic, bearing uronic acid residues and/or sulfate esters. There were, however, species‐specific differences in the degree and position of substitution on the mannan backbone, the type and substitution patterns of the anionic substituents, and the type and linkage patterns of sugars other than mannose. Although definitive functions for these polysaccharides in diatom biology remain uncertain, a possible role in biosilicification is discussed.     

Deletion of fucose residues in plant N‐glycans by repression of the GDP‐mannose 4,6‐dehydratase gene using virus‐induced gene silencing and RNA interference

Production of pharmaceutical glycoproteins in plants has many advantages in terms of safety and reduced costs. However, plant‐produced glycoproteins have N‐glycans with plant‐specific sugar residues (core β‐1,2‐xylose and α‐1,3‐fucose) and a Lewis a (Le^a) epitope, i.e., Galβ(1‐3)[Fucα(1‐4)]GlcNAc. Because these sugar residues and glycan structures seemed to be immunogenic, several attempts have been made to delete them by repressing their respective glycosyltransferase genes. However, until date, such deletions have not been successful in completely eliminating the fucose residues. In this study, we simultaneously reduced the plant‐specific core α‐1,3‐fucose and α‐1,4‐fucose residues in the Le^a epitopes by repressing the Guanosine 5′‐diphosphate (GDP)‐D‐mannose 4,6‐dehydratase (GMD) gene, which is associated with GDP‐L‐fucose biosynthesis, in Nicotiana benthamiana plants. Repression of GMD was achieved using virus‐induced gene silencing (VIGS) and RNA interference (RNAi). The proportion of fucose‐free N‐glycans found in total soluble protein from GMD gene‐repressed plants increased by 80% and 95% following VIGS and RNAi, respectively, compared to wild‐type plants. A small amount of putative galactose substitution in N‐glycans from the NbGMD gene‐repressed plants was observed, similar to what has been previously reported GMD‐knockout Arabidopsis mutant. On the other hand, the recombinant mouse granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) with fucose‐deleted N‐glycans was successfully produced in NbGMD‐RNAi transgenic N. benthamiana plants. Thus, repression of the GMD gene is thus very useful for deleting immunogenic total fucose residues and facilitating the production of pharmaceutical glycoproteins in plants.     

Synthesis,structure, and biological active evaluation of a new cyclic tetranuclear copper(II) complex bridged both by oxamido and carboxylate groups

A new tetracopper(II) complex bridged both by oxamido and carboxylato groups, namely [Cu₄(dmaepox)₂(bpy)₂](NO₃)₂·2H₂O, where H₃dmaepox and bpy represent N‐benzoato‐N′‐ (3‐methylaminopropyl)oxamide and 2,2′‐bipyridine, was synthesized, and its structure reveals the presence of a centrosymmetric cyclic tetracopper(II) cation assembled by a pair of cis‐dmaepox^3–‐ bridged dicopper(II) units through the carboxylato groups, in which the endo‐ and exo‐copper(II) ions bridged by the oxamido group have a square‐planar and a square‐pyramidal coordination geometries, respectively. The aromatic packing interactions assemble the complex molecules to a two‐dimensional supramolecular structure. The reactivity toward DNA and protein bovine serum albumin (BSA) indicates that the complex can interact with herring sperm DNA through the intercalation mode and the binding affinity is dominated by the hydrophobicity and chelate ring arrangement around copper(II) ions and quenches the intrinsic fluorescence of BSA via a static process. The cytotoxicity of the complex shows selective cancer cell antiproliferative activity.     

