Enzyme labeling in steroid enzyme immunoassays. Comparison of the p-nitrophenyl ester and N-succinimidyl ester methods.

Enzyme labeling of steroids by the p-nitrophenyl ester method was investigated in comparison with the N-succinimidyl ester method. The active ester of a testosterone or 11-deoxycortisol derivative was treated with beta-galactosidase and horseradish peroxidase to give labeled antigens. Various molar ratios of steroid to enzyme and pH conditions were tested. Satisfactory immunoreactivities with an anti-steroid antibody in each enzyme immunoassay system were obtained with the labeled antigens prepared at pH 8.5 by the use of molar ratios higher than 30. The enzyme labeling method should be useful in the case of polar steroids or drugs, since the p-nitrophenyl ester is relatively stable when compared with the N-succinimidyl ester.     

Reactivity of polymer substituents. Aminolysis of p-nitrophenyl ester residues attached to various polymer backbones

Styrene, methyl acrylate, and N,N-dimethylacrylamide were copolymerized with small proportions of p-nitrophenyl acrylate or p-nitrophenyl esters of CH₂?CHCONH(CH₂)_nCOOH (n = 1,3,5). The aminolysis of these copolymers in dioxane and chlorobenzene solution was compared with the corresponding reaction of low molecular weight analogs. The ratio of the reactivity of the polymer substituent and its analog was found to be insensitive to the nature of the amine but strongly dependent on the nature of the polymer backbone. Poly(dimethylacrylamide) chains carrying active ester substituents were in some cases much more reactive than their analogs due to the activating effect of the dimethylamide groups. In the case of the p-nitrophenyl acrylate–styrene copolymer, the aminolysis exhibited a dispersion of the rate constant due to its sensitivity to stereoisomerism in the vicinity of the active ester, but no similar effects were observed with the other copolymers.     

Catalysis of ester aminolysis by cyclodextrins. The reaction of alkylamines with p-nitrophenyl alkanoates

The effects of four cyclodextrins (alpha-CD, beta-CD, hydroxypropyl-beta-CD, and gamma-CD) on the aminolysis of p-nitrophenyl alkanoates (acetate to heptanoate) by primary amines (n-propyl to n-octyl, isobutyl, isopentyl, cyclopentyl, cyclohexyl, benzyl) in aqueous solution have been investigated. Rate constants for amine attack on the free and CD-bound esters (k(N) and k(cN)) have ratios (k(cN)/k(N)) varying from 0.08 (retardation) to 180 (catalysis). For the kinetically equivalent process of free ester reacting with CD-bound amine (k(Nc)), the ratios k(Nc)/k(N) vary from 0.2 to 28. Either way, there is evidence of catalysis in some cases and retardation in others. Changes in reactivity parameters with structure indicate more than one mode of transition state binding to the CDs. Short esters react with short alkylamines by attack of free amine on the ester bound by its aryl group, but for longer amines, free ester reacts with CD-bound amine. Reaction of long esters with long amines, which is catalyzed by beta-CD and gamma-CD, involves inclusion of the alkylamino group and possibly the ester acyl group. The larger cavity of gamma-CD may allow the inclusion of the ester aryl group, as well as the alkylamino group, in the transition state. Reaction between an ester bound to the CD by its acyl group and free amine appears not to be important.     

Tail ordering due to headgroup hydrogen bonding interactions in surfactant monolayers at the water-oil interface

Interactions between surfactants, and the resultant ordering of surfactant assemblies, can be tuned by the appropriate choice of head- and tailgroups. Detailed studies of the ordering of monolayers of long-chain n-alkanoic and n-alkanol monolayers at the water-vapor interface have demonstrated that rigid-rod all-trans ordering of the tailgroups is maintained upon replacing the alcohol with a carboxylic acid headgroup. In contrast, at the water-hexane liquid-liquid interface, we demonstrate that substitution of the -CH(2)OH with the -COOH headgroup produces a major conformational change of the tailgroup from disordered to ordered. This is demonstrated by the electron density profiles of triacontanol (CH(3)(CH(2))(29)OH) and triacontanoic acid (CH(3)(CH(2))(28)COOH) monolayers at the water-hexane interface, as determined by X-ray reflectivity measurements. Molecular dynamics simulations illustrate the presence of hydrogen bonding between the triacontanoic acid headgroups that is likely responsible for the tail ordering. A simple free energy illustrates the interplay between the attractive hydrogen bonding and the ordering of the tailgroup.     

