A STUDY OF AQUEOUS SOLUTIONS OF NUCLEIC ACID CONSTITUENTS EXPOSED TO MONOCHROMATIC 160 nm VACUUM-UV LIGHT BY SPIN-TRAPPING METHOD

Spin-trapping technique was employed to detect and identify free radical intermediates produced in aqueous solutions of nucleic acid constituents (pyrimidine bases and pyrimidine nucleosides) after irradiation by monochromatic 160 nm vacuum-UV light from the electron storage ring. Short-lived free radicals produced in the molecules were converted into relatively long-lived free radicals (spin-adducts) by the reaction with MNP used as a spin trap. The resulting nitroxide radicals were subsequently analysed by esr.
The esr measurements were made after irradiating H₂O solution containing sample and the spin trap and once freeze-drying it and re-dissolving the residual powder in D₂O to get the spectrum with a well-resolved hyperfine structure. Thus, clear evidence that most of the radicals were not formed by H-addition but formed by OH-addition at the C5 position of the 5, 6 double bond were obtained for pyrimidine bases. For pyrimidine nucleosides, although the effect of H₂O-D₂O exchange was not recognized on resolution improvement of the hyperfine structure of the esr spectra, careful analysis of the hyperfine structure made it possible to identify the radical structures; OH-addition radicals at the C6 of the double bond of the base moiety in addition to the OH-addition radicals at the C5 position for all except for 2-deoxycytidine. Evidence for the formation of free radicals at the sugar moiety was not clear.     

Influence of Pb(II) on the radical properties of humic substances and model compounds

The influence of Pb(II) ions on the properties of the free radicals formed in humic acids and fulvic acids was investigated by electron paramagnetic resonance spectroscopy. It is shown that, in both humic acid and fulvic acid, Pb(II) ions shift the radical formation equilibrium by increasing the concentration of stable radicals. Moreover, in both humic acid and fulvic acid, Pb(II) ions cause a characteristic lowering of the stable radicals' g-values to g = 2.0010, which is below the free electron g-value. This effect is unique for Pb ions and is not observed with other dications. Gallic acid (3,4,5-trihydroxybenzoic acid) and tannic acid are shown to be appropriate models for the free radical properties, i.e., g-values, Pb effect, pH dependence, of humic and fulvic acid, respectively. On the basis of density functional theory calculations for the model system (gallic acid-Pb), the observed characteristic g-value reduction upon Pb binding is attributed to the delocalization of the unpaired spin density onto the Pb atom. The present data reveal a novel environmental role of Pb(II) ions on the formation and stabilization of free radicals in natural organic matter.     

Effect of copper(II) ions on the stability of spin adducts formed by the reaction of 5,5-dimethylpyrroline-N-oxide with hydroxyl and hydroxylalkyl radicals

The reactions of copper(II) species with spin adducts formed by reaction of OH·, CH₃CHOH or (CH₃)₂COH with DMPO (5,5-dimethylpyrroline-N-oxide) have been investigated. The spin adducts prepared by -irradiation of appropriate solutions were detected using esr. Copper(II) in the form of aqua, phen, or 1-alanine complexes reacted rapidly with the OH· adduct to give products that gave no esr signal, but with the hydroxylalkyl adducts, reacted to give a new 9-line esr signal, identified as due to the H· adduct, (DMPO-H·). Measurement of the yield of Cu(phen) ₂ ⁺ formed by reaction of the copper(II) complex with the spin adduct showed that approximately 25% of the radiation-produced radicals formed esr detectable spin adducts. The implications of these findings for the use of spin trapping to detect radical intermediates in biological systems is discussed.     

The Effects of a Strong Disaggregating Agent on Sec-Page of Aquatic and Soil Humic Matter

Abstract

Size-exclusion chromatographic (SEC) fractionation and electrophoretic separation of aquatic humic matter samples from a Finnish lake using Sephadex G-75 with 7 M urea solution as eluent and 10% polyacrylamide gel (PAGE) with urea and sodium dodecyl sulphate solution (SDS), respectively, were performed and compared to similar analyses performed on a Russian chernozem soil humic acid sample and Nordic reference fulvic and humic acid samples. The integrated whole of aquatic humic solutes and soil humic acids were found to exhibit similar SEC-PAGE behaviours. Humic matter was not excessively disaggregated by the 7 M urea and hence SEC-PAGE can with confidence be applied as a coarse, initial fractionation procedure or for certain predeterminations of the structural composition.     

