A dual-column HPLC method for the simultaneous determination of DHPG (9-[(1,3-dihydroxy-2-propoxy)methyl]guanine) and its mono and diesters in biological samples

A convenient method for the simultaneous determination of various DHPG species present in biological samples is presented. This method utilizes a cation exchange column (25 cm X 4.6 mm i.d.) coupled in series to a short reversed-phase column (5 cm X 4.6 mm i.d.). The mobile phase consists of methanol:0.005M ammonium phosphate buffer, pH 2.5. There is a large polarity difference between DHPG and its esters due to the non-polar side chain of the ester moiety. The simultaneous determination of the diesters, monoesters, and DHPG in these samples using only the cation exchange or the reversed-phase column is not possible without time-consuming gradient elution. In the reversed-phase mode alone, the esters are highly retained relative to DHPG, whereas the esters are only slightly retained on a cation exchange column and are insensitive to changes in pH and ionic strength of the mobile phase. However, a combination of these two columns provides interesting selectivity for these compounds and offers a unique way of controlling the retention times of these species relative to each other. The retention time of esters can be selectively altered (with respect to DHPG) by changing the composition of methanol in the mobile phase. In contrast, the retention time of DHPG is controlled by changing the buffer strength and pH of the mobile phase.     

Extreme stability of an unsolvated alpha-helix

High-temperature ion mobility measurements have been performed for alpha-helical Ac-A15K+H+ and globular Ac-KA15+H+ peptides. The alpha-helical and globular conformations do not melt into random coils as the temperature is raised. Instead, both conformations survive to the point where the peptide signals vanishes due to fragmentation. This occurs at 600 K for the globular Ac-KA15+H+ peptide and at 725 K for the alpha-helical Ac-A15K+H+. For the helical Ac-A15K+H+ peptide it appears that fragmentation is triggered by disruption of the helical conformation.     

Zur Kenntnis des Natrium-tetrahydroxoaluminat-chlorids Na2[Al(OH)4]Cl

On the Sodium Tetrahydroxoaluminate Chloride Na₂[Al(OH)₄]Cl The hitherto unknown compound Na₂[Al(OH)₄]Cl was prepared by crystallisation from a NaCl containing sodium aluminate solution. According to the X-ray single crystal investigation (tetragonal, space group P4/nmm, a = 7.541 Å, c = 5.059 Å, Z = 2) the compound represents the first example of a crystalline hydroxoaluminate with monomeric [Al(OH)₄]^? anions. Cl^? shows a quadratic anti prismatic coordination to 4 Na⁺ and over hydrogen bonds to 4 O^2? while Na⁺ is octahedrally coordinated by 4 O^2? and 2 Cl^? (axial). The results of the crystal structure analysis are confirmed by ²⁷Al and ²³Na MAS NMR investigations. Na₂[Al(OH)₄]Cl decomposes at about 200°C without intermediates under formation of β-NaAlO₂ and NaCl.     

Effects of As(III) binding on alpha-helical structure

As(III) displays a wide range of effects in cellular chemistry. Surprisingly, the structural consequences of arsenic binding to peptides and proteins are poorly understood. This study utilizes model alpha-helical peptides containing two cysteine (Cys) residues in various sequential arrangements and spatial locations to study the structural effects of arsenic binding. With i, and i + 1, i + 2, or i + 3 arrangements, CD spectroscopy shows that As(III) coordination causes helical destabilization when Cys residues are located at central or C-terminal regions of the helix. Interestingly, arsenic binding to i, i + 3 positions results in the elimination of helical structure and the formation of a relatively stable alternate fold. In contrast, helical stabilization is observed for peptides containing i, i + 4 Cys residues, with corresponding pseudo pairwise interaction energies (Delta G(pw) degrees) of -1.0 and -0.7 kcal/mol for C-terminal and central placements, respectively. Binding affinities and association rate constants show that As(III) binding is comparatively insensitive to the location of the Cys residues within these moderately stable helices. These data demonstrate that As(III) binding can be a significant modulator of helical secondary structure.     