O‐Glycosylated 24 kDa human growth hormone has a mucin‐like biantennary disialylated tetrasaccharide attached at Thr‐60

MS was used to characterize the 24 kDa human growth hormone (hGH) glycoprotein isoform and determine the locus of O‐linked oligosaccharide attachment, the oligosaccharide branching topology, and the monosaccharide sequence. MALDI‐TOF/MS and ESI‐MS/MS analyses of glycosylated 24 kDa hGH tryptic peptides showed that this hGH isoform is a product of the hGH normal gene. Analysis of the glycoprotein hydrolysate by high‐performance anion‐exchange chromatography with pulsed amperometric detection and HPLC with fluorescent detection for N‐acetyl neuraminic acid (NeuAc) yielded the oligosaccharide composition (NeuAc₂, N‐acetyl galactosamine₁, Gal₁). After β‐elimination to release the oligosaccharide from glycosylated 24 kDa hGH, collision‐induced dissociation of tryptic glycopeptide T6 indicated that there had been an O‐linked oligosaccharide attached to Thr‐60. The sequence and branching structure of the oligosaccharide were determined by ESI‐MS/MS analysis of tryptic glycopeptide T6. The mucin‐like O‐oligosaccharide sequence linked to Thr‐60 begins with N‐acetyl galactosamine and branches in a bifurcated topology with one appendage consisting of galactose followed by NeuAc and the other consisting of a single NeuAc. The oligosaccharide moiety lies in the high‐affinity binding site 1 structural epitope of hGH that interfaces with both the growth hormone and the prolactin receptors and is predicted to sterically affect receptor interactions and alter the biological actions of hGH.     

Reproductive biology of the guitarfish Rhinobatos percellens (Chondrichthyes,Rhinobatidae) from the São Paulo Coast,Brazil, western South Atlantic Ocean

The reproductive biology of the guitarfish Rhinobatos percellens was studied from 751 specimens caught by bottom pair trawlers off the coast of São Paulo, Brazil, between c. 24° 00′ S; 45° 15′ W and c. 25° 10′ S; 47° 52′ W, from September 2007 to August 2009. The total length (L_T) and total mass (M_T) relationship for males and females combined was M_T = 1·29E‐06 L_T^3·15 (r = 0·99, n = 751). The mean L_T of sexually mature specimens was 548 mm for males and 583 mm for females. Clasper growth was allometric and showed three distinct phases. Most claspers were calcified in specimens of c. 550 mm L_T. The mean diameter of the largest oocyte was 29·8 mm, the mean ovarian fecundity was seven oocytes and ovulation occurred between August and November. Uterine fecundity ranged from two to 13 embryos (mean of five embryos). Larger females had higher litter sizes and larger embryos; the size‐at‐birth was c. 200 mm L_T. The hepato‐somatic index oscillated seasonally for males and females; the gonado‐somatic index had little variation in males, but varied seasonally in females. The presence of many non‐pregnant adult females and of encapsulated eggs during two consecutive seasons suggests a resting period between gestations and the possibility of diapause.     

Evidence for the enzymatic transfer of N-acetylglucosamine from UDP-N-acetylglucosamine into dolichol derivatives and glycoproteins by calf brain membranes

Incubating white matter membranes with UDP-N-acetyl-[¹⁴C]glucosamine in the presence of Mg²⁺ and AMP resulted in the labeling of two major glycolipids, a minor glycolipid and several membrane-associated glycoproteins. The addition of AMP protected the labeled sugar nucleotide from degradation by a membrane-bound sugar nucleotide pyrophosphatase activity. While no labeled oligosaccharide lipid was recovered in a CHCl₃CH₃OHH₂O (10:10:3) extract after incubating with only UDP-N-acetyl-[¹⁴C] glucosamine, Mg²⁺, and AMP, the inclusion of unlabeled GDP-mannose led to the formation of an N-acetyl-[¹⁴C]glucosamine-labeled oligosaccharide lipid that was soluble in CHCl₃CH₃OHH₂O (10:10:3). The [GlcNAc-¹⁴C]oligosaccharide unit was released by treatment with 0.1 N HCl in 80% tetrahydrofuran at 50 °C for 30 min and appears to have the same molecular size as the lipid-linked [mannose-¹⁴C] oligosaccharide, formed enzymatically by white matter membranes as judged by their elution behavior on Bio-Gel P-6. The incorporation of N-acetyl-[¹⁴C]glucosamine into glycolipid was stimulated by exogenous dolichol monophosphate, but inhibited by UMP or tunicamycin, a glucosamine-containing antibiotic. Although UMP and tunicamycin drastically inhibited the labeling of glycolipid, these compounds had very little effect on the labeling of glycoproteins. The major glycolipids have the chemical and Chromatographic characteristics of N-acetylglucosaminylpyrophosphoryldolichol and N,N′-diacetylchitobiosylpyrophosphoryldolichol. When the labeled glycoproteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, four labeled polypeptides were observed, having apparent molecular weights of 145,000, 105,000, 54,000, and 35,000. Virtually all of the N-acetyl-[¹⁴C]glucosamine was released when the labeled glycopeptides, produced by pronase digestion, were incubated with an exo-β-N-acetylglucosaminidase, indicating that all of the N-acetyl-[¹⁴C]glucosamine incorporated under these conditions is attached to white matter membrane glycoproteins at nonreducing termini.     