Prediction of Properties of Soil Humic Substances from FTIR Spectra using Partial Least Squares Regression

Abstract

Humic substances (HS) play a key role in aquatic and terrestrial ecosystems. The understanding of the ecological functionalities of HS is based on the analysis of their properties, which is normally a very time-consuming procedure. Therefore we tested the possibility to apply the partial least squares regression (PLSR) method in connection with the mid infrared Fourier transform (FTIR) spectra of a series of soil humic substances for the prediction of different HS properties. The results with humic acid (HA) and fulvic acid (FA) fractions of soil HS from different environments show the possibility to predict several properties of an unknown soil HA with satisfying reliability above all the elemental composition.     

Preparation of N-aryl-ketonitrone spin traps

The syntheses of seven N-aryl-C,C-dialkoxycarbonylnitrones 1–7, six of which were original, were achieved from the appropriate aryl-nitroso compounds. These ketonitrones were found to trap efficiently carbon-centred free radicals in aqueous media, yielding stable aminoxyl radicals whose EPR spectra lasted several days. The two penta-deuterated compounds 6 and 7 were also found to be efficient at trapping methoxyl radical. Their various spin adducts showed simple three line signals, very sensitive to the polarity of the environment. This study represents the very first use of linear ketonitrones as spin traps.     

Separation and determination of dissolved and particulate humic substances in river water

Distribution of humic and fulvic acids in participate or dissolved form is studied by using simple leaching and sorption techniques. After filtration of water sample (100–200 ml), the filter along with suspended particles is treated with 5 ml of chloroform and 3 ml of 0.1 mol/l sodium hydroxide solution. The filter dissolves completely in the organic phase, while the suspended particles remain in the aqueous phase enabling a leaching of humic substances. The leaching is repeated once more with 2 ml of 0.1 mol/l sodium hydroxide solution. The humic and fulvic acids in the combined solution are fractionated at pH l by filtration, where the membrane filter is preliminarily coated with sodium dodecyl sulfate. On the other hand, dissolved humic substances are concentrated from a 50-ml filtered sample by sorption on a DEAE-cellulose column. They are desorbed with 5 ml of 0.1 mol/l sodium hydroxide solution and fractionated at pH 1. The spectrophotometric analysis of river water reveals that fulvic acid is predominant in suspended particles as well as in filtered samples. The concentration of dissolved humic and fulvic acids is approximately ten times that of suspended particles.     

Quantitative EPR study on free radicals in the natural polyphenols interacting with metal ions and other environmental pollutants

Quantitative electron paramagnetic resonance (EPR) method was applied to characterise radicals stabilised in polyphenolic matrices of various biogenic materials: lichens, mosses, composts, soils, peats, brown coals and sewage sludge sediments. The investigations were carried out on raw materials and extracted fractions of humic acids (HA). General trends of g value and spin concentration changes were found. These parameters in lichens strongly depend on lichen species and air pollution. Determination of the g value and semiquinone spin concentration allows to assess degree of transformation of organic matter in compost, soil, peat and lignite. Application of gaseous ammonia as a base penetrating the organic matrices extends the possibilities and usefulness of the method. Interaction of metal ions with humic materials is illustrated by interaction of VO2+ ion with peat and lignite HA as well as demineralised (raw and carbonised) brown coal. Our investigations demonstrate that quantitative EPR is a rapid and effective monitoring method to study the influence of various environmental factors on substances containing polyphenolic matrices.     

On the application of electron paramagnetic resonance in the study of naturally occurring quinones and quinols

From electron paramagnetic resonance (EPR) spectra of numerous semiquinone anion radicals we propose that the spin density distributions in benzo-, naphtho- and anthrasemiquinones are changed under the influence of various substituent in a comparable fashion, electron donating substituents having the opposite effect of withdrawing ones. These findings facilitate the work in obtaining correct assignments, further leading to safe identification of new radicals. Semiquinone anion radicals of para- and ortho-hydroquinones are demonstrated to exist at physiological pH in buffered solutions under continuous access to molecular oxygen. Reaction schemes are proposed for the formation of primary and tertiary radicals and hydroxylated products of ten quinols. From studies on crude plant extracts are three quinols, known as precursors in the biogenesis of chimaphilin, observed and identified in Chimaphila umbellata. Other studies demonstrate the power of using semiquinone radicals as chemotaxonomic markers.     