Enzyme-modulated cleavage of dsDNA for studying interfacial biomolecular interactions.

This work describes the chemistry and methodology for constructing multilayers of bis-biotinylated dsDNA on metal substrates after enzyme cleavage and demonstrates its use for amplified microgravimetric and impedimetric analyses of anticancer drug, cisplatin. Specific chemical modification of dsDNA prior to immobilization was achieved via a bisulfite-catalyzed transamination of cytosine after endonuclease cleavage of plasmid DNA. The specificity of the reaction of cytosine residues at ss- versus dsDNA loci after endonuclease cleavage was characterized using circular dichroism, mass spectrometry, and absorption spectrophotometry. The biotinylated dsDNA consisting of 2961 base pairs was then used as a ligand at avidin-modified gold electrodes. Ac impedance spectroscopy and quartz crystal microbalance measurements clearly showed that the response to cisplatin increased linearly with target concentrations. The impedance spectroscopy resulted in a detection limit of 1 nM and a surface density of 4.8 x 10(13) molecules/0.1 cm(2). The immobilization of dsDNA on surfaces is a significant improvement over existing approaches in that it enables the attachment of long pieces of unmodified double-stranded DNA via a simple biotinylation step. The immobilization technique provides a generic approach for dsDNA-based sensor development and for monitoring DNA-analyte interactions.     

Enzyme labeling in steroid enzyme immunoassays. The p-nitrophenyl ester method for alkaline phosphatase and glucose oxidase labelings.

The p-nitrophenyl ester method was assessed as an enzyme labeling technique. The active ester of a carboxylated testosterone derivative was treated with alkaline phosphatase and glucose oxidase to give labeled antigens, using various molar ratios of steroid to enzyme. Satisfactory immunoreactivities with an anti-testosterone antibody in an enzyme immunoassay system were obtained with the labeled antigens prepared at pH 8.5 by the use of molar ratios higher than 30 and 10, respectively, in the alkaline phosphatase and glucose oxidase labelings.     

Intramolecular interactions I. The use of someMannich bases of naphthols as model compounds for intramolecular hydrogen bonding

Some model compounds for intramolecular hydrogen bonding are presented. The nmr, IR, UV and fluorescence spectra are shown to be suited for the study of such hydrogen bonds.

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Intramolekulare Wechselwirkungen, I. Einige Mannich-Basen von Naphtholen als Modellverbindungen für intramolekulare Wasserstoffbrückenverbindung

Zusammenfassung Eine Klasse von Verbindungen zur Untersuchung von intramolekularen Wasserstoffbrücken wird vorgestellt. NMR-, IR-, UV- und Fluoreszenzspektroskopie sind bestens geeignet, derartige Wasserstoffbrücken zu studieren.

     

An exactly solvable model for a ternary solution with three-body interactions and orientationally dependent bonding

A model is considered in which the bonds of a honeycomb lattice are covered by rodlike molecules of types AA, BB, and AB. Neighboring molecular ends have three-body and orientation-dependent interactions. The model is shown to be equivalent to a spin-1/2 Ising model on the same lattice with a field, but with only pairwise interactions. Symmetric and asymmetric coexistence surfaces for the separation into an AA-rich and a BB-rich phase are calculated exactly.     

Dilithium tetrachlorocuprate. A reagent for regioselective cleavage of epoxides to chlorohydrins.

The title compound reacts with epoxides to selectively afford the vicinal chlorohydrin resulting from attack by chloride at the less substituted carbon atom.     