Characterization of Secondary Metabolites in Flowers of Sanguisorba officinalis L. by HPLC‐DAD‐MSn and GC/MS

The investigations reported here focus on an in‐depth characterization of the secondary metabolite profile of Sanguisorba officinalis flowers. For this purpose, fresh flowers were extracted with MeOH/H₂O and EtOH/H₂O and the resulting crude extracts fractionated using CH₂Cl₂, AcOEt, and BuOH. Individual compounds were characterized by high performance liquid chromatography and gas chromatography coupled with mass spectrometric detection (HPLC‐DAD‐MSⁿ and GC/MS). MeOH/H₂O extraction and LC/MSⁿ investigations revealed the occurrence of flavonoid glycosides (quercetin, kaempferol), ellagitannin glycosides and four anthocyanins. Among the latter, two components, i. e., cyanidin‐malonyl‐glucose and cyanidin‐galloyl‐hexose, have not been reported for S. officinalis so far. Furthermore, phenylethylamine was characterized for the first time in Sanguisorba by pH value dependent extraction with CH₂Cl₂. In addition, AcOEt and BuOH extracts were analyzed by GC/MS both prior to and after acid hydrolysis of secondary metabolites. For this purpose, the extracts were treated with 1 n HCl solution (105 °C, 1 h) and derivatized with BSTFA. Analyses revealed the occurrence of several classes of phenolic compounds, such as gallic acid, hydroxybenzoic acid, hydroxycinnamic acid and ellagic acid derivatives. Additionally, the most prominent ursane‐type triterpenoid (ziyu‐glycoside I) from Sanguisorba and its corresponding aglycone isomers were detected and assigned based on their characteristic fragmentation patterns.     

Carbon dioxide,methane, and nitrous oxide exchanges in an age‐sequence of temperate pine forests

We investigated soil carbon dioxide (CO₂), methane (CH₄), and nitrous oxide (N₂O) exchanges in an age‐sequence (4, 17, 32, 67 years old) of eastern white pine (Pinus strobus L.) forests in southern Ontario, Canada, for the period of mid‐April to mid‐December in 2006 and 2007. For both CH₄ and N₂O, we observed uptake and emission ranging from ?160 to 245 μg CH₄ m^?2 h^?1 and ?52 to 21 μg N₂O m^?2 h^?1, respectively (negative values indicate uptake). Mean fluxes from mid‐April to mid‐December across the 4, 17, 32, 67 years old stands were similar for CO₂ fluxes (259, 246, 220, and 250 mg CO₂ m^?2 h^?1, respectively), without pattern for N₂O fluxes (?3.7, 1.5, ?2.2, and ?7.6 μg N₂O m^?2 h^?1, respectively), whereas the uptake rates of CH₄ increased with stand age (6.4, ?7.9, ?10.8, and ?23.3 μg CH₄ m^?2 h^?1, respectively). For the same period, the combined contribution of CH₄ and N₂O exchanges to the global warming potential (GWP) calculated from net ecosystem exchange of CO₂ and aggregated soil exchanges of CH₄ and N₂O was on average 4%, <1%, <1%, and 2% for the 4, 17, 32, 67 years old stand, respectively. Soil CO₂ fluxes correlated positively with soil temperature but had no relationship with soil moisture. We found no control of soil temperature or soil moisture on CH₄ and N₂O fluxes, but CH₄ emission was observed following summer rainfall events. LFH layer removal reduced CO₂ emissions by 43%, increased CH₄ uptake during dry and warm soil conditions by more than twofold, but did not affect N₂O flux. We suggest that significant alternating sink and source potentials for both CH₄ and N₂O may occur in N‐ and soil water‐limited forest ecosystems, which constitute a large portion of forest cover in temperate areas.     