Pulse radiolysis studies of vitaminK1 in an aqueous–organic mixed solvent

Pulse radiolysis studies of VK₁ were carried out in a nitrogen-saturated aqueous–isopropanol–acetone mixed solvent. We studied the spectroscopic characteristics of semiquinone radicals of VK₁ and determined the kinetic parameters of formation and decay of the species. It was found that the decay process of semiquinone radicals of VK₁ was according to a first-order reaction, this was different from the case in ethanol solution, possible reasons were also analyzed.     

Rapid separation of humic acid in fresh waters by coprecipitation and flotation

After removal of suspended matter in 1 liter of water by flotation with a cationic surfactant, humic acid at theg/l level is separated from fulvic acid by coprecipitation with milligram quantities of iron(III) hydroxide at pH 7 followed by flotation with anionic surfactants. The iron(III) hydroxide is dissolved in 2M hydrochloric acid, and the acid-insoluble humic acid is filtered off on an ultrafilter and then dissolved in 10 ml of 0.1M potassium hydroxide solution for measurements of absorption spectra, molecular weight distribution and complexing ability. The time required for the separation is ca. 1 h.     

ESR molecular dynamics study at the oxidation sites of polypropylene with nitroxyl spin labels and spin probes

Novel methods suitable for the selective insertion of nitroxyl spin labels and spin probes at the oxidation sites of isotactic polypropylene have been developed based on reactions with the oxidation products, on spin trapping with nitroso and nitrone compounds and taking advantage of the spontaneous generation of nitroxyl radicals as intermediates in the stabilization mechanism by the HALS. The analysis of the esr spectra of the doxyl spin label and of the spin probes from the HALS, performed according to the stochastic Liouville theory, has allowed information to be obtained on the molecular mobility at the oxidation sites as a function of the temperature.     

Dynamic light scattering measurements of particle size development in aqueous humic materials

Dynamic light scattering (DLS) has been used to monitor changes in aggregate sizes of aqueous humic materials as a function of solution properties. Humic and fulvic acids were dissolved at relatively low concentrations (15-30 mg L(-1)) in solutions of different temperature, cation and ethanol content, and pH. The results could be explained in terms of intramolecular contraction and intermolecular aggregation of humic polymers. The former were prevalent in soil humic acids, and less so in aquatic humic acids and fulvic acids. Increasing the temperature of humic solutions generally led to an increase in particle sizes, which was ascribed to an effect akin to surfactant clouding. The addition of cations led to either contraction or expansion, depending on the charge and concentration of the ion, and the nature of the humic material. Reducing the pH initially caused contraction, followed by growth and precipitation in more highly acidic media.     

Carbon Black Detector,A Novel Approach to Visible Spectroscopy of Liquids Using Photoacoustic Detection

Abstract

A simple and reproducible method of recording visible spectra of solutions using photoacoustic spectrometry and carbon black as a thermal detector is introduced. Light intensity is proportionally measured by carbon black detector, a microphone system and a simple cell compartment placed after monochromator for liquid samples. Exact match with the absorption spectra is found with certain standard solutions. Aqueous solutions of Cr(III), KMnO₄, disodium fluorescein and chlorpromazine semiquinone cation radicals are tested.     

Dynamic conductivity measurements in humic and fulvic acid solutions

Conductivity changes of dilute aqueous humic and fulvic acids solutions were monitored after the addition of small quantities of Cu, Cd, Pb, and Zn. The solutions were stirred at a constant and reproducible rate, and measurements proceeded until stable conductivities were attained. The values were compared to KCl reference solutions treated in an identical manner, and the results showed that the humic materials significantly reduced metal ion mobilities. Pb and Cu were affected more than Cd and Zn, and especially Zn regained a significant portion of its mobility upon dispersal. Cu, in contrast, was strongly and irreversibly retarded in all cases. It was inferred that Zn underwent transient electrostatic interactions with humics at high local concentration, while associations between Cu and humics were rapid and permanent. Different humic materials showed these effects to different extents. Raising the temperature from 20 to 60 °C did not, as expected, lead to higher relative conductivities in metal-humate solutions, but showed enhanced retardation of the cations. It was noted that temperature induced aggregation (clouding) of humic solutions increased the effective sizes of the metal-humate complexes, reducing their mobilities in aqueous solution.     