A new cleavage strategy for ester linked polymer supports: generation of a tertiary alcohol library

The addition of different Grignard reagents to polymer bound esters resulting in the generation of a tertiary alcohol library with concomitant cleavage is described.     

Synthesis of 5-fluorouracil-imprinted polymers with multiple hydrogen bonding interactions.

The antitumor active compound 5-fluorouracil (5-FU) was used as a target molecule and 5-FU-imprinted polymers were synthesized using 2,6-bis(acrylamido)pyridine and/or 2-(trifluoromethyl)acrylic acid as functional monomers. The 5-FU-imprinted polymers showed a higher affinity for 5-FU than that for 5-FU derivatives. By using both functional monomers simultaneously, the affinity and separation for 5-FU were improved.     

Study of hydrogen bonding interactions relevant to biomolecular structure and function

Ab initio molecular orbital calculations were used to study hydrogen bonding interactions and interatomic distances of a number of hydrogen bonded complexes that are germane to biomolecular structure and function. The calculations were carried out at the STO-3G, 3-21G, 6-31G*, and MP2/6-31G* levels (geometries were fully optimized at each level). For anionic species, 6-31 + G* and MP2/6-31 + G* were also used. In some cases, more sophisticated calculations were also carried out. Whenever possible, the corresponding enthalpy, entropy, and free energy of complexation were calculated. The agreement with the limited quantity of experimental data is good. For comparison, we also carried out semiempirical molecular orbital calculations. In general, AM1 and PM3 give lower interaction enthalpies than the best ab initio results. With regard to structural results, AM1 tends to favor bifurcated structures for O? H-O and N? HO types of hydrogen bonds, but not for hydrogen bonds involving O-H? S and S-H? O, where the usual hydrogen bond patterns are observed. Overall, AM1 geometries are in general in poor agreement with ab initio structural results. On the other hand, PM3 gives geometries similar to the ab initio ones. Hence, from the structural point of view PM3 does show some improvement over AM1. Finally, insights into the formation of cyclic or open formate–water hydrogen bonded complexes are presented. © 1992 by John Wiley & Sons, Inc.     

A model for the appearance of statistical pores in membranes due to selfoscillations

The aim of this paper is to demonstrate the possibility of the appearance of stochastic pores in membranes, irrespective of external reasons. Fluctuations in thickness, due to structural inhomogeneities and the dynamics of amphiphile molecules, can determine the perforation of a membrane equivalent to the appearance of a pore. The height of the energy barrier for membrane perforation (pore formation) is equal to 91 kT. The geometric profile of the pore if of an elliptical toroidal form. Generally, after membrane perforation, the pore evolves to a stable state. Other possibilities are also discussed.     

A mathematical model for seasonal variability of vitamin D due to solar radiation

It is widely recognized that vitamin D deficiency has detrimental health consequences. The ultraviolet (UV) B radiation increases the serum vitamin D level, expressed by 25-hydroxyvitamin-D(3) [25(OH)D]. An analytical model is presented to calculate the serum 25(OH)D changes throughout a year, caused by the solar exposure variability due to geophysical and habitual factors. The model is tuned by taking into account recent experimental results of serum 25(OH)D changes, after a series of artificial (by fluorescent tubes) UV exposures. The model uses the erythemal and vitamin D weighted irradiances, inferred from the Brewer spectrophotometer and the Kipp and Zonen broad-band meter measurements, carried out in Belsk (52°N, 21°E), Poland, in 2010. The modeled seasonal pattern of the serum 25(OH)D concentration in Polish indoor workers is only slightly different, than in subjects with typical outdoor activity habits, and in those with sun-seeking behavior. A deep minimum in the serum 25(OH)D concentration appears in late winter, regardless of outdoor activity habits. An extra sunbathing to boost the vitamin D level is not worth taking, because of a minor improvement of the vitamin D status, and because of a greater erythema risk. It would be much safer and more effective to maintain an adequate vitamin D level through diet supplements, even in summer, for non sun-seeking subjects.