Mitsunobu Reactions of 5‐Fluorouridine with the Terpenols Phytol and Nerol: DNA Building Blocks for a Biomimetic Lipophilization of Nucleic Acids

The cancerostatic 5‐fluorouridine (5‐FUrd; 1 ) was sequentially sugar‐protected by introduction of a 2′,3′‐O‐heptylidene ketal group (→ 2 ), followed by 5′‐O‐monomethoxytritylation (→ 3 ). This fully protected derivative was submitted to Mitsunobu reactions with either phytol ((Z and E)‐isomer) or nerol ((Z)‐isomer) to yield the nucleoterpenes 4a and 4b . Both were 5′‐O‐deprotected with 2% Cl₂CHCOOH in CH₂Cl₂ to yield compounds 5a and 5b , respectively. These were converted to the 5′‐O‐cyanoethyl phosphoramidites 6a and 6b , respectively. Moreover, the 2′,3′‐O‐(1‐nonyldecylidene) derivative, 7a , of 5‐fluorouridine was resynthesized and labelled at C(5′) with an Eterneon‐480 fluorophor^® (→ 7b ). The resulting nucleolipid was studied with respect to its incorporation in an artificial bilayer, as well as to its aggregate formation. Additionally, two oligonucleotides carrying terminal phytol‐alkylated 5‐fluorouridine tags were prepared, one of which was studied concerning its incorporation in an artificial lipid bilayer.     

Specific, reversible inactivation of yeast beta-hydroxy-beta-methylglutraryl-CoA reductase by CoA.

A non-stoichiometric material [Na₄(5′-IMP)₂·15H₂O]_0.2[Na₂(Pt(5′-IMP)₂ (trimethylenediamine))·13.5H₂O]_0.8 has been prepared and investigated by single-crystal X-ray methods and ¹H and ¹³C nmr spectroscopy. The compound is isomorphous with the monosodium and disodium salts of 5′-IMP and two Pt(II)-5′-IMP compounds previously reported to be non-stoichiometric. However, the structural changes in the packing motif of the 5′-IMP molecules induced on Pt(II) coordination are uniform only if the 5′-IMP complex containing (NH₃)₂Pt(II) is stoichiometric. Preliminary studies on the latter complex, synthesized in our laboratories, demonstrate that the complex is indeed stoichiometric.     

Crystal Structure and Conformation of 5‐Fluorouridine: Conformational Preferences for 5‐Fluorinated Pyranosides

