Electrophoretic extraction and analysis of DNA from chitosan or poly-l-lysine-coated alginate beads

Alginate beads containing entrapped DNA were produced using both external and internal calcium sources, and coated with chitosan or poly-l-lysine membranes. The beads were assayed with DNase nuclease to determine formulation conditions offering the highest level of DNA protection fromnucleic acid hydrolysis, simulating gastrointestinal exposure. A method was developed to extract and assay intracapsular DNA through a modified agarose electrophoresis system. Both external and internally gelled beads were permeable to DNase (M_w=31 kDa), indicated by the absence of DNA after nuclease exposure. At low levels of DNase exposure, coated high guluronic content alginate beads offered a higher level of DNA protection compared with coated beads with low guluronic alginate. No apparent correlation was found with chitosan membrane molecular weight and degree of deacetylation; however, increasing poly-l-lysine molecular weight appeared to increase DNase exclusion from beads. At elevated levels of DNase exposure, DNA hydrolysis was evident within all coated beads with the exception of those coated with the highest molecular weight poly-l-lysine (M_w=197.1 kDa), which provided almost total nuclease protection. Optimal combination then for DNA protection from nucleases is a high guluronic alginate core, coated with high molecular weight poly-l-lysine.     

Reactions of pyridinium t-butoxycarbonylmethylide with perfluoropropene and some fluoroaromatics

Perfluorotoluene, pentafluoropyridine, 3-chlorotetrafluoropyridine, and 3,5-dichlorotrifluoropyridine react with pyridinium t-butoxycarbonylmethylide (I) in acetonitrile at 0 – 20 °C to yield, $\text{via}$ nucleophilic displacement of a 4-F substituent in each case, the new pyridinium methylides (II)–(V), respectively. Treatment of perfluoropropene with (I) gives 1-(t-butoxycarbonyl)-2-fluoro-3-(trifluoromethyl)- pyrrolo[1,2-$\text{a}$]pyridine (VI) and 1,3-bis(t-butoxycarbonyl)-2- (1,2,2,2-tetrafluoroethyl)pyrrolo[1,2-$\text{a}$]pyridine (VII), formation of the latter (minor) product providing evidence that the former arises $\text{via}$ a stepwise dipolar cycloaddition.

     

X=Y-ZH systems as potential 1,3-dipoles. The stereochemistry and regiochemistry of cycloaddition reactions of imines of α-amino-acid esters.

The stereochemistry and regiochemistry of cycloadditions of α-amino acid ester imines is dependent both on imine structure and on the reactivity of the dipolarophile. Phenylglycine imines undergo competing dipole stereomutation and cycloaddition with some dipolarophiles.     

Functionalized di- and tetrathia-crowns and their use to quantitatively separate and recover gold(III), palladium(II), silver(I) and mercury(II) Ions

Two new dithia-crowns containing a hydroxy group and 1,4,7,10-tetrathia-18-crown-6 containing an allyl-oxymethyl substituent were prepared in good yields. Two of these crowns were covalently attached to silica gel. The silica gel-bound thia-crowns were used to separate gold( III ), palladium( II ), silver( I ) and mercury( II ) ions from an aqueous 0.1 M nitric acid solution which also contained 1.0 M ferric chloride.     

Synthesis and conformational analysis by 1H NMR and restrained molecular dynamics simulations of the cyclic decapeptide [Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly]