Theoretical studies of electron and hydrogen transfer reactions between semiquinone radicals and oxygen

 To explore the interactions between ubiquinones and oxygen in living organisms, the thermodynamics of a series of electron and hydrogen transfer reactions between semiquinone radicals, as well as their corresponding protonated forms, and oxygen, singlet or triplet, were studied using the hybrid Hartree–Fock–density functional theory method Becke's three parameter hybrid method with the Lee, Yang, and Parr correlation functional. Effects of the solvent and of the isoprenyl tail on the electron and hydrogen transfer reactions were also investigated. It is found that semiquinone radicals (semiquinone anion radicals or protonated semiquinone radicals) cannot react with triplet oxygen to form the superoxide anion radical O₂ ⁻. In contrast, neutral quinones can scavenge O₂ ⁻ efficiently. In the gas phase, only protonated semiquinone radicals can react spontaneously with singlet oxygen to produce peroxyl radical (HO₂). However, both semiquinone anion radicals and protonated semiquinone radicals can react with singlet oxygen to produce harmful oxygen radicals (O₂ ^{− a l l b u l l} and HO₂, respectively) in aqueous and protein environments. The free-energy changes of the corresponding reactions obtained for different ubiquinone systems are very similar. It clearly shows that the isoprenyl tail does not influence the electron and hydrogen transfer reactions between semiquinone radicals and oxygen significantly. Results of electron affinities, vertical ionization potentials, and proton affinities also show that the isoprenyl tail has no substantial effect on the electronic properties of ubiquinones. Received: 3 July 2000 / Accepted: 6 September 2000 / Published online: 21 December 2000     

(Perhalogenmethylthio)heterocyclen. XXI. Reaktionen halogenierter perhalogenmethylthiopyrrole: Salz- und radikalbildung,dimerisierung

2-Chloro-3,4,5-tris(trifluoromethylthio)pyrrole ( 2a ), 3-Chloro-2,4,5-tris(trifluoromethylthio)pyarrole ( 2b ) and 3,4-dichloro-2,5-bis(trifluromoethylthio)pyrrole ( 2c ) react with silver nitrate/silver acetate in good yield to give the corresponding N-silver salts 3a-c . Compound 2b forms with an aqueous potassium hydroxide solution the N-potassium salt 4 . Compounds 3a and 3b react with iodine to give the dimeers 2,2′-dichloro-3,3,′ 4,4′5,5′-hexakis(trifluoromethylthio)-2,2′-bi-2H-pyrrolyl ( 5a ) and 3,3′-dichloro-2,2′,4,4′,5,5′-hexakis(trifluoromethylthio)-2,2′-bi-2H-pyrrolyl ( 5b ). The dimers dissociate in solution to the corresponding pyrrolayl radicals. The esr and endor spectra of 3-chloro-2,4,5-tris(trifluoromethylthio)pyrrolyl were measured; coupling constants are given. For the newly prepared substances melting-points, ¹⁹F-nmr and ir spectroscopical data are provided.     

Separation and determination of traces of heavy metals complexed with humic substances in fresh waters by sorption on diethylaminoethyl-sephadex a-25

Copper(II), lead and cadmium complexed with humic and fulvic acids in filtered 1-l samples of fresh water are sorbed on a column containing 0.5 ml of the macroreticular weak-base anion exchanger, diethylaminoethyl-Sephadex A-25 at a flow rate of 20 ml min^?1. Simple metal cations are not sorbed at all. The sorbed trace metals are quantitatively desorbed with 4 M nitric acid batchwise and determined by graphite-furnace atomic absorption spectrometry. For synthetic aqueous solutions containing traces of heavy metals and humic acid, the results are in conformity with those obtained by cationexchange separation. About 80% of the sorbed humic substances are eluted with 0.5 M sodium hydroxide solution from the A-25 column and its quantity is estimated spectrophotometrically at 400 nm.     

Interaction of Humic Substances with Organic Pollutants and its Influence on the Concentration of these Pollutants from Water

Abstract

The influence of fulvic acids (FAs) and humic acids (HAs) of different origin on liquid-liquid and solid-phase extraction of hydrophobic organic compounds (HOCs) with broadly varying values of octanol-water partition coefficients K_ow has been investigated. It is shown that the solubilization by dissolved FAs and HAs can lead to a significant decrease in recovery of some HOCs even from very dilute (10–20 mg/l) aqueous solutions of these acids. The results of these investigations and an analysis of the relevant literature data, demonstrate that the extent of solubilization of HOCs by humic substances is determined not only by K_ow, but also by the specific structure of the HOCs. The hypothesis of the formation of intramolecular micelles (IMMs) by unaggregated FAs and HAs, and the partition of HOCs between water and the IMMs is put forward to explain the solubilization of HOCs.