Crystals of 5‐fluorouridine (5FUrd) have unit cell dimensions a = 7.716(1), b = 5.861(2), c = 13.041(1)Å, α = γ = 90°, β = 96.70° (1), space group P2₁, Z = 2, ρ_obs = 1.56 gm/c.c and ρ_calc = 1574 gm/c.c The crystal structure was determined with diffractometric data and refined to a final reliability index of 0.042 for the observed 2205 reflections (I ≥ 3σ). The nucleoside has the anti conformation [χ = 53.1(4)°] with the furanose ring in the favorite C2′–endo conformation. The conformation across the sugar exocyclic bond is g⁺, with values of 49.1(4) and ? 69.3(4)° for Φ_θc and Φ_∞ respectively. The pseudorotational amplitude τ_m is 34.5 (2) with a phase angle of 171.6(4)°. The crystal structure is stabilized by a network of N–H…O and O–H…O involving the N3 of the uracil base and the sugar O3′ and O2′ as donors and the O2 and O4 of the uracil base and O3′ oxygen as acceptors respectively. Fluorine is neither involved in the hydrogen bonding nor in the stacking interactions. Our studies of several 5‐fluorinated nucleosides show the following preferred conformational features: 1) the most favored anti conformation for the nucleoside [χ varies from ? 20 to + 60°] 2) an inverse correlation between the glycosyl bond distance and the χ angle 3) a wide variation of conformations of the sugar ranging froni C2′–endo through C3′–endo to C4′–exo 4) the preferred g⁺ across the exocyclic C4′–C5′ bond and 5) no role for the fluorine atom in the hydrogen bonding or base stacking interactions.     

Soil–atmosphere exchange of greenhouse gases in a Eucalyptus marginata woodland, a clover-grass pasture, and Pinus radiata and Eucalyptus globulus plantations

Soils provide the largest terrestrial carbon store, the largest atmospheric CO₂ source, the largest terrestrial N₂O source and the largest terrestrial CH₄ sink, as mediated through root and soil microbial processes. A change in land use or management can alter these soil processes such that net greenhouse gas exchange may increase or decrease. We measured soil–atmosphere exchange of CO₂, N₂O and CH₄ in four adjacent land‐use systems (native eucalypt woodland, clover‐grass pasture, Pinus radiata and Eucalyptus globulus plantation) for short, but continuous, periods between October 2005 and June 2006 using an automated trace gas measurement system near Albany in southwest Western Australia. Mean N₂O emission in the pasture was 26.6 μg N m⁻² h⁻¹, significantly greater than in the natural and managed forests (< 2.0 μg N m⁻² h⁻¹). N₂O emission from pasture soil increased after rainfall events (up to 100 μg N m⁻² h⁻¹) and as soil water content increased into winter, whereas no soil water response was detected in the forest systems. Gross nitrification through ¹⁵N isotope dilution in all land‐use systems was small at water holding capacity < 30%, and under optimum soil water conditions gross nitrification ranged between < 0.1 and 1.0 mg N kg⁻¹ h⁻¹, being least in the native woodland/eucalypt plantation < pine plantation < pasture. Forest soils were a constant CH₄ sink, up to −20 μg C m⁻² h⁻¹ in the native woodland. Pasture soil was an occasional CH₄ source, but weak CH₄ sink overall (−3 μg C m⁻² h⁻¹). There were no strong correlations (R < 0.4) between CH₄ flux and soil moisture or temperature. Soil CO₂ emissions (35–55 mg C m⁻² h⁻¹) correlated with soil water content (R < 0.5) in all but the E. globulus plantation. Soil N₂O emissions from improved pastures can be considerable and comparable with intensively managed, irrigated and fertilised dairy pastures. In all land uses, soil N₂O emissions exceeded soil CH₄ uptake on a carbon dioxide equivalent basis. Overall, afforestation of improved pastures (i) decreases soil N₂O emissions and (ii) increases soil CH₄ uptake.     

Oligonucleotide N3′→P5′ Phosphoramidates and Thio‐Phoshoramidates as Potential Therapeutic Agents