Summary The design of enzyme mimics with therapeutic and industrial applications has interested both experimental and computational chemists for several decades. Recent advances in the computational methodology of restrained molecular dynamics, used in conjunction with data obtained from two-dimensional ¹H NMR spectroscopy, make it a promising method to study peptide and protein structure and function. Several issues, however, need to be addressed in order to assess the validity of this method for its explanatory and predictive value. Among the issues addressed in this study are: the accuracy and generizability of the GROMOS peptide molecular mechanics force field; the effect of inclusion of solvent on the simulations; and the effect of different types of restraining algorithms on the computational results. The decapeptide Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly, which corresponds to the sequence of ACTH_1–10, has been synthesized, cyclized, and studied by two-dimensional ¹H NMR spectroscopy. Restrained molecular dynamics (RMD) and time-averaged restrained molecular dynamics (TARMD) simulations were carried out on four different distance-geometry starting structures in order to determine and contrast the behavior of cyclic ACTH_1–10 in vacuum and in solution. For the RMD simulations, the structures did not fit the NOE data well, even at high values of the restraining potential. The TARMD simulation method, however, was able to give structures that fit the NOE data at high values of the restraining potential. In both cases, inclusion of explicit solvent molecules in the simulation had little effect on the quality of the fit, although it was found to dampen the motion of the cyclic peptide. For both simulation techniques, the number and size of the NOE violations increased as the restraining potential approached zero. This is due, presumably, to inadequacies in the force field. Additional TARMD vacuum-phase simulations, run with a larger memory length or with a larger sampling size (16 additional distance-geometry structures), yielded no significantly different results. The computed data were then analyzed to help explain the sparse NOE data and poor chymotryptic activity of the cyclic peptide. Cyclic ACTH_1–10, which contains the functional moieties of the catalytic triad of chymotrypsin, was evaluated as a potential mimic of chymotrypsin by measurement of the rate of hydrolysis of esters of L-and d-phenylalanine. The poor rate of hydrolysis is attributed to the flexibility of the decapeptide, the motion of the side chains, which result in the absence of long-range NOEs, the small size of the macrocycle relative to that of the substrate, and the inappropriate orientation of the Gly, His, and Ser residues. The results demonstrate the utility of this method in computer-aided molecular design of cyclic peptides and suggest structural modifications for future work based on a larger and more rigid peptide framework.     

Nioboxidfluoride mit Blockstrukturen — Elektronenmikroskopische Durchstrahlungsaufnahmen und ihre Simulation

Niobiumoxidefluorides Nb₅₉O₁₄₇F, Nb₃₁O₇₇F, Nb₆₅O₁₆₁F₃ and Nb₃₄O₈₄F₂ were prepared by reaction of Nb₂O₅ and Nb₃O₇F at 1 270°C. These niobiumoxidefluorides have blockstructures which were examined by high resolution electron microscopy. The observed images of the crystal structures were compared with computer simulated images.

     

Studies in azide chemistry. Part 12. One-pot conversion of 4-azidotetrafluoropyridine to 1,3,4-trifluoro-7,9-dimethyl-11H-pyrido[4,3-c]benzo[1,2]diazepine

Thermolysis of 4-azidotetrafluoropyridine in the presence of an excess of mesidine at 170 °C yields tetrafluoro-4-(2,4,6-trimethylphenylazo)- pyridine, which undergoes intramolecular dehydrofluorination $\text{in situ}$ to provide 1,3,4-trifluoro-7,9-dimethyl-11$\text{H}$-pyrido[4,3-$\text{c}$]benzo[1,2]diazepine.     

Mathematical properties of the vacuum polarization function

This paper is an investigation of certain mathematical properties of the vacuum polarization function (s). We show that (s) is a Herglotz function, has no complex zeroes, and belongs to the class of functions called typically real. In addition, we obtain upper bounds on the higher derivatives of (s), at s=0, given that we know the value of the first derivative at that point.Research supported in part by NASA Grant NSG-8035     

Filtered backprojection for the reconstruction of a high-resolution (4,2)D CH3-NH NOESY spectrum on a 29 kDa protein

Projection-reconstruction (PR) NMR enables rapid collection of multidimensional NMR data. NOESY represents a particularly difficult challenge for currently existing reconstruction algorithms, as it requires the quantitative reconstruction of an unknown number of peaks, at full sensitivity. We have demonstrated the successful application of PR-NMR to NOESY by determining the 4D methyl/amide NOESY spectrum of a 29 kDa protein, human carbonic anhydrase II, from 2D projections, using filtered backprojection for reconstruction. Compared with a 3D control spectrum, all expected peaks were faithfully reconstructed, with correct volumes and with no artifacts. Filtered backprojection thus provides a way to obtain high-resolution 4D NOESY data in the time required for conventional 3D data collection.     

Molecular orbital theoretical prediction of the isomeric products formed from reactions of arene oxides and related metabolites of polycyclic aromatic hydrocarbons

MO theoretical calculations based on the perturbational method of Dewar provide good correlation between predicted and observed structures of products formed during: (1) isomerization of arene oxides to phenols; (2) hydration and nucleophilic addition to arene oxides; and (3) dehydration of arene dihydrodiols. The method is equally applicable to the arene oxides, dihydrodiols, etc. derived from carcinogenic and noncarcinogenic polycyclic hydrocarbons. Extension to the related enzymatic reactions occurring during metabolism of carcinogenic hydrocarbons and to the reactions of the biologically active arene diolepoxides and aryloxirenes suggests the potential utility of this approach in predicting (a) metabolite structure and (b) the structural requirements for carcinogenic and mutagenic activity.