Quantification of free and total salicylic acid in plants by solid-phase extraction and isocratic high-performance anion-exchange chromatography

Salicylic acid (SA) is an important signaling compound in plants and is involved in various defense responses. Here we report a new method for quantification of free and total soluble SA in Arabidopsis thaliana with 5-fluorosalicylic acid (5-FSA) as internal standard. The SA was isolated from leaf extracts by solid-phase extraction with phenyl-phase cartridges and selectively eluted as the cationic iron(III)-complex. Recoveries of SA and 5-FSA were equal and exceeded 90%. Free SA was subsequently released from the iron(III)-complex by addition of 2,2-bipyridyl and high-performance anion-exchange chromatography was performed on an NH₂ column. The SA appeared as last peak with a retention time of 15 min, baseline-separated from other substances. On-line detection was performed fluorimetrically for both SA and 5-FSA at an excitation wavelength of 300 nm and an emission wavelength of 410 nm, because both substances give similar fluorescence spectra. The detection limit for SA was 5 ng g^–1 FW for a sample size of 100 mg. Thus the main advantages of the method are highly selective sample preparation, increased sensitivity, reduced analysis time compared with reversed-phase HPLC, and use of a novel internal standard detectable under the same conditions as SA. The techniques described are applicable to other plant materials.     

Synthesis and characterization of PAni–SiO2 and PTh–SiO2 nanocomposites' thin films by plasma polymerization

In the present work, we explore the possibility to deposit polyaniline–silicon dioxide (PAni–SiO₂) and polythiophene–silicon dioxide (PTh–SiO₂) nanocomposites through a plasma polymerization route. The films were generated by spraying of mixtures of nano-sized silica particles dispersed in the liquid monomer into a plasma stream of the DC-plasma discharge reactor. The silica in the resulted polymer matrix changes the conduction mechanisms varying from ohmic to ballistic and traps inducing the space charged limited currents (SCLC). The silica modifies the morphology and composition of the deposited films.     

A density functional theory study of the gas-phase elimination reactions of 4-arylideneimino-1,2,4-triazol-3(2H )-ones and their 3(2H )-thione analogues

Theoretical studies on the thermolysis in the gas phase of 4-arylideneimino-1,2,4-triazol-3(2H)-ones and 4-arylideneimino-1,2,4-triazol-3(2H)-thiones were carried out using density functional theory methods, at the B3LYP/6-31G(d) and B3LYP/6-311+G(2d,p) levels of theory. The proposed reaction mechanism occurs in one step, leading to the formation of 3-hydroxy-(2H)-1,2,4-triazole or 3-mercapto-(2H)-1,2,4-triazole and a 4-substituted benzonitrile, via a six-membered cyclic transition state. The progress of the reactions was followed by means of the Wiberg bond indices. The results indicate that the transition states have character intermediate between reactants and products, and the calculated synchronicities show that the reactions are slightly asynchronous, in the case of triazolones, and show a higher asynchronicity in the case of triazolthiones. The bond-breaking processes are slightly more advanced than the bond-forming ones, indicating a small bond deficiency in the transition states. Kinetic and activation parameters for the reactions studied have been calculated and compared with available experimental data.From the Proceedings of the 28th Congreso deQuímicos Teóricos de Expresión Latina (QUITEL 2002)     

Antimalarial activity of n(6)-substituted adenosine derivatives (part 2)

We have investigated the in vitro antimalarial activity of a new series of adenosine derivatives. The results show that N(6)-(1-naphthylmethyl)-5'-deoxy-5'-(amido)adenosines as well as N(6)-(4-phenylbenzyl)-5'-deoxy-5'-(amido)adenosines display significant activity against the malaria-causing parasites, with the sterically demanding bisubstituted species reported being active in most cases in the low-micromolar range. The novel compounds with unusual substitution pattern were obtained applying an efficient convergent polymer-assisted solution-phase (cPASP) synthesis protocol. Thus, we were able to prepare a series of substituted derivatives in parallel that would have been difficult to synthesize by standard techniques. The scope and limitations of the synthetic methodology are discussed.     

Histidine in enzyme active centers.

The presence of histidine in the active center of an enzyme can be demonstrated by kinetic measurements, chemical modification, NMR spectroscopy or X-ray structure analysis. Histidine is the only naturally occurring amino acid to contain an imidazole residue as a side chain. The role of histidine in enzyme catalysis depends, inter alia, upon the special features of the imidazole residue: it thus tends to form hydrogen bonds, combines donor and acceptor properties and can take part in either nucleophilic or base catalysis. In some of these enzymes the position of each atom is known; however, the theories as to how the catalysis proceeds at a molecular level are controversial.