Nucleic acids analogues, i.e., oligonucleotide N3′→P5′ phosphoramidates and N3′→P5′ thio‐phosphoramidates, containing 3′‐amino‐3′‐deoxy nucleosides with various 2′‐substituents were synthesized and extensively studied. These compounds resist nuclease hydrolysis and form stable duplexes with complementary native phosphodiester DNA and, particularly, RNA strands. An increase in duplexes' melting temperature, ΔT_m, relative to their phosphodiester counterparts, reaches 2.2–4.0° per modified nucleoside. 2′‐OH‐ (RNA‐like), 2′‐O‐Me‐, and 2′‐ribo‐F‐nucleoside substitutions result in the highest degree of duplex stabilization. Moreover, under close to physiological salt and pH conditions, the 2′‐deoxy‐ and 2′‐fluoro‐phosphoramidate compounds form extremely stable triple‐stranded complexes with either single‐ or double‐stranded phosphodiester DNA oligonucleotides. Melting temperature, T_m, of these triplexes exceeds T_m values for the isosequential phosphodiester counterparts by up to 35°. 2′‐Deoxy‐N3′→P5′ phosphoramidates adopt RNA‐like C3′‐endo or N‐type nucleoside sugar‐ring conformations and hence can be used as stable RNA mimetics. Duplexes formed by 2′‐deoxy phosphoramidates with complementary RNA strands are not substrates for RNase H‐mediated cleavage in vitro. Oligonucleotide phosphoramidates and especially thio‐phosphoramidates conjugated with lipid groups are cell‐permeable and demonstrate high biological target specific activity in vitro. In vivo, these compounds show good bioavailability and efficient biodistribution to all major organs, while exerting acceptable toxicity at therapeutically relevant doses. Short oligonucleotide N3′→P5′ thio‐phosphoramidate conjugated to 5′‐palmitoyl group, designated as GRN163L (Imetelstat), was recently introduced as a potent human telomerase inhibitor. GRN163L is not an antisense agent; it is a direct competitive inhibitor of human telomerase, which directly binds to the active site of the enzyme and thus inhibits its activity. This compound is currently in multiple Phase‐I and Phase‐I/II clinical trials as potential broad‐spectrum anticancer agent.     

New Flavonoid Glucuronate Esters with Anti‐inflammatory Activities from Scutellaria regeliana

Two new flavonoid glucuronate esters, named scuregeliosides A and B ( 1 and 2 ), as well as three known ones, chrysin‐7‐O‐β‐d ‐glucuronic acid methyl ester ( 3 ), 5,7,4′‐trihydroxyflavone‐8‐O‐β‐d ‐glucuronic acid methyl ester ( 4 ) and apigenin‐7‐O‐β‐d ‐glucuronic acid ethyl ester ( 5 ), were isolated from the ethanolic extract of the whole plant of Scutellaria regeliana. Their chemical structures were elucidated on the basis of comprehensive spectroscopic analyses. Five compounds were screened for anti‐inflammatory activity in vitro. As the results, the inhibition rates of release of β‐glucuronidase from rat polymorphonuclear leukocytes were in the range of 42.2 – 47.1% at a concentration of 10 μm .     

Guanosine Nucleolipids: Synthesis,Characterization, Aggregation and X‐Ray Crystallographic Identification of Electricity‐Conducting G‐Ribbons

The lipophilization of β‐d ‐riboguanosine ( 1 ) with various symmetric as well as asymmetric ketones is described (→ 3a – 3f ). The formation of the corresponding O‐2′,3′‐ketals is accompanied by the appearance of various fluorescent by‐products which were isolated chromatographically as mixtures and tentatively analyzed by ESI‐MS spectrometry. The mainly formed guanosine nucleolipids were isolated and characterized by elemental analyses, ¹H‐, ¹³C‐NMR and UV spectroscopy. For a drug profiling, static topological polar surface areas as well as ¹⁰logP_OW values were calculated by an increment‐based method as well as experimentally for the systems 1‐octanol‐H₂O and cyclohexane‐H₂O. The guanosine‐O‐2′,3′‐ketal derivatives 3b and 3a could be crystallized in (D₆)DMSO – the latter after one year of standing at ambient temperature. X‐ray analysis revealed the formation of self‐assembled ribbons consisting of two structurally similar 3b nucleolipid conformers as well as integrated (D₆)DMSO molecules. In the case of 3a ? DMSO, the ribbon is formed by a single type of guanosine nucleolipid molecules. The crystalline material 3b ? DMSO was further analyzed by differential scanning calorimetry (DSC) and temperature‐dependent polarization microscopy. Crystallization was also performed on interdigitated electrodes (Au, distance, 5 μm) and visualized by scanning electron microscopy. Resistance and amperage measurements clearly demonstrate that the electrode‐bridging 3b crystals are electrically conducting. All O‐2′,3′‐guanosine ketals were tested on their cytostatic/cytotoxic activity towards phorbol 12‐myristate 13‐acetate (PMA)‐differentiated human THP‐1 macrophages as well as against human astrocytoma/oligodendroglioma GOS‐3 cells and against rat malignant neuroectodermal BT4Ca cells.     

Antibacterial,antifungal and in vitro antileukaemia activity of metal complexes with thiosemicarbazones

1‐phenyl‐3‐methyl‐4‐benzoyl‐5‐pyrazolone 4‐ethyl‐thiosemicarbazone (HL) and its copper(II), vanadium(V) and nickel(II) complexes: [Cu(L)(Cl)]·C₂H₅OH·( 1 ), [Cu(L)₂]·H₂O ( 2 ), [Cu(L)(Br)]·H₂O·CH₃OH ( 3 ), [Cu(L)(NO₃)]·2C₂H₅OH ( 4 ), [VO₂(L)]·2H₂O ( 5 ), [Ni(L)₂]·H₂O ( 6 ), were synthesized and characterized. The ligand has been characterized by elemental analyses, IR, ¹H NMR and ¹³C NMR spectroscopy. The tridentate nature of the ligand is evident from the IR spectra. The copper(II), vanadium(V) and nickel(II) complexes have been characterized by different physico‐chemical techniques such as molar conductivity, magnetic susceptibility measurements and electronic, infrared and electron paramagnetic resonance spectral studies. The structures of the ligand and its copper(II) ( 2 , 4 ), and vanadium(V) ( 5 ) complexes have been determined by single‐crystal X‐ray diffraction. The composition of the coordination polyhedron of the central atom in 2 , 4 and 5 is different. The tetrahedral coordination geometry of Cu was found in complex 2 while in complex 4 , it is square planar, in complex 5 the coordination polyhedron of the central ion is distorted square pyramid. The in vitro antibacterial activity of the complexes against Escherichia coli, Salmonella abony, Staphylococcus aureus, Bacillus cereus and the antifungal activity against Candida albicans strains was higher for the metal complexes than for free ligand. The effect of the free ligand and its metal complexes on the proliferation of HL‐60 cells was tested.     

Synthesis and Crystal Structure of Binuclear Copper(II) Complex Bridged by N‐(2‐Hydroxyphenyl)‐N′‐[3‐(diethylamino)propyl]oxamide: In Vitro Anticancer Activity and Reactivity Toward DNA and Protein

A new oxamido‐bridged bicopper(II) complex, [Cu₂(pdpox)(bpy)(CH₃OH)](ClO₄), where H₃pdpox and bpy stand for N‐(2‐hydroxyphenyl)‐N′‐[3‐(diethylamino)propyl]oxamide and 2,2′‐bipyridine, respectively, has been synthesized and characterized by elemental analyses, molar conductivity measurements, infrared and electronic spectra studies, and X‐ray single crystal diffraction. In the crystal structure, the pdpox^3? ligand bridges two copper(II) ions as cisoid conformation. The inner copper(II) ion has a {N₃O} square‐planar coordination geometry, while the exo‐ one is in a {N₂O₃} square‐pyramidal environment. There are two sets of interpenetrating two‐dimensional hydrogen bonding networks parallel to the planes (2 1 0) and (), respectively, to form a three‐dimensional supramolecular structure. The bicopper(II) complex exhibits cytotoxic activity against the SMMC7721 and A549 cell lines. The reactivity toward herring sperm DNA and bovine serum albumin revealed that the bicopper(II) complex can interact with the DNA by intercalation mode, and the complex binds to protein BSA responsible for quenching of tryptophan fluorescence by static quenching mechanism. © 2013 Wiley Periodicals, Inc. J BiochemMol Toxicol 27:412‐424, 2013; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21504